@article {3046, title = {Engineering Geobacter pili to produce metal:organic filaments.}, journal = {Biosens Bioelectron}, volume = {222}, year = {2023}, month = {2023 Feb 15}, pages = {114993}, abstract = {
The organized self-assembly of conductive biological structures holds promise for creating new bioelectronic devices. In particular, Geobacter sulfurreducens type IVa pili have proven to be a versatile material for fabricating protein nanowire-based devices. To scale the production of conductive pili, we designed a strain of Shewanella oneidensis that heterologously expressed abundant, conductive Geobacter pili when grown aerobically in liquid culture. S. oneidensis expressing a cysteine-modified pilin, designed to enhance the capability to bind to gold, generated conductive pili that self-assembled into biohybrid filaments in the presence of gold nanoparticles. Elemental composition analysis confirmed the filament-metal interactions within the structures, which were several orders of magnitude larger than previously described metal:organic filaments. The results demonstrate that the S. oneidensis chassis significantly advances the possibilities for facile conductive protein nanowire design and fabrication.
}, keywords = {Biosensing Techniques, Electron Transport, Fimbriae, Bacterial, Geobacter, Gold, Metal Nanoparticles}, issn = {1873-4235}, doi = {10.1016/j.bios.2022.114993}, author = {Szmuc, Eric and Walker, David J F and Kireev, Dmitry and Akinwande, Deji and Lovley, Derek R and Keitz, Benjamin and Ellington, Andrew} } @article {3045, title = {H Is a Major Intermediate in Corrosion of Iron.}, journal = {mBio}, volume = {14}, year = {2023}, month = {2023 Apr 25}, pages = {e0007623}, abstract = {Desulfovibrio vulgaris has been a primary pure culture sulfate reducer for developing microbial corrosion concepts. Multiple mechanisms for how it accepts electrons from Fe have been proposed. We investigated Fe oxidation with a mutant of in which hydrogenase genes were deleted. The hydrogenase mutant grew as well as the parental strain with lactate as the electron donor, but unlike the parental strain, it was not able to grow on H. The parental strain reduced sulfate with Fe as the sole electron donor, but the hydrogenase mutant did not. H accumulated over time in Fe cultures of the hydrogenase mutant and sterile controls but not in parental strain cultures. Sulfide stimulated H production in uninoculated controls apparently by both reacting with Fe to generate H and facilitating electron transfer from Fe to H. Parental strain supernatants did not accelerate H production from Fe, ruling out a role for extracellular hydrogenases. Previously proposed electron transfer between Fe and via soluble electron shuttles was not evident. The hydrogenase mutant did not reduce sulfate in the presence of Fe and either riboflavin or anthraquinone-2,6-disulfonate, and these potential electron shuttles did not stimulate parental strain sulfate reduction with Fe as the electron donor. The results demonstrate that primarily accepts electrons from Fe via H as an intermediary electron carrier. These findings clarify the interpretation of previous corrosion studies and suggest that H-mediated electron transfer is an important mechanism for iron corrosion under sulfate-reducing conditions. Microbial corrosion of iron in the presence of sulfate-reducing microorganisms is economically significant. There is substantial debate over how microbes accelerate iron corrosion. Tools for genetic manipulation have only been developed for a few Fe(III)-reducing and methanogenic microorganisms known to corrode iron and in each case those microbes were found to accept electrons from Fe via direct electron transfer. However, iron corrosion is often most intense in the presence of sulfate-reducing microbes. The finding that Desulfovibrio vulgaris relies on H to shuttle electrons between Fe and cells revives the concept, developed in some of the earliest studies on microbial corrosion, that sulfate reducers consumption of H is a major microbial corrosion mechanism. The results further emphasize that direct Fe-to-microbe electron transfer has yet to be rigorously demonstrated in sulfate-reducing microbes.
}, keywords = {Corrosion, Desulfovibrio, Desulfovibrio vulgaris, Hydrogenase, Iron, Lactic Acid, Oxidation-Reduction, Sulfates}, issn = {2150-7511}, doi = {10.1128/mbio.00076-23}, author = {Woodard, Trevor L and Ueki, Toshiyuki and Lovley, Derek R} } @article {3044, title = {Microbial nanowires with genetically modified peptide ligands to sustainably fabricate electronic sensing devices.}, journal = {Biosens Bioelectron}, volume = {226}, year = {2023}, month = {2023 Apr 15}, pages = {115147}, abstract = {Nanowires have substantial potential as the sensor component in electronic sensing devices. However, surface functionalization of traditional nanowire and nanotube materials with short peptides that increase sensor selectivity and sensitivity requires complex chemistries with toxic reagents. In contrast, microorganisms can assemble pilin monomers into protein nanowires with intrinsic conductivity from renewable feedstocks, yielding an electronic material that is robust and stable in applications, but also biodegradable. Here we report that the sensitivity and selectivity of protein nanowire-based sensors can be modified with a simple plug and play genetic approach in which a short peptide sequence, designed to bind the analyte of interest, is incorporated into the pilin protein that is microbially assembled into nanowires. We employed a scalable Escherichia coli chassis to fabricate protein nanowires that displayed either a peptide previously demonstrated to effectively bind ammonia, or a peptide known to bind acetic acid. Sensors comprised of thin films of the nanowires amended with the ammonia-specific peptide had a ca. 100-fold greater response to ammonia than sensors made with unmodified protein nanowires. Protein nanowires with the peptide that binds acetic acid yielded a 4-fold higher response than nanowires without the peptide. The protein nanowire-based sensors had greater responses than previously reported sensors fabricated with other nanomaterials. The results demonstrate that protein nanowires with enhanced sensor response for analytes of interest can be fabricated with a flexible genetic strategy that sustainably eliminates the energy, environmental, and health concerns associated with other common nanomaterials.
}, keywords = {Acetic Acid, Ammonia, Biosensing Techniques, Electronics, Fimbriae Proteins, Ligands, Nanowires, Peptides}, issn = {1873-4235}, doi = {10.1016/j.bios.2023.115147}, author = {Lekbach, Yassir and Ueki, Toshiyuki and Liu, Xiaomeng and Woodard, Trevor and Yao, Jun and Lovley, Derek R} } @article {3043, title = {Response to Wang et al.: evidence contradicting the cytochrome-only model.}, journal = {Trends Microbiol}, volume = {31}, year = {2023}, month = {2023 Jun}, pages = {548-549}, keywords = {Cytochromes, Electron Transport}, issn = {1878-4380}, doi = {10.1016/j.tim.2023.03.006}, author = {Lovley, Derek R} } @article {3051, title = {Direct microbial electron uptake as a mechanism for stainless steel corrosion in aerobic environments.}, journal = {Water Res}, volume = {219}, year = {2022}, month = {2022 Jul 01}, pages = {118553}, abstract = {Shewanella oneidensis MR-1 is an attractive model microbe for elucidating the biofilm-metal interactions that contribute to the billions of dollars in corrosion damage to industrial applications each year. Multiple mechanisms for S. oneidensis-enhanced corrosion have been proposed, but none of these mechanisms have previously been rigorously investigated with methods that rule out alternative routes for electron transfer. We found that S. oneidensis grown under aerobic conditions formed thick biofilms (\~{}50~{\textmu}m) on stainless steel coupons, accelerating corrosion over sterile controls. H and flavins were ruled out as intermediary electron carriers because stainless steel did not reduce riboflavin and previous studies have demonstrated stainless does not generate H. Strain ∆mtrCBA, in which the genes for the most abundant porin-cytochrome conduit in S. oneidensis were deleted, corroded stainless steel substantially less than wild-type in aerobic cultures. Wild-type biofilms readily reduced nitrate with stainless steel as the sole electron donor under anaerobic conditions, but strain ∆mtrCBA did not. These results demonstrate that S. oneidensis can directly consume electrons from iron-containing metals and illustrate how direct metal-to-microbe electron transfer can be an important route for corrosion, even in aerobic environments.
}, keywords = {Biofilms, Corrosion, Electron Transport, Electrons, Metals, Oxidation-Reduction, Stainless Steel, Steel}, issn = {1879-2448}, doi = {10.1016/j.watres.2022.118553}, author = {Zhou, Enze and Li, Feng and Zhang, Dawei and Xu, Dake and Li, Zhong and Jia, Ru and Jin, Yuting and Song, Hao and Li, Huabing and Wang, Qiang and Wang, Jianjun and Li, Xiaogang and Gu, Tingyue and Homborg, Axel M and Mol, Johannes M C and Smith, Jessica A and Wang, Fuhui and Lovley, Derek R} } @article {3059, title = {Electromicrobiology: the ecophysiology of phylogenetically diverse electroactive microorganisms.}, journal = {Nat Rev Microbiol}, volume = {20}, year = {2022}, month = {2022 Jan}, pages = {5-19}, abstract = {Electroactive microorganisms markedly affect many environments in which they establish outer-surface electrical contacts with other cells and minerals or reduce soluble extracellular redox-active molecules such as flavins and humic substances. A growing body of research emphasizes their broad phylogenetic diversity and shows that these microorganisms have key roles in multiple biogeochemical cycles, as well as the microbiome of the gut, anaerobic waste digesters and metal corrosion. Diverse bacteria and archaea have independently evolved cytochrome-based strategies for electron exchange between the outer cell surface and the cell interior, but cytochrome-free mechanisms are also prevalent. Electrically conductive protein filaments, soluble electron shuttles and non-biological conductive materials can substantially extend the electronic reach of microorganisms beyond the surface of the cell. The growing appreciation of the diversity of electroactive microorganisms and their unique electronic capabilities is leading to a broad range of applications.
}, keywords = {Archaea, Bacteria, Bacterial Physiological Phenomena, Cytochromes, Electron Transport, Oxidation-Reduction, Phylogeny}, issn = {1740-1534}, doi = {10.1038/s41579-021-00597-6}, author = {Lovley, Derek R and Holmes, Dawn E} } @article {3054, title = {Electrotrophy: Other microbial species, iron, and electrodes as electron donors for microbial respirations.}, journal = {Bioresour Technol}, volume = {345}, year = {2022}, month = {2022 Feb}, pages = {126553}, abstract = {Electrotrophy, the growth of microbes on extracellular electron donors, drives important biogeochemical cycles and has practical applications. Studies of Fe(II)-based electrotrophy have provided foundational cytochrome-based mechanistic models for electron transport into cells. Direct electron uptake from other microbial species, Fe(0), or cathodes is of intense interest due to its potential roles in the production and anaerobic oxidation of methane, corrosion, and bioelectrochemical technologies. Other cells or Fe(0) can serve as the sole electron donor supporting the growth of several Geobacter and methanogen strains that are unable to use H as an electron donor, providing strong evidence for electrotrophy. Additional evidence for electrotrophy in Geobacter strains and Methanosarcina acetivorans is a requirement for outer-surface c-type cytochromes. However, in most instances claims for electrotrophy in anaerobes are based on indirect inference and the possibility that H is actually the electron donor supporting growth has not been rigorously excluded.
}, keywords = {Electrodes, Electron Transport, Electrons, Geobacter, Iron, Oxidation-Reduction, Soil Microbiology}, issn = {1873-2976}, doi = {10.1016/j.biortech.2021.126553}, author = {Lovley, Derek R} } @article {3049, title = {On the Existence of Pilin-Based Microbial Nanowires.}, journal = {Front Microbiol}, volume = {13}, year = {2022}, month = {2022}, pages = {872610}, issn = {1664-302X}, doi = {10.3389/fmicb.2022.872610}, author = {Lovley, Derek R} } @article {3047, title = {Genetic Manipulation of Desulfovibrio ferrophilus and Evaluation of Fe(III) Oxide Reduction Mechanisms.}, journal = {Microbiol Spectr}, volume = {10}, year = {2022}, month = {2022 Dec 21}, pages = {e0392222}, abstract = {The sulfate-reducing microbe Desulfovibrio ferrophilus is of interest due to its relatively rare ability to also grow with Fe(III) oxide as an electron acceptor and its rapid corrosion of metallic iron. Previous studies have suggested multiple agents for extracellular electron exchange including a soluble electron shuttle, electrically conductive pili, and outer surface multiheme -type cytochromes. However, the previous lack of a strategy for genetic manipulation of limited mechanistic investigations. We developed an electroporation-mediated transformation method that enabled replacement of genes of interest with an antibiotic resistance gene via double-crossover homologous recombination. Genes were identified that are essential for flagellum-based motility and the expression of the two types of pili. Disrupting flagellum-based motility or expression of either of the two pili did not inhibit Fe(III) oxide reduction, nor did deleting genes for multiheme -type cytochromes predicted to be associated with the outer membrane. Although redundancies in cytochrome or pilus function might explain some of these phenotypes, overall, the results are consistent with primarily reducing Fe(III) oxide via an electron shuttle. The finding that is genetically tractable not only will aid in elucidating further details of its mechanisms for Fe(III) oxide reduction but also provides a new experimental approach for developing a better understanding of some of its other unique features, such as the ability to corrode metallic iron at high rates and accept electrons from negatively poised electrodes. is an important pure culture model for Fe(III) oxide reduction and the corrosion of iron-containing metals in anaerobic marine environments. This study demonstrates that is genetically tractable, an important advance for elucidating the mechanisms by which it interacts with extracellular electron acceptors and donors. The results demonstrate that there is not one specific outer surface multiheme -type cytochrome that is essential for Fe(III) oxide reduction. This finding, coupled with the lack of apparent porin-cytochrome conduits encoded in the genome and the finding that deleting genes for pilus and flagellum expression did not inhibit Fe(III) oxide reduction, suggests that has adopted strategies for extracellular electron exchange that are different from those of intensively studied electroactive microbes like and species. Thus, the ability to genetically manipulate is likely to lead to new mechanistic concepts in electromicrobiology.
}, keywords = {Cytochromes, Electron Transport, Ferric Compounds, Iron, Oxidation-Reduction, Oxides}, issn = {2165-0497}, doi = {10.1128/spectrum.03922-22}, author = {Ueki, Toshiyuki and Woodard, Trevor L and Lovley, Derek R} } @article {3052, title = {Microbe Profile: : a model for novel physiologies of biogeochemical and technological significance.}, journal = {Microbiology (Reading)}, volume = {168}, year = {2022}, month = {2022 Feb}, abstract = {has served as the initial model for a substantial number of newly recognized microbial physiologies that play an important role in biogeochemical cycling of carbon, metals and nutrients. The strategies used by for microbial interaction with minerals, contaminants, other microbes and electrodes have led to new technologies for bioremediation, bioenergy conversion and the sustainable production of {\textquoteright}green{\textquoteright} electronics.
}, keywords = {Biodegradation, Environmental, Electron Transport, Geobacter, Microbial Interactions, Oxidation-Reduction}, issn = {1465-2080}, doi = {10.1099/mic.0.001138}, author = {Lovley, Derek R} } @article {3048, title = {Microbial biofilms for electricity generation from water evaporation and power to wearables.}, journal = {Nat Commun}, volume = {13}, year = {2022}, month = {2022 Jul 28}, pages = {4369}, abstract = {Employing renewable materials for fabricating clean energy harvesting devices can further improve sustainability. Microorganisms can be mass produced with renewable feedstocks. Here, we demonstrate that it is possible to engineer microbial biofilms as a cohesive, flexible material for long-term continuous electricity production from evaporating water. Single biofilm sheet (~40 {\textmu}m thick) serving as the functional component in an electronic device continuously produces power density (~1 μW/cm) higher than that achieved with thicker engineered materials. The energy output is comparable to that achieved with similar sized biofilms catalyzing current production in microbial fuel cells, without the need for an organic feedstock or maintaining cell viability. The biofilm can be sandwiched between a pair of mesh electrodes for scalable device integration and current production. The devices maintain the energy production in ionic solutions and can be used as skin-patch devices to harvest electricity from sweat and moisture on skin to continuously power wearable devices. Biofilms made from different microbial species show generic current production from water evaporation. These results suggest that we can harness the ubiquity of biofilms in nature as additional sources of biomaterial for evaporation-based electricity generation in diverse aqueous environments.
}, keywords = {Bioelectric Energy Sources, Biofilms, Electricity, Electrodes, Water, Wearable Electronic Devices}, issn = {2041-1723}, doi = {10.1038/s41467-022-32105-6}, author = {Liu, Xiaomeng and Ueki, Toshiyuki and Gao, Hongyan and Woodard, Trevor L and Nevin, Kelly P and Fu, Tianda and Fu, Shuai and Sun, Lu and Lovley, Derek R and Yao, Jun} } @article {3053, title = {Microbial nanowires.}, journal = {Curr Biol}, volume = {32}, year = {2022}, month = {2022 Feb 07}, pages = {R110-R112}, abstract = {In this Quick guide, Derek Lovley introduces microbial nanowires-conductive extracellular appendages made by some bacteria and archaea.
}, keywords = {Bacteria, Electric Conductivity, Electron Transport, Fimbriae, Bacterial, Nanowires}, issn = {1879-0445}, doi = {10.1016/j.cub.2021.12.019}, author = {Lovley, Derek R} } @article {3050, title = {Untangling Geobacter sulfurreducens Nanowires.}, journal = {mBio}, volume = {13}, year = {2022}, month = {2022 Jun 28}, pages = {e0085022}, keywords = {Electron Transport, Fimbriae Proteins, Fimbriae, Bacterial, Geobacter, Nanowires, Oxidation-Reduction}, issn = {2150-7511}, doi = {10.1128/mbio.00850-22}, author = {Lovley, Derek R} } @article {2624, title = {Chemically Induced Cell Wall Stapling in Bacteria.}, journal = {Cell Chem Biol}, volume = {28}, year = {2021}, month = {2021 Feb 18}, pages = {213-220.e4}, abstract = {Transpeptidation reinforces the structure of cell-wall peptidoglycan, an extracellular heteropolymer that protects bacteria from osmotic lysis. The clinical success of transpeptidase-inhibiting \β-lactam antibiotics illustrates the essentiality of these cross-linkages for cell-wall integrity, but the presence of multiple, seemingly redundant transpeptidases in many species makes it challenging to determine cross-link function. Here, we present a technique to link peptide strands by chemical rather than enzymatic reaction. We employ biocompatible click chemistry to induce triazole formation between azido- and alkynyl-d-alanine residues that are metabolically installed in the peptidoglycan of Gram-positive or Gram-negative bacteria. Synthetic triazole cross-links can be visualized using azidocoumarin-d-alanine, an amino acid derivative that undergoes fluorescent enhancement upon reaction with terminal alkynes. Cell-wall stapling protects Escherichia coli from treatment with the broad-spectrum \β-lactams ampicillin and carbenicillin. Chemical control of cell-wall structure in live bacteria can provide functional insights that are orthogonal to those obtained by genetics.
}, issn = {2451-9448}, doi = {10.1016/j.chembiol.2020.11.006}, author = {Rivera, Sylvia L and Espaillat, Akbar and Aditham, Arjun K and Shieh, Peyton and Muriel-Mundo, Chris and Kim, Justin and Cava, Felipe and Siegrist, M Sloan} } @article {3063, title = {Correlation of Key Physiological Properties of Isolates with Environment of Origin.}, journal = {Appl Environ Microbiol}, volume = {87}, year = {2021}, month = {2021 Jun 11}, pages = {e0073121}, abstract = {It is known that the physiology of species can differ significantly, but the ecological impact of these differences is unclear. We recovered two strains of from two different ecosystems with a similar enrichment and isolation method. Both strains had the same ability to metabolize organic substrates and participate in direct interspecies electron transfer but also had major physiological differences. Strain DH-1, which was isolated from an anaerobic digester, used H as an electron donor. Genome analysis indicated that it lacks an Rnf complex and conserves energy from acetate metabolism via intracellular H cycling. In contrast, strain DH-2, a subsurface isolate, lacks hydrogenases required for H uptake and cycling and has an Rnf complex for energy conservation when growing on acetate. Further analysis of the genomes of previously described isolates, as well as phylogenetic and metagenomic data on uncultured in anaerobic digesters and diverse soils and sediments, revealed a physiological dichotomy that corresponded with environment of origin. The physiology of type I revolves around H production and consumption. In contrast, type II species eschew H and have genes for an Rnf complex and the multiheme, membrane-bound -type cytochrome MmcA, shown to be essential for extracellular electron transfer. The distribution of species in diverse environments suggests that the type I H-based physiology is well suited for high-energy environments, like anaerobic digesters, whereas type II Rnf/cytochrome-based physiology is an adaptation to the slower, steady-state carbon and electron fluxes common in organic-poor anaerobic soils and sediments. Biogenic methane is a significant greenhouse gas, and the conversion of organic wastes to methane is an important bioenergy process. species play an important role in methane production in many methanogenic soils and sediments as well as anaerobic waste digesters. The studies reported here emphasize that the genus is composed of two physiologically distinct groups. This is important to recognize when interpreting the role of in methanogenic environments, especially regarding H metabolism. Furthermore, the finding that type I species predominate in environments with high rates of carbon and electron flux and that type II species predominate in lower-energy environments suggests that evaluating the relative abundance of type I and type II may provide further insights into rates of carbon and electron flux in methanogenic environments.
}, keywords = {Acetates, Anaerobiosis, Bioreactors, Ecosystem, Electron Transport, Ethanol, Genome, Archaeal, Hydrogen, Methane, Methanosarcina, Phylogeny}, issn = {1098-5336}, doi = {10.1128/AEM.00731-21}, author = {Zhou, Jinjie and Holmes, Dawn E and Tang, Hai-Yan and Lovley, Derek R} } @article {3058, title = {Direct Observation of Electrically Conductive Pili Emanating from .}, journal = {mBio}, volume = {12}, year = {2021}, month = {2021 Aug 31}, pages = {e0220921}, abstract = {Geobacter sulfurreducens is a model microbe for elucidating the mechanisms for extracellular electron transfer in several biogeochemical cycles, bioelectrochemical applications, and microbial metal corrosion. Multiple lines of evidence previously suggested that electrically conductive pili (e-pili) are an essential conduit for long-range extracellular electron transport in G. sulfurreducens. However, it has recently been reported that G. sulfurreducens does not express e-pili and that filaments comprised of multi-heme -type cytochromes are responsible for long-range electron transport. This possibility was directly investigated by examining cells, rather than filament preparations, with atomic force microscopy. Approximately 90\% of the filaments emanating from wild-type cells had a diameter (3 nm) and conductance consistent with previous reports of e-pili harvested from G. sulfurreducens or heterologously expressed in Escherichia coli from the G. sulfurreducens pilin gene. The remaining 10\% of filaments had a morphology consistent with filaments comprised of the -type cytochrome OmcS. A strain expressing a modified pilin gene designed to yield poorly conductive pili expressed 90\% filaments with a 3-nm diameter, but greatly reduced conductance, further indicating that the 3-nm diameter conductive filaments in the wild-type strain were e-pili. A strain in which genes for five of the most abundant outer-surface -type cytochromes, including OmcS, were deleted yielded only 3-nm-diameter filaments with the same conductance as in the wild type. These results demonstrate that e-pili are the most abundant conductive filaments expressed by G. sulfurreducens, consistent with previous functional studies demonstrating the need for e-pili for long-range extracellular electron transfer. Electroactive microbes have significant environmental impacts, as well as applications in bioenergy and bioremediation. The composition, function, and even existence of electrically conductive pili (e-pili) has been one of the most contentious areas of investigation in electromicrobiology, in part because e-pili offer a mechanism for long-range electron transport that does not involve the metal cofactors common in much of biological electron transport. This study demonstrates that e-pili are abundant filaments emanating from Geobacter sulfurreducens, which serves as a model for long-range extracellular electron transfer in direct interspecies electron transfer, dissimilatory metal reduction, microbe-electrode exchange, and corrosion caused by direct electron uptake from Fe(0). The methods described in this study provide a simple strategy for evaluating the distribution of conductive filaments throughout the microbial world with an approach that avoids artifactual production and/or enrichment of filaments that may not be physiologically relevant.
}, keywords = {Electric Conductivity, Electrons, Escherichia coli, Fimbriae Proteins, Fimbriae, Bacterial, Geobacter, Microscopy, Atomic Force, Oxidation-Reduction}, issn = {2150-7511}, doi = {10.1128/mBio.02209-21}, author = {Liu, Xinying and Walker, David J F and Nonnenmann, Stephen S and Sun, Dezhi and Lovley, Derek R} } @article {3055, title = {Extracellular Electron Exchange Capabilities of and .}, journal = {Environ Sci Technol}, volume = {55}, year = {2021}, month = {2021 Dec 07}, pages = {16195-16203}, abstract = {Microbial extracellular electron transfer plays an important role in diverse biogeochemical cycles, metal corrosion, bioelectrochemical technologies, and anaerobic digestion. Evaluation of electron uptake from pure Fe(0) and stainless steel indicated that, in contrast to previous speculation in the literature, and are not able to directly extract electrons from solid-phase electron-donating surfaces. grew with Fe(III) as the electron acceptor, but did not. reduced Fe(III) oxide occluded within porous alginate beads, suggesting that it released a soluble electron shuttle to promote Fe(III) oxide reduction. Conductive atomic force microscopy revealed that the pili are electrically conductive and the expression of a gene encoding an aromatics-rich putative pilin was upregulated during growth on Fe(III) oxide. The expression of genes for multi-heme -type cytochromes was not upregulated during growth with Fe(III) as the electron acceptor, and genes for a porin-cytochrome conduit across the outer membrane were not apparent in the genome. The results suggest that has adopted a novel combination of strategies to enable extracellular electron transport, which may be of biogeochemical and technological significance.
}, keywords = {Desulfovibrio, Electron Transport, Electrons, Ferric Compounds, Geobacter, Oxidation-Reduction}, issn = {1520-5851}, doi = {10.1021/acs.est.1c04071}, author = {Liang, Dandan and Liu, Xinying and Woodard, Trevor L and Holmes, Dawn E and Smith, Jessica A and Nevin, Kelly P and Feng, Yujie and Lovley, Derek R} } @article {3057, title = {Generation of High Current Densities in Geobacter sulfurreducens Lacking the Putative Gene for the PilB Pilus Assembly Motor.}, journal = {Microbiol Spectr}, volume = {9}, year = {2021}, month = {2021 Oct 31}, pages = {e0087721}, abstract = {Geobacter sulfurreducens is commonly employed as a model for the study of extracellular electron transport mechanisms in the species. Deletion of , which is known to encode the pilus assembly motor protein for type IV pili in other bacteria, has been proposed as an effective strategy for evaluating the role of electrically conductive pili (e-pili) in G. sulfurreducens extracellular electron transfer. In those studies, the inhibition of e-pili expression associated with deletion was not demonstrated directly but was inferred from the observation that deletion mutants produced lower current densities than wild-type cells. Here, we report that deleting did not diminish current production. Conducting probe atomic force microscopy revealed filaments with the same diameter and similar current-voltage response as e-pili harvested from wild-type G. sulfurreducens or when e-pili are expressed heterologously from the G. sulfurreducens pilin gene in Escherichia coli. Immunogold labeling demonstrated that a G. sulfurreducens strain expressing a pilin monomer with a His tag continued to express His tag-labeled filaments when was deleted. These results suggest that a reinterpretation of the results of previous studies on G. sulfurreducens deletion strains may be necessary. Geobacter sulfurreducens is a model microbe for the study of biogeochemically and technologically significant processes, such as the reduction of Fe(III) oxides in soils and sediments, bioelectrochemical applications that produce electric current from waste organic matter or drive useful processes with the consumption of renewable electricity, direct interspecies electron transfer in anaerobic digestors and methanogenic soils and sediments, and metal corrosion. Elucidating the phenotypes associated with gene deletions is an important strategy for determining the mechanisms for extracellular electron transfer in G. sulfurreducens. The results reported here demonstrate that we cannot replicate the key phenotype reported for a gene deletion that has been central to the development of models for long-range electron transport in G. sulfurreducens.
}, keywords = {Bacterial Proteins, Electric Conductivity, Electron Transport, Fimbriae Proteins, Fimbriae, Bacterial, Gene Deletion, Geobacter, Geologic Sediments, Microscopy, Atomic Force, Oxidoreductases}, issn = {2165-0497}, doi = {10.1128/Spectrum.00877-21}, author = {Ueki, Toshiyuki and Walker, David J F and Nevin, Kelly P and Ward, Joy E and Woodard, Trevor L and Nonnenmann, Stephen S and Lovley, Derek R} } @article {3065, title = {Intrinsically Conductive Microbial Nanowires for {\textquoteright}Green{\textquoteright} Electronics with Novel Functions.}, journal = {Trends Biotechnol}, volume = {39}, year = {2021}, month = {2021 Sep}, pages = {940-952}, abstract = {Intrinsically conductive protein nanowires, microbially produced from inexpensive, renewable feedstocks, are a sustainable alternative to traditional nanowire electronic materials, which require high energy inputs and hazardous conditions/chemicals for fabrication and can be highly toxic. Pilin-based nanowires can be tailored for specific functions via the design of synthetic pilin genes to tune wire conductivity or introduce novel functionalities. Other microbially produced nanowire options for electronics may include cytochrome wires, curli fibers, and the conductive fibers of cable bacteria. Proof-of-concept protein nanowire electronics that have been successfully demonstrated include biomedical sensors, neuromorphic devices, and a device that generates electricity from ambient humidity. Further development of applications will require interdisciplinary teams of engineers, biophysicists, and synthetic biologists.
}, keywords = {Electric Conductivity, Electronics, Nanowires, Proteins}, issn = {1879-3096}, doi = {10.1016/j.tibtech.2020.12.005}, author = {Lovley, Derek R and Yao, Jun} } @article {3056, title = {Mechanisms for Electron Uptake by Methanosarcina acetivorans during Direct Interspecies Electron Transfer.}, journal = {mBio}, volume = {12}, year = {2021}, month = {2021 Oct 26}, pages = {e0234421}, abstract = {Direct interspecies electron transfer (DIET) between bacteria and methanogenic archaea appears to be an important syntrophy in both natural and engineered methanogenic environments. However, the electrical connections on the outer surface of methanogens and the subsequent processing of electrons for carbon dioxide reduction to methane are poorly understood. Here, we report that the genetically tractable methanogen Methanosarcina acetivorans can grow via DIET in coculture with Geobacter metallireducens serving as the electron-donating partner. Comparison of gene expression patterns in grown in coculture versus pure-culture growth on acetate revealed that transcripts for the outer-surface multiheme type cytochrome MmcA were higher during DIET-based growth. Deletion of inhibited DIET. The high aromatic amino acid content of archaellins suggests that they might assemble into electrically conductive archaella. A mutant that could not express archaella was deficient in DIET. However, this mutant grew in DIET-based coculture as well as the archaellum-expressing parental strain in the presence of granular activated carbon, which was previously shown to serve as a substitute for electrically conductive pili as a conduit for long-range interspecies electron transfer in other DIET-based cocultures. Transcriptomic data suggesting that the membrane-bound Rnf, Fpo, and HdrED complexes also play a role in DIET were incorporated into a charge-balanced model illustrating how electrons entering the cell through MmcA can yield energy to support growth from carbon dioxide reduction. The results are the first genetics-based functional demonstration of likely outer-surface electrical contacts for DIET in a methanogen. The conversion of organic matter to methane plays an important role in the global carbon cycle and is an effective strategy for converting wastes to a useful biofuel. The reduction of carbon dioxide to methane accounts for approximately a third of the methane produced in anaerobic soils and sediments as well as waste digesters. Potential electron donors for carbon dioxide reduction are H or electrons derived from direct interspecies electron transfer (DIET) between bacteria and methanogens. Elucidating the relative importance of these electron donors has been difficult due to a lack of information on the electrical connections on the outer surfaces of methanogens and how they process the electrons received from DIET. Transcriptomic patterns and gene deletion phenotypes reported here provide insight into how a group of organisms that play an important role in methane production in soils and sediments participate in DIET.
}, keywords = {Archaeal Proteins, Electron Transport, Electrons, Methane, Methanosarcina, Transcriptome}, issn = {2150-7511}, doi = {10.1128/mBio.02344-21}, author = {Holmes, Dawn E and Zhou, Jinjie and Ueki, Toshiyuki and Woodard, Trevor and Lovley, Derek R} } @article {2626, title = {Membrane-partitioned cell wall synthesis in mycobacteria.}, journal = {Elife}, volume = {10}, year = {2021}, month = {2021 Feb 05}, abstract = {Many antibiotics target the assembly of cell wall peptidoglycan, an essential, heteropolymeric mesh that encases most bacteria. In rod-shaped bacteria, cell wall elongation is spatially precise yet relies on limited pools of lipid-linked precursors that generate and are attracted to membrane disorder. By tracking enzymes, substrates, and products of peptidoglycan biosynthesis in , we show that precursors are made in plasma membrane domains that are laterally and biochemically distinct from sites of cell wall assembly. Membrane partitioning likely contributes to robust, orderly peptidoglycan synthesis, suggesting that these domains help template peptidoglycan synthesis. The cell wall-organizing protein DivIVA and the cell wall itself promote domain homeostasis. These data support a model in which the peptidoglycan polymer feeds back on its membrane template to maintain an environment conducive to directional synthesis. Our findings are applicable to rod-shaped bacteria that are phylogenetically distant from , indicating that horizontal compartmentalization of precursors may be a general feature of bacillary cell wall biogenesis.
}, issn = {2050-084X}, doi = {10.7554/eLife.60263}, author = {Garc{\'\i}a-Heredia, Alam and Kado, Takehiro and Sein, Caralyn E and Puffal, Julia and Osman, Sarah H and Judd, Julius and Gray, Todd A and Morita, Yasu S and Siegrist, M Sloan} } @article {3060, title = {Microbial corrosion of metals: The corrosion microbiome.}, journal = {Adv Microb Physiol}, volume = {78}, year = {2021}, month = {2021}, pages = {317-390}, abstract = {Microbially catalyzed corrosion of metals is a substantial economic concern. Aerobic microbes primarily enhance Fe oxidation through indirect mechanisms and their impact appears to be limited compared to anaerobic microbes. Several anaerobic mechanisms are known to accelerate Fe oxidation. Microbes can consume H abiotically generated from the oxidation of Fe. Microbial H removal makes continued Fe oxidation more thermodynamically favorable. Extracellular hydrogenases further accelerate Fe oxidation. Organic electron shuttles such as flavins, phenazines, and possibly humic substances may replace H as the electron carrier between Fe and cells. Direct Fe-to-microbe electron transfer is also possible. Which of these anaerobic mechanisms predominates in model pure culture isolates is typically poorly documented because of a lack of functional genetic studies. Microbial mechanisms for Fe oxidation may also apply to some other metals. An ultimate goal of microbial metal corrosion research is to develop molecular tools to diagnose the occurrence, mechanisms, and rates of metal corrosion to guide the implementation of the most effective mitigation strategies. A systems biology approach that includes innovative isolation and characterization methods, as well as functional genomic investigations, will be required in order to identify the diagnostic features to be gleaned from meta-omic analysis of corroding materials. A better understanding of microbial metal corrosion mechanisms is expected to lead to new corrosion mitigation strategies. The understanding of the corrosion microbiome is clearly in its infancy, but interdisciplinary electrochemical, microbiological, and molecular tools are available to make rapid progress in this field.
}, keywords = {Corrosion, Electron Transport, Metals, Microbiota, Oxidation-Reduction}, issn = {2162-5468}, doi = {10.1016/bs.ampbs.2021.01.002}, author = {Lekbach, Yassir and Liu, Tao and Li, Yingchao and Moradi, Masoumeh and Dou, Wenwen and Xu, Dake and Smith, Jessica A and Lovley, Derek R} } @article {3061, title = {Self-sustained green neuromorphic interfaces.}, journal = {Nat Commun}, volume = {12}, year = {2021}, month = {2021 Jun 07}, pages = {3351}, abstract = {Incorporating neuromorphic electronics in bioelectronic interfaces can provide intelligent responsiveness to environments. However, the signal mismatch between the environmental stimuli and driving amplitude in neuromorphic devices has limited the functional versatility and energy sustainability. Here we demonstrate multifunctional, self-sustained neuromorphic interfaces by achieving signal matching at the biological level. The advances rely on the unique properties of microbially produced protein nanowires, which enable both bio-amplitude (e.g., <100 mV) signal processing and energy harvesting from ambient humidity. Integrating protein nanowire-based sensors, energy devices and memristors of bio-amplitude functions yields flexible, self-powered neuromorphic interfaces that can intelligently interpret biologically relevant stimuli for smart responses. These features, coupled with the fact that protein nanowires are a green biomaterial of potential diverse functionalities, take the interfaces a step closer to biological integration.
}, keywords = {Biocompatible Materials, Electronics, Nanotechnology, Nanowires, Neural Networks, Computer, Proteins, Synapses}, issn = {2041-1723}, doi = {10.1038/s41467-021-23744-2}, author = {Fu, Tianda and Liu, Xiaomeng and Fu, Shuai and Woodard, Trevor and Gao, Hongyan and Lovley, Derek R and Yao, Jun} } @article {3064, title = {Solvent-Induced Assembly of Microbial Protein Nanowires into Superstructured Bundles.}, journal = {Biomacromolecules}, volume = {22}, year = {2021}, month = {2021 Mar 08}, pages = {1305-1311}, abstract = {Protein-based electronic biomaterials represent an attractive alternative to traditional metallic and semiconductor materials due to their environmentally benign production and purification. However, major challenges hindering further development of these materials include (1) limitations associated with processing proteins in organic solvents and (2) difficulties in forming higher-order structures or scaffolds with multilength scale control. This paper addresses both challenges, resulting in the formation of one-dimensional bundles composed of electrically conductive protein nanowires harvested from the microbes and . Processing these bionanowires from common organic solvents, such as hexane, cyclohexane, and DMF, enabled the production of multilength scale structures composed of distinctly visible pili. Transmission electron microscopy revealed striking images of bundled protein nanowires up to 10 μm in length and with widths ranging from 50-500 nm (representing assembly of tens to hundreds of nanowires). Conductive atomic force microscopy confirmed the presence of an appreciable nanowire conductivity in their bundled state. These results greatly expand the possibilities for fabricating a diverse array of protein nanowire-based electronic device architectures.
}, keywords = {Electric Conductivity, Electron Transport, Geobacter, Nanowires, Solvents}, issn = {1526-4602}, doi = {10.1021/acs.biomac.0c01790}, author = {Sun, Yun-Lu and Montz, Brian J and Selhorst, Ryan and Tang, Hai-Yan and Zhu, Jiaxin and Nevin, Kelly P and Woodard, Trevor L and Ribbe, Alexander E and Russell, Thomas P and Nonnenmann, Stephen S and Lovley, Derek R and Emrick, Todd} } @article {3062, title = {Stainless steel corrosion via direct iron-to-microbe electron transfer by Geobacter species.}, journal = {ISME J}, volume = {15}, year = {2021}, month = {2021 Oct}, pages = {3084-3093}, abstract = {Microbial corrosion of iron-based materials is a substantial economic problem. A mechanistic understanding is required to develop mitigation strategies, but previous mechanistic studies have been limited to investigations with relatively pure Fe(0), which is not a common structural material. We report here that the mechanism for microbial corrosion of stainless steel, the metal of choice for many actual applications, can be significantly different from that~for Fe(0). Although H is often an intermediary electron carrier between the metal and microbes during Fe(0) corrosion, we found that H is not abiotically produced from stainless steel, making this corrosion mechanism unlikely. Geobacter sulfurreducens and Geobacter metallireducens, electrotrophs that are known to directly accept electrons from other microbes or electrodes, extracted electrons from stainless steel via direct iron-to-microbe electron transfer. Genetic modification to prevent H consumption did not negatively impact on stainless steel corrosion. Corrosion was inhibited when genes for outer-surface cytochromes that are key electrical contacts were deleted. These results indicate that a common model of microbial Fe(0) corrosion~by hydrogenase-positive microbes, in which H serves as an intermediary electron carrier between the metal surface and the microbe, may not apply to the microbial corrosion of stainless steel. However, direct iron-to-microbe electron transfer is a~feasible route for stainless steel corrosion.
}, keywords = {Corrosion, Electrons, Geobacter, Iron, Stainless Steel}, issn = {1751-7370}, doi = {10.1038/s41396-021-00990-2}, author = {Tang, Hai-Yan and Yang, Chuntian and Ueki, Toshiyuki and Pittman, Conor C and Xu, Dake and Woodard, Trevor L and Holmes, Dawn E and Gu, Tingyue and Wang, Fuhui and Lovley, Derek R} } @article {2625, title = {Trehalose Recycling Promotes Energy-Efficient Biosynthesis of the Mycobacterial Cell Envelope.}, journal = {mBio}, volume = {12}, year = {2021}, month = {2021 01 19}, abstract = {The mycomembrane layer of the mycobacterial cell envelope is a barrier to environmental, immune, and antibiotic insults. There is considerable evidence of mycomembrane plasticity during infection and in response to host-mimicking stresses. Since mycobacteria are resource and energy limited under these conditions, it is likely that remodeling has distinct requirements from those of the well-characterized biosynthetic program that operates during unrestricted growth. Unexpectedly, we found that mycomembrane remodeling in nutrient-starved, nonreplicating mycobacteria includes synthesis in addition to turnover. Mycomembrane synthesis under these conditions occurs along the cell periphery, in contrast to the polar assembly of actively growing cells, and both liberates and relies on the nonmammalian disaccharide trehalose. In the absence of trehalose recycling, trehalose synthesis fuels mycomembrane remodeling. However, mycobacteria experience ATP depletion, enhanced respiration, and redox stress, hallmarks of futile cycling and the collateral dysfunction elicited by some bactericidal antibiotics. Inefficient energy metabolism compromises the survival of trehalose recycling mutants in macrophages. Our data suggest that trehalose recycling alleviates the energetic burden of mycomembrane remodeling under stress. Cell envelope recycling pathways are emerging targets for sensitizing resource-limited bacterial pathogens to host and antibiotic pressure. The glucose-based disaccharide trehalose is a stress protectant and carbon source in many nonmammalian cells. Mycobacteria are relatively unique in that they use trehalose for an additional, extracytoplasmic purpose: to build their outer \"myco\" membrane. In these organisms, trehalose connects mycomembrane biosynthesis and turnover to central carbon metabolism. Key to this connection is the retrograde transporter LpqY-SugABC. Unexpectedly, we found that nongrowing mycobacteria synthesize mycomembrane under carbon limitation but do not require LpqY-SugABC. In the absence of trehalose recycling, compensatory anabolism allows mycomembrane biosynthesis to continue. However, this workaround comes at a cost, namely, ATP consumption, increased respiration, and oxidative stress. Strikingly, these phenotypes resemble those elicited by futile cycles and some bactericidal antibiotics. We demonstrate that inefficient energy metabolism attenuates trehalose recycling mutant in macrophages. Energy-expensive macromolecule biosynthesis triggered in the absence of recycling may be a new paradigm for boosting host activity against bacterial pathogens.
}, issn = {2150-7511}, doi = {10.1128/mBio.02801-20}, author = {Pohane, Amol Arunrao and Carr, Caleb R and Garhyan, Jaishree and Swarts, Benjamin M and Siegrist, M Sloan} } @article {3069, title = {Bioinspired bio-voltage memristors.}, journal = {Nat Commun}, volume = {11}, year = {2020}, month = {2020 Apr 20}, pages = {1861}, abstract = {Memristive devices are promising candidates to emulate biological computing. However, the typical switching voltages (0.2-2 V) in previously described devices are much higher than the amplitude in biological counterparts. Here we demonstrate a type of diffusive memristor, fabricated from the protein nanowires harvested from the bacterium Geobacter sulfurreducens, that functions at the biological voltages of 40-100 mV. Memristive function at biological voltages is possible because the protein nanowires catalyze metallization. Artificial neurons built from these memristors not only function at biological action potentials (e.g., 100 mV, 1 ms) but also exhibit temporal integration close to that in biological neurons. The potential of using the memristor to directly process biosensing signals is also demonstrated.
}, keywords = {Action Potentials, Biosensing Techniques, Electricity, Electronics, Equipment Design, Geobacter, Humans, Molecular Dynamics Simulation, Nanotechnology, Nanowires, Neural Networks, Computer, Neurons, Synapses, Wearable Electronic Devices}, issn = {2041-1723}, doi = {10.1038/s41467-020-15759-y}, author = {Fu, Tianda and Liu, Xiaomeng and Gao, Hongyan and Ward, Joy E and Liu, Xiaorong and Yin, Bing and Wang, Zhongrui and Zhuo, Ye and Walker, David J F and Joshua Yang, J and Chen, Jianhan and Lovley, Derek R and Yao, Jun} } @article {3067, title = { Capable of Direct Interspecies Electron Transfer.}, journal = {Environ Sci Technol}, volume = {54}, year = {2020}, month = {2020 Dec 01}, pages = {15347-15354}, abstract = {Direct interspecies electron transfer (DIET) from bacteria to methanogens is a revolutionary concept for syntrophic metabolism in methanogenic soils/sediments and anaerobic digestion. Previous studies have indicated that the potential for DIET is limited to methanogens in the , leading to the assumption that an abundance of other types of methanogens, such as species, indicates a lack of DIET. We report here on a strain of , designated strain YSL, that grows via DIET in defined cocultures with . The cocultures formed aggregates, in which cells of strain YSL and were uniformly dispersed throughout. This close association of the two species is the likely explanation for the ability of a strain of that could not express electrically conductive pili to grow in coculture with strain YSL. Granular activated carbon promoted the initial formation of the DIET-based cocultures. The discovery of DIET in , the genus of methanogens that has been the exemplar for interspecies electron transfer H, suggests that the capacity for DIET is much more broadly distributed among methanogens than previously considered. More innovative approaches to microbial isolation and characterization are needed in order to better understand how methanogenic communities function.
}, keywords = {Electron Transport, Electrons, Geobacter, Methane, Methanobacterium}, issn = {1520-5851}, doi = {10.1021/acs.est.0c05525}, author = {Zheng, Shiling and Liu, Fanghua and Wang, Bingchen and Zhang, Yuechao and Lovley, Derek R} } @article {3070, title = {An Chassis for Production of Electrically Conductive Protein Nanowires.}, journal = {ACS Synth Biol}, volume = {9}, year = {2020}, month = {2020 Mar 20}, pages = {647-654}, abstract = {pilin-based electrically conductive protein nanowires (e-PNs) are a revolutionary electronic material. They offer novel options for electronic sensing applications and have the remarkable ability to harvest electrical energy from atmospheric humidity. However, technical constraints limit mass cultivation and genetic manipulation of . Therefore, we designed a strain of to express e-PNs by introducing a plasmid that contained an inducible operon with genes for type IV pili biogenesis machinery and a synthetic gene designed to yield a peptide monomer that could be assembled into e-PNs. The e-PNs expressed in and harvested with a simple filtration method had the same diameter (3 nm) and conductance as e-PNs expressed in . These results, coupled with the robustness of for mass cultivation and the extensive toolbox for genetic manipulation, greatly expand the opportunities for large-scale fabrication of novel e-PNs.
}, keywords = {Electric Conductivity, Escherichia coli, Fimbriae Proteins, Fimbriae, Bacterial, Geobacter, Graphite, Microorganisms, Genetically-Modified, Microscopy, Atomic Force, Nanowires, Operon, Protein Engineering}, issn = {2161-5063}, doi = {10.1021/acssynbio.9b00506}, author = {Ueki, Toshiyuki and Walker, David J F and Woodard, Trevor L and Nevin, Kelly P and Nonnenmann, Stephen S and Lovley, Derek R} } @article {2622, title = {Chemical Biology Tools for Examining the Bacterial Cell Wall.}, journal = {Cell Chem Biol}, volume = {27}, year = {2020}, month = {2020 08 20}, pages = {1052-1062}, abstract = {Bacteria surround themselves with cell walls to maintain cell rigidity and protect against environmental insults. Here we review chemical and biochemical techniques employed to study bacterial cell wall biogenesis. Recent advances including the ability to isolate critical intermediates, metabolic approaches for probe incorporation, and isotopic labeling techniques have provided critical insight into the biochemistry of cell walls. Fundamental manuscripts that have used these techniques to discover cell wall-interacting proteins, flippases, and cell wall stoichiometry are discussed in detail. The review highlights that these powerful methods and techniques have exciting potential to identify and characterize new targets for antibiotic development.
}, issn = {2451-9448}, doi = {10.1016/j.chembiol.2020.07.024}, author = {Brown, Ashley R and Gordon, Rebecca A and Hyland, Stephen N and Siegrist, M Sloan and Grimes, Catherine L} } @article {2621, title = {Photoactivatable Glycolipid Probes for Identifying Mycolate-Protein Interactions in Live Mycobacteria.}, journal = {J Am Chem Soc}, volume = {142}, year = {2020}, month = {2020 04 29}, pages = {7725-7731}, abstract = {Mycobacteria have a distinctive glycolipid-rich outer membrane, the mycomembrane, which is a critical target for tuberculosis drug development. However, proteins that associate with the mycomembrane, or that are involved in its metabolism and host interactions, are not well-characterized. To facilitate the study of mycomembrane-related proteins, we developed photoactivatable trehalose monomycolate analogues that metabolically incorporate into the mycomembrane in live mycobacteria, enabling photo-cross-linking and click-chemistry-mediated analysis of mycolate-interacting proteins. When deployed in with quantitative proteomics, this strategy enriched over 100 proteins, including the mycomembrane porin (MspA), several proteins with known mycomembrane synthesis or remodeling functions (CmrA, MmpL3, Ag85, Tdmh), and numerous candidate mycolate-interacting proteins. Our approach is highly versatile, as it (i) enlists click chemistry for flexible protein functionalization; (ii) in principle can be applied to any mycobacterial species to identify endogenous bacterial proteins or host proteins that interact with mycolates; and (iii) can potentially be expanded to investigate protein interactions with other mycobacterial lipids. This tool is expected to help elucidate fundamental physiological and pathological processes related to the mycomembrane and may reveal novel diagnostic and therapeutic targets.
}, issn = {1520-5126}, doi = {10.1021/jacs.0c01065}, author = {Kavunja, Herbert W and Biegas, Kyle J and Banahene, Nicholas and Stewart, Jessica A and Piligian, Brent F and Groenevelt, Jessica M and Sein, Caralyn E and Morita, Yasu S and Niederweis, Michael and Siegrist, M Sloan and Swarts, Benjamin M} } @article {3071, title = {Power generation from ambient humidity using protein nanowires.}, journal = {Nature}, volume = {578}, year = {2020}, month = {2020 Feb}, pages = {550-554}, abstract = {Harvesting energy from the environment offers the promise of clean power for self-sustained systems. Known technologies-such as solar cells, thermoelectric devices and mechanical generators-have specific environmental requirements that restrict where they can be deployed and limit their potential for continuous energy production. The ubiquity of atmospheric moisture offers an alternative. However, existing moisture-based energy-harvesting technologies can produce only intermittent, brief (shorter than 50 seconds) bursts of power in the ambient environment, owing to the lack of a sustained conversion mechanism. Here we show that thin-film devices made from nanometre-scale protein wires harvested from the microbe Geobacter sulfurreducens can generate continuous electric power in the ambient environment. The devices produce a sustained voltage of around 0.5~volts across a 7-micrometre-thick film, with a current density of around 17 microamperes per square centimetre. We find the driving force behind this energy generation to be a self-maintained moisture gradient that forms within the film when the film is exposed to the humidity that is naturally present in air. Connecting several devices linearly scales up the voltage and current to power electronics. Our results demonstrate the feasibility of a continuous energy-harvesting strategy that is less restricted by location or environmental conditions than other sustainable approaches.
}, issn = {1476-4687}, doi = {10.1038/s41586-020-2010-9}, author = {Liu, Xiaomeng and Gao, Hongyan and Ward, Joy E and Liu, Xiaorong and Yin, Bing and Fu, Tianda and Chen, Jianhan and Lovley, Derek R and Yao, Jun} } @article {3068, title = {Protein Nanowires: the Electrification of the Microbial World and Maybe Our Own.}, journal = {J Bacteriol}, volume = {202}, year = {2020}, month = {2020 Sep 23}, abstract = {Electrically conductive protein nanowires appear to be widespread in the microbial world and are a revolutionary "green" material for the fabrication of electronic devices. Electrically conductive pili (e-pili) assembled from type IV pilin monomers have independently evolved multiple times in microbial history as have electrically conductive archaella (e-archaella) assembled from homologous archaellin monomers. A role for e-pili in long-range (micrometer) extracellular electron transport has been demonstrated in some microbes. The surprising finding of e-pili in syntrophic bacteria and the role of e-pili as conduits for direct interspecies electron transfer have necessitated a reassessment of routes for electron flux in important methanogenic environments, such as anaerobic digesters and terrestrial wetlands. Pilin monomers similar to those found in e-pili may also be a major building block of the conductive "cables" that transport electrons over centimeter distances through continuous filaments of cable bacteria consisting of a thousand cells or more. Protein nanowires harvested from microbes have many functional and sustainability advantages over traditional nanowire materials and have already yielded novel electronic devices for sustainable electricity production, neuromorphic memory, and sensing. e-pili can be mass produced with an chassis, providing a ready source of material for electronics as well as for studies on the basic mechanisms for long-range electron transport along protein nanowires. Continued exploration is required to better understand the electrification of microbial communities with microbial nanowires and to expand the "green toolbox" of sustainable materials for wiring and powering the emerging "Internet of things."
}, keywords = {Electric Conductivity, Electron Transport, Fimbriae Proteins, Fimbriae, Bacterial, Geobacter, Microscopy, Electron, Nanowires, Protein Engineering}, issn = {1098-5530}, doi = {10.1128/JB.00331-20}, author = {Lovley, Derek R and Holmes, Dawn E} } @article {3066, title = {Sparking Anaerobic Digestion: Promoting Direct Interspecies Electron Transfer to Enhance Methane Production.}, journal = {iScience}, volume = {23}, year = {2020}, month = {2020 Dec 18}, pages = {101794}, abstract = {Anaerobic digestion was one of the first bioenergy strategies developed, yet the interactions of the microbial community that is responsible for the production of methane are still poorly understood. For example, it has only recently been recognized that the bacteria that oxidize organic waste components can forge electrical connections with methane-producing microbes through biologically produced, protein-based, conductive circuits. This direct interspecies electron transfer (DIET) is faster than interspecies electron exchange via diffusive electron carriers, such as H. DIET is also more resilient to perturbations such as increases in organic load inputs or toxic compounds. However, with current digester practices DIET rarely predominates. Improvements in anaerobic digestion associated with the addition of electrically conductive materials have been attributed to increased DIET, but experimental verification has been lacking. This deficiency may soon be overcome with improved understanding of the diversity of microbes capable of DIET, which is leading to molecular tools for determining the extent of DIET. Here we review the microbiology of DIET, suggest molecular strategies for monitoring DIET in anaerobic digesters, and propose approaches for re-engineering digester design and practices to encourage DIET.
}, issn = {2589-0042}, doi = {10.1016/j.isci.2020.101794}, author = {Zhao, Zhiqiang and Li, Yang and Zhang, Yaobin and Lovley, Derek R} } @article {2623, title = {Supramolecular antibiotics: a strategy for conversion of broad-spectrum to narrow-spectrum antibiotics for .}, journal = {Nanoscale}, volume = {12}, year = {2020}, month = {2020 Oct 22}, pages = {20693-20698}, abstract = {The propensity of broad-spectrum antibiotics to indiscriminately kill both pathogenic and beneficial bacteria has a profound impact on the spread of resistance across multiple bacterial species. Alternative approaches that narrow antibacterial specificity towards desired pathogenic bacterial population are of great interest. Here, we report an enzyme-responsive antibiotic-loaded nanoassembly strategy for narrow delivery of otherwise broad-spectrum antibiotics. We specifically target Staphylococcus aureus (S. aureus), an important blood pathogen that secretes PC1 \β-lactamases. Our nanoassemblies selectively eradicate S. aureus grown in vitro with other bacteria, highlighting its potential capability in targeting the desired pathogenic bacterial population.
}, issn = {2040-3372}, doi = {10.1039/d0nr04886k}, author = {Koyasseril-Yehiya, Thameez M and Garc{\'\i}a-Heredia, Alam and Anson, Francesca and Rangadurai, Poornima and Siegrist, M Sloan and Thayumanavan, S} } @article {3072, title = {Syntrophus conductive pili demonstrate that common hydrogen-donating syntrophs can have a direct electron transfer option.}, journal = {ISME J}, volume = {14}, year = {2020}, month = {2020 Mar}, pages = {837-846}, abstract = {Syntrophic interspecies electron exchange is essential for the stable functioning of diverse anaerobic microbial communities. Hydrogen/formate interspecies electron transfer (HFIT), in which H and/or formate function as diffusible electron carriers, has been considered to be the primary mechanism for electron transfer because most common syntrophs were thought to lack biochemical components, such as electrically conductive pili (e-pili), necessary for direct interspecies electron transfer (DIET). Here we report that Syntrophus aciditrophicus, one of the most intensively studied microbial models for HFIT, produces e-pili and can grow via DIET. Heterologous expression of the putative S. aciditrophicus type IV pilin gene in Geobacter sulfurreducens yielded conductive pili of the same diameter (4 nm) and conductance of the native S. aciditrophicus pili and enabled long-range electron transport in G. sulfurreducens. S. aciditrophicus lacked abundant c-type cytochromes often associated with DIET. Pilin genes likely to yield e-pili were found in other genera of hydrogen/formate-producing syntrophs. The finding that DIET is a likely option for diverse syntrophs that are abundant in many anaerobic environments necessitates a reexamination of the paradigm that HFIT is the predominant mechanism for syntrophic electron exchange within anaerobic microbial communities of biogeochemical and practical significance.
}, keywords = {Deltaproteobacteria, Electric Conductivity, Electron Transport, Electrons, Fimbriae Proteins, Fimbriae, Bacterial, Formates, Geobacter, Hydrogen}, issn = {1751-7370}, doi = {10.1038/s41396-019-0575-9}, author = {Walker, David J F and Nevin, Kelly P and Holmes, Dawn E and Rotaru, Amelia-Elena and Ward, Joy E and Woodard, Trevor L and Zhu, Jiaxin and Ueki, Toshiyuki and Nonnenmann, Stephen S and McInerney, Michael J and Lovley, Derek R} } @article {3079, title = {The Archaellum of Methanospirillum hungatei Is Electrically Conductive.}, journal = {mBio}, volume = {10}, year = {2019}, month = {2019 Apr 16}, abstract = {Microbially produced electrically conductive protein filaments are of interest because they can function as conduits for long-range biological electron transfer. They also show promise as sustainably produced electronic materials. Until now, microbially produced conductive protein filaments have been reported only for bacteria. We report here that the archaellum of is electrically conductive. This is the first demonstration that electrically conductive protein filaments have evolved in Furthermore, the structure of the archaellum was previously determined (N. Poweleit, P. Ge, H. N. Nguyen, R. R. O. Loo, et al., Nat Microbiol 2:16222, 2016, https://doi.org/10.1038/nmicrobiol.2016.222). Thus, the archaellum of is the first microbially produced electrically conductive protein filament for which a structure is known. We analyzed the previously published structure and identified a core of tightly packed phenylalanines that is one likely route for electron conductance. The availability of the archaellum structure is expected to substantially advance mechanistic evaluation of long-range electron transport in microbially produced electrically conductive filaments and to aid in the design of "green" electronic materials that can be microbially produced with renewable feedstocks. Microbially produced electrically conductive protein filaments are a revolutionary, sustainably produced, electronic material with broad potential applications. The design of new protein nanowires based on the known archaellum structure could be a major advance over the current empirical design of synthetic protein nanowires from electrically conductive bacterial pili. An understanding of the diversity of outer-surface protein structures capable of electron transfer is important for developing models for microbial electrical communication with other cells and minerals in natural anaerobic environments. Extracellular electron exchange is also essential in engineered environments such as bioelectrochemical devices and anaerobic digesters converting wastes to methane. The finding that the archaellum of is electrically conductive suggests that some archaea might be able to make long-range electrical connections with their external environment.
}, keywords = {Electric Conductivity, Electricity, Electron Transport, Flagella, Methanospirillum, Phenylalanine}, issn = {2150-7511}, doi = {10.1128/mBio.00579-19}, author = {Walker, David J F and Martz, Eric and Holmes, Dawn E and Zhou, Zimu and Nonnenmann, Stephen S and Lovley, Derek R} } @article {3076, title = {Cryo-EM reveals the structural basis of long-range electron transport in a cytochrome-based bacterial nanowire.}, journal = {Commun Biol}, volume = {2}, year = {2019}, month = {2019}, pages = {219}, abstract = {Electrically conductive pili from species, termed bacterial nanowires, are intensely studied for their biological significance and potential in the development of new materials. Using cryo-electron microscopy, we have characterized nanowires from conductive pili preparations that are composed solely of head-to-tail stacked monomers of the six-heme C-type cytochrome OmcS. The unique fold of OmcS - closely wrapped around a continuous stack of hemes that can serve as an uninterrupted path for electron transport - generates a scaffold that supports the unbranched chain of hemes along the central axis of the filament. We present here, at 3.4 {\r A} resolution, the structure of this cytochrome-based filament and discuss its possible role in long-range biological electron transport.
}, keywords = {Cryoelectron Microscopy, Cytochromes c, Electron Transport, Fimbriae, Bacterial, Geobacter, Nanowires}, issn = {2399-3642}, doi = {10.1038/s42003-019-0448-9}, author = {Filman, David J and Marino, Stephen F and Ward, Joy E and Yang, Lu and Mester, Zolt{\'a}n and Bullitt, Esther and Lovley, Derek R and Strauss, Mike} } @article {3075, title = {Decorating the Outer Surface of Microbially Produced Protein Nanowires with Peptides.}, journal = {ACS Synth Biol}, volume = {8}, year = {2019}, month = {2019 Aug 16}, pages = {1809-1817}, abstract = {The potential applications of electrically conductive protein nanowires (e-PNs) harvested from might be greatly expanded if the outer surface of the wires could be modified to confer novel sensing capabilities or to enhance binding to other materials. We developed a simple strategy for functionalizing e-PNs with surface-exposed peptides. The gene for the monomer that assembles into e-PNs was modified to add peptide tags at the carboxyl terminus of the monomer. Strains of were constructed that fabricated synthetic e-PNs with a six-histidine "His-tag" or both the His-tag and a nine-peptide "HA-tag" exposed on the outer surface. Addition of the peptide tags did not diminish e-PN conductivity. The abundance of HA-tag in e-PNs was controlled by placing expression of the gene for the synthetic monomer with the HA-tag under transcriptional regulation. These studies suggest broad possibilities for tailoring e-PN properties for diverse applications.
}, keywords = {Carboxy-Lyases, Ethylene Glycols, Molecular Structure, Nanowires, Oxygenases, Peptides, Phenylalanine Ammonia-Lyase, Plasmids, Proteins, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins, Styrenes}, issn = {2161-5063}, doi = {10.1021/acssynbio.9b00131}, author = {Ueki, Toshiyuki and Walker, David J F and Tremblay, Pier-Luc and Nevin, Kelly P and Ward, Joy E and Woodard, Trevor L and Nonnenmann, Stephen S and Lovley, Derek R} } @article {2620, title = {Engineering the Mycomembrane of Live Mycobacteria with an Expanded Set of Trehalose Monomycolate Analogues.}, journal = {Chembiochem}, volume = {20}, year = {2019}, month = {2019 05 15}, pages = {1282-1291}, abstract = {Mycobacteria and related organisms in the Corynebacterineae suborder are characterized by a distinctive outer membrane referred to as the mycomembrane. Biosynthesis of the mycomembrane occurs through an essential process called mycoloylation, which involves antigen 85 (Ag85)-catalyzed transfer of mycolic acids from the mycoloyl donor trehalose monomycolate (TMM) to acceptor carbohydrates and, in some organisms, proteins. We recently described an alkyne-modified TMM analogue (O-AlkTMM-C7) which, in conjunction with click chemistry, acted as a chemical reporter for mycoloylation in intact cells and allowed metabolic labeling of mycoloylated components of the mycomembrane. Here, we describe the synthesis and evaluation of a toolbox of TMM-based reporters bearing alkyne, azide, trans-cyclooctene, and fluorescent tags. These compounds gave further insight into the substrate tolerance of mycoloyltransferases (e.g., Ag85s) in a cellular context and they provide significantly expanded experimental versatility by allowing one- or two-step cell labeling, live cell labeling, and rapid cell labeling via tetrazine ligation. Such capabilities will facilitate research on mycomembrane composition, biosynthesis, and dynamics. Moreover, because TMM is exclusively metabolized by Corynebacterineae, the described probes may be valuable for the specific detection and cell-surface engineering of Mycobacterium tuberculosis and related pathogens. We also performed experiments to establish the dependence of probe incorporation on mycoloyltransferase activity, results from which suggested that cellular labeling is a function not only of metabolic incorporation (and likely removal) pathway(s), but also accessibility across the envelope. Thus, whole-cell labeling experiments with TMM reporters should be carefully designed and interpreted when envelope permeability may be compromised. On the other hand, this property of TMM reporters can potentially be exploited as a convenient way to probe changes in envelope integrity and permeability, facilitating drug development studies.
}, keywords = {Acyltransferases, Alkynes, Azides, Bacillus subtilis, Cell Engineering, Cell Membrane, Click Chemistry, Cord Factors, Corynebacterium, Escherichia coli, Fluorescent Dyes, Molecular Structure, Mycobacterium smegmatis, Mycobacterium tuberculosis}, issn = {1439-7633}, doi = {10.1002/cbic.201800687}, author = {Fiolek, Taylor J and Banahene, Nicholas and Kavunja, Herbert W and Holmes, Nathan J and Rylski, Adrian K and Pohane, Amol Arunrao and Siegrist, M Sloan and Swarts, Benjamin M} } @article {3077, title = {Iron Corrosion via Direct Metal-Microbe Electron Transfer.}, journal = {mBio}, volume = {10}, year = {2019}, month = {2019 May 14}, abstract = {The concept that anaerobic microorganisms can directly accept electrons from Fe(0) has been controversial because direct metal-microbe electron transfer has previously only been indirectly inferred. Fe(0) oxidation was studied with strain ACL, an autotrophic strain that was previously shown to grow with electrons derived from a graphite cathode as the sole electron donor. Strain ACL grew with Fe(0) as the sole electron donor and fumarate as the electron acceptor. However, it appeared that at least a portion of the electron transfer was via H produced nonenzymatically from the oxidation of Fe(0) to Fe(II). H, which accumulated in abiotic controls, was consumed during the growth of strain ACL, the cells were predominately planktonic, and genes for the uptake hydrogenase were highly expressed. Strain ACL was constructed to prevent growth on H or formate by deleting the genes for the uptake of hydrogenase and formate dehydrogenases from strain ACL. Strain ACL also grew with Fe(0) as the sole electron donor, but H accumulated in the culture, and cells heavily colonized Fe(0) surfaces with no visible planktonic growth. Transcriptomics suggested that the outer surface -type cytochromes OmcS and OmcZ were important during growth of strain ACL on Fe(0). Strain ACL did not grow on Fe(0) if the gene for either of these cytochromes was deleted. The specific attachment of strain ACL to Fe(0), coupled with requirements for known extracellular electrical contacts, suggest that direct metal-microbe electron transfer is the most likely option for Fe(0) serving as an electron donor. The anaerobic corrosion of iron structures is expensive to repair and can be a safety and environmental concern. It has been known for over 100 years that the presence of anaerobic respiratory microorganisms can accelerate iron corrosion. Multiple studies have suggested that there are sulfate reducers, methanogens, and acetogens that can directly accept electrons from Fe(0) to support sulfate or carbon dioxide reduction. However, all of the strains studied can also use H as an electron donor for growth, which is known to be abiotically produced from Fe(0). Furthermore, no proteins definitely shown to function as extracellular electrical contacts with Fe(0) were identified. The studies described here demonstrate that direct electron transfer from Fe(0) can support anaerobic respiration. They also map out a simple genetic approach to the study of iron corrosion mechanisms in other microorganisms. A better understanding of how microorganisms promote iron corrosion is expected to lead to the development of strategies that can help reduce adverse impacts from this process.
}, keywords = {Anaerobiosis, Corrosion, Cytochromes, Electron Transport, Formate Dehydrogenases, Geobacter, Iron, Oxidation-Reduction, Oxidoreductases, Transcriptome}, issn = {2150-7511}, doi = {10.1128/mBio.00303-19}, author = {Tang, Hai-Yan and Holmes, Dawn E and Ueki, Toshiyuki and Palacios, Paola A and Lovley, Derek R} } @article {3074, title = {A Membrane-Bound Cytochrome Enables To Conserve Energy from Extracellular Electron Transfer.}, journal = {mBio}, volume = {10}, year = {2019}, month = {2019 Aug 20}, abstract = {Extracellular electron exchange in species and closely related plays an important role in the global carbon cycle and enhances the speed and stability of anaerobic digestion by facilitating efficient syntrophic interactions. Here, we grew with methanol provided as the electron donor and the humic analogue, anthraquione-2,6-disulfonate (AQDS), provided as the electron acceptor when methane production was inhibited with bromoethanesulfonate. AQDS was reduced with simultaneous methane production in the absence of bromoethanesulfonate. Transcriptomics revealed that expression of the gene for the transmembrane, multiheme, -type cytochrome MmcA was higher in AQDS-respiring cells than in cells performing methylotrophic methanogenesis. A strain in which the gene for MmcA was deleted failed to grow via AQDS reduction but grew with the conversion of methanol or acetate to methane, suggesting that MmcA has a specialized role as a conduit for extracellular electron transfer. Enhanced expression of genes for methanol conversion to methyl-coenzyme M and the Rnf complex suggested that methanol is oxidized to carbon dioxide in AQDS-respiring cells through a pathway that is similar to methyl-coenzyme M oxidation in methanogenic cells. However, during AQDS respiration the Rnf complex and reduced methanophenazine probably transfer electrons to MmcA, which functions as the terminal reductase for AQDS reduction. Extracellular electron transfer may enable the survival of methanogens in dynamic environments in which oxidized humic substances and Fe(III) oxides are intermittently available. The availability of tools for genetic manipulation of makes it an excellent model microbe for evaluating -type cytochrome-dependent extracellular electron transfer in The discovery of a methanogen that can conserve energy to support growth solely from the oxidation of organic carbon coupled to the reduction of an extracellular electron acceptor expands the possible environments in which methanogens might thrive. The potential importance of -type cytochromes for extracellular electron transfer to syntrophic bacterial partners and/or Fe(III) minerals in some was previously proposed, but these studies with provide the first genetic evidence for cytochrome-based extracellular electron transfer in The results suggest parallels with Gram-negative bacteria, such as and species, in which multiheme outer-surface -type cytochromes are an essential component for electrical communication with the extracellular environment. offers an unprecedented opportunity to study mechanisms for energy conservation from the anaerobic oxidation of one-carbon organic compounds coupled to extracellular electron transfer in with implications not only for methanogens but possibly also for that anaerobically oxidize methane.
}, keywords = {Acetates, Anthraquinones, Cytochromes, Electron Transport, Electrons, Ferric Compounds, Gene Expression Regulation, Archaeal, Gram-Negative Bacteria, Membranes, Mesna, Methane, Methanol, Methanosarcina, Oxidation-Reduction, Oxidoreductases, Transcriptome}, issn = {2150-7511}, doi = {10.1128/mBio.00789-19}, author = {Holmes, Dawn E and Ueki, Toshiyuki and Tang, Hai-Yan and Zhou, Jinjie and Smith, Jessica A and Chaput, Gina and Lovley, Derek R} } @article {3078, title = {A pilin chaperone required for the expression of electrically conductive Geobacter sulfurreducens pili.}, journal = {Environ Microbiol}, volume = {21}, year = {2019}, month = {2019 Jul}, pages = {2511-2522}, abstract = {Mechanisms controlling the expression of the electrically conductive pili (e-pili) of Geobacter species are of interest because of the important role of e-pili in diverse biogeochemical processes, anaerobic digestion and electromicrobiological applications. We investigated the function of the protein, designated Spc (short pilin chaperone), encoded by the gene immediately downstream from the gene for PilA, the monomer that assembles into e-pili. Multiple lines of evidence suggest that Spc forms an oligomer that is associated with the inner membrane. Mutating the start codon of spc to prevent translation increased the transcript abundance of pilA but greatly diminished the abundance of PilA, and e-pili could no longer be detected. Cross-linking, protein capture and two-hybrid studies demonstrated that Spc and PilA interacted. Two sites in PilA for electrostatic interaction with Spc were identified. The results demonstrate that Spc is required for PilA stability prior to incorporation into e-pili, suggesting that Spc has a chaperone function that may be specific to the relatively short PilA monomers that assemble into e-pili. These results are important for identifying microorganisms likely to express e-pili from (meta)genomic data and for the construction of microbial strains expressing e-pili.
}, keywords = {Electric Conductivity, Fimbriae Proteins, Fimbriae, Bacterial, Geobacter, Molecular Chaperones}, issn = {1462-2920}, doi = {10.1111/1462-2920.14638}, author = {Liu, Xing and Zhan, Ji and Jing, Xianyue and Zhou, Shungui and Lovley, Derek R} } @article {3073, title = { Protein Nanowires.}, journal = {Front Microbiol}, volume = {10}, year = {2019}, month = {2019}, pages = {2078}, abstract = {The study of electrically conductive protein nanowires in has led to new concepts for long-range extracellular electron transport, as well as for the development of sustainable conductive materials and electronic devices with novel functions. Until recently, electrically conductive pili (e-pili), assembled from the PilA pilin monomer, were the only known protein nanowires. However, filaments comprised of the multi-heme -type cytochrome, OmcS, are present in some preparations of outer-surface proteins. The purpose of this review is to evaluate the available evidence on the expression of e-pili and OmcS filaments and their biological function. Abundant literature demonstrates that expresses e-pili, which are required for long-range electron transport to Fe (III) oxides and through conductive biofilms. In contrast, there is no definitive evidence yet that wild-type express long filaments of OmcS extending from the cells, and deleting the gene for OmcS actually biofilm conductivity. The literature does not support the concern that many previous studies on e-pili were mistakenly studying OmcS filaments. For example, heterologous expression of the aromatic-rich pilin monomer of in increases the conductivity of individual nanowires more than 5,000-fold, whereas expression of an aromatic-poor pilin reduced conductivity more than 1,000-fold. This more than million-fold range in nanowire conductivity was achieved while maintaining the 3-nm diameter characteristic of e-pili. Purification methods that eliminate all traces of OmcS yield highly conductive e-pili, as does heterologous expression of the e-pilin monomer in microbes that do not produce OmcS or any other outer-surface cytochromes. Future studies of expression of protein nanowires need to be cognizant of the importance of maintaining environmentally relevant growth conditions because artificial laboratory culture conditions can rapidly select against e-pili expression. Principles derived from the study of e-pili have enabled identification of non-cytochrome protein nanowires in diverse bacteria and archaea. A similar search for cytochrome appendages is warranted. Both e-pili and OmcS filaments offer design options for the synthesis of protein-based "green" electronics, which may be the primary driving force for the study of these structures in the near future.
}, issn = {1664-302X}, doi = {10.3389/fmicb.2019.02078}, author = {Lovley, Derek R and Walker, David J F} } @article {2372, title = {Cell cycle localization dynamics of mitochondrial DNA polymerase IC in African trypanosomes.}, journal = {Mol Biol Cell}, volume = {29}, year = {2018}, month = {10/2018}, pages = {2540-2552}, abstract = {Trypanosoma brucei has a unique catenated mitochondrial DNA (mtDNA) network called kinetoplast DNA (kDNA). Replication of kDNA occurs once per cell cycle in near synchrony with nuclear S phase and requires the coordination of many proteins. Among these are three essential DNA polymerases (TbPOLIB, IC, and ID). Localization dynamics of these proteins with respect to kDNA replication stages and how they coordinate their functions during replication are not well understood. We previously demonstrated that TbPOLID undergoes dynamic localization changes that are coupled to kDNA replication events. Here, we report the localization of TbPOLIC, a second essential DNA polymerase, and demonstrate the accumulation of TbPOLIC foci at active kDNA replication sites (antipodal sites) during stage II of the kDNA duplication cycle. While TbPOLIC was undetectable by immunofluorescence during other cell cycle stages, steady-state protein levels measured by Western blot remained constant. TbPOLIC foci colocalized with the fraction of TbPOLID that localized to the antipodal sites. However, the partial colocalization of the two essential DNA polymerases suggests a highly dynamic environment at the antipodal sites to coordinate the trafficking of replication proteins during kDNA synthesis. These data indicate that cell cycle-dependent localization is a major regulatory mechanism for essential mtDNA polymerases during kDNA replication.
}, keywords = {Cell Cycle, DNA Replication, DNA, Kinetoplast, DNA-Directed DNA Polymerase, Gene Knockdown Techniques, Gene Silencing, Mitochondria, Protozoan Proteins, S Phase, Trypanosoma brucei brucei}, issn = {1939-4586}, doi = {10.1091/mbc.E18-02-0127}, author = {Concepci{\'o}n-Acevedo, Jeniffer and Miller, Jonathan C and Boucher, Michael J and Klingbeil, Michele M} } @article {3081, title = {Conductive Composite Materials Fabricated from Microbially Produced Protein Nanowires.}, journal = {Small}, volume = {14}, year = {2018}, month = {2018 Nov}, pages = {e1802624}, abstract = {Protein-based electronic materials have numerous potential advantages with respect to sustainability and biocompatibility over electronic materials that are synthesized using harsh chemical processes and/or which contain toxic components. The microorganism Geobacter sulfurreducens synthesizes electrically conductive protein nanowires (e-PNs) with high aspect ratios (3 nm {\texttimes} 10-30 {\textmu}m) from renewable organic feedstocks. Here, the integration of G. Sulfurreducens e-PNs into poly(vinyl alcohol) (PVA) as a host polymer matrix is described. The resultant e-PN/PVA composites exhibit conductivities comparable to PVA-based composites containing synthetic nanowires. The relationship between e-PN density and conductivity of the resultant composites is consistent with percolation theory. These e-PNs confer conductivity to the composites even under extreme conditions, with the highest conductivities achieved from materials prepared at pH 1.5 and temperatures greater than 100 {\textdegree}C. These results demonstrate that e-PNs represent viable and sustainable nanowire compositions for the fabrication of electrically conductive composite materials.
}, keywords = {Geobacter, Nanocomposites, Nanowires, Polymers}, issn = {1613-6829}, doi = {10.1002/smll.201802624}, author = {Sun, Yun-Lu and Tang, Hai-Yan and Ribbe, Alexander and Duzhko, Volodimyr and Woodard, Trevor L and Ward, Joy E and Bai, Ying and Nevin, Kelly P and Nonnenmann, Stephen S and Russell, Thomas and Emrick, Todd and Lovley, Derek R} } @article {3083, title = {Construction of a Strain With Exceptional Growth on Cathodes.}, journal = {Front Microbiol}, volume = {9}, year = {2018}, month = {2018}, pages = {1512}, abstract = {Insoluble extracellular electron donors are important sources of energy for anaerobic respiration in biogeochemical cycling and in diverse practical applications. The previous lack of a genetically tractable model microorganism that could be grown to high densities under anaerobic conditions in pure culture with an insoluble extracellular electron donor has stymied efforts to better understand this form of respiration. We report here on the design of a strain of , designated strain ACL, which grows as thick (ca. 35 μm) confluent biofilms on graphite cathodes poised at -500 mV ( Ag/AgCl) with fumarate as the electron acceptor. Sustained maximum current consumption rates were >0.8 A/m, which is >10-fold higher than the current consumption of the wild-type strain. The improved function on the cathode was achieved by introducing genes for an ATP-dependent citrate lyase, completing the complement of enzymes needed for a reverse TCA cycle for the synthesis of biosynthetic precursors from carbon dioxide. Strain ACL provides an important model organism for elucidating the mechanisms for effective anaerobic growth with an insoluble extracellular electron donor and may offer unique possibilities as a chassis for the introduction of synthetic metabolic pathways for the production of commodities with electrons derived from electrodes.
}, issn = {1664-302X}, doi = {10.3389/fmicb.2018.01512}, author = {Ueki, Toshiyuki and Nevin, Kelly P and Woodard, Trevor L and Aklujkar, Muktak A and Holmes, Dawn E and Lovley, Derek R} } @article {2219, title = {DivIVA concentrates mycobacterial cell envelope assembly for initiation and stabilization of polar growth.}, journal = {Cytoskeleton}, volume = {75}, year = {2018}, month = {11/2018}, chapter = {498}, doi = {10.1002/cm.21490}, author = {Melzer, ES and Sein, CE and Siegrist, MS and Chambers, JJ} } @article {3087, title = {Electrically conductive pili from pilin genes of phylogenetically diverse microorganisms.}, journal = {ISME J}, volume = {12}, year = {2018}, month = {2018 Jan}, pages = {48-58}, abstract = {The possibility that bacteria other than Geobacter species might contain genes for electrically conductive pili (e-pili) was investigated by heterologously expressing pilin genes of interest in Geobacter sulfurreducens. Strains of G. sulfurreducens producing high current densities, which are only possible with e-pili, were obtained with pilin genes from Flexistipes sinusarabici, Calditerrivibrio nitroreducens and Desulfurivibrio alkaliphilus. The conductance of pili from these strains was comparable to native G. sulfurreducens e-pili. The e-pili derived from C. nitroreducens, and D. alkaliphilus pilin genes are the first examples of relatively long (>100 amino acids) pilin monomers assembling into e-pili. The pilin gene from Candidatus Desulfofervidus auxilii did not yield e-pili, suggesting that the hypothesis that this sulfate reducer wires itself with e-pili to methane-oxidizing archaea to enable anaerobic methane oxidation should be reevaluated. A high density of aromatic amino acids and a lack of substantial aromatic-free gaps along the length of long pilins may be important characteristics leading to e-pili. This study demonstrates a simple method to screen pilin genes from difficult-to-culture microorganisms for their potential to yield e-pili; reveals new sources for biologically based electronic materials; and suggests that a wide phylogenetic diversity of microorganisms may use e-pili for extracellular electron exchange.
}, keywords = {Deltaproteobacteria, Electric Conductivity, Fimbriae Proteins, Fimbriae, Bacterial, Methane, Oxidation-Reduction, Phylogeny}, issn = {1751-7370}, doi = {10.1038/ismej.2017.141}, author = {Walker, David Jf and Adhikari, Ramesh Y and Holmes, Dawn E and Ward, Joy E and Woodard, Trevor L and Nevin, Kelly P and Lovley, Derek R} } @article {3080, title = {Electron and Proton Flux for Carbon Dioxide Reduction in During Direct Interspecies Electron Transfer.}, journal = {Front Microbiol}, volume = {9}, year = {2018}, month = {2018}, pages = {3109}, abstract = {Direct interspecies electron transfer (DIET) is important in diverse methanogenic environments, but how methanogens participate in DIET is poorly understood. Therefore, the transcriptome of grown via DIET in co-culture with was compared with its transcriptome when grown via H interspecies transfer (HIT) with . Notably, transcripts for the FH dehydrogenase, Fpo, and the heterodisulfide reductase, HdrABC, were more abundant during growth on DIET. A model for CO reduction was developed from these results in which electrons delivered to methanophenazine in the cell membrane are transferred to Fpo. The external proton gradient necessary to drive the otherwise thermodynamically unfavorable reverse electron transport for Fpo-catalyzed F reduction is derived from protons released from metabolism. Reduced F is a direct electron donor in the carbon dioxide reduction pathway and also serves as the electron donor for the proposed HdrABC-catalyzed electron bifurcation reaction in which reduced ferredoxin (also required for carbon dioxide reduction) is generated with simultaneous reduction of CoM-S-S-CoB. Expression of genes for putative redox-active proteins predicted to be localized on the outer cell surface was higher during growth on DIET, but further analysis will be required to identify the electron transfer route to methanophenazine. The results indicate that the pathways for electron and proton flux for CO reduction during DIET are substantially different than for HIT and suggest that gene expression patterns may also be useful for determining whether are directly accepting electrons from other extracellular electron donors, such as corroding metals or electrodes.
}, issn = {1664-302X}, doi = {10.3389/fmicb.2018.03109}, author = {Holmes, Dawn E and Rotaru, Amelia-Elena and Ueki, Toshiyuki and Shrestha, Pravin M and Ferry, James G and Lovley, Derek R} } @article {3082, title = {The Hydrogen Economy of Methanosarcina barkeri: Life in the Fast Lane.}, journal = {J Bacteriol}, volume = {200}, year = {2018}, month = {2018 Oct 15}, abstract = {Two recent studies (T. D. Mand, G. Kulkarni, and W. W. Metcalf, J. Bacteriol 200:e00342-18, 2018, https://doi.org/10.1128/JB.00342-18, and G. Kulkarni, T. D. Mand, and W. W. Metcalf, mBio 9:e01256-18, 2018, https://doi.org/10.1128/mBio.01256-18) analyzed an impressive array of hydrogenase-deficient mutant strains of not only to describe H-based growth but also to demonstrate the conservation of energy with intracellular hydrogen cycling, a novel strategy for creating a proton motive force to support ATP synthesis.
}, keywords = {Hydrogen, Hydrogenase, Methane, Methanosarcina barkeri, Proton-Motive Force}, issn = {1098-5530}, doi = {10.1128/JB.00445-18}, author = {Lovley, Derek R} } @article {2216, title = {Peptidoglycan precursor synthesis along the sidewall of pole-growing mycobacteria}, journal = {{eLife}}, volume = {7}, year = {2018}, month = {sep}, doi = {10.7554/elife.37243}, url = {https://doi.org/10.7554/elife.37243}, author = {Alam Garc{\'\i}a-Heredia and Amol Arunrao Pohane and Emily S Melzer and Caleb R Carr and Taylor J Fiolek and Sarah R Rundell and Hoong Chuin Lim and Jeffrey C Wagner and Yasu S Morita and Benjamin M Swarts and M Sloan Siegrist} } @article {3085, title = {Potential for Methanosarcina to Contribute to Uranium Reduction during Acetate-Promoted Groundwater Bioremediation.}, journal = {Microb Ecol}, volume = {76}, year = {2018}, month = {2018 Oct}, pages = {660-667}, abstract = {Previous studies of acetate-promoted bioremediation of uranium-contaminated aquifers focused on Geobacter because no other microorganisms that can couple the oxidation of acetate with U(VI) reduction had been detected in situ. Monitoring the levels of methyl CoM reductase subunit A (mcrA) transcripts during an acetate-injection field experiment demonstrated that acetoclastic methanogens from the genus Methanosarcina were enriched after 40~days of acetate amendment. The increased abundance of Methanosarcina corresponded with an accumulation of methane in the groundwater. In order to determine whether Methanosarcina species could be participating in U(VI) reduction in the subsurface, cell suspensions of Methanosarcina barkeri were incubated in the presence of U(VI) with acetate provided as the electron donor. U(VI) was reduced by metabolically active M. barkeri cells; however, no U(VI) reduction was observed in inactive controls. These results demonstrate that Methanosarcina species could play an important role in the long-term bioremediation of uranium-contaminated aquifers after depletion of Fe(III) oxides limits the growth of Geobacter species. The results also suggest that Methanosarcina have the potential to influence uranium geochemistry in a diversity of anaerobic sedimentary environments.
}, keywords = {Acetates, Biodegradation, Environmental, Geobacter, Groundwater, Methane, Methanosarcina, Oxidation-Reduction, Uranium, Water Pollutants, Chemical}, issn = {1432-184X}, doi = {10.1007/s00248-018-1165-5}, author = {Holmes, Dawn E and Orelana, Roberto and Giloteaux, Ludovic and Wang, Li-Ying and Shrestha, Pravin and Williams, Kenneth and Lovley, Derek R and Rotaru, Amelia-Elena} } @article {2218, title = {Spatial control of cell envelope biosynthesis in mycobacteria}, journal = {Pathogens and Disease}, volume = {76}, year = {2018}, month = {mar}, doi = {10.1093/femspd/fty027}, url = {https://doi.org/10.1093/femspd/fty027}, author = {Julia Puffal and Alam Garc{\'\i}a-Heredia and Kathryn C Rahlwes and M Sloan Siegrist and Yasu S Morita} } @article {3084, title = { Strains Expressing Poorly Conductive Pili Reveal Constraints on Direct Interspecies Electron Transfer Mechanisms.}, journal = {mBio}, volume = {9}, year = {2018}, month = {2018 Jul 10}, abstract = {Cytochrome-to-cytochrome electron transfer and electron transfer along conduits of multiple extracellular magnetite grains are often proposed as strategies for direct interspecies electron transfer (DIET) that do not require electrically conductive pili (e-pili). However, physical evidence for these proposed DIET mechanisms has been lacking. To investigate these possibilities further, we constructed strain Aro-5, in which the wild-type pilin gene was replaced with the pilin gene that was previously shown to yield poorly conductive pili in strain Aro-5. strain Aro-5 did not reduce Fe(III) oxide and produced only low current densities, phenotypes consistent with expression of poorly conductive pili. Like strain Aro-5, strain Aro-5 displayed abundant outer surface cytochromes. Cocultures initiated with wild-type as the electron-donating strain and strain Aro-5 as the electron-accepting strain grew via DIET. However, Aro-5/ wild-type cocultures did not. Cocultures initiated with the Aro-5 strains of both species grew only when amended with granular activated carbon (GAC), a conductive material known to be a conduit for DIET. Magnetite could not substitute for GAC. The inability of the two Aro-5 strains to adapt for DIET in the absence of GAC suggests that there are physical constraints on establishing DIET solely through cytochrome-to-cytochrome electron transfer or along chains of magnetite. The finding that DIET is possible with electron-accepting partners that lack highly conductive pili greatly expands the range of potential electron-accepting partners that might participate in DIET. DIET is thought to be an important mechanism for interspecies electron exchange in natural anaerobic soils and sediments in which methane is either produced or consumed, as well as in some photosynthetic mats and anaerobic digesters converting organic wastes to methane. Understanding the potential mechanisms for DIET will not only aid in modeling carbon and electron flow in these geochemically significant environments but will also be helpful for interpreting meta-omic data from as-yet-uncultured microbes in DIET-based communities and for designing strategies to promote DIET in anaerobic digesters. The results demonstrate the need to develop a better understanding of the diversity of types of e-pili in the microbial world to identify potential electron-donating partners for DIET. Novel methods for recovering as-yet-uncultivated microorganisms capable of DIET in culture will be needed to further evaluate whether DIET is possible without e-pili in the absence of conductive materials such as GAC.
}, keywords = {Cytochromes, Electron Transport, Ferric Compounds, Fimbriae, Bacterial, Geobacter, Microbial Interactions, Oxidation-Reduction}, issn = {2150-7511}, doi = {10.1128/mBio.01273-18}, author = {Ueki, Toshiyuki and Nevin, Kelly P and Rotaru, Amelia-Elena and Wang, Li-Ying and Ward, Joy E and Woodard, Trevor L and Lovley, Derek R} } @article {2217, title = {Stress-Induced Reorganization of the Mycobacterial Membrane Domain}, journal = {{mBio}}, volume = {9}, year = {2018}, month = {jan}, doi = {10.1128/mbio.01823-17}, url = {https://doi.org/10.1128/mbio.01823-17}, author = {Jennifer M. Hayashi and Kirill Richardson and Emily S. Melzer and Steven J. Sandler and Bree B. Aldridge and M. Sloan Siegrist and Yasu S. Morita}, editor = {Christina L. Stallings} } @article {3093, title = {Biofilm Formation by Clostridium ljungdahlii Is Induced by Sodium Chloride Stress: Experimental Evaluation and Transcriptome Analysis.}, journal = {PLoS One}, volume = {12}, year = {2017}, month = {2017}, pages = {e0170406}, abstract = {The acetogen Clostridium ljungdahlii is capable of syngas fermentation and microbial electrosynthesis. Biofilm formation could benefit both these applications, but was not yet reported for C. ljungdahlii. Biofilm formation does not occur under standard growth conditions, but attachment or aggregation could be induced by different stresses. The strongest biofilm formation was observed with the addition of sodium chloride. After 3 days of incubation, the biomass volume attached to a plastic surface was 20 times higher with than without the addition of 200 mM NaCl to the medium. The addition of NaCl also resulted in biofilm formation on glass, graphite and glassy carbon, the latter two being often used electrode materials for microbial electrosynthesis. Biofilms were composed of extracellular proteins, polysaccharides, as well as DNA, while pilus-like appendages were observed with, but not without, the addition of NaCl. A transcriptome analysis comparing planktonic (no NaCl) and biofilm (NaCl addition) cells showed that C. ljungdahlii coped with the salt stress by the upregulation of the general stress response, Na+ export and osmoprotectant accumulation. A potential role for poly-N-acetylglucosamines and D-alanine in biofilm formation was found. Flagellar motility was downregulated, while putative type IV pili biosynthesis genes were not expressed. Moreover, the gene expression analysis suggested the involvement of the transcriptional regulators LexA, Spo0A and CcpA in stress response and biofilm formation. This study showed that NaCl addition might be a valuable strategy to induce biofilm formation by C. ljungdahlii, which can improve the efficacy of syngas fermentation and microbial electrosynthesis applications.
}, keywords = {Biofilms, Biomass, Carbon, Clostridium, Culture Media, Culture Techniques, Fimbriae, Bacterial, Flagella, Gene Expression Profiling, Gene Expression Regulation, Bacterial, Glass, Graphite, Osmotic Pressure, RNA, Bacterial, RNA, Ribosomal, Sodium Chloride, Spores, Bacterial, Stress, Physiological}, issn = {1932-6203}, doi = {10.1371/journal.pone.0170406}, author = {Philips, Jo and Rabaey, Korneel and Lovley, Derek R and Vargas, Madeline} } @article {3089, title = {e-Biologics: Fabrication of Sustainable Electronics with "Green" Biological Materials.}, journal = {mBio}, volume = {8}, year = {2017}, month = {2017 Jun 27}, abstract = {The growing ubiquity of electronic devices is increasingly consuming substantial energy and rare resources for materials fabrication, as well as creating expansive volumes of toxic waste. This is not sustainable. Electronic biological materials (e-biologics) that are produced with microbes, or designed with microbial components as the guide for synthesis, are a potential green solution. Some e-biologics can be fabricated from renewable feedstocks with relatively low energy inputs, often while avoiding the harsh chemicals used for synthesizing more traditional electronic materials. Several are completely free of toxic components, can be readily recycled, and offer unique features not found in traditional electronic materials in terms of size, performance, and opportunities for diverse functionalization. An appropriate investment in the concerted multidisciplinary collaborative research required to identify and characterize e-biologics and to engineer materials and devices based on e-biologics could be rewarded with a new "green age" of sustainable electronic materials and devices.
}, keywords = {Biological Products, Electrical Equipment and Supplies, Electronics, Green Chemistry Technology}, issn = {2150-7511}, doi = {10.1128/mBio.00695-17}, author = {Lovley, Derek R} } @article {3094, title = {Expressing the Geobacter metallireducens PilA in Geobacter sulfurreducens Yields Pili with Exceptional Conductivity.}, journal = {mBio}, volume = {8}, year = {2017}, month = {2017 Jan 17}, abstract = {UNLABELLED: The electrically conductive pili (e-pili) of Geobacter sulfurreducens serve as a model for a novel strategy for long-range extracellular electron transfer. e-pili are also a new class of bioelectronic materials. However, the only other Geobacter pili previously studied, which were from G.~uraniireducens, were poorly conductive. In order to obtain more information on the range of pili conductivities in Geobacter species, the pili of G.~metallireducens were investigated. Heterologously expressing the PilA gene of G.~metallireducens in G.~sulfurreducens yielded a G.~sulfurreducens strain, designated strain MP, that produced abundant pili. Strain MP exhibited phenotypes consistent with the presence of e-pili, such as high rates of Fe(III) oxide reduction and high current densities on graphite anodes. Individual pili prepared at physiologically relevant pH~7 had conductivities of 277 {\textpm} 18.9 S/cm (mean {\textpm} standard deviation), which is 5,000-fold higher than the conductivity of G.~sulfurreducens pili at pH~7 and nearly 1 million-fold higher than the conductivity of G.~uraniireducens pili at the same pH. A potential explanation for the higher conductivity of the G.~metallireducens pili is their greater density of aromatic amino acids, which are known to be important components in electron transport along the length of the pilus. The G.~metallireducens pili represent the most highly conductive pili found to date and suggest strategies for designing synthetic pili with even higher conductivities.
IMPORTANCE: e-pili are a remarkable electrically conductive material that can be sustainably produced without harsh chemical processes from renewable feedstocks and that contain no toxic components in the final product. Thus, e-pili offer an unprecedented potential for developing novel materials, electronic devices, and sensors for diverse applications with a new "green" technology. Increasing e-pili conductivity will even further expand their potential applications. A proven strategy is to design synthetic e-pili that contain tryptophan, an aromatic amino acid not found in previously studied e-pili. The studies reported here demonstrate that a productive alternative approach is to search more broadly in the microbial world. Surprisingly, even though G.~metallireducens and G.~sulfurreducens are closely related, the conductivities of their e-pili differ by more than 3 orders of magnitude. The ability to produce e-pili with high conductivity without generating a genetically modified product enhances the attractiveness of this novel electronic material.
}, keywords = {Electric Conductivity, Electrodes, Electron Transport, Ferric Compounds, Fimbriae Proteins, Gene Expression, Geobacter, Oxidation-Reduction, Recombinant Proteins}, issn = {2150-7511}, doi = {10.1128/mBio.02203-16}, author = {Tan, Yang and Adhikari, Ramesh Y and Malvankar, Nikhil S and Ward, Joy E and Woodard, Trevor L and Nevin, Kelly P and Lovley, Derek R} } @article {3097, title = {Happy together: microbial communities that hook up to swap electrons.}, journal = {ISME J}, volume = {11}, year = {2017}, month = {2017 Feb}, pages = {327-336}, abstract = {The discovery of direct interspecies electron transfer (DIET) and cable bacteria has demonstrated that microbial cells can exchange electrons over long distances (μm-cm) through electrical connections. For example, in the presence of cable bacteria electrons are rapidly transported over centimeter distances, coupling the oxidation of reduced sulfur compounds in anoxic sediments to oxygen reduction in overlying surficial sediments. Bacteria and archaea wired for DIET are found in anaerobic methane-producing and methane-consuming communities. Electrical connections between gut microbes and host cells have also been proposed. Iterative environmental and defined culture studies on methanogenic communities revealed the importance of electrically conductive pili and c-type cytochromes in natural electrical grids, and demonstrated that conductive carbon materials and magnetite can substitute for these biological connectors to facilitate DIET. This understanding has led to strategies to enhance and stabilize anaerobic digestion. Key unknowns warranting further investigation include elucidation of the archaeal electrical connections facilitating DIET-based methane production and consumption; and the mechanisms for long-range electron transfer through cable bacteria. A better understanding of mechanisms for cell-to-cell electron transfer could facilitate the hunt for additional electrically connected microbial communities with omics approaches and could advance spin-off applications such as the development of sustainable bioelectronics materials and bioelectrochemical technologies.
}, keywords = {Archaea, Bacteria, Electron Transport, Methane, Microbial Consortia, Microbial Interactions, Models, Biological, Oxidation-Reduction}, issn = {1751-7370}, doi = {10.1038/ismej.2016.136}, author = {Lovley, Derek R} } @article {3092, title = {Metatranscriptomic Evidence for Direct Interspecies Electron Transfer between Geobacter and Methanothrix Species in Methanogenic Rice Paddy Soils.}, journal = {Appl Environ Microbiol}, volume = {83}, year = {2017}, month = {2017 May 01}, abstract = {The possibility that (formerly ) and species cooperate via direct interspecies electron transfer (DIET) in terrestrial methanogenic environments was investigated in rice paddy soils. Genes with high sequence similarity to the gene for the PilA pilin monomer of the electrically conductive pili (e-pili) of accounted for over half of the PilA gene sequences in metagenomic libraries and 42\% of the mRNA transcripts in RNA sequencing (RNA-seq) libraries. This abundance of e-pilin genes and transcripts is significant because e-pili can serve as conduits for DIET. Most of the e-pilin genes and transcripts were affiliated with species, but sequences most closely related to putative e-pilin genes from genera such as , , , and , were also detected. Approximately 17\% of all metagenomic and metatranscriptomic bacterial sequences clustered with species, and the finding that spp. were actively transcribing growth-related genes indicated that they were metabolically active in the soils. Genes coding for e-pilin were among the most highly transcribed genes. In addition, homologs of genes encoding OmcS, a -type cytochrome associated with the e-pili of and required for DIET, were also highly expressed in the soils. species in the soils highly expressed genes for enzymes involved in the reduction of carbon dioxide to methane. DIET is the only electron donor known to support CO reduction in Thus, these results are consistent with a model in which species were providing electrons to species for methane production through electrical connections of e-pili. species are some of the most important microbial contributors to global methane production, but surprisingly little is known about their physiology and ecology. The possibility that DIET is a source of electrons for in methanogenic rice paddy soils is important because it demonstrates that the contribution that makes to methane production in terrestrial environments may extend beyond the conversion of acetate to methane. Furthermore, defined coculture studies have suggested that when species receive some of their energy from DIET, they grow faster than when acetate is their sole energy source. Thus, growth and metabolism in methanogenic soils may be faster and more robust than generally considered. The results also suggest that the reason that species are repeatedly found to be among the most metabolically active microorganisms in methanogenic soils is that they grow syntrophically in cooperation with spp., and possibly other methanogens, via DIET.
}, keywords = {Carbon Dioxide, Electron Transport, Fimbriae Proteins, Gene Expression Profiling, Geobacter, Metagenome, Methane, Methanosarcinaceae, Oryza, Soil Microbiology}, issn = {1098-5336}, doi = {10.1128/AEM.00223-17}, author = {Holmes, Dawn E and Shrestha, Pravin M and Walker, David J F and Dang, Yan and Nevin, Kelly P and Woodard, Trevor L and Lovley, Derek R} } @article {1963, title = {Nuclease escape elements protect messenger RNA against cleavage by multiple viral endonucleases}, journal = {PLOS Pathogens}, volume = {13}, year = {2017}, month = {aug}, pages = {e1006593}, doi = {10.1371/journal.ppat.1006593}, url = {https://doi.org/10.1371/journal.ppat.1006593}, author = {Mandy Muller and Britt A. Glaunsinger}, editor = {James R. Smiley} } @article {3088, title = {Syntrophy Goes Electric: Direct Interspecies Electron Transfer.}, journal = {Annu Rev Microbiol}, volume = {71}, year = {2017}, month = {2017 Sep 08}, pages = {643-664}, abstract = {Direct interspecies electron transfer (DIET) has biogeochemical significance, and practical applications that rely on DIET or DIET-based aspects of microbial physiology are growing. Mechanisms for DIET have primarily been studied in defined cocultures in which Geobacter species are one of the DIET partners. Electrically conductive pili (e-pili) can be an important electrical conduit for DIET. However, there may be instances in which electrical contacts are made between electron transport proteins associated with the outer membranes of the partners. Alternatively, DIET partners can plug into conductive carbon materials, such as granular activated carbon, carbon cloth, and biochar, for long-range electron exchange without the need for e-pili. Magnetite promotes DIET, possibly by acting as a substitute for outer-surface c-type cytochromes. DIET is the primary mode of interspecies electron exchange in some anaerobic digesters converting wastes to methane. Promoting DIET with conductive materials shows promise for stabilizing and accelerating methane production in digesters, permitting higher organic loading rates. Various lines of evidence suggest that DIET is important in terrestrial wetlands, which are an important source of atmospheric methane. DIET may also have a role in anaerobic methane oxidation coupled to sulfate reduction, an important control on methane releases. The finding that DIET can serve as the source of electrons for anaerobic photosynthesis further broadens its potential environmental significance. Microorganisms capable of DIET are good catalysts for several bioelectrochemical technologies and e-pili are a promising renewable source of electronic materials. The study of DIET is in its early stages, and additional investigation is required to better understand the diversity of microorganisms that are capable of DIET, the importance of DIET to carbon and electron flow in anaerobic environments, and the biochemistry and physiology of DIET.
}, keywords = {Anaerobiosis, Cytochromes, Electron Transport, Environmental Microbiology, Geobacter, Industrial Microbiology, Methane, Oxidation-Reduction}, issn = {1545-3251}, doi = {10.1146/annurev-micro-030117-020420}, author = {Lovley, Derek R} } @article {3090, title = {Toward establishing minimum requirements for extracellular electron transfer in Geobacter sulfurreducens.}, journal = {FEMS Microbiol Lett}, volume = {364}, year = {2017}, month = {2017 May 01}, abstract = {The highly redundant pathways for extracellular electron transfer in Geobacter sulfurreducens must be simplified for this microorganism to serve as an effective chassis for applications such as the development of sensors and biocomputing. Five homologs of the periplasmic c-type cytochromes, PpcA-E, offer the possibility of multiple routes of electron transfer across the periplasm. The presence of a large number of outer membrane c-type cytochromes allows G. sulfurreducens to adapt to disruption of an electron transfer pathway in the outer membrane. A strain in which genes for all five periplasmic cytochromes, PpcA-E, were deleted did not reduce Fe(III). Introducing ppcA under the control of an IPTG-inducible system in the quintuple deletion strain yielded a strain that reduced Fe(III) only in the presence of IPTG. A strain lacking known major outer membrane cytochromes, OmcB, OmcE, OmcS and OmcT, and putative functional homologs of OmcB, did not reduce Fe(III). Introduction of omcB in this septuple deletion strain restored the ability to reduce Fe(III). These results demonstrate that it is possible to trim redundancy from the extracellular electron transfer pathways in G. sulfurreducens in order to construct strains with defined extracellular electron transfer routes.
}, keywords = {Bacterial Outer Membrane Proteins, Bacterial Proteins, Cytochromes c, Electron Transport, Ferric Compounds, Gene Expression Regulation, Bacterial, Geobacter, Oxidation-Reduction, Periplasm}, issn = {1574-6968}, doi = {10.1093/femsle/fnx093}, author = {Ueki, Toshiyuki and DiDonato, Laurie N and Lovley, Derek R} } @article {3086, title = {Transcriptomic profiles of Clostridium ljungdahlii during lithotrophic growth with syngas or H and CO compared to organotrophic growth with fructose.}, journal = {Sci Rep}, volume = {7}, year = {2017}, month = {2017 Oct 13}, pages = {13135}, abstract = {Clostridium ljungdahlii derives energy by lithotrophic and organotrophic acetogenesis. C. ljungdahlii was grown organotrophically with fructose and also lithotrophically, either with syngas - a gas mixture containing hydrogen (H), carbon dioxide (CO), and carbon monoxide (CO), or with H and CO. Gene expression was compared quantitatively by microarrays using RNA extracted from all three conditions. Gene expression with fructose and with H/CO was compared by RNA-Seq. Upregulated genes with both syngas and H/CO (compared to fructose) point to the urea cycle, uptake and degradation of peptides and amino acids, response to sulfur starvation, potentially NADPH-producing pathways involving (S)-malate and ornithine, quorum sensing, sporulation, and cell wall remodeling, suggesting a global and multicellular response to lithotrophic conditions. With syngas, the upregulated (R)-lactate dehydrogenase gene represents a route of electron transfer from ferredoxin to NAD. With H/CO, flavodoxin and histidine biosynthesis genes were upregulated. Downregulated genes corresponded to an intracytoplasmic microcompartment for disposal of methylglyoxal, a toxic byproduct of glycolysis, as 1-propanol. Several cytoplasmic and membrane-associated redox-active protein genes were differentially regulated. The transcriptomic profiles of C. ljungdahlii in lithotrophic and organotrophic growth modes indicate large-scale physiological and metabolic differences, observations that may guide biofuel and commodity chemical production with this species.
}, keywords = {Carbon Dioxide, Carbon Monoxide, Clostridium, Fructose, Hydrogen, NADP, Transcriptome}, issn = {2045-2322}, doi = {10.1038/s41598-017-12712-w}, author = {Aklujkar, Muktak and Leang, Ching and Shrestha, Pravin M and Shrestha, Minita and Lovley, Derek R} } @article {3091, title = {The electrically conductive pili of pecies are a recently evolved feature for extracellular electron transfer.}, journal = {Microb Genom}, volume = {2}, year = {2016}, month = {2016 Aug}, pages = {e000072}, abstract = {The electrically conductive pili (e-pili) of have environmental and practical significance because they can facilitate electron transfer to insoluble Fe(III) oxides; to other microbial species; and through electrically conductive biofilms. E-pili conductivity has been attributed to the truncated PilA monomer, which permits tight packing of aromatic amino acids to form a conductive path along the length of e-pili. In order to better understand the evolution and distribution of e-pili in the microbial world, type IVa PilA proteins from various Gram-negative and Gram-positive bacteria were examined with a particular emphasis on Fe(III)-respiring bacteria. E-pilin genes are primarily restricted to a tight phylogenetic group in the order Desulfuromonadales. The downstream gene in all but one of the Desulfuromonadales that possess an e-pilin gene is a gene previously annotated as {\textquoteright}{\textquoteright} that has characteristics suggesting that it may encode an outer-membrane protein. Other genes associated with pilin function are clustered with e-pilin and {\textquoteright}{\textquoteright} genes in the Desulfuromonadales. In contrast, in the few bacteria outside the Desulfuromonadales that contain e-pilin genes, the other genes required for pilin function may have been acquired through horizontal gene transfer. Of the 95 known Fe(III)-reducing micro-organisms for which genomes are available, 80 \% lack e-pilin genes, suggesting that e-pili are just one of several mechanisms involved in extracellular electron transport. These studies provide insight into where and when e-pili are likely to contribute to extracellular electron transport processes that are biogeochemically important and involved in bioenergy conversions.
}, keywords = {Electromagnetic Phenomena, Electron Transport, Ferric Compounds, Fimbriae Proteins, Fimbriae, Bacterial, Geobacter, Phylogeny}, issn = {2057-5858}, doi = {10.1099/mgen.0.000072}, author = {Holmes, Dawn E and Dang, Yan and Walker, David J F and Lovley, Derek R} } @article {3099, title = {Enhancing anaerobic digestion of complex organic waste with carbon-based conductive materials.}, journal = {Bioresour Technol}, volume = {220}, year = {2016}, month = {2016 Nov}, pages = {516-522}, abstract = {The aim of this work was to study the methanogenic metabolism of dog food, a food waste surrogate, in laboratory-scale reactors with different carbon-based conductive materials. Carbon cloth, carbon felt, and granular activated carbon all permitted higher organic loading rates and promoted faster recovery of soured reactors than the control reactors. Microbial community analysis revealed that specific and substantial enrichments of Sporanaerobacter and Methanosarcina were present on the carbon cloth surface. These results, and the known ability of Sporanaerobacter species to transfer electrons to elemental sulfur, suggest that Sporanaerobacter species can participate in direct interspecies electron transfer with Methanosarcina species when carbon cloth is available as an electron transfer mediator.
}, keywords = {Anaerobiosis, Animals, Bacteria, Bioreactors, Carbon, Carbon Fiber, Charcoal, Dogs, Electric Conductivity, Fatty Acids, Volatile, Hydrogen-Ion Concentration, Methane, Organic Chemicals, Waste Products}, issn = {1873-2976}, doi = {10.1016/j.biortech.2016.08.114}, author = {Dang, Yan and Holmes, Dawn E and Zhao, Zhiqiang and Woodard, Trevor L and Zhang, Yaobin and Sun, Dezhi and Wang, Li-Ying and Nevin, Kelly P and Lovley, Derek R} } @article {3103, title = {Expanding the Diet for DIET: Electron Donors Supporting Direct Interspecies Electron Transfer (DIET) in Defined Co-Cultures.}, journal = {Front Microbiol}, volume = {7}, year = {2016}, month = {2016}, pages = {236}, abstract = {Direct interspecies electron transfer (DIET) has been recognized as an alternative to interspecies H2 transfer as a mechanism for syntrophic growth, but previous studies on DIET with defined co-cultures have only documented DIET with ethanol as the electron donor in the absence of conductive materials. Co-cultures of Geobacter metallireducens and Geobacter sulfurreducens metabolized propanol, butanol, propionate, and butyrate with the reduction of fumarate to succinate. G. metallireducens utilized each of these substrates whereas only electrons available from DIET supported G. sulfurreducens respiration. A co-culture of G. metallireducens and a strain of G. sulfurreducens that could not metabolize acetate oxidized acetate with fumarate as the electron acceptor, demonstrating that acetate can also be syntrophically metabolized via DIET. A co-culture of G. metallireducens and Methanosaeta harundinacea previously shown to syntrophically convert ethanol to methane via DIET metabolized propanol or butanol as the sole electron donor, but not propionate or butyrate. The stoichiometric accumulation of propionate or butyrate in the propanol- or butanol-fed cultures demonstrated that M. harundinaceae could conserve energy to support growth solely from electrons derived from DIET. Co-cultures of G. metallireducens and Methanosarcina barkeri could also incompletely metabolize propanol and butanol and did not metabolize propionate or butyrate as sole electron donors. These results expand the range of substrates that are known to be syntrophically metabolized through DIET, but suggest that claims of propionate and butyrate metabolism via DIET in mixed microbial communities warrant further validation.
}, issn = {1664-302X}, doi = {10.3389/fmicb.2016.00236}, author = {Wang, Li-Ying and Nevin, Kelly P and Woodard, Trevor L and Mu, Bo-Zhong and Lovley, Derek R} } @article {3106, title = {Functional environmental proteomics: elucidating the role of a c-type cytochrome abundant during uranium bioremediation.}, journal = {ISME J}, volume = {10}, year = {2016}, month = {2016 Feb}, pages = {310-20}, abstract = {Studies with pure cultures of dissimilatory metal-reducing microorganisms have demonstrated that outer-surface c-type cytochromes are important electron transfer agents for the reduction of metals, but previous environmental proteomic studies have typically not recovered cytochrome sequences from subsurface environments in which metal reduction is important. Gel-separation, heme-staining and mass spectrometry of proteins in groundwater from in situ uranium bioremediation experiments identified a putative c-type cytochrome, designated Geobacter subsurface c-type cytochrome A (GscA), encoded within the genome of strain M18, a Geobacter isolate previously recovered from the site. Homologs of GscA were identified in the genomes of other Geobacter isolates in the phylogenetic cluster known as subsurface clade 1, which predominates in a diversity of Fe(III)-reducing subsurface environments. Most of the gscA sequences recovered from groundwater genomic DNA clustered in a tight phylogenetic group closely related to strain M18. GscA was most abundant in groundwater samples in which Geobacter sp. predominated. Expression of gscA in a strain of Geobacter sulfurreducens that lacked the gene for the c-type cytochrome OmcS, thought to facilitate electron transfer from conductive pili to Fe(III) oxide, restored the capacity for Fe(III) oxide reduction. Atomic force microscopy provided evidence that GscA was associated with the pili. These results demonstrate that a c-type cytochrome with an apparent function similar to that of OmcS is abundant when Geobacter sp. are abundant in the subsurface, providing insight into the mechanisms for the growth of subsurface Geobacter sp. on Fe(III) oxide and suggesting an approach for functional analysis of other Geobacter proteins found in the subsurface.
}, keywords = {Amino Acid Sequence, Bacterial Proteins, Biodegradation, Environmental, Cytochrome c Group, Electron Transport, Ferric Compounds, Geobacter, Groundwater, Molecular Sequence Data, Oxidation-Reduction, Phylogeny, Proteomics, Sequence Alignment, Uranium}, issn = {1751-7370}, doi = {10.1038/ismej.2015.113}, author = {Yun, Jiae and Malvankar, Nikhil S and Ueki, Toshiyuki and Lovley, Derek R} } @article {3098, title = {Genetic switches and related tools for controlling gene expression and electrical outputs of Geobacter sulfurreducens.}, journal = {J Ind Microbiol Biotechnol}, volume = {43}, year = {2016}, month = {2016 Nov}, pages = {1561-1575}, abstract = {Physiological studies and biotechnology applications of Geobacter species have been limited by a lack of genetic tools. Therefore, potential additional molecular strategies for controlling metabolism were explored. When the gene for citrate synthase, or acetyl-CoA transferase, was placed under the control of a LacI/IPTG regulator/inducer system, cells grew on acetate only in the presence of IPTG. The TetR/AT system could also be used to control citrate synthase gene expression and acetate metabolism. A strain that required IPTG for growth on D-lactate was constructed by placing the gene for D-lactate dehydrogenase under the control of the LacI/IPTG system. D-Lactate served as an inducer in a strain in which a D-lactate responsive promoter and transcription repressor were used to control citrate synthase expression. Iron- and potassium-responsive systems were successfully incorporated to regulate citrate synthase expression and growth on acetate. Linking the appropriate degradation tags on the citrate synthase protein made it possible to control acetate metabolism with either the endogenous ClpXP or exogenous Lon protease and tag system. The ability to control current output from Geobacter biofilms and the construction of an AND logic gate for acetate metabolism suggested that the tools developed may be applicable for biosensor and biocomputing applications.
}, keywords = {Acetates, Acetyl Coenzyme A, Citrate (si)-Synthase, Electric Conductivity, Gene Expression Regulation, Geobacter, Isopropyl Thiogalactoside, L-Lactate Dehydrogenase, Lac Repressors, Promoter Regions, Genetic, Transferases}, issn = {1476-5535}, doi = {10.1007/s10295-016-1836-5}, author = {Ueki, Toshiyuki and Nevin, Kelly P and Woodard, Trevor L and Lovley, Derek R} } @article {3095, title = {How to Sustainably Feed a Microbe: Strategies for Biological Production of Carbon-Based Commodities with Renewable Electricity.}, journal = {Front Microbiol}, volume = {7}, year = {2016}, month = {2016}, pages = {1879}, abstract = {As interest and application of renewable energy grows, strategies are needed to align the asynchronous supply and demand. Microbial metabolisms are a potentially sustainable mechanism for transforming renewable electrical energy into biocommodities that are easily stored and transported. Acetogens and methanogens can reduce carbon dioxide to organic products including methane, acetic acid, and ethanol. The library of biocommodities is expanded when engineered metabolisms of acetogens are included. Typically, electrochemical systems are employed to integrate renewable energy sources with biological systems for production of carbon-based commodities. Within these systems, there are three prevailing mechanisms for delivering electrons to microorganisms for the conversion of carbon dioxide to reduce organic compounds: (1) electrons can be delivered to microorganisms via H produced separately in a electrolyzer, (2) H produced at a cathode can convey electrons to microorganisms supported on the cathode surface, and (3) a cathode can directly feed electrons to microorganisms. Each of these strategies has advantages and disadvantages that must be considered in designing full-scale processes. This review considers the evolving understanding of each of these approaches and the state of design for advancing these strategies toward viability.
}, issn = {1664-302X}, doi = {10.3389/fmicb.2016.01879}, author = {Butler, Caitlyn S and Lovley, Derek R} } @article {1718, title = {Hydrogen and thiosulfate limits for growth of a thermophilic, autotrophic Desulfurobacterium species from a deep-sea hydrothermal vent.}, journal = {Environ Microbiol Rep}, volume = {8}, year = {2016}, month = {2016 Apr}, pages = {196-200}, abstract = {Hydrothermal fluids (341{\textdegree}C and 19{\textdegree}C) were collected < 1 m apart from a black smoker chimney and a tubeworm mound on the Boardwalk edifice at the Endeavour Segment in the northeastern Pacific Ocean to study anaerobic microbial growth in hydrothermal mineral deposits. Geochemical modelling of mixed vent fluid and seawater suggests the mixture was anoxic above 55{\textdegree}C and that low H2 concentrations (79 μmol kg(-1) in end-member hydrothermal fluid) limit anaerobic hydrogenotrophic growth above this temperature. A thermophilic, hydrogenotrophic sulfur reducer, Desulfurobacterium strain HR11, was isolated from the 19{\textdegree}C fluid raising questions about its H2 -dependent growth kinetics. Strain HR11 grew at 40-77{\textdegree}C (Topt 72-75{\textdegree}C), pH 5-8.5 (pHopt 6-7) and 1-5\% (wt vol(-1) ) NaCl (NaClopt 3-4\%). The highest growth rates occurred when S2 O3 (2-) and S{\textdegree} were reduced to H2 S. Modest growth occurred by NO3 (-) reduction. Monod constants for its growth were Ks of 30 μM for H2 and Ks of 20 μM for S2 O3 (2-) with a μmax of 2.0 h(-1) . The minimum H2 and S2 O3 (2-) concentrations for growth were 3 μM and 5 μM respectively. Possible sources of S2 O3 (2-) and S{\textdegree} are from abiotic dissolved sulfide and pyrite oxidation by O2 .
}, issn = {1758-2229}, doi = {10.1111/1758-2229.12368}, author = {Stewart, Lucy C and Llewellyn, James G and Butterfield, David A and Lilley, Marvin D and Holden, James F} } @article {1708, title = {Hydrogen Production and Enzyme Activities in the Hyperthermophile Thermococcus paralvinellae Grown on Maltose, Tryptone, and Agricultural Waste.}, journal = {Front Microbiol}, volume = {7}, year = {2016}, month = {2016}, pages = {167}, abstract = {Thermococcus may be an important alternative source of H2 in the hot subseafloor in otherwise low H2 environments such as some hydrothermal vents and oil reservoirs. It may also be useful in industry for rapid agricultural waste treatment and concomitant H2 production. Thermococcus paralvinellae grown at 82{\textdegree}C without sulfur produced up to 5 mmol of H2 L(-1) at rates of 5-36 fmol H2 cell(-1) h(-1) on 0.5\% (wt vol(-1)) maltose, 0.5\% (wt vol(-1)) tryptone, and 0.5\% maltose + 0.05\% tryptone media. Two potentially inhibiting conditions, the presence of 10 mM acetate and low pH (pH 5) in maltose-only medium, did not significantly affect growth or H2 production. Growth rates, H2 production rates, and cell yields based on H2 production were the same as those for Pyrococcus furiosus grown at 95{\textdegree}C on the same media for comparison. Acetate, butyrate, succinate, isovalerate, and formate were also detected as end products. After 100 h, T. paralvinellae produced up to 5 mmol of H2 L(-1) of medium when grown on up to 70\% (vol vol(-1)) waste milk from cows undergoing treatment for mastitis with the bacterial antibiotic Ceftiofur and from untreated cows. The amount of H2 produced by T. paralvinellae increased with increasing waste concentrations, but decreased in P. furiosus cultures supplemented with waste milk above 1\% concentration. All mesophilic bacteria from the waste milk that grew on Luria Bertani, Sheep{\textquoteright}s Blood (selective for Staphylococcus, the typical cause of mastitis), and MacConkey (selective for Gram-negative enteric bacteria) agar plates were killed by heat during incubation at 82{\textdegree}C. Ceftiofur, which is heat labile, was below the detection limit following incubation at 82{\textdegree}C. T. paralvinellae also produced up to 6 mmol of H2 L(-1) of medium when grown on 0.1-10\% (wt vol(-1)) spent brewery grain while P. furiosus produced < 1 mmol of H2 L(-1). Twelve of 13 enzyme activities in T. paralvinellae showed significant (p < 0.05) differences across six different growth conditions; however, methyl viologen-dependent membrane hydrogenase activity remained constant across all media types. The results demonstrate the potential of at least some Thermococcus species to produce H2 if protein and α-glucosides are present as substrates.
}, issn = {1664-302X}, doi = {10.3389/fmicb.2016.00167}, author = {Hensley, Sarah A and Moreira, Emily and Holden, James F} } @article {2373, title = {Leishmania DNA Replication Timing: A Stochastic Event?}, journal = {Trends Parasitol}, volume = {32}, year = {2016}, month = {2016 10}, pages = {755-757}, abstract = {For eukaryotic genomes, DNA synthesis initiates at multiple discrete regions known as replication origins in a dynamic yet regulated manner to ensure genomic stability. Two recent studies using different approaches reveal few Leishmania origins and that origin firing may proceed in a mainly stochastic manner.
}, keywords = {Animals, DNA Replication Timing, Drug Delivery Systems, Genomic Instability, Humans, Leishmania, Leishmaniasis, Replication Origin}, issn = {1471-5007}, doi = {10.1016/j.pt.2016.05.011}, author = {Rocha-Granados, Maria C and Klingbeil, Michele M} } @article {3100, title = {The Low Conductivity of Geobacter uraniireducens Pili Suggests a Diversity of Extracellular Electron Transfer Mechanisms in the Genus Geobacter.}, journal = {Front Microbiol}, volume = {7}, year = {2016}, month = {2016}, pages = {980}, abstract = {Studies on the mechanisms for extracellular electron transfer in Geobacter species have primarily focused on Geobacter sulfurreducens, but the poor conservation of genes for some electron transfer components within the Geobacter genus suggests that there may be a diversity of extracellular electron transport strategies among Geobacter species. Examination of the gene sequences for PilA, the type IV pilus monomer, in Geobacter species revealed that the PilA sequence of Geobacter uraniireducens was much longer than that of G. sulfurreducens. This is of interest because it has been proposed that the relatively short PilA sequence of G. sulfurreducens is an important feature conferring conductivity to G. sulfurreducens pili. In order to investigate the properties of the G. uraniireducens pili in more detail, a strain of G. sulfurreducens that expressed pili comprised the PilA of G. uraniireducens was constructed. This strain, designated strain GUP, produced abundant pili, but generated low current densities and reduced Fe(III) very poorly. At pH 7, the conductivity of the G. uraniireducens pili was 3 {\texttimes} 10(-4) S/cm, much lower than the previously reported 5 {\texttimes} 10(-2) S/cm conductivity of G. sulfurreducens pili at the same pH. Consideration of the likely voltage difference across pili during Fe(III) oxide reduction suggested that G. sulfurreducens pili can readily accommodate maximum reported rates of respiration, but that G. uraniireducens pili are not sufficiently conductive to be an effective mediator of long-range electron transfer. In contrast to G. sulfurreducens and G. metallireducens, which require direct contact with Fe(III) oxides in order to reduce them, G. uraniireducens reduced Fe(III) oxides occluded within microporous beads, demonstrating that G. uraniireducens produces a soluble electron shuttle to facilitate Fe(III) oxide reduction. The results demonstrate that Geobacter species may differ substantially in their mechanisms for long-range electron transport and that it is important to have information beyond a phylogenetic affiliation in order to make conclusions about the mechanisms by which Geobacter species are transferring electrons to extracellular electron acceptors.
}, issn = {1664-302X}, doi = {10.3389/fmicb.2016.00980}, author = {Tan, Yang and Adhikari, Ramesh Y and Malvankar, Nikhil S and Ward, Joy E and Nevin, Kelly P and Woodard, Trevor L and Smith, Jessica A and Snoeyenbos-West, Oona L and Franks, Ashley E and Tuominen, Mark T and Lovley, Derek R} } @article {3102, title = {Low Energy Atomic Models Suggesting a Pilus Structure that could Account for Electrical Conductivity of Geobacter sulfurreducens Pili.}, journal = {Sci Rep}, volume = {6}, year = {2016}, month = {2016 Mar 22}, pages = {23385}, abstract = {The metallic-like electrical conductivity of Geobacter sulfurreducens pili has been documented with multiple lines of experimental evidence, but there is only a rudimentary understanding of the structural features which contribute to this novel mode of biological electron transport. In order to determine if it was feasible for the pilin monomers of G. sulfurreducens to assemble into a conductive filament, theoretical energy-minimized models of Geobacter pili were constructed with a previously described approach, in which pilin monomers are assembled using randomized structural parameters and distance constraints. The lowest energy models from a specific group of predicted structures lacked a central channel, in contrast to previously existing pili models. In half of the no-channel models the three N-terminal aromatic residues of the pilin monomer are arranged in a potentially electrically conductive geometry, sufficiently close to account for the experimentally observed metallic like conductivity of the pili that has been attributed to overlapping pi-pi orbitals of aromatic amino acids. These atomic resolution models capable of explaining the observed conductive properties of Geobacter pili are a valuable tool to guide further investigation of the metallic-like conductivity of the pili, their role in biogeochemical cycling, and applications in bioenergy and bioelectronics.
}, keywords = {Electric Conductivity, Fimbriae, Bacterial, Geobacter, Models, Theoretical}, issn = {2045-2322}, doi = {10.1038/srep23385}, author = {Xiao, Ke and Malvankar, Nikhil S and Shu, Chuanjun and Martz, Eric and Lovley, Derek R and Sun, Xiao} } @article {3104, title = {Potential enhancement of direct interspecies electron transfer for syntrophic metabolism of propionate and butyrate with biochar in up-flow anaerobic sludge blanket reactors.}, journal = {Bioresour Technol}, volume = {209}, year = {2016}, month = {2016 Jun}, pages = {148-56}, abstract = {Promoting direct interspecies electron transfer (DIET) to enhance syntrophic metabolism may be a strategy for accelerating the conversion of organic wastes to methane, but microorganisms capable of metabolizing propionate and butyrate via DIET under methanogenic conditions have yet to be identified. In an attempt to establish methanogenic communities metabolizing propionate or butyrate with DIET, enrichments were initiated with up-flow anaerobic sludge blanket (UASB), similar to those that were previously reported to support communities that metabolized ethanol with DIET that relied on direct biological electrical connections. In the absence of any amendments, microbial communities enriched were dominated by microorganisms closely related to pure cultures that are known to metabolize propionate or butyrate to acetate with production of H2. When biochar was added to the reactors there was a substantial enrichment on the biochar surface of 16S rRNA gene sequences closely related to Geobacter and Methanosaeta species known to participate in DIET.
}, keywords = {Acetates, Bioreactors, Butyric Acid, Charcoal, Electron Transport, Geobacter, Methane, Microbial Consortia, Propionates, RNA, Ribosomal, 16S, Sewage, Waste Disposal, Fluid}, issn = {1873-2976}, doi = {10.1016/j.biortech.2016.03.005}, author = {Zhao, Zhiqiang and Zhang, Yaobin and Holmes, Dawn E and Dang, Yan and Woodard, Trevor L and Nevin, Kelly P and Lovley, Derek R} } @article {3096, title = {Reply to {\textquoteright}Measuring conductivity of living Geobacter sulfurreducens biofilms{\textquoteright}.}, journal = {Nat Nanotechnol}, volume = {11}, year = {2016}, month = {2016 Nov 08}, pages = {913-914}, keywords = {Biofilms, Electric Conductivity, Geobacter}, issn = {1748-3395}, doi = {10.1038/nnano.2016.191}, author = {Malvankar, Nikhil S and Rotello, Vincent M and Tuominen, Mark T and Lovley, Derek R} } @article {3101, title = {Synthetic Biological Protein Nanowires with High Conductivity.}, journal = {Small}, volume = {12}, year = {2016}, month = {2016 Sep}, pages = {4481-5}, abstract = {Genetic modification to add tryptophan to PilA, the monomer for the electrically conductive pili of Geobacter sulfurreducens, yields conductive protein filaments 2000-fold more conductive than the wild-type pili while cutting the diameter in half to 1.5 nm.
}, keywords = {Amino Acid Sequence, Electric Conductivity, Fimbriae, Bacterial, Geobacter, Nanowires, Proteins, Tryptophan}, issn = {1613-6829}, doi = {10.1002/smll.201601112}, author = {Tan, Yang and Adhikari, Ramesh Y and Malvankar, Nikhil S and Pi, Shuang and Ward, Joy E and Woodard, Trevor L and Nevin, Kelly P and Xia, Qiangfei and Tuominen, Mark T and Lovley, Derek R} } @article {1416, title = {Antimicrobial surfaces containing cationic nanoparticles: how immobilized, clustered, and protruding cationic charge presentation affects killing activity and kinetics.}, journal = {Colloids Surf B Biointerfaces}, volume = {125}, year = {2015}, month = {2015 Jan 1}, pages = {255-63}, abstract = {This work examines how the antimicrobial (killing) activity of net-negative surfaces depends on the presentation of antimicrobial cationic functionality: distributed versus clustered, and flat clusters versus raised clusters. Specifically, the ability to kill Staphylococcus aureus by sparsely distributed 10 nm cationic nanoparticles, immobilized on a negative surface and backfilled with a PEG (polyethylene glycol) brush, was compared with that for a dense layer of the same immobilized nanoparticles. Additionally, sparsely distributed 10 nm poly-L-lysine (PLL) coils, adsorbed to a surface to produce flat cationic \"patches\" and backfilled with a PEG brush were compared to a saturated adsorbed layer of PLL. The latter resembled classical uniformly cationic antimicrobial surfaces. The protrusion of the cationic clusters substantially influenced killing but the surface concentration of the clusters had minor impact, as long as bacteria adhered. When surfaces were functionalized at the minimum nanoparticle and patch densities needed for bacterial adhesion, killing activity was substantial within 30 min and nearly complete within 2 h. Essentially identical killing was observed on more densely functionalized surfaces. Surfaces containing protruding (by about 8 nm) nanoparticles accomplished rapid killing (at 30 min) compared with surfaces containing similarly cationic but flat features (PLL patches). Importantly, the overall surface density of cationic functionality within the clusters was lower than reported thresholds for antimicrobial action. Also surprising, the nanoparticles were far more deadly when surface-immobilized compared with free in solution. These findings support a killing mechanism involving interfacial stress.
}, issn = {1873-4367}, doi = {10.1016/j.colsurfb.2014.10.043}, author = {Fang, Bing and Jiang, Ying and N{\"u}sslein, Klaus and Rotello, Vincent M and Santore, Maria M} } @article {3121, title = {Centimeter-long electron transport in marine sediments via conductive minerals.}, journal = {ISME J}, volume = {9}, year = {2015}, month = {2015 Feb}, pages = {527-31}, abstract = {Centimeter-long electron conduction through marine sediments, in which electrons derived from sulfide in anoxic sediments are transported to oxygen in surficial sediments, may have an important influence on sediment geochemistry. Filamentous bacteria have been proposed to mediate the electron transport, but the filament conductivity could not be verified and other mechanisms are possible. Surprisingly, previous investigations have never actually measured the sediment conductivity or its basic physical properties. Here we report direct measurements that demonstrate centimeter-long electron flow through marine sediments, with conductivities sufficient to account for previously estimated electron fluxes. Conductivity was lost for oxidized sediments, which contrasts with the previously described increase in the conductivity of microbial biofilms upon oxidation. Adding pyrite to the sediments significantly enhanced the conductivity. These results suggest that the role of conductive minerals, which are more commonly found in sediments than centimeter-long microbial filaments, need to be considered when modeling marine sediment biogeochemistry.
}, keywords = {Electron Transport, Geologic Sediments, Iron, Minerals, Oxidation-Reduction, Sulfides}, issn = {1751-7370}, doi = {10.1038/ismej.2014.131}, author = {Malvankar, Nikhil S and King, Gary M and Lovley, Derek R} } @article {1728, title = {Complete genome sequence of the hyperthermophilic methanogen Methanocaldococcus bathoardescens JH146(T) isolated from the basalt subseafloor.}, journal = {Mar Genomics}, volume = {24 Pt 3}, year = {2015}, month = {2015 Dec}, pages = {229-30}, abstract = {Methanocaldococcus bathoardescens JH146(T) is a hyperthermophilic and obligate hydrogenotrophic methanogen isolated from low-temperature (26 {\textdegree}C) hydrothermal vent fluid at Axial Seamount in the northeastern Pacific Ocean. It is most closely related to the N2-fixing methanogen Methanocaldococcus sp. FS406-22; however, they differ in that JH146 cannot fix N2 or reductively assimilate nitrate. In this study, we present the complete genome sequence of strain JH146(T) (1,607,556 bp) with its 1635 protein coding genes, and 41 RNA genes. Our analysis focuses on its methane production via the acetyl-CoA pathway and its deleted gene clusters related to nitrogen assimilation. This study extends our understanding of methanogenesis at high temperatures and the impact of these organisms on the biogeochemistry of subseafloor hydrothermal environments and the deep sea.
}, issn = {1876-7478}, doi = {10.1016/j.margen.2015.06.002}, author = {Kim, You-Tae and Jung, Jong-Hyun and Stewart, Lucy C and Kwon, Soon-Wo and Holden, James F and Park, Cheon-Seok} } @article {3109, title = {Enhancing syntrophic metabolism in up-flow anaerobic sludge blanket reactors with conductive carbon materials.}, journal = {Bioresour Technol}, volume = {191}, year = {2015}, month = {2015 Sep}, pages = {140-5}, abstract = {Syntrophic metabolism of alcohols and fatty acids is a critical step in anaerobic digestion, which if enhanced can better stabilize the process and enable shorter retention times. Direct interspecies electron transfer (DIET) has recently been recognized as an alternative route to hydrogen interspecies transfer as a mechanism for interspecies syntrophic electron exchange. Therefore, the possibility of accelerating syntrophic metabolism of ethanol in up-flow anaerobic sludge blanket (UASB) reactors by incorporating conductive materials in reactor design was investigated. Graphite, biochar, and carbon cloth all immediately enhanced methane production and COD removal. As the hydraulic retention time was decreased the increased effectiveness of treatment in reactors with conductive materials increased versus the control reactor. When these conductive materials were removed from the reactors rates of syntrophic metabolism declined to rates comparable to the control reactor. These results suggest that incorporating conductive materials in the design of UASB reactors may enhance digester effectiveness.
}, keywords = {Anaerobiosis, Bioreactors, Carbon, Charcoal, Electron Transport, Graphite, Hydrogen, Methane, Sewage, Waste Disposal, Fluid}, issn = {1873-2976}, doi = {10.1016/j.biortech.2015.05.007}, author = {Zhao, Zhiqiang and Zhang, Yaobin and Woodard, T L and Nevin, K P and Lovley, D R} } @article {3120, title = {Evidence of Geobacter-associated phage in a uranium-contaminated aquifer.}, journal = {ISME J}, volume = {9}, year = {2015}, month = {2015 Feb}, pages = {333-46}, abstract = {Geobacter species may be important agents in the bioremediation of organic and metal contaminants in the subsurface, but as yet unknown factors limit the in situ growth of subsurface Geobacter well below rates predicted by analysis of gene expression or in silico metabolic modeling. Analysis of the genomes of five different Geobacter species recovered from contaminated subsurface sites indicated that each of the isolates had been infected with phage. Geobacter-associated phage sequences were also detected by metagenomic and proteomic analysis of samples from a uranium-contaminated aquifer undergoing in situ bioremediation, and phage particles were detected by microscopic analysis in groundwater collected from sediment enrichment cultures. Transcript abundance for genes from the Geobacter-associated phage structural proteins, tail tube Gp19 and baseplate J, increased in the groundwater in response to the growth of Geobacter species when acetate was added, and then declined as the number of Geobacter decreased. Western blot analysis of a Geobacter-associated tail tube protein Gp19 in the groundwater demonstrated that its abundance tracked with the abundance of Geobacter species. These results suggest that the enhanced growth of Geobacter species in the subsurface associated with in situ uranium bioremediation increased the abundance and activity of Geobacter-associated phage and show that future studies should focus on how these phages might be influencing the ecology of this site.
}, keywords = {Bacteriophages, Biodegradation, Environmental, Genes, Viral, Geobacter, Groundwater, Metagenome, Proteomics, Transcriptome, Uranium, Viral Proteins, Water Pollutants, Radioactive}, issn = {1751-7370}, doi = {10.1038/ismej.2014.128}, author = {Holmes, Dawn E and Giloteaux, Ludovic and Chaurasia, Akhilesh K and Williams, Kenneth H and Luef, Birgit and Wilkins, Michael J and Wrighton, Kelly C and Thompson, Courtney A and Comolli, Luis R and Lovley, Derek R} } @article {1503, title = {Host Actin Polymerization Tunes the Cell Division Cycle of an Intracellular Pathogen.}, journal = {Cell Rep}, year = {2015}, month = {2015 Apr 15}, abstract = {Growth and division are two of the most fundamental capabilities of a bacterial cell. While they are well described for model organisms growing in broth culture, very little is known about the cell division cycle of bacteria replicating in more complex environments. Using a D-alanine reporter strategy, we found that intracellular Listeria monocytogenes (Lm) spend a smaller proportion of their cell cycle dividing compared to Lm growing in broth culture. This alteration to the cell division cycle is independent of bacterial doubling time. Instead, polymerization of host-derived actin at the bacterial cell surface extends the non-dividing elongation period and compresses the division period. By decreasing the relative proportion of dividing Lm, actin polymerization biases the population toward cells with the highest propensity to form actin tails. Thus, there is a positive-feedback loop between the Lm cell division\ cycle and a physical interaction with the host cytoskeleton.
}, issn = {2211-1247}, doi = {10.1016/j.celrep.2015.03.046}, author = {Siegrist, M Sloan and Aditham, Arjun K and Espaillat, Akbar and Cameron, Todd A and Whiteside, Sarah A and Cava, Felipe and Portnoy, Daniel A and Bertozzi, Carolyn R} } @article {1559, title = {Identification of 4-Hydroxycumyl Alcohol As the Major MnO2-Mediated Bisphenol A Transformation Product and Evaluation of Its Environmental Fate.}, journal = {Environ Sci Technol}, volume = {49}, year = {2015}, month = {2015 May 19}, pages = {6214-21}, abstract = {Bisphenol A (BPA), an environmental contaminant with weak estrogenic activity, resists microbial degradation under anoxic conditions but is susceptible to abiotic transformation by manganese dioxide (MnO2). BPA degradation followed pseudo-first-order kinetics with a rate constant of 0.96 ({\textpm}0.03) min(-1) in the presence of 2 mM MnO2 (0.017\% w/w) at pH 7.2. 4-hydroxycumyl alcohol (HCA) was the major transformation product, and, on a molar basis, up to 64\% of the initial amount of BPA was recovered as HCA. MnO2 was also reactive toward HCA, albeit at 5-fold lower rates, and CO2 evolution (i.e., mineralization) occurred. In microcosms established with freshwater sediment, HCA was rapidly biodegraded under oxic, but not anoxic conditions. With a measured octanol-water partition coefficient (Log K(ow)) of 0.76 and an aqueous solubility of 2.65 g L(-1), HCA is more mobile in saturated media than BPA (Log K(ow) = 2.76; aqueous solubility = 0.31 g L(-1)), and therefore more likely to encounter oxic zones and undergo aerobic biodegradation. These findings corroborate that BPA is not inert under anoxic conditions and suggest that MnO2-mediated coupled abiotic-biotic processes may be relevant for controlling the fate and longevity of BPA in sediments and aquifers.
}, issn = {1520-5851}, doi = {10.1021/acs.est.5b00372}, author = {Im, Jeongdae and Prevatte, Carson W and Campagna, Shawn R and L{\"o}ffler, Frank E} } @article {1437, title = {Illumination of growth, division and secretion by metabolic labeling of the bacterial cell surface.}, journal = {FEMS Microbiol Rev}, volume = {39}, year = {2015}, month = {2015 Mar}, pages = {184-202}, abstract = {The cell surface is the essential interface between a bacterium and its surroundings. Composed primarily of molecules that are not directly genetically encoded, this highly dynamic structure accommodates the basic cellular processes of growth and division as well as the transport of molecules between the cytoplasm and the extracellular milieu. In this review, we describe aspects of bacterial growth, division and secretion that have recently been uncovered by metabolic labeling of the cell envelope. Metabolite derivatives can be used to label a variety of macromolecules, from proteins to non-genetically-encoded glycans and lipids. The embedded metabolite enables precise tracking in time and space, and the versatility of newer chemoselective detection methods offers the ability to execute multiple experiments concurrently. In addition to reviewing the discoveries enabled by metabolic labeling of the bacterial cell envelope, we also discuss the potential of these techniques for translational applications. Finally, we offer some guidelines for implementing this emerging technology.
}, issn = {1574-6976}, doi = {10.1093/femsre/fuu012}, author = {Siegrist, M Sloan and Swarts, Benjamin M and Fox, Douglas M and Lim, Shion An and Bertozzi, Carolyn R} } @article {1439, title = {Land use, soil and litter chemistry drive bacterial community structures in samples of the rainforest and Cerrado (Brazilian Savannah) biomes in Southern Amazonia.}, journal = {European Journal of Soil Biology}, volume = {66}, year = {2015}, month = {2014 Nov}, pages = {32-29}, doi = {10.1016/j.ejsobi.2014.11.001}, url = {http://www.sciencedirect.com/science/article/pii/S1164556314001022}, author = {N{\"u}sslein, K and Lammel, D and Tsai, SM and Cerri, CC} } @article {3105, title = {Link between capacity for current production and syntrophic growth in Geobacter species.}, journal = {Front Microbiol}, volume = {6}, year = {2015}, month = {2015}, pages = {744}, abstract = {Electrodes are unnatural electron acceptors, and it is yet unknown how some Geobacter species evolved to use electrodes as terminal electron acceptors. Analysis of different Geobacter species revealed that they varied in their capacity for current production. Geobacter metallireducens and G. hydrogenophilus generated high current densities (ca. 0.2 mA/cm(2)), comparable to G. sulfurreducens. G. bremensis, G. chapellei, G. humireducens, and G. uraniireducens, produced much lower currents (ca. 0.05 mA/cm(2)) and G. bemidjiensis was previously found to not produce current. There was no correspondence between the effectiveness of current generation and Fe(III) oxide reduction rates. Some high-current-density strains (G. metallireducens and G. hydrogenophilus) reduced Fe(III)-oxides as fast as some low-current-density strains (G. bremensis, G. humireducens, and G. uraniireducens) whereas other low-current-density strains (G. bemidjiensis and G. chapellei) reduced Fe(III) oxide as slowly as G. sulfurreducens, a high-current-density strain. However, there was a correspondence between the ability to produce higher currents and the ability to grow syntrophically. G. hydrogenophilus was found to grow in co-culture with Methanosarcina barkeri, which is capable of direct interspecies electron transfer (DIET), but not with Methanospirillum hungatei capable only of H2 or formate transfer. Conductive granular activated carbon (GAC) stimulated metabolism of the G. hydrogenophilus - M. barkeri co-culture, consistent with electron exchange via DIET. These findings, coupled with the previous finding that G. metallireducens and G. sulfurreducens are also capable of DIET, suggest that evolution to optimize DIET has fortuitously conferred the capability for high-density current production to some Geobacter species.
}, issn = {1664-302X}, doi = {10.3389/fmicb.2015.00744}, author = {Rotaru, Amelia-Elena and Woodard, Trevor L and Nevin, Kelly P and Lovley, Derek R} } @article {1451, title = {Long-term warming drives loss of dominant taxa and rRNA copy number among soil bacteria.}, journal = {Frontiers in Microbiology}, year = {2015}, month = {2015 Jan}, doi = {10.3389/fmicb.2015.00104}, author = {DeAngelis, KM and Pold, G and Topcuoglu, B and van Diepen, L and Blanchard, J and Melillo, J and Frey, S} } @article {3128, title = {Magnetite compensates for the lack of a pilin-associated c-type cytochrome in extracellular electron exchange.}, journal = {Environ Microbiol}, volume = {17}, year = {2015}, month = {2015 Mar}, pages = {648-55}, abstract = {Nanoscale magnetite can facilitate microbial extracellular electron transfer that plays an important role in biogeochemical cycles, bioremediation and several bioenergy strategies, but the mechanisms for the stimulation of extracellular electron transfer are poorly understood. Further investigation revealed that magnetite attached to the electrically conductive pili of Geobacter species in a manner reminiscent of the association of the multi-heme c-type cytochrome OmcS with the pili of Geobacter sulfurreducens. Magnetite conferred extracellular electron capabilities on an OmcS-deficient strain unable to participate in interspecies electron transfer or Fe(III) oxide reduction. In the presence of magnetite wild-type cells repressed expression of the OmcS gene, suggesting that cells might need to produce less OmcS when magnetite was available. The finding that magnetite can compensate for the lack of the electron transfer functions of a multi-heme c-type cytochrome has implications not only for the function of modern microbes, but also for the early evolution of microbial electron transport mechanisms.
}, keywords = {Cytochrome c Group, Electron Transport, Electrons, Ferrosoferric Oxide, Fimbriae Proteins, Fimbriae, Bacterial, Gene Expression Regulation, Bacterial, Geobacter, Heme, Oxides}, issn = {1462-2920}, doi = {10.1111/1462-2920.12485}, author = {Liu, Fanghua and Rotaru, Amelia-Elena and Shrestha, Pravin M and Malvankar, Nikhil S and Nevin, Kelly P and Lovley, Derek R} } @article {1737, title = {Methane cycling. Nonequilibrium clumped isotope signals in microbial methane.}, journal = {Science}, volume = {348}, year = {2015}, month = {2015 Apr 24}, pages = {428-31}, abstract = {Methane is a key component in the global carbon cycle, with a wide range of anthropogenic and natural sources. Although isotopic compositions of methane have traditionally aided source identification, the abundance of its multiply substituted "clumped" isotopologues (for example, (13)CH3D) has recently emerged as a proxy for determining methane-formation temperatures. However, the effect of biological processes on methane{\textquoteright}s clumped isotopologue signature is poorly constrained. We show that methanogenesis proceeding at relatively high rates in cattle, surface environments, and laboratory cultures exerts kinetic control on (13)CH3D abundances and results in anomalously elevated formation-temperature estimates. We demonstrate quantitatively that H2 availability accounts for this effect. Clumped methane thermometry can therefore provide constraints on the generation of methane in diverse settings, including continental serpentinization sites and ancient, deep groundwaters.
}, keywords = {Animals, Carbon Cycle, Carbon Isotopes, Cattle, Groundwater, Hydrogen, Methane, Methanomicrobiales, Temperature}, issn = {1095-9203}, doi = {10.1126/science.aaa4326}, author = {Wang, David T and Gruen, Danielle S and Lollar, Barbara Sherwood and Hinrichs, Kai-Uwe and Stewart, Lucy C and Holden, James F and Hristov, Alexander N and Pohlman, John W and Morrill, Penny L and K{\"o}nneke, Martin and Delwiche, Kyle B and Reeves, Eoghan P and Sutcliffe, Chelsea N and Ritter, Daniel J and Seewald, Jeffrey S and McIntosh, Jennifer C and Hemond, Harold F and Kubo, Michael D and Cardace, Dawn and Hoehler, Tori M and Ono, Shuhei} } @article {1743, title = {Methanocaldococcus bathoardescens sp. nov., a hyperthermophilic methanogen isolated from a volcanically active deep-sea hydrothermal vent.}, journal = {Int J Syst Evol Microbiol}, volume = {65}, year = {2015}, month = {2015 Apr}, pages = {1280-3}, abstract = {A hyperthermophilic methanogen, strain JH146(T), was isolated from 26 {\textdegree}C hydrothermal vent fluid emanating from a crack in basaltic rock at Marker 113 vent, Axial Seamount in the northeastern Pacific Ocean. It was identified as an obligate anaerobe that uses only H2 and CO2 for growth. Phylogenetic analysis based on 16S rRNA gene sequences showed that the strain is more than 97\% similar to other species of the genus Methanocaldococcus . Therefore, overall genome relatedness index analyses were performed to establish that strain JH146(T) represents a novel species. For each analysis, strain JH146(T) was most similar to Methanocaldococcus sp. FS406-22, which can fix N2 and also comes from Marker 113 vent. However, strain JH146(T) differs from strain FS406-22 in that it cannot fix N2. The average nucleotide identity score for strain JH146(T) was 87\%, the genome-to-genome direct comparison score was 33-55\% and the species identification score was 93\%. For each analysis, strain JH146(T) was below the species delineation cut-off. Full-genome gene synteny analysis showed that strain JH146(T) and strain FS406-22 have 97\% genome synteny, but strain JH146(T) was missing the operons necessary for N2 fixation and assimilatory nitrate reduction that are present in strain FS406-22. Based on its whole genome sequence, strain JH146(T) is suggested to represent a novel species of the genus Methanocaldococcus for which the name Methanocaldococcus bathoardescens is proposed. The type strain is JH146(T) ( = DSM 27223(T) = KACC 18232(T)).
}, keywords = {DNA, Bacterial, Hydrothermal Vents, Methanocaldococcus, Molecular Sequence Data, Pacific Ocean, Phylogeny, RNA, Ribosomal, 16S, Seawater, Sequence Analysis, DNA}, issn = {1466-5034}, doi = {10.1099/ijs.0.000097}, author = {Stewart, Lucy C and Jung, Jong-Hyun and Kim, You-Tae and Kwon, Soon-Wo and Park, Cheon-Seok and Holden, James F} } @article {1414, title = {Mitochondrial heat shock protein machinery hsp70/hsp40 is indispensable for proper mitochondrial DNA maintenance and replication.}, journal = {MBio}, volume = {6}, year = {2015}, month = {2015}, abstract = {UNLABELLED: Mitochondrial chaperones have multiple functions that are essential for proper functioning of mitochondria. In the human-pathogenic protist Trypanosoma brucei, we demonstrate a novel function of the highly conserved machinery composed of mitochondrial heat shock proteins 70 and 40 (mtHsp70/mtHsp40) and the ATP exchange factor Mge1. The mitochondrial DNA of T. brucei, also known as kinetoplast DNA (kDNA), is represented by a single catenated network composed of thousands of minicircles and dozens of maxicircles packed into an electron-dense kDNA disk. The chaperones mtHsp70 and mtHsp40 and their cofactor Mge1 are uniformly distributed throughout the single mitochondrial network and are all essential for the parasite. Following RNA interference (RNAi)-mediated depletion of each of these proteins, the kDNA network shrinks and eventually disappears. Ultrastructural analysis of cells depleted for mtHsp70 or mtHsp40 revealed that the otherwise compact kDNA network becomes severely compromised, a consequence of decreased maxicircle and minicircle copy numbers. Moreover, we show that the replication of minicircles is impaired, although the lack of these proteins has a bigger impact on the less abundant maxicircles. We provide additional evidence that these chaperones are indispensable for the maintenance and replication of kDNA, in addition to their already known functions in Fe-S cluster synthesis and protein import.
IMPORTANCE: Impairment or loss of mitochondrial DNA is associated with mitochondrial dysfunction and a wide range of neural, muscular, and other diseases. We present the first evidence showing that the entire mtHsp70/mtHsp40 machinery plays an important role in mitochondrial DNA replication and maintenance, a function likely retained from prokaryotes. These abundant, ubiquitous, and multifunctional chaperones share phenotypes with enzymes engaged in the initial stages of replication of the mitochondrial DNA in T. brucei.
}, issn = {2150-7511}, doi = {10.1128/mBio.02425-14}, author = {T{\'y}{\v c}, Ji{\v r}{\'\i} and Klingbeil, Michele M and Luke{\v s}, Julius} } @article {1425, title = {Novel mechanisms of controlling the activities of the transcription factors Spo0A and ComA by the plasmid-encoded quorum sensing regulators Rap60-Phr60 in Bacillus subtilis.}, journal = {Mol Microbiol}, year = {2015}, month = {2015 Jan 19}, abstract = {Bacillus subtilis and its closest relatives have multiple rap-phr quorum sensing gene pairs that coordinate a variety of physiological processes with population density. Extra-chromosomal rap-phr genes are also present on mobile genetic elements, yet relatively little is known about their function. In this work, we demonstrate that Rap60-Phr60 from plasmid pTA1060 coordinates a variety of biological processes with population density including sporulation, cannibalism, biofilm formation and genetic competence. Similar to other Rap proteins that control sporulation, Rap60 modulates phosphorylation of the transcription factor Spo0A by acting as a phosphatase of Spo0F\∼P, an intermediate of the sporulation phosphorelay system. Additionally, Rap60 plays a noncanonical role in regulating the autophosphorylation of the sporulation-specific kinase KinA, a novel activity for Rap proteins. In contrast, Rap proteins that modulate genetic competence interfere with DNA binding by the transcription factor ComA. Rap60 regulates the activity of ComA in a unique manner by forming a Rap60-ComA-DNA ternary complex that inhibits transcription of target genes. Taken together, this work provides new insight into two novel mechanisms of regulating Spo0A and ComA by Rap60 and expands our general understanding of how plasmid-encoded quorum sensing pairs regulate important biological processes.
}, issn = {1365-2958}, doi = {10.1111/mmi.12939}, author = {Boguslawski, Kristina M and Hill, Patrick A and Griffith, Kevin L} } @article {2374, title = {Orientation of DNA Minicircles Balances Density and Topological Complexity in Kinetoplast DNA.}, journal = {PLoS One}, volume = {10}, year = {2015}, month = {2015}, pages = {e0130998}, abstract = {Kinetoplast DNA (kDNA), a unique mitochondrial structure common to trypanosomatid parasites, contains thousands of DNA minicircles that are densely packed and can be topologically linked into a chain mail-like network. Experimental data indicate that every minicircle in the network is, on average, singly linked to three other minicircles (i.e., has mean valence 3) before replication and to six minicircles in the late stages of replication. The biophysical factors that determine the topology of the network and its changes during the cell cycle remain unknown. Using a mathematical modeling approach, we previously showed that volume confinement alone can drive the formation of the network and that it induces a linear relationship between mean valence and minicircle density. Our modeling also predicted a minicircle valence two orders of magnitude greater than that observed in kDNA. To determine the factors that contribute to this discrepancy we systematically analyzed the relationship between the topological properties of the network (i.e., minicircle density and mean valence) and its biophysical properties such as DNA bending, electrostatic repulsion, and minicircle relative position and orientation. Significantly, our results showed that most of the discrepancy between the theoretical and experimental observations can be accounted for by the orientation of the minicircles with volume exclusion due to electrostatic interactions and DNA bending playing smaller roles. Our results are in agreement with the three dimensional kDNA organization model, initially proposed by Delain and Riou, in which minicircles are oriented almost perpendicular to the horizontal plane of the kDNA disk. We suggest that while minicircle confinement drives the formation of kDNA networks, it is minicircle orientation that regulates the topological complexity of the network.
}, keywords = {Cell Cycle, Crithidia fasciculata, DNA Replication, DNA, Kinetoplast, DNA, Mitochondrial}, issn = {1932-6203}, doi = {10.1371/journal.pone.0130998}, author = {Diao, Yuanan and Rodriguez, Victor and Klingbeil, Michele and Arsuaga, Javier} } @article {1457, title = {A pleiotropic drug resistance transporter is involved in reduced sensitivity to multiple fungicide classes in Sclerotinia homoeocarpa (F.T. Bennett).}, journal = {Mol Plant Pathol}, volume = {16}, year = {2015}, month = {2015 Apr}, pages = {251-61}, abstract = {Dollar spot, caused by Sclerotinia homoeocarpa, is a prevalent turfgrass disease, and the fungus exhibits widespread fungicide resistance in North America. In a previous study, an ABC-G transporter, ShatrD, was associated with practical field resistance to demethylation inhibitor (DMI) fungicides. Mining of ABC-G transporters, also known as pleiotropic drug resistance (PDR) transporters, from RNA-Seq data gave an assortment of transcripts, several with high sequence similarity to functionally characterized transporters from Botrytis cinerea, and others with closest blastx hits from Aspergillus and Monilinia. In addition to ShatrD, another PDR transporter showed significant over-expression in replicated RNA-Seq data, and in a collection of field-resistant isolates, as measured by quantitative polymerase chain reaction. These isolates also showed reduced sensitivity to unrelated fungicide classes. Using a yeast complementation system, we sought to test the hypothesis that this PDR transporter effluxes DMI as well as chemically unrelated fungicides. The transporter (ShPDR1) was cloned into the Gal1 expression vector and transformed into a yeast PDR transporter deletion mutant, AD12345678. Complementation assays indicated that ShPDR1 complemented the mutant in the presence of propiconazole (DMI), iprodione (dicarboximide) and boscalid (SDHI, succinate dehydrogenase inhibitor). Our results indicate that the over-expression of ShPDR1 is correlated with practical field resistance to DMI fungicides and reduced sensitivity to dicarboximide and SDHI fungicides. These findings highlight the potential for the eventual development of a multidrug resistance phenotype in this pathogen. In addition, this study presents a pipeline for the discovery and validation of fungicide resistance genes using de\ novo next-generation sequencing and molecular biology techniques in an unsequenced plant pathogenic fungus.
}, issn = {1364-3703}, doi = {10.1111/mpp.12174}, author = {Sang, Hyunkyu and Hulvey, Jon and Popko, James T and Lopes, John and Swaminathan, Aishwarya and Chang, Taehyun and Jung, Geunhwa} } @article {3107, title = {Protozoan grazing reduces the current output of microbial fuel cells.}, journal = {Bioresour Technol}, volume = {193}, year = {2015}, month = {2015 Oct}, pages = {8-14}, abstract = {Several experiments were conducted to determine whether protozoan grazing can reduce current output from sediment microbial fuel cells. When marine sediments were amended with eukaryotic inhibitors, the power output from the fuel cells increased 2-5-fold. Quantitative PCR showed that Geobacteraceae sequences were 120 times more abundant on anodes from treated fuel cells compared to untreated fuel cells, and that Spirotrichea sequences in untreated fuel cells were 200 times more abundant on anode surfaces than in the surrounding sediments. Defined studies with current-producing biofilms of Geobacter sulfurreducens and pure cultures of protozoa demonstrated that protozoa that were effective in consuming G. sulfurreducens reduced current production up to 91\% when added to G. sulfurreducens fuel cells. These results suggest that anode biofilms are an attractive food source for protozoa and that protozoan grazing can be an important factor limiting the current output of sediment microbial fuel cells.
}, keywords = {Bioelectric Energy Sources, Biofilms, Electricity, Electrodes, Eukaryota, Geobacter, Geologic Sediments}, issn = {1873-2976}, doi = {10.1016/j.biortech.2015.06.056}, author = {Holmes, Dawn E and Nevin, Kelly P and Snoeyenbos-West, Oona L and Woodard, Trevor L and Strickland, Justin N and Lovley, Derek R} } @article {1622, title = {Regulation of Clostridium difficile Spore Formation by the SpoIIQ and SpoIIIA Proteins.}, journal = {PLoS Genet}, volume = {11}, year = {2015}, month = {2015 Oct}, pages = {e1005562}, abstract = {Sporulation is an ancient developmental process that involves the formation of a highly resistant endospore within a larger mother cell. In the model organism Bacillus subtilis, sporulation-specific sigma factors activate compartment-specific transcriptional programs that drive spore morphogenesis. \σG activity in the forespore depends on the formation of a secretion complex, known as the \"feeding tube,\" that bridges the mother cell and forespore and maintains forespore integrity. Even though these channel components are conserved in all spore formers, recent studies in the major nosocomial pathogen Clostridium difficile suggested that these components are dispensable for \σG activity. In this study, we investigated the requirements of the SpoIIQ and SpoIIIA proteins during C. difficile sporulation. C. difficile spoIIQ, spoIIIA, and spoIIIAH mutants exhibited defects in engulfment, tethering of coat to the forespore, and heat-resistant spore formation, even though they activate \σG at wildtype levels. Although the spoIIQ, spoIIIA, and spoIIIAH mutants were defective in engulfment, metabolic labeling studies revealed that they nevertheless actively transformed the peptidoglycan at the leading edge of engulfment. In vitro pull-down assays further demonstrated that C. difficile SpoIIQ directly interacts with SpoIIIAH. Interestingly, mutation of the conserved Walker A ATP binding motif, but not the Walker B ATP hydrolysis motif, disrupted SpoIIIAA function during C. difficile spore formation. This finding contrasts with B. subtilis, which requires both Walker A and B motifs for SpoIIIAA function. Taken together, our findings suggest that inhibiting SpoIIQ, SpoIIIAA, or SpoIIIAH function could prevent the formation of infectious C. difficile spores and thus disease transmission.
}, issn = {1553-7404}, doi = {10.1371/journal.pgen.1005562}, author = {Fimlaid, Kelly A and Jensen, Owen and Donnelly, M Lauren and Siegrist, M Sloan and Shen, Aimee} } @article {1558, title = {Response to Comment on "Environmental Fate of the Next Generation Refrigerant 2,3,3,3-Tetrafluoropropene (HFO-1234yf)".}, journal = {Environ Sci Technol}, volume = {49}, year = {2015}, month = {2015 Jul 7}, pages = {8265-6}, issn = {1520-5851}, doi = {10.1021/acs.est.5b01970}, author = {Im, Jeongdae and Walshe-Langford, Gillian E and Moon, Ji-Won and L{\"o}ffler, Frank E} } @article {1964, title = {A Ribonucleoprotein Complex Protects the Interleukin-6 mRNA from Degradation by Distinct Herpesviral Endonucleases}, journal = {PLOS Pathogens}, volume = {11}, year = {2015}, month = {may}, pages = {e1004899}, doi = {10.1371/journal.ppat.1004899}, url = {https://doi.org/10.1371/journal.ppat.1004899}, author = {Mandy Muller and Stephanie Hutin and Oliver Marigold and Kathy H. Li and Al Burlingame and Britt A. Glaunsinger}, editor = {Nicholas K Conrad} } @article {3114, title = {Seeing is believing: novel imaging techniques help clarify microbial nanowire structure and function.}, journal = {Environ Microbiol}, volume = {17}, year = {2015}, month = {2015 Jul}, pages = {2209-15}, abstract = {Novel imaging approaches have recently helped to clarify the properties of {\textquoteright}microbial nanowires{\textquoteright}. Geobacter sulfurreducens pili are actual wires. They possess metallic-like conductivity, which can be attributed to overlapping pi-pi orbitals of key aromatic amino acids. Electrostatic force microscopy recently confirmed charge propagation along the pili, in a manner similar to carbon nanotubes. The pili are essential for long-range electron transport to insoluble electron acceptors and interspecies electron transfer. Previous claims that Shewanella oneidensis also produce conductive pili have recently been recanted, based on novel live-imaging studies. The putative pili are, in fact, long extensions of the cytochrome-rich outer membrane and periplasm that, when dried, collapse to form filaments with dimensions similar to pili. It has yet to be demonstrated whether the cytochrome-to-cytochrome electron hopping documented in the dried membrane extensions takes place in intact hydrated membrane extensions or whether the membrane extensions enhance electron transport to insoluble electron acceptors such as Fe(III) oxides or electrodes. These findings demonstrate that G. sulfurreducens conductive pili and the outer membrane extensions of S. oneidensis are fundamentally different in composition, mechanism of electron transport and physiological role. New methods for evaluating filament conductivity will facilitate screening the microbial world for nanowires and elucidating their function.
}, keywords = {Cytochromes, Electric Conductivity, Electron Transport, Electrons, Ferric Compounds, Fimbriae, Bacterial, Geobacter, Microscopy, Atomic Force, Nanowires, Oxides, Periplasm, Shewanella}, issn = {1462-2920}, doi = {10.1111/1462-2920.12708}, author = {Lovley, Derek R and Malvankar, Nikhil S} } @article {3108, title = {Simplifying microbial electrosynthesis reactor design.}, journal = {Front Microbiol}, volume = {6}, year = {2015}, month = {2015}, pages = {468}, abstract = {Microbial electrosynthesis, an artificial form of photosynthesis, can efficiently convert carbon dioxide into organic commodities; however, this process has only previously been demonstrated in reactors that have features likely to be a barrier to scale-up. Therefore, the possibility of simplifying reactor design by both eliminating potentiostatic control of the cathode and removing the membrane separating the anode and cathode was investigated with biofilms of Sporomusa ovata. S. ovata reduces carbon dioxide to acetate and acts as the microbial catalyst for plain graphite stick cathodes as the electron donor. In traditional {\textquoteright}H-cell{\textquoteright} reactors, where the anode and cathode chambers were separated with a proton-selective membrane, the rates and columbic efficiencies of microbial electrosynthesis remained high when electron delivery at the cathode was powered with a direct current power source rather than with a potentiostat-poised cathode utilized in previous studies. A membrane-less reactor with a direct-current power source with the cathode and anode positioned to avoid oxygen exposure at the cathode, retained high rates of acetate production as well as high columbic and energetic efficiencies. The finding that microbial electrosynthesis is feasible without a membrane separating the anode from the cathode, coupled with a direct current power source supplying the energy for electron delivery, is expected to greatly simplify future reactor design and lower construction costs.
}, issn = {1664-302X}, doi = {10.3389/fmicb.2015.00468}, author = {Giddings, Cloelle G S and Nevin, Kelly P and Woodward, Trevor and Lovley, Derek R and Butler, Caitlyn S} } @article {3111, title = {Structural basis for metallic-like conductivity in microbial nanowires.}, journal = {mBio}, volume = {6}, year = {2015}, month = {2015 Mar 03}, pages = {e00084}, abstract = {UNLABELLED: Direct measurement of multiple physical properties of Geobacter sulfurreducens pili have demonstrated that they possess metallic-like conductivity, but several studies have suggested that metallic-like conductivity is unlikely based on the structures of the G.~sulfurreducens pilus predicted from homology models. In order to further evaluate this discrepancy, pili were examined with synchrotron X-ray microdiffraction and rocking-curve X-ray diffraction. Both techniques revealed a periodic 3.2-{\r A} spacing in conductive, wild-type G.~sulfurreducens pili that was missing in the nonconductive pili of strain Aro5, which lack key aromatic acids required for conductivity. The intensity of the 3.2-{\r A} peak increased 100-fold when the pH was shifted from 10.5 to 2, corresponding with a previously reported 100-fold increase in pilus conductivity with this pH change. These results suggest a clear structure-function correlation for metallic-like conductivity that can be attributed to overlapping π-orbitals of aromatic amino acids. A homology model of the G.~sulfurreducens pilus was constructed with a Pseudomonas aeruginosa pilus model as a template as an alternative to previous models, which were based on a Neisseria gonorrhoeae pilus structure. This alternative model predicted that aromatic amino acids in G.~sulfurreducens pili are packed within 3 to 4~{\r A}, consistent with the experimental results. Thus, the predictions of homology modeling are highly sensitive to assumptions inherent in the model construction. The experimental results reported here further support the concept that the pili of G.~sulfurreducens represent a novel class of electronically functional proteins in which aromatic amino acids promote long-distance electron transport.
IMPORTANCE: The mechanism for long-range electron transport along the conductive pili of Geobacter sulfurreducens is of interest because these "microbial nanowires" are important in biogeochemical cycling as well as applications in bioenergy and bioelectronics. Although proteins are typically insulators, G.~sulfurreducens pilus proteins possess metallic-like conductivity. The studies reported here provide important structural insights into the mechanism of the metallic-like conductivity of G.~sulfurreducens pili. This information is expected to be useful in the design of novel bioelectronic materials.
}, keywords = {Amino Acids, Aromatic, Chemical Phenomena, Electrophysiological Phenomena, Fimbriae, Bacterial, Geobacter, Hydrogen-Ion Concentration, Models, Molecular, Nanowires, X-Ray Diffraction}, issn = {2150-7511}, doi = {10.1128/mBio.00084-15}, author = {Malvankar, Nikhil S and Vargas, Madeline and Nevin, Kelly and Tremblay, Pier-Luc and Evans-Lutterodt, Kenneth and Nykypanchuk, Dmytro and Martz, Eric and Tuominen, Mark T and Lovley, Derek R} } @article {3110, title = {Syntrophic growth via quinone-mediated interspecies electron transfer.}, journal = {Front Microbiol}, volume = {6}, year = {2015}, month = {2015}, pages = {121}, abstract = {The mechanisms by which microbial species exchange electrons are of interest because interspecies electron transfer can expand the metabolic capabilities of microbial communities. Previous studies with the humic substance analog anthraquinone-2,6-disulfonate (AQDS) suggested that quinone-mediated interspecies electron transfer (QUIET) is feasible, but it was not determined if sufficient energy is available from QUIET to support the growth of both species. Furthermore, there have been no previous studies on the mechanisms for the oxidation of anthrahydroquinone-2,6-disulfonate (AHQDS). A co-culture of Geobacter metallireducens and G. sulfurreducens metabolized ethanol with the reduction of fumarate much faster in the presence of AQDS, and there was an increase in cell protein. G. sulfurreducens was more abundant, consistent with G. sulfurreducens obtaining electrons from acetate that G. metallireducens produced from ethanol, as well as from AHQDS. Co-cultures initiated with a citrate synthase-deficient strain of G. sulfurreducens that was unable to use acetate as an electron donor also metabolized ethanol with the reduction of fumarate and cell growth, but acetate accumulated over time. G. sulfurreducens and G. metallireducens were equally abundant in these co-cultures reflecting the inability of the citrate synthase-deficient strain of G. sulfurreducens to metabolize acetate. Evaluation of the mechanisms by which G. sulfurreducens accepts electrons from AHQDS demonstrated that a strain deficient in outer-surface c-type cytochromes that are required for AQDS reduction was as effective at QUIET as the wild-type strain. Deletion of additional genes previously implicated in extracellular electron transfer also had no impact on QUIET. These results demonstrate that QUIET can yield sufficient energy to support the growth of both syntrophic partners, but that the mechanisms by which electrons are derived from extracellular hydroquinones require further investigation.
}, issn = {1664-302X}, doi = {10.3389/fmicb.2015.00121}, author = {Smith, Jessica A and Nevin, Kelly P and Lovley, Derek R} } @article {1438, title = {Variation in diazotrophic community structure in forest soils reflects land use history.}, journal = {Soil Biology and Biochemistry}, volume = {80}, year = {2015}, month = {2014 Sep}, pages = {1-8}, doi = {10.1016/j.soilbio.2014.09.017}, url = {http://www.sciencedirect.com/science/article/pii/S0038071714003253}, author = {N{\"u}sslein, K and Izquierdo, J} } @article {1561, title = {4-methylphenol produced in freshwater sediment microcosms is not a bisphenol A metabolite.}, journal = {Chemosphere}, volume = {117}, year = {2014}, month = {2014 Dec}, pages = {521-6}, abstract = {4-Methylphenol (4-MP), a putative bisphenol A (BPA) degradation intermediate, was detected at concentrations reaching 2.1 mg L(-1) in anoxic microcosms containing 10 mg L(-1) BPA and 5 g of freshwater sediment material collected from four geographically distinct locations and amended with nitrate, nitrite, ferric iron, or bicarbonate as electron acceptors. 4-MP accumulation was transient, and 4-MP degradation was observed under all redox conditions tested. 4-MP was not detected in microcosms not amended with BPA. Unexpectedly, incubations with (13)C-labeled BPA failed to produce (13)C-labeled 4-MP suggesting that 4-MP was not derived from BPA. The detection of 4-MP in live microcosms amended with lactate, but not containing BPA corroborated that BPA was not the source of 4-MP. These findings demonstrate that the transient formation of 4-MP as a possible BPA degradation intermediate must be interpreted cautiously, as microbial activity in streambed microcosms may generate 4-MP from sediment-associated organic material.
}, keywords = {Anaerobiosis, Benzhydryl Compounds, Biodegradation, Environmental, Chromatography, High Pressure Liquid, Cresols, Environmental Monitoring, Fresh Water, Gas Chromatography-Mass Spectrometry, Geologic Sediments, Phenols, Water Pollutants, Chemical}, issn = {1879-1298}, doi = {10.1016/j.chemosphere.2014.09.008}, author = {Im, Jeongdae and Prevatte, Carson W and Lee, Hong Geun and Campagna, Shawn R and L{\"o}ffler, Frank E} } @article {1421, title = {Biodegradation of low concentrations of 1,2-dibromoethane in groundwater is enhanced by phenol.}, journal = {Appl Microbiol Biotechnol}, volume = {98}, year = {2014}, month = {2014 Feb}, pages = {1329-38}, abstract = {The lead scavenger 1,2-dibromoethane (EDB), a former additive to leaded gasoline, is a common groundwater contaminant, yet not much knowledge is available for its targeted bioremediation, especially under in situ conditions. The study site was an aviation gas spill site, which, although all hydrocarbons and most of the EDB were remediated in the mid-1990s, still exhibits low levels of EDB remaining in the groundwater (about 11 \μg EDB/l). To evaluate the effect of phenol on biostimulation of low concentration of EDB, microcosms were established from an EDB-contaminated aquifer. After 300 days at environmentally relevant conditions (12\ \±\ 2 \°C, static incubation), EDB was not significantly removed from unamended microcosms compared to the abiotic control. However, in treatments amended with phenol, up to 80 \% of the initial EDB concentration had been degraded, while added phenol was removed completely. Microbial community composition in unamended and phenol-amended microcosms remained unchanged, and Polaromonas sp. dominated both types of microcosms, but total bacterial abundance and numbers of the gene for phenol hydroxylase were higher in phenol-amended microcosms. Dehalogenase, an indicator suggesting targeted aerobic biodegradation of EDB, was not detected in either treatment. This finding suggests phenol hydroxylase, rather than a dehalogenation reaction, may be responsible for 1,2-dibromoethane oxidation under in situ conditions. In addition, biostimulation of EDB is possible through the addition of low levels of phenol in aerobic groundwater sites.
}, keywords = {Bacteria, Biota, Ethylene Dibromide, Groundwater, Metabolic Networks and Pathways, Phenol, Water Pollutants}, issn = {1432-0614}, doi = {10.1007/s00253-013-4963-1}, author = {Baek, Kyunghwa and Wang, Meng and McKeever, Robert and Rieber, Kahlil and Park, Chul and N{\"u}sslein, Klaus} } @article {1413, title = {Bioinformatic and biochemical analysis of a novel maltose-forming α-amylase of the GH57 family in the hyperthermophilic archaeon Thermococcus sp. CL1.}, journal = {Enzyme Microb Technol}, volume = {60}, year = {2014}, month = {2014 Jun 10}, pages = {9-15}, abstract = {Maltose-forming \α-amylase is a glycoside hydrolase family 57 (GH57) member that is unique because it displays dual hydrolysis activity toward \α-1,4- and \α-1,6-glycosidic linkages and only recognizes maltose. This enzyme was previously identified only in Pyrococcus sp. ST04 (PSMA); however, we recently found two homologs subgroups in Thermococcus species. One subgroup (subgroup A) showed relatively high amino acid sequence similarity to PSMA (\>71\%), while the other subgroup (subgroup B) showed lower homology with PSMA (\<59\%). To characterize the subgroup B maltose-forming \α-amylase from Thermococcus species (TCMA), we cloned the CL1_0868 gene from Thermococcus sp. CL1 and then successfully expressed the gene in Escherichia coli. Although TCMA has a different oligomeric state relative to PSMA, TCMA showed similar substrate specificity. However, TCMA was shown to hydrolyze maltooligosaccharides more easily than PSMA. Also, TCMA displayed different optimum conditions depending on the glycosidic linkage of the substrate. TCMA had the highest activity at 85\°C and at pH 5.0 for \α-1,4-glycosidic linkage hydrolysis whereas it showed its maximal activity to cleave \α-1,6-glycosidic linkages at 98\°C and pH 6.0.
}, issn = {1879-0909}, doi = {10.1016/j.enzmictec.2014.03.009}, author = {Jeon, Eun-Jung and Jung, Jong-Hyun and Seo, Dong-Ho and Jung, Dong-Hyun and Holden, James F and Park, Cheon-Seok} } @article {3117, title = {Carbon cloth stimulates direct interspecies electron transfer in syntrophic co-cultures.}, journal = {Bioresour Technol}, volume = {173}, year = {2014}, month = {2014 Dec}, pages = {82-86}, abstract = {This study investigated the possibility that the electrical conductivity of carbon cloth accelerates direct interspecies electron transfer (DIET) in co-cultures. Carbon cloth accelerated metabolism of DIET co-cultures (Geobacter metallireducens-Geobacter sulfurreducens and G.metallireducens-Methanosarcina barkeri) but did not promote metabolism of co-cultures performing interspecies H2 transfer (Desulfovibrio vulgaris-G.sulfurreducens). On the other hand, DIET co-cultures were not stimulated by poorly conductive cotton cloth. Mutant strains lacking electrically conductive pili, or pili-associated cytochromes participated in DIET only in the presence of carbon cloth. In co-cultures promoted by carbon cloth, cells were primarily associated with the cloth although the syntrophic partners were too far apart for cell-to-cell biological electrical connections to be feasible. Carbon cloth seemingly mediated interspecies electron transfer between the distant syntrophic partners. These results suggest that the ability of carbon cloth to accelerate DIET should be considered in anaerobic digester designs that incorporate carbon cloth.
}, keywords = {Carbon, Cell Communication, Coculture Techniques, Electric Conductivity, Electron Transport, Geobacter, Materials Testing, Membranes, Artificial, Microbial Consortia, Oxidation-Reduction, Symbiosis}, issn = {1873-2976}, doi = {10.1016/j.biortech.2014.09.009}, author = {Chen, Shanshan and Rotaru, Amelia-Elena and Liu, Fanghua and Philips, Jo and Woodard, Trevor L and Nevin, Kelly P and Lovley, Derek R} } @article {1965, title = {CCHCR1 Interacts Specifically with the E2 Protein of Human Papillomavirus Type 16 on a Surface Overlapping BRD4 Binding}, journal = {PLoS ONE}, volume = {9}, year = {2014}, month = {mar}, pages = {e92581}, doi = {10.1371/journal.pone.0092581}, url = {https://doi.org/10.1371/journal.pone.0092581}, author = {Mandy Muller and Caroline Demeret}, editor = {Lawrence Banks} } @article {1200, title = {Complete genome sequence of hyperthermophilic archaeon Thermococcus sp. ES1.}, journal = {J Biotechnol}, volume = {174C}, year = {2014}, month = {2014 Jan 25}, pages = {14-15}, abstract = {Thermococcus sp. strain ES1 is an anaerobic, hyperthermophilic archaeon from a hydrothermal vent that catabolizes sugars and peptides and produces H2S from S{\textdegree}, H2, acetate and CO2 as its primary metabolites. We present the complete genome sequence of this strain (1,957,742bp) with a focus on its substrate utilization and metabolite production capabilities. The sequence will contribute to the development of heterotrophic archaea for bioenergy production and biogeochemical modeling in hydrothermal environments.
}, issn = {1873-4863}, doi = {10.1016/j.jbiotec.2014.01.022}, author = {Jung, Jong-Hyun and Kim, You-Tae and Jeon, Eun-Jung and Seo, Dong-Ho and Hensley, Sarah A and Holden, James F and Lee, Ju-Hoon and Park, Cheon-Seok} } @article {3127, title = {Constraint-based modeling of carbon fixation and the energetics of electron transfer in Geobacter metallireducens.}, journal = {PLoS Comput Biol}, volume = {10}, year = {2014}, month = {2014 Apr}, pages = {e1003575}, abstract = {Geobacter species are of great interest for environmental and biotechnology applications as they can carry out direct electron transfer to insoluble metals or other microorganisms and have the ability to assimilate inorganic carbon. Here, we report on the capability and key enabling metabolic machinery of Geobacter metallireducens GS-15 to carry out CO2 fixation and direct electron transfer to iron. An updated metabolic reconstruction was generated, growth screens on targeted conditions of interest were performed, and constraint-based analysis was utilized to characterize and evaluate critical pathways and reactions in G. metallireducens. The novel capability of G. metallireducens to grow autotrophically with formate and Fe(III) was predicted and subsequently validated in vivo. Additionally, the energetic cost of transferring electrons to an external electron acceptor was determined through analysis of growth experiments carried out using three different electron acceptors (Fe(III), nitrate, and fumarate) by systematically isolating and examining different parts of the electron transport chain. The updated reconstruction will serve as a knowledgebase for understanding and engineering Geobacter and similar species.
}, keywords = {Carbon, Electron Transport, Energy Metabolism, Genome, Bacterial, Geobacter, Models, Biological}, issn = {1553-7358}, doi = {10.1371/journal.pcbi.1003575}, author = {Feist, Adam M and Nagarajan, Harish and Rotaru, Amelia-Elena and Tremblay, Pier-Luc and Zhang, Tian and Nevin, Kelly P and Lovley, Derek R and Zengler, Karsten} } @article {3115, title = {Converting carbon dioxide to butyrate with an engineered strain of Clostridium ljungdahlii.}, journal = {mBio}, volume = {5}, year = {2014}, month = {2014 Oct 21}, pages = {e01636-14}, abstract = {Microbial conversion of carbon dioxide to organic commodities via syngas metabolism or microbial electrosynthesis is an attractive option for production of renewable biocommodities. The recent development of an initial genetic toolbox for the acetogen Clostridium ljungdahlii has suggested that C. ljungdahlii may be an effective chassis for such conversions. This possibility was evaluated by engineering a strain to produce butyrate, a valuable commodity that is not a natural product of C. ljungdahlii metabolism. Heterologous genes required for butyrate production from acetyl-coenzyme A (CoA) were identified and introduced initially on plasmids and in subsequent strain designs integrated into the C. ljungdahlii chromosome. Iterative strain designs involved increasing translation of a key enzyme by modifying a ribosome binding site, inactivating the gene encoding the first step in the conversion of acetyl-CoA to acetate, disrupting the gene which encodes the primary bifunctional aldehyde/alcohol dehydrogenase for ethanol production, and interrupting the gene for a CoA transferase that potentially represented an alternative route for the production of acetate. These modifications yielded a strain in which ca. 50 or 70\% of the carbon and electron flow was diverted to the production of butyrate with H2 or CO as the electron donor, respectively. These results demonstrate the possibility of producing high-value commodities from carbon dioxide with C. ljungdahlii as the catalyst. Importance: The development of a microbial chassis for efficient conversion of carbon dioxide directly to desired organic products would greatly advance the environmentally sustainable production of biofuels and other commodities. Clostridium ljungdahlii is an effective catalyst for microbial electrosynthesis, a technology in which electricity generated with renewable technologies, such as solar or wind, powers the conversion of carbon dioxide and water to organic products. Other electron donors for C. ljungdahlii include carbon monoxide, which can be derived from industrial waste gases or the conversion of recalcitrant biomass to syngas, as well as hydrogen, another syngas component. The finding that carbon and electron flow in C. ljungdahlii can be diverted from the production of acetate to butyrate synthesis is an important step toward the goal of renewable commodity production from carbon dioxide with this organism.
}, keywords = {Acetyl Coenzyme A, Butyrates, Carbon Dioxide, Clostridium, Metabolic Engineering, Metabolic Flux Analysis, Metabolic Networks and Pathways, Recombinant Proteins}, issn = {2150-7511}, doi = {10.1128/mBio.01636-14}, author = {Ueki, Toshiyuki and Nevin, Kelly P and Woodard, Trevor L and Lovley, Derek R} } @article {3113, title = {Correlation between microbial community and granule conductivity in anaerobic bioreactors for brewery wastewater treatment.}, journal = {Bioresour Technol}, volume = {174}, year = {2014}, month = {2014 Dec}, pages = {306-10}, abstract = {Prior investigation of an upflow anaerobic sludge blanket (UASB) reactor treating brewery wastes suggested that direct interspecies electron transfer (DIET) significantly contributed to interspecies electron transfer to methanogens. To investigate DIET in granules further, the electrical conductivity and bacterial community composition of granules in fourteen samples from four different UASB reactors treating brewery wastes were investigated. All of the UASB granules were electrically conductive whereas control granules from ANAMMOX (ANaerobic AMMonium OXidation) reactors and microbial granules from an aerobic bioreactor designed for phosphate removal were not. There was a moderate correlation (r=0.67) between the abundance of Geobacter species in the UASB granules and granule conductivity, suggesting that Geobacter contributed to granule conductivity. These results, coupled with previous studies, which have demonstrated that Geobacter species can donate electrons to methanogens that are typically predominant in anaerobic digesters, suggest that DIET may be a widespread phenomenon in UASB reactors treating brewery wastes.
}, keywords = {Alcoholic Beverages, Anaerobiosis, Bacteria, Bioreactors, Electric Conductivity, Ethanol, Sequence Analysis, DNA, Sewage, Waste Disposal, Fluid, Wastewater, Water Purification}, issn = {1873-2976}, doi = {10.1016/j.biortech.2014.10.004}, author = {Shrestha, Pravin Malla and Malvankar, Nikhil S and Werner, Jeffrey J and Franks, Ashley E and Elena-Rotaru, Amelia and Shrestha, Minita and Liu, Fanghua and Nevin, Kelly P and Angenent, Largus T and Lovley, Derek R} } @article {3125, title = {Direct interspecies electron transfer between Geobacter metallireducens and Methanosarcina barkeri.}, journal = {Appl Environ Microbiol}, volume = {80}, year = {2014}, month = {2014 Aug}, pages = {4599-605}, abstract = {Direct interspecies electron transfer (DIET) is potentially an effective form of syntrophy in methanogenic communities, but little is known about the diversity of methanogens capable of DIET. The ability of Methanosarcina barkeri to participate in DIET was evaluated in coculture with Geobacter metallireducens. Cocultures formed aggregates that shared electrons via DIET during the stoichiometric conversion of ethanol to methane. Cocultures could not be initiated with a pilin-deficient G. metallireducens strain, suggesting that long-range electron transfer along pili was important for DIET. Amendments of granular activated carbon permitted the pilin-deficient G. metallireducens isolates to share electrons with M. barkeri, demonstrating that this conductive material could substitute for pili in promoting DIET. When M. barkeri was grown in coculture with the H2-producing Pelobacter carbinolicus, incapable of DIET, M. barkeri utilized H2 as an electron donor but metabolized little of the acetate that P.carbinolicus produced. This suggested that H2, but not electrons derived from DIET, inhibited acetate metabolism. P. carbinolicus-M. barkeri cocultures did not aggregate, demonstrating that, unlike DIET, close physical contact was not necessary for interspecies H2 transfer. M. barkeri is the second methanogen found to accept electrons via DIET and the first methanogen known to be capable of using either H2 or electrons derived from DIET for CO2 reduction. Furthermore, M. barkeri is genetically tractable,making it a model organism for elucidating mechanisms by which methanogens make biological electrical connections with other cells.
}, keywords = {Biological Transport, Electron Transport, Ethanol, Fimbriae Proteins, Fimbriae, Bacterial, Geobacter, Hydrogen, Methane, Methanosarcina barkeri}, issn = {1098-5336}, doi = {10.1128/AEM.00895-14}, author = {Rotaru, Amelia-Elena and Shrestha, Pravin Malla and Liu, Fanghua and Markovaite, Beatrice and Chen, Shanshan and Nevin, Kelly P and Lovley, Derek R} } @article {3129, title = {The Dnmt2 RNA methyltransferase homolog of Geobacter sulfurreducens specifically methylates tRNA-Glu.}, journal = {Nucleic Acids Res}, volume = {42}, year = {2014}, month = {2014 Jun}, pages = {6487-96}, abstract = {Dnmt2 enzymes are conserved in eukaryotes, where they methylate C38 of tRNA-Asp with high activity. Here, the activity of one of the very few prokaryotic Dnmt2 homologs from Geobacter species (GsDnmt2) was investigated. GsDnmt2 was observed to methylate tRNA-Asp from flies and mice. Unexpectedly, it had only a weak activity toward its matching Geobacter tRNA-Asp, but methylated Geobacter tRNA-Glu with good activity. In agreement with this result, we show that tRNA-Glu is methylated in Geobacter while the methylation is absent in tRNA-Asp. The activities of Dnmt2 enzymes from Homo sapiens, Drosophila melanogaster, Schizosaccharomyces pombe and Dictyostelium discoideum for methylation of the Geobacter tRNA-Asp and tRNA-Glu were determined showing that all these Dnmt2s preferentially methylate tRNA-Asp. Hence, the GsDnmt2 enzyme has a swapped transfer ribonucleic acid (tRNA) specificity. By comparing the different tRNAs, a characteristic sequence pattern was identified in the variable loop of all preferred tRNA substrates. An exchange of two nucleotides in the variable loop of murine tRNA-Asp converted it to the corresponding variable loop of tRNA-Glu and led to a strong reduction of GsDnmt2 activity. Interestingly, the same loss of activity was observed with human DNMT2, indicating that the variable loop functions as a specificity determinant in tRNA recognition of Dnmt2 enzymes.
}, keywords = {Animals, Bacterial Proteins, Geobacter, Humans, Methylation, Mice, Nucleic Acid Conformation, RNA, Transfer, Asp, RNA, Transfer, Glu, Substrate Specificity, tRNA Methyltransferases}, issn = {1362-4962}, doi = {10.1093/nar/gku256}, author = {Shanmugam, Raghuvaran and Aklujkar, Muktak and Sch{\"a}fer, Matthias and Reinhardt, Richard and Nickel, Olaf and Reuter, Gunter and Lovley, Derek R and Ehrenhofer-Murray, Ann and Nellen, Wolfgang and Ankri, Serge and Helm, Mark and Jurkowski, Tomasz P and Jeltsch, Albert} } @article {1418, title = {Easy come easy go: surfaces containing immobilized nanoparticles or isolated polycation chains facilitate removal of captured Staphylococcus aureus by retarding bacterial bond maturation.}, journal = {ACS Nano}, volume = {8}, year = {2014}, month = {2014 Feb 25}, pages = {1180-90}, abstract = {Adhesion of bacteria is a key step in the functioning of antimicrobial surfaces or certain types of on-line sensors. The subsequent removal of these bacteria, within a \∼ 10-30 min time frame, is equally important but complicated by the tendency of bacterial adhesion to strengthen within minutes of initial capture. This study uses Staphylococcus aureus as a model bacterium to demonstrate the general strategy of clustering adhesive surface functionality (at length scales smaller than the bacteria themselves) on otherwise nonadhesive surfaces to capture and retain bacteria (easy come) while limiting the progressive strengthening of adhesion. The loose attachment facilitates bacteria removal by moderate shearing flow (easy go). This strategy is demonstrated using surfaces containing sparsely and randomly arranged immobilized amine-functionalized nanoparticles or poly-l-lysine chains, about 10 nm in size. The rest of the surface is backfilled with a nonadhesive polyethylene glycol (PEG) brush that, by itself, repels S. aureus. The nanoparticles or polymer chains cluster cationic functionality, providing small regions that attract negatively charged S. aureus cells. Compared with surfaces of nearly uniform cationic character where S. aureus adhesion quickly becomes strong (on a time scale less than 5 min), placement of cationic charge in small clusters retards or prevents processes that increase bacteria adhesion on a time scale of \∼ 30 min, providing \"easy go\" surfaces.
}, keywords = {Nanoparticles, Polyamines, Staphylococcus aureus, Surface Properties}, issn = {1936-086X}, doi = {10.1021/nn405845y}, author = {Fang, Bing and Jiang, Ying and Rotello, Vincent M and N{\"u}sslein, Klaus and Santore, Maria M} } @article {1560, title = {Environmental fate of the next generation refrigerant 2,3,3,3-tetrafluoropropene (HFO-1234yf).}, journal = {Environ Sci Technol}, volume = {48}, year = {2014}, month = {2014 Nov 18}, pages = {13181-7}, abstract = {The hydrofluoroolefin 2,3,3,3-tetrafluoropropene (HFO-1234yf) has been introduced to replace 1,1,1,2-tetrafluoroethane (HFC-134a) as refrigerant in mobile, including vehicle, air conditioning systems because of its lower global warming potential. HFO-1234yf is volatile at ambient temperatures; however, high production volumes and widespread handling are expected to release this fluorocarbon into terrestrial and aquatic environments, including groundwater. Laboratory experiments explored HFO-1234yf degradation by (i) microbial processes under oxic and anoxic conditions, (ii) abiotic processes mediated by reactive mineral phases and zerovalent iron (Fe(0), ZVI), and (iii) cobalamin-catalyzed biomimetic transformation. These investigations demonstrated that HFO-1234yf was recalcitrant to microbial (co)metabolism and no transformation was observed in incubations with ZVI, makinawite (FeS), sulfate green rust (GR(SO4)), magnetite (Fe(3)O(4)), and manganese oxide (MnO2). Sequential reductive defluorination of HFO-1234yf to 3,3,3-trifluoropropene and 3,3-dichloropropene with concomitant stoichiometric release of fluoride occurred in incubations with reduced cobalamins (e.g., vitamin B12) indicating that biomolecules can transform HFO-1234yf at circumneutral pH and at ambient temperature. Taken together, these findings suggest that HFO-1234yf recalcitrance in aquifers should be expected; however, HFO-1234yf is not inert and a biomolecule may mediate reductive transformation in low redox environments, albeit at low rates.
}, issn = {1520-5851}, doi = {10.1021/es5032147}, author = {Im, Jeongdae and Walshe-Langford, Gillian E and Moon, Ji-Won and L{\"o}ffler, Frank E} } @article {3131, title = {A Geobacter sulfurreducens strain expressing pseudomonas aeruginosa type IV pili localizes OmcS on pili but is deficient in Fe(III) oxide reduction and current production.}, journal = {Appl Environ Microbiol}, volume = {80}, year = {2014}, month = {2014 Feb}, pages = {1219-24}, abstract = {The conductive pili of Geobacter species play an important role in electron transfer to Fe(III) oxides, in long-range electron transport through current-producing biofilms, and in direct interspecies electron transfer. Although multiple lines of evidence have indicated that the pili of Geobacter sulfurreducens have a metal-like conductivity, independent of the presence of c-type cytochromes, this claim is still controversial. In order to further investigate this phenomenon, a strain of G. sulfurreducens, designated strain PA, was constructed in which the gene for the native PilA, the structural pilin protein, was replaced with the PilA gene of Pseudomonas aeruginosa PAO1. Strain PA expressed and properly assembled P. aeruginosa PilA subunits into pili and exhibited a profile of outer surface c-type cytochromes similar to that of a control strain expressing the G. sulfurreducens PilA. Surprisingly, the strain PA pili were decorated with the c-type cytochrome OmcS in a manner similar to the control strain. However, the strain PA pili were 14-fold less conductive than the pili of the control strain, and strain PA was severely impaired in Fe(III) oxide reduction and current production. These results demonstrate that the presence of OmcS on pili is not sufficient to confer conductivity to pili and suggest that there are unique structural features of the G. sulfurreducens PilA that are necessary for conductivity.
}, keywords = {Amino Acid Sequence, Cytochromes c, Electricity, Ferric Compounds, Fimbriae Proteins, Fimbriae, Bacterial, Geobacter, Methanosarcinaceae, Molecular Sequence Data, Oxidation-Reduction, Pseudomonas aeruginosa, Sequence Alignment}, issn = {1098-5336}, doi = {10.1128/AEM.02938-13}, author = {Liu, Xing and Tremblay, Pier-Luc and Malvankar, Nikhil S and Nevin, Kelly P and Lovley, Derek R and Vargas, Madeline} } @article {3126, title = {Going wireless: Fe(III) oxide reduction without pili by Geobacter sulfurreducens strain JS-1.}, journal = {Appl Environ Microbiol}, volume = {80}, year = {2014}, month = {2014 Jul}, pages = {4331-40}, abstract = {Previous studies have suggested that the conductive pili of Geobacter sulfurreducens are essential for extracellular electron transfer to Fe(III) oxides and for optimal long-range electron transport through current-producing biofilms. The KN400 strain of G. sulfurreducens reduces poorly crystalline Fe(III) oxide more rapidly than the more extensively studied DL-1 strain. Deletion of the gene encoding PilA, the structural pilin protein, in strain KN400 inhibited Fe(III) oxide reduction. However, low rates of Fe(III) reduction were detected after extended incubation (>30 days) in the presence of Fe(III) oxide. After seven consecutive transfers, the PilA-deficient strain adapted to reduce Fe(III) oxide as fast as the wild type. Microarray, whole-genome resequencing, proteomic, and gene deletion studies indicated that this adaptation was associated with the production of larger amounts of the c-type cytochrome PgcA, which was released into the culture medium. It is proposed that the extracellular cytochrome acts as an electron shuttle, promoting electron transfer from the outer cell surface to Fe(III) oxides. The adapted PilA-deficient strain competed well with the wild-type strain when both were grown together on Fe(III) oxide. However, when 50\% of the culture medium was replaced with fresh medium every 3 days, the wild-type strain outcompeted the adapted strain. A possible explanation for this is that the necessity to produce additional PgcA, to replace the PgcA being continually removed, put the adapted strain at a competitive disadvantage, similar to the apparent selection against electron shuttle-producing Fe(III) reducers in many anaerobic soils and sediments. Despite increased extracellular cytochrome production, the adapted PilA-deficient strain produced low levels of current, consistent with the concept that long-range electron transport through G. sulfurreducens biofilms is more effective via pili.
}, keywords = {Adaptation, Physiological, Biofilms, DNA, Bacterial, Electron Transport, Ferric Compounds, Fimbriae Proteins, Fimbriae, Bacterial, Gene Deletion, Geobacter, Oligonucleotide Array Sequence Analysis, Proteomics, Sequence Analysis, DNA}, issn = {1098-5336}, doi = {10.1128/AEM.01122-14}, author = {Smith, Jessica A and Tremblay, Pier-Luc and Shrestha, Pravin Malla and Snoeyenbos-West, Oona L and Franks, Ashley E and Nevin, Kelly P and Lovley, Derek R} } @article {1201, title = {Identification and Characterization of an Archaeal Kojibiose Catabolic Pathway in the Hyperthermophilic Pyrococcus sp. Strain ST04.}, journal = {J Bacteriol}, volume = {196}, year = {2014}, month = {2014 Mar}, pages = {1122-31}, abstract = {A unique gene cluster responsible for kojibiose utilization was identified in the genome of Pyrococcus sp. strain ST04. The proteins it encodes hydrolyze kojibiose, a disaccharide product of glucose caramelization, and form glucose-6-phosphate (G6P) in two steps. Heterologous expression of the kojibiose-related enzymes in Escherichia coli revealed that two genes, Py04_1502 and Py04_1503, encode kojibiose phosphorylase (designated PsKP, for Pyrococcus sp. strain ST04 kojibiose phosphorylase) and β-phosphoglucomutase (PsPGM), respectively. Enzymatic assays show that PsKP hydrolyzes kojibiose to glucose and β-glucose-1-phosphate (β-G1P). The Km values for kojibiose and phosphate were determined to be 2.53 {\textpm} 0.21 mM and 1.34 {\textpm} 0.04 mM, respectively. PsPGM then converts β-G1P into G6P in the presence of 6 mM MgCl2. Conversion activity from β-G1P to G6P was 46.81 {\textpm} 3.66 U/mg, and reverse conversion activity from G6P to β-G1P was 3.51 {\textpm} 0.13 U/mg. The proteins are highly thermostable, with optimal temperatures of 90{\textdegree}C for PsKP and 95{\textdegree}C for PsPGM. These results indicate that Pyrococcus sp. strain ST04 converts kojibiose into G6P, a substrate of the glycolytic pathway. This is the first report of a disaccharide utilization pathway via phosphorolysis in hyperthermophilic archaea.
}, issn = {1098-5530}, doi = {10.1128/JB.01222-13}, author = {Jung, Jong-Hyun and Seo, Dong-Ho and Holden, James F and Park, Cheon-Seok} } @article {3122, title = {Identification of genes specifically required for the anaerobic metabolism of benzene in Geobacter metallireducens.}, journal = {Front Microbiol}, volume = {5}, year = {2014}, month = {2014}, pages = {245}, abstract = {Although the biochemical pathways for the anaerobic degradation of many of the hydrocarbon constituents in petroleum reservoirs have been elucidated, the mechanisms for anaerobic activation of benzene, a very stable molecule, are not known. Previous studies have demonstrated that Geobacter metallireducens can anaerobically oxidize benzene to carbon dioxide with Fe(III) as the sole electron acceptor and that phenol is an intermediate in benzene oxidation. In an attempt to identify enzymes that might be involved in the conversion of benzene to phenol, whole-genome gene transcript abundance was compared in cells metabolizing benzene and cells metabolizing phenol. Eleven genes had significantly higher transcript abundance in benzene-metabolizing cells. Five of these genes had annotations suggesting that they did not encode proteins that could be involved in benzene metabolism and were not further studied. Strains were constructed in which one of the remaining six genes was deleted. The strain in which the monocistronic gene Gmet 0232 was deleted metabolized phenol, but not benzene. Transcript abundance of the adjacent monocistronic gene, Gmet 0231, predicted to encode a zinc-containing oxidoreductase, was elevated in cells metabolizing benzene, although not at a statistically significant level. However, deleting Gmet 0231 also yielded a strain that could metabolize phenol, but not benzene. Although homologs of Gmet 0231 and Gmet 0232 are found in microorganisms not known to anaerobically metabolize benzene, the adjacent localization of these genes is unique to G. metallireducens. The discovery of genes that are specifically required for the metabolism of benzene, but not phenol in G. metallireducens is an important step in potentially identifying the mechanisms for anaerobic benzene activation.
}, issn = {1664-302X}, doi = {10.3389/fmicb.2014.00245}, author = {Zhang, Tian and Tremblay, Pier-Luc and Chaurasia, Akhilesh K and Smith, Jessica A and Bain, Timothy S and Lovley, Derek R} } @article {1442, title = {Imaging bacterial peptidoglycan with near-infrared fluorogenic azide probes.}, journal = {Proc Natl Acad Sci U S A}, volume = {111}, year = {2014}, month = {2014 Apr 15}, pages = {5456-61}, abstract = {Fluorescent probes designed for activation by bioorthogonal chemistry have enabled the visualization of biomolecules in living systems. Such activatable probes with near-infrared (NIR) emission would be ideal for in vivo imaging but have proven difficult to engineer. We present the development of NIR fluorogenic azide probes based on the Si-rhodamine scaffold that undergo a fluorescence enhancement of up to 48-fold upon reaction with terminal or strained alkynes. We used the probes for mammalian cell surface imaging and, in conjunction with a new class of cyclooctyne D-amino acids, for visualization of bacterial peptidoglycan without the need to wash away unreacted probe.
}, keywords = {Azides, Fluorescent Dyes, Molecular Imaging, Molecular Probe Techniques, Molecular Structure, Peptidoglycan}, issn = {1091-6490}, doi = {10.1073/pnas.1322727111}, author = {Shieh, Peyton and Siegrist, M Sloan and Cullen, Andrew J and Bertozzi, Carolyn R} } @article {3130, title = {Lactose-inducible system for metabolic engineering of Clostridium ljungdahlii.}, journal = {Appl Environ Microbiol}, volume = {80}, year = {2014}, month = {2014 Apr}, pages = {2410-6}, abstract = {The development of tools for genetic manipulation of Clostridium ljungdahlii has increased its attractiveness as a chassis for autotrophic production of organic commodities and biofuels from syngas and microbial electrosynthesis and established it as a model organism for the study of the basic physiology of acetogenesis. In an attempt to expand the genetic toolbox for C. ljungdahlii, the possibility of adapting a lactose-inducible system for gene expression, previously reported for Clostridium perfringens, was investigated. The plasmid pAH2, originally developed for C. perfringens with a gusA reporter gene, functioned as an effective lactose-inducible system in C. ljungdahlii. Lactose induction of C. ljungdahlii containing pB1, in which the gene for the aldehyde/alcohol dehydrogenase AdhE1 was downstream of the lactose-inducible promoter, increased expression of adhE1 30-fold over the wild-type level, increasing ethanol production 1.5-fold, with a corresponding decrease in acetate production. Lactose-inducible expression of adhE1 in a strain in which adhE1 and the adhE1 homolog adhE2 had been deleted from the chromosome restored ethanol production to levels comparable to those in the wild-type strain. Inducing expression of adhE2 similarly failed to restore ethanol production, suggesting that adhE1 is the homolog responsible for ethanol production. Lactose-inducible expression of the four heterologous genes necessary to convert acetyl coenzyme A (acetyl-CoA) to acetone diverted ca. 60\% of carbon flow to acetone production during growth on fructose, and 25\% of carbon flow went to acetone when carbon monoxide was the electron donor. These studies demonstrate that the lactose-inducible system described here will be useful for redirecting carbon and electron flow for the biosynthesis of products more valuable than acetate. Furthermore, this tool should aid in optimizing microbial electrosynthesis and for basic studies on the physiology of acetogenesis.
}, keywords = {Acetic Acid, Acetone, Acetyl Coenzyme A, Alcohol Dehydrogenase, Carbon, Clostridium, Ethanol, Fructose, Gene Expression, Gene Expression Regulation, Bacterial, Lactose, Metabolic Engineering, Metabolic Flux Analysis, Transcriptional Activation}, issn = {1098-5336}, doi = {10.1128/AEM.03666-13}, author = {Banerjee, Areen and Leang, Ching and Ueki, Toshiyuki and Nevin, Kelly P and Lovley, Derek R} } @article {1417, title = {Land use change alters functional gene diversity, composition and abundance in Amazon forest soil microbial communities.}, journal = {Mol Ecol}, volume = {23}, year = {2014}, month = {2014 Jun}, pages = {2988-99}, abstract = {Land use change in the Amazon rainforest alters the taxonomic structure of soil microbial communities, but whether it alters their functional gene composition is unknown. We used the highly parallel microarray technology GeoChip 4.0, which contains 83,992 probes specific for genes linked nutrient cycling and other processes, to evaluate how the diversity, abundance and similarity of the targeted genes responded to forest-to-pasture conversion. We also evaluated whether these parameters were reestablished with secondary forest growth. A spatially nested scheme was employed to sample a primary forest, two pastures (6 and 38 years old) and a secondary forest. Both pastures had significantly lower microbial functional genes richness and diversity when compared to the primary forest. Gene composition and turnover were also significantly modified with land use change. Edaphic traits associated with soil acidity, iron availability, soil texture and organic matter concentration were correlated with these gene changes. Although primary and secondary forests showed similar functional gene richness and diversity, there were differences in gene composition and turnover, suggesting that community recovery was not complete in the secondary forest. Gene association analysis revealed that response to ecosystem conversion varied significantly across functional gene groups, with genes linked to carbon and nitrogen cycling mostly altered. This study indicates that diversity and abundance of numerous environmentally important genes respond to forest-to-pasture conversion and hence have the potential to affect the related processes at an ecosystem scale.
}, keywords = {Agriculture, Carbon Cycle, Ecosystem, Genes, Bacterial, Genes, Fungal, Genetic Variation, Metagenome, Multigene Family, Nitrogen Cycle, Oligonucleotide Array Sequence Analysis, Soil Microbiology, Trees, Tropical Climate}, issn = {1365-294X}, doi = {10.1111/mec.12786}, author = {Paula, Fabiana S and Rodrigues, Jorge L M and Zhou, Jizhong and Wu, Liyou and Mueller, Rebecca C and Mirza, Babur S and Bohannan, Brendan J M and N{\"u}sslein, Klaus and Deng, Ye and Tiedje, James M and Pellizari, Vivian H} } @article {1419, title = {Links between plant and fungal communities across a deforestation chronosequence in the Amazon rainforest.}, journal = {ISME J}, volume = {8}, year = {2014}, month = {2014 Jul}, pages = {1548-50}, abstract = {Understanding the interactions among microbial communities, plant communities and soil properties following deforestation could provide insights into the long-term effects of land-use change on ecosystem functions, and may help identify approaches that promote the recovery of degraded sites. We combined high-throughput sequencing of fungal rDNA and molecular barcoding of plant roots to estimate fungal and plant community composition in soil sampled across a chronosequence of deforestation. We found significant effects of land-use change on fungal community composition, which was more closely correlated to plant community composition than to changes in soil properties or geographic distance, providing evidence for strong links between above- and below-ground communities in tropical forests.
}, keywords = {Brazil, Conservation of Natural Resources, DNA Barcoding, Taxonomic, DNA, Fungal, DNA, Ribosomal, Ecosystem, Fungi, Phylogeny, Plant Roots, Soil Microbiology, Trees, Tropical Climate}, issn = {1751-7370}, doi = {10.1038/ismej.2013.253}, author = {Mueller, Rebecca C and Paula, Fabiana S and Mirza, Babur S and Rodrigues, Jorge L M and N{\"u}sslein, Klaus and Bohannan, Brendan J M} } @article {1411, title = {Magnetite formation from ferrihydrite by hyperthermophilic archaea from Endeavour Segment, Juan de Fuca Ridge hydrothermal vent chimneys.}, journal = {Geobiology}, volume = {12}, year = {2014}, month = {2014 May}, pages = {200-11}, abstract = {Hyperthermophilic iron reducers are common in hydrothermal chimneys found along the Endeavour Segment in the northeastern Pacific Ocean based on culture-dependent estimates. However, information on the availability of Fe(III) (oxyhydr) oxides within these chimneys, the types of Fe(III) (oxyhydr) oxides utilized by the organisms, rates and environmental constraints of hyperthermophilic iron reduction, and mineral end products is needed to determine their biogeochemical significance and are addressed in this study. Thin-section petrography on the interior of a hydrothermal chimney from the Dante edifice at Endeavour showed a thin coat of Fe(III) (oxyhydr) oxide associated with amorphous silica on the exposed outer surfaces of pyrrhotite, sphalerite, and chalcopyrite in pore spaces, along with anhydrite precipitation in the pores that is indicative of seawater ingress. The iron sulfide minerals were likely oxidized to Fe(III) (oxyhydr) oxide with increasing pH and Eh due to cooling and seawater exposure, providing reactants for bioreduction. Culture-dependent estimates of hyperthermophilic iron reducer abundances in this sample were 1740 and 10 cells per gram (dry weight) of material from the outer surface and the marcasite-sphalerite-rich interior, respectively. Two hyperthermophilic iron reducers, Hyperthermus sp. Ro04 and Pyrodictium sp. Su06, were isolated from other active hydrothermal chimneys on the Endeavour Segment. Strain Ro04 is a neutrophilic (pH opt 7-8) heterotroph, while strain Su06 is a mildly acidophilic (pH opt 5), hydrogenotrophic autotroph, both with optimal growth temperatures of 90-92 \°C. M\össbauer spectroscopy of the iron oxides before and after growth demonstrated that both organisms form nanophase (\<12 nm) magnetite [Fe3 O4 ] from laboratory-synthesized ferrihydrite [Fe10 O14 (OH)2 ] with no detectable mineral intermediates. They produced up to 40 mm Fe(2+) in a growth-dependent manner, while all abiotic and biotic controls produced \<3 mm Fe(2+) . Hyperthermophilic iron reducers may have a growth advantage over other hyperthermophiles in hydrothermal systems that are mildly acidic where mineral weathering at increased temperatures occurs.
}, keywords = {Ferric Compounds, Ferrosoferric Oxide, Hydrothermal Vents, Iron, Oxidation-Reduction, Pacific Ocean, Photomicrography, Pyrodictiaceae, Spectroscopy, Mossbauer, Sulfides}, issn = {1472-4669}, doi = {10.1111/gbi.12083}, author = {Lin, T Jennifer and Breves, E A and Dyar, M D and Ver Eecke, H C and Jamieson, J W and Holden, J F} } @article {1203, title = {Maltose-forming α-amylase from the hyperthermophilic archaeon Pyrococcus sp. ST04.}, journal = {Appl Microbiol Biotechnol}, volume = {98}, year = {2014}, month = {2014 Mar}, pages = {2121-31}, abstract = {The deduced amino acid sequence from a gene of the hyperthermophilic archaeon Pyrococcus sp. ST04 (Py04_0872) contained a conserved glycoside hydrolase family 57 (GH57) motif, but showed <13~\% sequence identity with other known Pyrococcus GH57 enzymes, such as 4-α-glucanotransferase (EC 2.4.1.25), amylopullulanase (EC 3.2.1.41), and branching enzyme (EC 2.4.1.18). This gene was cloned and expressed in Escherichia coli, and the recombinant product (P yrococcus sp. ST04 maltose-forming α-amylase, PSMA) was a novel 70-kDa maltose-forming α-amylase. PSMA only recognized maltose (G2) units with α-1,4 and α-1,6 linkages in polysaccharides (e.g., starch, amylopectin, and glycogen) and hydrolyzed pullulan very poorly. G2 was the primary end product of hydrolysis. Branched cyclodextrin (CD) was only hydrolyzed along its branched maltooligosaccharides. 6-O-glucosyl-β-cyclodextrin (G1-β-CD) and β-cyclodextrin (β-CD) were resistant to PSMA suggesting that PSMA is an exo-type glucan hydrolase with α-1,4- and α-1,6-glucan hydrolytic activities. The half-saturation value (K m) for the α-1,4 linkage of maltotriose (G3) was 8.4~mM while that of the α-1,6 linkage of 6-O-maltosyl-β-cyclodextrin (G2-β-CD) was 0.3~mM. The k cat values were 381.0~min(-1) for G3 and 1,545.0~min(-1) for G2-β-CD. The enzyme was inhibited competitively by the reaction product G2, and the K i constant was 0.7~mM. PSMA bridges the gap between amylases that hydrolyze larger maltodextrins and α-glucosidase that feeds G2 into glycolysis by hydrolyzing smaller glucans into G2 units.
}, issn = {1432-0614}, doi = {10.1007/s00253-013-5068-6}, author = {Jung, Jong-Hyun and Seo, Dong-Ho and Holden, James F and Park, Cheon-Seok} } @article {3119, title = {Methane production from protozoan endosymbionts following stimulation of microbial metabolism within subsurface sediments.}, journal = {Front Microbiol}, volume = {5}, year = {2014}, month = {2014}, pages = {366}, abstract = {Previous studies have suggested that protozoa prey on Fe(III)- and sulfate-reducing bacteria that are enriched when acetate is added to uranium contaminated subsurface sediments to stimulate U(VI) reduction. In order to determine whether protozoa continue to impact subsurface biogeochemistry after these acetate amendments have stopped, 18S rRNA and {\ss}-tubulin sequences from this phase of an in situ uranium bioremediation field experiment were analyzed. Sequences most similar to Metopus species predominated, with the majority of sequences most closely related to M. palaeformis, a cilitated protozoan known to harbor methanogenic symbionts. Quantification of mcrA mRNA transcripts in the groundwater suggested that methanogens closely related to Metopus endosymbionts were metabolically active at this time. There was a strong correlation between the number of mcrA transcripts from the putative endosymbiotic methanogen and Metopus {\ss}-tubulin mRNA transcripts during the course of the field experiment, suggesting that the activity of the methanogens was dependent upon the activity of the Metopus species. Addition of the eukaryotic inhibitors cyclohexamide and colchicine to laboratory incubations of acetate-amended subsurface sediments significantly inhibited methane production and there was a direct correlation between methane concentration and Metopus {\ss}-tubulin and putative symbiont mcrA gene copies. These results suggest that, following the stimulation of subsurface microbial growth with acetate, protozoa harboring methanogenic endosymbionts become important members of the microbial community, feeding on moribund biomass and producing methane.
}, issn = {1664-302X}, doi = {10.3389/fmicb.2014.00366}, author = {Holmes, Dawn E and Giloteaux, Ludovic and Orellana, Roberto and Williams, Kenneth H and Robbins, Mark J and Lovley, Derek R} } @article {3123, title = {Microbial nanowires for bioenergy applications.}, journal = {Curr Opin Biotechnol}, volume = {27}, year = {2014}, month = {2014 Jun}, pages = {88-95}, abstract = {Microbial nanowires are electrically conductive filaments that facilitate long-range extracellular electron transfer. The model for electron transport along Shewanella oneidensis nanowires is electron hopping/tunneling between cytochromes adorning the filaments. Geobacter sulfurreducens nanowires are comprised of pili that have metal-like conductivity attributed to overlapping pi-pi orbitals of aromatic amino acids. The nanowires of Geobacter species have been implicated in direct interspecies electron transfer (DIET), which may be an important mode of syntrophy in the conversion of organic wastes to methane. Nanowire networks confer conductivity to Geobacter biofilms converting organic compounds to electricity in microbial fuel cells (MFCs) and increasing nanowire production is the only genetic manipulation shown to yield strains with improved current-producing capabilities. Introducing nanowires, or nanowire mimetics, might improve other bioenergy strategies that rely on extracellular electron exchange, such as microbial electrosynthesis. Similarities between microbial nanowires and synthetic conducting polymers suggest additional energy-related applications.
}, keywords = {Anaerobiosis, Bioelectric Energy Sources, Biofilms, Electric Conductivity, Electricity, Electron Transport, Electrons, Fimbriae, Bacterial, Geobacter, Methane, Nanowires, Shewanella, Synthetic Biology}, issn = {1879-0429}, doi = {10.1016/j.copbio.2013.12.003}, author = {Malvankar, Nikhil S and Lovley, Derek R} } @article {1441, title = {Mycobacterial Esx-3 requires multiple components for iron acquisition.}, journal = {MBio}, volume = {5}, year = {2014}, month = {2014}, pages = {e01073-14}, abstract = {ABSTRACT The type VII secretion systems are conserved across mycobacterial species and in many Gram-positive bacteria. While the well-characterized Esx-1 pathway is required for the virulence of pathogenic mycobacteria and conjugation in the model organism Mycobacterium smegmatis, Esx-3 contributes to mycobactin-mediated iron acquisition in these bacteria. Here we show that several Esx-3 components are individually required for function under low-iron conditions but that at least one, the membrane-bound protease MycP3 of M. smegmatis, is partially expendable. All of the esx-3 mutants tested, including the ΔmycP3ms mutant, failed to export the native Esx-3 substrates EsxHms and EsxGms to quantifiable levels, as determined by targeted mass spectrometry. Although we were able to restore low-iron growth to the esx-3 mutants by genetic complementation, we found a wide range of complementation levels for protein export. Indeed, minute quantities of extracellular EsxHms and EsxGms were sufficient for iron acquisition under our experimental conditions. The apparent separation of Esx-3 function in iron acquisition from robust EsxGms and EsxHms secretion in the ΔmycP3ms mutant and in some of the complemented esx-3 mutants compels reexamination of the structure-function relationships for type VII secretion systems. IMPORTANCE Mycobacteria have several paralogous type VII secretion systems, Esx-1 through Esx-5. Whereas Esx-1 is required for pathogenic mycobacteria to grow within an infected host, Esx-3 is essential for growth in vitro. We and others have shown that Esx-3 is required for siderophore-mediated iron acquisition. In this work, we identify individual Esx-3 components that contribute to this process. As in the Esx-1 system, most mutations that abolish Esx-3 protein export also disrupt its function. Unexpectedly, however, ultrasensitive quantitation of Esx-3 secretion by multiple-reaction-monitoring mass spectrometry (MRM-MS) revealed that very low levels of export were sufficient for iron acquisition under similar conditions. Although protein export clearly contributes to type VII function, the relationship is not absolute.
}, keywords = {Bacterial Secretion Systems, Gene Order, Genetic Loci, Iron, Mycobacterium, Oxazoles}, issn = {2150-7511}, doi = {10.1128/mBio.01073-14}, author = {Siegrist, M Sloan and Steigedal, Magnus and Ahmad, Rushdy and Mehra, Alka and Dragset, Marte S and Schuster, Brian M and Philips, Jennifer A and Carr, Steven A and Rubin, Eric J} } @article {1443, title = {Mycobacterial lipid logic.}, journal = {Cell Host Microbe}, volume = {15}, year = {2014}, month = {2014 Jan 15}, pages = {1-2}, abstract = {During infection of the lung epithelium, Mycobacterium tuberculosis must infect and survive within macrophages long enough to be transported into deeper lung tissues. Cambier et~al. (2013) show that pathogenic mycobacteria use the coordinated action of two cell wall glycolipids to regulate macrophage recruitment to initial infection sites.
}, keywords = {Animals, Female, Immune Evasion, Macrophages, Membrane Lipids, Mycobacterium}, issn = {1934-6069}, doi = {10.1016/j.chom.2013.12.005}, author = {Siegrist, M Sloan and Bertozzi, Carolyn R} } @article {3124, title = {Promoting interspecies electron transfer with biochar.}, journal = {Sci Rep}, volume = {4}, year = {2014}, month = {2014 May 21}, pages = {5019}, abstract = {Biochar, a charcoal-like product of the incomplete combustion of organic materials, is an increasingly popular soil amendment designed to improve soil fertility. We investigated the possibility that biochar could promote direct interspecies electron transfer (DIET) in a manner similar to that previously reported for granular activated carbon (GAC). Although the biochars investigated were 1000 times less conductive than GAC, they stimulated DIET in co-cultures of Geobacter metallireducens with Geobacter sulfurreducens or Methanosarcina barkeri in which ethanol was the electron donor. Cells were attached to the biochar, yet not in close contact, suggesting that electrons were likely conducted through the biochar, rather than biological electrical connections. The finding that biochar can stimulate DIET may be an important consideration when amending soils with biochar and can help explain why biochar may enhance methane production from organic wastes under anaerobic conditions.
}, keywords = {Charcoal, Coculture Techniques, Electron Transport, Electrons, Ethanol, Geobacter, Methanosarcina barkeri, Soil}, issn = {2045-2322}, doi = {10.1038/srep05019}, author = {Chen, Shanshan and Rotaru, Amelia-Elena and Shrestha, Pravin Malla and Malvankar, Nikhil S and Liu, Fanghua and Fan, Wei and Nevin, Kelly P and Lovley, Derek R} } @article {1450, title = {Properties of Topological Networks of Flexible Polygonal Chains.}, journal = {Mol Based Math Biol}, volume = {2}, year = {2014}, pages = {98-106}, author = {Arsuaga, J and Diao, Y and Klingbeil, M and Rodriguez, V} } @article {3118, title = {Proteome of Geobacter sulfurreducens in the presence of U(VI).}, journal = {Microbiology (Reading)}, volume = {160}, year = {2014}, month = {2014 Dec}, pages = {2607-2617}, abstract = {Geobacter species often play an important role in the in situ bioremediation of uranium-contaminated groundwater, but little is known about how these microbes avoid uranium toxicity. To evaluate this further, the proteome of Geobacter sulfurreducens exposed to 100 {\textmu}M U(VI) acetate was compared to control cells not exposed to U(VI). Of the 1363 proteins detected from these cultures, 203 proteins had higher abundance during exposure to U(VI) compared with the control cells and 148 proteins had lower abundance. U(VI)-exposed cultures expressed lower levels of proteins involved in growth, protein and amino acid biosynthesis, as well as key central metabolism enzymes as a result of the deleterious effect of U(VI) on the growth of G. sulfurreducens. In contrast, proteins involved in detoxification, such as several efflux pumps belonging to the RND (resistance-nodulation-cell division) family, and membrane protection, and other proteins, such as chaperones and proteins involved in secretion systems, were found in higher abundance in cells exposed to U(VI). Exposing G. sulfurreducens to U(VI) resulted in a higher abundance of many proteins associated with the oxidative stress response, such as superoxide dismutase and superoxide reductase. A strain in which the gene for superoxide dismutase was deleted grew more slowly than the WT strain in the presence of U(VI), but not in its absence. The results suggested that there is no specific mechanism for uranium detoxification. Rather, multiple general stress responses are induced, which presumably enable Geobacter species to tolerate high uranium concentrations.
}, keywords = {Bacterial Proteins, Gene Expression Regulation, Bacterial, Geobacter, Organometallic Compounds, Proteome}, issn = {1465-2080}, doi = {10.1099/mic.0.081398-0}, author = {Orellana, Roberto and Hixson, Kim K and Murphy, Sean and Mester, T{\"u}nde and Sharma, Manju L and Lipton, Mary S and Lovley, Derek R} } @article {3112, title = {Real-time monitoring of subsurface microbial metabolism with graphite electrodes.}, journal = {Front Microbiol}, volume = {5}, year = {2014}, month = {2014}, pages = {621}, abstract = {Monitoring in situ microbial activity in anoxic submerged soils and aquatic sediments can be labor intensive and technically difficult, especially in dynamic environments in which a record of changes in microbial activity over time is desired. Microbial fuel cell concepts have previously been adapted to detect changes in the availability of relatively high concentrations of organic compounds in waste water but, in most soils and sediments, rates of microbial activity are not linked to the concentrations of labile substrates, but rather to the turnover rates of the substrate pools with steady state concentrations in the nM-μM range. In order to determine whether levels of current produced at a graphite anode would correspond to the rates of microbial metabolism in anoxic sediments, small graphite anodes were inserted in sediment cores and connected to graphite brush cathodes in the overlying water. Currents produced were compared with the rates of [2-(14)C]-acetate metabolism. There was a direct correlation between current production and the rate that [2-(14)C]-acetate was metabolized to (14)CO2 and (14)CH4 in sediments in which Fe(III) reduction, sulfate reduction, or methane production was the predominant terminal electron-accepting process. At comparable acetate turnover rates, currents were higher in the sediments in which sulfate-reduction or Fe(III) reduction predominated than in methanogenic sediments. This was attributed to reduced products (Fe(II), sulfide) produced at distance from the anode contributing to current production in addition to the current that was produced from microbial oxidation of organic substrates with electron transfer to the anode surface in all three sediment types. The results demonstrate that inexpensive graphite electrodes may provide a simple strategy for real-time monitoring of microbial activity in a diversity of anoxic soils and sediments.
}, issn = {1664-302X}, doi = {10.3389/fmicb.2014.00621}, author = {Wardman, Colin and Nevin, Kelly P and Lovley, Derek R} } @article {1562, title = {Regulation of ethanol-related behavior and ethanol metabolism by the Corazonin neurons and Corazonin receptor in Drosophila melanogaster.}, journal = {PLoS One}, volume = {9}, year = {2014}, month = {2014}, pages = {e87062}, abstract = {Impaired ethanol metabolism can lead to various alcohol-related health problems. Key enzymes in ethanol metabolism are alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH); however, neuroendocrine pathways that regulate the activities of these enzymes are largely unexplored. Here we identified a neuroendocrine system involving Corazonin (Crz) neuropeptide and its receptor (CrzR) as important physiological regulators of ethanol metabolism in Drosophila. Crz-cell deficient (Crz-CD) flies displayed significantly delayed recovery from ethanol-induced sedation that we refer to as hangover-like phenotype. Newly generated mutant lacking Crz Receptor (CrzR(01) ) and CrzR-knockdown flies showed even more severe hangover-like phenotype, which is causally associated with fast accumulation of acetaldehyde in the CrzR(01) mutant following ethanol exposure. Higher levels of acetaldehyde are likely due to 30\% reduced ALDH activity in the mutants. Moreover, increased ADH activity was found in the CrzR(01) mutant, but not in the Crz-CD flies. Quantitative RT-PCR revealed transcriptional upregulation of Adh gene in the CrzR(01) . Transgenic inhibition of cyclic AMP-dependent protein kinase (PKA) also results in significantly increased ADH activity and Adh mRNA levels, indicating PKA-dependent transcriptional regulation of Adh by CrzR. Furthermore, inhibition of PKA or cAMP response element binding protein (CREB) in CrzR cells leads to comparable hangover-like phenotype to the CrzR(01) mutant. These findings suggest that CrzR-associated signaling pathway is critical for ethanol detoxification via Crz-dependent regulation of ALDH activity and Crz-independent transcriptional regulation of ADH. Our study provides new insights into the neuroendocrine-associated ethanol-related behavior and metabolism.
}, keywords = {Acetaldehyde, Alcohol Dehydrogenase, Aldehyde Dehydrogenase, Alleles, Animals, Cyclic AMP-Dependent Protein Kinases, Drosophila melanogaster, Drosophila Proteins, Ethanol, Genes, Reporter, Male, Mutation, Neurons, Neuropeptides, Phenotype, Receptors, Neuropeptide, RNA, Messenger, Transcription, Genetic}, issn = {1932-6203}, doi = {10.1371/journal.pone.0087062}, author = {Sha, Kai and Choi, Seung-Hoon and Im, Jeongdae and Lee, Gyunghee G and Loeffler, Frank and Park, Jae H} } @article {1420, title = {Response of free-living nitrogen-fixing microorganisms to land use change in the Amazon rainforest.}, journal = {Appl Environ Microbiol}, volume = {80}, year = {2014}, month = {2014 Jan}, pages = {281-8}, abstract = {The Amazon rainforest, the largest equatorial forest in the world, is being cleared for pasture and agricultural use at alarming rates. Tropical deforestation is known to cause alterations in microbial communities at taxonomic and phylogenetic levels, but it is unclear whether microbial functional groups are altered. We asked whether free-living nitrogen-fixing microorganisms (diazotrophs) respond to deforestation in the Amazon rainforest, using analysis of the marker gene nifH. Clone libraries were generated from soil samples collected from a primary forest, a 5-year-old pasture originally converted from primary forest, and a secondary forest established after pasture abandonment. Although diazotroph richness did not significantly change among the three plots, diazotroph community composition was altered with forest-to-pasture conversion, and phylogenetic similarity was higher among pasture communities than among those in forests. There was also 10-fold increase in nifH gene abundance following conversion from primary forest to pasture. Three environmental factors were associated with the observed changes: soil acidity, total N concentration, and C/N ratio. Our results suggest a partial restoration to initial levels of abundance and community structure of diazotrophs following pasture abandonment, with primary and secondary forests sharing similar communities. We postulate that the response of diazotrophs to land use change is a direct consequence of changes in plant communities, particularly the higher N demand of pasture plant communities for supporting aboveground plant growth.
}, keywords = {Agriculture, Bacteria, Biota, Carbon, Cluster Analysis, Conservation of Natural Resources, Human Activities, Hydrogen-Ion Concentration, Molecular Sequence Data, Nitrogen, Nitrogen Fixation, Oxidoreductases, Phylogeny, Sequence Analysis, DNA, Soil, Soil Microbiology, South America, Trees}, issn = {1098-5336}, doi = {10.1128/AEM.02362-13}, author = {Mirza, Babur S and Potisap, Chotima and N{\"u}sslein, Klaus and Bohannan, Brendan J M and Rodrigues, Jorge L M} } @article {1456, title = {Saccharomyces cerevisiae essential genes with an Opi- phenotype.}, journal = {G3 (Bethesda)}, volume = {4}, year = {2014}, month = {2014 Apr}, pages = {761-7}, abstract = {The overproduction and secretion of inositol (i.e., Opi(-)) phenotype is associated with defects in regulation of phospholipid biosynthesis in yeast. Here we report a screen of the essential yeast gene set using a conditional-expression library. This screen identified novel functions previously unknown to affect phospholipid synthesis.
}, keywords = {Cluster Analysis, Genes, Essential, Histone Acetyltransferases, Inositol, Phenotype, Phospholipids, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins}, issn = {2160-1836}, doi = {10.1534/g3.113.010140}, author = {Salas-Santiago, Bryan and Lopes, John M} } @article {1412, title = {Structural features underlying the selective cleavage of a novel exo-type maltose-forming amylase from Pyrococcus sp. ST04.}, journal = {Acta Crystallogr D Biol Crystallogr}, volume = {70}, year = {2014}, month = {2014 Jun}, pages = {1659-68}, abstract = {A novel maltose-forming \α-amylase (PSMA) was recently found in the hyperthermophilic archaeon Pyrococcus sp. ST04. This enzyme shows \<13\% amino-acid sequence identity to other known \α-amylases and displays a unique enzymatic property in that it hydrolyzes both \α-1,4-glucosidic and \α-1,6-glucosidic linkages of substrates, recognizing only maltose units, in an exo-type manner. Here, the crystal structure of PSMA at a resolution of 1.8 \Å is reported, showing a tight ring-shaped tetramer with monomers composed of two domains: an N-domain (amino acids 1-341) with a typical GH57 family (\β/\α)7-barrel fold and a C-domain (amino acids 342-597) composed of \α-helical bundles. A small closed cavity observed in proximity to the catalytic residues Glu153 and Asp253 at the domain interface has the appropriate volume and geometry to bind a maltose unit, accounting for the selective exo-type maltose hydrolysis of the enzyme. A narrow gate at the putative subsite +1 formed by residue Phe218 and Phe452 is essential for specific cleavage of glucosidic bonds. The closed cavity at the active site is connected to a short substrate-binding channel that extends to the central hole of the tetramer, exhibiting a geometry that is significantly different from classical maltogenic amylases or \β-amylases. The structural features of this novel exo-type maltose-forming \α-amylase provide a molecular basis for its unique enzymatic characteristics and for its potential use in industrial applications and protein engineering.
}, keywords = {Amylases, Catalytic Domain, Maltose, Models, Molecular, Mutagenesis, Site-Directed, Protein Conformation, Proteolysis, Pyrococcus, Substrate Specificity}, issn = {1399-0047}, doi = {10.1107/S1399004714006567}, author = {Park, Kwang-Hyun and Jung, Jong-Hyun and Park, Sung-Goo and Lee, Myeong-Eun and Holden, James F and Park, Cheon-Seok and Woo, Eui-Jeon} } @article {1440, title = {Subpolar addition of new cell wall is directed by DivIVA in mycobacteria.}, journal = {Proc Natl Acad Sci U S A}, volume = {111}, year = {2014}, month = {2014 Aug 5}, pages = {E3243-51}, abstract = {Mycobacteria are surrounded by a complex multilayered envelope and elongate at the poles. The principles that organize the coordinated addition of chemically diverse cell wall layers during polar extension remain unclear. We show that enzymes mediating the terminal cytosolic steps of peptidoglycan, arabinogalactan, and mycolic acid synthesis colocalize at sites of cell growth or division. The tropomyosin-like protein, DivIVA, is targeted to the negative curvature of the pole, is enriched at the growing end, and determines cell shape from this site. In contrast, cell wall synthetic complexes are concentrated at a distinct subpolar location. When viewed at subdiffraction resolution, new peptidoglycan is deposited at this subpolar site, and inert cell wall covers the DivIVA-marked tip. The differentiation between polar tip and cell wall synthetic complexes is also apparent at the biochemical level. Enzymes that generate mycolate precursors interact with DivIVA, but the final condensation of mycolic acids occurs in a distinct protein complex at the site of nascent cell wall addition. We propose an ultrastructural model of mycobacterial polar growth where new cell wall is added in an annular zone below the cell tip. This model may be broadly applicable to other bacterial and fungal organisms that grow via polar extension.
}, keywords = {Bacterial Proteins, Cell Cycle Proteins, Cell Membrane, Cell Polarity, Cell Wall, Models, Biological, Mycobacterium smegmatis, Mycolic Acids, Protein Binding}, issn = {1091-6490}, doi = {10.1073/pnas.1402158111}, author = {Meniche, Xavier and Otten, Renee and Siegrist, M Sloan and Baer, Christina E and Murphy, Kenan C and Bertozzi, Carolyn R and Sassetti, Christopher M} } @article {3135, title = {Sulfur oxidation to sulfate coupled with electron transfer to electrodes by Desulfuromonas strain TZ1.}, journal = {Microbiology (Reading)}, volume = {160}, year = {2014}, month = {2014 Jan}, pages = {123-129}, abstract = {Microbial oxidation of elemental sulfur with an electrode serving as the electron acceptor is of interest because this may play an important role in the recovery of electrons from sulfidic wastes and for current production in marine benthic microbial fuel cells. Enrichments initiated with a marine sediment inoculum, with elemental sulfur as the electron donor and a positively poised (+300 mV versus Ag/AgCl) anode as the electron acceptor, yielded an anode biofilm with a diversity of micro-organisms, including Thiobacillus, Sulfurimonas, Pseudomonas, Clostridium and Desulfuromonas species. Further enrichment of the anode biofilm inoculum in medium with elemental sulfur as the electron donor and Fe(III) oxide as the electron acceptor, followed by isolation in solidified sulfur/Fe(III) medium yielded a strain of Desulfuromonas, designated strain TZ1. Strain TZ1 effectively oxidized elemental sulfur to sulfate with an anode serving as the sole electron acceptor, at rates faster than Desulfobulbus propionicus, the only other organism in pure culture previously shown to oxidize S{\textdegree} with current production. The abundance of Desulfuromonas species enriched on the anodes of marine benthic fuel cells has previously been interpreted as acetate oxidation driving current production, but the results presented here suggest that sulfur-driven current production is a likely alternative.
}, keywords = {Bioelectric Energy Sources, Desulfuromonas, DNA, Bacterial, Electricity, Electrodes, Geologic Sediments, Molecular Sequence Data, Oxidation-Reduction, Sequence Analysis, DNA, Sulfates, Sulfur}, issn = {1465-2080}, doi = {10.1099/mic.0.069930-0}, author = {Zhang, Tian and Bain, Timothy S and Barlett, Melissa A and Dar, Shabir A and Snoeyenbos-West, Oona L and Nevin, Kelly P and Lovley, Derek R} } @article {1410, title = {Thermococcus paralvinellae sp. nov. and Thermococcus cleftensis sp. nov. of hyperthermophilic heterotrophs from deep-sea hydrothermal vents.}, journal = {Int J Syst Evol Microbiol}, volume = {64}, year = {2014}, month = {2014 Nov}, pages = {3655-9}, abstract = {Two heterotrophic hyperthermophilic strains, ES1(T) and CL1(T), were isolated from Paralvinella sp. polychaete worms collected from active hydrothermal vent chimneys in the north-eastern Pacific Ocean. Both were obligately anaerobic and produced H2S in the presence of elemental sulfur and H2. Complete genome sequences are available for both strains. Phylogenetic analyses based on 16S rRNA gene sequences showed that the strains are more than 97\% similar to most other species of the genus Thermococcus. Therefore, overall genome relatedness index analyses were performed to establish that these strains are novel species. For each analysis, strain ES1(T) was determined to be most similar to Thermococcus barophilus MP(T), while strain CL1(T) was determined to be most similar to Thermococcus sp. 4557. The average nucleotide identity scores for these strains were 84\% for strain ES1(T) and 81\% for strain CL1(T), genome-to-genome direct comparison scores were 23\% for strain ES1(T) and 47\% for strain CL1(T), and the species identification scores were 89\% for strain ES1(T) and 88\% for strain CL1(T). For each analysis, strains ES1(T) and CL1(T) were below the species delineation cut-off. Therefore, based on their whole genome sequences, strains ES1(T) and CL1(T) are suggested to represent novel species of the genus Thermococcus for which the names Thermococcus paralvinellae sp. nov. and Thermococcus cleftensis sp. nov. are proposed, respectively. The type strains are ES1(T) ( =DSM 27261(T) =KACC 17923(T)) and CL1(T) ( =DSM 27260(T) =KACC 17922(T)).
}, keywords = {Animals, DNA, Archaeal, Hydrothermal Vents, Molecular Sequence Data, Pacific Ocean, Phylogeny, Polychaeta, RNA, Ribosomal, 16S, Sequence Analysis, DNA, Thermococcus}, issn = {1466-5034}, doi = {10.1099/ijs.0.066100-0}, author = {Hensley, Sarah A and Jung, Jong-Hyun and Park, Cheon-Seok and Holden, James F} } @article {3116, title = {Visualization of charge propagation along individual pili proteins using ambient electrostatic force microscopy.}, journal = {Nat Nanotechnol}, volume = {9}, year = {2014}, month = {2014 Dec}, pages = {1012-7}, abstract = {The nanoscale imaging of charge flow in proteins is crucial to understanding several life processes, including respiration, metabolism and photosynthesis. However, existing imaging methods are only effective under non-physiological conditions or are limited to photosynthetic proteins. Here, we show that electrostatic force microscopy can be used to directly visualize charge propagation along pili of Geobacter sulfurreducens with nanometre resolution and under ambient conditions. Charges injected at a single point into individual, untreated pili, which are still attached to cells, propagated over the entire filament. The mobile charge density in the pili, as well as the temperature and pH dependence of the charge density, were similar to those of carbon nanotubes and other organic conductors. These findings, coupled with a lack of charge propagation in mutated pili that were missing key aromatic amino acids, suggest that the pili of G. sulfurreducens function as molecular wires with transport via delocalized charges, rather than the hopping mechanism that is typical of biological electron transport.
}, keywords = {Fimbriae Proteins, Fimbriae, Bacterial, Geobacter, Microscopy, Electrochemical, Scanning}, issn = {1748-3395}, doi = {10.1038/nnano.2014.236}, author = {Malvankar, Nikhil S and Yalcin, Sibel Ebru and Tuominen, Mark T and Lovley, Derek R} } @article {3136, title = {Anaerobic benzene oxidation via phenol in Geobacter metallireducens.}, journal = {Appl Environ Microbiol}, volume = {79}, year = {2013}, month = {2013 Dec}, pages = {7800-6}, abstract = {Anaerobic activation of benzene is expected to represent a novel biochemistry of environmental significance. Therefore, benzene metabolism was investigated in Geobacter metallireducens, the only genetically tractable organism known to anaerobically degrade benzene. Trace amounts (<0.5 μM) of phenol accumulated in cultures of Geobacter metallireducens anaerobically oxidizing benzene to carbon dioxide with the reduction of Fe(III). Phenol was not detected in cell-free controls or in Fe(II)- and benzene-containing cultures of Geobacter sulfurreducens, a Geobacter species that cannot metabolize benzene. The phenol produced in G. metallireducens cultures was labeled with (18)O during growth in H2(18)O, as expected for anaerobic conversion of benzene to phenol. Analysis of whole-genome gene expression patterns indicated that genes for phenol metabolism were upregulated during growth on benzene but that genes for benzoate or toluene metabolism were not, further suggesting that phenol was an intermediate in benzene metabolism. Deletion of the genes for PpsA or PpcB, subunits of two enzymes specifically required for the metabolism of phenol, removed the capacity for benzene metabolism. These results demonstrate that benzene hydroxylation to phenol is an alternative to carboxylation for anaerobic benzene activation and suggest that this may be an important metabolic route for benzene removal in petroleum-contaminated groundwaters, in which Geobacter species are considered to play an important role in anaerobic benzene degradation.
}, keywords = {Anaerobiosis, Benzene, Carbon Dioxide, Gene Deletion, Gene Expression Profiling, Geobacter, Iron, Metabolic Networks and Pathways, Oxidation-Reduction, Phenol, Water}, issn = {1098-5336}, doi = {10.1128/AEM.03134-13}, author = {Zhang, Tian and Tremblay, Pier-Luc and Chaurasia, Akhilesh Kumar and Smith, Jessica A and Bain, Timothy S and Lovley, Derek R} } @article {3142, title = {Aromatic amino acids required for pili conductivity and long-range extracellular electron transport in Geobacter sulfurreducens.}, journal = {mBio}, volume = {4}, year = {2013}, month = {2013 Mar 12}, pages = {e00105-13}, abstract = {UNLABELLED: It has been proposed that Geobacter sulfurreducens requires conductive pili for long-range electron transport to Fe(III) oxides and for high-density current production in microbial fuel cells. In order to investigate this further, we constructed a strain of G. sulfurreducens, designated Aro-5, which produced pili with diminished conductivity. This was accomplished by modifying the amino acid sequence of PilA, the structural pilin protein. An alanine was substituted for each of the five aromatic amino acids in the carboxyl terminus of PilA, the region in which G. sulfurreducens PilA differs most significantly from the PilAs of microorganisms incapable of long-range extracellular electron transport. Strain Aro-5 produced pili that were properly decorated with the multiheme c-type cytochrome OmcS, which is essential for Fe(III) oxide reduction. However, pili preparations of the Aro-5 strain had greatly diminished conductivity and Aro-5 cultures were severely limited in their capacity to reduce Fe(III) compared to the control strain. Current production of the Aro-5 strain, with a graphite anode serving as the electron acceptor, was less than 10\% of that of the control strain. The conductivity of the Aro-5 biofilms was 10-fold lower than the control strain{\textquoteright}s. These results demonstrate that the pili of G. sulfurreducens must be conductive in order for the cells to be effective in extracellular long-range electron transport.
IMPORTANCE: Extracellular electron transfer by Geobacter species plays an important role in the biogeochemistry of soils and sediments and has a number of bioenergy applications. For example, microbial reduction of Fe(III) oxide is one of the most geochemically significant processes in anaerobic soils, aquatic sediments, and aquifers, and Geobacter organisms are often abundant in such environments. Geobacter sulfurreducens produces the highest current densities of any known pure culture, and close relatives are often the most abundant organisms colonizing anodes in microbial fuel cells that harvest electricity from wastewater or aquatic sediments. The finding that a strain of G. sulfurreducens that produces pili with low conductivity is limited in these extracellular electron transport functions provides further insight into these environmentally significant processes.
}, keywords = {Amino Acids, Aromatic, Bioelectric Energy Sources, Biofilms, Electricity, Electrodes, Electron Transport, Ferric Compounds, Fimbriae Proteins, Fimbriae, Bacterial, Geobacter, Graphite}, issn = {2150-7511}, doi = {10.1128/mBio.00105-13}, author = {Vargas, Madeline and Malvankar, Nikhil S and Tremblay, Pier-Luc and Leang, Ching and Smith, Jessica A and Patel, Pranav and Snoeyenbos-West, Oona and Nevin, Kelly P and Lovley, Derek R} } @article {3153, title = {Bioremediation of uranium-contaminated groundwater: a systems approach to subsurface biogeochemistry.}, journal = {Curr Opin Biotechnol}, volume = {24}, year = {2013}, month = {2013 Jun}, pages = {489-97}, abstract = {Adding organic electron donors to stimulate microbial reduction of highly soluble U(VI) to less soluble U(IV) is a promising strategy for immobilizing uranium in contaminated subsurface environments. Studies suggest that diagnosing the in situ physiological status of the subsurface community during uranium bioremediation with environmental transcriptomic and proteomic techniques can identify factors potentially limiting U(VI) reduction activity. Models which couple genome-scale in silico representations of the metabolism of key microbial populations with geochemical and hydrological models may be able to predict the outcome of bioremediation strategies and aid in the development of new approaches. Concerns remain about the long-term stability of sequestered U(IV) minerals and the release of co-contaminants associated with Fe(III) oxides, which might be overcome through targeted delivery of electrons to select microorganisms using in situ electrodes.
}, keywords = {Biodegradation, Environmental, Electrodes, Ferric Compounds, Genomics, Groundwater, Proteomics, Uranium, Water Pollutants, Radioactive}, issn = {1879-0429}, doi = {10.1016/j.copbio.2012.10.008}, author = {Williams, Kenneth H and Bargar, John R and Lloyd, Jonathan R and Lovley, Derek R} } @article {1452, title = {Changes in microbial dynamics during long-term decomposition in tropical forests.}, journal = {Soil Biology and Biochemistry}, volume = {66}, year = {2013}, month = {2013 Jun}, pages = {60-68}, doi = {10.1016/j.soilbio.2013.06.010}, author = {DeAngelis, KM and Chivian, D and Fortney, J and Simmons, BA and Hazen, TC and Silver, WL} } @article {3133, title = {Characterization and modelling of interspecies electron transfer mechanisms and microbial community dynamics of a syntrophic association.}, journal = {Nat Commun}, volume = {4}, year = {2013}, month = {2013}, pages = {2809}, abstract = {Syntrophic associations are central to microbial communities and thus have a fundamental role in the global carbon cycle. Despite biochemical approaches describing the physiological activity of these communities, there has been a lack of a mechanistic understanding of the relationship between complex nutritional and energetic dependencies and their functioning. Here we apply a multi-omic modelling workflow that combines genomic, transcriptomic and physiological data with genome-scale models to investigate dynamics and electron flow mechanisms in the syntrophic association of Geobacter metallireducens and Geobacter sulfurreducens. Genome-scale modelling of direct interspecies electron transfer reveals insights into the energetics of electron transfer mechanisms. While G. sulfurreducens adapts to rapid syntrophic growth by changes at the genomic and transcriptomic level, G. metallireducens responds only at the transcriptomic level. This multi-omic approach enhances our understanding of adaptive responses and factors that shape the evolution of syntrophic communities.
}, keywords = {Adaptation, Biological, Biological Evolution, Electron Transport, Genome, Bacterial, Geobacter, Microbial Interactions, Models, Biological, Symbiosis, Transcriptome}, issn = {2041-1723}, doi = {10.1038/ncomms3809}, author = {Nagarajan, Harish and Embree, Mallory and Rotaru, Amelia-Elena and Shrestha, Pravin M and Feist, Adam M and Palsson, Bernhard {\O} and Lovley, Derek R and Zengler, Karsten} } @article {3154, title = {Characterization and transcription of arsenic respiration and resistance genes during in situ uranium bioremediation.}, journal = {ISME J}, volume = {7}, year = {2013}, month = {2013 Feb}, pages = {370-83}, abstract = {The possibility of arsenic release and the potential role of Geobacter in arsenic biogeochemistry during in situ uranium bioremediation was investigated because increased availability of organic matter has been associated with substantial releases of arsenic in other subsurface environments. In a field experiment conducted at the Rifle, CO study site, groundwater arsenic concentrations increased when acetate was added. The number of transcripts from arrA, which codes for the α-subunit of dissimilatory As(V) reductase, and acr3, which codes for the arsenic pump protein Acr3, were determined with quantitative reverse transcription-PCR. Most of the arrA (>60\%) and acr3-1 (>90\%) sequences that were recovered were most similar to Geobacter species, while the majority of acr3-2 (>50\%) sequences were most closely related to Rhodoferax ferrireducens. Analysis of transcript abundance demonstrated that transcription of acr3-1 by the subsurface Geobacter community was correlated with arsenic concentrations in the groundwater. In contrast, Geobacter arrA transcript numbers lagged behind the major arsenic release and remained high even after arsenic concentrations declined. This suggested that factors other than As(V) availability regulated the transcription of arrA in situ, even though the presence of As(V) increased the transcription of arrA in cultures of Geobacter lovleyi, which was capable of As(V) reduction. These results demonstrate that subsurface Geobacter species can tightly regulate their physiological response to changes in groundwater arsenic concentrations. The transcriptomic approach developed here should be useful for the study of a diversity of other environments in which Geobacter species are considered to have an important influence on arsenic biogeochemistry.
}, keywords = {Acetates, Arsenate Reductases, Arsenic, Biodegradation, Environmental, Colorado, Gene Expression Regulation, Bacterial, Genes, Bacterial, Geobacter, Groundwater, Transcriptome, Uranium}, issn = {1751-7370}, doi = {10.1038/ismej.2012.109}, author = {Giloteaux, Ludovic and Holmes, Dawn E and Williams, Kenneth H and Wrighton, Kelly C and Wilkins, Michael J and Montgomery, Alison P and Smith, Jessica A and Orellana, Roberto and Thompson, Courtney A and Roper, Thomas J and Long, Philip E and Lovley, Derek R} } @article {3132, title = {Characterizing acetogenic metabolism using a genome-scale metabolic reconstruction of Clostridium ljungdahlii.}, journal = {Microb Cell Fact}, volume = {12}, year = {2013}, month = {2013 Nov 25}, pages = {118}, abstract = {BACKGROUND: The metabolic capabilities of acetogens to ferment a wide range of sugars, to grow autotrophically on H2/CO2, and more importantly on synthesis gas (H2/CO/CO2) make them very attractive candidates as production hosts for biofuels and biocommodities. Acetogenic metabolism is considered one of the earliest modes of bacterial metabolism. A thorough understanding of various factors governing the metabolism, in particular energy conservation mechanisms, is critical for metabolic engineering of acetogens for targeted production of desired chemicals.
RESULTS: Here, we present the genome-scale metabolic network of Clostridium ljungdahlii, the first such model for an acetogen. This genome-scale model (iHN637) consisting of 637 genes, 785 reactions, and 698 metabolites captures all the major central metabolic and biosynthetic pathways, in particular pathways involved in carbon fixation and energy conservation. A combination of metabolic modeling, with physiological and transcriptomic data provided insights into autotrophic metabolism as well as aided the characterization of a nitrate reduction pathway in C. ljungdahlii. Analysis of the iHN637 metabolic model revealed that flavin based electron bifurcation played a key role in energy conservation during autotrophic growth and helped identify genes for some of the critical steps in this mechanism.
CONCLUSIONS: iHN637 represents a predictive model that recapitulates experimental data, and provides valuable insights into the metabolic response of C. ljungdahlii to genetic perturbations under various growth conditions. Thus, the model will be instrumental in guiding metabolic engineering of C. ljungdahlii for the industrial production of biocommodities and biofuels.
}, keywords = {Acetates, Biofuels, Clostridium, Genome, Metabolic Engineering, Metabolic Networks and Pathways}, issn = {1475-2859}, doi = {10.1186/1475-2859-12-118}, author = {Nagarajan, Harish and Sahin, Merve and Nogales, Juan and Latif, Haythem and Lovley, Derek R and Ebrahim, Ali and Zengler, Karsten} } @article {3140, title = {Characterizing the interplay between multiple levels of organization within bacterial sigma factor regulatory networks.}, journal = {Nat Commun}, volume = {4}, year = {2013}, month = {2013}, pages = {1755}, abstract = {Bacteria contain multiple sigma factors, each targeting diverse, but often overlapping sets of promoters, thereby forming a complex network. The layout and deployment of such a sigma factor network directly impacts global transcriptional regulation and ultimately dictates the phenotype. Here we integrate multi-omic data sets to determine the topology, the operational, and functional states of the sigma factor network in Geobacter sulfurreducens, revealing a unique network topology of interacting sigma factors. Analysis of the operational state of the sigma factor network shows a highly modular structure with σ(N) being the major regulator of energy metabolism. Surprisingly, the functional state of the network during the two most divergent growth conditions is nearly static, with sigma factor binding profiles almost invariant to environmental stimuli. This first comprehensive elucidation of the interplay between different levels of the sigma factor network organization is fundamental to characterize transcriptional regulatory mechanisms in bacteria.
}, keywords = {Energy Metabolism, Gene Expression Profiling, Gene Expression Regulation, Bacterial, Gene Regulatory Networks, Genes, Bacterial, Geobacter, Models, Biological, Regulon, Sigma Factor}, issn = {2041-1723}, doi = {10.1038/ncomms2743}, author = {Qiu, Yu and Nagarajan, Harish and Embree, Mallory and Shieu, Wendy and Abate, Elisa and Ju{\'a}rez, Katy and Cho, Byung-Kwan and Elkins, James G and Nevin, Kelly P and Barrett, Christian L and Lovley, Derek R and Palsson, Bernhard O and Zengler, Karsten} } @article {1966, title = {A Comparative Approach to Characterize the Landscape of Host-Pathogen Protein-Protein Interactions}, journal = {Journal of Visualized Experiments}, year = {2013}, month = {jul}, doi = {10.3791/50404}, url = {https://doi.org/10.3791/50404}, author = {Mandy Muller and Patricia Cassonnet and Michel Favre and Yves Jacob and Caroline Demeret} } @article {1423, title = {Conversion of the Amazon rainforest to agriculture results in biotic homogenization of soil bacterial communities.}, journal = {Proc Natl Acad Sci U S A}, volume = {110}, year = {2013}, month = {2013 Jan 15}, pages = {988-93}, abstract = {The Amazon rainforest is the Earth\&$\#$39;s largest reservoir of plant and animal diversity, and it has been subjected to especially high rates of land use change, primarily to cattle pasture. This conversion has had a strongly negative effect on biological diversity, reducing the number of plant and animal species and homogenizing communities. We report here that microbial biodiversity also responds strongly to conversion of the Amazon rainforest, but in a manner different from plants and animals. Local taxonomic and phylogenetic diversity of soil bacteria increases after conversion, but communities become more similar across space. This homogenization is driven by the loss of forest soil bacteria with restricted ranges (endemics) and results in a net loss of diversity. This study shows homogenization of microbial communities in response to human activities. Given that soil microbes represent the majority of biodiversity in terrestrial ecosystems and are intimately involved in ecosystem functions, we argue that microbial biodiversity loss should be taken into account when assessing the impact of land use change in tropical forests.
}, keywords = {Agriculture, Animals, Bacteria, Biodiversity, Brazil, Cattle, Ecosystem, Humans, Phylogeny, Rain, Soil Microbiology, Trees, Tropical Climate}, issn = {1091-6490}, doi = {10.1073/pnas.1220608110}, author = {Rodrigues, Jorge L M and Pellizari, Vivian H and Mueller, Rebecca and Baek, Kyunghwa and Jesus, Ederson da C and Paula, Fabiana S and Mirza, Babur and Hamaoui, George S and Tsai, Siu Mui and Feigl, Brigitte and Tiedje, James M and Bohannan, Brendan J M and N{\"u}sslein, Klaus} } @article {935, title = {Critical roles for lipomannan and lipoarabinomannan in cell wall integrity of mycobacteria and pathogenesis of tuberculosis.}, journal = {MBio}, volume = {4}, year = {2013}, month = {2013}, abstract = {ABSTRACT Lipomannan (LM) and lipoarabinomannan (LAM) are mycobacterial glycolipids containing a long mannose polymer. While they are implicated in immune modulations, the significance of LM and LAM as structural components of the mycobacterial cell wall remains unknown. We have previously reported that a branch-forming mannosyltransferase plays a critical role in controlling the sizes of LM and LAM and that deletion or overexpression of this enzyme results in gross changes in LM/LAM structures. Here, we show that such changes in LM/LAM structures have a significant impact on the cell wall integrity of mycobacteria. In Mycobacterium smegmatis, structural defects in LM and LAM resulted in loss of acid-fast staining, increased sensitivity to \β-lactam antibiotics, and faster killing by THP-1 macrophages. Furthermore, equivalent Mycobacterium\ tuberculosis mutants became more sensitive to \β-lactams, and one mutant showed attenuated virulence in mice. Our results revealed previously unknown structural roles for LM and LAM and further demonstrated that they are important for the pathogenesis of tuberculosis. IMPORTANCE Tuberculosis (TB) is a global burden, affecting millions of people worldwide. Mycobacterium tuberculosis is a causative agent of TB, and understanding the biology of M.\ tuberculosis is essential for tackling this devastating disease. The cell wall of M.\ tuberculosis is highly impermeable and plays a protective role in establishing infection. Among the cell wall components, LM and LAM are major glycolipids found in all Mycobacterium species, show various immunomodulatory activities, and have been thought to play roles in TB pathogenesis. However, the roles of LM and LAM as integral parts of the cell wall structure have not been elucidated. Here we show that LM and LAM play critical roles in the integrity of mycobacterial cell wall and the pathogenesis of TB. These findings will now allow us to seek the possibility that the LM/LAM biosynthetic pathway is a chemotherapeutic target.
}, issn = {2150-7511}, doi = {10.1128/mBio.00472-12}, author = {Fukuda, Takeshi and Matsumura, Takayuki and Ato, Manabu and Hamasaki, Maho and Nishiuchi, Yukiko and Murakami, Yoshiko and Maeda, Yusuke and Yoshimori, Tamotsu and Matsumoto, Sohkichi and Kobayashi, Kazuo and Kinoshita, Taroh and Morita, Yasu S} } @article {1445, title = {(D)-Amino acid chemical reporters reveal peptidoglycan dynamics of an intracellular pathogen.}, journal = {ACS Chem Biol}, volume = {8}, year = {2013}, month = {2013 Mar 15}, pages = {500-5}, abstract = {Peptidoglycan (PG) is an essential component of the bacterial cell wall. Although experiments with organisms in vitro have yielded a wealth of information on PG synthesis and maturation, it is unclear how these studies translate to bacteria replicating within host cells. We report a chemical approach for probing PG in vivo via metabolic labeling and bioorthogonal chemistry. A wide variety of bacterial species incorporated azide and alkyne-functionalized d-alanine into their cell walls, which we visualized by covalent reaction with click chemistry probes. The d-alanine analogues were specifically incorporated into nascent PG of the intracellular pathogen Listeria monocytogenes both in vitro and during macrophage infection. Metabolic incorporation of d-alanine derivatives and click chemistry detection constitute a facile, modular platform that facilitates unprecedented spatial and temporal resolution of PG dynamics in vivo.
}, keywords = {Alanine, Click Chemistry, Listeria monocytogenes, Molecular Conformation, Molecular Dynamics Simulation, Peptidoglycan}, issn = {1554-8937}, doi = {10.1021/cb3004995}, author = {Siegrist, M Sloan and Whiteside, Sarah and Jewett, John C and Aditham, Arjun and Cava, Felipe and Bertozzi, Carolyn R} } @article {3144, title = {Electrobiocommodities: powering microbial production of fuels and commodity chemicals from carbon dioxide with electricity.}, journal = {Curr Opin Biotechnol}, volume = {24}, year = {2013}, month = {2013 Jun}, pages = {385-90}, abstract = {Electricity can be an energy source for microbially catalyzed production of fuels and other organic commodities from carbon dioxide. These electrobiocommodities (E-BCs) can be produced directly via electrode-to-microbe electron transfer or indirectly with electrochemically generated electron donors such as H2 or formate. Producing E-BCs may be a more efficient and environmentally sustainable strategy for converting solar energy to biocommodities than approaches that rely on biological photosynthesis. A diversity of microbial physiologies could potentially be adapted for E-BC production, but to date acetogenic microorganisms are the only organisms shown to covert electrically generated low potential electrons and carbon dioxide into multi-carbon organic products with high recovery of electrons in product. Substantial research and development will be required for E-BC commercialization.
}, keywords = {Bioelectric Energy Sources, Carbon Dioxide, Cell Respiration, Chemical Industry, Electricity, Methane, Microbiology, Oxygen, Renewable Energy}, issn = {1879-0429}, doi = {10.1016/j.copbio.2013.02.012}, author = {Lovley, Derek R and Nevin, Kelly P} } @article {3145, title = {Enrichment of specific protozoan populations during in situ bioremediation of uranium-contaminated groundwater.}, journal = {ISME J}, volume = {7}, year = {2013}, month = {2013 Jul}, pages = {1286-98}, abstract = {The importance of bacteria in the anaerobic bioremediation of groundwater polluted with organic and/or metal contaminants is well recognized and in some instances so well understood that modeling of the in situ metabolic activity of the relevant subsurface microorganisms in response to changes in subsurface geochemistry is feasible. However, a potentially significant factor influencing bacterial growth and activity in the subsurface that has not been adequately addressed is protozoan predation of the microorganisms responsible for bioremediation. In field experiments at a uranium-contaminated aquifer located in Rifle, CO, USA, acetate amendments initially promoted the growth of metal-reducing Geobacter species, followed by the growth of sulfate reducers, as observed previously. Analysis of 18S rRNA gene sequences revealed a broad diversity of sequences closely related to known bacteriovorous protozoa in the groundwater before the addition of acetate. The bloom of Geobacter species was accompanied by a specific enrichment of sequences most closely related to the ameboid flagellate, Breviata anathema, which at their peak accounted for over 80\% of the sequences recovered. The abundance of Geobacter species declined following the rapid emergence of B. anathema. The subsequent growth of sulfate-reducing Peptococcaceae was accompanied by another specific enrichment of protozoa, but with sequences most similar to diplomonadid flagellates from the family Hexamitidae, which accounted for up to 100\% of the sequences recovered during this phase of the bioremediation. These results suggest a prey-predator response with specific protozoa responding to increased availability of preferred prey bacteria. Thus, quantifying the influence of protozoan predation on the growth, activity and composition of the subsurface bacterial community is essential for predictive modeling of in situ uranium bioremediation strategies.
}, keywords = {Acetates, Biodegradation, Environmental, Eukaryota, Geobacter, Groundwater, Molecular Sequence Data, Oxidation-Reduction, Phylogeny, RNA, Ribosomal, 16S, RNA, Ribosomal, 18S, Uranium}, issn = {1751-7370}, doi = {10.1038/ismej.2013.20}, author = {Holmes, Dawn E and Giloteaux, Ludovic and Williams, Kenneth H and Wrighton, Kelly C and Wilkins, Michael J and Thompson, Courtney A and Roper, Thomas J and Long, Philip E and Lovley, Derek R} } @article {1415, title = {Evidence supporting dissimilatory and assimilatory lignin degradation in Enterobacter lignolyticus SCF1.}, journal = {Front Microbiol}, volume = {4}, year = {2013}, month = {2013}, pages = {280}, abstract = {Lignocellulosic biofuels are promising as sustainable alternative fuels, but lignin inhibits access of enzymes to cellulose, and by-products of lignin degradation can be toxic to cells. The fast growth, high efficiency and specificity of enzymes employed in the anaerobic litter deconstruction carried out by tropical soil bacteria make these organisms useful templates for improving biofuel production. The facultative anaerobe Enterobacter lignolyticus SCF1 was initially cultivated from Cloud Forest soils in the Luquillo Experimental Forest in Puerto Rico, based on anaerobic growth on lignin as sole carbon source. The source of the isolate was tropical forest soils that decompose litter rapidly with low and fluctuating redox potentials, where bacteria using oxygen-independent enzymes likely play an important role in decomposition. We have used transcriptomics and proteomics to examine the observed increased growth of SCF1 grown on media amended with lignin compared to unamended growth. Proteomics suggested accelerated xylose uptake and metabolism under lignin-amended growth, with up-regulation of proteins involved in lignin degradation via the 4-hydroxyphenylacetate degradation pathway, catalase/peroxidase enzymes, and the glutathione biosynthesis and glutathione S-transferase (GST) proteins. We also observed increased production of NADH-quinone oxidoreductase, other electron transport chain proteins, and ATP synthase and ATP-binding cassette (ABC) transporters. This suggested the use of lignin as terminal electron acceptor. We detected significant lignin degradation over time by absorbance, and also used metabolomics to demonstrate moderately significant decreased xylose concentrations as well as increased metabolic products acetate and formate in stationary phase in lignin-amended compared to unamended growth conditions. Our data show the advantages of a multi-omics approach toward providing insights as to how lignin may be used in nature by microorganisms coping with poor carbon availability.
}, issn = {1664-302X}, doi = {10.3389/fmicb.2013.00280}, author = {Deangelis, Kristen M and Sharma, Deepak and Varney, Rebecca and Simmons, Blake and Isern, Nancy G and Markilllie, LM and Nicora, Carrie and Norbeck, Angela D and Taylor, Ronald C and Aldrich, Joshua T and Robinson, Errol W} } @article {3138, title = {Field evidence of selenium bioreduction in a uranium-contaminated aquifer.}, journal = {Environ Microbiol Rep}, volume = {5}, year = {2013}, month = {2013 Jun}, pages = {444-52}, abstract = {Removal of selenium from groundwater was documented during injection of acetate into a uranium-contaminated aquifer near Rifle, Colorado (USA). Bioreduction of aqueous selenium to its elemental form (Se0) concentrated it within mineralized biofilms affixed to tubing used to circulate acetate-amended groundwater. Scanning and transmission electron microscopy revealed close association between Se0 precipitates and cell surfaces, with Se0 aggregates having a diameter of 50-60 nm. Accumulation of Se0 within biofilms occurred over a three-week interval at a rate of c. 9 mg Se0 m(-2) tubing day(-1). Removal was inferred to result from the activity of a mixed microbial community within the biofilms capable of coupling acetate oxidation to the reduction of oxygen, nitrate and selenate. Phylogenetic analysis of the biofilm revealed a community dominated by strains of Dechloromonas sp. and Thauera sp., with isolates exhibiting genetic similarity to the latter known to reduce selenate to Se0. Enrichment cultures of selenate-respiring microorganisms were readily established using Rifle site groundwater and acetate, with cultures dominated by strains closely related to D. aromatica (96-99\% similarity). Predominance of Dechloromonas sp. in recovered biofilms and enrichments suggests this microorganism may play a role in the removal of selenium oxyanions present in Se-impacted groundwaters and sediments.
}, keywords = {Acetates, Betaproteobacteria, Biodegradation, Environmental, Biofilms, Colorado, Groundwater, Humans, Microbial Consortia, Oxidation-Reduction, Oxygen, Phylogeny, RNA, Ribosomal, 16S, Selenic Acid, Selenium, Selenium Compounds, Thauera, Uranium, Water Pollutants, Chemical}, issn = {1758-2229}, doi = {10.1111/1758-2229.12032}, author = {Williams, Kenneth H and Wilkins, Michael J and N{\textquoteright}Guessan, A Lucie and Arey, Bruce and Dodova, Elena and Dohnalkova, Alice and Holmes, Dawn and Lovley, Derek R and Long, Philip E} } @article {1564, title = {A field trial of nutrient stimulation of methanotrophs to reduce greenhouse gas emissions from landfill cover soils.}, journal = {J Air Waste Manag Assoc}, volume = {63}, year = {2013}, month = {2013 Mar}, pages = {300-9}, abstract = {UNLABELLED: Landfills are among the major sources of anthropogenic methane (CH4) estimated to reach 40 x 10(9) kg per year worldwide by 2015 (IPCC, 2007). A 2 1/2-year field experiment was conducted at a closed landfill in western Michigan where methanotrophs, methane-consuming bacteria, were stimulated by nutrient addition to the soil without significantly increasing biogenic nitrous oxide (N2O) production. The effects of the nitrogen amendments (KNO3 and NH4Cl), phenylacetylene (a selective inhibitor of nitrifying bacteria that contribute to N2O production), and a canopy (to reduce direct water infiltration) on the vertical soil gas profiles of CH4, CO2, and O2 were measured in the top meter of the soil. Methane and nitrous oxide fluxes were calculated from the corresponding soil gas concentration gradients with respect to depth and a Millington-Quirk diffusivity coefficient in soil derived empirically from soil porosity, water content, and diffusivity coefficients in air from the literature. Methane flux estimates were as high as 218.4 g m(-2) day(-1) in the fall and 12.8 g/m(-2) day(-1) in the summer. During the spring and summer CH4 fluxes were reduced by more than half by adding KNO3 and NH4Cl into the soil as compared to control plots, while N2O fluxes increased substantially. The concurrent addition of phenylacetylene to the amendment decreased peak N2O production by half and the rate of peak methane oxidation by about one-third. The seasonal average methane and N2O flux data were extrapolated to estimate the reduction of CH4 and N2O fluxes into the atmosphere by nitrogen and inhibitor addition to the cover soils. The results suggest that such additions coupled with soil moisture management may provide a potential strategy to significantly reduce greenhouse gas emissions from landfills.
IMPLICATIONS: The results of a 2 1/2-year study of effects of nutrient stimulation on methane oxidation in landfill cover soils demonstrates that nutrient addition does decrease methane emissions. The work further underscores the control which soil moisture exerts on methane oxidation. Water management is critical to the success of methane oxidation strategies.
}, keywords = {Biodegradation, Environmental, Gases, Greenhouse Effect, Methane, Oxidation-Reduction, Soil, Soil Microbiology, Waste Management}, issn = {1096-2247}, author = {Lizik, William and Im, Jeongdae and Semrau, Jeremy D and Barcelona, Michael J} } @article {3143, title = {Fluctuations in species-level protein expression occur during element and nutrient cycling in the subsurface.}, journal = {PLoS One}, volume = {8}, year = {2013}, month = {2013}, pages = {e57819}, abstract = {While microbial activities in environmental systems play a key role in the utilization and cycling of essential elements and compounds, microbial activity and growth frequently fluctuates in response to environmental stimuli and perturbations. To investigate these fluctuations within a saturated aquifer system, we monitored a carbon-stimulated in situ Geobacter population while iron reduction was occurring, using 16S rRNA abundances and high-resolution tandem mass spectrometry proteome measurements. Following carbon amendment, 16S rRNA analysis of temporally separated samples revealed the rapid enrichment of Geobacter-like environmental strains with strong similarity to G. bemidjiensis. Tandem mass spectrometry proteomics measurements suggest high carbon flux through Geobacter respiratory pathways, and the synthesis of anapleurotic four carbon compounds from acetyl-CoA via pyruvate ferredoxin oxidoreductase activity. Across a 40-day period where Fe(III) reduction was occurring, fluctuations in protein expression reflected changes in anabolic versus catabolic reactions, with increased levels of biosynthesis occurring soon after acetate arrival in the aquifer. In addition, localized shifts in nutrient limitation were inferred based on expression of nitrogenase enzymes and phosphate uptake proteins. These temporal data offer the first example of differing microbial protein expression associated with changing geochemical conditions in a subsurface environment.
}, keywords = {Biomass, Carbon, Environment, Gene Expression Regulation, Bacterial, Geobacter, Groundwater, Humic Substances, Iron, Oxidation-Reduction, Phosphates, Plankton, Proteomics, RNA, Ribosomal, 16S, Tandem Mass Spectrometry, Uranium, Vanadium, Water Microbiology}, issn = {1932-6203}, doi = {10.1371/journal.pone.0057819}, author = {Wilkins, Michael J and Wrighton, Kelly C and Nicora, Carrie D and Williams, Kenneth H and McCue, Lee Ann and Handley, Kim M and Miller, Chris S and Giloteaux, Ludovic and Montgomery, Alison P and Lovley, Derek R and Banfield, Jillian F and Long, Philip E and Lipton, Mary S} } @article {3150, title = {A genetic system for Clostridium ljungdahlii: a chassis for autotrophic production of biocommodities and a model homoacetogen.}, journal = {Appl Environ Microbiol}, volume = {79}, year = {2013}, month = {2013 Feb}, pages = {1102-9}, abstract = {Methods for genetic manipulation of Clostridium ljungdahlii are of interest because of the potential for production of fuels and other biocommodities from carbon dioxide via microbial electrosynthesis or more traditional modes of autotrophy with hydrogen or carbon monoxide as the electron donor. Furthermore, acetogenesis plays an important role in the global carbon cycle. Gene deletion strategies required for physiological studies of C. ljungdahlii have not previously been demonstrated. An electroporation procedure for introducing plasmids was optimized, and four different replicative origins for plasmid propagation in C. ljungdahlii were identified. Chromosomal gene deletion via double-crossover homologous recombination with a suicide vector was demonstrated initially with deletion of the gene for FliA, a putative sigma factor involved in flagellar biogenesis and motility in C. ljungdahlii. Deletion of fliA yielded a strain that lacked flagella and was not motile. To evaluate the potential utility of gene deletions for functional genomic studies and to redirect carbon and electron flow, the genes for the putative bifunctional aldehyde/alcohol dehydrogenases, adhE1 and adhE2, were deleted individually or together. Deletion of adhE1, but not adhE2, diminished ethanol production with a corresponding carbon recovery in acetate. The double deletion mutant had a phenotype similar to that of the adhE1-deficient strain. Expression of adhE1 in trans partially restored the capacity for ethanol production. These results demonstrate the feasibility of genetic investigations of acetogen physiology and the potential for genetic manipulation of C. ljungdahlii to optimize autotrophic biocommodity production.
}, keywords = {Clostridium, Electroporation, Gene Deletion, Genetic Complementation Test, Genetic Vectors, Genetics, Microbial, Metabolic Engineering, Molecular Biology, Plasmids, Transformation, Bacterial}, issn = {1098-5336}, doi = {10.1128/AEM.02891-12}, author = {Leang, Ching and Ueki, Toshiyuki and Nevin, Kelly P and Lovley, Derek R} } @article {1202, title = {Growth kinetics and energetics of a deep-sea hyperthermophilic methanogen under varying environmental conditions.}, journal = {Environ Microbiol Rep}, volume = {5}, year = {2013}, month = {2013 Oct}, pages = {665-71}, abstract = {A hyperthermophilic deep-sea methanogen, Methanocaldococcus strain JH146, was isolated from 26{\textdegree}C hydrothermal fluid at Axial Volcano to model high temperature methanogenesis in the subseafloor. Emphasis was placed on defining growth kinetics, cell yields and growth energy demand (GE) across a range of conditions. The organism uses H2 and CO2 as its sole carbon and energy sources. At various temperatures, pHs, and chlorinities, its growth rates and cell yields co-varied while GE remained uniform at 1.69 {\texttimes} 10(-11) J cell(-1)s(-1) {\textpm} 0.68 {\texttimes} 10(-11) J cell(-1)s(-1) (s.d., n = 23). An exception was at superoptimal growth temperatures where GE increased to 7.25 {\texttimes} 10(-11) J cell(-1)s(-1) presumably due to heat shock. GE also increased from 5.1 {\texttimes} 10(-12) J cell(-1)s(-1) to 7.61 {\texttimes} 10(-11) J cell(-1)s(-1) as NH4 (+) concentrations decreased from 9.4 mM to 0.14 mM. JH146 did not fix N2 or assimilate NO3 (-), lacked the N2-fixing (cluster II) nifH gene, and became nitrogen limited below 0.14 mM NH4Cl. Nitrogen availability may impact growth in situ since ammonia concentrations at Axial Volcano are < 18 μM. Our approach contributes to refining bioenergetic and carbon flux models for methanogens and other organisms in hydrothermal vents and other environments.
}, issn = {1758-2229}, doi = {10.1111/1758-2229.12065}, author = {Ver Eecke, Helene C and Akerman, Nancy H and Huber, Julie A and Butterfield, David A and Holden, James F} } @article {1565, title = {Guided cobalamin biosynthesis supports Dehalococcoides mccartyi reductive dechlorination activity.}, journal = {Philos Trans R Soc Lond B Biol Sci}, volume = {368}, year = {2013}, month = {2013 Apr 19}, pages = {20120320}, abstract = {Dehalococcoides mccartyi strains are corrinoid-auxotrophic Bacteria and axenic cultures that require vitamin B12 (CN-Cbl) to conserve energy via organohalide respiration. Cultures of D. mccartyi strains BAV1, GT and FL2 grown with limiting amounts of 1 {\textmu}g l(-1) CN-Cbl quickly depleted CN-Cbl, and reductive dechlorination of polychlorinated ethenes was incomplete leading to vinyl chloride (VC) accumulation. In contrast, the same cultures amended with 25 {\textmu}g l(-1) CN-Cbl exhibited up to 2.3-fold higher dechlorination rates, 2.8-9.1-fold increased growth yields, and completely consumed growth-supporting chlorinated ethenes. To explore whether known cobamide-producing microbes supply Dehalococcoides with the required corrinoid cofactor, co-culture experiments were performed with the methanogen Methanosarcina barkeri strain Fusaro and two acetogens, Sporomusa ovata and Sporomusa sp. strain KB-1, as Dehalococcoides partner populations. During growth with H2/CO2, M. barkeri axenic cultures produced 4.2 {\textpm} 0.1 {\textmu}g l(-1) extracellular cobamide (factor III), whereas the Sporomusa cultures produced phenolyl- and p-cresolyl-cobamides. Neither factor III nor the phenolic cobamides supported Dehalococcoides reductive dechlorination activity suggesting that M. barkeri and the Sporomusa sp. cannot fulfil Dehalococcoides{\textquoteright} nutritional requirements. Dehalococcoides dechlorination activity and growth occurred in M. barkeri and Sporomusa sp. co-cultures amended with 10 {\textmu}M 5{\textquoteright},6{\textquoteright}-dimethylbenzimidazole (DMB), indicating that a cobalamin is a preferred corrinoid cofactor of strains BAV1, GT and FL2 when grown with chlorinated ethenes as electron acceptors. Even though the methanogen and acetogen populations tested did not produce cobalamin, the addition of DMB enabled guided biosynthesis and generated a cobalamin that supported Dehalococcoides{\textquoteright} activity and growth. Guided cobalamin biosynthesis may offer opportunities to sustain and enhance Dehalococcoides activity in contaminated subsurface environments.
}, keywords = {Bacteriological Techniques, Benzimidazoles, Biodegradation, Environmental, Chloroflexi, Chromatography, High Pressure Liquid, Coculture Techniques, Culture Media, Dichloroethylenes, Halogenation, Oxidation-Reduction, Trichloroethylene, Vitamin B 12}, issn = {1471-2970}, doi = {10.1098/rstb.2012.0320}, author = {Yan, Jun and Im, Jeongdae and Yang, Yi and L{\"o}ffler, Frank E} } @article {1422, title = {Identification and characterization of a NaCl-responsive genetic locus involved in survival during desiccation in Sinorhizobium meliloti.}, journal = {Appl Environ Microbiol}, volume = {79}, year = {2013}, month = {2013 Sep}, pages = {5693-700}, abstract = {The Rhizobiaceae are a bacterial family of enormous agricultural importance due to the ability of its members to fix atmospheric nitrogen in an intimate relationship with plants. Their survival as naturally occurring soil bacteria in agricultural soils as well as popular seed inocula is affected directly by drought and salinity. Survival after desiccation in the presence of NaCl is enabled by underlying genetic mechanisms in the model organism Sinorhizobium meliloti 1021. Since salt stress parallels a loss in water activity, the identification of NaCl-responsive loci may identify loci involved in survival during desiccation. This approach enabled identification of the loci asnO and ngg by their reduced ability to grow on increased NaCl concentrations, likely due to their inability to produce the osmoprotectant N-acetylglutaminylglutamine (NAGGN). In addition, the mutant harboring ngg::Tn5luxAB was affected in its ability to survive desiccation and responded to osmotic stress. The desiccation sensitivity may have been due to secondary functions of Ngg (N-acetylglutaminylglutamine synthetase)-like cell wall metabolism as suggested by the presence of a d-alanine-d-alanine ligase (dAla-dAla) domain and by sensitivity of the mutant to \β-lactam antibiotics. asnO::Tn5luxAB is expressed during the stationary phase under normal growth conditions. Amino acid sequence similarity to enzymes producing \β-lactam inhibitors and increased resistance to \β-lactam antibiotics may indicate that asnO is involved in the production of a \β-lactam inhibitor.
}, keywords = {Desiccation, DNA Transposable Elements, Gene Expression Regulation, Bacterial, Gene Knockout Techniques, Genes, Bacterial, Genetic Loci, Microbial Viability, Mutagenesis, Insertional, Osmotic Pressure, Sinorhizobium meliloti, Sodium Chloride, Stress, Physiological}, issn = {1098-5336}, doi = {10.1128/AEM.01037-13}, author = {Vriezen, Jan A C and de Bruijn, Frans J and N{\"u}sslein, Klaus} } @article {3139, title = {Improved cathode for high efficient microbial-catalyzed reduction in microbial electrosynthesis cells.}, journal = {Phys Chem Chem Phys}, volume = {15}, year = {2013}, month = {2013 Sep 14}, pages = {14290-4}, abstract = {Microbial electrosynthesis cells (MECs) are devices wherein microorganisms can electrochemically interact with electrodes, directly donating or accepting electrons from electrode surfaces. Here, we developed a novel cathode by using nickel nanowires anchored to graphite for the improvement of microbial-catalyzed reduction in MEC cathode chamber. This porous nickel-nanowire-network-coated graphite electrode increased the interfacial area and interfacial interactions between the cathode surface and the microbial biofilm. A 2.3 fold increase in bio-reduction rate over the untreated graphite was observed. Around 282 mM day(-1) m(-2) of acetate resulting from the bio-reduction of carbon dioxide by Sporomusa was produced with 82 {\textpm} 14\% of the electrons consumed being recovered in acetate.
}, keywords = {Biocatalysis, Bioelectric Energy Sources, Biofilms, Carbon Dioxide, Electrochemical Techniques, Electrodes, Graphite, Nanowires, Nickel, Oxidation-Reduction, Veillonellaceae}, issn = {1463-9084}, doi = {10.1039/c3cp52697f}, author = {Nie, Huarong and Zhang, Tian and Cui, Mengmeng and Lu, Haiyun and Lovley, Derek R and Russell, Thomas P} } @article {1563, title = {Interference of ferric ions with ferrous iron quantification using the ferrozine assay.}, journal = {J Microbiol Methods}, volume = {95}, year = {2013}, month = {2013 Dec}, pages = {366-7}, abstract = {The ferrozine assay is a widely used colorimetric method for determining soluble iron concentrations. We provide evidence for a heretofore unrecognized interference of ferric ions (Fe(3+)) on ferrous iron (Fe(2+)) measurements performed in the dark. Fe(3+) concentrations affected the absorbance measurements, which linearly increased with incubation time.
}, keywords = {Chemistry Techniques, Analytical, Colorimetry, Darkness, Diagnostic Errors, Ferric Compounds, Ferrous Compounds, Ferrozine, Ions, Iron, Time Factors}, issn = {1872-8359}, doi = {10.1016/j.mimet.2013.10.005}, author = {Im, Jeongdae and Lee, Jaejin and L{\"o}ffler, Frank E} } @article {1030, title = {Metagenomes of tropical soil-derived anaerobic switchgrass-adapted consortia with and without iron.}, journal = {Stand Genomic Sci}, volume = {7}, year = {2013}, month = {2013}, pages = {382-98}, abstract = {Tropical forest soils decompose litter rapidly with frequent episodes of anoxia, making it likely that bacteria using alternate terminal electron acceptors (TEAs) such as iron play a large role in supporting decomposition under these conditions. The prevalence of many types of metabolism in litter deconstruction makes these soils useful templates for improving biofuel production. To investigate how iron availability affects decomposition, we cultivated feedstock-adapted consortia (FACs) derived from iron-rich tropical forest soils accustomed to experiencing frequent episodes of anaerobic conditions and frequently fluctuating redox. One consortium was propagated under fermenting conditions, with switchgrass as the sole carbon source in minimal media (SG only FACs), and the other consortium was treated the same way but received poorly crystalline iron as an additional terminal electron acceptor (SG + Fe FACs). We sequenced the metagenomes of both consortia to a depth of about 150 Mb each, resulting in a coverage of 26\× for the more diverse SG + Fe FACs, and 81\× for the relatively less diverse SG only FACs. Both consortia were able to quickly grow on switchgrass, and the iron-amended consortium exhibited significantly higher microbial diversity than the unamended consortium. We found evidence of higher stress in the unamended FACs and increased sugar transport and utilization in the iron-amended FACs. This work provides metagenomic evidence that supplementation of alternative TEAs may improve feedstock deconstruction in biofuel production.
}, issn = {1944-3277}, doi = {10.4056/sigs.3377516}, author = {Deangelis, Kristen M and D{\textquoteright}haeseleer, Patrik and Chivian, Dylan and Simmons, Blake and Arkin, Adam P and Mavromatis, Konstantinos and Malfatti, Stephanie and Tringe, Susannah and Hazen, Terry C} } @article {3147, title = {Molecular analysis of the in situ growth rates of subsurface Geobacter species.}, journal = {Appl Environ Microbiol}, volume = {79}, year = {2013}, month = {2013 Mar}, pages = {1646-53}, abstract = {Molecular tools that can provide an estimate of the in situ growth rate of Geobacter species could improve understanding of dissimilatory metal reduction in a diversity of environments. Whole-genome microarray analyses of a subsurface isolate of Geobacter uraniireducens, grown under a variety of conditions, identified a number of genes that are differentially expressed at different specific growth rates. Expression of two genes encoding ribosomal proteins, rpsC and rplL, was further evaluated with quantitative reverse transcription-PCR (qRT-PCR) in cells with doubling times ranging from 6.56 h to 89.28 h. Transcript abundance of rpsC correlated best (r(2) = 0.90) with specific growth rates. Therefore, expression patterns of rpsC were used to estimate specific growth rates of Geobacter species during an in situ uranium bioremediation field experiment in which acetate was added to the groundwater to promote dissimilatory metal reduction. Initially, increased availability of acetate in the groundwater resulted in higher expression of Geobacter rpsC, and the increase in the number of Geobacter cells estimated with fluorescent in situ hybridization compared well with specific growth rates estimated from levels of in situ rpsC expression. However, in later phases, cell number increases were substantially lower than predicted from rpsC transcript abundance. This change coincided with a bloom of protozoa and increased attachment of Geobacter species to solid phases. These results suggest that monitoring rpsC expression may better reflect the actual rate that Geobacter species are metabolizing and growing during in situ uranium bioremediation than changes in cell abundance.
}, keywords = {Acetates, Biodegradation, Environmental, DNA, Bacterial, Gene Expression Profiling, Geobacter, Groundwater, In Situ Hybridization, Fluorescence, Molecular Sequence Data, Ribosomal Proteins, Sequence Analysis, DNA, Uranium}, issn = {1098-5336}, doi = {10.1128/AEM.03263-12}, author = {Holmes, Dawn E and Giloteaux, Ludovic and Barlett, Melissa and Chavan, Milind A and Smith, Jessica A and Williams, Kenneth H and Wilkins, Michael and Long, Philip and Lovley, Derek R} } @article {1204, title = {Molecular cloning and enzymatic characterization of cyclomaltodextrinase from hyperthermophilic archaeon Thermococcus sp. CL1.}, journal = {J Microbiol Biotechnol}, volume = {23}, year = {2013}, month = {2013 Aug}, pages = {1060-9}, abstract = {Genome organization near cyclomaltodextrinases (CDases) was analyzed and compared for four different hyperthermophilic archaea: Thermococcus, Pyrococcus, Staphylothermus, and Thermofilum. A gene (CL1_0884) encoding a putative CDase from Thermococcus sp. CL1 (tccd) was cloned and expressed in Escherichia coli. TcCD was confirmed to be highly thermostable, with optimal activity at 85{\textcelsius}. The melting temperature of TcCD was determined to be 93oC by both differential scanning calorimetry and differential scanning fluorimetry. A size-exclusion chromatography experiment showed that TcCD exists as a monomer. TcCD preferentially hydrolyzed α-cyclodextrin (α-CD), and at the initial stage catalyzed a ring-opening reaction by cleaving one α-1,4-glycosidic linkage of the CD ring to produce the corresponding single maltooligosaccharide. Furthermore, TcCD could hydrolyze branched CDs (G1-α-CD, G1-β- CD, and G2-β-CD) to yield significant amounts (45\%, 40\%, and 46\%) of isomaltooligosaccharides (panose and 6(2)-α-maltosylmaltose) in addition to glucose and maltose. This enzyme is one of the most thermostable maltogenic amylases reported, and might be of potential value in the production of isomaltooligosaccharides in the food industry.
}, issn = {1738-8872}, author = {Lee, Jae-Eun and Kim, In-Hwan and Jung, Jong-Hyun and Seo, Dong-Ho and Kang, Sung-Gyun and Holden, James F and Cha, Jaeho and Park, Cheon-Seok} } @article {1444, title = {Osmosensory signaling in Mycobacterium tuberculosis mediated by a eukaryotic-like Ser/Thr protein kinase.}, journal = {Proc Natl Acad Sci U S A}, volume = {110}, year = {2013}, month = {2013 Dec 24}, pages = {E5069-77}, abstract = {Bacteria are able to adapt to dramatically different microenvironments, but in many organisms, the signaling pathways, transcriptional programs, and downstream physiological changes involved in adaptation are not well-understood. Here, we discovered that osmotic stress stimulates a signaling network in Mycobacterium tuberculosis regulated by the eukaryotic-like receptor Ser/Thr protein kinase PknD. Expression of the PknD substrate Rv0516c was highly induced by osmotic stress. Furthermore, Rv0516c disruption modified peptidoglycan thickness, enhanced antibiotic resistance, and activated genes in the regulon of the alternative σ-factor SigF. Phosphorylation of Rv0516c regulated the abundance of EspA, a virulence-associated substrate of the type VII ESX-1 secretion system. These findings identify an osmosensory pathway orchestrated by PknD, Rv0516c, and SigF that enables adaptation to osmotic stress through cell wall remodeling and virulence factor production. Given the widespread occurrence of eukaryotic-like Ser/Thr protein kinases in bacteria, these proteins may play a broad role in bacterial osmosensing.
}, keywords = {Adaptation, Biological, Blotting, Western, Gene Expression Regulation, Bacterial, Green Fluorescent Proteins, Microarray Analysis, Mycobacterium tuberculosis, Osmolar Concentration, Osmotic Pressure, Phosphorylation, Protein Kinases, Signal Transduction}, issn = {1091-6490}, doi = {10.1073/pnas.1321205110}, author = {Hatzios, Stavroula K and Baer, Christina E and Rustad, Tige R and Siegrist, M Sloan and Pang, Jennifer M and Ortega, Corrie and Alber, Tom and Grundner, Christoph and Sherman, David R and Bertozzi, Carolyn R} } @article {3151, title = {Outer cell surface components essential for Fe(III) oxide reduction by Geobacter metallireducens.}, journal = {Appl Environ Microbiol}, volume = {79}, year = {2013}, month = {2013 Feb}, pages = {901-7}, abstract = {Geobacter species are important Fe(III) reducers in a diversity of soils and sediments. Mechanisms for Fe(III) oxide reduction have been studied in detail in Geobacter sulfurreducens, but a number of the most thoroughly studied outer surface components of G. sulfurreducens, particularly c-type cytochromes, are not well conserved among Geobacter species. In order to identify cellular components potentially important for Fe(III) oxide reduction in Geobacter metallireducens, gene transcript abundance was compared in cells grown on Fe(III) oxide or soluble Fe(III) citrate with whole-genome microarrays. Outer-surface cytochromes were also identified. Deletion of genes for c-type cytochromes that had higher transcript abundance during growth on Fe(III) oxides and/or were detected in the outer-surface protein fraction identified six c-type cytochrome genes, that when deleted removed the capacity for Fe(III) oxide reduction. Several of the c-type cytochromes which were essential for Fe(III) oxide reduction in G. metallireducens have homologs in G. sulfurreducens that are not important for Fe(III) oxide reduction. Other genes essential for Fe(III) oxide reduction included a gene predicted to encode an NHL (Ncl-1-HT2A-Lin-41) repeat-containing protein and a gene potentially involved in pili glycosylation. Genes associated with flagellum-based motility, chemotaxis, and pili had higher transcript abundance during growth on Fe(III) oxide, consistent with the previously proposed importance of these components in Fe(III) oxide reduction. These results demonstrate that there are similarities in extracellular electron transfer between G. metallireducens and G. sulfurreducens but the outer-surface c-type cytochromes involved in Fe(III) oxide reduction are different.
}, keywords = {Bacterial Proteins, Culture Media, Cytochromes c, Electron Transport, Ferric Compounds, Gene Deletion, Gene Expression Profiling, Geobacter, Microarray Analysis, Oxidation-Reduction}, issn = {1098-5336}, doi = {10.1128/AEM.02954-12}, author = {Smith, Jessica A and Lovley, Derek R and Tremblay, Pier-Luc} } @article {3134, title = {Syntrophic growth with direct interspecies electron transfer as the primary mechanism for energy exchange.}, journal = {Environ Microbiol Rep}, volume = {5}, year = {2013}, month = {2013 Dec}, pages = {904-10}, abstract = {Direct interspecies electron transfer (DIET) through biological electrical connections is an alternative to interspecies H2 transfer as a mechanism for electron exchange in syntrophic cultures. However, it has not previously been determined whether electrons received via DIET yield energy to support cell growth. In order to investigate this, co-cultures of Geobacter metallireducens, which can transfer electrons to wild-type G. sulfurreducens via DIET, were established with a citrate synthase-deficient G. sulfurreducens strain that can receive electrons for respiration through DIET only. In a medium with ethanol as the electron donor and fumarate as the electron acceptor, co-cultures with the citrate synthase-deficient G. sulfurreducens strain metabolized ethanol as fast as co-cultures with wild-type, but the acetate that G. metallireducens generated from ethanol oxidation accumulated. The lack of acetate metabolism resulted in less fumarate reduction and lower cell abundance of G. sulfurreducens. RNAseq analysis of transcript abundance was consistent with a lack of acetate metabolism in G. sulfurreducens and revealed gene expression levels for the uptake hydrogenase, formate dehydrogenase, the pilus-associated c-type cytochrome OmcS and pili consistent with electron transfer via DIET. These results suggest that electrons transferred via DIET can serve as the sole energy source to support anaerobic respiration.
}, keywords = {Acetates, Anaerobiosis, Citrate (si)-Synthase, Cytochrome c Group, Electron Transport, Electrons, Energy Metabolism, Ethanol, Fimbriae, Bacterial, Formate Dehydrogenases, Fumarates, Geobacter, Oxidation-Reduction}, issn = {1758-2229}, doi = {10.1111/1758-2229.12093}, author = {Shrestha, Pravin Malla and Rotaru, Amelia-Elena and Aklujkar, Muktak and Liu, Fanghua and Shrestha, Minita and Summers, Zarath M and Malvankar, Nikhil and Flores, Dan Carlo and Lovley, Derek R} } @article {1458, title = {Transcription regulation of a yeast gene from a downstream location.}, journal = {J Mol Biol}, volume = {425}, year = {2013}, month = {2013 Feb 8}, pages = {457-65}, abstract = {Mechanisms for coregulation of transcription of tandem genes in yeast remain largely speculative. This study focused on inositol-mediated regulation of the tandem gene pair SNA3-INO1. While the pattern of regulation of these two genes was similar, results showed that intermediate levels of inositol repressed INO1 and induced SNA3. Results also showed that inositol-mediated regulation of the SNA3 gene was not a function of its promoter but occurred from factors within the SNA3-INO1 intergenic region. The basic helix-loop-helix proteins, Ino2p and Ino4p, mediated this regulation through the upstream activation sequence (UAS)(INO) (E-box) sequences in the intergenic region. These results provide a model for studying coregulation of yeast tandem genes. This is especially significant given that many tandem gene pairs in yeast are coregulated even though context-specific UAS sequences are known only for one gene in the pair.
}, keywords = {Gene Expression Regulation, Fungal, Inositol, Membrane Proteins, Myo-Inositol-1-Phosphate Synthase, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins, Transcription, Genetic}, issn = {1089-8638}, doi = {10.1016/j.jmb.2012.11.018}, author = {Shetty, Ameet and Swaminathan, Aishwarya and Lopes, John M} } @article {3146, title = {Transcriptomic and genetic analysis of direct interspecies electron transfer.}, journal = {Appl Environ Microbiol}, volume = {79}, year = {2013}, month = {2013 Apr}, pages = {2397-404}, abstract = {The possibility that metatranscriptomic analysis could distinguish between direct interspecies electron transfer (DIET) and H2 interspecies transfer (HIT) in anaerobic communities was investigated by comparing gene transcript abundance in cocultures in which Geobacter sulfurreducens was the electron-accepting partner for either Geobacter metallireducens, which performs DIET, or Pelobacter carbinolicus, which relies on HIT. Transcript abundance for G. sulfurreducens uptake hydrogenase genes was 7-fold lower in cocultures with G. metallireducens than in cocultures with P. carbinolicus, consistent with DIET and HIT, respectively, in the two cocultures. Transcript abundance for the pilus-associated cytochrome OmcS, which is essential for DIET but not for HIT, was 240-fold higher in the cocultures with G. metallireducens than in cocultures with P. carbinolicus. The pilin gene pilA was moderately expressed despite a mutation that might be expected to repress pilA expression. Lower transcript abundance for G. sulfurreducens genes associated with acetate metabolism in the cocultures with P. carbinolicus was consistent with the repression of these genes by H2 during HIT. Genes for the biogenesis of pili and flagella and several c-type cytochrome genes were among the most highly expressed in G. metallireducens. Mutant strains that lacked the ability to produce pili, flagella, or the outer surface c-type cytochrome encoded by Gmet_2896 were not able to form cocultures with G. sulfurreducens. These results demonstrate that there are unique gene expression patterns that distinguish DIET from HIT and suggest that metatranscriptomics may be a promising route to investigate interspecies electron transfer pathways in more-complex environments.
}, keywords = {Acetates, Deltaproteobacteria, Electron Transport, Hydrogen, Metabolic Networks and Pathways, Transcriptome}, issn = {1098-5336}, doi = {10.1128/AEM.03837-12}, author = {Shrestha, Pravin Malla and Rotaru, Amelia-Elena and Summers, Zarath M and Shrestha, Minita and Liu, Fanghua and Lovley, Derek R} } @article {1274, title = {Trypanosoma brucei Orc1 is essential for nuclear DNA replication and affects both VSG silencing and VSG switching.}, journal = {Mol Microbiol}, volume = {87}, year = {2013}, month = {2013 Jan}, pages = {196-210}, abstract = {Binding of the Origin Recognition Complex (ORC) to replication origins is essential for initiation of DNA replication, but ORC has non-essential functions outside of DNA replication, including in heterochromatic gene silencing and telomere maintenance. Trypanosoma brucei, a protozoan parasite that causes human African trypanosomiasis, uses antigenic variation as a major virulence mechanism to evade the host\&$\#$39;s immune attack by expressing its major surface antigen, the Variant Surface Glycoprotein (VSG), in a monoallelic manner. An Orc1/Cdc6 homologue has been identified in T.\ brucei, but its role in DNA replication has not been directly confirmed and its potential involvement in VSG repression or switching has not been thoroughly investigated. In this study, we show that TbOrc1 is essential for nuclear DNA replication in mammalian-infectious bloodstream and tsetse procyclic forms (BF and PF). Depletion of TbOrc1 resulted in derepression of telomere-linked silent VSGs in both BF and PF, and increased VSG switching particularly through the in situ transcriptional switching mechanism. TbOrc1 associates with telomere repeats but appears to do so independently of two known T.\ brucei telomere proteins, TbRAP1 and TbTRF. We conclude that TbOrc1 has conserved functions in DNA replication and is also required to control telomere-linked VSG expression and VSG switching.
}, keywords = {Antigenic Variation, DNA Replication, DNA, Protozoan, Gene Silencing, Genes, Protozoan, Membrane Glycoproteins, Origin Recognition Complex, Promoter Regions, Genetic, Trypanosoma brucei brucei, Variant Surface Glycoproteins, Trypanosoma}, issn = {1365-2958}, doi = {10.1111/mmi.12093}, author = {Benmerzouga, Imaan and Concepci{\'o}n-Acevedo, Jeniffer and Kim, Hee-Sook and Vandoros, Anthula V and Cross, George A M and Klingbeil, Michele M and Li, Bibo} } @article {1453, title = {Up against the wall: the effects of climate warming on soil microbial diversity and the potential for feedbacks to the carbon cycle.}, journal = {Diversity}, volume = {5}, year = {2013}, pages = {409-425}, author = {Pold, G and DeAngelis, KM} } @article {3137, title = {U(VI) reduction by diverse outer surface c-type cytochromes of Geobacter sulfurreducens.}, journal = {Appl Environ Microbiol}, volume = {79}, year = {2013}, month = {2013 Oct}, pages = {6369-74}, abstract = {Early studies with Geobacter sulfurreducens suggested that outer-surface c-type cytochromes might play a role in U(VI) reduction, but it has recently been suggested that there is substantial U(VI) reduction at the surface of the electrically conductive pili known as microbial nanowires. This phenomenon was further investigated. A strain of G. sulfurreducens, known as Aro-5, which produces pili with substantially reduced conductivity reduced U(VI) nearly as well as the wild type, as did a strain in which the gene for PilA, the structural pilin protein, was deleted. In order to reduce rates of U(VI) reduction to levels less than 20\% of the wild-type rates, it was necessary to delete the genes for the five most abundant outer surface c-type cytochromes of G. sulfurreducens. X-ray absorption near-edge structure spectroscopy demonstrated that whereas 83\% {\textpm} 10\% of the uranium associated with wild-type cells correspond to U(IV) after 4 h of incubation, with the quintuple mutant, 89\% {\textpm} 10\% of uranium was U(VI). Transmission electron microscopy and X-ray energy dispersion spectroscopy revealed that wild-type cells did not precipitate uranium along pili as previously reported, but U(IV) was precipitated at the outer cell surface. These findings are consistent with those of previous studies, which have suggested that G. sulfurreducens requires outer-surface c-type cytochromes but not pili for the reduction of soluble extracellular electron acceptors.
}, keywords = {Cytochromes, Fimbriae, Bacterial, Gene Deletion, Geobacter, Microscopy, Electron, Transmission, Oxidation-Reduction, Uranium, X-Ray Absorption Spectroscopy}, issn = {1098-5336}, doi = {10.1128/AEM.02551-13}, author = {Orellana, Roberto and Leavitt, Janet J and Comolli, Luis R and Csencsits, Roseann and Janot, Noemie and Flanagan, Kelly A and Gray, Arianna S and Leang, Ching and Izallalen, Mounir and Mester, T{\"u}nde and Lovley, Derek R} } @article {3141, title = {When is a microbial culture "pure"? Persistent cryptic contaminant escapes detection even with deep genome sequencing.}, journal = {mBio}, volume = {4}, year = {2013}, month = {2013 Mar 12}, pages = {e00591-12}, abstract = {UNLABELLED: Geobacter sulfurreducens strain KN400 was recovered in previous studies in which a culture of the DL1 strain of G. sulfurreducens served as the inoculum in investigations of microbial current production at low anode potentials (-400 mV versus Ag/AgCl). Differences in the genome sequences of KN400 and DL1 were too great to have arisen from adaptive evolution during growth on the anode. Previous deep sequencing (80-fold coverage) of the DL1 culture failed to detect sequences specific to KN400, suggesting that KN400 was an external contaminant inadvertently introduced into the anode culturing system. In order to evaluate this further, a portion of the gene for OmcS, a c-type cytochrome that both KN400 and DL1 possess, was amplified from the DL1 culture. HiSeq-2000 Illumina sequencing of the PCR product detected the KN400 sequence, which differs from the DL1 sequence at 14 bp, at a frequency of ca. 1 in 10(5) copies of the DL1 sequence. A similar low frequency of KN400 was detected with quantitative PCR of a KN400-specific gene. KN400 persisted at this frequency after intensive restreaking of isolated colonies from the DL1 culture. However, a culture in which KN400 could no longer be detected was obtained by serial dilution to extinction in liquid medium. The KN400-free culture could not grow on an anode poised at -400 mV. Thus, KN400 cryptically persisted in the culture dominated by DL1 for more than a decade, undetected by even deep whole-genome sequencing, and was only fortuitously uncovered by the unnatural selection pressure of growth on a low-potential electrode.
IMPORTANCE: Repeated streaking of isolated colonies on solidified medium remains a common strategy for obtaining pure cultures, especially of difficult-to-cultivate microorganisms such as strict anaerobes. The results presented here demonstrate that verifying the purity of cultures obtained in this manner may be difficult because extremely rare variants can persist, undetectable with even deep genomic DNA sequencing. The only way to ensure that a culture is pure is to cultivate it from an initial single cell, which may be technically difficult for many environmentally significant microbes.
}, keywords = {Coinfection, Electrodes, Genes, Bacterial, Genotype, Geobacter, High-Throughput Nucleotide Sequencing, Microbial Interactions, Polymerase Chain Reaction}, issn = {2150-7511}, doi = {10.1128/mBio.00591-12}, author = {Shrestha, Pravin Malla and Nevin, Kelly P and Shrestha, Minita and Lovley, Derek R} } @article {3155, title = {Anaerobic benzene oxidation by Geobacter species.}, journal = {Appl Environ Microbiol}, volume = {78}, year = {2012}, month = {2012 Dec}, pages = {8304-10}, abstract = {The abundance of Geobacter species in contaminated aquifers in which benzene is anaerobically degraded has led to the suggestion that some Geobacter species might be capable of anaerobic benzene degradation, but this has never been documented. A strain of Geobacter, designated strain Ben, was isolated from sediments from the Fe(III)-reducing zone of a petroleum-contaminated aquifer in which there was significant capacity for anaerobic benzene oxidation. Strain Ben grew in a medium with benzene as the sole electron donor and Fe(III) oxide as the sole electron acceptor. Furthermore, additional evaluation of Geobacter metallireducens demonstrated that it could also grow in benzene-Fe(III) medium. In both strain Ben and G. metallireducens the stoichiometry of benzene metabolism and Fe(III) reduction was consistent with the oxidation of benzene to carbon dioxide with Fe(III) serving as the sole electron acceptor. With benzene as the electron donor, and Fe(III) oxide (strain Ben) or Fe(III) citrate (G. metallireducens) as the electron acceptor, the cell yields of strain Ben and G. metallireducens were 3.2 {\texttimes} 10(9) and 8.4 {\texttimes} 10(9) cells/mmol of Fe(III) reduced, respectively. Strain Ben also oxidized benzene with anthraquinone-2,6-disulfonate (AQDS) as the sole electron acceptor with cell yields of 5.9 {\texttimes} 10(9) cells/mmol of AQDS reduced. Strain Ben serves as model organism for the study of anaerobic benzene metabolism in petroleum-contaminated aquifers, and G. metallireducens is the first anaerobic benzene-degrading organism that can be genetically manipulated.
}, keywords = {Anaerobiosis, Benzene, Carbon Dioxide, Cluster Analysis, Culture Media, DNA, Bacterial, DNA, Ribosomal, Ferric Compounds, Geobacter, Groundwater, Molecular Sequence Data, Oxidation-Reduction, Phylogeny, RNA, Ribosomal, 16S, Sequence Analysis, DNA}, issn = {1098-5336}, doi = {10.1128/AEM.02469-12}, author = {Zhang, Tian and Bain, Timothy S and Nevin, Kelly P and Barlett, Melissa A and Lovley, Derek R} } @article {368, title = {Anaerobic decomposition of switchgrass by tropical soil-derived feedstock-adapted consortia.}, journal = {MBio}, volume = {3}, year = {2012}, month = {2012}, abstract = {Tropical forest soils decompose litter rapidly with frequent episodes of anoxic conditions, making it likely that bacteria using alternate terminal electron acceptors (TEAs) play a large role in decomposition. This makes these soils useful templates for improving biofuel production. To investigate how TEAs affect decomposition, we cultivated feedstock-adapted consortia (FACs) derived from two tropical forest soils collected from the ends of a rainfall gradient: organic matter-rich tropical cloud forest (CF) soils, which experience sustained low redox, and iron-rich tropical rain forest (RF) soils, which experience rapidly fluctuating redox. Communities were anaerobically passed through three transfers of 10 weeks each with switchgrass as a sole carbon (C) source; FACs were then amended with nitrate, sulfate, or iron oxide. C mineralization and cellulase activities were higher in CF-FACs than in RF-FACs. Pyrosequencing of the small-subunit rRNA revealed members of the Firmicutes, Bacteroidetes, and Alphaproteobacteria as dominant. RF- and CF-FAC communities were not different in microbial diversity or biomass. The RF-FACs, derived from fluctuating redox soils, were the most responsive to the addition of TEAs, while the CF-FACs were overall more efficient and productive, both on a per-gram switchgrass and a per-cell biomass basis. These results suggest that decomposing microbial communities in fluctuating redox environments are adapted to the presence of a diversity of TEAs and ready to take advantage of them. More importantly, these data highlight the role of local environmental conditions in shaping microbial community function that may be separate from phylogenetic structure. IMPORTANCE: After multiple transfers, we established microbial consortia derived from two tropical forest soils with different native redox conditions. Communities derived from the rapidly fluctuating redox environment maintained a capacity to use added terminal electron acceptors (TEAs) after multiple transfers, though they were not present during the enrichment. Communities derived from lower-redox soils were not responsive to TEA addition but were much more efficient at switchgrass decomposition. Though the communities were different, diversity was not, and both were dominated by many of the same species of clostridia. This reflects the inadequacy of rRNA for determining the function of microbial communities, in this case the retained ability to utilize TEAs that were not part of the selective growth conditions. More importantly, this suggests that microbial community function is shaped by life history, where environmental factors produce heritable traits through natural selection over time, creating variation in the community, a phenomenon not well documented for microbes.}, keywords = {Adaptation, Physiological, Alphaproteobacteria, Anaerobiosis, Bacteroidetes, Biota, Biotransformation, Carbon, Electron Transport, Enzyme Activation, Ferric Compounds, Genes, rRNA, Microbial Consortia, Nitrates, Oxidation-Reduction, Panicum, Phylogeny, Rain, Soil Microbiology, Sulfates, Trees, Tropical Climate}, issn = {2150-7511}, doi = {10.1128/mBio.00249-11}, author = {Deangelis, Kristen M and Fortney, Julian L and Borglin, Sharon and Silver, Whendee L and Simmons, Blake A and Hazen, Terry C} } @article {356, title = {Analysis of the genome of the sexually transmitted insect virus Helicoverpa zea nudivirus 2.}, journal = {Viruses}, volume = {4}, year = {2012}, month = {2012 Jan}, pages = {28-61}, abstract = {The sexually transmitted insect virus Helicoverpa zea nudivirus 2 (HzNV-2) was determined to have a circular double-stranded DNA genome of 231,621 bp coding for an estimated 113 open reading frames (ORFs). HzNV-2 is most closely related to the nudiviruses, a sister group of the insect baculoviruses. Several putative ORFs that share homology with the baculovirus core genes were identified in the viral genome. However, HzNV-2 lacks several key genetic features of baculoviruses including the late transcriptional regulation factor, LEF-1 and the palindromic hrs, which serve as origins of replication. The HzNV-2 genome was found to code for three ORFs that had significant sequence homology to cellular genes which are not generally found in viral genomes. These included a presumed juvenile hormone esterase gene, a gene coding for a putative zinc-dependent matrix metalloprotease, and a major facilitator superfamily protein gene; all of which are believed to play a role in the cellular proliferation and the tissue hypertrophy observed in the malformation of reproductive organs observed in HzNV-2 infected corn earworm moths, Helicoverpa zea.}, keywords = {Amino Acid Sequence, Animals, Baculoviridae, Base Sequence, Biological Evolution, Consensus Sequence, DNA Viruses, Female, Genome, Viral, Insect Viruses, Molecular Sequence Data, Moths, Open Reading Frames, Phylogeny, Sequence Alignment, Sequence Homology, Species Specificity, Viral Proteins}, issn = {1999-4915}, doi = {10.3390/v4010028}, author = {Burand, John P and Kim, Woojin and Afonso, Claudio L and Tulman, Edan R and Kutish, Gerald F and Lu, Zhiqiang and Rock, Daniel L} } @article {370, title = {Application of phenotypic microarrays to environmental microbiology.}, journal = {Curr Opin Biotechnol}, volume = {23}, year = {2012}, month = {2012 Feb}, pages = {41-8}, abstract = {Environmental organisms are extremely diverse and only a small fraction has been successfully cultured in the laboratory. Culture in micro wells provides a method for rapid screening of a wide variety of growth conditions and commercially available plates contain a large number of substrates, nutrient sources, and inhibitors, which can provide an assessment of the phenotype of an organism. This review describes applications of phenotype arrays to anaerobic and thermophilic microorganisms, use of the plates in stress response studies, in development of culture media for newly discovered strains, and for assessment of phenotype of environmental communities. Also discussed are considerations and challenges in data interpretation and visualization, including data normalization, statistics, and curve fitting.}, issn = {1879-0429}, doi = {10.1016/j.copbio.2011.12.006}, author = {Borglin, Sharon and Joyner, Dominique and Deangelis, Kristen M and Khudyakov, Jane and D{\textquoteright}haeseleer, Patrik and Joachimiak, Marcin P and Hazen, Terry} } @article {726, title = {Biodegradation of ethylene dibromide (1,2-dibromoethane [EDB]) in microcosms simulating in situ and biostimulated conditions.}, journal = {J Hazard Mater}, volume = {209-210}, year = {2012}, month = {2012 Mar 30}, pages = {92-8}, abstract = {Although 1,2-dibromoethane (EDB) is a common groundwater contaminant, there is the lack of knowledge surrounding EDB biodegradation, especially under aerobic conditions. We have performed an extensive microcosm study to investigate the biodegradation of EDB under simulated in situ and biostimulated conditions. The materials for soil microcosms were collected from an EDB-contaminated aquifer at the Massachusetts Military Reservation in Cape Cod, MA. This EDB plume has persisted for nearly 40 years in both aerobic and anaerobic EDB zones of the aquifer. Microcosms were constructed under environmentally relevant conditions (field EDB and DO concentrations; incubated at 12{\textdegree}C). The results showed that natural attenuation occurred under anaerobic conditions but not under aerobic conditions, explaining why aerobic EDB contamination is so persistent. EDB degradation rates were greater under biostimulated conditions for both the aerobic and anaerobic microcosms. Particularly for aerobic biostimulation, methane-amended microcosms degraded EDB, on average, at a first order rate eight times faster than unamended microcosms. The best performing replicate achieved an EDB degradation rate of 7.0 yr(-1) (half-life (t(1/2))=0.10 yr). Residual methane concentrations and the emergence of methanotrophic bacteria, measured by culture independent bacterial analysis, provided strong indications that EDB degradation in aerobic methane-amended microcosms occurred via cometabolic degradation. These results indicate the potential for enhanced natural attenuation of EDB and that methane could be considered co-substrate for EDB bioremediation for the EDB-contaminated groundwater in aerobic zone.}, keywords = {Aerobiosis, Anaerobiosis, Biodegradation, Environmental, Ethylene Dibromide, Water Microbiology, Water Pollutants, Chemical}, issn = {1873-3336}, doi = {10.1016/j.jhazmat.2011.12.067}, author = {McKeever, Robert and Sheppard, Diane and N{\"u}sslein, Klaus and Baek, Kyung-Hwa and Rieber, Khalil and Ergas, Sarina J and Forbes, Rose and Hilyard, Mark and Park, Chul} } @article {1969, title = {Comparative analysis of virus{\textendash}host interactomes with a mammalian high-throughput protein complementation assay based on Gaussia princeps luciferase}, journal = {Methods}, volume = {58}, year = {2012}, month = {dec}, pages = {349{\textendash}359}, doi = {10.1016/j.ymeth.2012.07.029}, url = {https://doi.org/10.1016/j.ymeth.2012.07.029}, author = {Gr{\'e}gory Neveu and Patricia Cassonnet and Pierre-Olivier Vidalain and Caroline Rolloy and Jos{\'e} Mendoza and Louis Jones and Fr{\'e}d{\'e}ric Tangy and Mandy Muller and Caroline Demeret and Lionel Tafforeau and Vincent Lotteau and Chantal Rabourdin-Combe and Gilles Trav{\'e} and Am{\'e}lie Dricot and David E. Hill and Marc Vidal and Michel Favre and Yves Jacob} } @article {3156, title = {Comparative genomic analysis of Geobacter sulfurreducens KN400, a strain with enhanced capacity for extracellular electron transfer and electricity production.}, journal = {BMC Genomics}, volume = {13}, year = {2012}, month = {2012 Sep 12}, pages = {471}, abstract = {BACKGROUND: A new strain of Geobacter sulfurreducens, strain KN400, produces more electrical current in microbial fuel cells and reduces insoluble Fe(III) oxides much faster than the wildtype strain, PCA. The genome of KN400 was compared to wildtype with the goal of discovering how the network for extracellular electron transfer has changed and how these two strains evolved.
RESULTS: Both genomes were re-annotated, resulting in 14 fewer genes (net) in the PCA genome; 28 fewer (net) in the KN400 genome; and ca. 400 gene start and stop sites moved. 96\% of genes in KN400 had clear orthologs with conserved synteny in PCA. Most of the remaining genes were in regions of genomic mobility and were strain-specific or conserved in other Geobacteraceae, indicating that the changes occurred post-divergence. There were 27,270 single nucleotide polymorphisms (SNP) between the genomes. There was significant enrichment for SNP locations in non-coding or synonymous amino acid sites, indicating significant selective pressure since the divergence. 25\% of orthologs had sequence differences, and this set was enriched in phosphorylation and ATP-dependent enzymes. Substantial sequence differences (at least 12 non-synonymous SNP/kb) were found in 3.6\% of the orthologs, and this set was enriched in cytochromes and integral membrane proteins. Genes known to be involved in electron transport, those used in the metabolic cell model, and those that exhibit changes in expression during growth in microbial fuel cells were examined in detail.
CONCLUSIONS: The improvement in external electron transfer in the KN400 strain does not appear to be due to novel gene acquisition, but rather to changes in the common metabolic network. The increase in electron transfer rate and yield in KN400 may be due to changes in carbon flux towards oxidation pathways and to changes in ATP metabolism, both of which indicate that the overall energy state of the cell may be different. The electrically conductive pili appear to be unchanged, but cytochrome folding, localization, and redox potentials may all be affected, which would alter the electrical connection between the cell and the substrate.
}, keywords = {Bioelectric Energy Sources, Comparative Genomic Hybridization, Electron Transport, Gene Expression Regulation, Bacterial, Genome, Bacterial, Geobacter, Metabolic Networks and Pathways, Molecular Sequence Annotation, Polymorphism, Single Nucleotide}, issn = {1471-2164}, doi = {10.1186/1471-2164-13-471}, author = {Butler, Jessica E and Young, Nelson D and Aklujkar, Muktak and Lovley, Derek R} } @article {1207, title = {Complete genome sequence of the hyperthermophilic archaeon Pyrococcus sp. strain ST04, isolated from a deep-sea hydrothermal sulfide chimney on the Juan de Fuca Ridge.}, journal = {J Bacteriol}, volume = {194}, year = {2012}, month = {2012 Aug}, pages = {4434-5}, abstract = {Pyrococcus sp. strain ST04 is a hyperthermophilic, anaerobic, and heterotrophic archaeon isolated from a deep-sea hydrothermal sulfide chimney on the Endeavour Segment of the Juan de Fuca Ridge in the northeastern Pacific Ocean. To further understand the distinct characteristics of this archaeon at the genome level (polysaccharide utilization at high temperature and ATP generation by a Na(+) gradient), the genome of strain ST04 was completely sequenced and analyzed. Here, we present the complete genome sequence analysis results of Pyrococcus sp. ST04 and report the major findings from the genome annotation, with a focus on its saccharolytic and metabolite production potential.
}, keywords = {Adenosine Triphosphate, Anaerobiosis, DNA, Archaeal, Genome, Archaeal, Heterotrophic Processes, Hydrothermal Vents, Molecular Sequence Data, Pacific Ocean, Polysaccharides, Pyrococcus, Seawater, Sequence Analysis, DNA, Sodium Chloride, Sulfides}, issn = {1098-5530}, doi = {10.1128/JB.00824-12}, author = {Jung, Jong-Hyun and Lee, Ju-Hoon and Holden, James F and Seo, Dong-Ho and Shin, Hakdong and Kim, Hae-Yeong and Kim, Wooki and Ryu, Sangryeol and Park, Cheon-Seok} } @article {1205, title = {Complete genome sequence of the hyperthermophilic archaeon Thermococcus sp. strain CL1, isolated from a Paralvinella sp. polychaete worm collected from a hydrothermal vent.}, journal = {J Bacteriol}, volume = {194}, year = {2012}, month = {2012 Sep}, pages = {4769-70}, abstract = {Thermococcus sp. strain CL1 is a hyperthermophilic, anaerobic, and heterotrophic archaeon isolated from a Paralvinella sp. polychaete worm living on an active deep-sea hydrothermal sulfide chimney on the Cleft Segment of the Juan de Fuca Ridge. To further understand the distinct characteristics of this archaeon at the genome level, its genome was completely sequenced and analyzed. Here, we announce the complete genome sequence (1,950,313 bp) of Thermococcus sp. strain CL1, with a focus on H(2)- and energy-producing capabilities and its amino acid biosynthesis and acquisition in an extreme habitat.
}, keywords = {Animals, Base Sequence, Chromosome Mapping, DNA, Archaeal, DNA, Ribosomal, Genome, Bacterial, Hydrothermal Vents, Molecular Sequence Data, Phylogeny, Polychaeta, Sequence Analysis, DNA, Thermococcus}, issn = {1098-5530}, doi = {10.1128/JB.01016-12}, author = {Jung, Jong-Hyun and Holden, James F and Seo, Dong-Ho and Park, Kwan-Hwa and Shin, Hakdong and Ryu, Sangryeol and Lee, Ju-Hoon and Park, Cheon-Seok} } @article {727, title = {Desiccation induces viable but Non-Culturable cells in Sinorhizobium meliloti 1021.}, journal = {AMB Express}, volume = {2}, year = {2012}, month = {2012}, pages = {6}, abstract = {ABSTRACT: Sinorhizobium meliloti is a microorganism commercially used in the production of e.g. Medicago sativa seed inocula. Many inocula are powder-based and production includes a drying step. Although S. meliloti survives drying well, the quality of the inocula is reduced during this process. In this study we determined survival during desiccation of the commercial strains 102F84 and 102F85 as well as the model strain USDA1021.The survival of S. meliloti 1021 was estimated during nine weeks at 22\% relative humidity. We found that after an initial rapid decline of colony forming units, the decline slowed to a steady 10-fold reduction in colony forming units every 22 days. In spite of the reduction in colony forming units, the fraction of the population identified as viable (42-54\%) based on the Baclight live/dead stain did not change significantly over time. This change in the ability of viable cells to form colonies shows (i) an underestimation of the survival of rhizobial cells using plating methods, and that (ii) in a part of the population desiccation induces a Viable But Non Culturable (VBNC)-like state, which has not been reported before. Resuscitation attempts did not lead to a higher recovery of colony forming units indicating the VBNC state is stable under the conditions tested. This observation has important consequences for the use of rhizobia. Finding methods to resuscitate this fraction may increase the quality of powder-based seed inocula.}, issn = {2191-0855}, doi = {10.1186/2191-0855-2-6}, author = {Vriezen, Jan Ac and de Bruijn, Frans J and N{\"u}sslein, Klaus R} } @article {410, title = {The design of long-term effective uranium bioremediation strategy using a community metabolic model.}, journal = {Biotechnol Bioeng}, volume = {109}, year = {2012}, month = {2012 Oct}, pages = {2475-83}, abstract = {Acetate amendment at uranium contaminated sites in Rifle, CO. leads to an initial bloom of Geobacter accompanied by the removal of U(VI) from the groundwater, followed by an increase of sulfate-reducing bacteria (SRBs) which are poor reducers of U(VI). One of the challenges associated with bioremediation is the decay in Geobacter abundance, which has been attributed to the depletion of bio-accessible Fe(III), motivating the investigation of simultaneous amendments of acetate and Fe(III) as an alternative bioremediation strategy. In order to understand the community metabolism of Geobacter and SRBs during artificial substrate amendment, we have created a genome-scale dynamic community model of Geobacter and SRBs using the previously described Dynamic Multi-species Metabolic Modeling framework. Optimization techniques are used to determine the optimal acetate and Fe(III) addition profile. Field-scale simulation of acetate addition accurately predicted the in situ data. The simulations suggest that batch amendment of Fe(III) along with continuous acetate addition is insufficient to promote long-term bioremediation, while continuous amendment of Fe(III) along with continuous acetate addition is sufficient to promote long-term bioremediation. By computationally minimizing the acetate and Fe(III) addition rates as well as the difference between the predicted and target uranium concentration, we showed that it is possible to maintain the uranium concentration below the environmental safety standard while minimizing the cost of chemical additions. These simulations show that simultaneous addition of acetate and Fe(III) has the potential to be an effective uranium bioremediation strategy. They also show that computational modeling of microbial community is an important tool to design effective strategies for practical applications in environmental biotechnology. Biotechnol. Bioeng. 2012; 109: 2475-2483. {\textcopyright} 2012 Wiley Periodicals, Inc.}, issn = {1097-0290}, doi = {10.1002/bit.24528}, author = {Zhuang, K and Ma, E and Lovley, Derek R and Mahadevan, Radhakrishnan} } @article {720, title = {Disassembly of simian virus 40 during passage through the endoplasmic reticulum and in the cytoplasm.}, journal = {J Virol}, volume = {86}, year = {2012}, month = {2012 Feb}, pages = {1555-62}, abstract = {The nonenveloped polyomavirus simian virus 40 (SV40) is taken up into cells by a caveola-mediated endocytic process that delivers the virus to the endoplasmic reticulum (ER). Within the ER lumen, the capsid undergoes partial disassembly, which exposes its internal capsid proteins VP2 and VP3 to immunostaining with antibodies. We demonstrate here that the SV40 genome does not become accessible to detection while the virus is in the ER. Instead, the genome becomes accessible two distinct detection procedures, one using anti-bromodeoxyuridine antibodies and the other using a 5-ethynyl-2-deoxyuridine-based chemical reaction, only after the emergence of partially disassembled SV40 particles in the cytoplasm. These cytoplasmic particles retain some of the SV40 capsid proteins, VP1, VP2, and VP3, in addition to the viral genome. Thus, SV40 particles undergo discrete disassembly steps during entry that are separated temporally and topologically. First, a partial disassembly of the particles occurs in the ER, which exposes internal capsid proteins VP2 and VP3. Then, in the cytoplasm, disassembly progresses further to also make the genomic DNA accessible to immune detection.}, keywords = {Animals, Cell Line, Cercopithecus aethiops, Cytoplasm, Endoplasmic Reticulum, Genome, Viral, Microscopy, Fluorescence, Simian virus 40, Virus Assembly}, issn = {1098-5514}, doi = {10.1128/JVI.05753-11}, author = {Kuksin, Dmitry and Norkin, Leonard C} } @article {721, title = {Disassociation of the SV40 Genome from Capsid Proteins Prior to Nuclear Entry.}, journal = {Virol J}, volume = {9}, year = {2012}, month = {2012 Aug 10}, pages = {158}, abstract = {ABSTRACT: BACKGROUND: Previously, we demonstrated that input SV40 particles undergo a partial disassembly in the endoplasmic reticulum, which exposes internal capsid proteins VP2 and VP3 to immunostaining. Then, in the cytoplasm, disassembly progresses further to also make the genomic DNA accessible to immune detection, as well as to detection by an ethynyl-2-deoxyuridine (EdU)-based chemical reaction. The cytoplasmic partially disassembled SV40 particles retain some of the SV40 capsid proteins, VP1, VP2, and VP3, in addition to the viral genome. FINDINGS: In the current study, we asked where in the cell the SV40 genome might disassociate from capsid components. We observed partially disassembled input SV40 particles around the nucleus and, beginning at 12 hours post-infection, 5-Bromo-2-deoxyuridine (BrdU)-labeled parental SV40 DNA in the nucleus, as detected using anti-BrdU antibodies. However, among the more than 1500 cells examined, we never detected input VP2/VP3 in the nucleus. Upon translocation of the BrdU-labeled SV40 genomes into nuclei, they were transcribed and, thus, are representative of productive infection CONCLUSIONS: Our findings imply that the SV40 genome disassociates from the capsid proteins before or at the point of entry into the nucleus, and then enters the nucleus devoid of VP2/3..}, issn = {1743-422X}, doi = {10.1186/1743-422X-9-158}, author = {Kuksin, Dmitry and Norkin, Leonard C} } @article {379, title = {Dynamic localization of Trypanosoma brucei mitochondrial DNA polymerase ID.}, journal = {Eukaryot Cell}, volume = {11}, year = {2012}, month = {2012 Jul}, pages = {844-55}, abstract = {Trypanosomes contain a unique form of mitochondrial DNA called kinetoplast DNA (kDNA) that is a catenated network composed of minicircles and maxicircles. Several proteins are essential for network replication, and most of these localize to the antipodal sites or the kinetoflagellar zone. Essential components for kDNA synthesis include three mitochondrial DNA polymerases TbPOLIB, TbPOLIC, and TbPOLID). In contrast to other kDNA replication proteins, TbPOLID was previously reported to localize throughout the mitochondrial matrix. This spatial distribution suggests that TbPOLID requires redistribution to engage in kDNA replication. Here, we characterize the subcellular distribution of TbPOLID with respect to the Trypanosoma brucei cell cycle using immunofluorescence microscopy. Our analyses demonstrate that in addition to the previously reported matrix localization, TbPOLID was detected as discrete foci near the kDNA. TbPOLID foci colocalized with replicating minicircles at antipodal sites in a specific subset of the cells during stages II and III of kDNA replication. Additionally, the TbPOLID foci were stable following the inhibition of protein synthesis, detergent extraction, and DNase treatment. Taken together, these data demonstrate that TbPOLID has a dynamic localization that allows it to be spatially and temporally available to perform its role in kDNA replication.}, issn = {1535-9786}, doi = {10.1128/EC.05291-11}, author = {Concepci{\'o}n-Acevedo, Jeniffer and Luo, Juemin and Klingbeil, Michele M} } @article {406, title = {Electrical conductivity in a mixed-species biofilm.}, journal = {Appl Environ Microbiol}, volume = {78}, year = {2012}, month = {2012 Aug}, pages = {5967-71}, abstract = {Geobacter sulfurreducens can form electrically conductive biofilms, but the potential for conductivity through mixed-species biofilms has not been examined. A current-producing biofilm grown from a wastewater sludge inoculum was highly conductive with low charge transfer resistance even though microorganisms other than Geobacteraceae accounted for nearly half the microbial community.
}, keywords = {Biofilms, Electric Conductivity, Microbial Consortia, Sewage}, issn = {1098-5336}, doi = {10.1128/AEM.01803-12}, author = {Malvankar, Nikhil S and Lau, Joanne and Nevin, Kelly P and Franks, Ashley E and Tuominen, Mark T and Lovley, Derek R} } @article {405, title = {Electromicrobiology.}, journal = {Annu Rev Microbiol}, year = {2012}, month = {2012 Jun 28}, abstract = {Electromicrobiology deals with the interactions between microorganisms and electronic devices and with the novel electrical properties of microorganisms. A diversity of microorganisms can donate electrons to, or accept electrons from, electrodes without the addition of artificial electron shuttles. However, the mechanisms for microbe-electrode electron exchange have been seriously studied in only a few microorganisms. Shewanella oneidensis interacts with electrodes primarily via flavins that function as soluble electron shuttles. Geobacter sulfurreducens makes direct electrical contacts with electrodes via outer-surface, c-type cytochromes. G. sulfurreducens is also capable of long-range electron transport along pili, known as microbial nanowires, that have metallic-like conductivity similar to that previously described in synthetic conducting polymers. Pili networks confer conductivity to G. sulfurreducens biofilms, which function as a conducting polymer, with supercapacitor and transistor functionalities. Conductive microorganisms and/or their nanowires have a number of potential practical applications, but additional basic research will be necessary for rational optimization. Expected final online publication date for the Annual Review of Microbiology Volume 66 is September 08, 2012. Please see http://www.annualreviews.org/catalog/pubdates.aspx for revised estimates.}, issn = {1545-3251}, doi = {10.1146/annurev-micro-092611-150104}, author = {Lovley, Derek R} } @article {802, title = {Factors Limiting SOS Expression in Log-Phase Cells of Escherichia coli.}, journal = {J Bacteriol}, volume = {194}, year = {2012}, month = {2012 Oct}, pages = {5325-33}, abstract = {In Escherichia coli, RecA-single-stranded DNA (RecA-ssDNA) filaments catalyze DNA repair, recombination, and induction of the SOS response. It has been shown that, while many (15 to 25\%) log-phase cells have RecA filaments, few (about 1\%) are induced for SOS. It is hypothesized that RecA{\textquoteright}s ability to induce SOS expression in log-phase cells is repressed because of the potentially detrimental effects of SOS mutagenesis. To test this, mutations were sought to produce a population where the number of cells with SOS expression more closely equaled the number of RecA filaments. Here, it is shown that deleting radA (important for resolution of recombination structures) and increasing recA transcription 2- to 3-fold with a recAo1403 operator mutation act independently to minimally satisfy this condition. This allows 24\% of mutant cells to have elevated levels of SOS expression, a percentage similar to that of cells with RecA-green fluorescent protein (RecA-GFP) foci. In an xthA (exonuclease III gene) mutant where there are 3-fold more RecA loading events, recX (a destabilizer of RecA filaments) must be additionally deleted to achieve a population of cells where the percentage having elevated SOS expression (91\%) nearly equals the percentage with at least one RecA-GFP focus (83\%). It is proposed that, in the xthA mutant, there are three independent mechanisms that repress SOS expression in log-phase cells. These are the rapid processing of RecA filaments by RadA, maintaining the concentration of RecA below a critical level, and the destabilizing of RecA filaments by RecX. Only the first two mechanisms operate independently in a wild-type cell.}, issn = {1098-5530}, doi = {10.1128/JB.00674-12}, author = {Massoni, Shawn C and Leeson, Michael C and Long, Jarukit Edward and Gemme, Kristin and Mui, Alice and Sandler, Steven J} } @article {756, title = {Genetic diversity of ixodid ticks parasitizing eastern mouse and dwarf lemurs in Madagascar, with descriptions of the larva, nymph and male of Ixodes lemuris (Acari: Ixodidae).}, journal = {J Parasitol}, year = {2012}, month = {2012 Aug 27}, abstract = {Abstract The ixodid ticks parasitizing small-bodied nocturnal mouse and dwarf lemurs (Primates, Cheirogaleidae) in Madagascar are poorly documented. At Tsinjoarivo, a high-altitude eastern rainforest, mouse and dwarf lemurs were parasitized by ticks of the genus Ixodes. At Ranomafana, a montane southeastern rainforest, dwarf lemurs hosted ticks of the genus Ixodes, whereas mouse lemurs were parasitized by ticks of the genus Haemaphysalis. Our Ixodes specimens represent all active stages and females are morphologically consistent with previous descriptions of Ixodes lemuris females, the only described stage in the literature. Morphological comparisons and genetic analysis using fragments of COI gene confirm all Ixodes ticks from Tsinjoarivo and Ranomafana forests belong to the same species, i.e., Ixodes lemuris. Thus, we are able to provide descriptions of the previously unknown larva, nymph and male. Mouse lemurs at both locations were parasitized only by immature stages of Ixodes lemuris (at Tsinjoarivo) or Haemaphysalis lemuris (at Ranomafana) whereas dwarf lemurs were parasitized by all stages of Ixodes lemuris. We suggest that ecological and biogeographical conditions may affect the pattern of tick infestation at Tsinjoarivo and Ranomafana. Additional studies are necessary to understand the tick-host associations of small-bodied nocturnal lemurs.}, issn = {1937-2345}, doi = {10.1645/GE-3183.1}, author = {Blanco, Marina Beatriz and Elfawal, Mostafa A and Durden, Lance A and Beati, Lorenza and Xu, Guang and Godfrey, Laurie R and Rich, Stephen M} } @article {3149, title = {The genome of Pelobacter carbinolicus reveals surprising metabolic capabilities and physiological features.}, journal = {BMC Genomics}, volume = {13}, year = {2012}, month = {2012 Dec 10}, pages = {690}, abstract = {BACKGROUND: The bacterium Pelobacter carbinolicus is able to grow by fermentation, syntrophic hydrogen/formate transfer, or electron transfer to sulfur from short-chain alcohols, hydrogen or formate; it does not oxidize acetate and is not known to ferment any sugars or grow autotrophically. The genome of P. carbinolicus was sequenced in order to understand its metabolic capabilities and physiological features in comparison with its relatives, acetate-oxidizing Geobacter species.
RESULTS: Pathways were predicted for catabolism of known substrates: 2,3-butanediol, acetoin, glycerol, 1,2-ethanediol, ethanolamine, choline and ethanol. Multiple isozymes of 2,3-butanediol dehydrogenase, ATP synthase and [FeFe]-hydrogenase were differentiated and assigned roles according to their structural properties and genomic contexts. The absence of asparagine synthetase and the presence of a mutant tRNA for asparagine encoded among RNA-active enzymes suggest that P. carbinolicus may make asparaginyl-tRNA in a novel way. Catabolic glutamate dehydrogenases were discovered, implying that the tricarboxylic acid (TCA) cycle can function catabolically. A phosphotransferase system for uptake of sugars was discovered, along with enzymes that function in 2,3-butanediol production. Pyruvate:ferredoxin/flavodoxin oxidoreductase was identified as a potential bottleneck in both the supply of oxaloacetate for oxidation of acetate by the TCA cycle and the connection of glycolysis to production of ethanol. The P. carbinolicus genome was found to encode autotransporters and various appendages, including three proteins with similarity to the geopilin of electroconductive nanowires.
CONCLUSIONS: Several surprising metabolic capabilities and physiological features were predicted from the genome of P. carbinolicus, suggesting that it is more versatile than anticipated.
}, keywords = {Base Pairing, Base Sequence, Butylene Glycols, Choline, Deltaproteobacteria, Ethanolamine, Ethylene Glycol, Genome, Bacterial, Glycerol, Metabolic Networks and Pathways, Molecular Sequence Annotation, Molecular Sequence Data, Mutation, Oxidation-Reduction, Oxidoreductases, Propylene Glycols, RNA, Transfer, Asn, Sequence Analysis, DNA}, issn = {1471-2164}, doi = {10.1186/1471-2164-13-690}, author = {Aklujkar, Muktak and Haveman, Shelley A and DiDonato, Raymond and Chertkov, Olga and Han, Cliff S and Land, Miriam L and Brown, Peter and Lovley, Derek R} } @article {419, title = {Genome-scale analysis of anaerobic benzoate and phenol metabolism in the hyperthermophilic archaeon Ferroglobus placidus.}, journal = {ISME J}, volume = {6}, year = {2012}, month = {2012 Jan}, pages = {146-57}, abstract = {Insight into the mechanisms for the anaerobic metabolism of aromatic compounds by the hyperthermophilic archaeon Ferroglobus placidus is expected to improve understanding of the degradation of aromatics in hot (>80{\textdegree} C) environments and to identify enzymes that might have biotechnological applications. Analysis of the F. placidus genome revealed genes predicted to encode enzymes homologous to those previously identified as having a role in benzoate and phenol metabolism in mesophilic bacteria. Surprisingly, F. placidus lacks genes for an ATP-independent class II benzoyl-CoA (coenzyme A) reductase (BCR) found in all strictly anaerobic bacteria, but has instead genes coding for a bzd-type ATP-consuming class I BCR, similar to those found in facultative bacteria. The lower portion of the benzoate degradation pathway appears to be more similar to that found in the phototroph Rhodopseudomonas palustris, than the pathway reported for all heterotrophic anaerobic benzoate degraders. Many of the genes predicted to be involved in benzoate metabolism were found in one of two gene clusters. Genes for phenol carboxylation proceeding through a phenylphosphate intermediate were identified in a single gene cluster. Analysis of transcript abundance with a whole-genome microarray and quantitative reverse transcriptase polymerase chain reaction demonstrated that most of the genes predicted to be involved in benzoate or phenol metabolism had higher transcript abundance during growth on those substrates vs growth on acetate. These results suggest that the general strategies for benzoate and phenol metabolism are highly conserved between microorganisms living in moderate and hot environments, and that anaerobic metabolism of aromatic compounds might be analyzed in a wide range of environments with similar molecular targets.}, keywords = {Acetates, Archaea, Bacteria, Anaerobic, Benzoates, Metabolic Networks and Pathways, Phenol, Rhodopseudomonas}, issn = {1751-7370}, doi = {10.1038/ismej.2011.88}, author = {Holmes, Dawn E and Risso, Carla and Smith, Jessica A and Lovley, Derek R} } @article {367, title = {Global transcriptome response to ionic liquid by a tropical rain forest soil bacterium, Enterobacter lignolyticus.}, journal = {Proc Natl Acad Sci U S A}, volume = {109}, year = {2012}, month = {2012 Aug 7}, pages = {E2173-82}, abstract = {To process plant-based renewable biofuels, pretreatment of plant feedstock with ionic liquids has significant advantages over current methods for deconstruction of lignocellulosic feedstocks. However, ionic liquids are often toxic to the microorganisms used subsequently for biomass saccharification and fermentation. We previously isolated Enterobacter lignolyticus strain SCF1, a lignocellulolytic bacterium from tropical rain forest soil, and report here that it can grow in the presence of 0.5 M 1-ethyl-3-methylimidazolium chloride, a commonly used ionic liquid. We investigated molecular mechanisms of SCF1 ionic liquid tolerance using a combination of phenotypic growth assays, phospholipid fatty acid analysis, and RNA sequencing technologies. Potential modes of resistance to 1-ethyl-3-methylimidazolium chloride include an increase in cyclopropane fatty acids in the cell membrane, scavenging of compatible solutes, up-regulation of osmoprotectant transporters and drug efflux pumps, and down-regulation of membrane porins. These findings represent an important first step in understanding mechanisms of ionic liquid resistance in bacteria and provide a basis for engineering microbial tolerance.}, issn = {1091-6490}, doi = {10.1073/pnas.1112750109}, author = {Khudyakov, Jane I and D{\textquoteright}haeseleer, Patrik and Borglin, Sharon E and Deangelis, Kristen M and Woo, Hannah and Lindquist, Erika A and Hazen, Terry C and Simmons, Blake A and Thelen, Michael P} } @article {1424, title = {How Bacteria Adhere to Brushy PEG Surfaces: Clinging to Flaws and Compressing the Brush.}, journal = {Macromolecules}, volume = {45}, year = {2012}, month = {2012 Oct 23}, pages = {8373-8381}, abstract = {This study examined the compression of solvated polymer brushes on bioengineered surfaces during the initial stages of Staphylococcus Aureus (S. aureus) adhesion from gentle flow. A series of PEG [poly(ethylene glycol)] brushes, 7 to 17 nm in height and completely non-adhesive to proteins and bacteria, were modified by the incorporation of sparse isolated ~10 nm cationic polymer \"patches\" at their bases. These nanoscale regions, which lacked PEG tethers, were electrostatically attractive towards negative bacteria or proteins. S. aureus drawn to the interface by multiple adhesive patches compressed the PEG brush in the remaining contact region. The observed onset of bacterial or fibrinogen capture with increases in patch content was compared with calculations. Balancing the attraction energy (proportional to the number of patches engaging a bacterium during capture) against steric forces (calculated using the Alexander-DeGennes treatment) provided perspective on the brush compression. The results were consistent with a bacteria-surface gap on the order of the Debye length in these studies. In this limit of strong brush compression, structural features (height, persistence length) of the brush were unimportant so that osmotic pressure dominated the steric repulsion. Thus, the dominant factor for bacterial repulsion was the mass of PEG in the brush. This result explains empirical reports in the literature that identify the total PEG content of a brush as a criteria for prevention of bioadhesion, independent of tether length and spacing, within a reasonable range for those parameters. Bacterial capture was also compared to that of protein capture. It was found, surprisingly, that the patchy brushes were more protein-than bacteria-resistant. S. aureus adhesion driven by patches within otherwise protein-resistant PEG brushes was explained by the bacteria\&$\#$39;s greater tendency to compress large areas of brush to interact with many patches. By contrast, proteins are thought to penetrate the brush at a few sites of PEO-free patches. The finding provides a mechanism for the literature reports that in-vitro protein resistance is a poor predictor of in-vitro implant failure related to cell-surface adhesion.
}, issn = {0024-9297}, doi = {10.1021/ma300981r}, author = {Gon, S and Kumar, Kushi-Nidhi and N{\"u}sslein, Klaus and Santore, Maria M} } @article {1968, title = {The {HPV} E2-Host Protein-Protein Interactions: A Complex Hijacking of the Cellular Network}, journal = {The Open Virology Journal}, volume = {6}, year = {2012}, month = {dec}, pages = {173{\textendash}189}, doi = {10.2174/1874357901206010173}, url = {https://doi.org/10.2174/1874357901206010173}, author = {Mandy Muller and C Demeret} } @article {1206, title = {Hydrogen-limited growth of hyperthermophilic methanogens at deep-sea hydrothermal vents.}, journal = {Proc Natl Acad Sci U S A}, volume = {109}, year = {2012}, month = {2012 Aug 21}, pages = {13674-9}, abstract = {Microbial productivity at hydrothermal vents is among the highest found anywhere in the deep ocean, but constraints on microbial growth and metabolism at vents are lacking. We used a combination of cultivation, molecular, and geochemical tools to verify pure culture H(2) threshold measurements for hyperthermophilic methanogenesis in low-temperature hydrothermal fluids from Axial Volcano and Endeavour Segment in the northeastern Pacific Ocean. Two Methanocaldococcus strains from Axial and Methanocaldococcus jannaschii showed similar Monod growth kinetics when grown in a bioreactor at varying H(2) concentrations. Their H(2) half-saturation value was 66 μM, and growth ceased below 17-23 μM H(2), 10-fold lower than previously predicted. By comparison, measured H(2) and CH(4) concentrations in fluids suggest that there was generally sufficient H(2) for Methanocaldococcus growth at Axial but not at Endeavour. Fluids from one vent at Axial (Marker 113) had anomalously high CH(4) concentrations and contained various thermal classes of methanogens based on cultivation and mcrA/mrtA analyses. At Endeavour, methanogens were largely undetectable in fluid samples based on cultivation and molecular screens, although abundances of hyperthermophilic heterotrophs were relatively high. Where present, Methanocaldococcus genes were the predominant mcrA/mrtA sequences recovered and comprised \~{}0.2-6\% of the total archaeal community. Field and coculture data suggest that H(2) limitation may be partly ameliorated by H(2) syntrophy with hyperthermophilic heterotrophs. These data support our estimated H(2) threshold for hyperthermophilic methanogenesis at vents and highlight the need for coupled laboratory and field measurements to constrain microbial distribution and biogeochemical impacts in the deep sea.
}, keywords = {Archaea, Biodiversity, Coculture Techniques, DNA, Ribosomal, Ecosystem, Gases, Geography, Hydrogen, Hydrothermal Vents, Kinetics, Methane, Molecular Sequence Data, Temperature, Time Factors, Water Microbiology}, issn = {1091-6490}, doi = {10.1073/pnas.1206632109}, author = {Ver Eecke, Helene C and Butterfield, David A and Huber, Julie A and Lilley, Marvin D and Olson, Eric J and Roe, Kevin K and Evans, Leigh J and Merkel, Alexandr Y and Cantin, Holly V and Holden, James F} } @article {412, title = {Identification of multicomponent histidine-aspartate phosphorelay system controlling flagellar and motility gene expression in Geobacter species.}, journal = {J Biol Chem}, volume = {287}, year = {2012}, month = {2012 Mar 30}, pages = {10958-66}, abstract = {Geobacter species play an important role in the natural biogeochemical cycles of aquatic sediments and subsurface environments as well as in subsurface bioremediation by oxidizing organic compounds with the reduction of insoluble Fe(III) oxides. Flagellum-based motility is considered to be critical for Geobacter species to locate fresh sources of Fe(III) oxides. Functional and comparative genomic approaches, coupled with genetic and biochemical methods, identified key regulators for flagellar gene expression in Geobacter species. A master transcriptional regulator, designated FgrM, is a member of the enhancer-binding protein family. The fgrM gene in the most studied strain of Geobacter species, Geobacter sulfurreducens strain DL-1, is truncated by a transposase gene, preventing flagellar biosynthesis. Integrating a functional FgrM homolog restored flagellar biosynthesis and motility in G. sulfurreducens DL-1 and enhanced the ability to reduce insoluble Fe(III) oxide. Interrupting the fgrM gene in G. sulfurreducens strain KN400, which is motile, removed the capacity for flagellar production and inhibited Fe(III) oxide reduction. FgrM, which is also a response regulator of the two-component His-Asp phosphorelay system, was phosphorylated by histidine kinase GHK4, which was essential for flagellar production and motility. GHK4, which is a hybrid kinase with a receiver domain at the N terminus, was phosphorylated by another histidine kinase, GHK3. Therefore, the multicomponent His-Asp phosphorelay system appears to control flagellar gene expression in Geobacter species.}, issn = {1083-351X}, doi = {10.1074/jbc.M112.345041}, author = {Ueki, Toshiyuki and Leang, Ching and Inoue, Kengo and Lovley, Derek R} } @article {848, title = {In vitro assessment of halobacterial gas vesicles as a Chlamydia vaccine display and delivery system.}, journal = {Vaccine}, volume = {30}, year = {2012}, month = {2012 Sep 7}, pages = {5942-8}, abstract = {Chlamydia trachomatis is the leading cause of bacterial sexually transmitted disease worldwide and while antibiotic treatment is effective in eliminating the pathogen, up to 70\% of all infections are asymptomatic. Despite sustained efforts over the past 2 decades, an effective chlamydial vaccine remains elusive, due in large part to the lack of an effective delivery system. We explored the use of gas vesicles derived from Halobacterium salinarium as a potential display and delivery vehicle for chlamydial antigens of vaccine interest. Various size gene fragments coding for the major outer membrane protein (MOMP), outer membrane complex B (OmcB) and polymorphic outer membrane protein D (PompD) were integrated into and expressed as part of the gas vesicle protein C (gvpC) on the surface of these stable structures. The presence of the recombinant proteins was confirmed by Western blots probed using anti-gvpC and anti-Chlamydia antibodies as well as sera from Chlamydia-positive patients. Tissue culture evaluation revealed stability and a time-dependent degradation of recombinant gas vesicles (r-Gv) in human and animal cell lines. In vitro assessment using human foreskin fibroblasts (HFF) confirmed Toll-like receptor (TLR) 4 and 5 engagement by wild type and r-Gv, leading to MyD88 activation, TNF-α, IL-6 and IL-12 production. The data suggest that r-GV could be an effective, naturally adjuvanting, time-release antigen delivery system for immunologically relevant Chlamydia vaccine antigens which are readily recognized by human immune sera.}, issn = {1873-2518}, doi = {10.1016/j.vaccine.2012.07.038}, author = {Childs, Tawanna S and Webley, Wilmore C} } @article {404, title = {Interspecies electron transfer via hydrogen and formate rather than direct electrical connections in cocultures of Pelobacter carbinolicus and Geobacter sulfurreducens.}, journal = {Appl Environ Microbiol}, volume = {78}, year = {2012}, month = {2012 Nov}, pages = {7645-51}, abstract = {Direct interspecies electron transfer (DIET) is an alternative to interspecies H(2)/formate transfer as a mechanism for microbial species to cooperatively exchange electrons during syntrophic metabolism. To understand what specific properties contribute to DIET, studies were conducted with Pelobacter carbinolicus, a close relative of Geobacter metallireducens, which is capable of DIET. P. carbinolicus grew in coculture with Geobacter sulfurreducens with ethanol as the electron donor and fumarate as the electron acceptor, conditions under which G. sulfurreducens formed direct electrical connections with G. metallireducens. In contrast to the cell aggregation associated with DIET, P. carbinolicus and G. sulfurreducens did not aggregate. Attempts to initiate cocultures with a genetically modified strain of G. sulfurreducens incapable of both H(2) and formate utilization were unsuccessful, whereas cocultures readily grew with mutant strains capable of formate but not H(2) uptake or vice versa. The hydrogenase mutant of G. sulfurreducens compensated, in cocultures, with significantly increased formate dehydrogenase gene expression. In contrast, the transcript abundance of a hydrogenase gene was comparable in cocultures with that for the formate dehydrogenase mutant of G. sulfurreducens or the wild type, suggesting that H(2) was the primary electron carrier in the wild-type cocultures. Cocultures were also initiated with strains of G. sulfurreducens that could not produce pili or OmcS, two essential components for DIET. The finding that P. carbinolicus exchanged electrons with G. sulfurreducens via interspecies transfer of H(2)/formate rather than DIET demonstrates that not all microorganisms that can grow syntrophically are capable of DIET and that closely related microorganisms may use significantly different strategies for interspecies electron exchange.
}, keywords = {Coculture Techniques, Deltaproteobacteria, Electricity, Electron Transport, Electrons, Formates, Geobacter, Hydrogen, Microbial Interactions}, issn = {1098-5336}, doi = {10.1128/AEM.01946-12}, author = {Rotaru, Amelia-Elena and Shrestha, Pravin M and Liu, Fanghua and Ueki, Toshiyuki and Nevin, Kelly and Summers, Zarath M and Lovley, Derek R} } @article {416, title = {Laboratory evolution of Geobacter sulfurreducens for enhanced growth on lactate via a single-base-pair substitution in a transcriptional regulator.}, journal = {ISME J}, volume = {6}, year = {2012}, month = {2012 May}, pages = {975-83}, abstract = {The addition of organic compounds to groundwater in order to promote bioremediation may represent a new selective pressure on subsurface microorganisms. The ability of Geobacter sulfurreducens, which serves as a model for the Geobacter species that are important in various types of anaerobic groundwater bioremediation, to adapt for rapid metabolism of lactate, a common bioremediation amendment, was evaluated. Serial transfer of five parallel cultures in a medium with lactate as the sole electron donor yielded five strains that could metabolize lactate faster than the wild-type strain. Genome sequencing revealed that all five strains had non-synonymous single-nucleotide polymorphisms in the same gene, GSU0514, a putative transcriptional regulator. Introducing the single-base-pair mutation from one of the five strains into the wild-type strain conferred rapid growth on lactate. This strain and the five adaptively evolved strains had four to eight-fold higher transcript abundance than wild-type cells for genes for the two subunits of succinyl-CoA synthase, an enzyme required for growth on lactate. DNA-binding assays demonstrated that the protein encoded by GSU0514 bound to the putative promoter of the succinyl-CoA synthase operon. The binding sequence was not apparent elsewhere in the genome. These results demonstrate that a single-base-pair mutation in a transcriptional regulator can have a significant impact on the capacity for substrate utilization and suggest that adaptive evolution should be considered as a potential response of microorganisms to environmental change(s) imposed during bioremediation.}, issn = {1751-7370}, doi = {10.1038/ismej.2011.166}, author = {Summers, Zarath M and Ueki, Toshiyuki and Ismail, Wael and Haveman, Shelley A and Lovley, Derek R} } @article {1970, title = {Large Scale Genotype Comparison of Human Papillomavirus E2-Host Interaction Networks Provides New Insights for E2 Molecular Functions}, journal = {{PLoS} Pathogens}, volume = {8}, year = {2012}, month = {jun}, pages = {e1002761}, doi = {10.1371/journal.ppat.1002761}, url = {https://doi.org/10.1371/journal.ppat.1002761}, author = {Mandy Muller and Yves Jacob and Louis Jones and Am{\'e}lie Weiss and Laurent Brino and Thibault Chantier and Vincent Lotteau and Michel Favre and Caroline Demeret}, editor = {Frederick P. Roth} } @article {3152, title = {Long-range electron transport to Fe(III) oxide via pili with metallic-like conductivity.}, journal = {Biochem Soc Trans}, volume = {40}, year = {2012}, month = {2012 Dec 01}, pages = {1186-90}, abstract = {The mechanisms for Fe(III) oxide reduction by Geobacter species are of interest because Geobacter species have been shown to play an important role in Fe(III) oxide reduction in a diversity of environments in which Fe(III) reduction is a geochemically significant process. Geobacter species specifically express pili during growth on Fe(III) oxide compared with growth on soluble chelated Fe(III), and mutants that cannot produce pili are unable to effectively reduce Fe(III) oxide. The pili of Geobacter sulfurreducens are electrically conductive along their length under physiologically relevant conditions and exhibit a metallic-like conductivity similar to that observed previously in synthetic organic metals. Metallic-like conductivity in a biological protein filament is a previously unrecognized mechanism for electron transport that differs significantly from the more well-known biological strategy of electron hopping/tunnelling between closely spaced redox-active proteins. The multihaem c-type cytochrome OmcS is specifically associated with pili and is necessary for Fe(III) oxide reduction. However, multiple lines of evidence, including the metallic-like conductivity of the pili and the fact that OmcS molecules are spaced too far apart for electron hopping/tunnelling, indicate that OmcS is not responsible for long-range electron conduction along the pili. The role of OmcS may be to facilitate electron transfer from the pili to Fe(III) oxide. Long-range electron transport via pili with metallic-like conductivity is a paradigm shift that has important implications not only for Fe(III) oxide reduction, but also for interspecies electron exchange in syntrophic microbial communities as well as microbe-electrode interactions and the emerging field of bioelectronics.
}, keywords = {Bacterial Outer Membrane Proteins, Biofilms, Cytochromes, Electric Conductivity, Electron Transport, Ferric Compounds, Fimbriae, Bacterial, Geobacter, Oxidation-Reduction}, issn = {1470-8752}, doi = {10.1042/BST20120131}, author = {Lovley, Derek R} } @article {413, title = {Microbial functional gene diversity with a shift of subsurface redox conditions during In Situ uranium reduction.}, journal = {Appl Environ Microbiol}, volume = {78}, year = {2012}, month = {2012 Apr}, pages = {2966-72}, abstract = {To better understand the microbial functional diversity changes with subsurface redox conditions during in situ uranium bioremediation, key functional genes were studied with GeoChip, a comprehensive functional gene microarray, in field experiments at a uranium mill tailings remedial action (UMTRA) site (Rifle, CO). The results indicated that functional microbial communities altered with a shift in the dominant metabolic process, as documented by hierarchical cluster and ordination analyses of all detected functional genes. The abundance of dsrAB genes (dissimilatory sulfite reductase genes) and methane generation-related mcr genes (methyl coenzyme M reductase coding genes) increased when redox conditions shifted from Fe-reducing to sulfate-reducing conditions. The cytochrome genes detected were primarily from Geobacter sp. and decreased with lower subsurface redox conditions. Statistical analysis of environmental parameters and functional genes indicated that acetate, U(VI), and redox potential (E(h)) were the most significant geochemical variables linked to microbial functional gene structures, and changes in microbial functional diversity were strongly related to the dominant terminal electron-accepting process following acetate addition. The study indicates that the microbial functional genes clearly reflect the in situ redox conditions and the dominant microbial processes, which in turn influence uranium bioreduction. Microbial functional genes thus could be very useful for tracking microbial community structure and dynamics during bioremediation.}, keywords = {Biodegradation, Environmental, Biota, Environmental Microbiology, Environmental Pollutants, Genetic Variation, Microarray Analysis, Oxidation-Reduction, Uranium}, issn = {1098-5336}, doi = {10.1128/AEM.06528-11}, author = {Liang, Yuting and Van Nostrand, Joy D and N{\textquoteright}guessan, Lucie A and Peacock, Aaron D and Deng, Ye and Long, Philip E and Resch, C Tom and Wu, Liyou and He, Zhili and Li, Guanghe and Hazen, Terry C and Lovley, Derek R and Zhou, Jizhong} } @article {407, title = {Microbial nanowires: a new paradigm for biological electron transfer and bioelectronics.}, journal = {ChemSusChem}, volume = {5}, year = {2012}, month = {2012 Jun}, pages = {1039-46}, abstract = {The discovery that Geobacter sulfurreducens can produce protein filaments with metallic-like conductivity, known as microbial nanowires, that facilitate long-range electron transport is a paradigm shift in biological electron transfer and has important implications for biogeochemistry, microbial ecology, and the emerging field of bioelectronics. Although filaments in a wide diversity of microorganisms have been called microbial nanowires, the type IV pili of G. sulfurreducens and G. metallireducens are the only filaments that have been shown to be required for extracellular electron transport to extracellular electron acceptors or for conduction of electrons through biofilms. Studies of G. sulfurreducens pili preparations and intact biofilms under physiologically relevant conditions have provided multiple lines of evidence for metallic-like conduction along the length of pili and for the possibility of pili networks to confer high conductivity within biofilms. This mechanism of electron conduction contrasts with the previously known mechanism for biological electron transfer via electron tunneling or hopping between closely associated molecules, a strategy unlikely to be well adapted for long-range electron transport outside the cell. In addition to promoting electron exchange with abiotic electron acceptors, microbial nanowires have recently been shown to be involved in direct interspecies electron transfer between syntrophic partners. An improved understanding of the mechanisms for metallic-like conductivity in microbial nanowires, as well as engineering microorganisms with desirable catalytic abilities with nanowires, could lead to new applications in microbial electrosynthesis and bioelectronics.
}, keywords = {Bacterial Physiological Phenomena, Biofilms, Biotechnology, Electric Conductivity, Ferric Compounds, Fimbriae Proteins, Fimbriae, Bacterial, Nanostructures, Oxidation-Reduction}, issn = {1864-564X}, doi = {10.1002/cssc.201100733}, author = {Malvankar, Nikhil S and Lovley, Derek R} } @article {722, title = {Molecular approach to evaluate biostimulation of 1,2-dibromoethane in contaminated groundwater.}, journal = {Bioresour Technol}, volume = {123C}, year = {2012}, month = {2012 May 29}, pages = {207-213}, abstract = {This study investigated the effect of co-substrate amendments on EDB biodegradation under aerobic conditions. Microcosms were established using contaminated soil and groundwater samples and maintained under in situ conditions to determine EDB degradation rates, and the diversity and abundance of EDB degrading indigenous bacteria. After 100days of incubation, between 25\% and 56\% of the initial EDB was degraded in the microcosms, with added jet fuel providing highest degradation rates (2.97{\textpm}0.49yr(-1)). In all microcosms, the quantity of dehalogenase genes did not change significantly, while the number of BTEX monooxygenase and phenol hydroxylase genes increased with jet fuel amendments. These results indicate that EDB was not degraded by prior dehalogenation, but rather by cometabolism with adapted indigenous microorganisms. This is also reflected in the history of the plume, which originated from an aviation gasoline pipeline leak. This study suggests that biostimulation of EDB is possible at aerobic groundwater sites.}, issn = {1873-2976}, doi = {10.1016/j.biortech.2012.05.119}, author = {Baek, Kyunghwa and McKeever, Robert and Rieber, Kahlil and Sheppard, Diane and Park, Chul and Ergas, Sarina J and N{\"u}sslein, Klaus} } @article {803, title = {Mu insertions are repaired by the double-strand break repair pathway of Escherichia coli.}, journal = {PLoS Genet}, volume = {8}, year = {2012}, month = {2012 Apr}, pages = {e1002642}, abstract = {Mu is both a transposable element and a temperate bacteriophage. During lytic growth, it amplifies its genome by replicative transposition. During infection, it integrates into the Escherichia coli chromosome through a mechanism not requiring extensive DNA replication. In the latter pathway, the transposition intermediate is repaired by transposase-mediated resecting of the 5{\textquoteright} flaps attached to the ends of the incoming Mu genome, followed by filling the remaining 5 bp gaps at each end of the Mu insertion. It is widely assumed that the gaps are repaired by a gap-filling host polymerase. Using the E. coli Keio Collection to screen for mutants defective in recovery of stable Mu insertions, we show in this study that the gaps are repaired by the machinery responsible for the repair of double-strand breaks in E. coli-the replication restart proteins PriA-DnaT and homologous recombination proteins RecABC. We discuss alternate models for recombinational repair of the Mu gaps.}, issn = {1553-7404}, doi = {10.1371/journal.pgen.1002642}, author = {Jang, Sooin and Sandler, Steven J and Harshey, Rasika M} } @article {723, title = {New bactericidal surgical suture coating.}, journal = {Langmuir}, volume = {28}, year = {2012}, month = {2012 Aug 21}, pages = {12134-9}, abstract = {This paper demonstrates the effectiveness of a new antimicrobial suture coating. An amphiphilic polymer, poly[(aminoethyl methacrylate)-co-(butyl methacrylate)] (PAMBM), inspired by antimicrobial peptides, was bactericidal against S. aureus in time-kill experiments. PAMBM was then evaluated in a variety of polymer blends using the Japanese Industrial Standard (JIS) method and showed excellent antimicrobial activity at a low concentration (0.5 wt \%). Using a similar antimicrobial coating formula to commercial Vicryl Plus sutures, disk samples of the coating material containing PAMBM effectively killed bacteria (98\% reduction at 0.75 wt \%). Triclosan, the active ingredient in Vicryl Plus coatings, did not kill the bacteria. Further Kirby-Bauer assays of these disk samples showed an increasing zone of inhibition with increasing concentration of PAMBM. Finally, the PAMBM-containing coating was applied to sutures, and the morphology of the coating surface was characterized by SEM, along with Vicryl and uncoated sutures. The PAMBM-containing sutures killed bacteria more effectively (3 log(10) reduction at 2.4 wt \%) than Vicryl Plus sutures (0.5 log(10) reduction).}, issn = {1520-5827}, doi = {10.1021/la302732w}, author = {Li, Yan and Kumar, Kushi N and Dabkowski, Jeffrey M and Corrigan, Meagan and Scott, Richard W and N{\"u}sslein, Klaus and Tew, Gregory N} } @article {728, title = {Performance of a pilot-scale packed bed reactor for perchlorate reduction using a sulfur oxidizing bacterial consortium.}, journal = {Biotechnol Bioeng}, volume = {109}, year = {2012}, month = {2012 Mar}, pages = {637-46}, abstract = {A novel sulfur-utilizing perchlorate reducing bacterial consortium successfully treated perchlorate (ClO$_{4}$$^{-}$) in prior batch and bench-scale packed bed reactor (PBR) studies. This study examined the scale up of this process for treatment of water from a ClO $_{4}$$^{-}$ and RDX contaminated aquifer in Cape Cod Massachusetts. A pilot-scale upflow PBR (\~{}250-L) was constructed with elemental sulfur and crushed oyster shell packing media. The reactor was inoculated with sulfur oxidizing ClO$_{4}$$^{-}$ reducing cultures enriched from a wastewater seed. Sodium sulfite provided a good method of dissolved oxygen removal in batch cultures, but was found to promote the growth of bacteria that carry out sulfur disproportionation and sulfate reduction, which inhibited ClO$_{4}$$^{-}$ reduction in the pilot system. After terminating sulfite addition, the PBR successfully removed 96\% of the influent ClO$_{4}$$^{-}$ in the groundwater at an empty bed contact time (EBCT) of 12 h (effluent ClO$_{4}$$^{-}$ of 4.2 {\textmu}g L(-1)). Simultaneous ClO$_{4}$$^{-}$ and NO$_{3}$$^{-}$ reduction was observed in the lower half of the reactor before reactions shifted to sulfur disproportionation and sulfate reduction. Analyses of water quality profiles were supported by molecular analysis, which showed distinct groupings of ClO$_{4}$$^{-}$ and NO$_{3}$$^{-}$ degrading organisms at the inlet of the PBR, while sulfur disproportionation was the primary biological process occurring in the top potion of the reactor.}, keywords = {Bioreactors, Cluster Analysis, DNA, Bacterial, DNA, Ribosomal, Massachusetts, Microbial Consortia, Molecular Sequence Data, Oxidation-Reduction, Perchloric Acid, Phylogeny, RNA, Ribosomal, 16S, Sequence Analysis, DNA, Sulfites, Sulfur, Water Microbiology, Water Pollutants, Chemical, Water Purification}, issn = {1097-0290}, doi = {10.1002/bit.24354}, author = {Boles, Amber R and Conneely, Teresa and McKeever, Robert and Nixon, Paul and N{\"u}sslein, Klaus R and Ergas, Sarina J} } @article {409, title = {Phylogenetic classification of diverse LysR-type transcriptional regulators of a model prokaryote Geobacter sulfurreducens.}, journal = {J Mol Evol}, volume = {74}, year = {2012}, month = {2012 Apr}, pages = {187-205}, abstract = {The protein family of LysR-type transcriptional regulators (LTTRs) is highly abundant among prokaryotes. We analyzed 10,145 non-redundant microbial sequences with homology to eight LysR family regulators of a model prokaryote, Geobacter sulfurreducens, and employed phylogenetic tree inference for LTTR classification. We also analyzed the arrangement of genome clusters containing G. sulfurreducens LTTR genes and searched for LTTR regulatory motifs, suggesting likely regulatory targets of G. sulfurreducens LTTRs. This is the first study to date providing a detailed classification of LTTRs in the deltaproteobacterial family Geobacteraceae.}, issn = {1432-1432}, doi = {10.1007/s00239-012-9498-z}, author = {Krushkal, Julia and Qu, Yanhua and Lovley, Derek R and Adkins, Ronald M} } @article {369, title = {Phylogenetic clustering of soil microbial communities by 16S rRNA but not 16S rRNA genes.}, journal = {Appl Environ Microbiol}, volume = {78}, year = {2012}, month = {2012 Apr}, pages = {2459-61}, abstract = {We evaluated phylogenetic clustering of bacterial and archaeal communities from redox-dynamic subtropical forest soils that were defined by 16S rRNA and rRNA gene sequences. We observed significant clustering for the RNA-based communities but not the DNA-based communities, as well as increasing clustering over time of the highly active taxa detected by only rRNA.}, keywords = {Archaea, Bacteria, Biodiversity, Cluster Analysis, Ecosystem, Genes, rRNA, Oligonucleotide Array Sequence Analysis, Oxidation-Reduction, Phylogeny, RNA, Ribosomal, 16S, Soil Microbiology, Trees, Tropical Climate}, issn = {1098-5336}, doi = {10.1128/AEM.07547-11}, author = {Deangelis, Kristen M and Firestone, Mary K} } @article {849, title = {The prevalence and identity of Chlamydia-specific IgE in children with asthma and other chronic respiratory symptoms.}, journal = {Respir Res}, volume = {13}, year = {2012}, month = {2012 Apr 18}, pages = {32}, abstract = {ABSTRACT: BACKGROUND: Recent studies have confirmed the presence of viable Chlamydia in the bronchoalveolar lavage (BAL) fluid of pediatric patients with airway hyperresponsiveness. While specific IgG and IgM responses to C. pneumoniae are well described, the response and potential contribution of Ag-specific IgE are not known. The current study sought to determine if infection with Chlamydia triggers the production of pathogen-specific IgE in children with chronic respiratory diseases which might contribute to inflammation and pathology. METHODS: We obtained BAL fluid and serum from pediatric respiratory disease patients who were generally unresponsive to corticosteroid treatment as well as sera from age-matched control patients who saw their doctor for wellness checkups. Chlamydia-specific IgE was isolated from BAL and serum samples and their specificity determined by Western blot techniques. The presence of Chlamydia was confirmed by species-specific PCR and BAL culture assays. RESULTS: Chlamydial DNA was detected in the BAL fluid of 134/197 (68\%) patients. Total IgE increased with age until 15 years old and then decreased. Chlamydia-specific IgE was detected in the serum and/or BAL of 107/197 (54\%) patients suffering from chronic respiratory disease, but in none of the 35 healthy control sera (p < 0.0001). Of the 74 BAL culture-positive patients, 68 (91.9\%, p = 0.0001) tested positive for Chlamydia-specific IgE. Asthmatic patients had significantly higher IgE levels compared to non-asthmatics (p = 0.0001). Patients who were positive for Chlamydia DNA or culture had significantly higher levels of serum IgE compared to negative patients (p = 0.0071 and p = 0.0001 respectively). Only 6 chlamydial antigens induced Chlamydia-specific IgE and patients with C. pneumoniae-specific IgE had significantly greater levels of total IgE compared to C. pneumoniae-specific IgE negative ones (p = 0.0001). CONCLUSIONS: IgE antibodies play a central role in allergic inflammation; therefore production of Chlamydia-specific IgE may prove significant in the exacerbation of chronic, allergic airway diseases, thus highlighting a direct role for Chlamydia in asthma pathogenesis.}, issn = {1465-993X}, doi = {10.1186/1465-9921-13-32}, author = {Patel, Katir K and Anderson, Erica A and Salva, Paul S and Webley, Wilmore C} } @article {1446, title = {Probing the mycobacterial trehalome with bioorthogonal chemistry.}, journal = {J Am Chem Soc}, volume = {134}, year = {2012}, month = {2012 Oct 3}, pages = {16123-6}, abstract = {Mycobacteria, including the pathogen Mycobacterium tuberculosis, use the non-mammalian disaccharide trehalose as a precursor for essential cell-wall glycolipids and other metabolites. Here we describe a strategy for exploiting trehalose metabolic pathways to label glycolipids in mycobacteria with azide-modified trehalose (TreAz) analogues. Subsequent bioorthogonal ligation with alkyne-functionalized probes enabled detection and visualization of cell-surface glycolipids. Characterization of the metabolic fates of four TreAz analogues revealed unique labeling routes that can be harnessed for pathway-targeted investigation of the mycobacterial trehalome.
}, keywords = {Alkynes, Azides, Fluorescent Dyes, Glycolipids, Mycobacterium, Trehalose}, issn = {1520-5126}, doi = {10.1021/ja3062419}, author = {Swarts, Benjamin M and Holsclaw, Cynthia M and Jewett, John C and Alber, Marina and Fox, Douglas M and Siegrist, M Sloan and Leary, Julie A and Kalscheuer, Rainer and Bertozzi, Carolyn R} } @article {408, title = {Real-time spatial gene expression analysis within current-producing biofilms.}, journal = {ChemSusChem}, volume = {5}, year = {2012}, month = {2012 Jun}, pages = {1092-8}, abstract = {The expression of genes involved in central metabolism and extracellular electron transfer was examined in real-time in current-producing anode biofilms of Geobacter sulfurreducens. Strains of G. sulfurreducens were generated, in which the expression of the gene for a short half-life fluorescent protein was placed under control of the promoter of the genes of interest. Anode biofilms were grown in a chamber that permitted direct examination of the cell fluorescence with confocal scanning laser microscopy. Studies on nifD and citrate synthase expression in response to environmental changes demonstrated that the reporter system revealed initiation and termination of gene transcription. Uniform expression throughout the biofilms was noted for the genes for citrate synthase; PilA, the structural protein of the conductive pili; and OmcZ, a c-type cytochrome essential for optimal current production, which was localized at the anode-biofilm interface. These results demonstrate that even cells at great distance from the anode, or within expected low-pH zones, are metabolically active and likely to contribute to current production and that there are factors other than gene expression differences influencing the distribution of OmcZ. This real-time reporter approach is likely to be a useful tool in optimizing the design of technologies relying on microbe-electrode interactions.
}, keywords = {Bacterial Proteins, Bioelectric Energy Sources, Biofilms, Citrate (si)-Synthase, Cytochrome c Group, Fimbriae Proteins, Fimbriae, Bacterial, Gene Expression Regulation, Bacterial, Geobacter, Quaternary Ammonium Compounds}, issn = {1864-564X}, doi = {10.1002/cssc.201100714}, author = {Franks, Ashley E and Glaven, Richard H and Lovley, Derek R} } @article {355, title = {RNAi: Future in insect management.}, journal = {J Invertebr Pathol}, year = {2012}, month = {2012 Jul 24}, abstract = {RNA interference is a post- transcriptional, gene regulation mechanism found in virtually all plants and animals including insects. The demonstration of RNAi in insects and its successful use as a tool in the study of functional genomics opened the door to the development of a variety of novel, environmentally sound approaches for insect pest management. Here the current understanding of the biogenesis of the two RNAi classes in insects is reviewed. These are microRNAs (miRNAs) and short interfering RNAs (siRNAs). Several other key approaches in RNAi -based for insect control, as well as for the prevention of diseases in insects are also reviewed. The problems and prospects for the future use of RNAi in insects are presented.}, issn = {1096-0805}, doi = {10.1016/j.jip.2012.07.012}, author = {Burand, John P and Hunter, Wayne B} } @article {3148, title = {The Rnf complex of Clostridium ljungdahlii is a proton-translocating ferredoxin:NAD+ oxidoreductase essential for autotrophic growth.}, journal = {mBio}, volume = {4}, year = {2012}, month = {2012 Dec 26}, pages = {e00406-12}, abstract = {UNLABELLED: It has been predicted that the Rnf complex of Clostridium ljungdahlii is a proton-translocating ferredoxin:NAD(+) oxidoreductase which contributes to ATP synthesis by an H(+)-translocating ATPase under both autotrophic and heterotrophic growth conditions. The recent development of methods for genetic manipulation of C. ljungdahlii made it possible to evaluate the possible role of the Rnf complex in energy conservation. Disruption of the C. ljungdahlii rnf operon inhibited autotrophic growth. ATP synthesis, proton gradient, membrane potential, and proton motive force collapsed in the Rnf-deficient mutant with H(2) as the electron source and CO(2) as the electron acceptor. Heterotrophic growth was hindered in the absence of a functional Rnf complex, as ATP synthesis, proton gradient, and proton motive force were significantly reduced with fructose as the electron donor. Growth of the Rnf-deficient mutant was also inhibited when no source of fixed nitrogen was provided. These results demonstrate that the Rnf complex of C. ljungdahlii is responsible for translocation of protons across the membrane to elicit energy conservation during acetogenesis and is a multifunctional device also implicated in nitrogen fixation.
IMPORTANCE: Mechanisms for energy conservation in the acetogen Clostridium ljungdahlii are of interest because of its potential value as a chassis for the production of biocommodities with novel electron donors such as carbon monoxide, syngas, and electrons derived from electrodes. Characterizing the components implicated in the chemiosmotic ATP synthesis during acetogenesis by C. ljungdahlii is a prerequisite for the development of highly productive strains. The Rnf complex has been considered the prime candidate to be the pump responsible for the formation of an ion gradient coupled with ATP synthesis in multiple acetogens. However, experimental evidence for a proton-pumping Rnf complex has been lacking. This study establishes the C. ljungdahlii Rnf complex as a proton-translocating ferredoxin:NAD(+) oxidoreductase and demonstrates that C. ljungdahlii has the potential of becoming a model organism to study proton translocation, electron transport, and other functions of the Rnf complex in energy conservation or other processes.
}, keywords = {Adenosine Triphosphate, Autotrophic Processes, Clostridium, Energy Metabolism, Fructose, Gene Knockout Techniques, Genes, Essential, Nitrogen, Operon, Oxidoreductases, Proton-Motive Force}, issn = {2150-7511}, doi = {10.1128/mBio.00406-12}, author = {Tremblay, Pier-Luc and Zhang, Tian and Dar, Shabir A and Leang, Ching and Lovley, Derek R} } @article {411, title = {Role of the NiFe hydrogenase Hya in oxidative stress defense in Geobacter sulfurreducens.}, journal = {J Bacteriol}, volume = {194}, year = {2012}, month = {2012 May}, pages = {2248-53}, abstract = {Geobacter sulfurreducens, an Fe(III)-reducing deltaproteobacterium found in anoxic subsurface environments, contains 4 NiFe hydrogenases. Hyb, a periplasmically oriented membrane-bound NiFe hydrogenase, is essential for hydrogen-dependent growth. The functions of the three other hydrogenases are unknown. We show here that the other periplasmically oriented membrane-bound NiFe hydrogenase, Hya, is necessary for growth after exposure to oxidative stress when hydrogen or a highly limiting concentration of acetate is the electron source. The beneficial impact of Hya on growth was dependent on the presence of H(2) in the atmosphere. Moreover, the Hya-deficient strain was more sensitive to the presence of superoxide or hydrogen peroxide. Hya was also required to safeguard Hyb hydrogen oxidation activity after exposure to O(2). Overexpression studies demonstrated that Hya was more resistant to oxidative stress than Hyb. Overexpression of Hya also resulted in the creation of a recombinant strain better fitted for exposure to oxidative stress than wild-type G. sulfurreducens. These results demonstrate that one of the physiological roles of the O(2)-resistant Hya is to participate in the oxidative stress defense of G. sulfurreducens.}, keywords = {Bacterial Proteins, Gene Expression Regulation, Bacterial, Geobacter, Hydrogen Peroxide, Hydrogenase, Mutation, Oxidative Stress, Oxygen, Reactive Oxygen Species, Reverse Transcriptase Polymerase Chain Reaction, Xanthine Oxidase}, issn = {1098-5530}, doi = {10.1128/JB.00044-12}, author = {Tremblay, Pier-Luc and Lovley, Derek R} } @article {699, title = {Spatial heterogeneity of bacterial communities in sediments from an infiltration basin receiving highway runoff.}, journal = {Microb Ecol}, volume = {64}, year = {2012}, month = {2012 Aug}, pages = {461-73}, abstract = {The bacterial community diversity of highway runoff-contaminated sediment that had undergone 19 years of acetate-based de-icing agents addition followed by three years of acetate-free de-icing agents was investigated. Analysis of 26 sediment samples from two drilled soil cores by means of 16S rDNA PCR generated 3,402 clones, indicating an overall high bacterial diversity, with no prominent members within the communities. Sequence analyses provided evidences that each sediment sample displayed a specific structure bacterial community. Proteobacteria-affiliated clones (58\% and 43\% for the two boreholes) predominated in all samples, followed by Actinobacteria (12\% and 16\%), Firmicutes (7\% and 12\%) and Chloroflexi (7\% and 11\%). The subsurface geochemistry complemented the molecular methods to further distinguish ambient and contaminant plume zones. Principal component analysis revealed that the levels of Fe(II) and dissolved oxygen were strongly correlated with bacterial communities. At elevated Fe(II) levels, sequences associated with anaerobic bacteria were detected in high levels. As iron levels declined and oxygen levels increased below the plume bottom, there was a gradual shift in the community structure toward the increase of aerobic bacteria.}, issn = {1432-184X}, doi = {10.1007/s00248-012-0026-x}, author = {Rotaru, Camelia and Woodard, Trevor L and Choi, Seokyoon and Nevin, Kelly P} } @article {414, title = {Supercapacitors based on c-type cytochromes using conductive nanostructured networks of living bacteria.}, journal = {Chemphyschem}, volume = {13}, year = {2012}, month = {2012 Feb}, pages = {463-8}, abstract = {Supercapacitors have attracted interest in energy storage because they have the potential to complement or replace batteries. Here, we report that c-type cytochromes, naturally immersed in a living, electrically conductive microbial biofilm, greatly enhance the device capacitance by over two orders of magnitude. We employ genetic engineering, protein unfolding and Nernstian modeling for in vivo demonstration of charge storage capacity of c-type cytochromes and perform electrochemical impedance spectroscopy, cyclic voltammetry and charge-discharge cycling to confirm the pseudocapacitive, redox nature of biofilm capacitance. The biofilms also show low self-discharge and good charge/discharge reversibility. The superior electrochemical performance of the biofilm is related to its high abundance of cytochromes, providing large electron storage capacity, its nanostructured network with metallic-like conductivity, and its porous architecture with hydrous nature, offering prospects for future low cost and environmentally sustainable energy storage devices.}, keywords = {Bacteria, Biofilms, Cytochrome c Group, Dielectric Spectroscopy, Electric Capacitance, Electrodes, Geobacter, Nanostructures, Oxidation-Reduction}, issn = {1439-7641}, doi = {10.1002/cphc.201100865}, author = {Malvankar, Nikhil S and Mester, T{\"u}nde and Tuominen, Mark T and Lovley, Derek R} } @article {724, title = {Trace elements affect methanogenic activity and diversity in enrichments from subsurface coal bed produced water.}, journal = {Front Microbiol}, volume = {3}, year = {2012}, month = {2012}, pages = {175}, abstract = {Microbial methane from coal beds accounts for a significant and growing percentage of natural gas worldwide. Our knowledge of physical and geochemical factors regulating methanogenesis is still in its infancy. We hypothesized that in these closed systems, trace elements (as micronutrients) are a limiting factor for methanogenic growth and activity. Trace elements are essential components of enzymes or cofactors of metabolic pathways associated with methanogenesis. This study examined the effects of eight trace elements (iron, nickel, cobalt, molybdenum, zinc, manganese, boron, and copper) on methane production, on mcrA transcript levels, and on methanogenic community structure in enrichment cultures obtained from coal bed methane (CBM) well produced water samples from the Powder River Basin, Wyoming. Methane production was shown to be limited both by a lack of additional trace elements as well as by the addition of an overly concentrated trace element mixture. Addition of trace elements at concentrations optimized for standard media enhanced methane production by 37\%. After 7 days of incubation, the levels of mcrA transcripts in enrichment cultures with trace element amendment were much higher than in cultures without amendment. Transcript levels of mcrA correlated positively with elevated rates of methane production in supplemented enrichments (R(2) = 0.95). Metabolically active methanogens, identified by clone sequences of mcrA mRNA retrieved from enrichment cultures, were closely related to Methanobacterium subterraneum and Methanobacterium formicicum. Enrichment cultures were dominated by M. subterraneum and had slightly higher predicted methanogenic richness, but less diversity than enrichment cultures without amendments. These results suggest that varying concentrations of trace elements in produced water from different subsurface coal wells may cause changing levels of CBM production and alter the composition of the active methanogenic community.}, issn = {1664-302X}, doi = {10.3389/fmicb.2012.00175}, author = {Unal, Burcu and Perry, Verlin Ryan and Sheth, Mili and Gomez-Alvarez, Vicente and Chin, Kuk-Jeong and N{\"u}sslein, Klaus} } @article {32, title = {Transcription regulation of the Saccharomyces cerevisiae PHO5 gene by the Ino2p and Ino4p basic helix-loop-helix proteins.}, journal = {Mol Microbiol}, volume = {83}, year = {2012}, month = {2012 Jan}, pages = {395-407}, abstract = {The Saccharomyces cerevisiae PHO5 gene product accounts for a majority of the acid phosphatase activity. Its expression is induced by the basic helix-loop-helix (bHLH) protein, Pho4p, in response to phosphate depletion. Pho4p binds predominantly to two UAS elements (UASp1 at -356 and UASp2 at -247) in the PHO5 promoter. Previous studies from our lab have shown cross-regulation of different biological processes by bHLH proteins. This study tested the ability of all yeast bHLH proteins to regulate PHO5 expression and identified inositol-mediated regulation via the Ino2p/Ino4p bHLH proteins. Ino2p/Ino4p are known regulators of phospholipid biosynthetic genes. Genetic epistasis experiments showed that regulation by inositol required a third UAS site (UASp3 at -194). ChIP assays showed that Ino2p:Ino4p bind the PHO5 promoter and that this binding is dependent on Pho4p binding. These results demonstrate that phospholipid biosynthesis is co-ordinated with phosphate utilization via the bHLH proteins.
}, keywords = {Acid Phosphatase, Basic Helix-Loop-Helix Transcription Factors, Chromatin Immunoprecipitation, DNA, Fungal, Enhancer Elements, Genetic, Gene Expression Regulation, Fungal, Inositol, Models, Biological, Phospholipids, Promoter Regions, Genetic, Protein Binding, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins, Transcription Factors, Transcription, Genetic}, issn = {1365-2958}, doi = {10.1111/j.1365-2958.2011.07941.x}, author = {He, Ying and Swaminathan, Aishwarya and Lopes, John M} } @article {804, title = {Two isoforms of Geobacter sulfurreducens PilA have distinct roles in pilus biogenesis, cytochrome localization, extracellular electron transfer, and biofilm formation.}, journal = {J Bacteriol}, volume = {194}, year = {2012}, month = {2012 May}, pages = {2551-63}, abstract = {Type IV pili of Geobacter sulfurreducens are composed of PilA monomers and are essential for long-range extracellular electron transfer to insoluble Fe(III) oxides and graphite anodes. A previous analysis of pilA expression indicated that transcription was initiated at two positions, with two predicted ribosome-binding sites and translation start codons, potentially producing two PilA preprotein isoforms. The present study supports the existence of two functional translation start codons for pilA and identifies two isoforms (short and long) of the PilA preprotein. The short PilA isoform is found predominantly in an intracellular fraction. It seems to stabilize the long isoform and to influence the secretion of several outer-surface c-type cytochromes. The long PilA isoform is required for secretion of PilA to the outer cell surface, a process that requires coexpression of pilA with nine downstream genes. The long isoform was determined to be essential for biofilm formation on certain surfaces, for optimum current production in microbial fuel cells, and for growth on insoluble Fe(III) oxides.}, keywords = {Amino Acid Sequence, Bacterial Adhesion, Base Sequence, Biofilms, Cytochromes, Electron Transport, Fimbriae Proteins, Fimbriae, Bacterial, Gene Expression Regulation, Bacterial, Geobacter, Molecular Sequence Data, Protein Isoforms, Protein Transport, Reverse Transcriptase Polymerase Chain Reaction, Surface Properties}, issn = {1098-5530}, doi = {10.1128/JB.06366-11}, author = {Richter, Lubna V and Sandler, Steven J and Weis, Robert M} } @article {725, title = {Using flow to switch the valency of bacterial capture on engineered surfaces containing immobilized nanoparticles.}, journal = {Langmuir}, volume = {28}, year = {2012}, month = {2012 May 22}, pages = {7803-10}, abstract = {Toward an understanding of nanoparticle-bacterial interactions and the development of sensors and other substrates for controlled bacterial adhesion, this article describes the influence of flow on the initial stages of bacterial capture (Staphylococcus aureus) on surfaces containing cationic nanoparticles. A PEG (poly(ethylene glycol)) brush on the surface around the nanoparticles sterically repels the bacteria. Variations in ionic strength tune the Debye length from 1 to 4 nm, increasing the strength and range of the nanoparticle attractions toward the bacteria. At relatively high ionic strengths (physiological conditions), bacterial capture requires several nanoparticle-bacterial contacts, termed "multivalent capture". At low ionic strength and gentle wall shear rates (on the order of 10 s(-1)), individual bacteria can be captured and held by single surface-immobilized nanoparticles. Increasing the flow rate to 50 s(-1) causes a shift from monovalent to divalent capture. A comparison of experimental capture efficiencies with statistically determined capture probabilities reveals the initial area of bacteria-surface interaction, here about 50 nm in diameter for a Debye length κ(-1) of 4 nm. Additionally, for κ(-1) = 4 nm, the net per nanoparticle binding energies are strong but highly shear-sensitive, as is the case for biological ligand-receptor interactions. Although these results have been obtained for a specific system, they represent a regime of behavior that could be achieved with different bacteria and different materials, presenting an opportunity for further tuning of selective interactions. These finding suggest the use of surface elements to manipulate individual bacteria and nonfouling designs with precise but finite bacterial interactions.}, issn = {1520-5827}, doi = {10.1021/la205080y}, author = {Fang, Bing and Gon, Saugata and Park, Myoung-Hwan and Kumar, Kushi-Nidhi and Rotello, Vincent M and N{\"u}sslein, Klaus and Santore, Maria M} } @article {421, title = {Anaerobic oxidation of benzene by the hyperthermophilic archaeon Ferroglobus placidus.}, journal = {Appl Environ Microbiol}, volume = {77}, year = {2011}, month = {2011 Sep}, pages = {5926-33}, abstract = {Anaerobic benzene oxidation coupled to the reduction of Fe(III) was studied in Ferroglobus placidus in order to learn more about how such a stable molecule could be metabolized under strict anaerobic conditions. F. placidus conserved energy to support growth at 85{\textdegree}C in a medium with benzene provided as the sole electron donor and Fe(III) as the sole electron acceptor. The stoichiometry of benzene loss and Fe(III) reduction, as well as the conversion of [(14)C]benzene to [(14)C]carbon dioxide, was consistent with complete oxidation of benzene to carbon dioxide with electron transfer to Fe(III). Benzoate, but not phenol or toluene, accumulated at low levels during benzene metabolism, and [(14)C]benzoate was produced from [(14)C]benzene. Analysis of gene transcript levels revealed increased expression of genes encoding enzymes for anaerobic benzoate degradation during growth on benzene versus growth on acetate, but genes involved in phenol degradation were not upregulated during growth on benzene. A gene for a putative carboxylase that was more highly expressed in benzene- than in benzoate-grown cells was identified. These results suggest that benzene is carboxylated to benzoate and that phenol is not an important intermediate in the benzene metabolism of F. placidus. This is the first demonstration of a microorganism in pure culture that can grow on benzene under strict anaerobic conditions and for which there is strong evidence for degradation of benzene via clearly defined anaerobic metabolic pathways. Thus, F. placidus provides a much-needed pure culture model for further studies on the anaerobic activation of benzene in microorganisms.}, keywords = {Anaerobiosis, Archaeoglobales, Benzene, Carbon Radioisotopes, Ferric Compounds, Gene Expression Profiling, Hot Temperature, Isotope Labeling, Oxidation-Reduction}, issn = {1098-5336}, doi = {10.1128/AEM.05452-11}, author = {Holmes, Dawn E and Risso, Carla and Smith, Jessica A and Lovley, Derek R} } @article {729, title = {Bacterial adhesion on hybrid cationic nanoparticle-polymer brush surfaces: ionic strength tunes capture from monovalent to multivalent binding.}, journal = {Colloids Surf B Biointerfaces}, volume = {87}, year = {2011}, month = {2011 Oct 1}, pages = {109-15}, abstract = {This paper describes the creation of hybrid surfaces containing cationic nanoparticles and biocompatible PEG (polyethylene glycol) brushes that manipulate bacterial adhesion for potential diagnostic and implant applications. Here, \~{}10 nm cationically functionalized gold nanoparticles are immobilized randomly on negative silica surfaces at tightly controlled surface loadings, and the remaining areas are functionalized with a hydrated PEG brush, using a graft copolymer of poly-l-lysine and PEG (PLL-PEG), containing 2000 molecular weight PEG chains and roughly 30\% functionalization of the PLL. The cationic nanoparticles attract the negative surfaces of suspended Staphylococcus aureus bacteria while the PEG brush exerts a steric repulsion. With the nanoparticle and PEG brush heights on the same lengthscale, variations in ionic strength are demonstrated to profoundly influence the capture of S. aureus on these surfaces. For bacteria captured from gentle flow, a crossover from multivalent to univalent binding is demonstrated as the Debye length is increased from 1 to 4 nm. In the univalent regime, 1 um diameter spherical bacteria are captured and held by single nanoparticles. In the multivalent regime, there is an adhesion threshold in the surface density of nanoparticles needed for bacterial capture. The paper also documents an interesting effect concerning the relaxations in the PLL-PEG brush itself. For brushy surfaces containing no nanoparticles, bacterial adhesion persists on newly formed brushes, but is nearly eliminated after these brushes relax, at constant mass in buffer for 12h. Thus brushy relaxations increase biocompatibility.}, keywords = {Bacterial Adhesion, Cations, Nanoparticles, Osmolar Concentration, Polyethylene Glycols, Polylysine, Silicon Dioxide, Staphylococcus aureus, Static Electricity, Surface Properties}, issn = {1873-4367}, doi = {10.1016/j.colsurfb.2011.05.010}, author = {Fang, Bing and Gon, Saugata and Park, Myoung and Kumar, Kushi-Nidhi and Rotello, Vincent M and Nusslein, Klaus and Santore, Maria M} } @article {1971, title = {Benchmarking a luciferase complementation assay for detecting protein complexes}, journal = {Nature Methods}, volume = {8}, year = {2011}, month = {nov}, pages = {990{\textendash}992}, doi = {10.1038/nmeth.1773}, url = {https://doi.org/10.1038/nmeth.1773}, author = {Patricia Cassonnet and Caroline Rolloy and Gr{\'e}gory Neveu and Pierre-Olivier Vidalain and Thibault Chantier and Johann Pellet and Louis Jones and Mandy Muller and Caroline Demeret and Guillaume Gaud and Fran{\c c}oise Vuillier and Vincent Lotteau and Fr{\'e}d{\'e}ric Tangy and Michel Favre and Yves Jacob} } @article {427, title = {Biochemical characterization of purified OmcS, a c-type cytochrome required for insoluble Fe(III) reduction in Geobacter sulfurreducens.}, journal = {Biochim Biophys Acta}, volume = {1807}, year = {2011}, month = {2011 Apr}, pages = {404-12}, abstract = {Previous studies with Geobacter sulfurreducens have demonstrated that OmcS, an abundant c-type cytochrome that is only loosely bound to the outer surface, plays an important role in electron transfer to Fe(III) oxides as well as other extracellular electron acceptors. In order to further investigate the function of OmcS, it was purified from a strain that overproduces the protein. Purified OmcS had a molecular mass of 47015 Da, and six low-spin bis-histidinyl hexacoordinated heme groups. Its midpoint redox potential was -212 mV. A thermal stability analysis showed that the cooperative melting of purified OmcS occurs in the range of 65-82 {\textdegree}C. Far UV circular dichroism spectroscopy indicated that the secondary structure of purified OmcS consists of about 10\% α-helix and abundant disordered structures. Dithionite-reduced OmcS was able to transfer electrons to a variety of substrates of environmental importance including insoluble Fe(III) oxide, Mn(IV) oxide and humic substances. Stopped flow analysis revealed that the reaction rate of OmcS oxidation has a hyperbolic dependence on the concentration of the studied substrates. A ten-fold faster reaction rate with anthraquinone-2,6-disulfonate (AQDS) (25.2 s$^{-}${\textonesuperior}) was observed as compared to that with Fe(III) citrate (2.9 s$^{-}${\textonesuperior}). The results, coupled with previous localization and gene deletion studies, suggest that OmcS is well-suited to play an important role in extracellular electron transfer.}, keywords = {Circular Dichroism, Cytochrome c Group, Geobacter, Heme, Iron, Kinetics, Molecular Weight, Oxidation-Reduction, Solubility}, issn = {0006-3002}, doi = {10.1016/j.bbabio.2011.01.003}, author = {Qian, Xinlei and Mester, T{\"u}nde and Morgado, Leonor and Arakawa, Tsutomu and Sharma, Manju L and Inoue, Kengo and Joseph, Crisjoe and Salgueiro, Carlos A and Maroney, Michael J and Lovley, Derek R} } @article {1567, title = {Characterization of a novel facultative Methylocystis species capable of growth on methane, acetate and ethanol.}, journal = {Environ Microbiol Rep}, volume = {3}, year = {2011}, month = {2011 Apr}, pages = {174-81}, abstract = {A non-motile strain of Methylocystis, strain SB2, isolated from a spring bog in southeast Michigan, had a curved rod morphology with a typical type II intracytoplasmic membrane system. This organism expressed the membrane-bound or particulate methane monooxygenase (pMMO) as well as a chalkophore with high affinity for copper and did not express the cytoplasmic or soluble methane monooxygenase (sMMO). Strain SB2 was found to grow within the pH range of 6-9, with optimal growth at 6.8. Growth was observed at temperatures ranging between 10{\textdegree}C and 30{\textdegree}C, with no growth at 37{\textdegree}C. The DNA G+C content was 62.9 mol\%. Predominant fatty acids were 18:1ω7c (72.7\%) and 18:1ω9c (24\%) when grown on methane. Phylogenetic comparisons based on both pmoA and 16S rRNA sequences indicated that this organism belonged to the Methylocystis genus, and was closely related to Methylocystis rosea SV97(T) and Methylocystis echinoides IMET10491(T) (98\% 16S rRNA gene sequence similarity to both strains). DNA : DNA hybridizations indicated that strain SB2 had 70\% similarity with M. rosea SV97(T) . Unlike M. rosea SV97(T) , strain SB2 was able to utilize not only methane for growth, but also ethanol and acetate. Furthermore, the predominant fatty acids in strain SB2 were different from those found in M. rosea SV97(T) , i.e. 54.2\% and 39.7\% of fatty acids are 18:1ω8 and 18:1ω7 in M. rosea SV97(T) , while 18:1ω8 is completely absent in strain SB2.
}, issn = {1758-2229}, doi = {10.1111/j.1758-2229.2010.00204.x}, author = {Im, Jeongdae and Lee, Sung-Woo and Yoon, Sukhwan and Dispirito, Alan A and Semrau, Jeremy D} } @article {375, title = {Characterization of trapped lignin-degrading microbes in tropical forest soil.}, journal = {PLoS One}, volume = {6}, year = {2011}, month = {2011}, pages = {e19306}, abstract = {Lignin is often the most difficult portion of plant biomass to degrade, with fungi generally thought to dominate during late stage decomposition. Lignin in feedstock plant material represents a barrier to more efficient plant biomass conversion and can also hinder enzymatic access to cellulose, which is critical for biofuels production. Tropical rain forest soils in Puerto Rico are characterized by frequent anoxic conditions and fluctuating redox, suggesting the presence of lignin-degrading organisms and mechanisms that are different from known fungal decomposers and oxygen-dependent enzyme activities. We explored microbial lignin-degraders by burying bio-traps containing lignin-amended and unamended biosep beads in the soil for 1, 4, 13 and 30 weeks. At each time point, phenol oxidase and peroxidase enzyme activity was found to be elevated in the lignin-amended versus the unamended beads, while cellulolytic enzyme activities were significantly depressed in lignin-amended beads. Quantitative PCR of bacterial communities showed more bacterial colonization in the lignin-amended compared to the unamended beads after one and four weeks, suggesting that the lignin supported increased bacterial abundance. The microbial community was analyzed by small subunit 16S ribosomal RNA genes using microarray (PhyloChip) and by high-throughput amplicon pyrosequencing based on universal primers targeting bacterial, archaeal, and eukaryotic communities. Community trends were significantly affected by time and the presence of lignin on the beads. Lignin-amended beads have higher relative abundances of representatives from the phyla Actinobacteria, Firmicutes, Acidobacteria and Proteobacteria compared to unamended beads. This study suggests that in low and fluctuating redox soils, bacteria could play a role in anaerobic lignin decomposition.}, keywords = {Biodiversity, Biomass, Ecosystem, Gases, Lignin, Oligonucleotide Array Sequence Analysis, Phylogeny, Plants, RNA, Ribosomal, 16S, Sequence Analysis, DNA, Soil Microbiology, Trees}, issn = {1932-6203}, doi = {10.1371/journal.pone.0019306}, author = {Deangelis, Kristen M and Allgaier, Martin and Chavarria, Yaucin and Fortney, Julian L and Hugenholtz, Phillip and Simmons, Blake and Sublette, Kerry and Silver, Whendee L and Hazen, Terry C} } @article {851, title = {Colonization of paediatric lower respiratory tract with genital Mycoplasma species.}, journal = {Respirology}, volume = {16}, year = {2011}, month = {2011 Oct}, pages = {1081-7}, abstract = {BACKGROUND AND OBJECTIVE: Recently, much attention has been given to the possible role played by pathogens that colonize neonatal or paediatric airway and their potential involvement in chronic respiratory disease. The goal of the current study was to evaluate the prevalence of Mycoplasma organisms in the BAL fluid of paediatric patients suffering from a variety of chronic respiratory diseases to determine if there was any clear disease association with bacterial presence. METHODS: We examined 319 paediatric BAL samples for the presence of M.genitalium, M.hominis, U.urealyticum, U.parvum and M.pneumoniae DNA with species-specific PCR. RESULTS: Mycoplasma DNA was found in 32.6\% (104/319) of patient samples; 10\% (32/319) for M.pneumoniae, 8.8\% for U.parvum, 2.8\% for U.urealyticum; 4.7\% for M.hominis and 9.1\% for M.genitalium. There were no significant clinical and laboratory differences except serum IgE in asthmatics according to Mycoplasma colonization or not. Elevated levels of IgE were found more often in Mycoplasma DNA-negative patients than patients with bacterial DNA, 85/109 versus 24/109 respectively (P<0.0001). There was no difference in the frequency of Mycoplasmas between the asthmatics and the non-asthmatics; 30.6\% (69/225) versus 37.2\% (35/94) for Mycoplasma 16S DNA, and 8\% versus 14.9\% for M.pneumoniae, respectively. CONCLUSIONS: Our data indicate that in addition to M.pneumoniae, urogenital Mycoplasma species may colonize the airway of patients with chronic respiratory diseases. There was, however, no association between chronic asthma diagnosis and Mycoplasma colonization in this study.}, keywords = {Adolescent, Asthma, Child, Child, Preschool, Cohort Studies, DNA, Bacterial, Female, Humans, Infant, Infant, Newborn, Male, Mycoplasma, Mycoplasma genitalium, Mycoplasma hominis, Mycoplasma Infections, Mycoplasma pneumoniae, Respiratory System, Species Specificity, Ureaplasma, Ureaplasma Infections, Ureaplasma urealyticum, Young Adult}, issn = {1440-1843}, doi = {10.1111/j.1440-1843.2011.02016.x}, author = {Patel, Katir K and Salva, Paul S and Webley, Wilmore C} } @article {371, title = {Complete genome sequence of "Enterobacter lignolyticus" SCF1.}, journal = {Stand Genomic Sci}, volume = {5}, year = {2011}, month = {2011 Oct 15}, pages = {69-85}, abstract = {In an effort to discover anaerobic bacteria capable of lignin degradation, we isolated "Enterobacter lignolyticus" SCF1 on minimal media with alkali lignin as the sole source of carbon. This organism was isolated anaerobically from tropical forest soils collected from the Short Cloud Forest site in the El Yunque National Forest in Puerto Rico, USA, part of the Luquillo Long-Term Ecological Research Station. At this site, the soils experience strong fluctuations in redox potential and are net methane producers. Because of its ability to grow on lignin anaerobically, we sequenced the genome. The genome of "E. lignolyticus" SCF1 is 4.81 Mbp with no detected plasmids, and includes a relatively small arsenal of lignocellulolytic carbohydrate active enzymes. Lignin degradation was observed in culture, and the genome revealed two putative laccases, a putative peroxidase, and a complete 4-hydroxyphenylacetate degradation pathway encoded in a single gene cluster.}, issn = {1944-3277}, doi = {10.4056/sigs.2104875}, author = {Deangelis, Kristen M and D{\textquoteright}haeseleer, Patrik and Chivian, Dylan and Fortney, Julian L and Khudyakov, Jane and Simmons, Blake and Woo, Hannah and Arkin, Adam P and Davenport, Karen Walston and Goodwin, Lynne and Chen, Amy and Ivanova, Natalia and Kyrpides, Nikos C and Mavromatis, Konstantinos and Woyke, Tanja and Hazen, Terry C} } @article {1566, title = {Constitutive expression of pMMO by Methylocystis strain SB2 when grown on multi-carbon substrates: implications for biodegradation of chlorinated ethenes.}, journal = {Environ Microbiol Rep}, volume = {3}, year = {2011}, month = {2011 Apr}, pages = {182-8}, abstract = {The particulate methane monooxygenase (pMMO) in Methylocystis strain SB2 was found to be constitutively expressed in the absence of methane when the strain was grown on either acetate or ethanol. Real-time quantitative polymerase chain reaction (PCR) and reverse transcription-PCR showed that the expression of pmoA decreased by one to two orders of magnitude when grown on acetate as compared with growth of strain SB2 on methane. The capability of strain SB2 to degrade a mixture of chlorinated ethenes in the absence of methane was examined to verify the presence and activity of pMMO under acetate-growth conditions as well determine the effectiveness of such conditions for bioremediation. It was found that when strain SB2 was grown on acetate and exposed to 40 {\textmu}M each of trichloroethylene (TCE), trans-dichloroethylene (t-DCE) and vinyl chloride (VC), approximately 30\% of VC and t-DCE was degraded but no appreciable TCE removal was measured after 216 h of incubation. The ability to degrade VC and t-DCE was lost when acetylene was added, confirming that pMMO was responsible for the degradation of these chlorinated ethenes by Methylocystis strain SB2 when the strain was grown on acetate.
}, issn = {1758-2229}, doi = {10.1111/j.1758-2229.2010.00205.x}, author = {Yoon, Sukhwan and Im, Jeongdae and Bandow, Nathan and Dispirito, Alan A and Semrau, Jeremy D} } @article {441, title = {A c-type cytochrome and a transcriptional regulator responsible for enhanced extracellular electron transfer in Geobacter sulfurreducens revealed by adaptive evolution.}, journal = {Environ Microbiol}, volume = {13}, year = {2011}, month = {2011 Jan}, pages = {13-23}, abstract = {The stimulation of subsurface microbial metabolism often associated with engineered bioremediation of groundwater contaminants presents subsurface microorganisms, which are adapted for slow growth and metabolism in the subsurface, with new selective pressures. In order to better understand how Geobacter species might adapt to selective pressure for faster metal reduction in the subsurface, Geobacter sulfurreducens was put under selective pressure for rapid Fe(III) oxide reduction. The genomes of two resultant strains with rates of Fe(III) oxide reduction that were 10-fold higher than those of the parent strain were resequenced. Both strains contain either a single base-pair change or a 1 nucleotide insertion in a GEMM riboswitch upstream of GSU1761, a gene coding for the periplasmic c-type cytochrome designated PgcA. GSU1771, a gene coding for a SARP regulator, was also mutated in both strains. Introduction of either of the GEMM riboswitch mutations upstream of pgcA in the wild-type increased the abundance of pgcA transcripts, consistent with increased expression of pgcA in the adapted strains. One of the mutations doubled the rate of Fe(III) oxide reduction. Interruption of GSU1771 doubled the Fe(III) oxide reduction rate. This was associated with an increased in expression of pilA, the gene encoding the structural protein for the pili thought to function as microbial nanowires. The combination of the GSU1771 interruption with either of the pgcA mutations resulted in a strain that reduced Fe(III) as fast as the comparable adapted strain. These results suggest that the accumulation of a small number of beneficial mutations under selective pressure, similar to that potentially present during bioremediation, can greatly enhance the capacity for Fe(III) oxide reduction in G. sulfurreducens. Furthermore, the results emphasize the importance of the c-type cytochrome PgcA and pili in Fe(III) oxide reduction and demonstrate how adaptive evolution studies can aid in the elucidation of complex mechanisms, such as extracellular electron transfer.}, keywords = {Adaptation, Physiological, Biodegradation, Environmental, Cytochrome c Group, DNA, Bacterial, Electron Transport, Evolution, Molecular, Ferric Compounds, Gene Expression Profiling, Genes, Bacterial, Genome, Bacterial, Geobacter, Mutagenesis, Insertional, Mutation, Oligonucleotide Array Sequence Analysis, Oxidation-Reduction, Riboswitch, Sequence Analysis, DNA}, issn = {1462-2920}, doi = {10.1111/j.1462-2920.2010.02302.x}, author = {Tremblay, Pier-Luc and Summers, Zarath M and Glaven, Richard H and Nevin, Kelly P and Zengler, Karsten and Barrett, Christian L and Qiu, Yu and Palsson, Bernhard O and Lovley, Derek R} } @article {426, title = {Development of a biomarker for Geobacter activity and strain composition; proteogenomic analysis of the citrate synthase protein during bioremediation of U(VI).}, journal = {Microb Biotechnol}, volume = {4}, year = {2011}, month = {2011 Jan}, pages = {55-63}, abstract = {Monitoring the activity of target microorganisms during stimulated bioremediation is a key problem for the development of effective remediation strategies. At the US Department of Energy{\textquoteright}s Integrated Field Research Challenge (IFRC) site in Rifle, CO, the stimulation of Geobacter growth and activity via subsurface acetate addition leads to precipitation of U(VI) from groundwater as U(IV). Citrate synthase (gltA) is a key enzyme in Geobacter central metabolism that controls flux into the TCA cycle. Here, we utilize shotgun proteomic methods to demonstrate that the measurement of gltA peptides can be used to track Geobacter activity and strain evolution during in situ biostimulation. Abundances of conserved gltA peptides tracked Fe(III) reduction and changes in U(VI) concentrations during biostimulation, whereas changing patterns of unique peptide abundances between samples suggested sample-specific strain shifts within the Geobacter population. Abundances of unique peptides indicated potential differences at the strain level between Fe(III)-reducing populations stimulated during in situ biostimulation experiments conducted a year apart at the Rifle IFRC. These results offer a novel technique for the rapid screening of large numbers of proteomic samples for Geobacter species and will aid monitoring of subsurface bioremediation efforts that rely on metal reduction for desired outcomes.}, keywords = {Amino Acid Sequence, Bacterial Proteins, Biodegradation, Environmental, Biological Markers, Citrate (si)-Synthase, Geobacter, Groundwater, Molecular Sequence Data, Phylogeny, Proteomics, Sequence Alignment, Uranium}, issn = {1751-7915}, doi = {10.1111/j.1751-7915.2010.00194.x}, author = {Wilkins, Michael J and Callister, Stephen J and Miletto, Marzia and Williams, Kenneth H and Nicora, Carrie D and Lovley, Derek R and Long, Philip E and Lipton, Mary S} } @article {429, title = {Direct coupling of a genome-scale microbial in silico model and a groundwater reactive transport model.}, journal = {J Contam Hydrol}, volume = {122}, year = {2011}, month = {2011 Mar 25}, pages = {96-103}, abstract = {The activity of microorganisms often plays an important role in dynamic natural attenuation or engineered bioremediation of subsurface contaminants, such as chlorinated solvents, metals, and radionuclides. To evaluate and/or design bioremediated systems, quantitative reactive transport models are needed. State-of-the-art reactive transport models often ignore the microbial effects or simulate the microbial effects with static growth yield and constant reaction rate parameters over simulated conditions, while in reality microorganisms can dynamically modify their functionality (such as utilization of alternative respiratory pathways) in response to spatial and temporal variations in environmental conditions. Constraint-based genome-scale microbial in silico models, using genomic data and multiple-pathway reaction networks, have been shown to be able to simulate transient metabolism of some well studied microorganisms and identify growth rate, substrate uptake rates, and byproduct rates under different growth conditions. These rates can be identified and used to replace specific microbially-mediated reaction rates in a reactive transport model using local geochemical conditions as constraints. We previously demonstrated the potential utility of integrating a constraint-based microbial metabolism model with a reactive transport simulator as applied to bioremediation of uranium in groundwater. However, that work relied on an indirect coupling approach that was effective for initial demonstration but may not be extensible to more complex problems that are of significant interest (e.g., communities of microbial species and multiple constraining variables). Here, we extend that work by presenting and demonstrating a method of directly integrating a reactive transport model (FORTRAN code) with constraint-based in silico models solved with IBM ILOG CPLEX linear optimizer base system (C library). The models were integrated with BABEL, a language interoperability tool. The modeling system is designed in such a way that constraint-based models targeting different microorganisms or competing organism communities can be easily plugged into the system. Constraint-based modeling is very costly given the size of a genome-scale reaction network. To save computation time, a binary tree is traversed to examine the concentration and solution pool generated during the simulation in order to decide whether the constraint-based model should be called. We also show preliminary results from the integrated model including a comparison of the direct and indirect coupling approaches and evaluated the ability of the approach to simulate field experiment.}, keywords = {Biodegradation, Environmental, Biological Transport, Colorado, Computer Simulation, Genome, Bacterial, Geobacter, Models, Biological, Uranium}, issn = {1873-6009}, doi = {10.1016/j.jconhyd.2010.11.007}, author = {Fang, Yilin and Scheibe, Timothy D and Mahadevan, Radhakrishnan and Garg, Srinath and Long, Philip E and Lovley, Derek R} } @article {374, title = {Effects of selected root exudate components on soil bacterial communities.}, journal = {FEMS Microbiol Ecol}, volume = {77}, year = {2011}, month = {2011 Sep}, pages = {600-10}, abstract = {Low-molecular-weight organic compounds in root exudates play a key role in plant-microorganism interactions by influencing the structure and function of soil microbial communities. Model exudate solutions, based on organic acids (OAs) (quinic, lactic, maleic acids) and sugars (glucose, sucrose, fructose), previously identified in the rhizosphere of Pinus radiata, were applied to soil microcosms. Root exudate compound solutions stimulated soil dehydrogenase activity and the addition of OAs increased soil pH. The structure of active bacterial communities, based on reverse-transcribed 16S rRNA gene PCR, was assessed by denaturing gradient gel electrophoresis and PhyloChip microarrays. Bacterial taxon richness was greater in all treatments than that in control soil, with a wide range of taxa (88-1043) responding positively to exudate solutions and fewer (<24) responding negatively. OAs caused significantly greater increases than sugars in the detectable richness of the soil bacterial community and larger shifts of dominant taxa. The greater response of bacteria to OAs may be due to the higher amounts of added carbon, solubilization of soil organic matter or shifts in soil pH. Our results indicate that OAs play a significant role in shaping soil bacterial communities and this may therefore have a significant impact on plant growth.}, keywords = {Bacteria, DNA, Bacterial, Molecular Sequence Data, Organic Chemicals, Phylogeny, Pinus, Plant Exudates, Plant Roots, Rhizosphere, RNA, Ribosomal, 16S, Soil, Soil Microbiology}, issn = {1574-6941}, doi = {10.1111/j.1574-6941.2011.01150.x}, author = {Shi, Shengjing and Richardson, Alan E and O{\textquoteright}Callaghan, Maureen and Deangelis, Kristen M and Jones, Eirian E and Stewart, Alison and Firestone, Mary K and Condron, Leo M} } @article {424, title = {Electrosynthesis of organic compounds from carbon dioxide is catalyzed by a diversity of acetogenic microorganisms.}, journal = {Appl Environ Microbiol}, volume = {77}, year = {2011}, month = {2011 May}, pages = {2882-6}, abstract = {Microbial electrosynthesis, a process in which microorganisms use electrons derived from electrodes to reduce carbon dioxide to multicarbon, extracellular organic compounds, is a potential strategy for capturing electrical energy in carbon-carbon bonds of readily stored and easily distributed products, such as transportation fuels. To date, only one organism, the acetogen Sporomusa ovata, has been shown to be capable of electrosynthesis. The purpose of this study was to determine if a wider range of microorganisms is capable of this process. Several other acetogenic bacteria, including two other Sporomusa species, Clostridium ljungdahlii, Clostridium aceticum, and Moorella thermoacetica, consumed current with the production of organic acids. In general acetate was the primary product, but 2-oxobutyrate and formate also were formed, with 2-oxobutyrate being the predominant identified product of electrosynthesis by C. aceticum. S. sphaeroides, C. ljungdahlii, and M. thermoacetica had high (>80\%) efficiencies of electrons consumed and recovered in identified products. The acetogen Acetobacterium woodii was unable to consume current. These results expand the known range of microorganisms capable of electrosynthesis, providing multiple options for the further optimization of this process.}, keywords = {Acetobacterium, Carbon Dioxide, Clostridium, Electrodes, Electrons, Moorella, Organic Chemicals, Oxidation-Reduction, Veillonellaceae}, issn = {1098-5336}, doi = {10.1128/AEM.02642-10}, author = {Nevin, Kelly P and Hensley, Sarah A and Franks, Ashley E and Summers, Zarath M and Ou, Jianhong and Woodard, Trevor L and Snoeyenbos-West, Oona L and Lovley, Derek R} } @article {1569, title = {Field application of nitrogen and phenylacetylene to mitigate greenhouse gas emissions from landfill cover soils: effects on microbial community structure.}, journal = {Appl Microbiol Biotechnol}, volume = {89}, year = {2011}, month = {2011 Jan}, pages = {189-200}, abstract = {Landfills are large sources of CH(4), but a considerable amount of CH(4) can be removed in situ by methanotrophs if their activity can be stimulated through the addition of nitrogen. Nitrogen can, however, lead to increased N(2)O production. To examine the effects of nitrogen and a selective inhibitor on CH(4) oxidation and N(2)O production in situ, 0.5 M of NH(4)Cl and 0.25 M of KNO(3), with and without 0.01\% (w/v) phenylacetylene, were applied to test plots at a landfill in Kalamazoo, MI from 2007 November to 2009 July. Nitrogen amendments stimulated N(2)O production but had no effect on CH(4) oxidation. The addition of phenylacetylene stimulated CH(4) oxidation while reducing N(2)O production. Methanotrophs possessing particulate methane monooxygenase and archaeal ammonia-oxidizers (AOAs) were abundant. The addition of nitrogen reduced methanotrophic diversity, particularly for type I methanotrophs. The simultaneous addition of phenylacetylene increased methanotrophic diversity and the presence of type I methanotrophs. Clone libraries of the archaeal amoA gene showed that the addition of nitrogen increased AOAs affiliated with Crenarchaeal group 1.1b, while they decreased with the simultaneous addition of phenylacetylene. These results suggest that the addition of phenylacetylene with nitrogen reduces N(2)O production by selectively inhibiting AOAs and/or type II methanotrophs.
}, keywords = {Acetylene, Archaea, Archaeal Proteins, Bacteria, Bacterial Proteins, Gases, Greenhouse Effect, Methane, Molecular Sequence Data, Nitrogen, Refuse Disposal, Soil, Soil Microbiology}, issn = {1432-0614}, doi = {10.1007/s00253-010-2811-0}, author = {Im, Jeongdae and Lee, Sung-Woo and Bodrossy, Levente and Barcelona, Michael J and Semrau, Jeremy D} } @article {437, title = {Gene expression and deletion analysis of mechanisms for electron transfer from electrodes to Geobacter sulfurreducens.}, journal = {Bioelectrochemistry}, volume = {80}, year = {2011}, month = {2011 Feb}, pages = {142-50}, abstract = {Geobacter sulfurreducens is one of the few microorganisms available in pure culture known to directly accept electrons from a negatively poised electrode. Microarray analysis was used to compare gene transcript abundance in biofilms of G. sulfurreducens using a graphite electrode as the sole electron donor for fumarate reduction compared with transcript abundance in biofilms growing on the same material, but not consuming current. Surprisingly, genes for putative cell-electrode connections, such as outer-surface cytochromes and pili, which are highly expressed in current-producing biofilms, were not highly expressed in current-consuming biofilms. Microarray analysis of G. sulfurreducens gene transcript abundance in current-consuming biofilms versus current-producing biofilms gave similar results. In both comparative studies current-consuming biofilms had greater transcript abundance for a gene (GSU3274) encoding a putative monoheme, c-type cytochrome. Deletion of genes for outer-surface proteins previously shown to be essential for optimal electron transfer to electrodes had no impact on electron transfer from electrodes. Deletion of GSU3274 completely inhibited electron transfer from electrodes, but had no impact on electron transfer to electrodes. These differences in gene expression patterns and the impact of gene deletions suggest that the mechanisms for electron transfer from electrodes to G. sulfurreducens differ significantly from the mechanisms for electron transfer to electrodes.}, keywords = {Bacterial Proteins, Biofilms, Cytochromes, Electrodes, Electron Transport, Electrons, Gene Expression, Geobacter, Graphite, Oligonucleotide Array Sequence Analysis, Oxidation-Reduction, Sequence Deletion}, issn = {1878-562X}, doi = {10.1016/j.bioelechem.2010.07.005}, author = {Strycharz, Sarah M and Glaven, Richard H and Coppi, Maddalena V and Gannon, Sarah M and Perpetua, Lorrie A and Liu, Anna and Nevin, Kelly P and Lovley, Derek R} } @article {422, title = {Genome diversity of the TetR family of transcriptional regulators in a metal-reducing bacterial family Geobacteraceae and other microbial species.}, journal = {OMICS}, volume = {15}, year = {2011}, month = {2011 Jul-Aug}, pages = {495-506}, abstract = {Members of the TetR family of bacterial transcriptional regulators affect expression of genes whose products are involved in a variety of important functions, including osmotic stress, catabolic pathways, homeostasis, biosynthesis of antibiotics, expression of efflux pumps, multidrug resistance, and virulence of pathogenic bacteria. We used genome sequence information to carry out phylogenetic classification of 864 TetR family members with a special focus on TetR regulators in Geobacteraceae, an environmentally important family of delta-Proteobacteria. The genome of Geobacter sulfurreducens, a model representative of Geobacteraceae, contains nine genes from the tetR family. Several of these genes are located immediately upstream of operons encoding functionally important c-type cytochromes. Computational analyses identified the presence of conserved promoters and other regulatory binding sites upstream of several G. sulfurreducens tetR genes. This suggests the possibility of an intermediary role of TetR family proteins in Geobacteraceae in regulatory cascades involving a variety of sigma factors. In order to understand the role of the TetR regulatory family in Geobacteraceae, we have inferred phylogenetic relationships among the Geobacteraceae TetR proteins and their homologs in other microbial species.}, keywords = {Bacterial Proteins, Binding Sites, Gram-Negative Bacteria, Metals, Oxidation-Reduction, Phylogeny, Promoter Regions, Genetic, Sigma Factor}, issn = {1557-8100}, doi = {10.1089/omi.2010.0117}, author = {Krushkal, Julia and Sontineni, Sreedhar and Leang, Ching and Qu, Yanhua and Adkins, Ronald M and Lovley, Derek R} } @article {438, title = {Genome-scale dynamic modeling of the competition between Rhodoferax and Geobacter in anoxic subsurface environments.}, journal = {ISME J}, volume = {5}, year = {2011}, month = {2011 Feb}, pages = {305-16}, abstract = {The advent of rapid complete genome sequencing, and the potential to capture this information in genome-scale metabolic models, provide the possibility of comprehensively modeling microbial community interactions. For example, Rhodoferax and Geobacter species are acetate-oxidizing Fe(III)-reducers that compete in anoxic subsurface environments and this competition may have an influence on the in situ bioremediation of uranium-contaminated groundwater. Therefore, genome-scale models of Geobacter sulfurreducens and Rhodoferax ferrireducens were used to evaluate how Geobacter and Rhodoferax species might compete under diverse conditions found in a uranium-contaminated aquifer in Rifle, CO. The model predicted that at the low rates of acetate flux expected under natural conditions at the site, Rhodoferax will outcompete Geobacter as long as sufficient ammonium is available. The model also predicted that when high concentrations of acetate are added during in situ bioremediation, Geobacter species would predominate, consistent with field-scale observations. This can be attributed to the higher expected growth yields of Rhodoferax and the ability of Geobacter to fix nitrogen. The modeling predicted relative proportions of Geobacter and Rhodoferax in geochemically distinct zones of the Rifle site that were comparable to those that were previously documented with molecular techniques. The model also predicted that under nitrogen fixation, higher carbon and electron fluxes would be diverted toward respiration rather than biomass formation in Geobacter, providing a potential explanation for enhanced in situ U(VI) reduction in low-ammonium zones. These results show that genome-scale modeling can be a useful tool for predicting microbial interactions in subsurface environments and shows promise for designing bioremediation strategies.}, keywords = {Acetates, Anaerobiosis, Biodegradation, Environmental, Biomass, Comamonadaceae, Genome, Genome, Bacterial, Geobacter, Models, Biological, Nitrogen Fixation, Quaternary Ammonium Compounds, RNA, Ribosomal, 16S, Uranium, Water Microbiology, Water Pollutants, Radioactive}, issn = {1751-7370}, doi = {10.1038/ismej.2010.117}, author = {Zhuang, Kai and Izallalen, Mounir and Mouser, Paula and Richter, Hanno and Risso, Carla and Mahadevan, Radhakrishnan and Lovley, Derek R} } @article {415, title = {Geobacter: the microbe electric{\textquoteright}s physiology, ecology, and practical applications.}, journal = {Adv Microb Physiol}, volume = {59}, year = {2011}, month = {2011}, pages = {1-100}, abstract = {Geobacter species specialize in making electrical contacts with extracellular electron acceptors and other organisms. This permits Geobacter species to fill important niches in a diversity of anaerobic environments. Geobacter species appear to be the primary agents for coupling the oxidation of organic compounds to the reduction of insoluble Fe(III) and Mn(IV) oxides in many soils and sediments, a process of global biogeochemical significance. Some Geobacter species can anaerobically oxidize aromatic hydrocarbons and play an important role in aromatic hydrocarbon removal from contaminated aquifers. The ability of Geobacter species to reductively precipitate uranium and related contaminants has led to the development of bioremediation strategies for contaminated environments. Geobacter species produce higher current densities than any other known organism in microbial fuel cells and are common colonizers of electrodes harvesting electricity from organic wastes and aquatic sediments. Direct interspecies electron exchange between Geobacter species and syntrophic partners appears to be an important process in anaerobic wastewater digesters. Functional and comparative genomic studies have begun to reveal important aspects of Geobacter physiology and regulation, but much remains unexplored. Quantifying key gene transcripts and proteins of subsurface Geobacter communities has proven to be a powerful approach to diagnose the in situ physiological status of Geobacter species during groundwater bioremediation. The growth and activity of Geobacter species in the subsurface and their biogeochemical impact under different environmental conditions can be predicted with a systems biology approach in which genome-scale metabolic models are coupled with appropriate physical/chemical models. The proficiency of Geobacter species in transferring electrons to insoluble minerals, electrodes, and possibly other microorganisms can be attributed to their unique "microbial nanowires," pili that conduct electrons along their length with metallic-like conductivity. Surprisingly, the abundant c-type cytochromes of Geobacter species do not contribute to this long-range electron transport, but cytochromes are important for making the terminal electrical connections with Fe(III) oxides and electrodes and also function as capacitors, storing charge to permit continued respiration when extracellular electron acceptors are temporarily unavailable. The high conductivity of Geobacter pili and biofilms and the ability of biofilms to function as supercapacitors are novel properties that might contribute to the field of bioelectronics. The study of Geobacter species has revealed a remarkable number of microbial physiological properties that had not previously been described in any microorganism. Further investigation of these environmentally relevant and physiologically unique organisms is warranted.}, keywords = {Biotechnology, Ecology, Environmental Remediation, Ferric Compounds, Geobacter}, issn = {0065-2911}, doi = {10.1016/B978-0-12-387661-4.00004-5}, author = {Lovley, Derek R and Ueki, Toshiyuki and Zhang, Tian and Malvankar, Nikhil S and Shrestha, Pravin M and Flanagan, Kelly A and Aklujkar, Muktak and Butler, Jessica E and Giloteaux, Ludovic and Rotaru, Amelia-Elena and Holmes, Dawn E and Franks, Ashley E and Orellana, Roberto and Risso, Carla and Nevin, Kelly P} } @article {357, title = {Identification of a nucleopolyhedrovirus in winter moth populations from Massachusetts.}, journal = {J Invertebr Pathol}, volume = {108}, year = {2011}, month = {2011 Nov}, pages = {217-9}, abstract = {Winter moth, Operophtera brumata, originally from Europe, has invaded eastern Massachusetts causing widespread defoliation and damage to many deciduous tree species and a variety of crop plants in the infested area. We identified O. brumata nucleopolyhedrovirus (OpbuNPV) in winter moth larvae collected from field sites in Massachusetts by using PCR to amplify a 482 bp region of the baculovirus polyhedrin gene. Viral sequences were also detected in winter moth pupae that failed to emerge, suggesting that these insects may have died as a result of viral infection. This represents the first report of OpbuNPV in winter moth populations in the US.}, keywords = {Animals, Base Sequence, Environmental Monitoring, Larva, Massachusetts, Molecular Sequence Data, Moths, Nucleopolyhedrovirus, Viral Proteins, Virus Diseases}, issn = {1096-0805}, doi = {10.1016/j.jip.2011.08.005}, author = {Burand, John P and Kim, Woojin and Welch, Anna and Elkinton, Joseph S} } @article {686, title = {Identification of a second catalytically active trans-sialidase in Trypanosoma brucei.}, journal = {Biochem Biophys Res Commun}, volume = {415}, year = {2011}, month = {2011 Nov 18}, pages = {421-5}, abstract = {The procyclic stage of Trypanosoma brucei is covered by glycosylphosphatidylinositol (GPI)-anchored surface proteins called procyclins. The procyclin GPI anchor contains a side chain of N-acetyllactosamine repeats terminated by sialic acids. Sialic acid modification is mediated by trans-sialidases expressed on the parasite{\textquoteright}s cell surface. Previous studies suggested the presence of more than one active trans-sialidases, but only one has so far been reported. Here we cloned and examined enzyme activities of four additional trans-sialidase homologs, and show that one of them, Tb927.8.7350, encodes another active trans-sialidase, designated as TbSA C2. In an in vitro assay, TbSA C2 utilized α2-3 sialyllactose as a donor, and produced an α2-3-sialylated product, suggesting that it is an α2-3 trans-sialidase. We suggest that TbSA C2 plays a role in the sialic acid modification of the trypanosome cell surface.}, keywords = {Amino Sugars, Catalysis, Cell Membrane, Cloning, Molecular, Glycoproteins, Glycosylphosphatidylinositols, Mutation, N-Acetylneuraminic Acid, Neuraminidase, Trypanosoma brucei brucei}, issn = {1090-2104}, doi = {10.1016/j.bbrc.2011.10.085}, author = {Nakatani, Fumiki and Morita, Yasu S and Ashida, Hisashi and Nagamune, Kisaburo and Maeda, Yusuke and Kinoshita, Taroh} } @article {430, title = {In situ to in silico and back: elucidating the physiology and ecology of Geobacter spp. using genome-scale modelling.}, journal = {Nat Rev Microbiol}, volume = {9}, year = {2011}, month = {2011 Jan}, pages = {39-50}, abstract = {There is a wide diversity of unexplored metabolism encoded in the genomes of microorganisms that have an important environmental role. Genome-scale metabolic modelling enables the individual reactions that are encoded in annotated genomes to be organized into a coherent whole, which can then be used to predict metabolic fluxes that will optimize cell function under a range of conditions. In this Review, we summarize a series of studies in which genome-scale metabolic modelling of Geobacter spp. has resulted in an in-depth understanding of their central metabolism and ecology. A similar iterative modelling and experimental approach could accelerate elucidation of the physiology and ecology of other microorganisms inhabiting a diversity of environments, and could guide optimization of the practical applications of these species.}, keywords = {Computer Simulation, Environment, Gene Expression Regulation, Bacterial, Genome, Bacterial, Geobacter, Models, Biological}, issn = {1740-1534}, doi = {10.1038/nrmicro2456}, author = {Mahadevan, Radhakrishnan and Palsson, Bernhard {\O} and Lovley, Derek R} } @article {687, title = {Inositol lipid metabolism in mycobacteria: biosynthesis and regulatory mechanisms.}, journal = {Biochim Biophys Acta}, volume = {1810}, year = {2011}, month = {2011 Jun}, pages = {630-41}, abstract = {BACKGROUND: The genus Mycobacterium includes a number of medically important pathogens. The cell walls of these bacteria have many unique features, including the abundance of various inositol lipids, such as phosphatidylinositol mannosides (PIMs), lipomannan (LM), and lipoarabinomannan (LAM). The biosynthesis of these lipids is believed to be prime drug targets, and has been clarified in detail over the past several years. SCOPE OF REVIEW: Here we summarize our current understanding of the inositol lipid metabolism in mycobacteria. We will highlight unsolved issues and future directions especially in the context of metabolic regulation. MAJOR CONCLUSIONS: Inositol is a building block of phosphatidylinositol (PI), which is further elaborated to become PIMs, LM and LAM. d-myo-inositol 3-phosphate is an intermediate of the de novo inositol synthesis, but it is also the starting substrate for mycothiol synthesis. Controlling the level of d-myo-inositol 3-phosphate appears to be important for maintaining the steady state levels of mycothiol and inositol lipids. Several additional control mechanisms must exist to control the complex biosynthetic pathways of PI, PIMs, LM and LAM. These may include regulatory proteins such as a lipoprotein LpqW, and spatial separation of enzymes, such as the amphipathic PimA mannosyltransferase and later enzymes in the PIMs/LM biosynthetic pathway. Finally, we discuss mechanisms that underlie control of LM/LAM glycan polymer elongation. GENERAL SIGNIFICANCE: Mycobacteria have evolved a complex network of inositol metabolism. Clarifying its metabolism will not only provide better understanding of bacterial pathogenesis, but also understanding of the evolution and general functions of inositol lipids in nature.}, keywords = {Inositol, Lipid Metabolism, Lipids, Models, Biological, Mycobacterium, Phosphatidylinositols}, issn = {0006-3002}, doi = {10.1016/j.bbagen.2011.03.017}, author = {Morita, Yasu S and Fukuda, Takeshi and Sena, Chubert B C and Yamaryo-Botte, Yoshiki and McConville, Malcolm J and Kinoshita, Taroh} } @article {372, title = {Metagenomic analysis of a permafrost microbial community reveals a rapid response to thaw.}, journal = {Nature}, volume = {480}, year = {2011}, month = {2011 Dec 15}, pages = {368-71}, abstract = {Permafrost contains an estimated 1672 Pg carbon (C), an amount roughly equivalent to the total currently contained within land plants and the atmosphere. This reservoir of C is vulnerable to decomposition as rising global temperatures cause the permafrost to thaw. During thaw, trapped organic matter may become more accessible for microbial degradation and result in greenhouse gas emissions. Despite recent advances in the use of molecular tools to study permafrost microbial communities, their response to thaw remains unclear. Here we use deep metagenomic sequencing to determine the impact of thaw on microbial phylogenetic and functional genes, and relate these data to measurements of methane emissions. Metagenomics, the direct sequencing of DNA from the environment, allows the examination of whole biochemical pathways and associated processes, as opposed to individual pieces of the metabolic puzzle. Our metagenome analyses reveal that during transition from a frozen to a thawed state there are rapid shifts in many microbial, phylogenetic and functional gene abundances and pathways. After one week of incubation at 5 {\textdegree}C, permafrost metagenomes converge to be more similar to each other than while they are frozen. We find that multiple genes involved in cycling of C and nitrogen shift rapidly during thaw. We also construct the first draft genome from a complex soil metagenome, which corresponds to a novel methanogen. Methane previously accumulated in permafrost is released during thaw and subsequently consumed by methanotrophic bacteria. Together these data point towards the importance of rapid cycling of methane and nitrogen in thawing permafrost.}, keywords = {Alaska, Arctic Regions, Bacteria, Carbon, Carbon Cycle, DNA, Freezing, Genes, rRNA, Metagenome, Metagenomics, Methane, Nitrogen, Nitrogen Cycle, Oxidation-Reduction, Phylogeny, RNA, Ribosomal, 16S, Soil, Soil Microbiology, Temperature, Time Factors}, issn = {1476-4687}, doi = {10.1038/nature10576}, author = {Mackelprang, Rachel and Waldrop, Mark P and Deangelis, Kristen M and David, Maude M and Chavarria, Krystle L and Blazewicz, Steven J and Rubin, Edward M and Jansson, Janet K} } @article {420, title = {Molecular analysis of the metabolic rates of discrete subsurface populations of sulfate reducers.}, journal = {Appl Environ Microbiol}, volume = {77}, year = {2011}, month = {2011 Sep}, pages = {6502-9}, abstract = {Elucidating the in situ metabolic activity of phylogenetically diverse populations of sulfate-reducing microorganisms that populate anoxic sedimentary environments is key to understanding subsurface ecology. Previous pure culture studies have demonstrated that the transcript abundance of dissimilatory (bi)sulfite reductase genes is correlated with the sulfate-reducing activity of individual cells. To evaluate whether expression of these genes was diagnostic for subsurface communities, dissimilatory (bi)sulfite reductase gene transcript abundance in phylogenetically distinct sulfate-reducing populations was quantified during a field experiment in which acetate was added to uranium-contaminated groundwater. Analysis of dsrAB sequences prior to the addition of acetate indicated that Desulfobacteraceae, Desulfobulbaceae, and Syntrophaceae-related sulfate reducers were the most abundant. Quantifying dsrB transcripts of the individual populations suggested that Desulfobacteraceae initially had higher dsrB transcripts per cell than Desulfobulbaceae or Syntrophaceae populations and that the activity of Desulfobacteraceae increased further when the metabolism of dissimilatory metal reducers competing for the added acetate declined. In contrast, dsrB transcript abundance in Desulfobulbaceae and Syntrophaceae remained relatively constant, suggesting a lack of stimulation by added acetate. The indication of higher sulfate-reducing activity in the Desulfobacteraceae was consistent with the finding that Desulfobacteraceae became the predominant component of the sulfate-reducing community. Discontinuing acetate additions resulted in a decline in dsrB transcript abundance in the Desulfobacteraceae. These results suggest that monitoring transcripts of dissimilatory (bi)sulfite reductase genes in distinct populations of sulfate reducers can provide insight into the relative rates of metabolism of different components of the sulfate-reducing community and their ability to respond to environmental perturbations.}, keywords = {Biodiversity, Deltaproteobacteria, DNA, Bacterial, Gene Expression Profiling, Hydrogensulfite Reductase, Molecular Sequence Data, Oxidation-Reduction, Sequence Analysis, DNA, Soil Microbiology, Sulfates, Water Microbiology}, issn = {1098-5336}, doi = {10.1128/AEM.00576-11}, author = {Miletto, M and Williams, K H and N{\textquoteright}Guessan, A L and Lovley, D R} } @article {423, title = {Monitoring the metabolic status of geobacter species in contaminated groundwater by quantifying key metabolic proteins with Geobacter-specific antibodies.}, journal = {Appl Environ Microbiol}, volume = {77}, year = {2011}, month = {2011 Jul}, pages = {4597-602}, abstract = {Simple and inexpensive methods for assessing the metabolic status and bioremediation activities of subsurface microorganisms are required before bioremediation practitioners will adopt molecular diagnosis of the bioremediation community as a routine practice for guiding the development of bioremediation strategies. Quantifying gene transcripts can diagnose important aspects of microbial physiology during bioremediation but is technically challenging and does not account for the impact of translational modifications on protein abundance. An alternative strategy is to directly quantify the abundance of key proteins that might be diagnostic of physiological state. To evaluate this strategy, an antibody-based quantification approach was developed to investigate subsurface Geobacter communities. The abundance of citrate synthase corresponded with rates of metabolism of Geobacter bemidjiensis in chemostat cultures. During in situ bioremediation of uranium-contaminated groundwater the quantity of Geobacter citrate synthase increased with the addition of acetate to the groundwater and decreased when acetate amendments stopped. The abundance of the nitrogen-fixation protein, NifD, increased as ammonium became less available in the groundwater and then declined when ammonium concentrations increased. In a petroleum-contaminated aquifer, the abundance of BamB, an enzyme subunit involved in the anaerobic degradation of mono-aromatic compounds by Geobacter species, increased in zones in which Geobacter were expected to play an important role in aromatic hydrocarbon degradation. These results suggest that antibody-based detection of key metabolic proteins, which should be readily adaptable to standardized kits, may be a feasible method for diagnosing the metabolic state of microbial communities responsible for bioremediation, aiding in the rational design of bioremediation strategies.}, keywords = {Acetates, Antibodies, Bacterial, Bacterial Proteins, Geobacter, Petroleum, Quaternary Ammonium Compounds, Soil Microbiology, Water Microbiology}, issn = {1098-5336}, doi = {10.1128/AEM.00114-11}, author = {Yun, Jiae and Ueki, Toshiyuki and Miletto, Marzia and Lovley, Derek R} } @article {373, title = {PCR amplification-independent methods for detection of microbial communities by the high-density microarray PhyloChip.}, journal = {Appl Environ Microbiol}, volume = {77}, year = {2011}, month = {2011 Sep}, pages = {6313-22}, abstract = {Environmental microbial community analysis typically involves amplification by PCR, despite well-documented biases. We have developed two methods of PCR-independent microbial community analysis using the high-density microarray PhyloChip: direct hybridization of 16S rRNA (dirRNA) or rRNA converted to double-stranded cDNA (dscDNA). We compared dirRNA and dscDNA communities to PCR-amplified DNA communities using a mock community of eight taxa, as well as experiments derived from three environmental sample types: chromium-contaminated aquifer groundwater, tropical forest soil, and secondary sewage in seawater. Community profiles by both direct hybridization methods showed differences that were expected based on accompanying data but that were missing in PCR-amplified communities. Taxon richness decreased in RNA compared to that in DNA communities, suggesting a subset of 20\% in soil and 60\% in groundwater that is active; secondary sewage showed no difference between active and inactive populations. Direct hybridization of dscDNA and RNA is thus a viable alternative to PCR-amplified microbial community analysis, providing identification of the active populations within microbial communities that attenuate pollutants, drive global biogeochemical cycles, or proliferate disease states.}, keywords = {Biodiversity, DNA, Complementary, Environmental Microbiology, Metagenomics, Microarray Analysis, Oligonucleotide Array Sequence Analysis, RNA, Ribosomal, 16S, Sensitivity and Specificity}, issn = {1098-5336}, doi = {10.1128/AEM.05262-11}, author = {Deangelis, Kristen M and Wu, Cindy H and Beller, Harry R and Brodie, Eoin L and Chakraborty, Romy and DeSantis, Todd Z and Fortney, Julian L and Hazen, Terry C and Osman, Shariff R and Singer, Mary E and Tom, Lauren M and Andersen, Gary L} } @article {1568, title = {Pollutant degradation by a Methylocystis strain SB2 grown on ethanol: bioremediation via facultative methanotrophy.}, journal = {FEMS Microbiol Lett}, volume = {318}, year = {2011}, month = {2011 May}, pages = {137-42}, abstract = {A facultative methanotroph, Methylocystis strain SB2, was examined for its ability to degrade chlorinated hydrocarbons when grown on methane or ethanol. Strain SB2 grown on methane degraded vinyl chloride (VC), trans-dichloroethylene (t-DCE), trichloroethylene (TCE), 1,1,1-trichloroethane (1,1,1-TCA), and chloroform (CF), but not dichloromethane (DCM). Growth on methane was reduced in the presence of any chlorinated hydrocarbon. Strain SB2 grown on ethanol degraded VC, t-DCE, and TCE, and 1,1,1-TCA, but not DCM or CF. With the exception of 1,1,1-TCA, the growth of strain SB2 on ethanol was not affected by any individual chlorinated hydrocarbon. No degradation of any chlorinated hydrocarbon was observed when acetylene was added to ethanol-grown cultures, indicating that this degradation was due to particulate methane monooxygenase (pMMO) activity. When mixtures of chlorinated alkanes or alkenes were added to cultures growing on methane or ethanol, chlorinated alkene degradation occurred, but chlorinated alkanes were not, and growth was reduced on both methane and ethanol. Collectively, these data indicate that competitive inhibition of pMMO activity limits methanotrophic growth and pollutant degradation. Facultative methanotrophy may thus be useful to extend the utility of methanotrophs for bioremediation as the use of alternative growth substrates allows for pMMO activity to be focused on pollutant degradation.
}, keywords = {Autotrophic Processes, Biodegradation, Environmental, Environmental Pollutants, Ethanol, Hydrocarbons, Chlorinated, Methane, Methylocystaceae}, issn = {1574-6968}, doi = {10.1111/j.1574-6968.2011.02249.x}, author = {Im, Jeongdae and Semrau, Jeremy D} } @article {1208, title = {Production of hydrogen from α-1,4- and β-1,4-linked saccharides by marine hyperthermophilic Archaea.}, journal = {Appl Environ Microbiol}, volume = {77}, year = {2011}, month = {2011 May}, pages = {3169-73}, abstract = {Nineteen hyperthermophilic heterotrophs from deep-sea hydrothermal vents, plus the control organism Pyrococcus furiosus, were examined for their ability to grow and produce H$_{2}$ on maltose, cellobiose, and peptides and for the presence of the genes encoding proteins that hydrolyze starch and cellulose. All of the strains grew on these disaccharides and peptides and converted maltose and peptides to H$_{2}$ even when elemental sulfur was present as a terminal electron acceptor. Half of the strains had at least one gene for an extracellular starch hydrolase, but only P. furiosus had a gene for an extracellular β-1,4-endoglucanase. P. furiosus was serially adapted for growth on CF11 cellulose and H$_{2}$ production, which is the first reported instance of hyperthermophilic growth on cellulose, with a doubling time of 64 min. Cell-specific H$_{2}$ production rates were 29 fmol, 37 fmol, and 54 fmol of H$_{2}$ produced cell$^{-}${\textonesuperior} doubling$^{-}${\textonesuperior} on α-1,4-linked sugars, β-1,4-linked sugars, and peptides, respectively. The highest total community H$_{2}$ production rate came from growth on starch (2.6 mM H$_{2}$ produced h$^{-}${\textonesuperior}). Hyperthermophilic heterotrophs may serve as an important alternate source of H$_{2}$ for hydrogenotrophic microorganisms in low-H$_{2}$ hydrothermal environments, and some are candidates for H$_{2}$ bioenergy production in bioreactors.
}, keywords = {Archaea, Carbohydrate Metabolism, Hot Springs, Hydro-Lyases, Hydrogen, Peptides, Seawater}, issn = {1098-5336}, doi = {10.1128/AEM.01366-10}, author = {Oslowski, Daniel M and Jung, Jong-Hyun and Seo, Dong-Ho and Park, Cheon-Seok and Holden, James F} } @article {676, title = {Proteopedia: a status report on the collaborative, 3D web-encyclopedia of proteins and other biomolecules.}, journal = {J Struct Biol}, volume = {175}, year = {2011}, month = {2011 Aug}, pages = {244-52}, abstract = {Proteopedia is a collaborative, 3D web-encyclopedia of protein, nucleic acid and other biomolecule structures. Created as a means for communicating biomolecule structures to a diverse scientific audience, Proteopedia (http://www.proteopedia.org) presents structural annotation in an intuitive, interactive format and allows members of the scientific community to easily contribute their own annotations. Here, we provide a status report on Proteopedia by describing advances in the web resource since its inception three and a half years ago, focusing on features of potential direct use to the scientific community. We discuss its progress as a collaborative 3D-encyclopedia of structures as well as its use as a complement to scientific publications and PowerPoint presentations. We also describe Proteopedia{\textquoteright}s use for 3D visualization in structure-related pedagogy.}, keywords = {Encyclopedias as Topic, Information Dissemination, Information Management, Information Services, Models, Molecular, Molecular Biology, Online Systems, Protein Conformation, Proteins, User-Computer Interface}, issn = {1095-8657}, doi = {10.1016/j.jsb.2011.04.011}, author = {Prilusky, Jaime and Hodis, Eran and Canner, David and Decatur, Wayne A and Oberholser, Karl and Martz, Eric and Berchanski, Alexander and Harel, Michal and Sussman, Joel L} } @article {760, title = {Resolving the phylogeny of malaria parasites.}, journal = {Proc Natl Acad Sci U S A}, volume = {108}, year = {2011}, month = {2011 Aug 9}, pages = {12973-4}, keywords = {Animals, Evolution, Molecular, Malaria, Parasites}, issn = {1091-6490}, doi = {10.1073/pnas.1110141108}, author = {Rich, Stephen M and Xu, Guang} } @article {850, title = {Respiratory Chlamydophyla pneumoniae resides primarily in the lower airway.}, journal = {Eur Respir J}, volume = {38}, year = {2011}, month = {2011 Oct}, pages = {994-5; author reply 995}, keywords = {Asthma, Humans, Pulmonary Medicine}, issn = {1399-3003}, doi = {10.1183/09031936.00071711}, author = {Webley, W C and Hahn, D L} } @article {425, title = {A shift in the current: new applications and concepts for microbe-electrode electron exchange.}, journal = {Curr Opin Biotechnol}, volume = {22}, year = {2011}, month = {2011 Jun}, pages = {441-8}, abstract = {Perceived applications of microbe-electrode interactions are shifting from production of electric power to other technologies, some of which even consume current. Electrodes can serve as stable, long-term electron acceptors for contaminant-degrading microbes to promote rapid degradation of organic pollutants in anaerobic subsurface environments. Solar and other forms of renewable electrical energy can be used to provide electrons extracted from water to microorganisms on electrodes at suitably low potentials for a number of groundwater bioremediation applications as well as for the production of fuels and other organic compounds from carbon dioxide. The understanding of how microorganisms exchange electrons with electrodes has improved substantially and is expected to be helpful in optimizing practical applications of microbe-electrode interactions, as well as yielding insights into related natural environmental phenomena.}, keywords = {Bacteria, Biodegradation, Environmental, Biofuels, Carbon Dioxide, Electricity, Electrodes, Electrons, Environmental Pollutants, Fungi, Microbiological Phenomena, Organic Chemicals}, issn = {1879-0429}, doi = {10.1016/j.copbio.2011.01.009}, author = {Lovley, Derek R and Nevin, Kelly P} } @article {381, title = {Silencing of a putative inner arm dynein heavy chain results in flagellar immotility in Trypanosoma brucei.}, journal = {Mol Biochem Parasitol}, volume = {175}, year = {2011}, month = {2011 Jan}, pages = {68-75}, abstract = {The Trypanosoma brucei flagellum controls motility and is crucial for cell polarity and division. Unique features of trypanosome motility suggest that flagellar beat regulation in this organism is unusual and worthy of study. The flagellar axoneme, required for motility, has a structure that is highly conserved among eukaryotes. Of the several dyneins in the axonemal inner arm complex, dynein f is thought to control flagellar waveform shape. A T. brucei gene predicted to encode the dynein f alpha heavy chain, TbDNAH10, was silenced using RNA interference in procyclic T. brucei cells. This resulted in immotile flagella, showing no movement except for occasional slight twitches at the tips. Cell growth slowed dramatically and cells were found in large clusters. Microscopic analysis of silenced cultures showed many cells with detached flagella, sometimes entangled between multiple cells. DAPI staining showed an increased frequency of mis-positioned kinetoplasts and multinucleate cells, suggesting that these cells experience disruption at an early cell cycle stage, probably secondary to the motility defect. TEM images showed apparently normal axonemes and no discernable defects in inner arm structure. This study demonstrates the use of RNAi as an effective method to study very large genes such as dynein heavy chains (HCs), and the immotility phenotype of these dynein knockdowns suggests that an intact inner arm is necessary for flagellar beating in T. brucei. Since analogous mutants in Chlamydomonas reinhardtii retain motility, this phenotype likely reflects differences in requirements for motility and/or dynein assembly between the two organisms and these comparative studies will help elucidate the mechanisms of flagellar beat regulation.}, keywords = {Cell Nucleus, Dyneins, Flagella, Locomotion, Microscopy, Electron, Transmission, Organelles, Protozoan Proteins, RNA Interference, Trypanosoma brucei brucei}, issn = {1872-9428}, doi = {10.1016/j.molbiopara.2010.09.005}, author = {Springer, Amy L and Bruhn, David F and Kinzel, Kathryn W and Rosenthal, No{\"e}l F and Zukas, Randi and Klingbeil, Michele M} } @article {805, title = {Structure of the SSB-DNA polymerase III interface and its role in DNA replication.}, journal = {EMBO J}, volume = {30}, year = {2011}, month = {2011 Oct 19}, pages = {4236-47}, abstract = {Interactions between single-stranded DNA-binding proteins (SSBs) and the DNA replication machinery are found in all organisms, but the roles of these contacts remain poorly defined. In Escherichia coli, SSB{\textquoteright}s association with the χ subunit of the DNA polymerase III holoenzyme has been proposed to confer stability to the replisome and to aid delivery of primers to the lagging-strand DNA polymerase. Here, the SSB-binding site on χ is identified crystallographically and biochemical and cellular studies delineate the consequences of destabilizing the χ/SSB interface. An essential role for the χ/SSB interaction in lagging-strand primer utilization is not supported. However, sequence changes in χ that block complex formation with SSB lead to salt-dependent uncoupling of leading- and lagging-strand DNA synthesis and to a surprising obstruction of the leading-strand DNA polymerase in vitro, pointing to roles for the χ/SSB complex in replisome establishment and maintenance. Destabilization of the χ/SSB complex in vivo produces cells with temperature-dependent cell cycle defects that appear to arise from replisome instability.}, keywords = {Amino Acid Sequence, Bacterial Proteins, Base Sequence, DNA Polymerase III, DNA Replication, DNA, Single-Stranded, DNA-Binding Proteins, Escherichia coli, Escherichia coli Proteins, Holoenzymes, Molecular Sequence Data}, issn = {1460-2075}, doi = {10.1038/emboj.2011.305}, author = {Marceau, Aimee H and Bahng, Soon and Massoni, Shawn C and George, Nicholas P and Sandler, Steven J and Marians, Kenneth J and Keck, James L} } @article {719, title = {Taxonomical developments in the family Polyomaviridae.}, journal = {Arch Virol}, volume = {156}, year = {2011}, month = {2011 Sep}, pages = {1627-34}, abstract = {The Polyomaviridae Study Group of the International Committee on Taxonomy of Viruses (ICTV) has recommended several taxonomical revisions, as follows: The family Polyomaviridae, which is currently constituted as a single genus (Polyomavirus), will be comprised of three genera: two containing mammalian viruses and one containing avian viruses. The two mammalian genera will be designated Orthopolyomavirus and Wukipolyomavirus, and the avian genus will be named Avipolyomavirus. These genera will be created by the redistribution of species from the current single genus (Polyomavirus) and by the inclusion of several new species. In addition, the names of several species will be changed to reflect current usage.}, keywords = {Phylogeny, Polyomaviridae, Terminology as Topic}, issn = {1432-8798}, doi = {10.1007/s00705-011-1008-x}, author = {Johne, Reimar and Buck, Christopher B and Allander, Tobias and Atwood, Walter J and Garcea, Robert L and Imperiale, Michael J and Major, Eugene O and Ramqvist, Torbjorn and Norkin, Leonard C} } @article {380, title = {Three mitochondrial DNA polymerases are essential for kinetoplast DNA replication and survival of bloodstream form Trypanosoma brucei.}, journal = {Eukaryot Cell}, volume = {10}, year = {2011}, month = {2011 Jun}, pages = {734-43}, abstract = {Trypanosoma brucei, the causative agent of human African trypanosomiasis, has a complex life cycle that includes multiple life cycle stages and metabolic changes as the parasite switches between insect vector and mammalian host. The parasite{\textquoteright}s single mitochondrion contains a unique catenated mitochondrial DNA network called kinetoplast DNA (kDNA) that is composed of minicircles and maxicircles. Long-standing uncertainty about the requirement of kDNA in bloodstream form (BF) T. brucei has recently eroded, with reports of posttranscriptional editing and subsequent translation of kDNA-encoded transcripts as essential processes for BF parasites. These studies suggest that kDNA and its faithful replication are indispensable for this life cycle stage. Here we demonstrate that three kDNA replication proteins (mitochondrial DNA polymerases IB, IC, and ID) are required for BF parasite viability. Silencing of each polymerase was lethal, resulting in kDNA loss, persistence of prereplication DNA monomers, and collapse of the mitochondrial membrane potential. These data demonstrate that kDNA replication is indeed crucial for BF T. brucei. The contributions of mitochondrial DNA polymerases IB, IC, and ID to BF parasite viability suggest that these and other kDNA replication proteins warrant further investigation as a new class of targets for the development of antitrypanosomal drugs.}, keywords = {Cell Survival, Cells, Cultured, DNA Replication, DNA, Kinetoplast, DNA-Directed DNA Polymerase, Humans, Membrane Potential, Mitochondrial, Mitochondria, Parasitemia, RNA Interference, Trypanosoma brucei brucei, Trypanosomiasis, African}, issn = {1535-9786}, doi = {10.1128/EC.05008-11}, author = {Bruhn, David F and Sammartino, Mark P and Klingbeil, Michele M} } @article {418, title = {Tunable metallic-like conductivity in microbial nanowire networks.}, journal = {Nat Nanotechnol}, volume = {6}, year = {2011}, month = {2011 Sep}, pages = {573-9}, abstract = {Electronic nanostructures made from natural amino acids are attractive because of their relatively low cost, facile processing and absence of toxicity. However, most materials derived from natural amino acids are electronically insulating. Here, we report metallic-like conductivity in films of the bacterium Geobacter sulfurreducens and also in pilin nanofilaments (known as microbial nanowires) extracted from these bacteria. These materials have electronic conductivities of \~{}5~mS~cm(-1), which are comparable to those of synthetic metallic nanostructures. They can also conduct over distances on the centimetre scale, which is thousands of times the size of a bacterium. Moreover, the conductivity of the biofilm can be tuned by regulating gene expression, and also by varying the gate voltage in a transistor configuration. The conductivity of the nanofilaments has a temperature dependence similar to that of a disordered metal, and the conductivity could be increased by processing.}, keywords = {Electric Conductivity, Geobacter, Nanowires, Transistors, Electronic}, issn = {1748-3395}, doi = {10.1038/nnano.2011.119}, author = {Malvankar, Nikhil S and Vargas, Madeline and Nevin, Kelly P and Franks, Ashley E and Leang, Ching and Kim, Byoung-Chan and Inoue, Kengo and Mester, T{\"u}nde and Covalla, Sean F and Johnson, Jessica P and Rotello, Vincent M and Tuominen, Mark T and Lovley, Derek R} } @article {446, title = {Alignment of the c-type cytochrome OmcS along pili of Geobacter sulfurreducens.}, journal = {Appl Environ Microbiol}, volume = {76}, year = {2010}, month = {2010 Jun}, pages = {4080-4}, abstract = {Immunogold localization revealed that OmcS, a cytochrome that is required for Fe(III) oxide reduction by Geobacter sulfurreducens, was localized along the pili. The apparent spacing between OmcS molecules suggests that OmcS facilitates electron transfer from pili to Fe(III) oxides rather than promoting electron conduction along the length of the pili.}, keywords = {Cytochromes c, Ferric Compounds, Fimbriae, Bacterial, Geobacter, Immunohistochemistry, Microscopy, Immunoelectron}, issn = {1098-5336}, doi = {10.1128/AEM.00023-10}, author = {Leang, Ching and Qian, Xinlei and Mester, T{\"u}nde and Lovley, Derek R} } @article {433, title = {Analysis of biostimulated microbial communities from two field experiments reveals temporal and spatial differences in proteome profiles.}, journal = {Environ Sci Technol}, volume = {44}, year = {2010}, month = {2010 Dec 1}, pages = {8897-903}, abstract = {Stimulated by an acetate-amendment field experiment conducted in 2007, anaerobic microbial populations in the aquifer at the Rifle Integrated Field Research Challenge site in Colorado reduced mobile U(VI) to insoluble U(IV). During this experiment, planktonic biomass was sampled at various time points to quantitatively evaluate proteomes. In 2008, an acetate-amended field experiment was again conducted in a similar manner to the 2007 experiment. As there was no comprehensive metagenome sequence available for use in proteomics analysis, we systematically evaluated 12 different organism genome sequences to generate sets of aggregate genomes, or "pseudo-metagenomes", for supplying relative quantitative peptide and protein identifications. Proteomics results support previous observations of the dominance of Geobacteraceae during biostimulation using acetate as sole electron donor, and revealed a shift from an early stage of iron reduction to a late stage of iron reduction. Additionally, a shift from iron reduction to sulfate reduction was indicated by changes in the contribution of proteome information contributed by different organism genome sequences within the aggregate set. In addition, the comparison of proteome measurements made between the 2007 field experiment and 2008 field experiment revealed differences in proteome profiles. These differences may be the result of alterations in abundance and population structure within the planktonic biomass samples collected for analysis.}, keywords = {Bacteria, Biodiversity, Biomass, Fresh Water, Plankton, Proteome, Water Microbiology}, issn = {1520-5851}, doi = {10.1021/es101029f}, author = {Callister, Stephen J and Wilkins, Michael J and Nicora, Carrie D and Williams, Kenneth H and Banfield, Jillian F and VerBerkmoes, Nathan C and Hettich, Robert L and N{\textquoteright}Guessan, Lucie and Mouser, Paula J and Elifantz, Hila and Smith, Richard D and Lovley, Derek R and Lipton, Mary S and Long, Philip E} } @article {731, title = {Biotechnological potential of aquatic plant-microbe interactions.}, journal = {Curr Opin Biotechnol}, volume = {21}, year = {2010}, month = {2010 Jun}, pages = {339-45}, abstract = {The rhizosphere in terrestrial systems is the region of soil surrounding plant roots where there is increased microbial activity; in aquatic plants, this definition may be less clear because of diffusion of nutrients in water, but there is still a zone of influence by plant roots in this environment [1]. Within that zone chemical conditions differ from those of the surrounding environment as a consequence of a range of processes that were induced either directly by the activity of plant roots or by the activity of rhizosphere microflora. Recently, there are a number of new studies related to rhizospheres of aquatic plants and specifically their increased potential for remediation of contaminants, especially remediation of metals through aquatic plant-microbial interaction.}, keywords = {Biodegradation, Environmental, Biotechnology, Hydrogen-Ion Concentration, Plant Roots, Plants}, issn = {1879-0429}, doi = {10.1016/j.copbio.2010.04.004}, author = {Stout, L and N{\"u}sslein, K} } @article {807, title = {Clinical applications of biomarkers in pediatric traumatic brain injury.}, journal = {Childs Nerv Syst}, volume = {26}, year = {2010}, month = {2010 Feb}, pages = {205-13}, abstract = {INTRODUCTION: The diagnosis, treatment, and prediction of outcome in pediatric traumatic brain injury (TBI) present significant challenges to the treating clinician. Clinical and radiological tools for assessing injury severity and predicting outcome, in particular, lack sensitivity and specificity. In patients with mild TBI, often there is uncertainty about which patients should undergo radiological imaging and who is at risk for long term neurological sequelae. In severe TBI, often there is uncertainty about which patients will experience secondary insults and what the outcome for individual patients will be. In several other clinical specialties, biomarkers are used to diagnose disease, direct treatment, and prognosticate. However, an ideal biomarker for brain injury has not been found. METHODS: In this review, we examine the various factors that must be taken into account in the search for a reliable biomarker in brain injury. We review the important studies that have investigated common biomarkers of structural brain injury, in particular S100B, neuron-specific enolase, myelin basic protein, and glial fibrillary acid protein. DISCUSSION: The potential uses and limitations of these biomarkers in the context of TBI are discussed.}, keywords = {Animals, Biological Markers, Brain Injuries, Child, Humans}, issn = {1433-0350}, doi = {10.1007/s00381-009-1009-1}, author = {Sandler, Simon J I and Figaji, Anthony A and Adelson, P David} } @article {428, title = {Constraint-based modeling analysis of the metabolism of two Pelobacter species.}, journal = {BMC Syst Biol}, volume = {4}, year = {2010}, month = {2010}, pages = {174}, abstract = {BACKGROUND: Pelobacter species are commonly found in a number of subsurface environments, and are unique members of the Geobacteraceae family. They are phylogenetically intertwined with both Geobacter and Desulfuromonas species. Pelobacter species likely play important roles in the fermentative degradation of unusual organic matters and syntrophic metabolism in the natural environments, and are of interest for applications in bioremediation and microbial fuel cells. RESULTS: In order to better understand the physiology of Pelobacter species, genome-scale metabolic models for Pelobacter carbinolicus and Pelobacter propionicus were developed. Model development was greatly aided by the availability of models of the closely related Geobacter sulfurreducens and G. metallireducens. The reconstructed P. carbinolicus model contains 741 genes and 708 reactions, whereas the reconstructed P. propionicus model contains 661 genes and 650 reactions. A total of 470 reactions are shared among the two Pelobacter models and the two Geobacter models. The different reactions between the Pelobacter and Geobacter models reflect some unique metabolic capabilities such as fermentative growth for both Pelobacter species. The reconstructed Pelobacter models were validated by simulating published growth conditions including fermentations, hydrogen production in syntrophic co-culture conditions, hydrogen utilization, and Fe(III) reduction. Simulation results matched well with experimental data and indicated the accuracy of the models. CONCLUSIONS: We have developed genome-scale metabolic models of P. carbinolicus and P. propionicus. These models of Pelobacter metabolism can now be incorporated into the growing repertoire of genome scale models of the Geobacteraceae family to aid in describing the growth and activity of these organisms in anoxic environments and in the study of their roles and interactions in the subsurface microbial community.}, keywords = {Anaerobiosis, Citric Acid Cycle, Desulfuromonas, Electron Transport, Energy Metabolism, Gene Expression Regulation, Bacterial, Models, Biological, Reproducibility of Results, Sulfur}, issn = {1752-0509}, doi = {10.1186/1752-0509-4-174}, author = {Sun, Jun and Haveman, Shelley A and Bui, Olivia and Fahland, Tom R and Lovley, Derek R} } @article {677, title = {ConSurf 2010: calculating evolutionary conservation in sequence and structure of proteins and nucleic acids.}, journal = {Nucleic Acids Res}, volume = {38}, year = {2010}, month = {2010 Jul}, pages = {W529-33}, abstract = {It is informative to detect highly conserved positions in proteins and nucleic acid sequence/structure since they are often indicative of structural and/or functional importance. ConSurf (http://consurf.tau.ac.il) and ConSeq (http://conseq.tau.ac.il) are two well-established web servers for calculating the evolutionary conservation of amino acid positions in proteins using an empirical Bayesian inference, starting from protein structure and sequence, respectively. Here, we present the new version of the ConSurf web server that combines the two independent servers, providing an easier and more intuitive step-by-step interface, while offering the user more flexibility during the process. In addition, the new version of ConSurf calculates the evolutionary rates for nucleic acid sequences. The new version is freely available at: http://consurf.tau.ac.il/.}, keywords = {Amino Acid Sequence, Base Sequence, Conserved Sequence, Evolution, Molecular, Internet, Nucleic Acid Conformation, Protein Conformation, Sequence Analysis, Sequence Homology, Amino Acid, Software}, issn = {1362-4962}, doi = {10.1093/nar/gkq399}, author = {Ashkenazy, Haim and Erez, Elana and Martz, Eric and Pupko, Tal and Ben-Tal, Nir} } @article {689, title = {Controlled expression of branch-forming mannosyltransferase is critical for mycobacterial lipoarabinomannan biosynthesis.}, journal = {J Biol Chem}, volume = {285}, year = {2010}, month = {2010 Apr 30}, pages = {13326-36}, abstract = {Lipomannan (LM) and lipoarabinomannan (LAM) are phosphatidylinositol-anchored glycans present in the mycobacterial cell wall. In Mycobacterium smegmatis, the mannan core of LM/LAM constitutes a linear chain of 20-25 alpha1,6-mannoses elaborated by 8-9 alpha1,2-monomannose side branches. At least two alpha1,6-mannosyltransferases mediate the linear mannose chain elongation, and one branching alpha1,2-mannosyltransferase (encoded by MSMEG_4247) transfers monomannose branches. An MSMEG_4247 deletion mutant accumulates branchless LAM and interestingly fails to accumulate LM, suggesting an unexpected role of mannose branching for LM synthesis or maintenance. To understand the roles of MSMEG_4247-mediated branching more clearly, we analyzed the MSMEG_4247 deletion mutant in detail. Our study showed that the deletion mutant restored the synthesis of wild-type LM and LAM upon the expression of MSMEG_4247 at wild-type levels. In striking contrast, overexpression of MSMEG_4247 resulted in the accumulation of dwarfed LM/LAM, although monomannose branching was restored. The dwarfed LAM carried a mannan chain less than half the length of wild-type LAM and was elaborated by an arabinan that was about 4 times smaller. Induced overexpression of an elongating alpha1,6-mannosyltransferase competed with the overexpressed branching enzyme, alleviating the dwarfing effect of the branching enzyme. In wild-type cells, LM and LAM decreased in quantity in the stationary phase, and the expression levels of branching and elongating mannosyltransferases were reduced in concert, presumably to avoid producing abnormal LM/LAM. These data suggest that the coordinated expressions of branching and elongating mannosyltransferases are critical for mannan backbone elongation.}, keywords = {Bacterial Proteins, Cell Wall, Gene Deletion, Lipopolysaccharides, Mannose, Mannosyltransferases, Mycobacterium smegmatis}, issn = {1083-351X}, doi = {10.1074/jbc.M109.077297}, author = {Sena, Chubert B C and Fukuda, Takeshi and Miyanagi, Kana and Matsumoto, Sohkichi and Kobayashi, Kazuo and Murakami, Yoshiko and Maeda, Yusuke and Kinoshita, Taroh and Morita, Yasu S} } @article {443, title = {De Novo assembly of the complete genome of an enhanced electricity-producing variant of Geobacter sulfurreducens using only short reads.}, journal = {PLoS One}, volume = {5}, year = {2010}, month = {2010}, pages = {e10922}, abstract = {State-of-the-art DNA sequencing technologies are transforming the life sciences due to their ability to generate nucleotide sequence information with a speed and quantity that is unapproachable with traditional Sanger sequencing. Genome sequencing is a principal application of this technology, where the ultimate goal is the full and complete sequence of the organism of interest. Due to the nature of the raw data produced by these technologies, a full genomic sequence attained without the aid of Sanger sequencing has yet to be demonstrated.We have successfully developed a four-phase strategy for using only next-generation sequencing technologies (Illumina and 454) to assemble a complete microbial genome de novo. We applied this approach to completely assemble the 3.7 Mb genome of a rare Geobacter variant (KN400) that is capable of unprecedented current production at an electrode. Two key components of our strategy enabled us to achieve this result. First, we integrated the two data types early in the process to maximally leverage their complementary characteristics. And second, we used the output of different short read assembly programs in such a way so as to leverage the complementary nature of their different underlying algorithms or of their different implementations of the same underlying algorithm.The significance of our result is that it demonstrates a general approach for maximizing the efficiency and success of genome assembly projects as new sequencing technologies and new assembly algorithms are introduced. The general approach is a meta strategy, wherein sequencing data are integrated as early as possible and in particular ways and wherein multiple assembly algorithms are judiciously applied such that the deficiencies in one are complemented by another.}, keywords = {Algorithms, Electricity, Genome, Bacterial, Geobacter, Polymerase Chain Reaction}, issn = {1932-6203}, doi = {10.1371/journal.pone.0010922}, author = {Nagarajan, Harish and Butler, Jessica E and Klimes, Anna and Qiu, Yu and Zengler, Karsten and Ward, Joy and Young, Nelson D and Meth{\'e}, Barbara A and Palsson, Bernhard {\O} and Lovley, Derek R and Barrett, Christian L} } @article {31, title = {Derepression of INO1 transcription requires cooperation between the Ino2p-Ino4p heterodimer and Cbf1p and recruitment of the ISW2 chromatin-remodeling complex.}, journal = {Eukaryot Cell}, volume = {9}, year = {2010}, month = {2010 Dec}, pages = {1845-55}, abstract = {The Saccharomyces cerevisiae INO1 gene encodes the structural enzyme inositol-3-phosphate synthase for the synthesis de novo of inositol and inositol-containing phospholipids. The transcription of INO1 is completely derepressed in the absence of inositol and choline (I(-) C(-)). Derepression requires the binding of the Ino2p-Ino4p basic helix-loop-helix (bHLH) heterodimer to the UAS(INO) promoter element. We report here the requirement of a third bHLH protein, centromere-binding factor 1 (Cbf1p), for the complete derepression of INO1 transcription. We found that Cbf1p regulates INO1 transcription by binding to sites distal to the INO1 promoter and encompassing the upstream SNA3 open reading frame (ORF) and promoter. The binding of Cbf1p requires Ino2p-Ino4p binding to the UAS(INO) sites in the INO1 promoter and vice versa, suggesting a cooperative mechanism. Furthermore, Cbf1p binding to the upstream sites was required for the binding of the ISW2 chromatin-remodeling complex to the Ino2p-Ino4p-binding sites on the INO1 promoter. Consistent with this, ISW2 was also required for the complete derepression of INO1 transcription.
}, keywords = {Adenosine Triphosphatases, Basic Helix-Loop-Helix Leucine Zipper Transcription Factors, Basic Helix-Loop-Helix Transcription Factors, Chromatin, Chromatin Assembly and Disassembly, Dimerization, Gene Expression Regulation, Fungal, Myo-Inositol-1-Phosphate Synthase, Promoter Regions, Genetic, Protein Binding, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins, Transcription Factors, Transcription, Genetic}, issn = {1535-9786}, doi = {10.1128/EC.00144-10}, author = {Shetty, Ameet and Lopes, John M} } @article {431, title = {Direct exchange of electrons within aggregates of an evolved syntrophic coculture of anaerobic bacteria.}, journal = {Science}, volume = {330}, year = {2010}, month = {2010 Dec 3}, pages = {1413-5}, abstract = {Microbial consortia that cooperatively exchange electrons play a key role in the anaerobic processing of organic matter. Interspecies hydrogen transfer is a well-documented strategy for electron exchange in dispersed laboratory cultures, but cooperative partners in natural environments often form multispecies aggregates. We found that laboratory evolution of a coculture of Geobacter metallireducens and Geobacter sulfurreducens metabolizing ethanol favored the formation of aggregates that were electrically conductive. Sequencing aggregate DNA revealed selection for a mutation that enhances the production of a c-type cytochrome involved in extracellular electron transfer and accelerates the formation of aggregates. Aggregate formation was also much faster in mutants that were deficient in interspecies hydrogen transfer, further suggesting direct interspecies electron transfer.}, keywords = {Anaerobiosis, Bacterial Proteins, Biological Evolution, Culture Media, Cytochrome c Group, Electron Transport, Electrons, Ethanol, Fimbriae Proteins, Geobacter, Hydrogen, Microbial Consortia, Microbial Interactions, Mutation, Oxidation-Reduction, Selection, Genetic}, issn = {1095-9203}, doi = {10.1126/science.1196526}, author = {Summers, Zarath M and Fogarty, Heather E and Leang, Ching and Franks, Ashley E and Malvankar, Nikhil S and Lovley, Derek R} } @article {453, title = {Electrode-based approach for monitoring in situ microbial activity during subsurface bioremediation.}, journal = {Environ Sci Technol}, volume = {44}, year = {2010}, month = {2010 Jan 1}, pages = {47-54}, abstract = {Current production by microorganisms colonizing subsurface electrodes and its relationship to substrate availability and microbial activity was evaluated in an aquifer undergoing bioremediation. Borehole graphite anodes were installed downgradient from a region of acetate injection designed to stimulate bioreduction of U(VI); cathodes consisted of graphite electrodes embedded at the ground surface. Significant increases in current density (< or =50 mA/m2) tracked delivery of acetate to the electrodes, dropping rapidly when acetate inputs were discontinued. An upgradient control electrode not exposed to acetate produced low, steady currents (< or =0.2 mA/m2). Elevated current was strongly correlated with uranium removal but minimal correlation existed with elevated Fe(II). Confocal laser scanning microscopy of electrodes revealed firmly attached biofilms, and analysis of 16S rRNA gene sequences indicated the electrode surfaces were dominated (67-80\%) by Geobacter species. This is the first demonstration that electrodes can produce readily detectable currents despite long-range (6 m) separation of anode and cathode, and these results suggest that oxidation of acetate coupled to electron transfer to electrodes by Geobacter species was the primary source of current. Thus it is expected that current production may serve as an effective proxy for monitoring in situ microbial activity in a variety of subsurface anoxic environments.}, keywords = {Electrodes, Environmental Monitoring, Environmental Remediation, Geobacter, RNA, Ribosomal, 16S, Water Pollutants, Chemical}, issn = {0013-936X}, doi = {10.1021/es9017464}, author = {Williams, Kenneth H and Nevin, Kelly P and Franks, Ashley and Englert, Andreas and Long, Philip E and Lovley, Derek R} } @article {449, title = {Evolution of electron transfer out of the cell: comparative genomics of six Geobacter genomes.}, journal = {BMC Genomics}, volume = {11}, year = {2010}, month = {2010}, pages = {40}, abstract = {BACKGROUND: Geobacter species grow by transferring electrons out of the cell--either to Fe(III)-oxides or to man-made substances like energy-harvesting electrodes. Study of Geobacter sulfurreducens has shown that TCA cycle enzymes, inner-membrane respiratory enzymes, and periplasmic and outer-membrane cytochromes are required. Here we present comparative analysis of six Geobacter genomes, including species from the clade that predominates in the subsurface. Conservation of proteins across the genomes was determined to better understand the evolution of Geobacter species and to create a metabolic model applicable to subsurface environments. RESULTS: The results showed that enzymes for acetate transport and oxidation, and for proton transport across the inner membrane were well conserved. An NADH dehydrogenase, the ATP synthase, and several TCA cycle enzymes were among the best conserved in the genomes. However, most of the cytochromes required for Fe(III)-reduction were not, including many of the outer-membrane cytochromes. While conservation of cytochromes was poor, an abundance and diversity of cytochromes were found in every genome, with duplications apparent in several species. CONCLUSIONS: These results indicate there is a common pathway for acetate oxidation and energy generation across the family and in the last common ancestor. They also suggest that while cytochromes are important for extracellular electron transport, the path of electrons across the periplasm and outer membrane is variable. This combination of abundant cytochromes with weak sequence conservation suggests they may not be specific terminal reductases, but rather may be important in their heme-bearing capacity, as sinks for electrons between the inner-membrane electron transport chain and the extracellular acceptor.}, keywords = {Acetates, Bacterial Proton-Translocating ATPases, Citric Acid Cycle, Cluster Analysis, Comparative Genomic Hybridization, Cytochromes, Electron Transport, Evolution, Molecular, Gene Duplication, Gene Expression Regulation, Bacterial, Gene Transfer, Horizontal, Genome, Bacterial, Genomics, Geobacter, NADH Dehydrogenase, Oxidation-Reduction, Phylogeny}, issn = {1471-2164}, doi = {10.1186/1471-2164-11-40}, author = {Butler, Jessica E and Young, Nelson D and Lovley, Derek R} } @article {444, title = {Expression of acetate permease-like (apl ) genes in subsurface communities of Geobacter species under fluctuating acetate concentrations.}, journal = {FEMS Microbiol Ecol}, volume = {73}, year = {2010}, month = {2010 Sep}, pages = {441-9}, abstract = {The addition of acetate to uranium-contaminated aquifers in order to stimulate the growth and activity of Geobacter species that reduce uranium is a promising in situ bioremediation option. Optimizing this bioremediation strategy requires that sufficient acetate be added to promote Geobacter species growth. We hypothesized that under acetate-limiting conditions, subsurface Geobacter species would increase the expression of either putative acetate symporters genes (aplI and aplII). Acetate was added to a uranium-contaminated aquifer (Rifle, CO) in two continuous amendments separated by 5 days of groundwater flush to create changing acetate concentrations. While the expression of aplI in monitoring well D04 (high acetate) weakly correlated with the acetate concentration over time, the transcript levels for this gene were relatively constant in well D08 (low acetate). At the lowest acetate concentrations during the groundwater flush, the transcript levels of aplII were the highest. The expression of aplII decreased 2-10-fold upon acetate reintroduction. However, the overall instability of acetate concentrations throughout the experiment could not support a robust conclusion regarding the role of apl genes in response to acetate limitation under field conditions, in contrast to previous chemostat studies, suggesting that the function of a microbial community cannot be inferred based on lab experiments alone.}, keywords = {Acetates, Bacterial Proteins, Biodegradation, Environmental, Fresh Water, Gene Expression Regulation, Bacterial, Gene Library, Geobacter, Membrane Transport Proteins, Multigene Family, RNA, Bacterial, Uranium, Water Pollutants, Radioactive}, issn = {1574-6941}, doi = {10.1111/j.1574-6941.2010.00907.x}, author = {Elifantz, Hila and N{\textquoteright}guessan, Lucie A and Mouser, Paula J and Williams, Kenneth H and Wilkins, Michael J and Risso, Carla and Holmes, Dawn E and Long, Philip E and Lovley, Derek R} } @article {434, title = {The genome of Geobacter bemidjiensis, exemplar for the subsurface clade of Geobacter species that predominate in Fe(III)-reducing subsurface environments.}, journal = {BMC Genomics}, volume = {11}, year = {2010}, month = {2010}, pages = {490}, abstract = {BACKGROUND: Geobacter species in a phylogenetic cluster known as subsurface clade 1 are often the predominant microorganisms in subsurface environments in which Fe(III) reduction is the primary electron-accepting process. Geobacter bemidjiensis, a member of this clade, was isolated from hydrocarbon-contaminated subsurface sediments in Bemidji, Minnesota, and is closely related to Geobacter species found to be abundant at other subsurface sites. This study examines whether there are significant differences in the metabolism and physiology of G. bemidjiensis compared to non-subsurface Geobacter species. RESULTS: Annotation of the genome sequence of G. bemidjiensis indicates several differences in metabolism compared to previously sequenced non-subsurface Geobacteraceae, which will be useful for in silico metabolic modeling of subsurface bioremediation processes involving Geobacter species. Pathways can now be predicted for the use of various carbon sources such as propionate by G. bemidjiensis. Additional metabolic capabilities such as carbon dioxide fixation and growth on glucose were predicted from the genome annotation. The presence of different dicarboxylic acid transporters and two oxaloacetate decarboxylases in G. bemidjiensis may explain its ability to grow by disproportionation of fumarate. Although benzoate is the only aromatic compound that G. bemidjiensis is known or predicted to utilize as an electron donor and carbon source, the genome suggests that this species may be able to detoxify other aromatic pollutants without degrading them. Furthermore, G. bemidjiensis is auxotrophic for 4-aminobenzoate, which makes it the first Geobacter species identified as having a vitamin requirement. Several features of the genome indicated that G. bemidjiensis has enhanced abilities to respire, detoxify and avoid oxygen. CONCLUSION: Overall, the genome sequence of G. bemidjiensis offers surprising insights into the metabolism and physiology of Geobacteraceae in subsurface environments, compared to non-subsurface Geobacter species, such as the ability to disproportionate fumarate, more efficient oxidation of propionate, enhanced responses to oxygen stress, and dependence on the environment for a vitamin requirement. Therefore, an understanding of the activity of Geobacter species in the subsurface is more likely to benefit from studies of subsurface isolates such as G. bemidjiensis than from the non-subsurface model species studied so far.}, keywords = {Aldehyde Oxidoreductases, Biodegradation, Environmental, Carbohydrate Metabolism, Carbon Dioxide, Cell Wall, Electrons, Environmental Microbiology, Fatty Acids, Frameshift Mutation, Fumarates, Genes, Bacterial, Genome, Bacterial, Geobacter, Glucose, Iron, Metabolic Networks and Pathways, Multienzyme Complexes, Multigene Family, Osmosis, Oxidation-Reduction, Oxo-Acid-Lyases, Propionic Acids, Pyruvic Acid, Species Specificity, Surface Properties}, issn = {1471-2164}, doi = {10.1186/1471-2164-11-490}, author = {Aklujkar, Muktak and Young, Nelson D and Holmes, Dawn and Chavan, Milind and Risso, Carla and Kiss, Hajnalka E and Han, Cliff S and Land, Miriam L and Lovley, Derek R} } @article {452, title = {Genome-wide gene regulation of biosynthesis and energy generation by a novel transcriptional repressor in Geobacter species.}, journal = {Nucleic Acids Res}, volume = {38}, year = {2010}, month = {2010 Jan}, pages = {810-21}, abstract = {Geobacter species play important roles in bioremediation of contaminated environments and in electricity production from waste organic matter in microbial fuel cells. To better understand physiology of Geobacter species, expression and function of citrate synthase, a key enzyme in the TCA cycle that is important for organic acid oxidation in Geobacter species, was investigated. Geobacter sulfurreducens did not require citrate synthase for growth with hydrogen as the electron donor and fumarate as the electron acceptor. Expression of the citrate synthase gene, gltA, was repressed by a transcription factor under this growth condition. Functional and comparative genomics approaches, coupled with genetic and biochemical assays, identified a novel transcription factor termed HgtR that acts as a repressor for gltA. Further analysis revealed that HgtR is a global regulator for genes involved in biosynthesis and energy generation in Geobacter species. The hgtR gene was essential for growth with hydrogen, during which hgtR expression was induced. These findings provide important new insights into the mechanisms by which Geobacter species regulate their central metabolism under different environmental conditions.}, keywords = {Bacterial Proteins, Base Sequence, Citrate (si)-Synthase, Gene Expression Regulation, Bacterial, Genome, Bacterial, Geobacter, Molecular Sequence Data, Promoter Regions, Genetic, Repressor Proteins, Transcription, Genetic}, issn = {1362-4962}, doi = {10.1093/nar/gkp1085}, author = {Ueki, Toshiyuki and Lovley, Derek R} } @article {436, title = {Genome-wide survey for PilR recognition sites of the metal-reducing prokaryote Geobacter sulfurreducens.}, journal = {Gene}, volume = {469}, year = {2010}, month = {2010 Dec 1}, pages = {31-44}, abstract = {Geobacter sulfurreducens is a species from the bacterial family Geobacteraceae, members of which participate in bioenergy production and in environmental bioremediation. G. sulfurreducens pili are electrically conductive and are required for Fe(III) oxide reduction and for optimal current production in microbial fuel cells. PilR is an enhancer binding protein, which is an activator acting together with the alternative sigma factor, RpoN, in transcriptional regulation. Both RpoN and PilR are involved in regulation of expression of the pilA gene, whose product is pilin, a structural component of a pilus. Using bioinformatic approaches, we predicted G. sulfurreducens sequence elements that are likely to be regulated by PilR. The functional importance of the genome region containing a PilR binding site predicted upstream of the pilA gene was experimentally validated. The predicted G. sulfurreducens PilR binding sites are similar to PilR binding sites of Pseudomonas and Moraxella. While the number of predicted PilR-regulated sites did not deviate from that expected by chance, multiple sites were predicted upstream of genes with roles in biosynthesis and function of pili and flagella, in secretory pathways, and in cell wall biogenesis, suggesting the possible involvement of G. sulfurreducens PilR in regulation of production and assembly of pili and flagella.}, keywords = {Bacterial Proteins, Base Sequence, Binding Sites, Conserved Sequence, Ferric Compounds, Fimbriae Proteins, Gene Expression Regulation, Bacterial, Genome, Bacterial, Geobacter, Molecular Sequence Data, Promoter Regions, Genetic, Transcription Factors, Transcription, Genetic}, issn = {1879-0038}, doi = {10.1016/j.gene.2010.08.005}, author = {Krushkal, Julia and Ju{\'a}rez, Katy and Barbe, Jose F and Qu, Yanhua and Andrade, Angel and Puljic, Marko and Adkins, Ronald M and Lovley, Derek R and Ueki, Toshiyuki} } @article {852, title = {Infectious Chlamydia pneumoniae is associated with elevated interleukin-8 and airway neutrophilia in children with refractory asthma.}, journal = {Pediatr Infect Dis J}, volume = {29}, year = {2010}, month = {2010 Dec}, pages = {1093-8}, abstract = {BACKGROUND: Neutrophilic asthma is thought to be less responsive than eosinophilic asthma to anti-inflammatory therapies including corticosteroids. Chlamydia pneumoniae has been implicated in asthma, possibly by induction of interleukin (IL-8). We hypothesized that IL-8 is increased in the bronchoalveolar lavage (BAL) fluid from children with asthma and C. pneumoniae. METHODS: BAL fluid was analyzed for C. pneumoniae and IL-8 using polymerase chain reaction (PCR) and enzyme-linked immunosorbent assay from 2 asthma patient populations in the Bronx, NY and Massachusetts with an average age of 8 and 8.7 years old, respectively. For comparison, samples were also analyzed for C. trachomatis and Mycoplasma 16s DNA. RESULTS: Of 18 Bronx samples analyzed, 6 (33\%) were PCR-positive for C. pneumoniae, 10 (56\%) for C. trachomatis, and 8 (44\%) for Mycoplasma 16s DNA. IL-8 from C. pneumoniae-positive samples was 3.3-fold higher compared with negative samples (P = 0.003). There was no difference between patients tested for C. trachomatis or Mycoplasma. Of 84 Massachusetts samples analyzed, 42 (50\%) were PCR-positive for C. pneumoniae, 42 (50\%) for C. trachomatis, and 13 (16\%) for Mycoplasma. IL-8 concentration from C. pneumoniae-positive samples was 10.49-fold higher compared with negative samples (P = 0.0001). As in the Bronx cohort, there were no differences between patients tested for C. trachomatis or Mycoplasma. Lastly, BAL neutrophilia predicted the presence of C. pneumoniae but not Mycoplasma or C. trachomatis. CONCLUSIONS: Children with asthma who were PCR-positive for C. pneumoniae demonstrated elevated concentrations of IL-8 and neutrophils in BAL fluid compared with similar patients who were positive for C. trachomatis or Mycoplasma organisms, but PCR-negative for C. pneumoniae. Undiagnosed C. pneumoniae infection in children may therefore contribute to poorly controlled asthma via induction of IL-8.}, keywords = {Asthma, Bronchoalveolar Lavage Fluid, Child, Chlamydia trachomatis, Chlamydophila Infections, Chlamydophila pneumoniae, DNA, Bacterial, DNA, Ribosomal, Enzyme-Linked Immunosorbent Assay, Female, Humans, Interleukin-8, Male, Massachusetts, Mycoplasma, Neutrophils, New York City, Polymerase Chain Reaction, Respiratory System, RNA, Ribosomal, 16S}, issn = {1532-0987}, author = {Patel, Katir K and Vicencio, Alfin G and Du, Zhongfang and Tsirilakis, Kalliope and Salva, Paul S and Webley, Wilmore C} } @article {439, title = {Interference with histidyl-tRNA synthetase by a CRISPR spacer sequence as a factor in the evolution of Pelobacter carbinolicus.}, journal = {BMC Evol Biol}, volume = {10}, year = {2010}, month = {2010}, pages = {230}, abstract = {BACKGROUND: Pelobacter carbinolicus, a bacterium of the family Geobacteraceae, cannot reduce Fe(III) directly or produce electricity like its relatives. How P. carbinolicus evolved is an intriguing problem. The genome of P. carbinolicus contains clustered regularly interspaced short palindromic repeats (CRISPR) separated by unique spacer sequences, which recent studies have shown to produce RNA molecules that interfere with genes containing identical sequences. RESULTS: CRISPR spacer $\#$1, which matches a sequence within hisS, the histidyl-tRNA synthetase gene of P. carbinolicus, was shown to be expressed. Phylogenetic analysis and genetics demonstrated that a gene paralogous to hisS in the genomes of Geobacteraceae is unlikely to compensate for interference with hisS. Spacer $\#$1 inhibited growth of a transgenic strain of Geobacter sulfurreducens in which the native hisS was replaced with that of P. carbinolicus. The prediction that interference with hisS would result in an attenuated histidyl-tRNA pool insufficient for translation of proteins with multiple closely spaced histidines, predisposing them to mutation and elimination during evolution, was investigated by comparative genomics of P. carbinolicus and related species. Several ancestral genes with high histidine demand have been lost or modified in the P. carbinolicus lineage, providing an explanation for its physiological differences from other Geobacteraceae. CONCLUSIONS: The disappearance of multiheme c-type cytochromes and other genes typical of a metal-respiring ancestor from the P. carbinolicus lineage may be the consequence of spacer $\#$1 interfering with hisS, a condition that can be reproduced in a heterologous host. This is the first successful co-introduction of an active CRISPR spacer and its target in the same cell, the first application of a chimeric CRISPR construct consisting of a spacer from one species in the context of repeats of another species, and the first report of a potential impact of CRISPR on genome-scale evolution by interference with an essential gene.}, keywords = {Base Sequence, Comparative Genomic Hybridization, Computational Biology, Deltaproteobacteria, DNA, Bacterial, DNA, Intergenic, Evolution, Molecular, Genes, Bacterial, Genome, Bacterial, Geobacillus, Histidine-tRNA Ligase, Inverted Repeat Sequences, Molecular Sequence Data, Phylogeny, Sequence Alignment, Sequence Analysis, DNA}, issn = {1471-2148}, doi = {10.1186/1471-2148-10-230}, author = {Aklujkar, Muktak and Lovley, Derek R} } @article {432, title = {Metabolic response of Geobacter sulfurreducens towards electron donor/acceptor variation.}, journal = {Microb Cell Fact}, volume = {9}, year = {2010}, month = {2010}, pages = {90}, abstract = {BACKGROUND: Geobacter sulfurreducens is capable of coupling the complete oxidation of organic compounds to iron reduction. The metabolic response of G. sulfurreducens towards variations in electron donors (acetate, hydrogen) and acceptors (Fe(III), fumarate) was investigated via (13)C-based metabolic flux analysis. We examined the (13)C-labeling patterns of proteinogenic amino acids obtained from G. sulfurreducens cultured with (13)C-acetate. RESULTS: Using (13)C-based metabolic flux analysis, we observed that donor and acceptor variations gave rise to differences in gluconeogenetic initiation, tricarboxylic acid cycle activity, and amino acid biosynthesis pathways. Culturing G. sulfurreducens cells with Fe(III) as the electron acceptor and acetate as the electron donor resulted in pyruvate as the primary carbon source for gluconeogenesis. When fumarate was provided as the electron acceptor and acetate as the electron donor, the flux analysis suggested that fumarate served as both an electron acceptor and, in conjunction with acetate, a carbon source. Growth on fumarate and acetate resulted in the initiation of gluconeogenesis by phosphoenolpyruvate carboxykinase and a slightly elevated flux through the oxidative tricarboxylic acid cycle as compared to growth with Fe(III) as the electron acceptor. In addition, the direction of net flux between acetyl-CoA and pyruvate was reversed during growth on fumarate relative to Fe(III), while growth in the presence of Fe(III) and acetate which provided hydrogen as an electron donor, resulted in decreased flux through the tricarboxylic acid cycle. CONCLUSIONS: We gained detailed insight into the metabolism of G. sulfurreducens cells under various electron donor/acceptor conditions using (13)C-based metabolic flux analysis. Our results can be used for the development of G. sulfurreducens as a chassis for a variety of applications including bioremediation and renewable biofuel production.}, keywords = {Acetic Acid, Acetyl Coenzyme A, Amino Acids, Carbon Isotopes, Citric Acid Cycle, Electrons, Ferric Compounds, Fumarates, Geobacter, Gluconeogenesis, Oxidation-Reduction, Phosphoenolpyruvate Carboxykinase (GTP), Pyruvates}, issn = {1475-2859}, doi = {10.1186/1475-2859-9-90}, author = {Yang, Tae Hoon and Coppi, Maddalena V and Lovley, Derek R and Sun, Jun} } @article {376, title = {Microbial communities acclimate to recurring changes in soil redox potential status.}, journal = {Environ Microbiol}, volume = {12}, year = {2010}, month = {2010 Dec}, pages = {3137-49}, abstract = {Rapidly fluctuating environmental conditions can significantly stress organisms, particularly when fluctuations cross thresholds of normal physiological tolerance. Redox potential fluctuations are common in humid tropical soils, and microbial community acclimation or avoidance strategies for survival will in turn shape microbial community diversity and biogeochemistry. To assess the extent to which indigenous bacterial and archaeal communities are adapted to changing in redox potential, soils were incubated under static anoxic, static oxic or fluctuating redox potential conditions, and the standing (DNA-based) and active (RNA-based) communities and biogeochemistry were determined. Fluctuating redox potential conditions permitted simultaneous CO$_{2}$ respiration, methanogenesis, N$_{2}$O production and iron reduction. Exposure to static anaerobic conditions significantly changed community composition, while 4-day redox potential fluctuations did not. Using RNA:DNA ratios as a measure of activity, 285 taxa were more active under fluctuating than static conditions, compared with three taxa that were more active under static compared with fluctuating conditions. These data suggest an indigenous microbial community adapted to fluctuating redox potential.}, keywords = {Acclimatization, Archaea, Bacteria, Carbon Dioxide, DNA, Archaeal, DNA, Bacterial, Iron, Oligonucleotide Array Sequence Analysis, Oxidation-Reduction, RNA, Archaeal, RNA, Bacterial, Soil, Soil Microbiology, Trees}, issn = {1462-2920}, doi = {10.1111/j.1462-2920.2010.02286.x}, author = {Deangelis, Kristen M and Silver, Whendee L and Thompson, Andrew W and Firestone, Mary K} } @article {435, title = {Microbial electrosynthesis: feeding microbes electricity to convert carbon dioxide and water to multicarbon extracellular organic compounds.}, journal = {MBio}, volume = {1}, year = {2010}, month = {2010}, abstract = {The possibility of providing the acetogenic microorganism Sporomusa ovata with electrons delivered directly to the cells with a graphite electrode for the reduction of carbon dioxide to organic compounds was investigated. Biofilms of S. ovata growing on graphite cathode surfaces consumed electrons with the reduction of carbon dioxide to acetate and small amounts of 2-oxobutyrate. Electrons appearing in these products accounted for over 85\% of the electrons consumed. These results demonstrate that microbial production of multicarbon organic compounds from carbon dioxide and water with electricity as the energy source is feasible.}, keywords = {Bioelectric Energy Sources, Carbon Dioxide, Electricity, Organic Chemicals, Veillonellaceae, Water}, issn = {2150-7511}, doi = {10.1128/mBio.00103-10}, author = {Nevin, Kelly P and Woodard, Trevor L and Franks, Ashley E and Summers, Zarath M and Lovley, Derek R} } @article {450, title = {Microtoming coupled to microarray analysis to evaluate the spatial metabolic status of Geobacter sulfurreducens biofilms.}, journal = {ISME J}, volume = {4}, year = {2010}, month = {2010 Apr}, pages = {509-19}, abstract = {Further insight into the metabolic status of cells within anode biofilms is essential for understanding the functioning of microbial fuel cells and developing strategies to optimize their power output. Cells throughout anode biofilms of Geobacter sulfurreducens reduced the metabolic stains: 5-cyano-2,3-ditolyl tetrazolium chloride and Redox Green, suggesting metabolic activity throughout the biofilm. To compare the metabolic status of cells growing close to the anode versus cells in the outer portion of the anode biofilm, anode biofilms were encased in resin and sectioned into inner (0-20 microm from anode surface) and outer (30-60 microm) fractions. Transcriptional analysis revealed that, at a twofold threshold, 146 genes had significant (P<0.05) differences in transcript abundance between the inner and outer biofilm sections. Only 1 gene, GSU0093, a hypothetical ATP-binding cassette transporter, had significantly higher transcript abundances in the outer biofilm. Genes with lower transcript abundance in the outer biofilm included genes for ribosomal proteins and NADH dehydrogenase, suggesting lower metabolic rates. However, differences in transcript abundance were relatively low (The abundances of hyperthermophilic heterotrophs, methanogens, and autotrophic reducers of amorphous Fe(III) oxide in 18 samples of deep-sea hydrothermal vent sulfide chimneys of the Endeavour Segment were measured. The results indicate that conditions favor the growth of iron reducers toward the interiors of these deposits and that of heterotrophs toward the outer surfaces near high-temperature polychaete worms (Paralvinella sulfincola).
}, keywords = {Animals, Biodiversity, Ferric Compounds, Hot Springs, Methane, Oxidation-Reduction, Pacific Ocean, Polychaeta, Seawater}, issn = {1098-5336}, doi = {10.1128/AEM.01462-08}, author = {Ver Eecke, Helene C and Kelley, Deborah S and Holden, James F} } @article {462, title = {Anode biofilm transcriptomics reveals outer surface components essential for high density current production in Geobacter sulfurreducens fuel cells.}, journal = {PLoS One}, volume = {4}, year = {2009}, month = {2009}, pages = {e5628}, abstract = {The mechanisms by which Geobacter sulfurreducens transfers electrons through relatively thick (>50 microm) biofilms to electrodes acting as a sole electron acceptor were investigated. Biofilms of Geobacter sulfurreducens were grown either in flow-through systems with graphite anodes as the electron acceptor or on the same graphite surface, but with fumarate as the sole electron acceptor. Fumarate-grown biofilms were not immediately capable of significant current production, suggesting substantial physiological differences from current-producing biofilms. Microarray analysis revealed 13 genes in current-harvesting biofilms that had significantly higher transcript levels. The greatest increases were for pilA, the gene immediately downstream of pilA, and the genes for two outer c-type membrane cytochromes, OmcB and OmcZ. Down-regulated genes included the genes for the outer-membrane c-type cytochromes, OmcS and OmcT. Results of quantitative RT-PCR of gene transcript levels during biofilm growth were consistent with microarray results. OmcZ and the outer-surface c-type cytochrome, OmcE, were more abundant and OmcS was less abundant in current-harvesting cells. Strains in which pilA, the gene immediately downstream from pilA, omcB, omcS, omcE, or omcZ was deleted demonstrated that only deletion of pilA or omcZ severely inhibited current production and biofilm formation in current-harvesting mode. In contrast, these gene deletions had no impact on biofilm formation on graphite surfaces when fumarate served as the electron acceptor. These results suggest that biofilms grown harvesting current are specifically poised for electron transfer to electrodes and that, in addition to pili, OmcZ is a key component in electron transfer through differentiated G. sulfurreducens biofilms to electrodes.}, keywords = {Amino Acid Sequence, Bacterial Outer Membrane Proteins, Bioelectric Energy Sources, Biofilms, Cytochromes, Electrodes, Electron Transport, Fumarates, Gene Deletion, Gene Expression Profiling, Gene Expression Regulation, Bacterial, Genetic Complementation Test, Geobacter, Microscopy, Confocal, Molecular Sequence Data, Oligonucleotide Array Sequence Analysis, Oxidation-Reduction, RNA, Messenger, Up-Regulation}, issn = {1932-6203}, doi = {10.1371/journal.pone.0005628}, author = {Nevin, Kelly P and Kim, Byoung-Chan and Glaven, Richard H and Johnson, Jessica P and Woodard, Trevor L and Meth{\'e}, Barbara A and Didonato, Raymond J and Covalla, Sean F and Franks, Ashley E and Liu, Anna and Lovley, Derek R} } @article {733, title = {Antimicrobial polymers prepared by ring-opening metathesis polymerization: manipulating antimicrobial properties by organic counterion and charge density variation.}, journal = {Chemistry}, volume = {15}, year = {2009}, month = {2009 Nov 2}, pages = {11715-22}, abstract = {The synthesis and characterization of a series of poly(oxanorbornene)-based synthetic mimics of antimicrobial peptides (SMAMPs) is presented. In the first part, the effect of different organic counterions on the antimicrobial properties of the SMAMPs was investigated. Unexpectedly, adding hydrophobicity by complete anion exchange did not increase the SMAMPs{\textquoteright} antimicrobial activity. It was found by dye-leakage studies that this was due to the loss of membrane activity of these polymers caused by the formation of tight ion pairs between the organic counterions and the polymer backbone. In the second part, the effect of molecular charge density on the biological properties of a SMAMP was investigated. The results suggest that, above a certain charge threshold, neither minimum inhibitory concentration (MIC90) nor hemolytic activity (HC50) is greatly affected by adding more cationic groups to the molecule. A SMAMP with an MIC90 of 4 microg mL(-1) against Staphylococcus aureus and a selectivity (=HC50/MIC90) of 650 was discovered, the most selective SMAMP to date.}, keywords = {Anti-Infective Agents, Biomimetic Materials, Cations, Diamines, Hemolysis, Hydrophobic and Hydrophilic Interactions, Microbial Sensitivity Tests, Polymers}, issn = {1521-3765}, doi = {10.1002/chem.200900606}, author = {Lienkamp, Karen and Madkour, Ahmad E and Kumar, Kushi-Nidhi and N{\"u}sslein, Klaus and Tew, Gregory N} } @article {377, title = {Bacterial diversity analysis of Huanglongbing pathogen-infected citrus, using PhyloChip arrays and 16S rRNA gene clone library sequencing.}, journal = {Appl Environ Microbiol}, volume = {75}, year = {2009}, month = {2009 Mar}, pages = {1566-74}, abstract = {The bacterial diversity associated with citrus leaf midribs was characterized for citrus groves that contained the Huanglongbing (HLB) pathogen, which has yet to be cultivated in vitro. We employed a combination of high-density phylogenetic 16S rRNA gene microarrays and 16S rRNA gene clone library sequencing to determine the microbial community composition for symptomatic and asymptomatic citrus midribs. Our results revealed that citrus leaf midribs can support a diversity of microbes. PhyloChip analysis indicated that 47 orders of bacteria in 15 phyla were present in the citrus leaf midribs, while 20 orders in 8 phyla were observed with the cloning and sequencing method. PhyloChip arrays indicated that nine taxa were significantly more abundant in symptomatic midribs than in asymptomatic midribs. "Candidatus Liberibacter asiaticus" was detected at a very low level in asymptomatic plants but was over 200 times more abundant in symptomatic plants. The PhyloChip analysis results were further verified by sequencing 16S rRNA gene clone libraries, which indicated the dominance of "Candidatus Liberibacter asiaticus" in symptomatic leaves. These data implicate "Candidatus Liberibacter asiaticus" as the pathogen responsible for HLB disease.}, keywords = {Bacteria, Biodiversity, Citrus, DNA, Bacterial, DNA, Ribosomal, Genes, rRNA, Microarray Analysis, Molecular Sequence Data, Phylogeny, Plant Diseases, Plant Leaves, Rhizobiaceae, RNA, Bacterial, RNA, Ribosomal, 16S, Sequence Analysis, DNA, Sequence Homology, Nucleic Acid}, issn = {1098-5336}, doi = {10.1128/AEM.02404-08}, author = {Sagaram, Uma Shankar and Deangelis, Kristen M and Trivedi, Pankaj and Andersen, Gary L and Lu, Shi-En and Wang, Nian} } @article {735, title = {Biological perchlorate reduction in packed bed reactors using elemental sulfur.}, journal = {Environ Sci Technol}, volume = {43}, year = {2009}, month = {2009 Jun 15}, pages = {4466-71}, abstract = {Sulfur-utilizing perchlorate (ClO4-)-reducing bacteria were enriched from a denitrifying wastewater seed with elemental sulfur (S0) as an electron donor. The enrichment was composed of a diverse microbial community, with the majority identified as members of the phylum Proteobacteria. Cultures were inoculated into bench-scale packed bed reactors (PBR) with S0 and crushed oyster shell packing media. High ClO4-concentrations (5-8 mg/L) were reduced to < 0.5 mg/L at an empty bed contact time (EBCT) of 13 h. Low C1O4- concentrations (60-120 microg/L), more typical of contaminated groundwater sites, were reduced to < 4 microg/L at an EBCT of 7.5 h. PBR performance decreased when effluent recirculation was applied or when smaller S0 particle sizes were used, indicating that mass transfer of ClO4- to the attached biofilm was not the limiting mechanism in this process, and that biofilm acclimation and growth were key factors in overall reactor performance. The presence of nitrate (6.5 mg N/L) inhibited ClO4- reduction. The microbial community composition was found to change with ClO4- availability from a majority of Beta-Proteobacteria near the influent end of the reactor to primarily sulfur-oxidizing bacteria near the effluent end of the reactor.}, keywords = {Bacteria, Bioreactors, Environmental Pollutants, Medical Waste Disposal, Oxidation-Reduction, Perchloric Acid, Sulfur}, issn = {0013-936X}, author = {Sahu, Ashish K and Conneely, Teresa and N{\"u}sslein, Klaus R and Ergas, Sarina J} } @article {739, title = {Comparison of facially amphiphilic versus segregated monomers in the design of antibacterial copolymers.}, journal = {Chemistry}, volume = {15}, year = {2009}, month = {2009}, pages = {433-9}, abstract = {A direct comparison of two strategies for designing antimicrobial polymers is presented. Previously, we published several reports on the use of facially amphiphilic (FA) monomers which led to polynorbornenes with excellent antimicrobial activities and selectivities. Our polymers obtained by copolymerization of structurally similar segregated monomers, in which cationic and non-polar moieties reside on separate repeat units, led to polymers with less pronounced activities. A wide range of polymer amphiphilicities was surveyed by pairing a cationic oxanorbornene with eleven different non-polar monomers and varying the comonomer feed ratios. Their properties were tested using antimicrobial assays and copolymers possessing intermediate hydrophobicities were the most active. Polymer-induced leakage of dye-filled liposomes and microscopy of polymer-treated bacteria support a membrane-based mode of action. From these results there appears to be profound differences in how a polymer made from FA monomers interacts with the phospholipid bilayer compared with copolymers from segregated monomers. We conclude that a well-defined spatial relationship of the whole polymer is crucial to obtain synthetic mimics of antimicrobial peptides (SMAMPs): charged and non-polar moieties need to be balanced locally, for example, at the monomer level, and not just globally. We advocate the use of FA monomers for better control of biological properties. It is expected that this principle will be usefully applied to other backbones such as the polyacrylates, polystyrenes, and non-natural polyamides.}, keywords = {Anti-Bacterial Agents, Bacteria, Cell Membrane, Drug Design, Hemolysis, Plastics, Polymers, Surface-Active Agents}, issn = {1521-3765}, doi = {10.1002/chem.200801233}, author = {Gabriel, Gregory J and Maegerlein, Janet A and Nelson, Christopher F and Dabkowski, Jeffrey M and Eren, Tarik and N{\"u}sslein, Klaus and Tew, Gregory N} } @article {768, title = {Correlation between tick density and pathogen endemicity, New Hampshire.}, journal = {Emerg Infect Dis}, volume = {15}, year = {2009}, month = {2009 Apr}, pages = {585-7}, abstract = {To assess the endemicity of tick-borne pathogens in New Hampshire, we surveyed adult tick vectors. Pathogens were more prevalent in areas of high tick density, suggesting a correlation between tick establishment and pathogen endemicity. Infection rates in ticks correlated with disease frequency in humans.}, keywords = {Anaplasma phagocytophilum, Animals, Arachnid Vectors, Babesia microti, Babesiosis, Biosurveillance, Borrelia burgdorferi, Communicable Diseases, Emerging, Ehrlichiosis, Humans, Ixodes, Lyme Disease, New Hampshire}, issn = {1080-6059}, doi = {10.3201/eid1504.080940}, author = {Walk, Seth T and Xu, Guang and Stull, Jason W and Rich, Stephen M} } @article {464, title = {Coupling a genome-scale metabolic model with a reactive transport model to describe in situ uranium bioremediation.}, journal = {Microb Biotechnol}, volume = {2}, year = {2009}, month = {2009 Mar}, pages = {274-86}, abstract = {The increasing availability of the genome sequences of microorganisms involved in important bioremediation processes makes it feasible to consider developing genome-scale models that can aid in predicting the likely outcome of potential subsurface bioremediation strategies. Previous studies of the in situ bioremediation of uranium-contaminated groundwater have demonstrated that Geobacter species are often the dominant members of the groundwater community during active bioremediation and the primary organisms catalysing U(VI) reduction. Therefore, a genome-scale, constraint-based model of the metabolism of Geobacter sulfurreducens was coupled with the reactive transport model HYDROGEOCHEM in an attempt to model in situ uranium bioremediation. In order to simplify the modelling, the influence of only three growth factors was considered: acetate, the electron donor added to stimulate U(VI) reduction; Fe(III), the electron acceptor primarily supporting growth of Geobacter; and ammonium, a key nutrient. The constraint-based model predicted that growth yields of Geobacter varied significantly based on the availability of these three growth factors and that there are minimum thresholds of acetate and Fe(III) below which growth and activity are not possible. This contrasts with typical, empirical microbial models that assume fixed growth yields and the possibility for complete metabolism of the substrates. The coupled genome-scale and reactive transport model predicted acetate concentrations and U(VI) reduction rates in a field trial of in situ uranium bioremediation that were comparable to the predictions of a calibrated conventional model, but without the need for empirical calibration, other than specifying the initial biomass of Geobacter. These results suggest that coupling genome-scale metabolic models with reactive transport models may be a good approach to developing models that can be truly predictive, without empirical calibration, for evaluating the probable response of subsurface microorganisms to possible bioremediation approaches prior to implementation.}, keywords = {Acetates, Biodegradation, Environmental, Biological Transport, Genome, Bacterial, Geobacter, Iron, Models, Biological, Uranium}, issn = {1751-7915}, doi = {10.1111/j.1751-7915.2009.00087.x}, author = {Scheibe, Timothy D and Mahadevan, Radhakrishnan and Fang, Yilin and Garg, Srinath and Long, Philip E and Lovley, Derek R} } @article {690, title = {Direct recognition of the mycobacterial glycolipid, trehalose dimycolate, by C-type lectin Mincle.}, journal = {J Exp Med}, volume = {206}, year = {2009}, month = {2009 Dec 21}, pages = {2879-88}, abstract = {Tuberculosis remains a fatal disease caused by Mycobacterium tuberculosis, which contains various unique components that affect the host immune system. Trehalose-6,6{\textquoteright}-dimycolate (TDM; also called cord factor) is a mycobacterial cell wall glycolipid that is the most studied immunostimulatory component of M. tuberculosis. Despite five decades of research on TDM, its host receptor has not been clearly identified. Here, we demonstrate that macrophage inducible C-type lectin (Mincle) is an essential receptor for TDM. Heat-killed mycobacteria activated Mincle-expressing cells, but the activity was lost upon delipidation of the bacteria; analysis of the lipid extracts identified TDM as a Mincle ligand. TDM activated macrophages to produce inflammatory cytokines and nitric oxide, which are completely suppressed in Mincle-deficient macrophages. In vivo TDM administration induced a robust elevation of inflammatory cytokines in sera and characteristic lung inflammation, such as granuloma formation. However, no TDM-induced lung granuloma was formed in Mincle-deficient mice. Whole mycobacteria were able to activate macrophages even in MyD88-deficient background, but the activation was significantly diminished in Mincle/MyD88 double-deficient macrophages. These results demonstrate that Mincle is an essential receptor for the mycobacterial glycolipid, TDM.}, keywords = {Animals, Cord Factors, Granuloma, Lectins, C-Type, Ligands, Lung Diseases, Macrophage Activation, Membrane Proteins, Mice, Mice, Inbred C57BL, Myeloid Differentiation Factor 88, Receptors, IgG}, issn = {1540-9538}, doi = {10.1084/jem.20091750}, author = {Ishikawa, Eri and Ishikawa, Tetsuaki and Morita, Yasu S and Toyonaga, Kenji and Yamada, Hisakata and Takeuchi, Osamu and Kinoshita, Taroh and Akira, Shizuo and Yoshikai, Yasunobu and Yamasaki, Sho} } @article {473, title = {Diversity of promoter elements in a Geobacter sulfurreducens mutant adapted to disruption in electron transfer.}, journal = {Funct Integr Genomics}, volume = {9}, year = {2009}, month = {2009 Feb}, pages = {15-25}, abstract = {The delta-proteobacterium, Geobacter sulfurreducens, can obtain energy by coupling the oxidation of organic matter to the reduction of insoluble Fe(III) or the anode of a microbial fuel cell. Because Fe(III) oxide or the anode surface, in contrast to oxygen, nitrate, or sulfate, is not soluble nor can it be reduced readily, Geobacter species have developed mechanisms which allow electrons to be delivered across outer membrane to the cell surface. OmcB is an outer-membrane c-type cytochrome important for G. sulfurreducens Fe(III) respiration. In the absence of OmcB, cells lost the ability to reduce soluble or insoluble Fe(III). However, the omcB deletion mutant can slowly adapt to growth on soluble Fe(III) over prolonged incubation in the medium with acetate as the electron donor. We discuss available information about predicted or experimentally validated promoters and transcription regulatory sites identified upstream of operons with transcriptional expression significantly changed in the adapted omcB mutant. DNA sequences of upstream regions of coregulated operons in the adapted mutant are divergent, suggesting the presence of recognition sites for different transcriptional regulators and indicating that adaptation of the omcB mutant to growth on soluble Fe(III) has shifted the relevant expression networks involved to a more diverse molecular basis.}, keywords = {Adaptation, Physiological, Electron Transport, Genetic Variation, Geobacter, Mutation, Promoter Regions, Genetic}, issn = {1438-7948}, doi = {10.1007/s10142-008-0094-7}, author = {Krushkal, Julia and Leang, Ching and Barbe, Jose F and Qu, Yanhua and Yan, Bin and Puljic, Marko and Adkins, Ronald M and Lovley, Derek R} } @article {734, title = {"Doubly selective" antimicrobial polymers: how do they differentiate between bacteria?}, journal = {Chemistry}, volume = {15}, year = {2009}, month = {2009 Nov 2}, pages = {11710-4}, abstract = {We have investigated how doubly selective synthetic mimics of antimicrobial peptides (SMAMPs), which can differentiate not only between bacteria and mammalian cells, but also between Gram-negative and Gram-positive bacteria, make the latter distinction. By dye-leakage experiments on model vesicles and complementary experiments on bacteria, we were able to relate the Gram selectivity to structural differences of these bacteria types. We showed that the double membrane of E. coli rather than the difference in lipid composition between E. coli and S. aureus was responsible for Gram selectivity. The molecular-weight-dependent antimicrobial activity of the SMAMPs was shown to be a sieving effect: while the 3000 g mol(-1) SMAMP was able to penetrate the peptidoglycan layer of the Gram-positive S. aureus bacteria, the 50000 g mol(-1) SMAMP got stuck and consequently did not have antimicrobial activity.}, keywords = {Anti-Infective Agents, Biomimetic Materials, Escherichia coli, Microbial Sensitivity Tests, Peptidoglycan, Polymers, Staphylococcus aureus}, issn = {1521-3765}, doi = {10.1002/chem.200802558}, author = {Lienkamp, Karen and Kumar, Kushi-Nidhi and Som, Abhigyan and N{\"u}sslein, Klaus and Tew, Gregory N} } @article {1570, title = {Effect of nutrient and selective inhibitor amendments on methane oxidation, nitrous oxide production, and key gene presence and expression in landfill cover soils: characterization of the role of methanotrophs, nitrifiers, and denitrifiers.}, journal = {Appl Microbiol Biotechnol}, volume = {85}, year = {2009}, month = {2009 Nov}, pages = {389-403}, abstract = {Methane and nitrous oxide are both potent greenhouse gasses, with global warming potentials approximately 25 and 298 times that of carbon dioxide. A matrix of soil microcosms was constructed with landfill cover soils collected from the King Highway Landfill in Kalamazoo, Michigan and exposed to geochemical parameters known to affect methane consumption by methanotrophs while also examining their impact on biogenic nitrous oxide production. It was found that relatively dry soils (5\% moisture content) along with 15 mg NH (4) (+) (kg soil)(-1) and 0.1 mg phenylacetylene(kg soil)(-1) provided the greatest stimulation of methane oxidation while minimizing nitrous oxide production. Microarray analyses of pmoA showed that the methanotrophic community structure was dominated by Type II organisms, but Type I genera were more evident with the addition of ammonia. When phenylacetylene was added in conjunction with ammonia, the methanotrophic community structure was more similar to that observed in the presence of no amendments. PCR analyses showed the presence of amoA from both ammonia-oxidizing bacteria and archaea, and that the presence of key genes associated with these cells was reduced with the addition of phenylacetylene. Messenger RNA analyses found transcripts of pmoA, but not of mmoX, nirK, norB, or amoA from either ammonia-oxidizing bacteria or archaea. Pure culture analyses showed that methanotrophs could produce significant amounts of nitrous oxide, particularly when expressing the particulate methane monooxygenase (pMMO). Collectively, these data suggest that methanotrophs expressing pMMO played a role in nitrous oxide production in these microcosms.
}, keywords = {Archaea, Bacteria, Base Sequence, DNA, DNA Primers, DNA, Archaeal, DNA, Bacterial, Global Warming, Greenhouse Effect, Inorganic Chemicals, Methane, Nitrites, Nitrogen, Nitrogen Oxides, Nitrous Oxide, Oxidation-Reduction, Polymerase Chain Reaction, Reverse Transcriptase Polymerase Chain Reaction, RNA, Bacterial, RNA, Messenger, Soil, Water Pollutants, Chemical}, issn = {1432-0614}, doi = {10.1007/s00253-009-2238-7}, author = {Lee, Sung-Woo and Im, Jeongdae and Dispirito, Alan A and Bodrossy, Levente and Barcelona, Michael J and Semrau, Jeremy D} } @article {678, title = {Epitopia: a web-server for predicting B-cell epitopes.}, journal = {BMC Bioinformatics}, volume = {10}, year = {2009}, month = {2009}, pages = {287}, abstract = {BACKGROUND: Detecting candidate B-cell epitopes in a protein is a basic and fundamental step in many immunological applications. Due to the impracticality of experimental approaches to systematically scan the entire protein, a computational tool that predicts the most probable epitope regions is desirable. RESULTS: The Epitopia server is a web-based tool that aims to predict immunogenic regions in either a protein three-dimensional structure or a linear sequence. Epitopia implements a machine-learning algorithm that was trained to discern antigenic features within a given protein. The Epitopia algorithm has been compared to other available epitope prediction tools and was found to have higher predictive power. A special emphasis was put on the development of a user-friendly graphical interface for displaying the results. CONCLUSION: Epitopia is a user-friendly web-server that predicts immunogenic regions for both a protein structure and a protein sequence. Its accuracy and functionality make it a highly useful tool. Epitopia is available at http://epitopia.tau.ac.il and includes extensive explanations and example predictions.}, keywords = {Algorithms, Artificial Intelligence, Computational Biology, Epitopes, B-Lymphocyte, Internet, Software}, issn = {1471-2105}, doi = {10.1186/1471-2105-10-287}, author = {Rubinstein, Nimrod D and Mayrose, Itay and Martz, Eric and Pupko, Tal} } @article {765, title = {Estimating and testing haplotype-trait associations in non-diploid populations.}, journal = {J R Stat Soc Ser C Appl Stat}, volume = {58}, year = {2009}, month = {2009 Dec}, pages = {663-678}, abstract = {Malaria is an infectious disease that is caused by a group of parasites of the genus Plasmodium. Characterizing the association between polymorphisms in the parasite genome and measured traits in an infected human host may provide insight into disease aetiology and ultimately inform new strategies for improved treatment and prevention. This, however, presents an analytic challenge since individuals are often multiply infected with a variable and unknown number of genetically diverse parasitic strains. In addition, data on the alignment of nucleotides on a single chromosome, which is commonly referred to as haplotypic phase, is not generally observed. An expectation-maximization algorithm for estimating and testing associations between haplotypes and quantitative traits has been described for diploid (human) populations. We extend this method to account for both the uncertainty in haplotypic phase and the variable and unknown number of infections in the malaria setting. Further extensions are described for the human immunodeficiency virus quasi-species setting. A simulation study is presented to characterize performance of the method. Application of this approach to data arising from a cross-sectional study of n=126 multiply infected children in Uganda reveals some interesting associations requiring further investigation.}, issn = {0035-9254}, doi = {10.1111/j.1467-9876.2009.00673.x}, author = {Li, X and Thomas, B N and Rich, S M and Ecker, D and Tumwine, J K and Foulkes, A S} } @article {463, title = {Evolution from a respiratory ancestor to fill syntrophic and fermentative niches: comparative fenomics of six Geobacteraceae species.}, journal = {BMC Genomics}, volume = {10}, year = {2009}, month = {2009}, pages = {103}, abstract = {BACKGROUND: The anaerobic degradation of organic matter in natural environments, and the biotechnical use of anaerobes in energy production and remediation of subsurface environments, both require the cooperative activity of a diversity of microorganisms in different metabolic niches. The Geobacteraceae family contains members with three important anaerobic metabolisms: fermentation, syntrophic degradation of fermentation intermediates, and anaerobic respiration. RESULTS: In order to learn more about the evolution of anaerobic microbial communities, the genome sequences of six Geobacteraceae species were analyzed. The results indicate that the last common Geobacteraceae ancestor contained sufficient genes for anaerobic respiration, completely oxidizing organic compounds with the reduction of external electron acceptors, features that are still retained in modern Geobacter and Desulfuromonas species. Evolution of specialization for fermentative growth arose twice, via distinct lateral gene transfer events, in Pelobacter carbinolicus and Pelobacter propionicus. Furthermore, P. carbinolicus gained hydrogenase genes and genes for ferredoxin reduction that appear to permit syntrophic growth via hydrogen production. The gain of new physiological capabilities in the Pelobacter species were accompanied by the loss of several key genes necessary for the complete oxidation of organic compounds and the genes for the c-type cytochromes required for extracellular electron transfer. CONCLUSION: The results suggest that Pelobacter species evolved parallel strategies to enhance their ability to compete in environments in which electron acceptors for anaerobic respiration were limiting. More generally, these results demonstrate how relatively few gene changes can dramatically transform metabolic capabilities and expand the range of environments in which microorganisms can compete.}, keywords = {Anaerobiosis, Bacteria, Anaerobic, Biological Evolution, Cluster Analysis, Deltaproteobacteria, DNA, Bacterial, Fermentation, Gene Transfer, Horizontal, Genome, Bacterial, Genomics, Multigene Family, Phylogeny, Sequence Analysis, DNA}, issn = {1471-2164}, doi = {10.1186/1471-2164-10-103}, author = {Butler, Jessica E and Young, Nelson D and Lovley, Derek R} } @article {766, title = {Examining landscape factors influencing relative distribution of mosquito genera and frequency of virus infection.}, journal = {Ecohealth}, volume = {6}, year = {2009}, month = {2009 Jun}, pages = {239-49}, abstract = {Mosquito-borne infections cause some of the most debilitating human diseases, including yellow fever and malaria, yet we lack an understanding of how disease risk scales with human-driven habitat changes. We present an approach to study variation in mosquito distribution and concomitant viral infections on the landscape level. In a pilot study we analyzed mosquito distribution along a 10-km transect of a West African rainforest area, which included primary forest, secondary forest, plantations, and human settlements. Variation was observed in the abundance of Anopheles, Aedes, Culex, and Uranotaenia mosquitoes between the different habitat types. Screening of trapped mosquitoes from the different habitats led to the isolation of five uncharacterized viruses of the families Bunyaviridae, Coronaviridae, Flaviviridae, and Rhabdoviridae, as well as an unclassified virus. Polymerase chain reaction screening for these five viruses in individual mosquitoes indicated a trend toward infection with specific viruses in specific mosquito genera that differed by habitat. Based on these initial analyses, we believe that further work is indicated to investigate the impact of anthropogenic landscape changes on mosquito distribution and accompanying arbovirus infection.}, keywords = {Africa, Western, Animals, Culicidae, Ecosystem, Humans, Insect Vectors, Polymerase Chain Reaction, Population Surveillance, RNA Viruses, Trees, Tropical Climate}, issn = {1612-9210}, doi = {10.1007/s10393-009-0260-y}, author = {Junglen, S and Kurth, A and Kuehl, H and Quan, P-L and Ellerbrok, H and Pauli, G and Nitsche, A and Nunn, C and Rich, S M and Lipkin, W I and Briese, T and Leendertz, F H} } @article {737, title = {Fast disinfecting antimicrobial surfaces.}, journal = {Langmuir}, volume = {25}, year = {2009}, month = {2009 Jan 20}, pages = {1060-7}, abstract = {Silicon wafers and glass surfaces were functionalized with facially amphiphilic antimicrobial copolymers using the "grafting from" technique. Surface-initiated atom transfer radical polymerization (ATRP) was used to grow poly(butylmethacrylate)-co-poly(Boc-aminoethyl methacrylate) from the surfaces. Upon Boc-deprotection, these surfaces became highly antimicrobial and killed S. aureus and E. coli 100\% in less than 5 min. The molecular weight and grafting density of the polymer were controlled by varying the polymerization time and initiator surface density. Antimicrobial studies showed that the killing efficiency of these surfaces was independent of polymer layer thickness or grafting density within the range of surfaces studied.}, keywords = {Anti-Bacterial Agents, Escherichia coli, Glass, Microbial Sensitivity Tests, Molecular Structure, Particle Size, Polymethacrylic Acids, Silicon, Staphylococcus aureus, Surface Properties}, issn = {0743-7463}, doi = {10.1021/la802953v}, author = {Madkour, Ahmad E and Dabkowski, Jeffery M and Nusslein, Klaus and Tew, Gregory N} } @article {465, title = {Future shock from the microbe electric.}, journal = {Microb Biotechnol}, volume = {2}, year = {2009}, month = {2009 Mar}, pages = {139-41}, keywords = {Bacteria, Bioelectric Energy Sources, Biotechnology, Electricity, Energy-Generating Resources}, issn = {1751-7915}, doi = {10.1111/j.1751-7915.2009.00090_9.x}, author = {Lovley, Derek R} } @article {461, title = {The genome sequence of Geobacter metallireducens: features of metabolism, physiology and regulation common and dissimilar to Geobacter sulfurreducens.}, journal = {BMC Microbiol}, volume = {9}, year = {2009}, month = {2009}, pages = {109}, abstract = {BACKGROUND: The genome sequence of Geobacter metallireducens is the second to be completed from the metal-respiring genus Geobacter, and is compared in this report to that of Geobacter sulfurreducens in order to understand their metabolic, physiological and regulatory similarities and differences. RESULTS: The experimentally observed greater metabolic versatility of G. metallireducens versus G. sulfurreducens is borne out by the presence of more numerous genes for metabolism of organic acids including acetate, propionate, and pyruvate. Although G. metallireducens lacks a dicarboxylic acid transporter, it has acquired a second putative succinate dehydrogenase/fumarate reductase complex, suggesting that respiration of fumarate was important until recently in its evolutionary history. Vestiges of the molybdate (ModE) regulon of G. sulfurreducens can be detected in G. metallireducens, which has lost the global regulatory protein ModE but retained some putative ModE-binding sites and multiplied certain genes of molybdenum cofactor biosynthesis. Several enzymes of amino acid metabolism are of different origin in the two species, but significant patterns of gene organization are conserved. Whereas most Geobacteraceae are predicted to obtain biosynthetic reducing equivalents from electron transfer pathways via a ferredoxin oxidoreductase, G. metallireducens can derive them from the oxidative pentose phosphate pathway. In addition to the evidence of greater metabolic versatility, the G. metallireducens genome is also remarkable for the abundance of multicopy nucleotide sequences found in intergenic regions and even within genes. CONCLUSION: The genomic evidence suggests that metabolism, physiology and regulation of gene expression in G. metallireducens may be dramatically different from other Geobacteraceae.}, keywords = {Bacterial Proteins, DNA, Bacterial, Gene Expression Regulation, Bacterial, Genome, Bacterial, Geobacter, Phylogeny, Sequence Analysis, DNA, Species Specificity, Transcription Factors}, issn = {1471-2180}, doi = {10.1186/1471-2180-9-109}, author = {Aklujkar, Muktak and Krushkal, Julia and DiBartolo, Genevieve and Lapidus, Alla and Land, Miriam L and Lovley, Derek R} } @article {456, title = {Genome-scale comparison and constraint-based metabolic reconstruction of the facultative anaerobic Fe(III)-reducer Rhodoferax ferrireducens.}, journal = {BMC Genomics}, volume = {10}, year = {2009}, month = {2009}, pages = {447}, abstract = {BACKGROUND: Rhodoferax ferrireducens is a metabolically versatile, Fe(III)-reducing, subsurface microorganism that is likely to play an important role in the carbon and metal cycles in the subsurface. It also has the unique ability to convert sugars to electricity, oxidizing the sugars to carbon dioxide with quantitative electron transfer to graphite electrodes in microbial fuel cells. In order to expand our limited knowledge about R. ferrireducens, the complete genome sequence of this organism was further annotated and then the physiology of R. ferrireducens was investigated with a constraint-based, genome-scale in silico metabolic model and laboratory studies. RESULTS: The iterative modeling and experimental approach unveiled exciting, previously unknown physiological features, including an expanded range of substrates that support growth, such as cellobiose and citrate, and provided additional insights into important features such as the stoichiometry of the electron transport chain and the ability to grow via fumarate dismutation. Further analysis explained why R. ferrireducens is unable to grow via photosynthesis or fermentation of sugars like other members of this genus and uncovered novel genes for benzoate metabolism. The genome also revealed that R. ferrireducens is well-adapted for growth in the subsurface because it appears to be capable of dealing with a number of environmental insults, including heavy metals, aromatic compounds, nutrient limitation and oxidative stress. CONCLUSION: This study demonstrates that combining genome-scale modeling with the annotation of a new genome sequence can guide experimental studies and accelerate the understanding of the physiology of under-studied yet environmentally relevant microorganisms.}, keywords = {Comamonadaceae, Comparative Genomic Hybridization, DNA, Bacterial, Ferric Compounds, Genome, Bacterial, Genomics, Models, Biological, Oxidation-Reduction, Sequence Analysis, DNA}, issn = {1471-2164}, doi = {10.1186/1471-2164-10-447}, author = {Risso, Carla and Sun, Jun and Zhuang, Kai and Mahadevan, Radhakrishnan and DeBoy, Robert and Ismail, Wael and Shrivastava, Susmita and Huot, Heather and Kothari, Sagar and Daugherty, Sean and Bui, Olivia and Schilling, Christophe H and Lovley, Derek R and Meth{\'e}, Barbara A} } @article {466, title = {Genome-scale constraint-based modeling of Geobacter metallireducens.}, journal = {BMC Syst Biol}, volume = {3}, year = {2009}, month = {2009}, pages = {15}, abstract = {BACKGROUND: Geobacter metallireducens was the first organism that can be grown in pure culture to completely oxidize organic compounds with Fe(III) oxide serving as electron acceptor. Geobacter species, including G. sulfurreducens and G. metallireducens, are used for bioremediation and electricity generation from waste organic matter and renewable biomass. The constraint-based modeling approach enables the development of genome-scale in silico models that can predict the behavior of complex biological systems and their responses to the environments. Such a modeling approach was applied to provide physiological and ecological insights on the metabolism of G. metallireducens. RESULTS: The genome-scale metabolic model of G. metallireducens was constructed to include 747 genes and 697 reactions. Compared to the G. sulfurreducens model, the G. metallireducens metabolic model contains 118 unique reactions that reflect many of G. metallireducens{\textquoteright} specific metabolic capabilities. Detailed examination of the G. metallireducens model suggests that its central metabolism contains several energy-inefficient reactions that are not present in the G. sulfurreducens model. Experimental biomass yield of G. metallireducens growing on pyruvate was lower than the predicted optimal biomass yield. Microarray data of G. metallireducens growing with benzoate and acetate indicated that genes encoding these energy-inefficient reactions were up-regulated by benzoate. These results suggested that the energy-inefficient reactions were likely turned off during G. metallireducens growth with acetate for optimal biomass yield, but were up-regulated during growth with complex electron donors such as benzoate for rapid energy generation. Furthermore, several computational modeling approaches were applied to accelerate G. metallireducens research. For example, growth of G. metallireducens with different electron donors and electron acceptors were studied using the genome-scale metabolic model, which provided a fast and cost-effective way to understand the metabolism of G. metallireducens. CONCLUSION: We have developed a genome-scale metabolic model for G. metallireducens that features both metabolic similarities and differences to the published model for its close relative, G. sulfurreducens. Together these metabolic models provide an important resource for improving strategies on bioremediation and bioenergy generation.}, keywords = {Biodegradation, Environmental, Biomass, Computer Simulation, Ecosystem, Electron Transport, Energy Metabolism, Genome, Bacterial, Geobacter, Iron, Metabolic Networks and Pathways, Models, Biological, Models, Genetic, Mutation, Phenotype, Species Specificity, Systems Biology}, issn = {1752-0509}, doi = {10.1186/1752-0509-3-15}, author = {Sun, Jun and Sayyar, Bahareh and Butler, Jessica E and Pharkya, Priti and Fahland, Tom R and Famili, Iman and Schilling, Christophe H and Lovley, Derek R and Mahadevan, Radhakrishnan} } @article {458, title = {Genome-wide analysis of the RpoN regulon in Geobacter sulfurreducens.}, journal = {BMC Genomics}, volume = {10}, year = {2009}, month = {2009}, pages = {331}, abstract = {BACKGROUND: The role of the RNA polymerase sigma factor RpoN in regulation of gene expression in Geobacter sulfurreducens was investigated to better understand transcriptional regulatory networks as part of an effort to develop regulatory modules for genome-scale in silico models, which can predict the physiological responses of Geobacter species during groundwater bioremediation or electricity production. RESULTS: An rpoN deletion mutant could not be obtained under all conditions tested. In order to investigate the regulon of the G. sulfurreducens RpoN, an RpoN over-expression strain was made in which an extra copy of the rpoN gene was under the control of a taclac promoter. Combining both the microarray transcriptome analysis and the computational prediction revealed that the G. sulfurreducens RpoN controls genes involved in a wide range of cellular functions. Most importantly, RpoN controls the expression of the dcuB gene encoding the fumarate/succinate exchanger, which is essential for cell growth with fumarate as the terminal electron acceptor in G. sulfurreducens. RpoN also controls genes, which encode enzymes for both pathways of ammonia assimilation that is predicted to be essential under all growth conditions in G. sulfurreducens. Other genes that were identified as part of the RpoN regulon using either the computational prediction or the microarray transcriptome analysis included genes involved in flagella biosynthesis, pili biosynthesis and genes involved in central metabolism enzymes and cytochromes involved in extracellular electron transfer to Fe(III), which are known to be important for growth in subsurface environment or electricity production in microbial fuel cells. The consensus sequence for the predicted RpoN-regulated promoter elements is TTGGCACGGTTTTTGCT. CONCLUSION: The G. sulfurreducens RpoN is an essential sigma factor and a global regulator involved in a complex transcriptional network controlling a variety of cellular processes.}, keywords = {Bacterial Proteins, DNA, Bacterial, Gene Expression Profiling, Gene Expression Regulation, Bacterial, Genome-Wide Association Study, Geobacter, Multigene Family, Oligonucleotide Array Sequence Analysis, Promoter Regions, Genetic, Regulon, RNA Polymerase Sigma 54}, issn = {1471-2164}, doi = {10.1186/1471-2164-10-331}, author = {Leang, Ching and Krushkal, Julia and Ueki, Toshiyuki and Puljic, Marko and Sun, Jun and Ju{\'a}rez, Katy and N{\'u}{\~n}ez, Cinthia and Reguera, Gemma and DiDonato, Raymond and Postier, Bradley and Adkins, Ronald M and Lovley, Derek R} } @article {455, title = {GSEL version 2, an online genome-wide query system of operon organization and regulatory sequence elements of Geobacter sulfurreducens.}, journal = {OMICS}, volume = {13}, year = {2009}, month = {2009 Oct}, pages = {439-49}, abstract = {Geobacter sulfurreducens is a model organism within the delta-Proteobacterial family Geobacteraceae, members of which can participate in environmental bioremediation of metal and organic waste contaminants and in production of bioenergy. In this report, we describe a new, significantly expanded and updated, version 2 of the GSEL (Geobacter Sequence Elements) database ( http://geobacter.org/research/gsel2/ and http://geobacter.org/refs/gsel2/ ) and its accompanying online query system, which compiles information on operon organization and regulatory sequence elements in the genome of G. sulfurreducens. It incorporates a new online graphical browser, provides novel search capabilities, and includes updated operon predictions along with new information on predicted and experimentally validated genome regulatory sites. The GSEL database and online search system provides a unique and comprehensive tool cataloging information about gene regulation in G. sulfurreducens, aiding in investigation of mechanisms that regulate its ability to generate electric power, bioremediate environmental waste, and adapt to environmental changes.}, keywords = {Base Sequence, Databases, Genetic, Gene Expression Regulation, Bacterial, Genome, Bacterial, Geobacter, Humans, Internet, Online Systems, Operon, Regulatory Sequences, Nucleic Acid, Software, User-Computer Interface}, issn = {1557-8100}, doi = {10.1089/omi.2009.0081}, author = {Qu, Yanhua and Brown, Peter and Barbe, Jose F and Puljic, Marko and Merino, Enrique and Adkins, Ronald M and Lovley, Derek R and Krushkal, Julia} } @article {736, title = {Hydrogenotrophic denitrification and perchlorate reduction in ion exchange brines using membrane biofilm reactors.}, journal = {Biotechnol Bioeng}, volume = {104}, year = {2009}, month = {2009 Oct 15}, pages = {483-91}, abstract = {Halophilic (salt loving), hydrogenotrophic (H(2) oxidizing) denitrifying bacteria were investigated for treatment of nitrate (NO3-) and perchlorate (ClO4-) contaminated groundwater and ion exchange (IX) brines. Hydrogenotrophic denitrifying bacteria were enriched from a denitrifying wastewater seed under both halophilc and non-halophilc conditions. The cultures were inoculated into bench-scale membrane biofilm reactors (MBfRs) with an "outside in" configuration, with contaminated water supplied to the lumen of the membranes and H(2) supplied to the shell. Abiotic mass transfer tests showed that H(2) mass transfer coefficients were lower in brines than in tap water at highest Reynolds number, possibly due to increased transport of salts and decreased H(2) solubility at the membrane/liquid interface. An average NO3- removal efficiency of 93\% was observed for the MBfR operated in continuous flow mode with synthetic contaminated groundwater. Removal efficiencies of 30\% for NO3- and 42\% for ClO4- were observed for the MBfR operated with synthetic IX brine in batch operating mode with a reaction time of 53 h. Phylogenetic analysis focused on the active microbial community and revealed that halotolerant, NO3- -reducing bacteria of the bacterial classes Gamma-Proteobacteria and Sphingobacteria were the metabolically dominant members within the stabilized biofilm. This study shows that, despite decreased H(2) transfer under high salt conditions, hydrogenotrophic biological reduction may be successfully used for the treatment of NO3- and ClO- in a MBfR.}, keywords = {Bacteroidetes, Biofilms, Cluster Analysis, DNA, Bacterial, DNA, Ribosomal, Gammaproteobacteria, Hydrogen, Ion Exchange, Membranes, Molecular Sequence Data, Nitrites, Oxidation-Reduction, Perchloric Acid, Phylogeny, RNA, Ribosomal, 16S, Sequence Analysis, DNA, Water Pollutants, Chemical, Water Purification}, issn = {1097-0290}, doi = {10.1002/bit.22414}, author = {Sahu, Ashish K and Conneely, Teresa and N{\"u}sslein, Klaus and Ergas, Sarina J} } @article {738, title = {Hydrophilic modifications of an amphiphilic polynorbornene and the effects on its hemolytic and antibacterial activity.}, journal = {Biomacromolecules}, volume = {10}, year = {2009}, month = {2009 Feb 9}, pages = {353-9}, abstract = {Here we report the modification of an amphiphilic antibacterial polynorbornene, Poly3, via incorporation of hydrophilic, biocompatible groups. The sugar, zwitterionic, and polyethylene glycol based moieties were incorporated in varying ratios by copolymerization and postpolymerization techniques. Well-defined copolymers with molecular weights of 3 kDa and narrow polydispersity indices ranging from 1.08 to 1.15 were obtained. The effects of these modifications on the biological activity of these polymers were analyzed by determining their minimum inhibitory concentrations (MIC) and their hemolytic activities (HC50).}, keywords = {Anti-Bacterial Agents, Bacteria, Biocompatible Materials, Carbohydrates, Hemolysis, Microbial Sensitivity Tests, Plastics, Polyethylene, Static Electricity}, issn = {1526-4602}, doi = {10.1021/bm801129y}, author = {Colak, Semra and Nelson, Christopher F and N{\"u}sslein, Klaus and Tew, Gregory N} } @article {359, title = {Hytrosaviridae: a proposal for classification and nomenclature of a new insect virus family.}, journal = {Arch Virol}, volume = {154}, year = {2009}, month = {2009}, pages = {909-18}, abstract = {Salivary gland hypertrophy viruses (SGHVs) have been identified from different dipteran species, such as the tsetse fly Glossina pallidipes (GpSGHV), the housefly Musca domestica (MdSGHV) and the narcissus bulbfly Merodon equestris (MeSGHV). These viruses share the following characteristics: (i) they produce non-occluded, enveloped, rod-shaped virions that measure 500-1,000 nm in length and 50-100 nm in diameter; (ii) they possess a large circular double-stranded DNA (dsDNA) genome ranging in size from 120 to 190 kbp and having G + C ratios ranging from 28 to 44\%; (iii) they cause overt salivary gland hypertrophy (SGH) symptoms in dipteran adults and partial to complete sterility. The available information on the complete genome sequence of GpSGHV and MdSGHV indicates significant co-linearity between the two viral genomes, whereas no co-linearity was observed with baculoviruses, ascoviruses, entomopoxviruses, iridoviruses and nudiviruses, other large invertebrate DNA viruses. The DNA polymerases encoded by the SGHVs are of the type B and closely related, but they are phylogenetically distant from DNA polymerases encoded by other large dsDNA viruses. The great majority of SGHV ORFs could not be assigned by sequence comparison. Phylogenetic analysis of conserved genes clustered both SGHVs, but distantly from the nudiviruses and baculoviruses. On the basis of the available morphological, (patho)biological, genomic and phylogenetic data, we propose that the two viruses are members of a new virus family named Hytrosaviridae. This proposed family currently comprises two unassigned species, G. pallidipes salivary gland hypertrophy virus and M. domestica salivary gland hypertrophy virus, and a tentative unassigned species, M. equestris salivary gland hypertrophy virus. Here, we present the characteristics and the justification for establishing this new virus family.}, keywords = {Animals, Diptera, DNA, Circular, DNA, Viral, Insect Viruses, Salivary Glands, Terminology as Topic, Virion}, issn = {1432-8798}, doi = {10.1007/s00705-009-0398-5}, author = {Abd-Alla, Adly M M and Vlak, J M and Bergoin, M and Maruniak, J E and Parker, A and Burand, J P and Jehle, J A and Boucias, D G} } @article {459, title = {Influence of heterogeneous ammonium availability on bacterial community structure and the expression of nitrogen fixation and ammonium transporter genes during in situ bioremediation of uranium-contaminated groundwater.}, journal = {Environ Sci Technol}, volume = {43}, year = {2009}, month = {2009 Jun 15}, pages = {4386-92}, abstract = {The influence of ammonium availability on bacterial community structure and the physiological status of Geobacter species during in situ bioremediation of uranium-contaminated groundwater was evaluated. Ammonium concentrations varied by 2 orders of magnitude (< 4 to 400 microM) across th study site. Analysis of 16S rRNA sequences suggested that ammonium may have been one factor influencing the community composition prior to acetate amendment with Rhodoferax species predominating over Geobacter species with higher ammonium and Dechloromonas species dominating at the site with lowest ammonium. However, once acetate was added and dissimilatory metal reduction was stimulated, Geobacter species became the predominant organisms at all locations. Rates of U(VI) reduction appeared to be more related to acetate concentrations rather than ammonium levels. In situ mRNA transcript abundance of the nitrogen fixation gene, nifD, and the ammonium transporter gene, amtB, in Geobacter species indicated that ammonium was the primary source of nitrogen during uranium reduction. The abundance of amtB was inversely correlated to ammonium levels, whereas nifD transcript levels were similar across all sites examined. These results suggest that nifD and amtB expression are closely regulated in response to ammonium availability to ensure an adequate supply of nitrogen while conserving cell resources. Thus, quantifying nifD and amtB transcript expression appears to be a useful approach for monitoring the nitrogen-related physiological status of subsurface Geobacter species. This study also emphasizes the need for more detailed analysis of geochemical and physiological interactions at the field scale in order to adequately model subsurface microbial processes during bioremediation.}, keywords = {Carrier Proteins, DNA, Bacterial, Environmental Remediation, Gene Expression Regulation, Bacterial, Gene Library, Geobacter, Nitrogen Fixation, Quaternary Ammonium Compounds, Time Factors, Uranium, Water, Water Pollutants, Radioactive}, issn = {0013-936X}, author = {Mouser, Paula J and N{\textquoteright}guessan, Lucie A and Elifantz, Hila and Holmes, Dawn E and Williams, Kenneth H and Wilkins, Michael J and Long, Philip E and Lovley, Derek R} } @article {1447, title = {Mycobacterial Esx-3 is required for mycobactin-mediated iron acquisition.}, journal = {Proc Natl Acad Sci U S A}, volume = {106}, year = {2009}, month = {2009 Nov 3}, pages = {18792-7}, abstract = {The Esx secretion pathway is conserved across Gram-positive bacteria. Esx-1, the best-characterized system, is required for virulence of Mycobacterium tuberculosis, although its precise function during infection remains unclear. Esx-3, a paralogous system present in all mycobacterial species, is required for growth in vitro. Here, we demonstrate that mycobacteria lacking Esx-3 are defective in acquiring iron. To compete for the limited iron available in the host and the environment, these organisms use mycobactin, high-affinity iron-binding molecules. In the absence of Esx-3, mycobacteria synthesize mycobactin but are unable to use the bound iron and are impaired severely for growth during macrophage infection. Mycobacteria thus require a specialized secretion system for acquiring iron from siderophores.
}, keywords = {Animals, Bacterial Proteins, Genome, Bacterial, Iron, Macrophages, Mice, Mutation, Mycobacterium, Mycobacterium Infections, Oxazoles, Protein Binding, Secretory Pathway, Siderophores, Transcription, Genetic, Up-Regulation}, issn = {1091-6490}, doi = {10.1073/pnas.0900589106}, author = {Siegrist, M Sloan and Unnikrishnan, Meera and McConnell, Matthew J and Borowsky, Mark and Cheng, Tan-Yun and Siddiqi, Noman and Fortune, Sarah M and Moody, D Branch and Rubin, Eric J} } @article {853, title = {Occurrence of Chlamydia trachomatis and Chlamydia pneumoniae in paediatric respiratory infections.}, journal = {Eur Respir J}, volume = {33}, year = {2009}, month = {2009 Feb}, pages = {360-7}, abstract = {An emerging body of evidence suggests that half of asthma in both children and adults is associated with chronic lung infection. The aim of the present study was to determine the frequency of viable Chlamydia pneumoniae (Cp) and C. trachomatis (Ct) in the respiratory tracts of paediatric patients with chronic respiratory diseases. Bronchoalveolar lavage fluid (BALF) samples obtained from 182 children undergoing bronchoscopy for clinical reasons were assayed using PCR analysis, in vitro tissue culture and immunofluorescence staining for the presence of Cp and Ct. Chlamydia-specific DNA was detected by PCR in 124 (68\%) out of 182 patients; 79 were positive for Cp, 77 positive for Ct and 32 for both organisms; 75 patients had cultivable Chlamydia. Ct DNA prevalence decreased, whereas Cp positivity generally increased with age. A total of 59 out of 128 asthma patients and 16 out of 54 nonasthmatics were Chlamydia culture positive. When the patients were divided into inflammatory versus noninflammatory airway disease, there were 69 (46\%) out of 150 and six (18\%) out of 32 BALF samples with cultivable Chlamydia, respectively. Viable Chlamydia pneumoniae and Chlamydia trachomatis occur frequently in children with chronic respiratory diseases and may be more prevalent in asthma patients. To the current authors{\textquoteright} knowledge, this is the first report of viable Chlamydia trachomatis in the lungs of children.}, keywords = {Adolescent, Bronchoalveolar Lavage Fluid, Bronchoscopy, Child, Child, Preschool, Chlamydia Infections, Chlamydia trachomatis, Chlamydophila pneumoniae, Cohort Studies, Female, Humans, Infant, Infant, Newborn, Male, Microscopy, Fluorescence, Prospective Studies, Respiratory Tract Infections}, issn = {1399-3003}, doi = {10.1183/09031936.00019508}, author = {Webley, W C and Tilahun, Y and Lay, K and Patel, K and Stuart, E S and Andrzejewski, C and Salva, P S} } @article {767, title = {The origin of malignant malaria.}, journal = {Proc Natl Acad Sci U S A}, volume = {106}, year = {2009}, month = {2009 Sep 1}, pages = {14902-7}, abstract = {Plasmodium falciparum, the causative agent of malignant malaria, is among the most severe human infectious diseases. The closest known relative of P. falciparum is a chimpanzee parasite, Plasmodium reichenowi, of which one single isolate was previously known. The co-speciation hypothesis suggests that both parasites evolved separately from a common ancestor over the last 5-7 million years, in parallel with the divergence of their hosts, the hominin and chimpanzee lineages. Genetic analysis of eight new isolates of P. reichenowi, from wild and wild-born captive chimpanzees in Cameroon and C{\^o}te d{\textquoteright}Ivoire, shows that P. reichenowi is a geographically widespread and genetically diverse chimpanzee parasite. The genetic lineage comprising the totality of global P. falciparum is fully included within the much broader genetic diversity of P. reichenowi. This finding is inconsistent with the co-speciation hypothesis. Phylogenetic analysis indicates that all extant P. falciparum populations originated from P. reichenowi, likely by a single host transfer, which may have occurred as early as 2-3 million years ago, or as recently as 10,000 years ago. The evolutionary history of this relationship may be explained by two critical genetic mutations. First, inactivation of the CMAH gene in the human lineage rendered human ancestors unable to generate the sialic acid Neu5Gc from its precursor Neu5Ac, and likely made humans resistant to P. reichenowi. More recently, mutations in the dominant invasion receptor EBA 175 in the P. falciparum lineage provided the parasite with preference for the overabundant Neu5Ac precursor, accounting for its extreme human pathogenicity.}, keywords = {Amino Acid Sequence, Animals, Glycoproteins, Humans, Malaria, Molecular Sequence Data, Mutation, N-Acetylneuraminic Acid, Pan troglodytes, Phylogeny, Plasmodium, Plasmodium falciparum, Protozoan Infections, Animal, Protozoan Proteins, Sequence Alignment}, issn = {1091-6490}, doi = {10.1073/pnas.0907740106}, author = {Rich, Stephen M and Leendertz, Fabian H and Xu, Guang and LeBreton, Matthew and Djoko, Cyrille F and Aminake, Makoah N and Takang, Eric E and Diffo, Joseph L D and Pike, Brian L and Rosenthal, Benjamin M and Formenty, Pierre and Boesch, Christophe and Ayala, Francisco J and Wolfe, Nathan D} } @article {1448, title = {Phage transposon mutagenesis.}, journal = {Methods Mol Biol}, volume = {465}, year = {2009}, month = {2009}, pages = {311-23}, abstract = {Phage transduction is an attractive method of genetic manipulation in mycobacteria. PhiMycoMarT7 is well suited for transposon mutagenesis as it is temperature sensitive for replication and contains T7 promoters that promote transcription, a highly active transposase gene, and an Escherichia coli oriR6 K origin of replication. Mycobacterial transposon mutant libraries produced by PhiMycoMarT7 transduction are amenable to both forward and reverse genetic studies. In this protocol, we detail the preparation of PhiMycoMarT7, including a description of the phage, reconstitution of the phage, purification of plaques, preparation of phage stock, and titering of phage stock. We then describe the transduction procedure and finally outline the isolation of individual transposon mutants.
}, keywords = {Bacteriophages, DNA Transposable Elements, Mutagenesis, Mutation, Mycobacterium, Transduction, Genetic}, issn = {1940-6029}, doi = {10.1007/978-1-59745-207-6_21}, author = {Siegrist, M Sloan and Rubin, Eric J} } @article {490, title = {PilR, a transcriptional regulator for pilin and other genes required for Fe(III) reduction in Geobacter sulfurreducens.}, journal = {J Mol Microbiol Biotechnol}, volume = {16}, year = {2009}, month = {2009}, pages = {146-58}, abstract = {Growth using Fe(III) as a terminal electron acceptor is a critical physiological process in Geobacter sulfurreducens. However, the mechanisms of electron transfer during Fe(III) reduction are only now being understood. It has been demonstrated that the pili in G. sulfurreducens function as microbial nanowires conducting electrons onto Fe(III) oxides. A number of c-type cytochromes have also been shown to play important roles in Fe(III) reduction. However, the regulatory networks controlling the expression of the genes involved in such processes are not well known. Here we report that the expression of pilA, which encodes the pilistructural protein, is directly regulated by a two-component regulatory system in which PilR functions as an RpoN-dependent enhancer binding protein. Surprisingly, a deletion of the pilR gene affected not only insoluble Fe(III) reduction, which requires pili, but also soluble Fe(III) reduction, which, in contrast, does not require pili. Gene expression profiling using whole-genome DNA microarray and quantitative RT-PCR analyses obtained with a PilR-deficient mutant revealed that the expression of pilA and other pilin-related genes are downregulated, while many c-type cytochromes involved in Fe(III) reduction were differentially regulated. This is the first instance of an enhancer binding protein implicated in regulating genes involved in Fe(III) respiratory functions.}, keywords = {Bacterial Proteins, Ferric Compounds, Fimbriae Proteins, Gene Expression Regulation, Bacterial, Genes, Regulator, Geobacter, Oxidation-Reduction, Transcription, Genetic}, issn = {1660-2412}, doi = {10.1159/000115849}, author = {Ju{\'a}rez, Katy and Kim, Byoung-Chan and Nevin, Kelly and Olvera, Leticia and Reguera, Gemma and Lovley, Derek R and Meth{\'e}, Barbara A} } @article {469, title = {Polar lipid fatty acids, LPS-hydroxy fatty acids, and respiratory quinones of three Geobacter strains, and variation with electron acceptor.}, journal = {J Ind Microbiol Biotechnol}, volume = {36}, year = {2009}, month = {2009 Feb}, pages = {205-9}, abstract = {The polar lipid fatty acids, lipopolysaccharide hydroxy-fatty acids, and respiratory quinones of Geobacter metallireducens str. GS-15, Geobacter sulfurreducens str. PCA, and Geobacter bemidjiensis str. Bem are reported. Also, the lipids of G. metallireducens were compared when grown with Fe(3+) or nitrate as electron acceptors and G. sulfurreducens with Fe(3+) or fumarate. In all experiments, the most abundant polar lipid fatty acids were 14:0, i15:0, 16:1 omega 7c, 16:1 omega 5c, and 16:0; lipopolysaccharide hydroxy-fatty acids were dominated by 3oh16:0, 3oh14:0, 9oh16:0, and 10oh16:0; and menaquinone-8 was the most abundant respiratory quinone. Some variation in lipid profiles with strain were observed, but not with electron acceptor.}, keywords = {Culture Media, Electrons, Environmental Microbiology, Fatty Acids, Ferrous Compounds, Geobacter, Lipids, Lipopolysaccharides, Nitrates, Quinones, Vitamin K 2}, issn = {1476-5535}, doi = {10.1007/s10295-008-0486-7}, author = {Hedrick, D B and Peacock, A D and Lovley, D R and Woodard, T L and Nevin, K P and Long, P E and White, D C} } @article {358, title = {Presence and prevalence of viruses in local and migratory honeybees (Apis mellifera) in Massachusetts.}, journal = {Appl Environ Microbiol}, volume = {75}, year = {2009}, month = {2009 Dec}, pages = {7862-5}, abstract = {Migratory and local bees in Massachusetts were analyzed for seven viruses. Three were detected: black queen cell virus (BQCV), deformed wing virus (DWV), and sacbrood virus (SBV). DWV was most common, followed closely by BQCV and then by SBV. BQCV and SBV were present at significantly higher rates in the migratory bees assayed, bringing into question the impact that these bees have on the health of local bee populations.}, keywords = {Animals, Beekeeping, Bees, Insect Viruses, Massachusetts, Reverse Transcriptase Polymerase Chain Reaction}, issn = {1098-5336}, doi = {10.1128/AEM.01319-09}, author = {Welch, Anna and Drummond, Francis and Tewari, Sunil and Averill, Anne and Burand, John P} } @article {457, title = {Proteogenomic monitoring of Geobacter physiology during stimulated uranium bioremediation.}, journal = {Appl Environ Microbiol}, volume = {75}, year = {2009}, month = {2009 Oct}, pages = {6591-9}, abstract = {Implementation of uranium bioremediation requires methods for monitoring the membership and activities of the subsurface microbial communities that are responsible for reduction of soluble U(VI) to insoluble U(IV). Here, we report a proteomics-based approach for simultaneously documenting the strain membership and microbial physiology of the dominant Geobacter community members during in situ acetate amendment of the U-contaminated Rifle, CO, aquifer. Three planktonic Geobacter-dominated samples were obtained from two wells down-gradient of acetate addition. Over 2,500 proteins from each of these samples were identified by matching liquid chromatography-tandem mass spectrometry spectra to peptides predicted from seven isolate Geobacter genomes. Genome-specific peptides indicate early proliferation of multiple M21 and Geobacter bemidjiensis-like strains and later possible emergence of M21 and G. bemidjiensis-like strains more closely related to Geobacter lovleyi. Throughout biostimulation, the proteome is dominated by enzymes that convert acetate to acetyl-coenzyme A and pyruvate for central metabolism, while abundant peptides matching tricarboxylic acid cycle proteins and ATP synthase subunits were also detected, indicating the importance of energy generation during the period of rapid growth following the start of biostimulation. Evolving Geobacter strain composition may be linked to changes in protein abundance over the course of biostimulation and may reflect changes in metabolic functioning. Thus, metagenomics-independent community proteogenomics can be used to diagnose the status of the subsurface consortia upon which remediation biotechnology relies.}, keywords = {Amino Acid Sequence, Bacterial Proteins, Biodegradation, Environmental, Genomics, Geobacter, Molecular Sequence Data, Oxidation-Reduction, Peptide Mapping, Plankton, Proteomics, Uranium, Water Microbiology, Water Pollutants, Radioactive}, issn = {1098-5336}, doi = {10.1128/AEM.01064-09}, author = {Wilkins, Michael J and VerBerkmoes, Nathan C and Williams, Kenneth H and Callister, Stephen J and Mouser, Paula J and Elifantz, Hila and N{\textquoteright}guessan, Lucie A and Thomas, Brian C and Nicora, Carrie D and Shah, Manesh B and Abraham, Paul and Lipton, Mary S and Lovley, Derek R and Hettich, Robert L and Long, Philip E and Banfield, Jillian F} } @article {679, title = {Proteopedia.Org: a scientific "Wiki" bridging the rift between 3D structure and function of biomacromolecules.}, journal = {Biopolymers}, volume = {92}, year = {2009}, month = {2009}, pages = {76-7}, keywords = {Computational Biology, Imaging, Three-Dimensional, Internet, Models, Molecular, Protein Structure, Quaternary, Protein Structure, Tertiary, Proteins, Structure-Activity Relationship}, issn = {0006-3525}, doi = {10.1002/bip.21126}, author = {Martz, Eric} } @article {467, title = {Quantifying expression of Geobacter spp. oxidative stress genes in pure culture and during in situ uranium bioremediation.}, journal = {ISME J}, volume = {3}, year = {2009}, month = {2009 Apr}, pages = {454-65}, abstract = {As part of an effort to diagnose the physiological status of Geobacter species during in situ bioremediation of uranium-contaminated groundwater, transcript levels for two genes potentially associated with oxidative stress, cydA and sodA, were quantified throughout a bioremediation field study in Rifle, CO, USA. Despite the accumulation of Fe(II) in the groundwater, which is inconsistent with the presence of dissolved oxygen, both genes were highly expressed during the bioremediation process. Therefore, the response to oxidative stress was further evaluated with Geobacter uraniireducens, an isolate from the Rifle site. When G. uraniireducens cultured with fumarate as the electron acceptor was exposed to 5\% oxygen for 8 h, there was a significant increase in cydA and sodA transcripts as well as other genes associated with oxygen respiration or oxidative stress. Oxygen-exposed cells had lower transcript abundance for genes associated with anaerobic respiration, metabolism and motility. Short-term oxygen exposure had little impact on cydA transcript levels, as more than 1 h was required for increases to levels comparable to the subsurface. Abundance of cydA and sodA transcripts for the isolate G. sulfurreducens were always higher in cells cultured with Fe(III) compared with fumarate as an electron acceptor, even when fumarate-grown cells were exposed to oxygen, and Fe(III)-grown cells were grown anaerobically. These results suggest that the apparently high Geobacter cydA and sodA expression during bioremediation cannot necessarily be attributed to oxidative stress and demonstrate that diagnosis of the metabolic status of subsurface microorganisms through transcript analysis should be coupled with appropriate geochemical analyses.}, keywords = {Anaerobiosis, Bacterial Proteins, Biodegradation, Environmental, Colorado, Ferric Compounds, Fumarates, Gene Expression Profiling, Geobacter, Oxidative Stress, Soil Microbiology, Uranium}, issn = {1751-7370}, doi = {10.1038/ismej.2008.126}, author = {Mouser, Paula J and Holmes, Dawn E and Perpetua, Lorrie A and DiDonato, Raymond and Postier, Brad and Liu, Anna and Lovley, Derek R} } @article {460, title = {Selection of a variant of Geobacter sulfurreducens with enhanced capacity for current production in microbial fuel cells.}, journal = {Biosens Bioelectron}, volume = {24}, year = {2009}, month = {2009 Aug 15}, pages = {3498-503}, abstract = {Geobacter sulfurreducens produces current densities in microbial fuel cells that are among the highest known for pure cultures. The possibility of adapting this organism to produce even higher current densities was evaluated. A system in which a graphite anode was poised at -400 mV (versus Ag/AgCl) was inoculated with the wild-type strain of G. sulfurreducens, strain DL-1. An isolate, designated strain KN400, was recovered from the biofilm after 5 months of growth on the electrode. KN400 was much more effective in current production than strain DL-1. This was apparent with anodes poised at -400 mV, as well as in systems run in true fuel cell mode. KN400 had current (7.6A/m(2)) and power (3.9 W/m(2)) densities that respectively were substantially higher than those of DL1 (1.4A/m(2) and 0.5 W/m(2)). On a per cell basis KN400 was more effective in current production than DL1, requiring thinner biofilms to make equivalent current. The enhanced capacity for current production in KN400 was associated with a greater abundance of electrically conductive microbial nanowires than DL1 and lower internal resistance (0.015 versus 0.130 Omega/m(2)) and mass transfer limitation in KN400 fuel cells. KN400 produced flagella, whereas DL1 does not. Surprisingly, KN400 had much less outer-surface c-type cytochromes than DL1. KN400 also had a greater propensity to form biofilms on glass or graphite than DL1, even when growing with the soluble electron acceptor, fumarate. These results demonstrate that it is possible to enhance the ability of microorganisms to electrochemically interact with electrodes with the appropriate selective pressure and that improved current production is associated with clear differences in the properties of the outer surface of the cell that may provide insights into the mechanisms for microbe-electrode interactions.}, keywords = {Bioelectric Energy Sources, Electrochemistry, Equipment Design, Equipment Failure Analysis, Geobacter, Species Specificity}, issn = {1873-4235}, doi = {10.1016/j.bios.2009.05.004}, author = {Yi, Hana and Nevin, Kelly P and Kim, Byoung-Chan and Franks, Ashely E and Klimes, Anna and Tender, Leonard M and Lovley, Derek R} } @article {378, title = {Selective progressive response of soil microbial community to wild oat roots.}, journal = {ISME J}, volume = {3}, year = {2009}, month = {2009 Feb}, pages = {168-78}, abstract = {Roots moving through soil induce physical and chemical changes that differentiate rhizosphere from bulk soil, and the effects of these changes on soil microorganisms have long been a topic of interest. The use of a high-density 16S rRNA microarray (PhyloChip) for bacterial and archaeal community analysis has allowed definition of the populations that respond to the root within the complex grassland soil community; this research accompanies compositional changes reported earlier, including increases in chitinase- and protease-specific activity, cell numbers and quorum sensing signal. PhyloChip results showed a significant change compared with bulk soil in relative abundance for 7\% of the total rhizosphere microbial community (147 of 1917 taxa); the 7\% response value was confirmed by16S rRNA terminal restriction fragment length polymorphism analysis. This PhyloChip-defined dynamic subset was comprised of taxa in 17 of the 44 phyla detected in all soil samples. Expected rhizosphere-competent phyla, such as Proteobacteria and Firmicutes, were well represented, as were less-well-documented rhizosphere colonizers including Actinobacteria, Verrucomicrobia and Nitrospira. Richness of Bacteroidetes and Actinobacteria decreased in soil near the root tip compared with bulk soil, but then increased in older root zones. Quantitative PCR revealed rhizosphere abundance of beta-Proteobacteria and Actinobacteria at about 10(8) copies of 16S rRNA genes per g soil, with Nitrospira having about 10(5) copies per g soil. This report demonstrates that changes in a relatively small subset of the soil microbial community are sufficient to produce substantial changes in functions observed earlier in progressively more mature rhizosphere zones.}, keywords = {Avena sativa, Bacteria, Biodiversity, Colony Count, Microbial, Microarray Analysis, Oligonucleotide Array Sequence Analysis, Plant Roots, Polymerase Chain Reaction, RNA, Bacterial, RNA, Ribosomal, 16S, Soil Microbiology}, issn = {1751-7370}, doi = {10.1038/ismej.2008.103}, author = {Deangelis, Kristen M and Brodie, Eoin L and DeSantis, Todd Z and Andersen, Gary L and Lindow, Steven E and Firestone, Mary K} } @article {809, title = {Single molecule analysis of a red fluorescent RecA protein reveals a defect in nucleoprotein filament nucleation that relates to its reduced biological functions.}, journal = {J Biol Chem}, volume = {284}, year = {2009}, month = {2009 Jul 10}, pages = {18664-73}, abstract = {Fluorescent fusion proteins are exceedingly useful for monitoring protein localization in situ or visualizing protein behavior at the single molecule level. Unfortunately, some proteins are rendered inactive by the fusion. To circumvent this problem, we fused a hyperactive RecA protein (RecA803 protein) to monomeric red fluorescent protein (mRFP1) to produce a functional protein (RecA-RFP) that is suitable for in vivo and in vitro analysis. In vivo, the RecA-RFP partially restores UV resistance, conjugational recombination, and SOS induction to recA(-) cells. In vitro, the purified RecA-RFP protein forms a nucleoprotein filament whose k(cat) for single-stranded DNA-dependent ATPase activity is reduced approximately 3-fold relative to wild-type protein, and which is largely inhibited by single-stranded DNA-binding protein. However, RecA protein is also a dATPase; dATP supports RecA-RFP nucleoprotein filament formation in the presence of single-stranded DNA-binding protein. Furthermore, as for the wild-type protein, the activities of RecA-RFP are further enhanced by shifting the pH to 6.2. As a consequence, RecA-RFP is proficient for DNA strand exchange with dATP or at lower pH. Finally, using single molecule visualization, RecA-RFP was seen to assemble into a continuous filament on duplex DNA, and to extend the DNA approximately 1.7-fold. Consistent with its attenuated activities, RecA-RFP nucleates onto double-stranded DNA approximately 3-fold more slowly than the wild-type protein, but still requires approximately 3 monomers to form the rate-limited nucleus needed for filament assembly. Thus, RecA-RFP reveals that its attenuated biological functions correlate with a reduced frequency of nucleoprotein filament nucleation at the single molecule level.}, keywords = {Cell Nucleus, DNA, DNA, Single-Stranded, Escherichia coli, Hydrogen-Ion Concentration, Kinetics, Luminescent Proteins, Nucleoproteins, Plasmids, Protein Binding, Rec A Recombinases, Recombination, Genetic, Sensitivity and Specificity, Ultraviolet Rays}, issn = {0021-9258}, doi = {10.1074/jbc.M109.004895}, author = {Handa, Naofumi and Amitani, Ichiro and Gumlaw, Nathan and Sandler, Steven J and Kowalczykowski, Stephen C} } @article {808, title = {Suppression of constitutive SOS expression by recA4162 (I298V) and recA4164 (L126V) requires UvrD and RecX in Escherichia coli K-12.}, journal = {Mol Microbiol}, volume = {73}, year = {2009}, month = {2009 Jul}, pages = {226-39}, abstract = {Sensing DNA damage and initiation of genetic responses to repair DNA damage are critical to cell survival. In Escherichia coli, RecA polymerizes on ssDNA produced by DNA damage creating a RecA-DNA filament that interacts with the LexA repressor inducing the SOS response. RecA filament stability is negatively modulated by RecX and UvrD. recA730 (E38K) and recA4142 (F217Y) constitutively express the SOS response. recA4162 (I298V) and recA4164 (L126V) are intragenic suppressors of the constitutive SOS phenotype of recA730. Herein, it is shown that these suppressors are not allele specific and can suppress SOS(C) expression of recA730 and recA4142 in cis and in trans. recA4162 and recA4164 single mutants (and the recA730 and recA4142 derivatives) are Rec(+), UV(R) and are able to induce the SOS response after UV treatment like wild-type. UvrD and RecX are required for the suppression in two (recA730,4164 and recA4142,4162) of the four double mutants tested. To explain the data, one model suggests that recA(C) alleles promote SOS(C) expression by mimicking RecA filament structures that induce SOS and the suppressor alleles mimic RecA filament at end of SOS. UvrD and RecX are attracted to these latter structures to help dismantle or destabilize the RecA filament.}, keywords = {Alleles, DNA Helicases, Escherichia coli, Escherichia coli Proteins, Gene Expression Regulation, Bacterial, Mutation, Missense, Rec A Recombinases, SOS Response (Genetics), Substrate Specificity}, issn = {1365-2958}, doi = {10.1111/j.1365-2958.2009.06765.x}, author = {Long, Jarukit E and Renzette, Nicholas and Sandler, Steven J} } @article {471, title = {Transcriptome of Geobacter uraniireducens growing in uranium-contaminated subsurface sediments.}, journal = {ISME J}, volume = {3}, year = {2009}, month = {2009 Feb}, pages = {216-30}, abstract = {To learn more about the physiological state of Geobacter species living in subsurface sediments, heat-sterilized sediments from a uranium-contaminated aquifer in Rifle, Colorado, were inoculated with Geobacter uraniireducens, a pure culture representative of the Geobacter species that predominates during in situ uranium bioremediation at this site. Whole-genome microarray analysis comparing sediment-grown G. uraniireducens with cells grown in defined culture medium indicated that there were 1084 genes that had higher transcript levels during growth in sediments. Thirty-four c-type cytochrome genes were upregulated in the sediment-grown cells, including several genes that are homologous to cytochromes that are required for optimal Fe(III) and U(VI) reduction by G. sulfurreducens. Sediment-grown cells also had higher levels of transcripts, indicative of such physiological states as nitrogen limitation, phosphate limitation and heavy metal stress. Quantitative reverse transcription PCR showed that many of the metabolic indicator genes that appeared to be upregulated in sediment-grown G. uraniireducens also showed an increase in expression in the natural community of Geobacter species present during an in situ uranium bioremediation field experiment at the Rifle site. These results demonstrate that it is feasible to monitor gene expression of a microorganism growing in sediments on a genome scale and that analysis of the physiological status of a pure culture growing in subsurface sediments can provide insights into the factors controlling the physiology of natural subsurface communities.}, keywords = {Colorado, DNA, Bacterial, Environmental Microbiology, Gene Expression Profiling, Geobacter, Geologic Sediments, Molecular Sequence Data, Oligonucleotide Array Sequence Analysis, Sequence Analysis, DNA, Uranium}, issn = {1751-7370}, doi = {10.1038/ismej.2008.89}, author = {Holmes, Dawn E and O{\textquoteright}Neil, Regina A and Chavan, Milind A and N{\textquoteright}guessan, Lucie A and Vrionis, Helen A and Perpetua, Lorrie A and Larrahondo, M Juliana and DiDonato, Raymond and Liu, Anna and Lovley, Derek R} } @article {393, title = {Two functions of the C-terminal domain of Escherichia coli Rob: mediating "sequestration-dispersal" as a novel off-on switch for regulating Rob{\textquoteright}s activity as a transcription activator and preventing degradation of Rob by Lon protease.}, journal = {J Mol Biol}, volume = {388}, year = {2009}, month = {2009 May 8}, pages = {415-30}, abstract = {In Escherichia coli, Rob activates transcription of the SoxRS/MarA/Rob regulon. Previous work revealed that Rob resides in three to four immunostainable foci, that dipyridyl and bile salts are inducers of its activity, and that inducers bind to Rob{\textquoteright}s C-terminal domain (CTD). We propose that sequestration inactivates Rob by blocking its access to the transcriptional machinery and that inducers activate Rob by mediating its dispersal, allowing interaction with RNA polymerase. To test "sequestration-dispersal" as a new mechanism for regulating the activity of transcriptional activators, we fused Rob{\textquoteright}s CTD to SoxS and used indirect immunofluorescence microscopy to determine the effect of inducers on SoxS-Rob{\textquoteright}s cellular localization. Unlike native SoxS, which is uniformly distributed throughout the cell, SoxS-Rob is sequestered without an inducer, but is rapidly dispersed when cells are treated with an inducer. In this manner, Rob{\textquoteright}s CTD serves as an anti-sigma factor in regulating the co-sigma-factor-like activity of SoxS when fused to it. Rob{\textquoteright}s CTD also protects its N-terminus from Lon protease, since Lon{\textquoteright}s normally rapid degradation of SoxS is blocked in the chimera. Accordingly, Rob{\textquoteright}s CTD has novel regulatory properties that can be bestowed on another E. coli protein.}, keywords = {Decanoic Acids, DNA-Binding Proteins, Escherichia coli, Escherichia coli Proteins, Gene Expression Regulation, Bacterial, Genes, Reporter, Microscopy, Fluorescence, Models, Biological, Protease La, Protein Structure, Tertiary, Pyridines, Recombinant Fusion Proteins, Trans-Activators, Transcription, Genetic}, issn = {1089-8638}, doi = {10.1016/j.jmb.2009.03.023}, author = {Griffith, Kevin L and Fitzpatrick, M Megan and Keen, Edward F and Wolf, Richard E} } @article {360, title = {Two viruses that cause salivary gland hypertrophy in Glossina pallidipes and Musca domestica are related and form a distinct phylogenetic clade.}, journal = {J Gen Virol}, volume = {90}, year = {2009}, month = {2009 Feb}, pages = {334-46}, abstract = {Glossina pallidipes and Musca domestica salivary gland hypertrophy viruses (GpSGHV and MdSGHV) replicate in the nucleus of salivary gland cells causing distinct tissue hypertrophy and reduction of host fertility. They share general characteristics with the non-occluded insect nudiviruses, such as being insect-pathogenic, having enveloped, rod-shaped virions, and large circular double-stranded DNA genomes. MdSGHV measures 65x550 nm and contains a 124 279 bp genome (approximately 44 mol\% G+C content) that codes for 108 putative open reading frames (ORFs). GpSGHV, measuring 50x1000 nm, contains a 190 032 bp genome (28 mol\% G+C content) with 160 putative ORFs. Comparative genomic analysis demonstrates that 37 MdSGHV ORFs have homology to 42 GpSGHV ORFs, as some MdSGHV ORFs have homology to two different GpSGHV ORFs. Nine genes with known functions (dnapol, ts, pif-1, pif-2, pif-3, mmp, p74, odv-e66 and helicase-2), a homologue of the conserved baculovirus gene Ac81 and at least 13 virion proteins are present in both SGHVs. The amino acid identity ranged from 19 to 39 \% among ORFs. An (A/T/G)TAAG motif, similar to the baculovirus late promoter motif, was enriched 100 bp upstream of the ORF transcription initiation sites of both viruses. Six and seven putative microRNA sequences were found in MdSGHV and GpSGHV genomes, respectively. There was genome. Collinearity between the two SGHVs, but not between the SGHVs and the nudiviruses. Phylogenetic analysis of conserved genes clustered both SGHVs in a single clade separated from the nudiviruses and baculoviruses. Although MdSGHV and GpSGHV are different viruses, their pathology, host range and genome composition indicate that they are related.}, keywords = {Animals, Chromosome Mapping, Conserved Sequence, Cytomegalovirus, DNA, Viral, Genes, Viral, Genome, Viral, Houseflies, Hypertrophy, Open Reading Frames, Salivary Glands, Tsetse Flies, Virion}, issn = {0022-1317}, doi = {10.1099/vir.0.006783-0}, author = {Garcia-Maruniak, Alejandra and Abd-Alla, Adly M M and Salem, Tamer Z and Parker, Andrew G and Lietze, Verena-Ulrike and van Oers, Monique M and Maruniak, James E and Kim, Woojin and Burand, John P and Cousserans, Fran{\c c}ois and Robinson, Alan S and Vlak, Just M and Bergoin, Max and Boucias, Drion G} } @article {383, title = {Unraveling the secrets of regulating mitochondrial DNA replication.}, journal = {Mol Cell}, volume = {35}, year = {2009}, month = {2009 Aug 28}, pages = {398-400}, abstract = {In this issue, Liu et al. (2009) report that maxicircle DNA copy number in trypanosomes is regulated by proteolysis of a helicase; the complex kinetoplast DNA system yields a clear view of how mitochondrial DNA replication can be regulated.}, keywords = {Animals, DNA Helicases, DNA Replication, DNA, Kinetoplast, DNA, Mitochondrial, DNA, Protozoan, Gene Expression Regulation, Mutation, Peptide Hydrolases, Protozoan Proteins, Time Factors, Trypanosoma brucei brucei}, issn = {1097-4164}, doi = {10.1016/j.molcel.2009.08.007}, author = {Klingbeil, Michele M and Shapiro, Theresa A} } @article {811, title = {UvrD303, a hyperhelicase mutant that antagonizes RecA-dependent SOS expression by a mechanism that depends on its C terminus.}, journal = {J Bacteriol}, volume = {191}, year = {2009}, month = {2009 Mar}, pages = {1429-38}, abstract = {Genomic integrity is critical for an organism{\textquoteright}s survival and ability to reproduce. In Escherichia coli, the UvrD helicase has roles in nucleotide excision repair and methyl-directed mismatch repair and can limit reactions by RecA under certain circumstances. UvrD303 (D403A D404A) is a hyperhelicase mutant, and when expressed from a multicopy plasmid, it results in UV sensitivity (UV(s)), recombination deficiency, and antimutability. In order to understand the molecular mechanism underlying the UV(s) phenotype of uvrD303 cells, this mutation was transferred to the E. coli chromosome and studied in single copy. It is shown here that uvrD303 mutants are UV sensitive, recombination deficient, and antimutable and additionally have a moderate defect in inducing the SOS response after UV treatment. The UV-sensitive phenotype is epistatic with recA and additive with uvrA and is partially suppressed by removing the LexA repressor. Furthermore, uvrD303 is able to inhibit constitutive SOS expression caused by the recA730 mutation. The ability of UvrD303 to antagonize SOS expression was dependent on its 40 C-terminal amino acids. It is proposed that UvrD303, via its C terminus, can decrease the levels of RecA activity in the cell.}, keywords = {DNA Helicases, DNA, Bacterial, Escherichia coli K12, Escherichia coli Proteins, Gene Expression Regulation, Bacterial, Mutation, Rec A Recombinases, Recombination, Genetic, SOS Response (Genetics), Ultraviolet Rays}, issn = {1098-5530}, doi = {10.1128/JB.01415-08}, author = {Centore, Richard C and Leeson, Michael C and Sandler, Steven J} } @article {742, title = {Antimicrobial polymers prepared by ROMP with unprecedented selectivity: a molecular construction kit approach.}, journal = {J Am Chem Soc}, volume = {130}, year = {2008}, month = {2008 Jul 30}, pages = {9836-43}, abstract = {Synthetic Mimics of Antimicrobial Peptides (SMAMPs) imitate natural host-defense peptides, a vital component of the body{\textquoteright}s immune system. This work presents a molecular construction kit that allows the easy and versatile synthesis of a broad variety of facially amphiphilic oxanorbornene-derived monomers. Their ring-opening metathesis polymerization (ROMP) and deprotection provide several series of SMAMPs. Using amphiphilicity, monomer feed ratio, and molecular weight as parameters, polymers with 533 times higher selectivitiy (selecitviy = hemolytic concentration/minimum inhibitory concentration) for bacteria over mammalian cells were discovered. Some of these polymers were 50 times more selective for Gram-positive over Gram-negative bacteria while other polymers surprisingly showed the opposite preference. This kind of "double selectivity" (bacteria over mammalian and one bacterial type over another) is unprecedented in other polymer systems and is attributed to the monomer{\textquoteright}s facial amphiphilicity.}, keywords = {Anti-Infective Agents, Antimicrobial Cationic Peptides, Biomimetic Materials, Erythrocytes, Escherichia coli, Hemolysis, Humans, Hydrophobic and Hydrophilic Interactions, Microbial Sensitivity Tests, Molecular Weight, Norbornanes, Polymers, Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization, Staphylococcus aureus, Structure-Activity Relationship}, issn = {1520-5126}, doi = {10.1021/ja801662y}, author = {Lienkamp, Karen and Madkour, Ahmad E and Musante, Ashlan and Nelson, Christopher F and N{\"u}sslein, Klaus and Tew, Gregory N} } @article {701, title = {Benefits of in-situ synthesized microarrays for analysis of gene expression in understudied microorganisms.}, journal = {J Microbiol Methods}, volume = {74}, year = {2008}, month = {2008 Jul}, pages = {26-32}, abstract = {Although the genome sequences of many microorganisms are now known, whole-genome DNA microarray platforms consisting of PCR amplicon, or oligonucleotide elements printed onto glass slides have been readily available for only a relatively few, highly studied microorganisms. For those microorganisms more recently cultured or studied by fewer investigators it has been difficult to justify the initial time and expense of developing such array platforms especially if only a limited number of gene expression studies are envisioned. However, in-situ synthesized oligonucleotide (ISO) arrays can be inexpensively fabricated on an {\textquoteright}as needed{\textquoteright} basis with a reduced initial investment in time, personnel, resources, and costs. To evaluate the performance of one ISO array platform, gene expression patterns in Geobacter sulfurreducens under nitrogen-fixing conditions were compared with results from quantitative reverse transcriptase PCR (qRT-PCR) and previously published data from a similar experiment using spotted PCR amplicon arrays. There were strong correlations between the results of the ISO arrays and the results from qRT-PCR (r(2)=0.762) and spotted array (r(2)=0.744) analyses. After initial use the ISO arrays could be successfully stripped and reused. The increased flexibility in array design and reusability coupled with a lower initial investment in terms of fabrication time and cost for the ISO arrays suggest that they may be the preferred approach when investigating gene expression in microorganisms, especially when only a few expression studies are required.}, keywords = {Bacterial Proteins, Computational Biology, Gene Expression, Geobacter, Nitrogen Fixation, Oligonucleotide Array Sequence Analysis, Oligonucleotides, Polymerase Chain Reaction}, issn = {0167-7012}, doi = {10.1016/j.mimet.2007.07.004}, author = {Postier, Bradley and DiDonato, Raymond and Nevin, Kelly P and Liu, Anna and Frank, Bryan and Lovley, Derek and Methe, Barbara A} } @article {486, title = {Characterization of extracellular minerals produced during dissimilatory Fe(III) and U(VI) reduction at 100 degrees C by Pyrobaculum islandicum.}, journal = {Geobiology}, volume = {6}, year = {2008}, month = {2008 Mar}, pages = {147-54}, abstract = {In order to gain insight into the significance of biotic metal reduction and mineral formation in hyperthermophilic environments, metal mineralization as a result of the dissimilatory reduction of poorly crystalline Fe(III) oxide, and U(VI) reduction at 100 degrees C by Pyrobaculum islandicum was investigated. When P. islandicum was grown in a medium with poorly crystalline Fe(III) oxide as an electron acceptor and hydrogen as an electron donor, the Fe(III) oxide was reduced to an extracellular, ultrafine-grained magnetite with characteristics similar to that found in some hot environments and that was previously thought to be of abiotic origin. Furthermore, cell suspensions of P. islandicum rapidly reduced the soluble and oxidized form of uranium, U(VI), to extracellular precipitates of the highly insoluble U(IV) mineral, uraninite (UO(2)). The reduction of U(VI) was dependent on the presence of hydrogen as the electron donor. These findings suggest that microbes may play a key role in metal deposition in hyperthermophilic environments and provide a plausible explanation for such phenomena as magnetite accumulation and formation of uranium deposits at ca. 100 degrees C.}, keywords = {Ferric Compounds, Ferumoxytol, Hot Temperature, Hydrogen, Minerals, Oxidation-Reduction, Pyrobaculum, Uranium, X-Ray Diffraction}, issn = {1472-4669}, doi = {10.1111/j.1472-4669.2007.00142.x}, author = {Kashefi, K and Moskowitz, B M and Lovley, D R} } @article {700, title = {Characterizing regulation of metabolism in Geobacter sulfurreducens through genome-wide expression data and sequence analysis.}, journal = {OMICS}, volume = {12}, year = {2008}, month = {2008 Mar}, pages = {33-59}, abstract = {Geobacteraceae are a family of metal reducing bacteria with important applications in bioremediation and electricity generation. G. sulfurreducens is a representative of Geobacteraceae that has been extensively studied with the goal of extending the understanding of this family of organisms for optimizing their practical applications. Here, we have analyzed gene expression data from 10 experiments involving environmental and genetic perturbations and have identified putative transcription factor binding sites (TFBS) involved in regulating key aspects of metabolism. Specifically, we considered data from both a subset of 10 microarray experiments (7 of 10) and all 10 experiments. The expression data from these two sets were independently clustered, and the upstream regions of genes and operons from the clusters in both sets were used to identify TFBS using the AlignACE program. This analysis resulted in the identification of motifs upstream of several genes involved in central metabolism, sulfate assimilation, and energy metabolism, as well as genes potentially encoding acetate permease. Further, similar TFBS were identified from the analysis of both sets, suggesting that these TFBS are significant in the regulation of metabolism in G. sulfurreducens. In addition, we have utilized microarray data to derive condition specific constraints on the capacity of key enzymes in central metabolism. We have incorporated these constraints into the metabolic model of G. sulfurreducens and simulated Fe(II)-limited growth. The resulting prediction was consistent with data, suggesting that regulatory constraints are important for simulating growth phenotypes in nonoptimal environments.}, keywords = {Gene Expression Regulation, Bacterial, Genome, Bacterial, Geobacter, Models, Genetic, Oligonucleotide Array Sequence Analysis, Sequence Analysis, DNA, Transcription, Genetic}, issn = {1536-2310}, doi = {10.1089/omi.2007.0043}, author = {Mahadevan, Radhakrishnan and Yan, Bin and Postier, Brad and Nevin, Kelly P and Woodard, Trevor L and O{\textquoteright}Neil, Regina and Coppi, Maddalena V and Meth{\'e}, Barbara A and Krushkal, Julia} } @article {470, title = {Comparative genomics of Geobacter chemotaxis genes reveals diverse signaling function.}, journal = {BMC Genomics}, volume = {9}, year = {2008}, month = {2008}, pages = {471}, abstract = {BACKGROUND: Geobacter species are delta-Proteobacteria and are often the predominant species in a variety of sedimentary environments where Fe(III) reduction is important. Their ability to remediate contaminated environments and produce electricity makes them attractive for further study. Cell motility, biofilm formation, and type IV pili all appear important for the growth of Geobacter in changing environments and for electricity production. Recent studies in other bacteria have demonstrated that signaling pathways homologous to the paradigm established for Escherichia coli chemotaxis can regulate type IV pili-dependent motility, the synthesis of flagella and type IV pili, the production of extracellular matrix material, and biofilm formation. The classification of these pathways by comparative genomics improves the ability to understand how Geobacter thrives in natural environments and better their use in microbial fuel cells. RESULTS: The genomes of G. sulfurreducens, G. metallireducens, and G. uraniireducens contain multiple (approximately 70) homologs of chemotaxis genes arranged in several major clusters (six, seven, and seven, respectively). Unlike the single gene cluster of E. coli, the Geobacter clusters are not all located near the flagellar genes. The probable functions of some Geobacter clusters are assignable by homology to known pathways; others appear to be unique to the Geobacter sp. and contain genes of unknown function. We identified large numbers of methyl-accepting chemotaxis protein (MCP) homologs that have diverse sensing domain architectures and generate a potential for sensing a great variety of environmental signals. We discuss mechanisms for class-specific segregation of the MCPs in the cell membrane, which serve to maintain pathway specificity and diminish crosstalk. Finally, the regulation of gene expression in Geobacter differs from E. coli. The sequences of predicted promoter elements suggest that the alternative sigma factors sigma28 and sigma54 play a role in regulating the Geobacter chemotaxis gene expression. CONCLUSION: The numerous chemoreceptors and chemotaxis-like gene clusters of Geobacter appear to be responsible for a diverse set of signaling functions in addition to chemotaxis, including gene regulation and biofilm formation, through functionally and spatially distinct signaling pathways.}, keywords = {Amino Acid Sequence, Bacterial Proteins, Chemotaxis, Escherichia coli, Gene Expression Regulation, Bacterial, Genome, Bacterial, Genomics, Geobacter, Membrane Proteins, Molecular Sequence Data, Multigene Family, Promoter Regions, Genetic, Sequence Alignment, Sequence Homology, Amino Acid}, issn = {1471-2164}, doi = {10.1186/1471-2164-9-471}, author = {Tran, Hoa T and Krushkal, Julia and Antommattei, Frances M and Lovley, Derek R and Weis, Robert M} } @article {489, title = {Computational and experimental analysis of redundancy in the central metabolism of Geobacter sulfurreducens.}, journal = {PLoS Comput Biol}, volume = {4}, year = {2008}, month = {2008 Feb}, pages = {e36}, abstract = {Previous model-based analysis of the metabolic network of Geobacter sulfurreducens suggested the existence of several redundant pathways. Here, we identified eight sets of redundant pathways that included redundancy for the assimilation of acetate, and for the conversion of pyruvate into acetyl-CoA. These equivalent pathways and two other sub-optimal pathways were studied using 5 single-gene deletion mutants in those pathways for the evaluation of the predictive capacity of the model. The growth phenotypes of these mutants were studied under 12 different conditions of electron donor and acceptor availability. The comparison of the model predictions with the resulting experimental phenotypes indicated that pyruvate ferredoxin oxidoreductase is the only activity able to convert pyruvate into acetyl-CoA. However, the results and the modeling showed that the two acetate activation pathways present are not only active, but needed due to the additional role of the acetyl-CoA transferase in the TCA cycle, probably reflecting the adaptation of these bacteria to acetate utilization. In other cases, the data reconciliation suggested additional capacity constraints that were confirmed with biochemical assays. The results demonstrate the need to experimentally verify the activity of key enzymes when developing in silico models of microbial physiology based on sequence-based reconstruction of metabolic networks.}, keywords = {Bacterial Proteins, Base Sequence, Computer Simulation, Gene Expression Regulation, Bacterial, Geobacter, Models, Biological, Molecular Sequence Data, Multienzyme Complexes, Signal Transduction}, issn = {1553-7358}, doi = {10.1371/journal.pcbi.0040036}, author = {Segura, Daniel and Mahadevan, Radhakrishnan and Ju{\'a}rez, Katy and Lovley, Derek R} } @article {1210, title = {Constraints on anaerobic respiration in the hyperthermophilic Archaea Pyrobaculum islandicum and Pyrobaculum aerophilum.}, journal = {Appl Environ Microbiol}, volume = {74}, year = {2008}, month = {2008 Jan}, pages = {396-402}, abstract = {Pyrobaculum islandicum uses iron, thiosulfate, and elemental sulfur for anaerobic respiration, while Pyrobaculum aerophilum uses iron and nitrate; however, the constraints on these processes and their physiological mechanisms for iron and sulfur reduction are not well understood. Growth rates on sulfur compounds are highest at pH 5 to 6 and highly reduced (<-420-mV) conditions, while growth rates on nitrate and iron are highest at pH 7 to 9 and more-oxidized (>-210-mV) conditions. Growth on iron expands the known pH range of growth for both organisms. P. islandicum differs from P. aerophilum in that it requires direct contact with insoluble iron oxide for growth, it did not produce any extracellular compounds when grown on insoluble iron, and it lacked 2,6-anthrahydroquinone disulfonate oxidase activity. Furthermore, iron reduction in P. islandicum appears to be completely independent of c-type cytochromes. Like that in P. aerophilum, NADH-dependent ferric reductase activity in P. islandicum increased significantly in iron-grown cultures relative to that in non-iron-grown cultures. Proteomic analyses showed that there were significant increases in the amounts of a putative membrane-bound thiosulfate reductase in P. islandicum cultures grown on thiosulfate relative to those in cultures grown on iron and elemental sulfur. This is the first evidence of this enzyme being used in either a hyperthermophile or an archaeon. Pyrobaculum arsenaticum and Pyrobaculum calidifontis also grew on Fe(III) citrate and insoluble iron oxide, but only P. arsenaticum could grow on insoluble iron without direct contact.
}, keywords = {Anaerobiosis, FMN Reductase, Hydrogen-Ion Concentration, Iron, Nitrates, Oxidation-Reduction, Pyrobaculum, Species Specificity, Sulfur, Sulfurtransferases, Thiosulfates}, issn = {1098-5336}, doi = {10.1128/AEM.02033-07}, author = {Feinberg, Lawrence F and Srikanth, R and Vachet, Richard W and Holden, James F} } @article {395, title = {A degenerate tripartite DNA-binding site required for activation of ComA-dependent quorum response gene expression in Bacillus subtilis.}, journal = {J Mol Biol}, volume = {381}, year = {2008}, month = {2008 Aug 29}, pages = {261-75}, abstract = {In Bacillus subtilis, the transcription factor ComA activates several biological processes in response to increasing population density. Extracellular peptide signaling is used to coordinate the activity of ComA with population density. At low culture densities, when the concentration of signaling peptides is lowest, ComA is largely inactive. At higher densities, when the concentration of signaling peptides is higher, ComA is active and activates the transcription of at least nine operons involved in the development of competence and in the production of degradative enzymes and antibiotics. We found that ComA binds a degenerate tripartite sequence consisting of three DNA-binding determinants or "recognition elements." Mutational analyses showed that all three recognition elements are required for transcription activation in vivo and for specific DNA binding by ComA in vitro. Degeneracy of the recognition elements in the ComA-binding site is an important regulatory feature for coordinating transcription with population density (i.e., promoters containing an optimized binding site have high activity at low culture density and are no longer regulated in the normal-density-dependent manner). We found that purified ComA forms a dimer in solution, and we propose a model for how two dimers of ComA bind to an odd number of DNA-binding determinants to activate transcription of target genes. This DNA-protein architecture for transcription activation appears to be conserved for ComA homologs in other Bacillus species.}, keywords = {Bacillus subtilis, Bacterial Proteins, Base Sequence, Binding Sites, Dimerization, DNA, Bacterial, DNA-Binding Proteins, Electrophoretic Mobility Shift Assay, Gene Expression Regulation, Bacterial, Peptide Synthases, Phosphoprotein Phosphatases, Promoter Regions, Genetic, Protein Binding, Quorum Sensing, Sequence Analysis, DNA, Transcription, Genetic, Transcriptional Activation}, issn = {1089-8638}, doi = {10.1016/j.jmb.2008.06.035}, author = {Griffith, Kevin L and Grossman, Alan D} } @article {498, title = {The degree of redundancy in metabolic genes is linked to mode of metabolism.}, journal = {Biophys J}, volume = {94}, year = {2008}, month = {2008 Feb 15}, pages = {1216-20}, abstract = {An understanding of the factors favoring the maintenance of duplicate genes in microbial genomes is essential for developing models of microbial evolution. A genome-scale flux-balance analysis of the metabolic network of Saccharomyces cerevisiae has suggested that gene duplications primarily provide increased enzyme dosage to enhance metabolic flux because the incidence of gene duplications in essential genes is no higher than that in nonessential genes. Here, we used genome-scale metabolic models to analyze the extent of genetic and biochemical redundancy in prokaryotes that are either specialists, with one major mode of energy generation, or generalists, which have multiple metabolic strategies for conservation of energy. Surprisingly, the results suggest that generalists, such as Escherichia coli and Bacillus subtilis, are similar to the eukaryotic generalist, S. cerevisiae, in having a low percentage (<10\%) of essential genes and few duplications of these essential genes, whereas metabolic specialists, such as Geobacter sulfurreducens and Methanosarcina barkeri, have a high percentage (>30\%) of essential genes and a high degree of genetic redundancy in these genes compared to nonessential genes. Furthermore, the specialist organisms appear to rely more on gene duplications rather than alternative-but-equivalent metabolic pathways to provide resilience to gene loss. Generalists rely more on alternative pathways. Thus, the concept that the role of gene duplications is to boost enzymatic flux rather than provide metabolic resilience may not be universal. Rather, the degree of gene duplication in microorganisms may be linked to mode of metabolism and environmental niche.}, keywords = {Bacterial Physiological Phenomena, Bacterial Proteins, Computer Simulation, Gene Expression Regulation, Bacterial, Genes, Duplicate, Models, Biological, Signal Transduction}, issn = {1542-0086}, doi = {10.1529/biophysj.107.118414}, author = {Mahadevan, R and Lovley, D R} } @article {810, title = {Differential requirements of two recA mutants for constitutive SOS expression in Escherichia coli K-12.}, journal = {PLoS One}, volume = {3}, year = {2008}, month = {2008}, pages = {e4100}, abstract = {BACKGROUND: Repairing DNA damage begins with its detection and is often followed by elicitation of a cellular response. In E. coli, RecA polymerizes on ssDNA produced after DNA damage and induces the SOS Response. The RecA-DNA filament is an allosteric effector of LexA auto-proteolysis. LexA is the repressor of the SOS Response. Not all RecA-DNA filaments, however, lead to an SOS Response. Certain recA mutants express the SOS Response (recA(C)) in the absence of external DNA damage in log phase cells. METHODOLOGY/PRINCIPAL FINDINGS: Genetic analysis of two recA(C) mutants was used to determine the mechanism of constitutive SOS (SOS(C)) expression in a population of log phase cells using fluorescence of single cells carrying an SOS reporter system (sulAp-gfp). SOS(C) expression in recA4142 mutants was dependent on its initial level of transcription, recBCD, recFOR, recX, dinI, xthA and the type of medium in which the cells were grown. SOS(C) expression in recA730 mutants was affected by none of the mutations or conditions tested above. CONCLUSIONS/SIGNIFICANCE: It is concluded that not all recA(C) alleles cause SOS(C) expression by the same mechanism. It is hypothesized that RecA4142 is loaded on to a double-strand end of DNA and that the RecA filament is stabilized by the presence of DinI and destabilized by RecX. RecFOR regulate the activity of RecX to destabilize the RecA filament. RecA730 causes SOS(C) expression by binding to ssDNA in a mechanism yet to be determined.}, keywords = {Escherichia coli K12, Escherichia coli Proteins, Gene Expression Regulation, Bacterial, Genes, Bacterial, Models, Biological, Mutation, Rec A Recombinases, SOS Response (Genetics)}, issn = {1932-6203}, doi = {10.1371/journal.pone.0004100}, author = {Long, Jarukit Edward and Renzette, Nicholas and Centore, Richard C and Sandler, Steven J} } @article {487, title = {Electricity generation by Geobacter sulfurreducens attached to gold electrodes.}, journal = {Langmuir}, volume = {24}, year = {2008}, month = {2008 Apr 15}, pages = {4376-9}, abstract = {The versatility of gold for electrode manufacture suggests that it could be an ideal material for some microbial fuel cell applications. However, previous studies have suggested that microorganisms that readily transfer electrons to graphite do not transfer electrons to gold. Investigations with Geobacter sulfurreducens demonstrated that it could grow on gold anodes producing current nearly as effectively as with graphite anodes. Current production was associated with the development of G. sulfurreducens biofilms up to 40 microm thick. No current was produced if pilA, the gene for the structural protein of the conductive pili of G. sulfurreducens, was deleted. The finding that gold is a suitable anode material for microbial fuel cells offers expanded possibilities for the construction of microbial fuel cells and the electrochemical analysis of microbe-electrode interactions.}, keywords = {Electrodes, Electrons, Geobacter, Gold, Microscopy, Confocal, Microscopy, Electron, Scanning, Surface Properties}, issn = {0743-7463}, doi = {10.1021/la703469y}, author = {Richter, Hanno and McCarthy, Kevin and Nevin, Kelly P and Johnson, Jessica P and Rotello, Vincent M and Lovley, Derek R} } @article {491, title = {Elucidation of an alternate isoleucine biosynthesis pathway in Geobacter sulfurreducens.}, journal = {J Bacteriol}, volume = {190}, year = {2008}, month = {2008 Apr}, pages = {2266-74}, abstract = {The central metabolic model for Geobacter sulfurreducens included a single pathway for the biosynthesis of isoleucine that was analogous to that of Escherichia coli, in which the isoleucine precursor 2-oxobutanoate is generated from threonine. 13C labeling studies performed in G. sulfurreducens indicated that this pathway accounted for a minor fraction of isoleucine biosynthesis and that the majority of isoleucine was instead derived from acetyl-coenzyme A and pyruvate, possibly via the citramalate pathway. Genes encoding citramalate synthase (GSU1798), which catalyzes the first dedicated step in the citramalate pathway, and threonine ammonia-lyase (GSU0486), which catalyzes the conversion of threonine to 2-oxobutanoate, were identified and knocked out. Mutants lacking both of these enzymes were auxotrophs for isoleucine, whereas single mutants were capable of growth in the absence of isoleucine. Biochemical characterization of the single mutants revealed deficiencies in citramalate synthase and threonine ammonia-lyase activity. Thus, in G. sulfurreducens, 2-oxobutanoate can be synthesized either from citramalate or threonine, with the former being the main pathway for isoleucine biosynthesis. The citramalate synthase of G. sulfurreducens constitutes the first characterized member of a phylogenetically distinct clade of citramalate synthases, which contains representatives from a wide variety of microorganisms.}, keywords = {Acetyl Coenzyme A, Bacterial Proteins, Biosynthetic Pathways, Butyric Acids, Carbon Isotopes, Geobacter, Isoleucine, Malates, Pyruvic Acid, Threonine, Threonine Dehydratase}, issn = {1098-5530}, doi = {10.1128/JB.01841-07}, author = {Risso, Carla and Van Dien, Stephen J and Orloff, Amber and Lovley, Derek R and Coppi, Maddalena V} } @article {485, title = {Extracellular electron transfer: wires, capacitors, iron lungs, and more.}, journal = {Geobiology}, volume = {6}, year = {2008}, month = {2008 Jun}, pages = {225-31}, keywords = {Cell Surface Extensions, Cytochromes, Electron Transport, Ferric Compounds, Models, Biological, Shewanella}, issn = {1472-4669}, doi = {10.1111/j.1472-4669.2008.00148.x}, author = {Lovley, Derek R} } @article {492, title = {Fluorescent properties of c-type cytochromes reveal their potential role as an extracytoplasmic electron sink in Geobacter sulfurreducens.}, journal = {Environ Microbiol}, volume = {10}, year = {2008}, month = {2008 Feb}, pages = {497-505}, abstract = {A novel fluorescence technique for monitoring the redox status of c-type cytochromes in Geobacter sulfurreducens was developed in order to evaluate the capacity of these extracytoplasmic cytochromes to store electrons during periods in which an external electron acceptor is not available. When intact cells in which the cytochromes were in a reduced state were excited at a wavelength of 350 nm, they fluoresced with maxima at 402 and 437 nm. Oxidation of the cytochromes resulted in a loss of fluorescence. This method was much more sensitive than the traditional approach of detecting c-type cytochromes via visible light absorbance. Furthermore, fluorescence of reduced cytochromes in individual cells could be detected via fluorescence microscopy, and the cytochromes in a G. sulfurreducens biofilm, remotely excited with an optical fibre, could be detected at distances as far as 5 cm. Fluorescence analysis of cytochrome oxidation and reduction of the external electron acceptor, anthraquinone-2,6-disulfonate, suggested that the extracytoplasmic cytochromes of G. sulfurreducens could store approximately 10(7) electrons per cell. Independent analysis of the haem content of the cells determined from analysis of incorporation of (55)Fe into cytochromes provided a similar estimate of cytochrome electron-storage capacity. This electron-storage capacity could, in the absence of an external electron acceptor, permit continued electron transfer across the inner membrane sufficient to supply the maintenance energy requirements for G. sulfurreducens for up to 8 min or enough proton motive force to power flagella motors for G. sulfurreducens motility. The fluorescence approach described here provides a sensitive method for evaluating the redox status of Geobacter species in culture and/or its environments. Furthermore, these results suggest that the periplasmic and outer-membrane cytochromes of Geobacter species act as capacitors, allowing continued electron transport, and thus viability and motility, for Geobacter species as they move between heterogeneously dispersed Fe(III) oxides during growth in the subsurface.}, keywords = {Anthraquinones, Cytochromes c, Electron Transport, Ferric Compounds, Flagella, Fluorescence, Geobacter, Microscopy, Fluorescence, Oxidation-Reduction}, issn = {1462-2920}, doi = {10.1111/j.1462-2920.2007.01470.x}, author = {Esteve-N{\'u}{\~n}ez, Abraham and Sosnik, Julian and Visconti, Pablo and Lovley, Derek R} } @article {488, title = {Gene transcript analysis of assimilatory iron limitation in Geobacteraceae during groundwater bioremediation.}, journal = {Environ Microbiol}, volume = {10}, year = {2008}, month = {2008 May}, pages = {1218-30}, abstract = {Limitations on the availability of Fe(III) as an electron acceptor are thought to play an important role in restricting the growth and activity of Geobacter species during bioremediation of contaminated subsurface environments, but the possibility that these organisms might also be limited in the subsurface by the availability of iron for assimilatory purposes was not previously considered because copious quantities of Fe(II) are produced as the result of Fe(III) reduction. Analysis of multiple Geobacteraceae genomes revealed the presence of a three-gene cluster consisting of homologues of two iron-dependent regulators, fur and dtxR (ideR), separated by a homologue of feoB, which encodes an Fe(II) uptake protein. This cluster appears to be conserved among members of the Geobacteraceae and was detected in several environments. Expression of the fur-feoB-ideR cluster decreased as Fe(II) concentrations increased in chemostat cultures. The number of Geobacteraceae feoB transcripts in groundwater samples from a site undergoing in situ uranium bioremediation was relatively high until the concentration of dissolved Fe(II) increased near the end of the field experiment. These results suggest that, because much of the Fe(II) is sequestered in solid phases, Geobacter species, which have a high requirement for iron for iron-sulfur proteins, may be limited by the amount of iron available for assimilatory purposes. These results demonstrate the ability of transcript analysis to reveal previously unsuspected aspects of the in situ physiology of microorganisms in subsurface environments.}, keywords = {Bacterial Proteins, Biodegradation, Environmental, Culture Media, Ferric Compounds, Ferrous Compounds, Fresh Water, Gene Expression Regulation, Bacterial, Geobacter, Iron, Multigene Family, Phylogeny, Polymerase Chain Reaction, Repressor Proteins, Reverse Transcriptase Polymerase Chain Reaction, Transcription, Genetic, Uranium, Water Pollution, Radioactive}, issn = {1462-2920}, doi = {10.1111/j.1462-2920.2007.01537.x}, author = {O{\textquoteright}Neil, Regina A and Holmes, Dawn E and Coppi, Maddalena V and Adams, Lorrie A and Larrahondo, M Juliana and Ward, Joy E and Nevin, Kelly P and Woodard, Trevor L and Vrionis, Helen A and N{\textquoteright}guessan, Lucie A and Lovley, Derek R} } @article {483, title = {Genes for two multicopper proteins required for Fe(III) oxide reduction in Geobacter sulfurreducens have different expression patterns both in the subsurface and on energy-harvesting electrodes.}, journal = {Microbiology}, volume = {154}, year = {2008}, month = {2008 May}, pages = {1422-35}, abstract = {Previous studies have shown that Geobacter sulfurreducens requires the outer-membrane, multicopper protein OmpB for Fe(III) oxide reduction. A homologue of OmpB, designated OmpC, which is 36 \% similar to OmpB, has been discovered in the G. sulfurreducens genome. Deletion of ompC inhibited reduction of insoluble, but not soluble Fe(III). Analysis of multiple Geobacter and Pelobacter genomes, as well as in situ Geobacter, indicated that genes encoding multicopper proteins are conserved in Geobacter species but are not found in Pelobacter species. Levels of ompB transcripts were similar in G. sulfurreducens at different growth rates in chemostats and during growth on a microbial fuel cell anode. In contrast, ompC transcript levels increased at higher growth rates in chemostats and with increasing current production in fuel cells. Constant levels of Geobacter ompB transcripts were detected in groundwater during a field experiment in which acetate was added to the subsurface to promote in situ uranium bioremediation. In contrast, ompC transcript levels increased during the rapid phase of growth of Geobacter species following addition of acetate to the groundwater and then rapidly declined. These results demonstrate that more than one multicopper protein is required for optimal Fe(III) oxide reduction in G. sulfurreducens and suggest that, in environmental studies, quantifying OmpB/OmpC-related genes could help alleviate the problem that Pelobacter genes may be inadvertently quantified via quantitative analysis of 16S rRNA genes. Furthermore, comparison of differential expression of ompB and ompC may provide insight into the in situ metabolic state of Geobacter species in environments of interest.}, keywords = {Acetates, Amino Acid Sequence, Bacterial Outer Membrane Proteins, Electrodes, Ferric Compounds, Gene Deletion, Gene Expression Profiling, Geobacter, Molecular Sequence Data, Oxidation-Reduction, Phylogeny, Sequence Alignment, Sequence Homology, Nucleic Acid, Soil Microbiology, Uranium}, issn = {1350-0872}, doi = {10.1099/mic.0.2007/014365-0}, author = {Holmes, Dawn E and Mester, T{\"u}nde and O{\textquoteright}Neil, Regina A and Perpetua, Lorrie A and Larrahondo, M Juliana and Glaven, Richard and Sharma, Manju L and Ward, Joy E and Nevin, Kelly P and Lovley, Derek R} } @article {481, title = {Genome-wide gene expression patterns and growth requirements suggest that Pelobacter carbinolicus reduces Fe(III) indirectly via sulfide production.}, journal = {Appl Environ Microbiol}, volume = {74}, year = {2008}, month = {2008 Jul}, pages = {4277-84}, abstract = {Although Pelobacter species are closely related to Geobacter species, recent studies suggested that Pelobacter carbinolicus may reduce Fe(III) via a different mechanism because it lacks the outer-surface c-type cytochromes that are required for Fe(III) reduction by Geobacter sulfurreducens. Investigation into the mechanisms for Fe(III) reduction demonstrated that P. carbinolicus had growth yields on both soluble and insoluble Fe(III) consistent with those of other Fe(III)-reducing bacteria. Comparison of whole-genome transcript levels during growth on Fe(III) versus fermentative growth demonstrated that the greatest apparent change in gene expression was an increase in transcript levels for four contiguous genes. These genes encode two putative periplasmic thioredoxins; a putative outer-membrane transport protein; and a putative NAD(FAD)-dependent dehydrogenase with homology to disulfide oxidoreductases in the N terminus, rhodanese (sulfurtransferase) in the center, and uncharacterized conserved proteins in the C terminus. Unlike G. sulfurreducens, transcript levels for cytochrome genes did not increase in P. carbinolicus during growth on Fe(III). P. carbinolicus could use sulfate as the sole source of sulfur during fermentative growth, but required elemental sulfur or sulfide for growth on Fe(III). The increased expression of genes potentially involved in sulfur reduction, coupled with the requirement for sulfur or sulfide during growth on Fe(III), suggests that P. carbinolicus reduces Fe(III) via an indirect mechanism in which (i) elemental sulfur is reduced to sulfide and (ii) the sulfide reduces Fe(III) with the regeneration of elemental sulfur. This contrasts with the direct reduction of Fe(III) that has been proposed for Geobacter species.}, keywords = {Acetoin, Cytochrome c Group, Deltaproteobacteria, Ethanol, Fermentation, Ferric Compounds, Gene Expression Profiling, Genome, Bacterial, Iron, Nitrilotriacetic Acid, Oligonucleotide Array Sequence Analysis, Oxidation-Reduction, Reverse Transcriptase Polymerase Chain Reaction, RNA, Bacterial, Substrate Specificity, Sulfides, Sulfur, Sulfur-Reducing Bacteria, Thioredoxins}, issn = {1098-5336}, doi = {10.1128/AEM.02901-07}, author = {Haveman, Shelley A and Didonato, Raymond J and Villanueva, Laura and Shelobolina, Evgenya S and Postier, Bradley L and Xu, Bo and Liu, Anna and Lovley, Derek R} } @article {476, title = {Geobacter sulfurreducens strain engineered for increased rates of respiration.}, journal = {Metab Eng}, volume = {10}, year = {2008}, month = {2008 Sep}, pages = {267-75}, abstract = {Geobacter species are among the most effective microorganisms known for the bioremediation of radioactive and toxic metals in contaminated subsurface environments and for converting organic compounds to electricity in microbial fuel cells. However, faster rates of electron transfer could aid in optimizing these processes. Therefore, the Optknock strain design methodology was applied in an iterative manner to the constraint-based, in silico model of Geobacter sulfurreducens to identify gene deletions predicted to increase respiration rates. The common factor in the Optknock predictions was that each resulted in a predicted increase in the cellular ATP demand, either by creating ATP-consuming futile cycles or decreasing the availability of reducing equivalents and inorganic phosphate for ATP biosynthesis. The in silico model predicted that increasing the ATP demand would result in higher fluxes of acetate through the TCA cycle and higher rates of NADPH oxidation coupled with decreases in flux in reactions that funnel acetate toward biosynthetic pathways. A strain of G. sulfurreducens was constructed in which the hydrolytic, F(1) portion of the membrane-bound F(0)F(1) (H(+))-ATP synthase complex was expressed when IPTG was added to the medium. Induction of the ATP drain decreased the ATP content of the cell by more than half. The cells with the ATP drain had higher rates of respiration, slower growth rates, and a lower cell yield. Genome-wide analysis of gene transcript levels indicated that when the higher rate of respiration was induced transcript levels were higher for genes involved in energy metabolism, especially in those encoding TCA cycle enzymes, subunits of the NADH dehydrogenase, and proteins involved in electron acceptor reduction. This was accompanied by lower transcript levels for genes encoding proteins involved in amino acid biosynthesis, cell growth, and motility. Several changes in gene expression that involve processes not included in the in silico model were also detected, including increased expression of a number of redox-active proteins, such as c-type cytochromes and a putative multicopper outer-surface protein. The results demonstrate that it is possible to genetically engineer increased respiration rates in G. sulfurreducens in accordance with predictions from in silico metabolic modeling. To our knowledge, this is the first report of metabolic engineering to increase the respiratory rate of a microorganism.}, keywords = {Adenosine Triphosphate, Bacterial Proteins, Biodegradation, Environmental, Citric Acid Cycle, Electron Transport, Geobacter, Metals, Models, Biological, NADH Dehydrogenase, NADP, Oxygen Consumption, Phosphates, Proton-Translocating ATPases, Radioactive Pollutants}, issn = {1096-7184}, doi = {10.1016/j.ymben.2008.06.005}, author = {Izallalen, Mounir and Mahadevan, Radhakrishnan and Burgard, Anthony and Postier, Bradley and DiDonato, Raymond and Sun, Jun and Schilling, Christopher H and Lovley, Derek R} } @article {484, title = {Geobacter uraniireducens sp. nov., isolated from subsurface sediment undergoing uranium bioremediation.}, journal = {Int J Syst Evol Microbiol}, volume = {58}, year = {2008}, month = {2008 May}, pages = {1075-8}, abstract = {A Gram-negative, rod-shaped, motile bacterium, strain Rf4T, which conserves energy from dissimilatory Fe(III) reduction concomitant with acetate oxidation, was isolated from subsurface sediment undergoing uranium bioremediation. The 16S rRNA gene sequence of strain Rf4T matched sequences recovered in 16S rRNA gene clone libraries constructed from DNA extracted from groundwater sampled at the same time as the source sediment. Cells of strain Rf4T were regular, motile rods, 1.2-2.0 microm long and 0.5-0.6 microm in diameter, with rounded ends. Cells had one lateral flagellum. Growth was optimal at pH 6.5-7.0 and 32 degrees C. With acetate as the electron donor, strain Rf4T used Fe(III), Mn(IV), anthraquinone-2,6-disulfonate, malate and fumarate as electron acceptors and reduced U(VI) in cell suspensions. With poorly crystalline Fe(III) oxide as the electron acceptor, strain Rf4T oxidized the following electron donors: acetate, lactate, pyruvate and ethanol. Phylogenetic analysis of the 16S rRNA gene sequence of strain Rf4T placed it in the genus Geobacter. Strain Rf4T was most closely related to {\textquoteright}Geobacter humireducens{\textquoteright} JW3 (95.9 \% sequence similarity), Geobacter bremensis Dfr1T (95.4 \%) and Geobacter bemidjiensis BemT (95.1 \%). Based on phylogenetic analysis and phenotypic differences between strain Rf4T and closely related Geobacter species, this strain is described as a representative of a novel species, Geobacter uraniireducens sp. nov. The type strain is Rf4T (=ATCC BAA-1134T =JCM 13001T).}, keywords = {Bacterial Typing Techniques, DNA, Bacterial, Genes, rRNA, Genotype, Geobacter, Geologic Sediments, Molecular Sequence Data, Oxidation-Reduction, Phenotype, Phylogeny, RNA, Ribosomal, 16S, Sequence Analysis, DNA, Species Specificity, Uranium}, issn = {1466-5026}, doi = {10.1099/ijs.0.65377-0}, author = {Shelobolina, Evgenya S and Vrionis, Helen A and Findlay, Robert H and Lovley, Derek R} } @article {475, title = {Graphite electrode as a sole electron donor for reductive dechlorination of tetrachlorethene by Geobacter lovleyi.}, journal = {Appl Environ Microbiol}, volume = {74}, year = {2008}, month = {2008 Oct}, pages = {5943-7}, abstract = {The possibility that graphite electrodes can serve as the direct electron donor for microbially catalyzed reductive dechlorination was investigated with Geobacter lovleyi. In an initial evaluation of whether G. lovleyi could interact electronically with graphite electrodes, cells were provided with acetate as the electron donor and an electrode as the sole electron acceptor. Current was produced at levels that were ca. 10-fold lower than those previously reported for Geobacter sulfurreducens under similar conditions, and G. lovleyi anode biofilms were correspondingly thinner. When an electrode poised at -300 mV (versus a standard hydrogen electrode) was provided as the electron donor, G. lovleyi effectively reduced fumarate to succinate. The stoichiometry of electrons consumed to succinate produced was 2:1, the ratio expected if the electrode served as the sole electron donor for fumarate reduction. G. lovleyi effectively reduced tetrachloroethene (PCE) to cis-dichloroethene with a poised electrode as the sole electron donor at rates comparable to those obtained when acetate serves as the electron donor. Cells were less abundant on the electrodes when the electrodes served as an electron donor than when they served as an electron acceptor. PCE was not reduced in controls without cells or when the current supply to cells was interrupted. These results demonstrate that G. lovleyi can use a poised electrode as a direct electron donor for reductive dechlorination of PCE. The ability to colocalize dechlorinating microorganisms with electrodes has several potential advantages for bioremediation of subsurface chlorinated contaminants, especially in source zones where electron donor delivery is challenging and often limits dechlorination.}, keywords = {Acetic Acid, Biofilms, Biomass, Electricity, Electrodes, Electrons, Ethylene Dichlorides, Fumarates, Geobacter, Graphite, Microscopy, Electron, Scanning, Succinic Acid, Tetrachloroethylene}, issn = {1098-5336}, doi = {10.1128/AEM.00961-08}, author = {Strycharz, Sarah M and Woodard, Trevor L and Johnson, Jessica P and Nevin, Kelly P and Sanford, Robert A and L{\"o}ffler, Frank E and Lovley, Derek R} } @article {497, title = {Growth of thermophilic and hyperthermophilic Fe(III)-reducing microorganisms on a ferruginous smectite as the sole electron acceptor.}, journal = {Appl Environ Microbiol}, volume = {74}, year = {2008}, month = {2008 Jan}, pages = {251-8}, abstract = {Recent studies have suggested that the structural Fe(III) within phyllosilicate minerals, including smectite and illite, is an important electron acceptor for Fe(III)-reducing microorganisms in sedimentary environments at moderate temperatures. The reduction of structural Fe(III) by thermophiles, however, has not previously been described. A wide range of thermophilic and hyperthermophilic Archaea and Bacteria from marine and freshwater environments that are known to reduce poorly crystalline Fe(III) oxides were tested for their ability to reduce structural (octahedrally coordinated) Fe(III) in smectite (SWa-1) as the sole electron acceptor. Two out of the 10 organisms tested, Geoglobus ahangari and Geothermobacterium ferrireducens, were not able to conserve energy to support growth by reduction of Fe(III) in SWa-1 despite the fact that both organisms were originally isolated with solid-phase Fe(III) as the electron acceptor. The other organisms tested were able to grow on SWa-1 and reduced 6.3 to 15.1\% of the Fe(III). This is 20 to 50\% less than the reported amounts of Fe(III) reduced in the same smectite (SWa-1) by mesophilic Fe(III) reducers. Two organisms, Geothermobacter ehrlichii and archaeal strain 140, produced copious amounts of an exopolysaccharide material, which may have played an active role in the dissolution of the structural iron in SWa-1 smectite. The reduction of structural Fe(III) in SWa-1 by archaeal strain 140 was studied in detail. Microbial Fe(III) reduction was accompanied by an increase in interlayer and octahedral charges and some incorporation of potassium and magnesium into the smectite structure. However, these changes in the major element chemistry of SWa-1 smectite did not result in the formation of an illite-like structure, as reported for a mesophilic Fe(III) reducer. These results suggest that thermophilic Fe(III)-reducing organisms differ in their ability to reduce and solubilize structural Fe(III) in SWa-1 smectite and that SWa-1 is not easily transformed to illite by these organisms.}, keywords = {Archaea, Bacteria, Ferric Compounds, Geologic Sediments, Hot Temperature, Oxidation-Reduction, Silicates}, issn = {1098-5336}, doi = {10.1128/AEM.01580-07}, author = {Kashefi, Kazem and Shelobolina, Evgenya S and Elliott, W Crawford and Lovley, Derek R} } @article {494, title = {Growth with high planktonic biomass in Shewanella oneidensis fuel cells.}, journal = {FEMS Microbiol Lett}, volume = {278}, year = {2008}, month = {2008 Jan}, pages = {29-35}, abstract = {Shewanella oneidensis MR-1 grew for over 50 days in microbial fuel cells, incompletely oxidizing lactate to acetate with high recovery of the electrons derived from this reaction as electricity. Electricity was produced with lactate or hydrogen and current was comparable to that of electricigens which completely oxidize organic substrates. However, unlike fuel cells with previously described electricigens, in which cells are primarily attached to the anode, at least as many of the S. oneidensis cells were planktonic as were attached to the anode. These results demonstrate that S. oneidensis may conserve energy for growth with an electrode serving as an electron acceptor and suggest that multiple strategies for electron transfer to fuel cell anodes exist.}, keywords = {Bioelectric Energy Sources, Electricity, Electron Transport, Lactic Acid, Plankton, Shewanella}, issn = {0378-1097}, doi = {10.1111/j.1574-6968.2007.00964.x}, author = {Lanthier, Martin and Gregory, Kelvin B and Lovley, Derek R} } @article {472, title = {Highly conserved genes in Geobacter species with expression patterns indicative of acetate limitation.}, journal = {Microbiology}, volume = {154}, year = {2008}, month = {2008 Sep}, pages = {2589-99}, abstract = {Analysis of the genome of Geobacter sulfurreducens revealed four genes encoding putative symporters with homology to ActP, an acetate transporter in Escherichia coli. Three of these genes, aplA, aplB and aplC, are highly similar (over 90 \% identical) and fell within a tight phylogenetic cluster (Group I) consisting entirely of Geobacter homologues. Transcript levels for all three genes increased in response to acetate limitation. The fourth gene, aplD, is phylogenetically distinct (Group II) and its expression was not influenced by acetate availability. Deletion of any one of the three genes in Group I did not significantly affect acetate-dependent growth, suggesting functional redundancy. Attempts to recover mutants in which various combinations of two of these genes were deleted were unsuccessful, suggesting that at least two of these three transporter genes are required to support growth. Closely related Group I apl genes were found in the genomes of other Geobacter species whose genome sequences are available. Furthermore, related genes could be detected in genomic DNA extracted from a subsurface environment undergoing in situ uranium bioremediation. The transporter genes recovered from the subsurface were most closely related to Group I apl genes found in the genomes of cultured Geobacter species that were isolated from contaminated subsurface environments. The increased expression of these genes in response to acetate limitation, their high degree of conservation among Geobacter species and the ease with which they can be detected in environmental samples suggest that Group I apl genes of the Geobacteraceae may be suitable biomarkers for acetate limitation. Monitoring the expression of these genes could aid in the design of strategies for acetate-mediated in situ bioremediation of uranium-contaminated groundwater.}, keywords = {Acetates, Biodegradation, Environmental, DNA, Bacterial, Escherichia coli, Escherichia coli Proteins, Gene Deletion, Gene Expression, Genes, Bacterial, Genome, Bacterial, Geobacter, Membrane Transport Proteins, Phylogeny, Uranium}, issn = {1350-0872}, doi = {10.1099/mic.0.2008/017244-0}, author = {Risso, Carla and Meth{\'e}, Barbara A and Elifantz, Hila and Holmes, Dawn E and Lovley, Derek R} } @article {394, title = {Inducible protein degradation in Bacillus subtilis using heterologous peptide tags and adaptor proteins to target substrates to the protease ClpXP.}, journal = {Mol Microbiol}, volume = {70}, year = {2008}, month = {2008 Nov}, pages = {1012-25}, abstract = {The ability to manipulate protein levels is useful for dissecting regulatory pathways, elucidating gene function and constructing synthetic biological circuits. We engineered an inducible protein degradation system for use in Bacillus subtilis based on Escherichia coli and Caulobacter crescentusssrA tags and SspB adaptors that deliver proteins to ClpXP for proteolysis. In this system, modified ssrA degradation tags are fused onto the 3{\textquoteright} end of the genes of interest. Unlike wild-type ssrA, these modified tags require the adaptor protein SspB to target tagged proteins for proteolysis. In the absence of SspB, the tagged proteins accumulate to near physiological levels. By inducing SspB expression from a regulated promoter, the tagged substrates are rapidly delivered to the B. subtilis ClpXP protease for degradation. We used this system to degrade the reporter GFP and several native B. subtilis proteins, including, the transcription factor ComA, two sporulation kinases (KinA, KinB) and the sporulation and chromosome partitioning protein Spo0J. We also used modified E. coli and C. crescentus ssrA tags to independently control the degradation of two different proteins in the same cell. These tools will be useful for studying biological processes in B. subtilis and can potentially be modified for use in other bacteria.}, keywords = {Alleles, Bacillus subtilis, Bacterial Proteins, Carrier Proteins, Caulobacter crescentus, DNA-Binding Proteins, Endopeptidase Clp, Escherichia coli, Escherichia coli Proteins, Gene Expression Regulation, Bacterial, Genetic Vectors, Green Fluorescent Proteins, Plasmids, Promoter Regions, Genetic, Protein Engineering, RNA, Bacterial, Substrate Specificity}, issn = {1365-2958}, doi = {10.1111/j.1365-2958.2008.06467.x}, author = {Griffith, Kevin L and Grossman, Alan D} } @article {480, title = {Insights into genes involved in electricity generation in Geobacter sulfurreducens via whole genome microarray analysis of the OmcF-deficient mutant.}, journal = {Bioelectrochemistry}, volume = {73}, year = {2008}, month = {2008 Jun}, pages = {70-5}, abstract = {Geobacter sulfurreducens effectively produces electricity in microbial fuel cells by oxidizing acetate with an electrode serving as the sole electron acceptor. Deletion of the gene encoding OmcF, a monoheme outer membrane c-type cytochrome, substantially decreased current production. Previous studies demonstrated that inhibition of Fe(III) reduction in the OmcF-deficient mutant could be attributed to poor transcription of the gene for OmcB, an outer membrane c-type cytochrome that is required for Fe(III) reduction. However, a mutant in which omcB was deleted produced electricity as well as wild type. Microarray analysis of the OmcF-deficient mutant versus the wild type revealed that many of the genes with the greatest decreases in transcript levels were genes whose expression was previously reported to be upregulated in cells grown with an electrode as the sole electron acceptor. These included genes with putative functions related to metal efflux and/or type I secretion and two hypothetical proteins. The outer membrane cytochromes, OmcS and OmcE, which previous studies have demonstrated are required for optimal current generation, were not detected on the outer surface of the OmcF-deficient mutant even though the omcS and omcE genes were still transcribed, suggesting that the putative secretion system could be involved in the export of outer membrane proteins necessary for electron transfer to the fuel cell anode. These results suggest that the requirement for OmcF for optimal current production is not because OmcF is directly involved in extracellular electron transfer but because OmcF is required for the appropriate transcription of other genes either directly or indirectly involved in electricity production.}, keywords = {Bacterial Outer Membrane Proteins, Cytochromes c, Down-Regulation, Electricity, Gene Expression Regulation, Bacterial, Genome, Bacterial, Geobacter, Mutation, Oligonucleotide Array Sequence Analysis, Transcription, Genetic}, issn = {1567-5394}, doi = {10.1016/j.bioelechem.2008.04.023}, author = {Kim, Byoung-Chan and Postier, Bradley L and Didonato, Raymond J and Chaudhuri, Swades K and Nevin, Kelly P and Lovley, Derek R} } @article {741, title = {Investigating the effect of increasing charge density on the hemolytic activity of synthetic antimicrobial polymers.}, journal = {Biomacromolecules}, volume = {9}, year = {2008}, month = {2008 Oct}, pages = {2805-10}, abstract = {The current study is aimed at investigating the effect of fine-tuning the cationic character of synthetic mimics of antimicrobial peptides (SMAMPs) on the hemolytic and antibacterial activities. A series of novel norbornene monomers that carry one, two, or three Boc-protected amine functionalities was prepared. Ring-opening metathesis polymerization (ROMP) of the monomers, followed by deprotection of the amine groups resulted in cationic antimicrobial polynorbornenes that carry one, two, and three charges per monomer repeat unit. Increasing the number of amine groups on the most hydrophobic polymer reduced its hemolytic activity significantly. To understand the membrane activity of these polymers, we conducted dye leakage experiments on lipid vesicles that mimic bacteria and red blood cell membranes, and these results showed a strong correlation with the hemolysis data.}, keywords = {Amines, Anti-Infective Agents, Antimicrobial Cationic Peptides, Chemistry, Pharmaceutical, Drug Design, Erythrocytes, Escherichia coli, Hemolysis, Humans, Lipids, Magnetic Resonance Spectroscopy, Models, Chemical, Peptides, Polymers, Staphylococcus aureus}, issn = {1526-4602}, doi = {10.1021/bm800569x}, author = {Al-Badri, Zoha M and Som, Abhigyan and Lyon, Sarah and Nelson, Christopher F and N{\"u}sslein, Klaus and Tew, Gregory N} } @article {478, title = {Investigation of direct vs. indirect involvement of the c-type cytochrome MacA in Fe(III) reduction by Geobacter sulfurreducens.}, journal = {FEMS Microbiol Lett}, volume = {286}, year = {2008}, month = {2008 Sep}, pages = {39-44}, abstract = {The electron transfer pathway to Fe(III) reduction in Geobacter sulfurreducens has been hypothesized to consist of a series of c-type cytochromes. Previous genetic studies suggested that the inner membrane-associated, c-type cytochrome, MacA, was a component of the electron transfer chain leading to Fe(III) reduction in the dissimilatory Fe(III)-reducer, G. sulfurreducens. However, investigation of the expression of OmcB, an outer-membrane c-type cytochrome demonstrated previously to be critical for optimal Fe(III) reduction, revealed that both omcB transcript and protein levels were dramatically reduced in the MacA-deficient mutant. Expression of the omcB gene in trans enabled the MacA-deficient mutant to reduce Fe(III) at a rate that was proportional to the level of omcB expression. These results suggest that MacA is not directly involved in electron transfer to Fe(III) and further confirm the importance of OmcB in Fe(III) reduction by G. sulfurreducens.}, keywords = {ATP-Binding Cassette Transporters, Bacterial Proteins, Cytochromes c, Ferric Compounds, Gene Expression Regulation, Bacterial, Geobacter, Oxidation-Reduction}, issn = {0378-1097}, doi = {10.1111/j.1574-6968.2008.01252.x}, author = {Kim, Byoung-Chan and Lovley, Derek R} } @article {468, title = {The microbe electric: conversion of organic matter to electricity.}, journal = {Curr Opin Biotechnol}, volume = {19}, year = {2008}, month = {2008 Dec}, pages = {564-71}, abstract = {Broad application of microbial fuel cells will require substantial increases in current density. A better understanding of the microbiology of these systems may help. Recent studies have greatly expanded the range of microorganisms known to function either as electrode-reducing microorganisms at the anode or as electrode-oxidizing microorganisms at the cathode. Microorganisms that can completely oxidize organic compounds with an electrode serving as the sole electron acceptor are expected to be the primary contributors to power production. Several mechanisms for electron transfer to anodes have been proposed including: direct electron transfer via outer-surface c-type cytochromes, long-range electron transfer via microbial nanowires, electron flow through a conductive biofilm matrix containing cytochromes, and soluble electron shuttles. Which mechanisms are most important depend on the microorganisms and the thickness of the anode biofilm. Emerging systems biology approaches to the study, design, and evolution of microorganisms interacting with electrodes are expected to contribute to improved microbial fuel cells.}, keywords = {Bacteria, Bioelectric Energy Sources, Electricity, Electrodes, Organic Chemicals, Oxidation-Reduction}, issn = {1879-0429}, doi = {10.1016/j.copbio.2008.10.005}, author = {Lovley, Derek R} } @article {691, title = {Mutations in pimE restore lipoarabinomannan synthesis and growth in a Mycobacterium smegmatis lpqW mutant.}, journal = {J Bacteriol}, volume = {190}, year = {2008}, month = {2008 May}, pages = {3690-9}, abstract = {Lipoarabinomannans (LAMs) and phosphatidylinositol mannosides (PIMs) are abundant glycolipids in the cell walls of all corynebacteria and mycobacteria, including the devastating human pathogen Mycobacterium tuberculosis. We have recently shown that M. smegmatis mutants of the lipoprotein-encoding lpqW gene have a profound defect in LAM biosynthesis. When these mutants are cultured in complex medium, spontaneous bypass mutants consistently evolve in which LAM biosynthesis is restored at the expense of polar PIM synthesis. Here we show that restoration of LAM biosynthesis in the lpqW mutant results from secondary mutations in the pimE gene. PimE is a mannosyltransferase involved in converting AcPIM4, a proposed branch point intermediate in the PIM and LAM biosynthetic pathways, to more polar PIMs. Mutations in pimE arose due to insertion of the mobile genetic element ISMsm1 and independent point mutations that were clustered in predicted extracytoplasmic loops of this polytopic membrane protein. Our findings provide the first strong evidence that LpqW is required to channel intermediates such as AcPIM4 into LAM synthesis and that loss of PimE function results in the accumulation of AcPIM4, bypassing the need for LpqW. These data highlight new mechanisms regulating the biosynthetic pathways of these essential cell wall components.}, keywords = {Cell Wall, Lipopolysaccharides, Lipoproteins, Mannosyltransferases, Mutation, Mycobacterium smegmatis, Phosphatidylinositols}, issn = {1098-5530}, doi = {10.1128/JB.00200-08}, author = {Crellin, Paul K and Kovacevic, Svetozar and Martin, Kirstee L and Brammananth, Rajini and Morita, Yasu S and Billman-Jacobe, Helen and McConville, Malcolm J and Coppel, Ross L} } @article {479, title = {Power output and columbic efficiencies from biofilms of Geobacter sulfurreducens comparable to mixed community microbial fuel cells.}, journal = {Environ Microbiol}, volume = {10}, year = {2008}, month = {2008 Oct}, pages = {2505-14}, abstract = {It has been previously noted that mixed communities typically produce more power in microbial fuel cells than pure cultures. If true, this has important implications for the design of microbial fuel cells and for studying the process of electron transfer on anode biofilms. To further evaluate this, Geobacter sulfurreducens was grown with acetate as fuel in a continuous flow {\textquoteright}ministack{\textquoteright} system in which the carbon cloth anode and cathode were positioned in close proximity, and the cation-selective membrane surface area was maximized in order to overcome some of the electrochemical limitations that were inherent in fuel cells previously employed for the study of pure cultures. Reducing the size of the anode in order to eliminate cathode limitation resulted in maximum current and power densities per m(2) of anode surface of 4.56 A m(-2) and 1.88 W m(-2) respectively. Electron recovery as current from acetate oxidation was c. 100\% when oxygen diffusion into the system was minimized. This performance is comparable to the highest levels previously reported for mixed communities in similar microbial fuel cells and slightly higher than the power output of an anaerobic sludge inoculum in the same ministack system. Minimizing the volume of the anode chamber yielded a volumetric power density of 2.15 kW m(-3), which is the highest power density per volume yet reported for a microbial fuel cell. Geobacter sulfurreducens formed relatively uniform biofilms 3-18 mum thick on the carbon cloth anodes. When graphite sticks served as the anode, the current density (3.10 A m(-2)) was somewhat less than with the carbon cloth anodes, but the biofilms were thicker (c. 50 mum) with a more complex pillar and channel structure. These results suggest that the previously observed disparity in power production in pure and mixed culture microbial fuel cell systems can be attributed more to differences in the fuel cell designs than to any inherent superior capability of mixed cultures to produce more power than pure cultures.}, keywords = {Acetic Acid, Biofilms, Carbon, Electricity, Electrodes, Geobacter, Oxidation-Reduction}, issn = {1462-2920}, doi = {10.1111/j.1462-2920.2008.01675.x}, author = {Nevin, K P and Richter, H and Covalla, S F and Johnson, J P and Woodard, T L and Orloff, A L and Jia, H and Zhang, M and Lovley, D R} } @article {477, title = {Proteome of Geobacter sulfurreducens grown with Fe(III) oxide or Fe(III) citrate as the electron acceptor.}, journal = {Biochim Biophys Acta}, volume = {1784}, year = {2008}, month = {2008 Dec}, pages = {1935-41}, abstract = {The mechanisms for Fe(III) oxide reduction in Geobacter species are of interest because Fe(III) oxides are the most abundant form of Fe(III) in many soils and sediments and Geobacter species are prevalent Fe(III)-reducing microorganisms in many of these environments. Protein abundance in G. sulfurreducens grown on poorly crystalline Fe(III) oxide or on soluble Fe(III) citrate was compared with a global accurate mass and time tag proteomic approach in order to identify proteins that might be specifically associated with Fe(III) oxide reduction. A total of 2991 proteins were detected in G. sulfurreducens grown with acetate as the electron donor and either Fe(III) oxide or soluble Fe(III) citrate as the electron acceptor, resulting in 86\% recovery of the genes predicted to encode proteins. Of the total expressed proteins 76\% were less abundant in Fe(III) oxide cultures than in Fe(III) citrate cultures, which is consistent with the overall slower rate of metabolism during growth with an insoluble electron acceptor. A total of 269 proteins were more abundant in Fe(III) oxide-grown cells than in cells grown on Fe(III) citrate. Most of these proteins were in the energy metabolism category: primarily electron transport proteins, including 13 c-type cytochromes and PilA, the structural protein for electrically conductive pili. Several of the cytochromes that were more abundant in Fe(III) oxide-grown cells were previously shown with genetic approaches to be essential for optimal Fe(III) oxide reduction. Other proteins that were more abundant during growth on Fe(III) oxide included transport and binding proteins, proteins involved in regulation and signal transduction, cell envelope proteins, and enzymes for amino acid and protein biosynthesis, among others. There were also a substantial number of proteins of unknown function that were more abundant during growth on Fe(III) oxide. These results indicate that electron transport to Fe(III) oxide requires additional and/or different proteins than electron transfer to soluble, chelated Fe(III) and suggest proteins whose functions should be further investigated in order to better understand the mechanisms of electron transfer to Fe(III) oxide in G. sulfurreducens.}, keywords = {Bacterial Proteins, Ferric Compounds, Gene Expression Regulation, Bacterial, Geobacter, Oxidation-Reduction, Proteome}, issn = {0006-3002}, doi = {10.1016/j.bbapap.2008.06.011}, author = {Ding, Yan-Huai R and Hixson, Kim K and Aklujkar, Muktak A and Lipton, Mary S and Smith, Richard D and Lovley, Derek R and Mester, T{\"u}nde} } @article {680, title = {Proteopedia - a scientific {\textquoteright}wiki{\textquoteright} bridging the rift between three-dimensional structure and function of biomacromolecules.}, journal = {Genome Biol}, volume = {9}, year = {2008}, month = {2008}, pages = {R121}, abstract = {Many scientists lack the background to fully utilize the wealth of solved three-dimensional biomacromolecule structures. Thus, a resource is needed to present structure/function information in a user-friendly manner to a broad scientific audience. Proteopedia http://www.proteopedia.org is an interactive, wiki web-resource whose pages have embedded three-dimensional structures surrounded by descriptive text containing hyperlinks that change the appearance (view, representations, colors, labels) of the adjacent three-dimensional structure to reflect the concept explained in the text.}, keywords = {Databases, Protein, Internet, Models, Molecular, Protein Conformation, Protein Structure, Tertiary, Proteins, Software}, issn = {1465-6914}, doi = {10.1186/gb-2008-9-8-r121}, author = {Hodis, Eran and Prilusky, Jaime and Martz, Eric and Silman, Israel and Moult, John and Sussman, Joel L} } @article {474, title = {Quantification of Desulfovibrio vulgaris dissimilatory sulfite reductase gene expression during electron donor- and electron acceptor-limited growth.}, journal = {Appl Environ Microbiol}, volume = {74}, year = {2008}, month = {2008 Sep}, pages = {5850-3}, abstract = {Previous studies have suggested that levels of transcripts for dsrA, a gene encoding a subunit of the dissimilatory sulfite reductase, are not directly related to the rates of sulfate reduction in sediments under all conditions. This phenomenon was further investigated with chemostat-grown Desulfovibrio vulgaris. Under sulfate-limiting conditions, dsrA transcript levels increased as the bulk rates of sulfate reduction in the chemostat increased, but transcript levels were similar at all sulfate reduction rates under electron donor-limiting conditions. When both electron donor- and electron acceptor-limiting conditions were considered, there was a direct correspondence between dsrA transcript levels and the rates of sulfate reduction per cell. These results suggest that dsrA transcript levels may provide important information on the metabolic state of sulfate reducers.}, keywords = {Desulfovibrio vulgaris, Electron Transport, Gene Expression, Geologic Sediments, Hydrogensulfite Reductase, Oxidation-Reduction, RNA, Bacterial, RNA, Messenger, Sulfates}, issn = {1098-5336}, doi = {10.1128/AEM.00399-08}, author = {Villanueva, Laura and Haveman, Shelley A and Summers, Zara M and Lovley, Derek R} } @article {500, title = {Quantifying expression of a dissimilatory (bi)sulfite reductase gene in petroleum-contaminated marine harbor sediments.}, journal = {Microb Ecol}, volume = {55}, year = {2008}, month = {2008 Apr}, pages = {489-99}, abstract = {The possibility of quantifying in situ levels of transcripts for dissimilatory (bi)sulfite reductase (dsr) genes to track the activity of sulfate-reducing microorganisms in petroleum-contaminated marine harbor sediments was evaluated. Phylogenetic analysis of the cDNA generated from mRNA for a ca. 1.4 kbp portion of the contiguous dsrA and dsrB genes suggested that Desulfosarcina species, closely related to cultures known to anaerobically oxidize aromatic hydrocarbons, were active sulfate reducers in the sediments. The levels of dsrA transcripts (per mug total mRNA) were quantified in sediments incubated anaerobically at the in situ temperature as well as in sediments incubated at higher temperatures and/or with added acetate to increase the rate of sulfate reduction. Levels of dsrA transcripts were low when there was no sulfate reduction because the sediments were depleted of sulfate or if sulfate reduction was inhibited with added molybdate. There was a direct correlation between dsrA transcript levels and rates of sulfate reduction when sulfate was at ca. 10 mM in the various sediment treatments, but it was also apparent that within a given sediment, dsrA levels increased over time as long as sulfate was available, even when sulfate reduction rates did not increase. These results suggest that phylogenetic analysis of dsr transcript sequences may provide insight into the active sulfate reducers in marine sediments and that quantifying levels of dsrA transcripts can indicate whether sulfate reducers are active in particular sediment. Furthermore, it may only be possible to use dsrA transcript levels to compare the relative rates of sulfate reduction in sediments when sulfate concentrations, and possibly other environmental conditions, are comparable.}, keywords = {Anaerobiosis, Desulfitobacterium, DNA, Bacterial, DNA, Ribosomal, Gene Expression, Geologic Sediments, Hydrogensulfite Reductase, Molecular Sequence Data, Petroleum, Phylogeny, RNA, Bacterial, RNA, Messenger, RNA, Ribosomal, 16S, Temperature}, issn = {0095-3628}, doi = {10.1007/s00248-007-9294-2}, author = {Chin, Kuk-Jeong and Sharma, Manju L and Russell, Lyndsey A and O{\textquoteright}Neill, Kathleen R and Lovley, Derek R} } @article {812, title = {RecA-mediated SOS induction requires an extended filament conformation but no ATP hydrolysis.}, journal = {Mol Microbiol}, volume = {69}, year = {2008}, month = {2008 Sep}, pages = {1165-79}, abstract = {The Escherichia coli SOS response to DNA damage is modulated by the RecA protein, a recombinase that forms an extended filament on single-stranded DNA and hydrolyzes ATP. The RecA K72R (recA2201) mutation eliminates the ATPase activity of RecA protein. The mutation also limits the capacity of RecA to form long filaments in the presence of ATP. Strains with this mutation do not undergo SOS induction in vivo. We have combined the K72R variant of RecA with another mutation, RecA E38K (recA730). In vitro, the double mutant RecA E38K/K72R (recA730,2201) mimics the K72R mutant protein in that it has no ATPase activity. The double mutant protein will form long extended filaments on ssDNA and facilitate LexA cleavage almost as well as wild-type, and do so in the presence of ATP. Unlike recA K72R, the recA E38K/K72R double mutant promotes SOS induction in vivo after UV treatment. Thus, SOS induction does not require ATP hydrolysis by the RecA protein, but does require formation of extended RecA filaments. The RecA E38K/K72R protein represents an improved reagent for studies of the function of ATP hydrolysis by RecA in vivo and in vitro.}, keywords = {Adenosine Triphosphate, Amino Acid Substitution, Bacterial Proteins, DNA, Bacterial, DNA, Single-Stranded, Escherichia coli, Escherichia coli Proteins, Hydrolysis, Rec A Recombinases, Serine Endopeptidases, SOS Response (Genetics), Ultraviolet Rays}, issn = {1365-2958}, doi = {10.1111/j.1365-2958.2008.06341.x}, author = {Gruenig, Marielle C and Renzette, Nicholas and Long, Edward and Chitteni-Pattu, Sindhu and Inman, Ross B and Cox, Michael M and Sandler, Steven J} } @article {813, title = {Requirements for ATP binding and hydrolysis in RecA function in Escherichia coli.}, journal = {Mol Microbiol}, volume = {67}, year = {2008}, month = {2008 Mar}, pages = {1347-59}, abstract = {RecA is essential for recombination, DNA repair and SOS induction in Escherichia coli. ATP hydrolysis is known to be important for RecA{\textquoteright}s roles in recombination and DNA repair. In vitro reactions modelling SOS induction minimally require ssDNA and non-hydrolyzable ATP analogues. This predicts that ATP hydrolysis will not be required for SOS induction in vivo. The requirement of ATP binding and hydrolysis for SOS induction in vivo is tested here through the study of recA4159 (K72A) and recA2201 (K72R). RecA4159 is thought to have reduced affinity for ATP. RecA2201 binds, but does not hydrolyse ATP. Neither mutant was able to induce SOS expression after UV irradiation. RecA2201, unlike RecA4159, could form filaments on DNA and storage structures as measured with RecA-GFP. RecA2201 was able to form hybrid filaments and storage structures and was either recessive or dominant to RecA(+), depending on the ratio of the two proteins. RecA4159 was unable to enter RecA(+) filaments on DNA or storage structures and was recessive to RecA(+). It is concluded that ATP hydrolysis is essential for SOS induction. It is proposed that ATP binding is essential for storage structure formation and ability to interact with other RecA proteins in a filament.}, keywords = {Adenosine Triphosphate, Chromosomes, Bacterial, Escherichia coli, Escherichia coli Proteins, Hydrolysis, Mutation, Protein Binding, Rec A Recombinases, Ultraviolet Rays}, issn = {1365-2958}, doi = {10.1111/j.1365-2958.2008.06130.x}, author = {Renzette, Nicholas and Sandler, Steven J} } @article {384, title = {Stem-loop silencing reveals that a third mitochondrial DNA polymerase, POLID, is required for kinetoplast DNA replication in trypanosomes.}, journal = {Eukaryot Cell}, volume = {7}, year = {2008}, month = {2008 Dec}, pages = {2141-6}, abstract = {Kinetoplast DNA (kDNA), the mitochondrial genome of trypanosomes, is a catenated network containing thousands of minicircles and tens of maxicircles. The topological complexity dictates some unusual features including a topoisomerase-mediated release-and-reattachment mechanism for minicircle replication and at least six mitochondrial DNA polymerases (Pols) for kDNA transactions. Previously, we identified four family A DNA Pols from Trypanosoma brucei with similarity to bacterial DNA Pol I and demonstrated that two (POLIB and POLIC) were essential for maintaining the kDNA network, while POLIA was not. Here, we used RNA interference to investigate the function of POLID in procyclic T. brucei. Stem-loop silencing of POLID resulted in growth arrest and the progressive loss of the kDNA network. Additional defects in kDNA replication included a rapid decline in minicircle and maxicircle abundance and a transient accumulation of minicircle replication intermediates before loss of the kDNA network. These results demonstrate that POLID is a third essential DNA Pol required for kDNA replication. While other eukaryotes utilize a single DNA Pol (Pol gamma) for replication of mitochondrial DNA, T. brucei requires at least three to maintain the complex kDNA network.}, keywords = {Animals, DNA Replication, DNA, Kinetoplast, DNA-Directed DNA Polymerase, Mitochondria, Mitochondrial Proteins, Protozoan Proteins, RNA Interference, RNA, Double-Stranded, Trypanosoma brucei brucei}, issn = {1535-9786}, doi = {10.1128/EC.00199-08}, author = {Chandler, Julian and Vandoros, Anthula V and Mozeleski, Brian and Klingbeil, Michele M} } @article {482, title = {Sustained removal of uranium from contaminated groundwater following stimulation of dissimilatory metal reduction.}, journal = {Environ Sci Technol}, volume = {42}, year = {2008}, month = {2008 Apr 15}, pages = {2999-3004}, abstract = {Previous field studies on in situ bioremediation of uranium-contaminated groundwater in an aquifer in Rifle, Colorado identified two distinct phases following the addition of acetate to stimulate microbial respiration. In phase I, Geobacter species are the predominant organisms, Fe(III) is reduced, and microbial reduction of soluble U(VI) to insoluble U(IV) removes uranium from the groundwater. In phase II, Fe(III) is depleted, sulfate is reduced, and sulfate-reducing bacteria predominate. Long-term monitoring revealed an unexpected third phase during which U(VI) removal continues even after acetate additions are stopped. All three of these phases were successfully reproduced in flow-through sediment columns. When sediments from the third phase were heat sterilized, the capacity for U(VI) removal was lost. In the live sediments U(VI) removed from the groundwater was recovered as U(VI) in the sediments. This contrasts to the recovery of U(IV) in sediments resulting from the reduction of U(VI) to U(IV) during the Fe(III) reduction phase in acetate-amended sediments. Analysis of 16S rRNA gene sequences in the sediments in which U(VI) was being adsorbed indicated that members of the Firmicutes were the predominant organisms whereas no Firmicutes sequences were detected in background sediments which did not have the capacity to sorb U(VI), suggesting that the U(VI) adsorption might be due to the presence of these living organisms or at least their intact cell components. This unexpected enhanced adsorption of U(VI) onto sediments following the stimulation of microbial growth in the subsurface may potentially enhance the cost effectiveness of in situ uranium bioremediation.}, keywords = {Acetates, Adsorption, Bacteria, Colorado, Geologic Sediments, Oxidation-Reduction, RNA, Ribosomal, 16S, Sulfates, Uranium, Water Pollutants, Radioactive, Water Supply}, issn = {0013-936X}, author = {N{\textquoteright}guessan, Lucie A and Vrionis, Helen A and Resch, Charles T and Long, Philip E and Lovley, Derek R} } @article {740, title = {Synthetic mimic of antimicrobial peptide with nonmembrane-disrupting antibacterial properties.}, journal = {Biomacromolecules}, volume = {9}, year = {2008}, month = {2008 Nov}, pages = {2980-3}, abstract = {Polyguanidinium oxanorbornene ( PGON) was synthesized from norbornene monomers via ring-opening metathesis polymerization. This polymer was observed to be strongly antibacterial against Gram-negative and Gram-positive bacteria as well as nonhemolytic against human red blood cells. Time-kill studies indicated that this polymer is lethal and not just bacteriostatic. In sharp contrast to previously reported SMAMPs (synthetic mimics of antimicrobial peptides), PGON did not disrupt membranes in vesicle-dye leakage assays and microscopy experiments. The unique biological properties of PGON, in same ways similar to cell-penetrating peptides, strongly encourage the examination of other novel guanidino containing macromolecules as powerful and selective antimicrobial agents.}, keywords = {Anti-Bacterial Agents, Antimicrobial Cationic Peptides, Erythrocytes, Gram-Negative Bacteria, Gram-Positive Bacteria, Guanidines, Humans, Microbial Sensitivity Tests, Molecular Mimicry, Polymers}, issn = {1526-4602}, doi = {10.1021/bm800855t}, author = {Gabriel, Gregory J and Madkour, Ahmad E and Dabkowski, Jeffrey M and Nelson, Christopher F and N{\"u}sslein, Klaus and Tew, Gregory N} } @article {34, title = {Transcription regulation of the Saccharomyces cerevisiae PIS1 gene by inositol and the pleiotropic regulator, Ume6p.}, journal = {Mol Microbiol}, volume = {70}, year = {2008}, month = {2008 Dec}, pages = {1529-39}, abstract = {In Saccharomyces cerevisiae, transcription of most of the phospholipid biosynthetic genes (e.g. INO1, CHO1, CHO2 and OPI3) is repressed by growth in the presence of inositol and choline and derepressed in their absence. This regulation requires the Ino2p and Ino4p activators and the Opi1p repressor. The PIS1 structural gene is required for the synthesis of the essential lipid phosphatidylinositol. Previous reports show that PIS1 expression is uncoupled from inositol/choline regulation, but is regulated by carbon source, hypoxia and zinc. However, in this study we found that the expression of PIS1 is induced twofold by inositol. This regulation did not require Ino2p and Ino4p, although Ino4p was required for full expression. Ino4p is a basic helix-loop-helix protein that requires a binding partner. Curiously, none of the other basic helix-loop-helix proteins affected PIS1 expression. Inositol induction did require another general regulator of phospholipid biosynthesis, Ume6p. Ume6p was found to be a positive regulator of PIS1 gene expression. Ume6p, and several associated factors, were required for inositol-mediated induction and chromatin immunoprecipitation analysis showed that Ume6p directly regulates PIS1 expression. Thus, we demonstrate novel regulation of the PIS1 gene by Ume6p.}, keywords = {Chromatin Immunoprecipitation, DNA-Binding Proteins, Gene Expression Regulation, Fungal, Histone Deacetylases, Inositol, Repressor Proteins, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins, Transcription, Genetic, Transferases (Other Substituted Phosphate Groups)}, issn = {1365-2958}, doi = {10.1111/j.1365-2958.2008.06506.x}, author = {Jani, Niketa M and Lopes, John M} } @article {814, title = {XthA (Exonuclease III) regulates loading of RecA onto DNA substrates in log phase Escherichia coli cells.}, journal = {Mol Microbiol}, volume = {67}, year = {2008}, month = {2008 Jan}, pages = {88-101}, abstract = {Exonucleases can modify DNA substrates created during DNA replication, recombination and repair. In Escherichia coli, the effects of several 3{\textquoteright}-5{\textquoteright} exonucleases on RecA loading were studied by assaying RecA-GFP foci formation. Mutations in xthA (ExoIII), xseAB (ExoVII), xni (ExoIX), exoX (ExoX) and tatD (ExoXI) increased the number of RecA-GFP foci twofold to threefold in a population of log phase cells grown in minimal medium. These increases depend on xonA. Epistasis analysis shows that ExoVII, ExoX, ExoIX and ExoXI function in a common pathway, distinct from ExoIII (and ExoI is upstream of both pathways). It is shown (paradoxically) that in xthA mutants, RecA-GFP loading is predominantly RecBCD-dependent and that xthA recB double mutants are viable. Experiments show that while log phase xthA cells have twofold more double-stranded breaks (DSBs) than wild type, they do not induce the SOS response. The increase in RecA loading is independent of the base excision repair (BER) proteins Nth, MutM and Nei. It is proposed that log phase cells produce DSBs that do not induce the SOS response. Furthermore, ExoI, ExoIII and the other 3{\textquoteright}-5{\textquoteright} exonucleases process these DSBs, antagonizing the RecBCD pathway of RecA loading, thus regulating the availability of these substrates for recombination.}, keywords = {DNA Breaks, Double-Stranded, DNA Repair, DNA, Bacterial, Epistasis, Genetic, Escherichia coli K12, Escherichia coli Proteins, Exodeoxyribonucleases, Green Fluorescent Proteins, Microbial Viability, Rec A Recombinases, Recombinant Fusion Proteins, SOS Response (Genetics)}, issn = {0950-382X}, doi = {10.1111/j.1365-2958.2007.06026.x}, author = {Centore, Richard C and Lestini, Roxane and Sandler, Steven J} } @article {743, title = {Arsenic sequestration by nitrate respiring microbial communities in urban lake sediments.}, journal = {Chemosphere}, volume = {70}, year = {2007}, month = {2007 Dec}, pages = {329-36}, abstract = {Changes in microbial community composition and activity were related to geochemical conditions favoring arsenic sequestration in sediments collected from the urban, arsenic-contaminated Upper Mystic Lake. After amendment with nitrate, >94\% total soluble arsenic is sequestered by Fe(III)-(oxy)hydroxides generated in live sediments. Of this sequestered arsenic, >75\% existed as As(III), indicating As redox state alone is not responsible for changes in mobility. Arsenic sequestration was concurrent with the microbial respiration of nitrate as indicated by steady state hydrogen concentration and the presence of organisms similar to nitrate-reducing, iron-oxidizing bacteria belonging to the genus Dechloromonas in 16S rDNA clone libraries.}, keywords = {Arsenic, Betaproteobacteria, Biodegradation, Environmental, DNA, Bacterial, DNA, Ribosomal, Fresh Water, Geologic Sediments, Massachusetts, Nitrates, Oxidation-Reduction, RNA, Ribosomal, 16S, Urbanization, Water Microbiology, Water Pollutants, Chemical}, issn = {0045-6535}, doi = {10.1016/j.chemosphere.2007.05.094}, author = {Gibney, Brian P and N{\"u}sslein, Klaus} } @article {1212, title = {Characterization of malate dehydrogenase from the hyperthermophilic archaeon Pyrobaculum islandicum.}, journal = {Extremophiles}, volume = {11}, year = {2007}, month = {2007 Sep}, pages = {741-6}, abstract = {Native and recombinant malate dehydrogenase (MDH) was characterized from the hyperthermophilic, facultatively autotrophic archaeon Pyrobaculum islandicum. The enzyme is a homotetramer with a subunit mass of 33 kDa. The activity kinetics of the native and recombinant proteins are the same. The apparent K ( m ) values of the recombinant protein for oxaloacetate (OAA) and NADH (at 80 degrees C and pH 8.0) were 15 and 86 microM, respectively, with specific activity as high as 470 U mg(-1). Activity decreased more than 90\% when NADPH was used. The catalytic efficiency of OAA reduction by P. islandicum MDH using NADH was significantly higher than that reported for any other archaeal MDH. Unlike other archaeal MDHs, specific activity of the P. islandicum MDH back-reaction also decreased more than 90\% when malate and NAD(+) were used as substrates and was not detected with NADP(+). A phylogenetic tree of 31 archaeal MDHs shows that they fall into 5 distinct groups separated largely along taxonomic lines suggesting minimal lateral mdh transfer between Archaea.
}, keywords = {Citric Acid Cycle, Hydrogen-Ion Concentration, Kinetics, Malate Dehydrogenase, Malates, Molecular Weight, NAD, NADP, Oxaloacetic Acid, Phylogeny, Protein Conformation, Protein Subunits, Pyrobaculum, Recombinant Proteins, Sequence Analysis, Protein, Substrate Specificity, Temperature}, issn = {1431-0651}, doi = {10.1007/s00792-007-0081-2}, author = {Yennaco, Lynda J and Hu, Yajing and Holden, James F} } @article {746, title = {Comparative bacterial diversity in recent Hawaiian volcanic deposits of different ages.}, journal = {FEMS Microbiol Ecol}, volume = {60}, year = {2007}, month = {2007 Apr}, pages = {60-73}, abstract = {Volcanic activity creates new landforms that can change dramatically over time as a consequence of biotic succession. Nonetheless, volcanic deposits present severe constraints for microbial colonization and activity. We have characterized bacterial diversity on four recent deposits at Kilauea volcano, Hawaii (KVD). Much of the diversity was either closely related to uncultured organisms or distinct from any reported 16S rRNA gene sequences. Diversity indices suggested that diversity was highest in a moderately vegetated 210-year-old ash deposit (1790-KVD), and lowest for a 79-year-old lava flow (1921-KVD). Diversity for a 41-year-old tephra deposit (1959-KVD) and a 300-year-old rainforest (1700-KVD) reached intermediate values. The 1959-KVD and 1790-KVD communities were dominated by Acidobacteria, Alpha- and Gammaproteobacteria, Actinobacteria, Cyanobacteria, and many unclassified phylotypes. The 1921-KVD, an unvegetated low pH deposit, was dominated by unclassified phylotypes. In contrast, 1700-KVD was primarily populated by Alphaproteobacteria with very few unclassified phylotypes. Similar diversity indices and levels of trace gas flux were found for 1959-KVD and 1790-KVD; however, statistical analyses indicated significantly different communities. This study not only showed that microorganisms colonize recent volcanic deposits and are able to establish diverse communities, but also that their composition is governed by variations in local deposit parameters.}, keywords = {Bacteria, DNA, Bacterial, Ecosystem, Genes, rRNA, Genetic Variation, Hawaii, Molecular Sequence Data, Phylogeny, Polymerase Chain Reaction, RNA, Ribosomal, 16S, Sequence Analysis, DNA, Soil Microbiology, Time Factors, Volcanic Eruptions}, issn = {0168-6496}, doi = {10.1111/j.1574-6941.2006.00253.x}, author = {Gomez-Alvarez, Vicente and King, Gary M and N{\"u}sslein, Klaus} } @article {818, title = {DinI and RecX modulate RecA-DNA structures in Escherichia coli K-12.}, journal = {Mol Microbiol}, volume = {63}, year = {2007}, month = {2007 Jan}, pages = {103-15}, abstract = {RecA plays a central role in recombination, DNA repair and SOS induction through forming a RecA-DNA helical filament. Biochemical observations show that at low ratios to RecA, DinI and RecX stabilize and destabilize RecA-DNA filaments, respectively, and that the C-terminal 17 residues of RecA are important for RecX function. RecA-DNA filament formation was assayed in vivo using RecA-GFP foci formation in log-phase and UV-irradiated cells. In log-phase cells, dinI mutants have fewer foci than wild type and that recX mutants have more foci than wild type. A recADelta17::gfp mutant had more foci like a recX mutant. dinI recX double mutants have the same number of foci as dinI mutants alone, suggesting that dinI is epistatic to recX. After UV treatment, the dinI, recX and dinI recX mutants differed in their ability to form foci. All three mutants had fewer foci than wild type. The dinI mutant{\textquoteright}s foci persisted longer than wild-type foci. Roles of DinI and RecX after UV treatment differed from those during log-phase growth and may reflect the different DNA substrates, population of proteins or amounts during the SOS response. These experiments give new insight into the roles of these proteins.}, keywords = {Bacterial Proteins, DNA, Bacterial, Escherichia coli, Escherichia coli Proteins, Gene Expression Regulation, Bacterial, Genes, Bacterial, Rec A Recombinases}, issn = {0950-382X}, doi = {10.1111/j.1365-2958.2006.05496.x}, author = {Renzette, Nicholas and Gumlaw, Nathan and Sandler, Steven J} } @article {496, title = {Evidence that OmcB and OmpB of Geobacter sulfurreducens are outer membrane surface proteins.}, journal = {FEMS Microbiol Lett}, volume = {277}, year = {2007}, month = {2007 Dec}, pages = {21-7}, abstract = {The c-type cytochrome (OmcB) and the multicopper protein (OmpB) required for Fe(III) oxide reduction by Geobacter sulfurreducens were predicted previously to be outer membrane proteins, but it is not clear whether they are positioned in a manner that permits the interaction with Fe(III). Treatment of whole cells with proteinase K inhibited Fe(III) reduction, but had no impact on the inner membrane-associated fumarate reduction. OmcB was digested by protease, resulting in a smaller peptide. However, immunogold labeling coupled with transmission electron microscopy did not detect OmcB, suggesting that it is only partially exposed on the cell surface. In contrast, OmpB was completely digested with protease. OmpB was loosely associated with the cell surface as a substantial portion of it was recovered in the culture supernatant. Immunogold labeling demonstrated that OmpB associated with the cell was evenly distributed on the cell surface rather than localized to one side of the cell like the conductive pili. Although several proteins required for Fe(III) oxide reduction are shown to be exposed on the outer surface of G. sulfurreducens, the finding that OmcB is also surface exposed is the first report of a protein required for optimal Fe(III) citrate reduction at least partially accessible on the cell surface.}, keywords = {Bacterial Outer Membrane Proteins, Cell Membrane, Cytochromes c, Ferric Compounds, Geobacter, Microscopy, Electron, Transmission, Oxidation-Reduction, Peptide Hydrolases}, issn = {0378-1097}, doi = {10.1111/j.1574-6968.2007.00915.x}, author = {Qian, Xinlei and Reguera, Gemma and Mester, T{\"u}nde and Lovley, Derek R} } @article {771, title = {An expectation maximization approach to estimate malaria haplotype frequencies in multiply infected children.}, journal = {Stat Appl Genet Mol Biol}, volume = {6}, year = {2007}, month = {2007}, pages = {Article33}, abstract = {Characterizing genetic variability in the human pathogenic Plasmodium species, the group of parasites that cause Malaria, may have broad global health implications. Specifically, discerning the combinations of mutations that lead to viable parasites and the population level frequencies of these clonal sequences will allow for targeted vaccine development and individualized treatment choices. This presents an analytical challenge, however, since haplotypic phase (i.e. the alignment of bases on a single DNA strand) is generally unobservable in multiply infected individuals. This manuscript describes an expectation maximization (EM) approach to maximum likelihood estimation of haplotype frequencies in this missing data setting. The approach is applied to a cohort of N=341 malaria infected children in Uganda, Cameroon and Sudan to characterize regional differences. A simulation study is also presented to characterize method performance and assess sensitivity to distributional assumptions.}, keywords = {Animals, Child, Gene Frequency, Genotype, Humans, Likelihood Functions, Malaria, Malaria Vaccines, Mutation, Plasmodium}, issn = {1544-6115}, doi = {10.2202/1544-6115.1321}, author = {Li, Xiaohong and Foulkes, Andrea S and Yucel, Recai M and Rich, Stephen M} } @article {503, title = {Genome-wide expression profiling in Geobacter sulfurreducens: identification of Fur and RpoS transcription regulatory sites in a relGsu mutant.}, journal = {Funct Integr Genomics}, volume = {7}, year = {2007}, month = {2007 Jul}, pages = {229-55}, abstract = {Rel(Gsu) is the single Geobacter sulfurreducens homolog of RelA and SpoT proteins found in many organisms. These proteins are involved in the regulation of levels of guanosine 3{\textquoteright}, 5{\textquoteright} bispyrophosphate, ppGpp, a molecule that signals slow growth and stress response under nutrient limitation in bacteria. We used information obtained from genome-wide expression profiling of the rel(Gsu) deletion mutant to identify putative regulatory sites involved in transcription networks modulated by Rel(Gsu) or ppGpp. Differential gene expression in the rel(Gsu) deletion mutant, as compared to the wild type, was available from two growth conditions, steady state chemostat cultures and stationary phase batch cultures. Hierarchical clustering analysis of these two datasets identified several groups of operons that are likely co-regulated. Using a search for conserved motifs in the upstream regions of these co-regulated operons, we identified sequences similar to Fur- and RpoS-regulated sites. These findings suggest that Fur- and RpoS-dependent gene expression in G. sulfurreducens is affected by Rel(Gsu)-mediated signaling.}, keywords = {Bacterial Proteins, Base Sequence, Gene Deletion, Gene Expression Profiling, Gene Expression Regulation, Bacterial, Genes, Bacterial, Genome, Bacterial, Geobacter, Ligases, Mutation, Operon, Promoter Regions, Genetic, Regulatory Sequences, Nucleic Acid, Repressor Proteins, Sigma Factor, Transcription, Genetic}, issn = {1438-793X}, doi = {10.1007/s10142-007-0048-5}, author = {Krushkal, Julia and Yan, Bin and DiDonato, Laurie N and Puljic, Marko and Nevin, Kelly P and Woodard, Trevor L and Adkins, Ronald M and Meth{\'e}, Barbara A and Lovley, Derek R} } @article {509, title = {Genome-wide similarity search for transcription factors and their binding sites in a metal-reducing prokaryote Geobacter sulfurreducens.}, journal = {Biosystems}, volume = {90}, year = {2007}, month = {2007 Sep-Oct}, pages = {421-41}, abstract = {The knowledge obtained from understanding individual elements involved in gene regulation is important for reconstructing gene regulatory networks, a key for understanding cellular behavior. To study gene regulatory interactions in a model microorganism, Geobacter sulfurreducens, which participates in metal reduction and energy harvesting, we investigated the presence of 59 known Escherichia coli transcription factors and predicted transcription regulatory sites in its genome. The supplementary material, available at http://www.geobacter.org/research/genomescan/, provides the results of similarity comparisons that identified regulatory proteins of G. sulfurreducens and the genome locations of the predicted regulatory sites, including the list of putative regulatory elements in the upstream regions of every predicted operon and singleton open reading frame. Regulatory sequence elements, predicted using genome similarity searches to matrices of established transcription regulatory elements from E. coli, provide an initial insight into regulation of genes and operons in G. sulfurreducens. The predicted regulatory elements were predominantly located in the upstream regions of operons and singleton open reading frames. The validity of the predictions was examined using a permutation approach. Sequence similarity searches indicate that E. coli transcription factors ArgR, CytR, DeoR, FlhCD (both FlhC and FlhD subunits), FruR, GalR, GlpR, H-NS, LacI, MetJ, PurR, TrpR, and Tus are likely missing from G. sulfurreducens. Phylogenetic analysis suggests that one HU subunit is present in G. sulfurreducens as compared to two subunits in E. coli, while each of the two E. coli IHF subunits, HimA and HimD, have two homologs in G. sulfurreducens. The closest homolog of E. coli RpoE in G. sulfurreducens may be more similar to FecI than to RpoE. These findings represent the first step in the understanding of the regulatory relationships in G. sulfurreducens on the genome scale.}, keywords = {Bacterial Proteins, Binding Sites, False Positive Reactions, Gene Expression Regulation, Bacterial, Genome, Bacterial, Geobacter, Metals, Models, Biological, Models, Genetic, Models, Statistical, Operon, Phylogeny, Sigma Factor, Systems Biology, Transcription Factors, Transcription, Genetic}, issn = {0303-2647}, doi = {10.1016/j.biosystems.2006.10.006}, author = {Yan, Bin and Lovley, Derek R and Krushkal, Julia} } @article {502, title = {Genomic and microarray analysis of aromatics degradation in Geobacter metallireducens and comparison to a Geobacter isolate from a contaminated field site.}, journal = {BMC Genomics}, volume = {8}, year = {2007}, month = {2007}, pages = {180}, abstract = {BACKGROUND: Groundwater and subsurface environments contaminated with aromatic compounds can be remediated in situ by Geobacter species that couple oxidation of these compounds to reduction of Fe(III)-oxides. Geobacter metallireducens metabolizes many aromatic compounds, but the enzymes involved are not well known. RESULTS: The complete G. metallireducens genome contained a 300 kb island predicted to encode enzymes for the degradation of phenol, p-cresol, 4-hydroxybenzaldehyde, 4-hydroxybenzoate, benzyl alcohol, benzaldehyde, and benzoate. Toluene degradation genes were encoded in a separate region. None of these genes was found in closely related species that cannot degrade aromatic compounds. Abundant transposons and phage-like genes in the island suggest mobility, but nucleotide composition and lack of synteny with other species do not suggest a recent transfer. The inferred degradation pathways are similar to those in species that anaerobically oxidize aromatic compounds with nitrate as an electron acceptor. In these pathways the aromatic compounds are converted to benzoyl-CoA and then to 3-hydroxypimelyl-CoA. However, in G. metallireducens there were no genes for the energetically-expensive dearomatizing enzyme. Whole-genome changes in transcript levels were identified in cells oxidizing benzoate. These supported the predicted pathway, identified induced fatty-acid oxidation genes, and identified an apparent shift in the TCA cycle to a putative ATP-yielding succinyl-CoA synthase. Paralogs to several genes in the pathway were also induced, as were several putative molybdo-proteins. Comparison of the aromatics degradation pathway genes to the genome of an isolate from a contaminated field site showed very similar content, and suggested this strain degrades many of the same compounds. This strain also lacked a classical dearomatizing enzyme, but contained two copies of an eight-gene cluster encoding redox proteins that was 30-fold induced during benzoate oxidation. CONCLUSION: G. metallireducens appears to convert aromatic compounds to benzoyl-CoA, then to acetyl-CoA via fatty acid oxidation, and then to carbon dioxide via the TCA cycle. The enzyme responsible for dearomatizing the aromatic ring may be novel, and energetic investments at this step may be offset by a change in succinate metabolism. Analysis of a field isolate suggests that the pathways inferred for G. metallireducens may be applicable to modeling in situ bioremediation.}, keywords = {Biodegradation, Environmental, Citric Acid Cycle, Gene Expression Regulation, Bacterial, Genomics, Geobacter, Hydrocarbons, Aromatic, Microarray Analysis, Models, Chemical, Models, Genetic, Multigene Family, Oligonucleotide Array Sequence Analysis, Oxidation-Reduction, Phylogeny, Soil Pollutants, Water Pollutants, Chemical}, issn = {1471-2164}, doi = {10.1186/1471-2164-8-180}, author = {Butler, Jessica E and He, Qiang and Nevin, Kelly P and He, Zhili and Zhou, Jizhong and Lovley, Derek R} } @article {508, title = {Geobacter pickeringii sp. nov., Geobacter argillaceus sp. nov. and Pelosinus fermentans gen. nov., sp. nov., isolated from subsurface kaolin lenses.}, journal = {Int J Syst Evol Microbiol}, volume = {57}, year = {2007}, month = {2007 Jan}, pages = {126-35}, abstract = {The goal of this project was to isolate representative Fe(III)-reducing bacteria from kaolin clays that may influence iron mineralogy in kaolin. Two novel dissimilatory Fe(III)-reducing bacteria, strains G12(T) and G13(T), were isolated from sedimentary kaolin strata in Georgia (USA). Cells of strains G12(T) and G13(T) were motile, non-spore-forming regular rods, 1-2 mum long and 0.6 mum in diameter. Cells had one lateral flagellum. Phylogenetic analyses using the 16S rRNA gene sequence of the novel strains demonstrated their affiliation to the genus Geobacter. Strain G12(T) was most closely related to Geobacter pelophilus (94.7 \%) and Geobacter chapellei (94.1 \%). Strain G13(T) was most closely related to Geobacter grbiciae (95.3 \%) and Geobacter metallireducens (95.1 \%). Based on phylogenetic analyses and phenotypic differences between the novel isolates and other closely related species of the genus Geobacter, the isolates are proposed as representing two novel species, Geobacter argillaceus sp. nov. (type strain G12(T)=ATCC BAA-1139(T)=JCM 12999(T)) and Geobacter pickeringii sp. nov. (type strain G13(T)=ATCC BAA-1140(T)=DSM 17153(T)=JCM 13000(T)). Another isolate, strain R7(T), was derived from a primary kaolin deposit in Russia. The cells of strain R7(T) were motile, spore-forming, slightly curved rods, 0.6 x 2.0-6.0 microm in size and with up to six peritrichous flagella. Strain R7(T) was capable of reducing Fe(III) only in the presence of a fermentable substrate. 16S rRNA gene sequence analysis demonstrated that this isolate is unique, showing less than 92 \% similarity to bacteria of the Sporomusa-Pectinatus-Selenomomas phyletic group, including {\textquoteright}Anaerospora hongkongensis{\textquoteright} (90.2 \%), Acetonema longum (90.6 \%), Dendrosporobacter quercicolus (90.9 \%) and Anaerosinus glycerini (91.5 \%). On the basis of phylogenetic analysis and physiological tests, strain R7(T) is proposed to represent a novel genus and species, Pelosinus fermentans gen. nov., sp. nov. (type strain R7(T)=DSM 17108(T)=ATCC BAA-1133(T)), in the Sporomusa-Pectinatus-Selenomonas group.}, keywords = {Bacterial Typing Techniques, Base Composition, DNA, Bacterial, DNA, Ribosomal, Ferric Compounds, Genes, rRNA, Geobacter, Geologic Sediments, Georgia, Kaolin, Molecular Sequence Data, Oxidation-Reduction, Phylogeny, RNA, Ribosomal, 16S, Russia, Sequence Analysis, DNA, Species Specificity}, issn = {1466-5026}, doi = {10.1099/ijs.0.64221-0}, author = {Shelobolina, Evgenya S and Nevin, Kelly P and Blakeney-Hayward, Jessie D and Johnsen, Claudia V and Plaia, Todd W and Krader, Paul and Woodard, Trevor and Holmes, Dawn E and Vanpraagh, Catherine Gaw and Lovley, Derek R} } @article {816, title = {A hand-off mechanism for primosome assembly in replication restart.}, journal = {Mol Cell}, volume = {26}, year = {2007}, month = {2007 Jun 22}, pages = {781-93}, abstract = {Collapsed DNA replication forks must be reactivated through origin-independent reloading of the replication machinery (replisome) to ensure complete duplication of cellular genomes. In E. coli, the PriA-dependent pathway is the major replication restart mechanism and requires primosome proteins PriA, PriB, and DnaT for replisome reloading. However, the molecular mechanisms that regulate origin-independent replisome loading are not fully understood. Here, we demonstrate that assembly of primosome protein complexes represents a key regulatory mechanism, as inherently weak PriA-PriB and PriB-DnaT interactions are strongly stimulated by single-stranded DNA. Furthermore, the binding site on PriB for single-stranded DNA partially overlaps the binding sites for PriA and DnaT, suggesting a dynamic primosome assembly process in which single-stranded DNA is handed off from one primosome protein to another as a repaired replication fork is reactivated. This model helps explain how origin-independent initiation of DNA replication is restricted to repaired replication forks, preventing overreplication of the genome.}, keywords = {Binding Sites, DNA Helicases, DNA Replication, DNA, Bacterial, DNA, Single-Stranded, DNA-Binding Proteins, DNA-Directed DNA Polymerase, Escherichia coli, Escherichia coli Proteins, Genome, Bacterial, Models, Biological, Multienzyme Complexes, Protein Binding, Replication Origin}, issn = {1097-2765}, doi = {10.1016/j.molcel.2007.05.012}, author = {Lopper, Matthew and Boonsombat, Ruethairat and Sandler, Steven J and Keck, James L} } @article {507, title = {Heat-shock sigma factor RpoH from Geobacter sulfurreducens.}, journal = {Microbiology}, volume = {153}, year = {2007}, month = {2007 Mar}, pages = {838-46}, abstract = {Recent studies with Myxococcus xanthus have suggested that homologues of the Escherichia coli heat-shock sigma factor, RpoH, may not be involved in the heat-shock response in this delta-proteobacterium. The genome of another delta-proteobacterium, Geobacter sulfurreducens, which is considered to be a representative of the Fe(III)-reducing Geobacteraceae that predominate in a diversity of subsurface environments, contains an rpoH homologue. Characterization of the G. sulfurreducens rpoH homologue revealed that it was induced by a temperature shift from 30 degrees C to 42 degrees C and that an rpoH-deficient mutant was unable to grow at 42 degrees C. The predicted heat-shock genes, hrcA, grpE, dnaK, groES and htpG, were heat-shock inducible in an rpoH-dependent manner, and comparison of promoter regions of these genes identified the consensus sequences for the -10 and -35 promoter elements. In addition, DNA elements identical to the CIRCE consensus sequence were found in promoters of rpoH, hrcA and groES, suggesting that these genes are regulated by a homologue of the repressor HrcA, which is known to bind the CIRCE element. These results suggest that the G. sulfurreducens RpoH homologue is the heat-shock sigma factor and that heat-shock response in G. sulfurreducens is regulated positively by RpoH as well as negatively by the HrcA/CIRCE system.}, keywords = {Adaptation, Physiological, Base Sequence, Binding Sites, Consensus Sequence, DNA, Bacterial, Gene Expression Regulation, Bacterial, Genome, Bacterial, Geobacter, Heat-Shock Proteins, Hot Temperature, Molecular Sequence Data, Promoter Regions, Genetic, RNA, Bacterial, RNA, Messenger, Sigma Factor, Transcription, Genetic}, issn = {1350-0872}, doi = {10.1099/mic.0.2006/000638-0}, author = {Ueki, Toshiyuki and Lovley, Derek R} } @article {505, title = {Importance of c-Type cytochromes for U(VI) reduction by Geobacter sulfurreducens.}, journal = {BMC Microbiol}, volume = {7}, year = {2007}, month = {2007}, pages = {16}, abstract = {BACKGROUND: In order to study the mechanism of U(VI) reduction, the effect of deleting c-type cytochrome genes on the capacity of Geobacter sulfurreducens to reduce U(VI) with acetate serving as the electron donor was investigated. RESULTS: The ability of several c-type cytochrome deficient mutants to reduce U(VI) was lower than that of the wild type strain. Elimination of two confirmed outer membrane cytochromes and two putative outer membrane cytochromes significantly decreased (ca. 50-60\%) the ability of G. sulfurreducens to reduce U(VI). Involvement in U(VI) reduction did not appear to be a general property of outer membrane cytochromes, as elimination of two other confirmed outer membrane cytochromes, OmcB and OmcC, had very little impact on U(VI) reduction. Among the periplasmic cytochromes, only MacA, proposed to transfer electrons from the inner membrane to the periplasm, appeared to play a significant role in U(VI) reduction. A subpopulation of both wild type and U(VI) reduction-impaired cells, 24-30\%, accumulated amorphous uranium in the periplasm. Comparison of uranium-accumulating cells demonstrated a similar amount of periplasmic uranium accumulation in U(VI) reduction-impaired and wild type G. sulfurreducens. Assessment of the ability of the various suspensions to reduce Fe(III) revealed no correlation between the impact of cytochrome deletion on U(VI) reduction and reduction of Fe(III) hydroxide and chelated Fe(III). CONCLUSION: This study indicates that c-type cytochromes are involved in U(VI) reduction by Geobacter sulfurreducens. The data provide new evidence for extracellular uranium reduction by G. sulfurreducens but do not rule out the possibility of periplasmic uranium reduction. Occurrence of U(VI) reduction at the cell surface is supported by the significant impact of elimination of outer membrane cytochromes on U(VI) reduction and the lack of correlation between periplasmic uranium accumulation and the capacity for uranium reduction. Periplasmic uranium accumulation may reflect the ability of uranium to penetrate the outer membrane rather than the occurrence of enzymatic U(VI) reduction. Elimination of cytochromes rarely had a similar impact on both Fe(III) and U(VI) reduction, suggesting that there are differences in the routes of electron transfer to U(VI) and Fe(III). Further studies are required to clarify the pathways leading to U(VI) reduction in G. sulfurreducens.}, keywords = {Biodegradation, Environmental, Cytochrome c Group, Ferric Compounds, Geobacter, Microscopy, Electron, Transmission, Mutation, Oxidation-Reduction, Periplasm, Uranium}, issn = {1471-2180}, doi = {10.1186/1471-2180-7-16}, author = {Shelobolina, Evgenya S and Coppi, Maddalena V and Korenevsky, Anton A and DiDonato, Laurie N and Sullivan, Sara A and Konishi, Hiromi and Xu, Huifang and Leang, Ching and Butler, Jessica E and Kim, Byoung-Chan and Lovley, Derek R} } @article {815, title = {Intracranial pressure and cerebral oxygenation changes after decompressive craniectomy in a child with traumatic brain swelling.}, journal = {Childs Nerv Syst}, volume = {23}, year = {2007}, month = {2007 Nov}, pages = {1331-5}, abstract = {CASE REPORT: The authors present the case of a 5-year-old child with severe traumatic brain injury in whom decompressive hemicraniectomy was performed for progressive increased intracranial pressure (ICP) unresponsive to medical treatment. Data from ICP and cerebral tissue oxygenation monitoring in the contralateral hemisphere were recorded, which demonstrated the immediate and delayed mechanical and physiological changes occurring after bony and dural decompression. DISCUSSION: The role of the procedure and that of the monitoring approach are discussed.}, keywords = {Brain Edema, Brain Injuries, Cerebrum, Child, Preschool, Craniotomy, Decompression, Surgical, Functional Laterality, Humans, Hypoxia, Brain, Intracranial Hypertension, Male, Oxygen, Treatment Outcome}, issn = {0256-7040}, doi = {10.1007/s00381-007-0388-4}, author = {Figaji, A A and Fieggen, A G and Sandler, S J I and Argent, A C and Le Roux, P D and Peter, J C} } @article {499, title = {Involvement of Geobacter sulfurreducens SfrAB in acetate metabolism rather than intracellular, respiration-linked Fe(III) citrate reduction.}, journal = {Microbiology}, volume = {153}, year = {2007}, month = {2007 Oct}, pages = {3572-85}, abstract = {A soluble ferric reductase, SfrAB, which catalysed the NADPH-dependent reduction of chelated Fe(III), was previously purified from the dissimilatory Fe(III)-reducing micro-organism Geobacter sulfurreducens, suggesting that reduction of chelated forms of Fe(III) might be cytoplasmic. However, metabolically active spheroplast suspensions could not catalyse acetate-dependent Fe(III) citrate reduction, indicating that periplasmic and/or outer-membrane components were required for Fe(III) citrate reduction. Furthermore, phenotypic analysis of an SfrAB knockout mutant suggested that SfrAB was involved in acetate metabolism rather than respiration-linked Fe(III) reduction. The mutant could not grow via the reduction of either Fe(III) citrate or fumarate when acetate was the electron donor but could grow with either acceptor if either hydrogen or formate served as the electron donor. Following prolonged incubation in acetate : fumarate medium in the absence of hydrogen and formate, an {\textquoteright}acetate-adapted{\textquoteright} SfrAB-null strain was isolated that was capable of growth on acetate : fumarate medium but not acetate : Fe(III) citrate medium. Comparison of gene expression in this strain with that of the wild-type revealed upregulation of a potential NADPH-dependent ferredoxin oxidoreductase as well as genes involved in energy generation and amino acid uptake, suggesting that NADPH homeostasis and the tricarboxylic acid (TCA) cycle were perturbed in the {\textquoteright}acetate-adapted{\textquoteright} SfrAB-null strain. Membrane and soluble fractions prepared from the {\textquoteright}acetate-adapted{\textquoteright} strain were depleted of NADPH-dependent Fe(III), viologen and quinone reductase activities. These results indicate that cytoplasmic, respiration-linked reduction of Fe(III) by SfrAB in vivo is unlikely and suggest that deleting SfrAB may interfere with growth via acetate oxidation by interfering with NADP regeneration.}, keywords = {Acetates, Amino Acid Transport Systems, Bacterial Proteins, Cell Membrane, Citric Acid Cycle, Cytoplasm, Energy Metabolism, Ferric Compounds, Formic Acids, Fumarates, Gene Deletion, Gene Expression Profiling, Geobacter, Hydrogen, NADH, NADPH Oxidoreductases, Oligonucleotide Array Sequence Analysis}, issn = {1350-0872}, doi = {10.1099/mic.0.2007/006478-0}, author = {Coppi, Maddalena V and O{\textquoteright}Neil, Regina A and Leang, Ching and Kaufmann, Franz and Meth{\'e}, Barbara A and Nevin, Kelly P and Woodard, Trevor L and Liu, Anna and Lovley, Derek R} } @article {501, title = {Lack of electricity production by Pelobacter carbinolicus indicates that the capacity for Fe(III) oxide reduction does not necessarily confer electron transfer ability to fuel cell anodes.}, journal = {Appl Environ Microbiol}, volume = {73}, year = {2007}, month = {2007 Aug}, pages = {5347-53}, abstract = {The ability of Pelobacter carbinolicus to oxidize electron donors with electron transfer to the anodes of microbial fuel cells was evaluated because microorganisms closely related to Pelobacter species are generally abundant on the anodes of microbial fuel cells harvesting electricity from aquatic sediments. P. carbinolicus could not produce current in a microbial fuel cell with electron donors which support Fe(III) oxide reduction by this organism. Current was produced using a coculture of P. carbinolicus and Geobacter sulfurreducens with ethanol as the fuel. Ethanol consumption was associated with the transitory accumulation of acetate and hydrogen. G. sulfurreducens alone could not metabolize ethanol, suggesting that P. carbinolicus grew in the fuel cell by converting ethanol to hydrogen and acetate, which G. sulfurreducens oxidized with electron transfer to the anode. Up to 83\% of the electrons available in ethanol were recovered as electricity and in the metabolic intermediate acetate. Hydrogen consumption by G. sulfurreducens was important for ethanol metabolism by P. carbinolicus. Confocal microscopy and analysis of 16S rRNA genes revealed that half of the cells growing on the anode surface were P. carbinolicus, but there was a nearly equal number of planktonic cells of P. carbinolicus. In contrast, G. sulfurreducens was primarily attached to the anode. P. carbinolicus represents the first Fe(III) oxide-reducing microorganism found to be unable to produce current in a microbial fuel cell, providing the first suggestion that the mechanisms for extracellular electron transfer to Fe(III) oxides and fuel cell anodes may be different.}, keywords = {Bacteriological Techniques, Deltaproteobacteria, Electricity, Electron Transport, Ferrous Compounds, In Situ Hybridization, Fluorescence, Microscopy, Confocal, Oxidation-Reduction, RNA, Ribosomal, 16S}, issn = {0099-2240}, doi = {10.1128/AEM.00804-07}, author = {Richter, Hanno and Lanthier, Martin and Nevin, Kelly P and Lovley, Derek R} } @article {35, title = {Multiple basic helix-loop-helix proteins regulate expression of the ENO1 gene of Saccharomyces cerevisiae.}, journal = {Eukaryot Cell}, volume = {6}, year = {2007}, month = {2007 May}, pages = {786-96}, abstract = {The basic helix-loop-helix (bHLH) eukaryotic transcription factors have the ability to form multiple dimer combinations. This property, together with limited DNA-binding specificity for the E box (CANNTG), makes them ideally suited for combinatorial control of gene expression. We tested the ability of all nine Saccharomyces cerevisiae bHLH proteins to regulate the enolase-encoding gene ENO1. ENO1 was known to be activated by the bHLH protein Sgc1p. Here we show that expression of an ENO1-lacZ reporter was also regulated by the other eight bHLH proteins, namely, Ino2p, Ino4p, Cbf1p, Rtg1p, Rtg3p, Pho4p, Hms1p, and Ygr290wp. ENO1-lacZ expression was also repressed by growth in inositol-choline-containing medium. Epistatic analysis and chromatin immunoprecipitation experiments showed that regulation by Sgc1p, Ino2p, Ino4p, and Cbf1p and repression by inositol-choline required three distal E boxes, E1, E2, and E3. The pattern of bHLH binding to the three E boxes and experiments with two dominant-negative mutant alleles of INO4 and INO2 support the model that bHLH dimer selection affects ENO1-lacZ expression. These results support the general model that bHLH proteins can coordinate different biological pathways via multiple mechanisms.}, keywords = {Basic Helix-Loop-Helix Transcription Factors, beta-Galactosidase, Chromatin Immunoprecipitation, E-Box Elements, Epistasis, Genetic, Evolution, Molecular, Gene Expression Regulation, Fungal, Genes, Dominant, Models, Biological, Mutation, Phosphopyruvate Hydratase, Promoter Regions, Genetic, Recombinant Fusion Proteins, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins}, issn = {1535-9778}, doi = {10.1128/EC.00383-06}, author = {Chen, Meng and Lopes, John M} } @article {854, title = {Notice of duplicate publication.}, journal = {Am J Respir Crit Care Med}, volume = {175}, year = {2007}, month = {2007 Jan 1}, pages = {94}, keywords = {Duplicate Publication as Topic}, issn = {1073-449X}, author = {Webley, Wilmore C and Salva, Paul S and Andrzejewski, Chester and Cirino, Frances and West, Corrie A and Tilahun, Yaphet and Stuart, Elizabeth S} } @article {681, title = {Pepitope: epitope mapping from affinity-selected peptides.}, journal = {Bioinformatics}, volume = {23}, year = {2007}, month = {2007 Dec 1}, pages = {3244-6}, abstract = {Identifying the epitope to which an antibody binds is central for many immunological applications such as drug design and vaccine development. The Pepitope server is a web-based tool that aims at predicting discontinuous epitopes based on a set of peptides that were affinity-selected against a monoclonal antibody of interest. The server implements three different algorithms for epitope mapping: PepSurf, Mapitope, and a combination of the two. The rationale behind these algorithms is that the set of peptides mimics the genuine epitope in terms of physicochemical properties and spatial organization. When the three-dimensional (3D) structure of the antigen is known, the information in these peptides can be used to computationally infer the corresponding epitope. A user-friendly web interface and a graphical tool that allows viewing the predicted epitopes were developed. Pepitope can also be applied for inferring other types of protein-protein interactions beyond the immunological context, and as a general tool for aligning linear sequences to a 3D structure. AVAILABILITY: http://pepitope.tau.ac.il/}, keywords = {Algorithms, Amino Acid Sequence, Binding Sites, Epitope Mapping, Molecular Sequence Data, Peptides, Protein Binding, Sequence Alignment, Sequence Analysis, Protein, Software}, issn = {1367-4811}, doi = {10.1093/bioinformatics/btm493}, author = {Mayrose, Itay and Penn, Osnat and Erez, Elana and Rubinstein, Nimrod D and Shlomi, Tomer and Freund, Natalia Tarnovitski and Bublil, Erez M and Ruppin, Eytan and Sharan, Roded and Gershoni, Jonathan M and Martz, Eric and Pupko, Tal} } @article {510, title = {Possible nonconductive role of Geobacter sulfurreducens pilus nanowires in biofilm formation.}, journal = {J Bacteriol}, volume = {189}, year = {2007}, month = {2007 Mar}, pages = {2125-7}, abstract = {Geobacter sulfurreducens required expression of electrically conductive pili to form biofilms on Fe(III) oxide surfaces, but pili were also essential for biofilm development on plain glass when fumarate was the sole electron acceptor. Furthermore, pili were needed for cell aggregation in agglutination studies. These results suggest that the pili of G. sulfurreducens also have a structural role in biofilm formation.}, keywords = {Agglutination, Biofilms, Electron Transport, Ferric Compounds, Fimbriae, Bacterial, Geobacter}, issn = {0021-9193}, doi = {10.1128/JB.01284-06}, author = {Reguera, Gemma and Pollina, Rachael B and Nicoll, Julie S and Lovley, Derek R} } @article {504, title = {Prolixibacter bellariivorans gen. nov., sp. nov., a sugar-fermenting, psychrotolerant anaerobe of the phylum Bacteroidetes, isolated from a marine-sediment fuel cell.}, journal = {Int J Syst Evol Microbiol}, volume = {57}, year = {2007}, month = {2007 Apr}, pages = {701-7}, abstract = {A Gram-negative, non-motile, filamentous, rod-shaped, non-spore-forming bacterium (strain F2(T)) was isolated from the surface of an electricity-harvesting electrode incubated in marine sediments. Strain F2(T) does not contain c-type cytochromes, flexirubin or carotenoids. It is a facultative anaerobe that can ferment sugars by using a mixed acid fermentation pathway and it can grow over a wide range of temperatures (4-42 degrees C). The DNA G+C (44.9 mol\%) content and chemotaxonomic characteristics (major fatty acids, a-15 : 0 and 15 : 0) were consistent with those of species within the phylum Bacteroidetes. Phylogenetic analysis of the 16S rRNA nucleotide and elongation factor G amino acid sequences indicated that strain F2(T) represents a unique phylogenetic cluster within the phylum Bacteroidetes. On the basis of 16S rRNA gene sequence phylogeny, the closest relative available in pure culture, Alkaliflexus imshenetskii, is only 87.5 \% similar to strain F2(T). Results from physiological, biochemical and phylogenetic analyses showed that strain F2(T) should be classified as a novel genus and species within the phylum Bacteroidetes, for which the name Prolixibacter bellariivorans gen. nov., sp. nov. is proposed. The type strain is F2(T) (=ATCC BAA-1284(T)=JCM 13498(T)).}, keywords = {Bacteroidetes, Carbohydrate Metabolism, Cold Temperature, DNA, Bacterial, DNA, Ribosomal, Energy-Generating Resources, Geologic Sediments, Molecular Sequence Data, Phylogeny, RNA, Ribosomal, 16S, Seawater}, issn = {1466-5026}, doi = {10.1099/ijs.0.64296-0}, author = {Holmes, Dawn E and Nevin, Kelly P and Woodard, Trevor L and Peacock, Aaron D and Lovley, Derek R} } @article {506, title = {Reclassification of Trichlorobacter thiogenes as Geobacter thiogenes comb. nov.}, journal = {Int J Syst Evol Microbiol}, volume = {57}, year = {2007}, month = {2007 Mar}, pages = {463-6}, abstract = {Reclassification of the species Trichlorobacter thiogenes as Geobacter thiogenes comb. nov. is proposed on the basis of physiological traits and phylogenetic position. Characteristics additional to those provided in the original description revealed that the type strain (strain K1(T)=ATCC BAA-34(T)=JCM 14045(T)) has the ability to use Fe(III) as an electron acceptor for acetate oxidation and has an electron donor and acceptor profile typical of a Geobacter species, contains abundant c-type cytochromes, and has a temperature optimum of 30 degrees C and a pH optimum near pH 7.0; traits typical of members of the genus Geobacter. Phylogenetic analysis of nifD, recA, gyrB, rpoB, fusA and 16S rRNA genes further indicated that T. thiogenes falls within the Geobacter cluster of the family Geobacteraceae. Based on extensive phylogenetic evidence and the fact that T. thiogenes has the hallmark physiological characteristics of a Geobacter species, Trichlorobacter thiogenes should be reclassified as a member of the genus Geobacter.}, keywords = {DNA, Bacterial, DNA, Ribosomal, Genes, Bacterial, Geobacter, Phylogeny, RNA, Ribosomal, 16S, Temperature}, issn = {1466-5026}, doi = {10.1099/ijs.0.63408-0}, author = {Nevin, Kelly P and Holmes, Dawn E and Woodard, Trevor L and Covalla, Sean F and Lovley, Derek R} } @article {745, title = {Responses of rhizobia to desiccation in relation to osmotic stress, oxygen, and temperature.}, journal = {Appl Environ Microbiol}, volume = {73}, year = {2007}, month = {2007 Jun}, pages = {3451-9}, keywords = {Adaptation, Physiological, Dehydration, Hot Temperature, Osmotic Pressure, Oxidative Stress, Rhizobiaceae}, issn = {0099-2240}, doi = {10.1128/AEM.02991-06}, author = {Vriezen, Jan Ac and de Bruijn, Frans J and N{\"u}sslein, K} } @article {744, title = {Salinity constraints on subsurface archaeal diversity and methanogenesis in sedimentary rock rich in organic matter.}, journal = {Appl Environ Microbiol}, volume = {73}, year = {2007}, month = {2007 Jul}, pages = {4171-9}, abstract = {The diversity of microorganisms active within sedimentary rocks provides important controls on the geochemistry of many subsurface environments. In particular, biodegradation of organic matter in sedimentary rocks contributes to the biogeochemical cycling of carbon and other elements and strongly impacts the recovery and quality of fossil fuel resources. In this study, archaeal diversity was investigated along a salinity gradient spanning 8 to 3,490 mM Cl(-) in a subsurface shale rich in CH(4) derived from biodegradation of sedimentary hydrocarbons. Shale pore waters collected from wells in the main CH(4)-producing zone lacked electron acceptors such as O(2), NO(3)(-), Fe(3+), or SO(4)(2-). Acetate was detected only in high-salinity waters, suggesting that acetoclastic methanogenesis is inhibited at Cl(-) concentrations above approximately 1,000 mM. Most-probable-number series revealed differences in methanogen substrate utilization (acetate, trimethylamine, or H(2)/CO(2)) associated with chlorinity. The greatest methane production in enrichment cultures was observed for incubations with salinity at or close to the native pore water salinity of the inoculum. Restriction fragment length polymorphism analyses of archaeal 16S rRNA genes from seven wells indicated that there were links between archaeal communities and pore water salinity. Archaeal clone libraries constructed from sequences from 16S rRNA genes isolated from two wells revealed phylotypes similar to a halophilic methylotrophic Methanohalophilus species and a hydrogenotrophic Methanoplanus species at high salinity and a single phylotype closely related to Methanocorpusculum bavaricum at low salinity. These results show that several distinct communities of methanogens persist in this subsurface, CH(4)-producing environment and that each community is adapted to particular conditions of salinity and preferential substrate use and each community induces distinct geochemical signatures in shale formation waters.}, keywords = {Archaea, Base Sequence, Cloning, Molecular, DNA Primers, Genes, Archaeal, Genetic Variation, Geologic Sediments, Methane, Michigan, Molecular Sequence Data, Organic Chemicals, Phylogeny, RNA, Archaeal, RNA, Ribosomal, 16S, Sodium Chloride, Soil, Soil Microbiology, Water Microbiology}, issn = {0099-2240}, doi = {10.1128/AEM.02810-06}, author = {Waldron, Patricia J and Petsch, Steven T and Martini, Anna M and N{\"u}sslein, Klaus and N{\"u}slein, Klaus} } @article {682, title = {Selecton 2007: advanced models for detecting positive and purifying selection using a Bayesian inference approach.}, journal = {Nucleic Acids Res}, volume = {35}, year = {2007}, month = {2007 Jul}, pages = {W506-11}, abstract = {Biologically significant sites in a protein may be identified by contrasting the rates of synonymous (K(s)) and non-synonymous (K(a)) substitutions. This enables the inference of site-specific positive Darwinian selection and purifying selection. We present here Selecton version 2.2 (http://selecton.bioinfo.tau.ac.il), a web server which automatically calculates the ratio between K(a) and K(s) (omega) at each site of the protein. This ratio is graphically displayed on each site using a color-coding scheme, indicating either positive selection, purifying selection or lack of selection. Selecton implements an assembly of different evolutionary models, which allow for statistical testing of the hypothesis that a protein has undergone positive selection. Specifically, the recently developed mechanistic-empirical model is introduced, which takes into account the physicochemical properties of amino acids. Advanced options were introduced to allow maximal fine tuning of the server to the user{\textquoteright}s specific needs, including calculation of statistical support of the omega values, an advanced graphic display of the protein{\textquoteright}s 3-dimensional structure, use of different genetic codes and inputting of a pre-built phylogenetic tree. Selecton version 2.2 is an effective, user-friendly and freely available web server which implements up-to-date methods for computing site-specific selection forces, and the visualization of these forces on the protein{\textquoteright}s sequence and structure.}, keywords = {Algorithms, Amino Acid Sequence, Amino Acid Substitution, Amino Acids, Animals, Bayes Theorem, Computational Biology, Computer Simulation, Humans, Internet, Models, Molecular, Molecular Sequence Data, Phylogeny, Proteins, Sequence Analysis, Protein, Software}, issn = {1362-4962}, doi = {10.1093/nar/gkm382}, author = {Stern, Adi and Doron-Faigenboim, Adi and Erez, Elana and Martz, Eric and Bacharach, Eran and Pupko, Tal} } @article {495, title = {Steady state protein levels in Geobacter metallireducens grown with iron (III) citrate or nitrate as terminal electron acceptor.}, journal = {Proteomics}, volume = {7}, year = {2007}, month = {2007 Nov}, pages = {4148-57}, abstract = {Geobacter species predominate in aquatic sediments and submerged soils where organic carbon sources are oxidized with the reduction of Fe(III). The natural occurrence of Geobacter in some waste sites suggests this microorganism could be useful for bioremediation if growth and metabolic activity can be regulated. 2-DE was used to monitor the steady state protein levels of Geobacter metallireducens grown with either Fe(III) citrate or nitrate to elucidate metabolic differences in response to different terminal electron acceptors present in natural environments populated by Geobacter. Forty-six protein spots varied significantly in abundance (p<0.05) between the two growth conditions; proteins were identified by tryptic peptide mass and peptide sequence determined by MS/MS. Enzymes involved in pyruvate metabolism and the tricarboxylic acid (TCA) cycle were more abundant in cells grown with Fe(III) citrate, while proteins associated with nitrate metabolism and sensing cellular redox status along with several proteins of unknown function were more abundant in cells grown with nitrate. These results indicate a higher level of flux through the TCA cycle in the presence of Fe(III) compared to nitrate. The oxidative stress response observed in previous studies of Geobacter sulfurreducens grown with Fe(III) citrate was not seen in G. metallireducens.}, keywords = {Bacterial Proteins, Cell Proliferation, Electrons, Electrophoresis, Gel, Two-Dimensional, Ferric Compounds, Geobacter, Hydrogen-Ion Concentration, Nitrates, Oxidation-Reduction, Proteomics, Tandem Mass Spectrometry}, issn = {1615-9853}, doi = {10.1002/pmic.200600955}, author = {Ahrendt, Angela J and Tollaksen, Sandra L and Lindberg, Carl and Zhu, Wenhong and Yates, John R and Nevin, Kelly P and Babnigg, Gy{\"o}rgy and Lovley, Derek R and Giometti, Carol S} } @article {493, title = {Subsurface clade of Geobacteraceae that predominates in a diversity of Fe(III)-reducing subsurface environments.}, journal = {ISME J}, volume = {1}, year = {2007}, month = {2007 Dec}, pages = {663-77}, abstract = {There are distinct differences in the physiology of Geobacter species available in pure culture. Therefore, to understand the ecology of Geobacter species in subsurface environments, it is important to know which species predominate. Clone libraries were assembled with 16S rRNA genes and transcripts amplified from three subsurface environments in which Geobacter species are known to be important members of the microbial community: (1) a uranium-contaminated aquifer located in Rifle, CO, USA undergoing in situ bioremediation; (2) an acetate-impacted aquifer that serves as an analog for the long-term acetate amendments proposed for in situ uranium bioremediation and (3) a petroleum-contaminated aquifer in which Geobacter species play a role in the oxidation of aromatic hydrocarbons coupled with the reduction of Fe(III). The majority of Geobacteraceae 16S rRNA sequences found in these environments clustered in a phylogenetically coherent subsurface clade, which also contains a number of Geobacter species isolated from subsurface environments. Concatamers constructed with 43 Geobacter genes amplified from these sites also clustered within this subsurface clade. 16S rRNA transcript and gene sequences in the sediments and groundwater at the Rifle site were highly similar, suggesting that sampling groundwater via monitoring wells can recover the most active Geobacter species. These results suggest that further study of Geobacter species in the subsurface clade is necessary to accurately model the behavior of Geobacter species during subsurface bioremediation of metal and organic contaminants.}, keywords = {Biodegradation, Environmental, Ecosystem, Ferric Compounds, Geobacter, Hydrocarbons, Aromatic, Molecular Sequence Data, Oxidation-Reduction, Petroleum, Phylogeny, Polymerase Chain Reaction, RNA, Ribosomal, 16S, Sequence Analysis, DNA, Uranium}, issn = {1751-7362}, doi = {10.1038/ismej.2007.85}, author = {Holmes, Dawn E and O{\textquoteright}Neil, Regina A and Vrionis, Helen A and N{\textquoteright}guessan, Lucie A and Ortiz-Bernad, Irene and Larrahondo, Maria J and Adams, Lorrie A and Ward, Joy A and Nicoll, Julie S and Nevin, Kelly P and Chavan, Milind A and Johnson, Jessica P and Long, Philip E and Lovley, Derek R} } @article {36, title = {Transcriptional regulation of yeast phospholipid biosynthetic genes.}, journal = {Biochim Biophys Acta}, volume = {1771}, year = {2007}, month = {2007 Mar}, pages = {310-21}, abstract = {The last several years have been witness to significant developments in understanding transcriptional regulation of the yeast phospholipid structural genes. The response of most phospholipid structural genes to inositol is now understood on a mechanistic level. The roles of specific activators and repressors are also well established. The knowledge of specific regulatory factors that bind the promoters of phospholipid structural genes serves as a foundation for understanding the role of chromatin modification complexes. Collectively, these findings present a complex picture for transcriptional regulation of the phospholipid biosynthetic genes. The INO1 gene is an ideal example of the complexity of transcriptional control and continues to serve as a model for studying transcription in general. Furthermore, transcription of the regulatory genes is also subject to complex and essential regulation. In addition, databases resulting from a plethora of genome-wide studies have identified regulatory signals that control one of the essential phospholipid biosynthetic genes, PIS1. These databases also provide significant clues for other regulatory signals that may affect phospholipid biosynthesis. Here, we have tried to present a complete summary of the transcription factors and mechanisms that regulate the phospholipid biosynthetic genes.}, keywords = {CDP-Diacylglycerol-Inositol 3-Phosphatidyltransferase, Fungal Proteins, Gene Expression Regulation, Enzymologic, Gene Expression Regulation, Fungal, Intramolecular Lyases, Myo-Inositol-1-Phosphate Synthase, Phospholipids, Saccharomyces cerevisiae Proteins, Transcription Factors, Transcription, Genetic, Yeasts}, issn = {0006-3002}, doi = {10.1016/j.bbalip.2006.05.017}, author = {Chen, Meng and Hancock, Leandria C and Lopes, John M} } @article {817, title = {UvrD limits the number and intensities of RecA-green fluorescent protein structures in Escherichia coli K-12.}, journal = {J Bacteriol}, volume = {189}, year = {2007}, month = {2007 Apr}, pages = {2915-20}, abstract = {RecA is important for recombination, DNA repair, and SOS induction. In Escherichia coli, RecBCD, RecFOR, and RecJQ prepare DNA substrates onto which RecA binds. UvrD is a 3{\textquoteright}-to-5{\textquoteright} helicase that participates in methyl-directed mismatch repair and nucleotide excision repair. uvrD deletion mutants are sensitive to UV irradiation, hypermutable, and hyper-rec. In vitro, UvrD can dissociate RecA from single-stranded DNA. Other experiments suggest that UvrD removes RecA from DNA where it promotes unproductive reactions. To test if UvrD limits the number and/or the size of RecA-DNA structures in vivo, an uvrD mutation was combined with recA-gfp. This recA allele allows the number of RecA structures and the amount of RecA at these structures to be assayed in living cells. uvrD mutants show a threefold increase in the number of RecA-GFP foci, and these foci are, on average, nearly twofold higher in relative intensity. The increased number of RecA-green fluorescent protein foci in the uvrD mutant is dependent on recF, recO, recR, recJ, and recQ. The increase in average relative intensity is dependent on recO and recQ. These data support an in vivo role for UvrD in removing RecA from the DNA.}, keywords = {DNA Helicases, DNA, Bacterial, DNA-Binding Proteins, Escherichia coli K12, Escherichia coli Proteins, Green Fluorescent Proteins, Molecular Sequence Data, Mutagenesis, Rec A Recombinases, Recombinant Fusion Proteins}, issn = {0021-9193}, doi = {10.1128/JB.01777-06}, author = {Centore, Richard C and Sandler, Steven J} } @article {38, title = {Analysis of Opi1p repressor mutants.}, journal = {Curr Genet}, volume = {49}, year = {2006}, month = {2006 Jan}, pages = {30-8}, abstract = {Opi1p is the only known repressor protein specific to the phospholipid biosynthetic pathway. Opi1p is required for repression in response to inositol and choline supplementation. However, the mechanism of Opi1p repression is not completely understood. In part, this is because previously identified opi1 mutants contained nonsense mutations and thus provided little insight into the mechanism of Opi1p function. We have recently reported isolating novel opi1 mutants (rum and dim mutants) that contain missense mutations. Here, we show that these opi1 mutants produce Opi1p product at levels comparable to a wild-type strain. However, these mutants mis-regulate expression of two target genes, INO2-HIS3 and INO1-lacZ, and are also defective in autoregulation. An opi1-S339F mutant is particularly interesting because it completely eliminated autoregulation, but only abated regulation of an INO1-lacZ reporter. Two of the mutations in OPI1 (V343Q and S339F) provide genetic evidence for an interaction between Opi1p and the Ino2p activator since they reside in a region of Opi1p recently shown to interact with Ino2p in vitro. A third mutation (L252F) resides in a region of Opi1p with no known function.}, keywords = {Amino Acid Sequence, Basic Helix-Loop-Helix Transcription Factors, Conserved Sequence, Molecular Sequence Data, Mutation, Missense, Myo-Inositol-1-Phosphate Synthase, Repressor Proteins, Saccharomyces cerevisiae Proteins, Transcription Factors}, issn = {0172-8083}, doi = {10.1007/s00294-005-0021-6}, author = {Kaadige, Mohan R and Lopes, John M} } @article {514, title = {Biofilm and nanowire production leads to increased current in Geobacter sulfurreducens fuel cells.}, journal = {Appl Environ Microbiol}, volume = {72}, year = {2006}, month = {2006 Nov}, pages = {7345-8}, abstract = {Geobacter sulfurreducens developed highly structured, multilayer biofilms on the anode surface of a microbial fuel cell converting acetate to electricity. Cells at a distance from the anode remained viable, and there was no decrease in the efficiency of current production as the thickness of the biofilm increased. Genetic studies demonstrated that efficient electron transfer through the biofilm required the presence of electrically conductive pili. These pili may represent an electronic network permeating the biofilm that can promote long-range electrical transfer in an energy-efficient manner, increasing electricity production more than 10-fold.}, keywords = {Acetates, Bioelectric Energy Sources, Biofilms, Electricity, Electrodes, Electron Transport, Fimbriae, Bacterial, Geobacter, Microscopy, Confocal, Mutation, Nanowires}, issn = {0099-2240}, doi = {10.1128/AEM.01444-06}, author = {Reguera, Gemma and Nevin, Kelly P and Nicoll, Julie S and Covalla, Sean F and Woodard, Trevor L and Lovley, Derek R} } @article {747, title = {Broad-spectrum antibacterial activity by a novel abiogenic peptide mimic.}, journal = {Microbiology}, volume = {152}, year = {2006}, month = {2006 Jul}, pages = {1913-8}, abstract = {The human-mediated use and abuse of classical antibiotics has created a strong selective pressure for the rapid evolution of antibiotic resistance. As resistance levels rise, and the efficacy of classical antibiotics wanes, the intensity of the search for alternative antimicrobials has increased. One class of molecules that has attracted much attention is the antimicrobial peptides (AMPs). They exhibit broad-spectrum activity, they are potent and they are widespread as part of the innate defence system of both vertebrates and invertebrates. However, peptides are complex molecules that suffer from proteolytic degradation. The ability to capture the essential properties of antimicrobial peptides in simple easy-to-prepare molecules that are abiotic in origin and non-proteolytic offers many advantages. Mechanistic and structural knowledge of existing AMPs was used to design a novel compound that mimics the biochemical activity of an AMP. This report describes the development and in vitro characterization of a small peptide mimic that exhibited quick-acting and selective antibacterial activity against a broad range of bacteria, including numerous clinically relevant strains, at low MIC values.}, keywords = {Anti-Bacterial Agents, Drug Design, Humans, Microbial Sensitivity Tests, Peptides}, issn = {1350-0872}, doi = {10.1099/mic.0.28812-0}, author = {N{\"u}sslein, Klaus and Arnt, Lachelle and Rennie, Jason and Owens, Cullen and Tew, Gregory N} } @article {519, title = {Bug juice: harvesting electricity with microorganisms.}, journal = {Nat Rev Microbiol}, volume = {4}, year = {2006}, month = {2006 Jul}, pages = {497-508}, abstract = {It is well established that some reduced fermentation products or microbially reduced artificial mediators can abiotically react with electrodes to yield a small electrical current. This type of metabolism does not typically result in an efficient conversion of organic compounds to electricity because only some metabolic end products will react with electrodes, and the microorganisms only incompletely oxidize their organic fuels. A new form of microbial respiration has recently been discovered in which microorganisms conserve energy to support growth by oxidizing organic compounds to carbon dioxide with direct quantitative electron transfer to electrodes. These organisms, termed electricigens, offer the possibility of efficiently converting organic compounds into electricity in self-sustaining systems with long-term stability.}, keywords = {Bacteria, Bioelectric Energy Sources, Electricity, Electrodes, Electron Transport, Oxidation-Reduction}, issn = {1740-1526}, doi = {10.1038/nrmicro1442}, author = {Lovley, Derek R} } @article {1214, title = {Characterization of dissimilatory Fe(III) versus NO3- reduction in the hyperthermophilic archaeon Pyrobaculum aerophilum.}, journal = {J Bacteriol}, volume = {188}, year = {2006}, month = {2006 Jan}, pages = {525-31}, abstract = {The hyperthermophilic archaeon Pyrobaculum aerophilum used 20 mM Fe(III) citrate, 100 mM poorly crystalline Fe(III) oxide, and 10 mM KNO3 as terminal electron acceptors. The two forms of iron were reduced at different rates but with equal growth yields. The insoluble iron was reduced when segregated spatially by dialysis tubing, indicating that direct contact with the iron was not necessary for growth. When partitioned, there was no detectable Fe(III) or Fe(II) outside of the tubing after growth, suggesting that an electron shuttle, not a chelator, may be used as an extracellular mediator of iron reduction. The addition of 25 and 50\% (vol vol(-1)) cell-free spent insoluble iron media to fresh media led to growth without a lag phase. Liquid chromatography analysis of spent media showed that cultures grown in iron, especially insoluble iron, produced soluble extracellular compounds that were absent or less abundant in spent nitrate medium. NADH-dependent ferric reductase activity increased approximately 100-fold, while nitrate reductase activity decreased 10-fold in whole-cell extracts from iron-grown cells relative to those from nitrate-grown cells, suggesting that dissimilatory iron reduction was regulated. A novel 2,6-anthrahydroquinone disulfonate oxidase activity was more than 580-fold higher in iron-grown cells than in nitrate-grown cells. The activity was primarily (>95\%) associated with the membrane cellular fraction, but its physiological function is unknown. Nitrate-grown cultures produced two membrane-bound, c-type cytochromes that are predicted to be monoheme and part of nitrite reductase and a bc1 complex using genome analyses. Only one cytochrome was present in cells grown on Fe(III) citrate whose relative abundance was unchanged.
}, keywords = {Cytochromes c, Ferric Compounds, FMN Reductase, Hot Temperature, NAD, Nitrates, Oxidation-Reduction, Pyrobaculum}, issn = {0021-9193}, doi = {10.1128/JB.188.2.525-531.2006}, author = {Feinberg, Lawrence F and Holden, James F} } @article {524, title = {Characterization of metabolism in the Fe(III)-reducing organism Geobacter sulfurreducens by constraint-based modeling.}, journal = {Appl Environ Microbiol}, volume = {72}, year = {2006}, month = {2006 Feb}, pages = {1558-68}, abstract = {Geobacter sulfurreducens is a well-studied representative of the Geobacteraceae, which play a critical role in organic matter oxidation coupled to Fe(III) reduction, bioremediation of groundwater contaminated with organics or metals, and electricity production from waste organic matter. In order to investigate G. sulfurreducens central metabolism and electron transport, a metabolic model which integrated genome-based predictions with available genetic and physiological data was developed via the constraint-based modeling approach. Evaluation of the rates of proton production and consumption in the extracellular and cytoplasmic compartments revealed that energy conservation with extracellular electron acceptors, such as Fe(III), was limited relative to that associated with intracellular acceptors. This limitation was attributed to lack of cytoplasmic proton consumption during reduction of extracellular electron acceptors. Model-based analysis of the metabolic cost of producing an extracellular electron shuttle to promote electron transfer to insoluble Fe(III) oxides demonstrated why Geobacter species, which do not produce shuttles, have an energetic advantage over shuttle-producing Fe(III) reducers in subsurface environments. In silico analysis also revealed that the metabolic network of G. sulfurreducens could synthesize amino acids more efficiently than that of Escherichia coli due to the presence of a pyruvate-ferredoxin oxidoreductase, which catalyzes synthesis of pyruvate from acetate and carbon dioxide in a single step. In silico phenotypic analysis of deletion mutants demonstrated the capability of the model to explore the flexibility of G. sulfurreducens central metabolism and correctly predict mutant phenotypes. These results demonstrate that iterative modeling coupled with experimentation can accelerate the understanding of the physiology of poorly studied but environmentally relevant organisms and may help optimize their practical applications.}, keywords = {Amino Acids, Electron Transport, Escherichia coli, Fumarates, Geobacter, Iron, Models, Biological, Mutation, Oxidation-Reduction, Phenotype, Protons, Quinones, Species Specificity}, issn = {0099-2240}, doi = {10.1128/AEM.72.2.1558-1568.2006}, author = {Mahadevan, R and Bond, D R and Butler, J E and Esteve-Nu{\~n}ez, A and Coppi, M V and Palsson, B O and Schilling, C H and Lovley, D R} } @article {1213, title = {Citric acid cycle in the hyperthermophilic archaeon Pyrobaculum islandicum grown autotrophically, heterotrophically, and mixotrophically with acetate.}, journal = {J Bacteriol}, volume = {188}, year = {2006}, month = {2006 Jun}, pages = {4350-5}, abstract = {The hyperthermophilic archaeon Pyrobaculum islandicum uses the citric acid cycle in the oxidative and reductive directions for heterotrophic and autotrophic growth, respectively, but the control of carbon flow is poorly understood. P. islandicum was grown at 95 degrees C autotrophically, heterotrophically, and mixotrophically with acetate, H2, and small amounts of yeast extract and with thiosulfate as the terminal electron acceptor. The autotrophic growth rates and maximum concentrations of cells were significantly lower than those in other media. The growth rates on H2 and 0.001\% yeast extract with and without 0.05\% acetate were the same, but the maximum concentration of cells was fourfold higher with acetate. There was no growth with acetate if 0.001\% yeast extract was not present, and addition of H2 to acetate-containing medium greatly increased the growth rates and maximum concentrations of cells. P. islandicum cultures assimilated 14C-labeled acetate in the presence of H2 and yeast extract with an efficiency of 55\%. The activities of 11 of 19 enzymes involved in the central metabolism of P. islandicum were regulated under the three different growth conditions. Pyruvate synthase and acetate:coenzyme A (CoA) ligase (ADP-forming) activities were detected only in heterotrophically grown cultures. Citrate synthase activity decreased in autotrophic and acetate-containing cultures compared to the activity in heterotrophic cultures. Acetylated citrate lyase, acetate:CoA ligase (AMP forming), and phosphoenolpyruvate carboxylase activities increased in autotrophic and acetate-containing cultures. Citrate lyase activity was higher than ATP citrate synthase activity in autotrophic cultures. These data suggest that citrate lyase and AMP-forming acetate:CoA ligase, but not ATP citrate synthase, work opposite citrate synthase to control the direction of carbon flow in the citric acid cycle.
}, keywords = {Acetates, Carbon Dioxide, Citric Acid, Citric Acid Cycle, Culture Media, Deuterium, Ligases, Lyases, Pyrobaculum, Pyruvate Synthase, Substrate Specificity}, issn = {0021-9193}, doi = {10.1128/JB.00138-06}, author = {Hu, Yajing and Holden, James F} } @article {512, title = {Computational prediction of RpoS and RpoD regulatory sites in Geobacter sulfurreducens using sequence and gene expression information.}, journal = {Gene}, volume = {384}, year = {2006}, month = {2006 Dec 15}, pages = {73-95}, abstract = {RpoS, the sigma S subunit of RNA polymerase, is vital during the growth and survival of Geobacter sulfurreducens under conditions typically encountered in its native subsurface environments. We investigated the conservation of sites that may be important for RpoS function in G. sulfurreducens. We also employed sequence information and expression microarray data to predict G. sulfurreducens genome sites that may be related to RpoS regulation. Hierarchical clustering identified three clusters of significantly downregulated genes in the rpoS deletion mutant. The search for conserved overrepresented motifs in co-regulated operons identified likely -35 and -10 promoter elements upstream of a number of functionally important G. sulfurreducens operons that were downregulated in the rpoS deletion mutant. Putative -35/-10 promoter elements were also identified in the G. sulfurreducens genome using sequence similarity searches to matrices of -35/-10 promoter elements found in G. sulfurreducens and in Escherichia coli. Due to a sufficient degree of sequence similarity between -35/-10 promoter elements for RpoS, RpoD, and other sigma factors, both the sequence similarity searches and the search for conserved overrepresented motifs using microarray data may identify promoter elements for both RpoS and other sigma factors.}, keywords = {Amino Acid Sequence, Bacterial Proteins, Base Sequence, Citrates, Computational Biology, Conserved Sequence, DNA-Directed RNA Polymerases, Escherichia coli, Escherichia coli Proteins, Gene Deletion, Gene Expression Profiling, Gene Expression Regulation, Bacterial, Genes, Bacterial, Genome, Bacterial, Geobacter, Molecular Sequence Data, Oligonucleotide Array Sequence Analysis, Operon, Promoter Regions, Genetic, Sigma Factor, Transcription, Genetic}, issn = {0378-1119}, doi = {10.1016/j.gene.2006.06.025}, author = {Yan, Bin and N{\'u}{\~n}ez, Cinthia and Ueki, Toshiyuki and Esteve-N{\'u}{\~n}ez, Abraham and Puljic, Marko and Adkins, Ronald M and Meth{\'e}, Barbara A and Lovley, Derek R and Krushkal, Julia} } @article {515, title = {c-Type cytochromes in Pelobacter carbinolicus.}, journal = {Appl Environ Microbiol}, volume = {72}, year = {2006}, month = {2006 Nov}, pages = {6980-5}, abstract = {Previous studies failed to detect c-type cytochromes in Pelobacter species despite the fact that other close relatives in the Geobacteraceae, such as Geobacter and Desulfuromonas species, have abundant c-type cytochromes. Analysis of the recently completed genome sequence of Pelobacter carbinolicus revealed 14 open reading frames that could encode c-type cytochromes. Transcripts for all but one of these open reading frames were detected in acetoin-fermenting and/or Fe(III)-reducing cells. Three putative c-type cytochrome genes were expressed specifically during Fe(III) reduction, suggesting that the encoded proteins may participate in electron transfer to Fe(III). One of these proteins was a periplasmic triheme cytochrome with a high level of similarity to PpcA, which has a role in Fe(III) reduction in Geobacter sulfurreducens. Genes for heme biosynthesis and system II cytochrome c biogenesis were identified in the genome and shown to be expressed. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis gels of protein extracted from acetoin-fermenting P. carbinolicus cells contained three heme-staining bands which were confirmed by mass spectrometry to be among the 14 predicted c-type cytochromes. The number of cytochrome genes, the predicted amount of heme c per protein, and the ratio of heme-stained protein to total protein were much smaller in P. carbinolicus than in G. sulfurreducens. Furthermore, many of the c-type cytochromes that genetic studies have indicated are required for optimal Fe(III) reduction in G. sulfurreducens were not present in the P. carbinolicus genome. These results suggest that further evaluation of the functions of c-type cytochromes in the Geobacteraceae is warranted.}, keywords = {Bacterial Proteins, Cytochromes c, Deltaproteobacteria, Heme, Polymerase Chain Reaction, Proteomics, Reverse Transcriptase Polymerase Chain Reaction, RNA, Messenger}, issn = {0099-2240}, doi = {10.1128/AEM.01128-06}, author = {Haveman, Shelley A and Holmes, Dawn E and Ding, Yan-Huai R and Ward, Joy E and Didonato, Raymond J and Lovley, Derek R} } @article {749, title = {Desiccation responses and survival of Sinorhizobium meliloti USDA 1021 in relation to growth phase, temperature, chloride and sulfate availability.}, journal = {Lett Appl Microbiol}, volume = {42}, year = {2006}, month = {2006 Feb}, pages = {172-8}, abstract = {AIMS: To identify physical and physiological conditions that affect the survival of Sinorhizobium meliloti USDA 1021 during desiccation. METHODS AND RESULTS: An assay was developed to study desiccation response of S. meliloti USDA 1021 over a range of environmental conditions. We determined the survival during desiccation in relation to (i) matrices and media, (ii) growth phase, (iii) temperature, and (iv) chloride and sulfate availability. CONCLUSIONS: This study indicates that survival of S. meliloti USDA 1021 during desiccation is enhanced: (i) when cells were dried in the stationary phase, (ii) with increasing drying temperature at an optimum of 37 degrees C, and (iii) during an increase of chloride and sulfate, but not sodium or potassium availability. In addition, we resolved that the best matrix to test survival was nitrocellulose filters. SIGNIFICANCE AND IMPACT OF THE STUDY: The identification of physical and physiological factors that determine the survival during desiccation of S. meliloti USDA 1021 may aid in (i) the strategic development of improved seed inocula, (ii) the isolation, and (iii) the development of rhizobial strains with improved ability to survive desiccation. Furthermore, this work may provide insights into the survival of rhizobia under drought conditions.}, keywords = {Bacteriological Techniques, Chlorides, Desiccation, Models, Biological, Sinorhizobium meliloti, Sulfur Oxides, Temperature}, issn = {0266-8254}, doi = {10.1111/j.1472-765X.2005.01808.x}, author = {Vriezen, J A C and de Bruijn, F J and N{\"u}sslein, K} } @article {855, title = {Detection of Chlamydia in the peripheral blood cells of normal donors using in vitro culture, immunofluorescence microscopy and flow cytometry techniques.}, journal = {BMC Infect Dis}, volume = {6}, year = {2006}, month = {2006}, pages = {23}, abstract = {BACKGROUND: Chlamydia trachomatis (Ct) and Chlamydia pneumoniae (Cp) are medically significant infectious agents associated with various chronic human pathologies. Nevertheless, specific roles in disease progression or initiation are incompletely defined. Both pathogens infect established cell lines in vitro and polymerase chain reaction (PCR) has detected Chlamydia DNA in various clinical specimens as well as in normal donor peripheral blood monocytes (PBMC). However, Chlamydia infection of other blood cell types, quantification of Chlamydia infected cells in peripheral blood and transmission of this infection in vitro have not been examined. METHODS: Cp specific titers were assessed for sera from 459 normal human donor blood (NBD) samples. Isolated white blood cells (WBC) were assayed by in vitro culture to evaluate infection transmission of blood cell borne chlamydiae. Smears of fresh blood samples (FB) were dual immunostained for microscopic identification of Chlamydia-infected cell types and aliquots also assessed using Flow Cytometry (FC). RESULTS: ELISA demonstrated that 219 (47.7\%) of the NBD samples exhibit elevated anti-Cp antibody titers. Imunofluorescence microscopy of smears demonstrated 113 (24.6\%) of samples contained intracellular Chlamydia and monoclonals to specific CD markers showed that in vivo infection of neutrophil and eosinophil/basophil cells as well as monocytes occurs. In vitro culture established WBCs of 114 (24.8\%) of the NBD samples harbored infectious chlamydiae, clinically a potentially source of transmission, FC demonstrated both Chlamydia infected and uninfected cells can be readily identified and quantified. CONCLUSION: NBD can harbor infected neutrophils, eosinophil/basophils and monocytes. The chlamydiae are infectious in vitro, and both total, and cell type specific Chlamydia carriage is quantifiable by FC.}, keywords = {Adult, Blood Donors, Chlamydophila pneumoniae, Female, Flow Cytometry, Humans, Leukocytes, Male, Microscopy, Fluorescence}, issn = {1471-2334}, doi = {10.1186/1471-2334-6-23}, author = {Cirino, Frances and Webley, Wilmore C and West, Corrie and Croteau, Nancy L and Andrzejewski, Chester and Stuart, Elizabeth S} } @article {702, title = {Differential protein expression in the metal-reducing bacterium Geobacter sulfurreducens strain PCA grown with fumarate or ferric citrate.}, journal = {Proteomics}, volume = {6}, year = {2006}, month = {2006 Jan}, pages = {632-40}, abstract = {Geobacter sulfurreducens, generally considered to be a strict anaerobe, is a predominant microbe in subsurface environments, where it utilizes available metals as electron acceptors. To better understand the metabolic processes involved in the metal-reduction capability of this microbe, the proteins expressed by cells grown anaerobically with either fumarate or ferric citrate as electron acceptor were compared. Proteins were separated by 2-DE under denaturing or nondenaturing conditions, and proteins varying in abundance with a high level of statistical significance (p<0.0001) were identified by peptide mass analysis. Denaturing 2-DE revealed significant differences in the relative abundance of the membrane proteins OmpA and peptidoglycan-associated lipoprotein, several metabolic enzymes, and, in addition, superoxide dismutase and rubredoxin oxidoreductase. Nondenaturing 2-DE revealed elevated catalase in cells grown with ferric citrate. These results suggest that, in addition to adjustments in membrane transport and specific metabolic pathways in response to these two different electron acceptors, distinct differences exist in the oxidative environment within the cell when fumarate or soluble ferric citrate is provided as electron acceptor. Although an anaerobe, G. sulfurreducens appears to have alternate mechanisms for dealing with reactive oxygen species in response to increased intracellular soluble iron.}, keywords = {Anaerobiosis, Bacterial Proteins, Electrophoresis, Gel, Two-Dimensional, Ferric Compounds, Fumarates, Geobacter, Oxidation-Reduction, Proteomics, Reactive Oxygen Species, Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization}, issn = {1615-9853}, doi = {10.1002/pmic.200500137}, author = {Khare, Tripti and Esteve-N{\'u}{\~n}ez, Abraham and Nevin, Kelly P and Zhu, Wenhong and Yates, John R and Lovley, Derek and Giometti, Carol S} } @article {748, title = {Distribution of extensive nifH gene diversity across physical soil microenvironments.}, journal = {Microb Ecol}, volume = {51}, year = {2006}, month = {2006 May}, pages = {441-52}, abstract = {The diversity of nitrogen-fixing bacteria is well described for aquatic environments; however, terrestrial analyses remain mostly biased to rhizobial plant-microbe associations. We maximized the level of resolution for this study through the use of nucleotide sequence information extracted from a series of soil microenvironments, ranging from macroaggregates at 2000 microm to the clay fraction at < 75 microm in diameter. In addition, we attempted to create an overview of the distribution of terrestrial nitrogen fixers across such microenvironments by combining culture-independent techniques with a suite of natural soil environments from uniquely different origins. Soil diazotroph diversity was analyzed phylogenetically for 600 terrestrial nifH sequences from 12 midsized clone libraries based on microenvironments of three separate soils across a global scale. Statistical analyses of nifH gene clone libraries were used to estimate coverage, establish degrees of sequence overlap, and compare cluster distributions. These analyses revealed an extensive diversity in a tropical (19 phylotypes) and an arctic soil (17 phylotypes), and moderate diversity in a temperate soil (11 phylotypes). Within each soil, comparisons across aggregate size fractions delineated nifH gene cluster shifts within populations and degrees of sequence overlap that ranged from significantly different (arctic, tropical) to significantly similar (temperate). We suggest that this is due to population separation across aggregates of different size classes, which results from differences in the temporal stability of aggregates as niches for microbial communities. This study not only provides new knowledge of the arrangement of diazotrophic communities at the soil microscale, but it also contributes to the underrepresented knowledge of soil nifH sequences in the public databases.}, keywords = {Bacteria, Base Sequence, DNA Primers, Genes, Bacterial, Multigene Family, Oxidoreductases, Phylogeny, Polymerase Chain Reaction, Soil Microbiology}, issn = {0095-3628}, doi = {10.1007/s00248-006-9044-x}, author = {Izquierdo, Javier A and N{\"u}sslein, Klaus} } @article {522, title = {DNA microarray and proteomic analyses of the RpoS regulon in Geobacter sulfurreducens.}, journal = {J Bacteriol}, volume = {188}, year = {2006}, month = {2006 Apr}, pages = {2792-800}, abstract = {The regulon of the sigma factor RpoS was defined in Geobacter sulfurreducens by using a combination of DNA microarray expression profiles and proteomics. An rpoS mutant was examined under steady-state conditions with acetate as an electron donor and fumarate as an electron acceptor and with additional transcriptional profiling using Fe(III) as an electron acceptor. Expression analysis revealed that RpoS acts as both a positive and negative regulator. Many of the RpoS-dependent genes determined play roles in energy metabolism, including the tricarboxylic acid cycle, signal transduction, transport, protein synthesis and degradation, and amino acid metabolism and transport. As expected, RpoS activated genes involved in oxidative stress resistance and adaptation to nutrient limitation. Transcription of the cytochrome c oxidase operon, necessary for G. sulfurreducens growth using oxygen as an electron acceptor, and expression of at least 13 c-type cytochromes, including one previously shown to participate in Fe(III) reduction (MacA), were RpoS dependent. Analysis of a subset of the rpoS mutant proteome indicated that 15 major protein species showed reproducible differences in abundance relative to those of the wild-type strain. Protein identification using mass spectrometry indicated that the expression of seven of these proteins correlated with the microarray data. Collectively, these results indicate that RpoS exerts global effects on G. sulfurreducens physiology and that RpoS is vital to G. sulfurreducens survival under conditions typically encountered in its native subsurface environments.}, keywords = {Adaptation, Physiological, Amino Acids, Bacterial Proteins, Biological Transport, Citric Acid Cycle, Cytochromes, Electrophoresis, Gel, Two-Dimensional, Gene Deletion, Gene Expression Regulation, Bacterial, Geobacter, Mass Spectrometry, Mutagenesis, Insertional, Oligonucleotide Array Sequence Analysis, Oxidative Stress, Protein Biosynthesis, Proteome, Regulon, Sigma Factor, Signal Transduction}, issn = {0021-9193}, doi = {10.1128/JB.188.8.2792-2800.2006}, author = {N{\'u}{\~n}ez, Cinthia and Esteve-N{\'u}{\~n}ez, Abraham and Giometti, Carol and Tollaksen, Sandra and Khare, Tripti and Lin, Winston and Lovley, Derek R and Meth{\'e}, Barbara A} } @article {525, title = {Genetic characterization of a single bifunctional enzyme for fumarate reduction and succinate oxidation in Geobacter sulfurreducens and engineering of fumarate reduction in Geobacter metallireducens.}, journal = {J Bacteriol}, volume = {188}, year = {2006}, month = {2006 Jan}, pages = {450-5}, abstract = {The mechanism of fumarate reduction in Geobacter sulfurreducens was investigated. The genome contained genes encoding a heterotrimeric fumarate reductase, FrdCAB, with homology to the fumarate reductase of Wolinella succinogenes and the succinate dehydrogenase of Bacillus subtilis. Mutation of the putative catalytic subunit of the enzyme resulted in a strain that lacked fumarate reductase activity and was unable to grow with fumarate as the terminal electron acceptor. The mutant strain also lacked succinate dehydrogenase activity and did not grow with acetate as the electron donor and Fe(III) as the electron acceptor. The mutant strain could grow with acetate as the electron donor and Fe(III) as the electron acceptor if fumarate was provided to alleviate the need for succinate dehydrogenase activity in the tricarboxylic acid cycle. The growth rate of the mutant strain under these conditions was faster and the cell yields were higher than for wild type grown under conditions requiring succinate dehydrogenase activity, suggesting that the succinate dehydrogenase reaction consumes energy. An orthologous frdCAB operon was present in Geobacter metallireducens, which cannot grow with fumarate as the terminal electron acceptor. When a putative dicarboxylic acid transporter from G. sulfurreducens was expressed in G. metallireducens, growth with fumarate as the sole electron acceptor was possible. These results demonstrate that, unlike previously described organisms, G. sulfurreducens and possibly G. metallireducens use the same enzyme for both fumarate reduction and succinate oxidation in vivo.}, keywords = {Culture Media, Dicarboxylic Acids, Fumarates, Geobacter, Molecular Sequence Data, Operon, Oxidation-Reduction, Recombinant Proteins, Substrate Specificity, Succinate Dehydrogenase, Succinic Acid}, issn = {0021-9193}, doi = {10.1128/JB.188.2.450-455.2006}, author = {Butler, Jessica E and Glaven, Richard H and Esteve-N{\'u}{\~n}ez, Abraham and N{\'u}{\~n}ez, Cinthia and Shelobolina, Evgenya S and Bond, Daniel R and Lovley, Derek R} } @article {37, title = {Genomic analysis of the Opi- phenotype.}, journal = {Genetics}, volume = {173}, year = {2006}, month = {2006 Jun}, pages = {621-34}, abstract = {Most of the phospholipid biosynthetic genes of Saccharomyces cerevisiae are coordinately regulated in response to inositol and choline. Inositol affects the intracellular levels of phosphatidic acid (PA). Opi1p is a repressor of the phospholipid biosynthetic genes and specifically binds PA in the endoplasmic reticulum. In the presence of inositol, PA levels decrease, releasing Opi1p into the nucleus where it represses transcription. The opi1 mutant overproduces and excretes inositol into the growth medium in the absence of inositol and choline (Opi(-) phenotype). To better understand the mechanism of Opi1p repression, the viable yeast deletion set was screened to identify Opi(-) mutants. In total, 89 Opi(-) mutants were identified, of which 7 were previously known to have the Opi(-) phenotype. The Opi(-) mutant collection included genes with roles in phospholipid biosynthesis, transcription, protein processing/synthesis, and protein trafficking. Included in this set were all nonessential components of the NuA4 HAT complex and six proteins in the Rpd3p-Sin3p HDAC complex. It has previously been shown that defects in phosphatidylcholine synthesis (cho2 and opi3) yield the Opi(-) phenotype because of a buildup of PA. However, in this case the Opi(-) phenotype is conditional because PA can be shuttled through a salvage pathway (Kennedy pathway) by adding choline to the growth medium. Seven new mutants present in the Opi(-) collection (fun26, kex1, nup84, tps1, mrpl38, mrpl49, and opi10/yol032w) were also suppressed by choline, suggesting that these affect PC synthesis. Regulation in response to inositol is also coordinated with the unfolded protein response (UPR). Consistent with this, several Opi(-) mutants were found to affect the UPR (yhi9, ede1, and vps74).}, keywords = {Gene Expression Regulation, Fungal, Genes, Fungal, Genes, Reporter, Genome, Fungal, Genomics, Inositol, Lac Operon, Models, Biological, Mutation, Phenotype, Phospholipids, Protein Folding, Repressor Proteins, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins}, issn = {0016-6731}, doi = {10.1534/genetics.106.057489}, author = {Hancock, Leandria C and Behta, Ryan P and Lopes, John M} } @article {523, title = {Harvesting energy from the marine sediment-water interface II. Kinetic activity of anode materials.}, journal = {Biosens Bioelectron}, volume = {21}, year = {2006}, month = {2006 May 15}, pages = {2058-63}, abstract = {Here, we report a comparative study on the kinetic activity of various anodes of a recently described microbial fuel cell consisting of an anode imbedded in marine sediment and a cathode in overlying seawater. Using plain graphite anodes, it was demonstrated that a significant portion of the anodic current results from oxidation of sediment organic matter catalyzed by microorganisms colonizing the anode and capable of directly reducing the anode without added exogenous electron-transfer mediators. Here, graphite anodes incorporating microbial oxidants are evaluated in the laboratory relative to plain graphite with the goal of increasing power density by increasing current density. Anodes evaluated include graphite modified by adsorption of anthraquinone-1,6-disulfonic acid (AQDS) or 1,4-naphthoquinone (NQ), a graphite-ceramic composite containing Mn2+ and Ni2+, and graphite modified with a graphite paste containing Fe3O4 or Fe3O4 and Ni2+. It was found that these anodes possess between 1.5- and 2.2-fold greater kinetic activity than plain graphite. Fuel cells were deployed in a coastal site near Tuckerton, NJ (USA) that utilized two of these anodes. These fuel cells generated ca. 5-fold greater current density than a previously characterized fuel cell equipped with a plain graphite anode, and operated at the same site.}, keywords = {Electrochemistry, Electrodes, Energy-Generating Resources, Ferumoxytol, Geologic Sediments, Kinetics, Oceans and Seas, Seawater}, issn = {0956-5663}, doi = {10.1016/j.bios.2006.01.033}, author = {Lowy, Daniel A and Tender, Leonard M and Zeikus, J Gregory and Park, Doo Hyun and Lovley, Derek R} } @article {694, title = {Identification of a novel protein with a role in lipoarabinomannan biosynthesis in mycobacteria.}, journal = {J Biol Chem}, volume = {281}, year = {2006}, month = {2006 Apr 7}, pages = {9011-7}, abstract = {All species of Mycobacteria synthesize distinctive cell walls that are rich in phosphatidylinositol mannosides (PIMs), lipomannan (LM), and lipoarabinomannan (LAM). PIM glycolipids, having 2-4 mannose residues, can either be channeled into polar PIM species (with 6 Man residues) or hypermannosylated to form LM and LAM. In this study, we have identified a Mycobacterium smegmatis gene, termed lpqW, that is required for the conversion of PIMs to LAM and is highly conserved in all mycobacteria. A transposon mutant, Myco481, containing an insertion near the 3{\textquoteright} end of lpqW exhibited altered colony morphology on complex agar medium. This mutant was unstable and was consistently overgrown by a second mutant, represented by Myco481.1, that had normal growth and colony characteristics. Biochemical analysis and metabolic labeling studies showed that Myco481 synthesized the complete spectrum of apolar and polar PIMs but was unable to make LAM. LAM biosynthesis was restored to near wild type levels in Myco481.1. However, this mutant was unable to synthesize the major polar PIM (AcPIM6) and accumulated a smaller intermediate, AcPIM4. Targeted disruption of the lpqW gene and complementation of the initial Myco481 mutant with the wild type gene confirmed that the phenotype of this mutant was due to loss of LpqW. These studies suggest that LpqW has a role in regulating the flux of early PIM intermediates into polar PIM or LAM biosynthesis. They also suggest that AcPIM4 is the likely branch point intermediate in polar PIM and LAM biosynthesis.}, keywords = {Cell Membrane, Gene Expression Regulation, Bacterial, Genes, Bacterial, Lipopolysaccharides, Mutation, Mycobacterium smegmatis, Phenotype, Phosphatidylinositols, Virulence}, issn = {0021-9258}, doi = {10.1074/jbc.M511709200}, author = {Kovacevic, Svetozar and Anderson, Dianne and Morita, Yasu S and Patterson, John and Haites, Ruth and McMillan, Benjamin N I and Coppel, Ross and McConville, Malcolm J and Billman-Jacobe, Helen} } @article {513, title = {Microarray and genetic analysis of electron transfer to electrodes in Geobacter sulfurreducens.}, journal = {Environ Microbiol}, volume = {8}, year = {2006}, month = {2006 Oct}, pages = {1805-15}, abstract = {Whole-genome analysis of gene expression in Geobacter sulfurreducens revealed 474 genes with transcript levels that were significantly different during growth with an electrode as the sole electron acceptor versus growth on Fe(III) citrate. The greatest response was a more than 19-fold increase in transcript levels for omcS, which encodes an outer-membrane cytochrome previously shown to be required for Fe(III) oxide reduction. Quantitative reverse transcription polymerase chain reaction and Northern analyses confirmed the higher levels of omcS transcripts, which increased as power production increased. Deletion of omcS inhibited current production that was restored when omcS was expressed in trans. Transcript expression and genetic analysis suggested that OmcE, another outer-membrane cytochrome, is also involved in electron transfer to electrodes. Surprisingly, genes for other proteins known to be important in Fe(III) reduction such as the outer-membrane c-type cytochrome, OmcB, and the electrically conductive pilin "nanowires" did not have higher transcript levels on electrodes, and deletion of the relevant genes did not inhibit power production. Changes in the transcriptome suggested that cells growing on electrodes were subjected to less oxidative stress than cells growing on Fe(III) citrate and that a number of genes annotated as encoding metal efflux proteins or proteins of unknown function may be important for growth on electrodes. These results demonstrate for the first time that it is possible to evaluate gene expression, and hence the metabolic state, of microorganisms growing on electrodes on a genome-wide basis and suggest that OmcS, and to a lesser extent OmcE, are important in electron transfer to electrodes. This has important implications for the design of electrode materials and the genetic engineering of microorganisms to improve the function of microbial fuel cells.}, keywords = {Bacterial Outer Membrane Proteins, Blotting, Northern, Cytochromes c, Electrodes, Electrophysiology, Gene Expression Regulation, Bacterial, Geobacter, Oligonucleotide Array Sequence Analysis, Oxidation-Reduction, Reverse Transcriptase Polymerase Chain Reaction, RNA, Bacterial, RNA, Messenger}, issn = {1462-2912}, doi = {10.1111/j.1462-2920.2006.01065.x}, author = {Holmes, Dawn E and Chaudhuri, Swades K and Nevin, Kelly P and Mehta, Teena and Meth{\'e}, Barbara A and Liu, Anna and Ward, Joy E and Woodard, Trevor L and Webster, Jennifer and Lovley, Derek R} } @article {520, title = {Microbial fuel cells: novel microbial physiologies and engineering approaches.}, journal = {Curr Opin Biotechnol}, volume = {17}, year = {2006}, month = {2006 Jun}, pages = {327-32}, abstract = {The possibility of generating electricity with microbial fuel cells has been recognized for some time, but practical applications have been slow to develop. The recent development of a microbial fuel cell that can harvest electricity from the organic matter stored in marine sediments has demonstrated the feasibility of producing useful amounts of electricity in remote environments. Further study of these systems has led to the discovery of microorganisms that conserve energy to support their growth by completely oxidizing organic compounds to carbon dioxide with direct electron transfer to electrodes. This suggests that self-sustaining microbial fuel cells that can effectively convert a diverse range of waste organic matter or renewable biomass to electricity are feasible. Significant progress has recently been made to increase the power output of systems designed to convert organic wastes to electricity, but substantial additional optimization will be required for large-scale electricity production.}, keywords = {Bacterial Physiological Phenomena, Bioelectric Energy Sources, Biomass, Biosensing Techniques, Genetic Engineering}, issn = {0958-1669}, doi = {10.1016/j.copbio.2006.04.006}, author = {Lovley, Derek R} } @article {819, title = {A novel dnaC mutation that suppresses priB rep mutant phenotypes in Escherichia coli K-12.}, journal = {Mol Microbiol}, volume = {60}, year = {2006}, month = {2006 May}, pages = {973-83}, abstract = {The loading of a replisome in prokaryotic and eukaryotic cells at an origin of DNA replication and during replication restart is a highly ordered and regulated process. During replication restart in Escherichia coli, the PriA, PriB, PriC, DnaT and Rep proteins form multiple pathways that bind to repaired replication forks. These complexes are then recognized by DnaC as sites to load DnaB, the replicative helicase. Several dnaC mutations have been isolated that suppress phenotypes of some replication restart mutants. A new dnaC mutation (dnaC824) is reported here that efficiently suppresses priB rep mutant phenotypes. Furthermore, it is shown that dnaC824 will suppress phenotypes of priB priA300, rep priA300 and priB priC strains. Unlike other dnaC suppressors, it can only weakly suppress the absence of priA. Others have reported a different type of dnaC mutation, dnaC1331, is able to mimic priB mutant phenotypes. This is supported herein by showing that like dnaC1331, a priB mutation is synthetically lethal with a dam mutation and this can be rescued by a mutH mutation. Furthermore, priB dam lethality can also be suppressed by dnaC824. Like a priB mutation, a dnaC1331 mutation causes a priA2::kan-like phenotype when combined with priA300. Lastly, we show that dnaC824 is dominant to wild type and that dnaC1331 is recessive to wild type. Several models are discussed for the action of these mutant dnaC proteins in replication restart.}, keywords = {DNA Replication, DNA-Binding Proteins, Escherichia coli, Escherichia coli Proteins, Genes, Dominant, Genes, Lethal, Genes, Recessive, Mutant Proteins, Mutation, Phenotype, Suppression, Genetic}, issn = {0950-382X}, doi = {10.1111/j.1365-2958.2006.05147.x}, author = {Boonsombat, Ruethairat and Yeh, Su-Ping and Milne, Amy and Sandler, Steven J} } @article {692, title = {PimE is a polyprenol-phosphate-mannose-dependent mannosyltransferase that transfers the fifth mannose of phosphatidylinositol mannoside in mycobacteria.}, journal = {J Biol Chem}, volume = {281}, year = {2006}, month = {2006 Sep 1}, pages = {25143-55}, abstract = {Phosphatidylinositol mannosides (PIMs) are a major class of glycolipids in all mycobacteria. AcPIM2, a dimannosyl PIM, is both an end product and a precursor for polar PIMs, such as hexamannosyl PIM (AcPIM6) and the major cell wall lipoglycan, lipoarabinomannan (LAM). The mannosyltransferases that convert AcPIM2 to AcPIM6 or LAM are dependent on polyprenol-phosphate-mannose (PPM), but have not yet been characterized. Here, we identified a gene, termed pimE that is present in all mycobacteria, and is required for AcPIM6 biosynthesis. PimE was initially identified based on homology with eukaryotic PIG-M mannosyltransferases. PimE-deleted Mycobacterium smegmatis was defective in AcPIM6 synthesis, and accumulated the tetramannosyl PIM, AcPIM4. Loss of PimE had no affect on cell growth or viability, or the biosynthesis of other intracellular and cell wall glycans. However, changes in cell wall hydrophobicity and plasma membrane organization were detected, suggesting a role for AcPIM6 in the structural integrity of the cell wall and plasma membrane. These defects were corrected by ectopic expression of the pimE gene. Metabolic pulse-chase radiolabeling and cell-free PIM biosynthesis assays indicated that PimE catalyzes the alpha1,2-mannosyl transfer for the AcPIM5 synthesis. Mutation of an Asp residue in PimE that is conserved in and required for the activity of human PIG-M resulted in loss of PIM-biosynthetic activity, indicating that PimE is the catalytic component. Finally, PimE was localized to a distinct membrane fraction enriched in AcPIM4-6 biosynthesis. Taken together, PimE represents the first PPM-dependent mannosyl-transferase shown to be involved in PIM biosynthesis, where it mediates the fifth mannose transfer.}, keywords = {Amino Acid Sequence, Cell Proliferation, Cell Wall, Cell-Free System, Genome, Bacterial, Humans, Mannose, Mannosides, Mannosyltransferases, Molecular Sequence Data, Mycobacterium, Mycobacterium smegmatis, Phosphates, Phosphatidylinositols, Sequence Homology, Amino Acid}, issn = {0021-9258}, doi = {10.1074/jbc.M604214200}, author = {Morita, Yasu S and Sena, Chubert B C and Waller, Ross F and Kurokawa, Ken and Sernee, M Fleur and Nakatani, Fumiki and Haites, Ruth E and Billman-Jacobe, Helen and McConville, Malcolm J and Maeda, Yusuke and Kinoshita, Taroh} } @article {518, title = {The proteome of dissimilatory metal-reducing microorganism Geobacter sulfurreducens under various growth conditions.}, journal = {Biochim Biophys Acta}, volume = {1764}, year = {2006}, month = {2006 Jul}, pages = {1198-206}, abstract = {The proteome of Geobacter sulfurreducens, a model for the Geobacter species that predominate in many Fe(III)-reducing subsurface environments, was characterized with ultra high-pressure liquid chromatography and mass spectrometry using accurate mass and time (AMT) tags as well as with more traditional two-dimensional polyacrylamide gel electrophoresis (2-D PAGE). Cells were grown under six different growth conditions in order to enhance the potential that a wide range of genes would be expressed. The AMT tag approach was able to identify a much greater number of proteins than could be detected with the 2-D PAGE approach. With the AMT approach over 3,000 gene products were identified, representing about 90\% of the total predicted gene products in the genome. A high proportion of predicted proteins in most protein role categories were detected; the highest number of proteins was identified in the hypothetical protein role category. Furthermore, 91 c-type cytochromes of 111 predicted genes in the G. sulfurreducens genome were identified. Differences in the abundance of cytochromes and other proteins under different growth conditions provided information for future functional analysis of these proteins. These results demonstrate that a high percentage of the predicted proteins in the G. sulfurreducens genome are produced and that the AMT tag approach provides a rapid method for comparing differential expression of proteins under different growth conditions in this organism.}, keywords = {Bacterial Proteins, Bacteriological Techniques, Chromatography, High Pressure Liquid, Cytochrome c Group, Electrophoresis, Gel, Two-Dimensional, Ferric Compounds, Fumarates, Geobacter, Peptide Fragments, Proteome, Spectrometry, Mass, Electrospray Ionization}, issn = {0006-3002}, doi = {10.1016/j.bbapap.2006.04.017}, author = {Ding, Yan-Huai R and Hixson, Kim K and Giometti, Carol S and Stanley, Ann and Esteve-N{\'u}{\~n}ez, Abraham and Khare, Tripti and Tollaksen, Sandra L and Zhu, Wenhong and Adkins, Joshua N and Lipton, Mary S and Smith, Richard D and Mester, T{\"u}nde and Lovley, Derek R} } @article {517, title = {A putative multicopper protein secreted by an atypical type II secretion system involved in the reduction of insoluble electron acceptors in Geobacter sulfurreducens.}, journal = {Microbiology}, volume = {152}, year = {2006}, month = {2006 Aug}, pages = {2257-64}, abstract = {Extracellular electron transfer onto Fe(III) oxides in Geobacter sulfurreducens is considered to require proteins that must be exported to the outer surface of the cell. In order to investigate this, the putative gene for OxpG, the pseudopilin involved in a type II general secretion pathway of Gram-negative bacteria, was deleted. The mutant was unable to grow with insoluble Fe(III) oxide as the electron acceptor. Growth on soluble Fe(III) was not affected. An analysis of proteins that accumulated in the periplasm of the oxpG mutant, but not in the wild-type, led to the identification of a secreted protein, OmpB. OmpB is predicted to be a multicopper protein, with highest homology to the manganese oxidase, MofA, from Leptothrix discophora. OmpB contains a potential Fe(III)-binding site and a fibronectin type III domain, suggesting a possible role for this protein in accessing Fe(III) oxides. OmpB was localized to the membrane fraction of G. sulfurreducens and in the supernatant of growing cultures, consistent with the type II secretion system exporting OmpB. A mutant in which ompB was deleted had the same phenotype as the oxpG mutant, suggesting that the failure to export OmpB was responsible for the inability of the oxpG-deficient mutant to reduce Fe(III) oxide. This is the first report that proposes a role for a multicopper oxidase-like protein in an anaerobic organism. These results further emphasize the importance of outer-membrane proteins in Fe(III) oxide reduction and suggest that outer-membrane proteins other than c-type cytochromes are required for Fe(III) oxide reduction in Geobacter species.}, keywords = {Bacterial Outer Membrane Proteins, Electron Transport, Ferric Compounds, Fimbriae Proteins, Geobacter, Manganese Compounds, Mutation, Oxidation-Reduction, Oxides}, issn = {1350-0872}, doi = {10.1099/mic.0.28864-0}, author = {Mehta, Teena and Childers, Susan E and Glaven, Richard and Lovley, Derek R and Mester, T{\"u}nde} } @article {693, title = {Removal or maintenance of inositol-linked acyl chain in glycosylphosphatidylinositol is critical in trypanosome life cycle.}, journal = {J Biol Chem}, volume = {281}, year = {2006}, month = {2006 Apr 28}, pages = {11595-602}, abstract = {The protozoan parasite Trypanosoma brucei is coated by glycosylphosphatidylinositol (GPI)-anchored proteins. During GPI biosynthesis, inositol in phosphatidylinositol becomes acylated. Inositol is deacylated prior to attachment to variant surface glycoproteins in the bloodstream form, whereas it remains acylated in procyclins in the procyclic form. We have cloned a T. brucei GPI inositol deacylase (GPIdeAc2). In accordance with the acylation/deacylation profile, the level of GPIdeAc2 mRNA was 6-fold higher in the bloodstream form than in the procyclic form. Knockdown of GPIdeAc2 in the bloodstream form caused accumulation of an inositol-acylated GPI, a decreased VSG expression on the cell surface and slower growth, indicating that inositol-deacylation is essential for the growth of the bloodstream form. Overexpression of GPIdeAc2 in the procyclic form caused an accumulation of GPI biosynthetic intermediates lacking inositol-linked acyl chain and decreased cell surface procyclins because of release into the culture medium, indicating that overexpression of GPIdeAc2 is deleterious to the surface coat of the procyclic form. Therefore, the GPI inositol deacylase activity must be tightly regulated in trypanosome life cycle.}, keywords = {Acylation, Animals, Cloning, Molecular, Glycosylphosphatidylinositols, Inositol, Life Cycle Stages, Membrane Glycoproteins, Phosphoric Monoester Hydrolases, Protozoan Proteins, RNA, Messenger, Trypanosoma brucei brucei}, issn = {0021-9258}, doi = {10.1074/jbc.M513061200}, author = {Hong, Yeonchul and Nagamune, Kisaburo and Morita, Yasu S and Nakatani, Fumiki and Ashida, Hisashi and Maeda, Yusuke and Kinoshita, Taroh} } @article {511, title = {Role of RelGsu in stress response and Fe(III) reduction in Geobacter sulfurreducens.}, journal = {J Bacteriol}, volume = {188}, year = {2006}, month = {2006 Dec}, pages = {8469-78}, abstract = {Geobacter species are key members of the microbial community in many subsurface environments in which dissimilatory metal reduction is an important process. The genome of Geobacter sulfurreducens contains a gene designated rel(Gsu), which encodes a RelA homolog predicted to catalyze both the synthesis and the degradation of guanosine 3{\textquoteright},5{\textquoteright}-bispyrophosphate (ppGpp), a regulatory molecule that signals slow growth in response to nutrient limitation in bacteria. To evaluate the physiological role of Rel(Gsu) in G. sulfurreducens, a rel(Gsu) mutant was constructed and characterized, and ppGpp levels were monitored under various conditions in both the wild-type and rel(Gsu) mutant strains. In the wild-type strain, ppGpp and ppGp were produced in response to acetate and nitrogen deprivation, whereas exposure to oxygen resulted in an accumulation of ppGpp alone. Neither ppGpp nor ppGp could be detected in the rel(Gsu) mutant. The rel(Gsu) mutant consistently grew to a higher cell density than the wild type in acetate-fumarate medium and was less tolerant of oxidative stress than the wild type. The capacity for Fe(III) reduction was substantially diminished in the mutant. Microarray and quantitative reverse transcription-PCR analyses indicated that during stationary-phase growth, protein synthesis genes were up-regulated in the rel(Gsu) mutant and genes involved in stress responses and electron transport, including several implicated in Fe(III) reduction, were down-regulated in the mutant. The results are consistent with a role for Rel(Gsu) in regulating growth, stress responses, and Fe(III) reduction in G. sulfurreducens under conditions likely to be prevalent in subsurface environments.}, keywords = {Bacterial Proteins, Culture Media, Ferric Compounds, Gene Expression Regulation, Bacterial, Geobacter, Guanosine Tetraphosphate, Heat-Shock Response, Ligases, Mutation, Oligonucleotide Array Sequence Analysis, Oxidation-Reduction, Reverse Transcriptase Polymerase Chain Reaction, Sulfur}, issn = {0021-9193}, doi = {10.1128/JB.01278-06}, author = {DiDonato, Laurie N and Sullivan, Sara A and Meth{\'e}, Barbara A and Nevin, Kelly P and England, Reg and Lovley, Derek R} } @article {718, title = {Simian virus 40 late proteins possess lytic properties that render them capable of permeabilizing cellular membranes.}, journal = {J Virol}, volume = {80}, year = {2006}, month = {2006 Jul}, pages = {6575-87}, abstract = {Many nonenveloped viruses have evolved an infectious cycle that culminates in the lysis or permeabilization of the host to enable viral release. How these viruses initiate the lytic event is largely unknown. Here, we demonstrated that the simian virus 40 progeny accumulated at the nuclear envelope prior to the permeabilization of the nuclear, endoplasmic reticulum, and plasma membranes at a time which corresponded with the release of the progeny. The permeabilization of these cellular membranes temporally correlated with late protein expression and was not observed upon the inhibition of their synthesis. To address whether one or more of the late proteins possessed an inherent capacity to induce membrane permeabilization, we examined the permeability of Escherichia coli that separately expressed the late proteins. VP2 and VP3, but not VP1, caused the permeabilization of bacterial membranes. Additionally, VP3 expression resulted in bacterial cell lysis. These findings demonstrate that VP3 possesses an inherent lytic property that is independent of eukaryotic signaling or cell death pathways.}, keywords = {Animals, Capsid Proteins, Cell Death, Cell Line, Cell Membrane Permeability, Cercopithecus aethiops, Endoplasmic Reticulum, Escherichia coli, Gene Expression Regulation, Viral, Nuclear Envelope, Permeability, Protein Biosynthesis, Simian virus 40, Virus Replication}, issn = {0022-538X}, doi = {10.1128/JVI.00347-06}, author = {Daniels, Robert and Rusan, Nasser M and Wilbuer, Anne-Kathrin and Norkin, Leonard C and Wadsworth, Patricia and Hebert, Daniel N} } @article {695, title = {TbGPI16 is an essential component of GPI transamidase in Trypanosoma brucei.}, journal = {FEBS Lett}, volume = {580}, year = {2006}, month = {2006 Jan 23}, pages = {603-6}, abstract = {Glycosylphosphatidylinositol (GPI) is widely used by eukaryotic cell surface proteins for membrane attachment. De novo synthesized GPI precursors are attached to proteins post-translationally by the enzyme complex, GPI transamidase. TbGPI16, a component of the trypanosome transamidase, shares similarity with human PIG-T. Here, we show that TbGPI16 is the orthologue of PIG-T and an essential component of GPI transamidase by creating a TbGPI16 knockout. TbGPI16 forms a disulfide-linked complex with TbGPI8. A cysteine to serine mutant of TbGPI16 was unable to fully restore the surface expression of GPI-anchored proteins upon transfection into the knockout cells, indicating that its disulfide linkage with TbGPI8 is important for the full transamidase activity.}, keywords = {Acyltransferases, Animals, Disulfides, Gene Targeting, Humans, Membrane Glycoproteins, Multiprotein Complexes, Mutation, Protein Subunits, Protozoan Proteins, Trypanosoma brucei brucei}, issn = {0014-5793}, doi = {10.1016/j.febslet.2005.12.075}, author = {Hong, Yeonchul and Nagamune, Kisaburo and Ohishi, Kazuhito and Morita, Yasu S and Ashida, Hisashi and Maeda, Yusuke and Kinoshita, Taroh} } @article {521, title = {Two putative c-type multiheme cytochromes required for the expression of OmcB, an outer membrane protein essential for optimal Fe(III) reduction in Geobacter sulfurreducens.}, journal = {J Bacteriol}, volume = {188}, year = {2006}, month = {2006 Apr}, pages = {3138-42}, abstract = {Deletion of two homologous Geobacter sulfurreducens c-type cytochrome genes, omcG and omcH, decreased the rate of Fe(III) reduction and decreased the level of an outer membrane cytochrome critical for Fe(III) reduction, OmcB, without affecting its transcription. Expression of either gene restored Fe(III) reduction and OmcB expression, suggesting functional similarity.}, keywords = {Bacterial Outer Membrane Proteins, Bacterial Proteins, Blotting, Northern, Blotting, Western, Cytochromes c, Ferric Compounds, Gene Deletion, Gene Expression, Genes, Bacterial, Geobacter, Oxidation-Reduction, RNA, Bacterial, RNA, Messenger}, issn = {0021-9193}, doi = {10.1128/JB.188.8.3138-3142.2006}, author = {Kim, Byoung-Chan and Qian, Xinlei and Leang, Ching and Coppi, Maddalena V and Lovley, Derek R} } @article {531, title = {Adaptation to disruption of the electron transfer pathway for Fe(III) reduction in Geobacter sulfurreducens.}, journal = {J Bacteriol}, volume = {187}, year = {2005}, month = {2005 Sep}, pages = {5918-26}, abstract = {Previous studies demonstrated that an outer membrane c-type cytochrome, OmcB, was involved in Fe(III) reduction in Geobacter sulfurreducens. An OmcB-deficient mutant was greatly impaired in its ability to reduce both soluble and insoluble Fe(III). Reintroducing omcB restored the capacity for Fe(III) reduction at a level proportional to the level of OmcB production. Here, we report that the OmcB-deficient mutant gradually adapted to grow on soluble Fe(III) but not insoluble Fe(III). The adapted OmcB-deficient mutant reduced soluble Fe(III) at a rate comparable to that of the wild type, but the cell yield of the mutant was only ca. 60\% of that of the wild type under steady-state culturing conditions. Analysis of proteins and transcript levels demonstrated that expression of several membrane-associated cytochromes was higher in the adapted mutant than in the wild type. Further comparison of transcript levels during steady-state growth on Fe(III) citrate with a whole-genome DNA microarray revealed a significant shift in gene expression in an apparent attempt to adapt metabolism to the impaired electron transport to Fe(III). These results demonstrate that, although there are many other membrane-bound c-type cytochromes in G. sulfurreducens, increased expression of these cytochromes cannot completely compensate for the loss of OmcB. The concept that outer membrane cytochromes are promiscuous reductases that are interchangeable in function appears to be incorrect. Furthermore, the results indicate that there may be different mechanisms for electron transfer to soluble Fe(III) and insoluble Fe(III) oxides in G. sulfurreducens, which emphasizes the importance of studying electron transport to the environmentally relevant Fe(III) oxides.}, keywords = {Adaptation, Physiological, Bacterial Outer Membrane Proteins, Base Sequence, Cytochromes c, DNA Primers, Electron Transport, Ferric Compounds, Gene Expression Regulation, Bacterial, Geobacter, Kinetics, Oxidation-Reduction, Reverse Transcriptase Polymerase Chain Reaction}, issn = {0021-9193}, doi = {10.1128/JB.187.17.5918-5926.2005}, author = {Leang, Ching and Adams, L A and Chin, K-J and Nevin, K P and Meth{\'e}, B A and Webster, J and Sharma, M L and Lovley, D R} } @article {856, title = {The bronchial lavage of pediatric patients with asthma contains infectious Chlamydia.}, journal = {Am J Respir Crit Care Med}, volume = {171}, year = {2005}, month = {2005 May 15}, pages = {1083-8}, abstract = {There has been a worldwide increase in the incidence of asthma, and the disease has greatly impacted the public health care system. Chlamydia pneumoniae has been reported as a possible contributing factor in asthma. The organism has been detected by polymerase chain reaction (PCR) in bronchial tissue, but there has been no direct evidence of viability. To determine the frequency of viable Chlamydia in children, blood and bronchoalveolar lavage were collected from 70 pediatric patients undergoing flexible fiberoptic bronchoscopy. Forty-two of these patients had asthma, whereas the remaining patients had various respiratory disorders. Fifty-four percent (38) of the bronchoalveolar lavage samples were PCR-positive for Chlamydia, and 31\% (22) of the PCR-positive samples were positive when cultured on macrophages. Twenty-eight samples (40\%) and 14 samples (20\%) of the PCR- and culture-positive samples, respectively, were from patients with asthma. Culture of the blood samples revealed that 24 (34.3\%) of 70 were positive for Chlamydia compared with 8 (11\%) of 70 matched nonrespiratory control subjects (p < 0.01); 17 (24\%) of the positive blood cultures from the respiratory group were from patients with asthma. Elevation of total IgE was strongly associated with lavage culture positivity for Chlamydia. We therefore conclude that viable Chlamydia pneumoniae organisms are frequently present in the lung lavage fluid from this cohort of predominantly asthmatic pediatric patients.}, keywords = {Adolescent, Adult, Asthma, Bronchoalveolar Lavage Fluid, Child, Child, Preschool, Chlamydia, Chlamydia Infections, Cohort Studies, Female, Humans, Immunoglobulin E, Infant, Infant, Newborn, Male, Polymerase Chain Reaction, Tissue Culture Techniques}, issn = {1073-449X}, doi = {10.1164/rccm.200407-917OC}, author = {Webley, Wilmore C and Salva, Paul S and Andrzejewski, Chester and Cirino, Frances and West, Corrie A and Tilahun, Yaphet and Stuart, Elizabeth S} } @article {717, title = {The caveolae-mediated sv40 entry pathway bypasses the golgi complex en route to the endoplasmic reticulum.}, journal = {Virol J}, volume = {2}, year = {2005}, month = {2005}, pages = {38}, abstract = {BACKGROUND: Simian virus 40 (SV40) enters cells via an atypical caveolae-mediated endocytic pathway, which delivers the virus to a new intermediary compartment, the caveosome. The virus then is believed to go directly from the caveosome to the endoplasmic reticulum. Cholera toxin likewise enters via caveolae and traffics to caveosomes. But, in contrast to SV40, cholera toxin is transported from caveosomes to the endoplasmic reticulum via the Golgi. For that reason, and because the caveosome and Golgi may have some common markers, we revisited the issue of whether SV40 might access the endoplasmic reticulum via the Golgi. RESULTS: We confirmed our earlier finding that SV40 co localizes with the Golgi marker beta-COP. However, we show that the virus does not co localize with the more discriminating Golgi markers, golgin 97 and BODIPY-ceramide. CONCLUSION: The caveolae-mediated SV40 entry pathway does not intersect the Golgi. SV40 is seen to co localize with beta-COP because that protein is a marker for caveosomes as well as the Golgi. Moreover, these results are consistent with the likelihood that the caveosome is a sorting organelle. In addition, there are at least two distinct but related routes by which a ligand might traffic from the caveosome to the ER; one route involving transport through the Golgi, and another pathway that does not involve the Golgi.}, keywords = {Animals, Caveolae, Cercopithecus aethiops, Endoplasmic Reticulum, Fibroblasts, Golgi Apparatus, Immunohistochemistry, Simian virus 40}, issn = {1743-422X}, doi = {10.1186/1743-422X-2-38}, author = {Norkin, Leonard C and Kuksin, Dmitry} } @article {533, title = {Characterization of citrate synthase from Geobacter sulfurreducens and evidence for a family of citrate synthases similar to those of eukaryotes throughout the Geobacteraceae.}, journal = {Appl Environ Microbiol}, volume = {71}, year = {2005}, month = {2005 Jul}, pages = {3858-65}, abstract = {Members of the family Geobacteraceae are commonly the predominant Fe(III)-reducing microorganisms in sedimentary environments, as well as on the surface of energy-harvesting electrodes, and are able to effectively couple the oxidation of acetate to the reduction of external electron acceptors. Citrate synthase activity of these organisms is of interest due to its key role in acetate metabolism. Prior sequencing of the genome of Geobacter sulfurreducens revealed a putative citrate synthase sequence related to the citrate synthases of eukaryotes. All citrate synthase activity in G. sulfurreducens could be resolved to a single 49-kDa protein via affinity chromatography. The enzyme was successfully expressed at high levels in Escherichia coli with similar properties as the native enzyme, and kinetic parameters were comparable to related citrate synthases (kcat= 8.3 s(-1); Km= 14.1 and 4.3 microM for acetyl coenzyme A and oxaloacetate, respectively). The enzyme was dimeric and was slightly inhibited by ATP (Ki= 1.9 mM for acetyl coenzyme A), which is a known inhibitor for many eukaryotic, dimeric citrate synthases. NADH, an allosteric inhibitor of prokaryotic hexameric citrate synthases, did not affect enzyme activity. Unlike most prokaryotic dimeric citrate synthases, the enzyme did not have any methylcitrate synthase activity. A unique feature of the enzyme, in contrast to citrate synthases from both eukaryotes and prokaryotes, was a lack of stimulation by K+ ions. Similar citrate synthase sequences were detected in a diversity of other Geobacteraceae members. This first characterization of a eukaryotic-like citrate synthase from a prokaryote provides new insight into acetate metabolism in Geobacteraceae members and suggests a molecular target for tracking the presence and activity of these organisms in the environment.}, keywords = {Amino Acid Sequence, Citrate (si)-Synthase, Culture Media, Deltaproteobacteria, DNA, Bacterial, Eukaryotic Cells, Geobacter, Kinetics, Molecular Sequence Data, Phylogeny, Sequence Analysis, DNA}, issn = {0099-2240}, doi = {10.1128/AEM.71.7.3858-3865.2005}, author = {Bond, Daniel R and Mester, T{\"u}nde and Nesb{\o}, Camilla L and Izquierdo-Lopez, Andrea V and Collart, Frank L and Lovley, Derek R} } @article {396, title = {Characterization of TetD as a transcriptional activator of a subset of genes of the Escherichia coli SoxS/MarA/Rob regulon.}, journal = {Mol Microbiol}, volume = {56}, year = {2005}, month = {2005 May}, pages = {1103-17}, abstract = {In Escherichia coli, SoxS, MarA and Rob form a closely related subset of the AraC/XylS family of positive regulators, sharing approximately 42\% amino acid sequence identity over the length of SoxS and the ability to activate transcription of a common set of target genes that provide resistance to redox-cycling compounds and antibiotics. On the basis of its approximately 43\% amino acid sequence identity with SoxS, MarA and Rob, TetD, encoded by transposon Tn10, appears to be a fourth member of the subset. However, although its expression has been shown to be negatively regulated by TetC and not inducible by tetracycline, the physiological function of TetD is unknown. Accordingly, in the work presented here, we initiate a molecular characterization of TetD. We show that expression of TetD activates transcription of a subset of the SoxS/MarA/Rob regulon genes and confers resistance to redox-cycling compounds and antibiotics. We show that mutations in the putative TetD binding site of a TetD-activatable promoter and a mutation in the protein{\textquoteright}s N-terminal DNA recognition helix interfere with transcription activation, thereby indicating that TetD directly activates target gene transcription. Finally, we show that TetD, like SoxS and MarA, is intrinsically unstable; however, unlike SoxS and MarA, TetD is not degraded by Lon or any of the cell{\textquoteright}s known cytoplasmic ATP-dependent proteases. Thus, we conclude that TetD is a bona fide member of the SoxS/MarA/Rob subfamily of positive regulators.}, keywords = {Amino Acid Sequence, Binding Sites, DNA, DNA-Binding Proteins, Escherichia coli, Escherichia coli Proteins, Gene Expression Regulation, Bacterial, Multigene Family, Promoter Regions, Genetic, Regulon, Tetracycline Resistance, Trans-Activators, Transcription Factors, Transcription, Genetic}, issn = {0950-382X}, doi = {10.1111/j.1365-2958.2005.04599.x}, author = {Griffith, Kevin L and Becker, Stephen M and Wolf, Richard E} } @article {696, title = {Compartmentalization of lipid biosynthesis in mycobacteria.}, journal = {J Biol Chem}, volume = {280}, year = {2005}, month = {2005 Jun 3}, pages = {21645-52}, abstract = {The plasma membrane of Mycobacterium sp. is the site of synthesis of several distinct classes of lipids that are either retained in the membrane or exported to the overlying cell envelope. Here, we provide evidence that enzymes involved in the biosynthesis of two major lipid classes, the phosphatidylinositol mannosides (PIMs) and aminophospholipids, are compartmentalized within the plasma membrane. Enzymes involved in the synthesis of early PIM intermediates were localized to a membrane subdomain termed PMf, that was clearly resolved from the cell wall by isopyknic density centrifugation and amplified in rapidly dividing Mycobacterium smegmatis. In contrast, the major pool of apolar PIMs and enzymes involved in polar PIM biosynthesis were localized to a denser fraction that contained both plasma membrane and cell wall markers (PM-CW). Based on the resistance of the PIMs to solvent extraction in live but not lysed cells, we propose that polar PIM biosynthesis occurs in the plasma membrane rather than the cell wall component of the PM-CW. Enzymes involved in phosphatidylethanolamine biosynthesis also displayed a highly polarized distribution between the PMf and PM-CW fractions. The PMf was greatly reduced in non-dividing cells, concomitant with a reduction in the synthesis and steady-state levels of PIMs and amino-phospholipids and the redistribution of PMf marker enzymes to non-PM-CW fractions. The formation of the PMf and recruitment of enzymes to this domain may thus play a role in regulating growth-specific changes in the biosynthesis of membrane and cell wall lipids.}, keywords = {Bacterial Proteins, Biochemistry, Cell Membrane, Cell Wall, Hemagglutinins, Lipid Metabolism, Lipids, Mannosides, Mannosyltransferases, Microscopy, Electron, Models, Biological, Mycobacterium smegmatis, Phosphatidylethanolamines, Phosphatidylinositols, Phospholipids, Protein Structure, Tertiary, Subcellular Fractions}, issn = {0021-9258}, doi = {10.1074/jbc.M414181200}, author = {Morita, Yasu S and Velasquez, Ren{\'e} and Taig, Ellen and Waller, Ross F and Patterson, John H and Tull, Dedreia and Williams, Spencer J and Billman-Jacobe, Helen and McConville, Malcolm J} } @article {683, title = {ConSurf 2005: the projection of evolutionary conservation scores of residues on protein structures.}, journal = {Nucleic Acids Res}, volume = {33}, year = {2005}, month = {2005 Jul 1}, pages = {W299-302}, abstract = {Key amino acid positions that are important for maintaining the 3D structure of a protein and/or its function(s), e.g. catalytic activity, binding to ligand, DNA or other proteins, are often under strong evolutionary constraints. Thus, the biological importance of a residue often correlates with its level of evolutionary conservation within the protein family. ConSurf (http://consurf.tau.ac.il/) is a web-based tool that automatically calculates evolutionary conservation scores and maps them on protein structures via a user-friendly interface. Structurally and functionally important regions in the protein typically appear as patches of evolutionarily conserved residues that are spatially close to each other. We present here version 3.0 of ConSurf. This new version includes an empirical Bayesian method for scoring conservation, which is more accurate than the maximum-likelihood method that was used in the earlier release. Various additional steps in the calculation can now be controlled by a number of advanced options, thus further improving the accuracy of the calculation. Moreover, ConSurf version 3.0 also includes a measure of confidence for the inferred amino acid conservation scores.}, keywords = {Amino Acid Substitution, Amino Acids, Bacterial Proteins, Bayes Theorem, Evolution, Molecular, Internet, Models, Molecular, Potassium Channels, Protein Conformation, Software}, issn = {1362-4962}, doi = {10.1093/nar/gki370}, author = {Landau, Meytal and Mayrose, Itay and Rosenberg, Yossi and Glaser, Fabian and Martz, Eric and Pupko, Tal and Ben-Tal, Nir} } @article {539, title = {Crystal ball. In silico biology meets in situ phenomenology.}, journal = {Environ Microbiol}, volume = {7}, year = {2005}, month = {2005 Apr}, pages = {478-9}, keywords = {Bacterial Physiological Phenomena, Computational Biology, Models, Biological}, issn = {1462-2912}, doi = {10.1111/j.1462-2920.2005.803_8.x}, author = {Lovley, Derek R} } @article {538, title = {DNA microarray analysis of nitrogen fixation and Fe(III) reduction in Geobacter sulfurreducens.}, journal = {Appl Environ Microbiol}, volume = {71}, year = {2005}, month = {2005 May}, pages = {2530-8}, abstract = {A DNA microarray representing the genome of Geobacter sulfurreducens was constructed for use in global gene expression profiling of cells under steady-state conditions with acetate as the electron donor and Fe(III) or fumarate as the electron acceptor. Reproducible differences in transcript levels were also observed in comparisons between cells grown with ammonia and those fixing atmospheric nitrogen. There was a high correlation between changes in transcript levels determined with microarray analyses and an evaluation of a subset of the genome with quantitative PCR. As expected, cells required to fix nitrogen had higher levels of transcripts of genes associated with nitrogen fixation, further demonstrating that the microarray approach could reliably detect important physiological changes. Cells grown with Fe(III) as the electron acceptor had higher levels of transcripts for omcB, a gene coding for an outer membrane c-type cytochrome that is essential for Fe(III) reduction. Several other c-type cytochrome genes also appeared to be up-regulated. An unexpected result was significantly higher levels of transcripts for genes which have a role in metal efflux, potentially suggesting the importance of maintaining metal homeostasis during release of soluble metals when reducing Fe(III). A substantial proportion (30\%) of significantly expressed genes during Fe(III) reduction were genes of unknown function or hypothetical proteins, suggesting differences in Fe(III) reduction physiology among microorganisms which perform this metabolic process.}, keywords = {Ferric Compounds, Fumarates, Geobacter, Microarray Analysis, Nitrogen Fixation, Oxidation-Reduction, Polymerase Chain Reaction}, issn = {0099-2240}, doi = {10.1128/AEM.71.5.2530-2538.2005}, author = {Meth{\'e}, Barbara A and Webster, Jennifer and Nevin, Kelly and Butler, Jessica and Lovley, Derek R} } @article {540, title = {Evidence for involvement of an electron shuttle in electricity generation by Geothrix fermentans.}, journal = {Appl Environ Microbiol}, volume = {71}, year = {2005}, month = {2005 Apr}, pages = {2186-9}, abstract = {In experiments performed using graphite electrodes poised by a potentiostat (+200 mV versus Ag/AgCl) or in a microbial fuel cell (with oxygen as the electron acceptor), the Fe(III)-reducing organism Geothrix fermentans conserved energy to support growth by coupling the complete oxidation of acetate to reduction of a graphite electrode. Other organic compounds, such as lactate, malate, propionate, and succinate as well as components of peptone and yeast extract, were utilized for electricity production. However, electrical characteristics and the results of shuttling assays indicated that unlike previously described electrode-reducing microorganisms, G. fermentans produced a compound that promoted electrode reduction. This is the first report of complete oxidation of organic compounds linked to electrode reduction by an isolate outside of the Proteobacteria.}, keywords = {Acetates, Bacteria, Bioelectric Energy Sources, Electricity, Electrodes, Electron Transport, Graphite, Microscopy, Electron, Scanning, Oxidation-Reduction}, issn = {0099-2240}, doi = {10.1128/AEM.71.4.2186-2189.2005}, author = {Bond, Daniel R and Lovley, Derek R} } @article {535, title = {Extracellular electron transfer via microbial nanowires.}, journal = {Nature}, volume = {435}, year = {2005}, month = {2005 Jun 23}, pages = {1098-101}, abstract = {Microbes that can transfer electrons to extracellular electron acceptors, such as Fe(iii) oxides, are important in organic matter degradation and nutrient cycling in soils and sediments. Previous investigations on electron transfer to Fe(iii) have focused on the role of outer-membrane c-type cytochromes. However, some Fe(iii) reducers lack c-cytochromes. Geobacter species, which are the predominant Fe(iii) reducers in many environments, must directly contact Fe(iii) oxides to reduce them, and produce monolateral pili that were proposed, on the basis of the role of pili in other organisms, to aid in establishing contact with the Fe(iii) oxides. Here we report that a pilus-deficient mutant of Geobacter sulfurreducens could not reduce Fe(iii) oxides but could attach to them. Conducting-probe atomic force microscopy revealed that the pili were highly conductive. These results indicate that the pili of G. sulfurreducens might serve as biological nanowires, transferring electrons from the cell surface to the surface of Fe(iii) oxides. Electron transfer through pili indicates possibilities for other unique cell-surface and cell-cell interactions, and for bioengineering of novel conductive materials.}, keywords = {Biotechnology, Electric Conductivity, Electron Transport, Ferric Compounds, Fimbriae Proteins, Fimbriae, Bacterial, Genes, Bacterial, Geobacter, Microscopy, Atomic Force, Microscopy, Electron, Transmission, Mutation, Nanostructures, Phylogeny}, issn = {1476-4687}, doi = {10.1038/nature03661}, author = {Reguera, Gemma and McCarthy, Kevin D and Mehta, Teena and Nicoll, Julie S and Tuominen, Mark T and Lovley, Derek R} } @article {697, title = {Function of phosphatidylinositol in mycobacteria.}, journal = {J Biol Chem}, volume = {280}, year = {2005}, month = {2005 Mar 25}, pages = {10981-7}, abstract = {Phosphatidylinositol (PI) is an abundant phospholipid in the cytoplasmic membrane of mycobacteria and the precursor for more complex glycolipids, such as the PI mannosides (PIMs) and lipoarabinomannan (LAM). To investigate whether the large steady-state pools of PI and apolar PIMs are required for mycobacterial growth, we have generated a Mycobacterium smegmatis inositol auxotroph by disruption of the ino1 gene. The ino1 mutant displayed wild-type growth rates and steady-state levels of PI, PIM, and LAM when grown in the presence of 1 mM inositol. The non-dividing ino1 mutant was highly resistant to inositol starvation, reflecting the slow turnover of inositol lipids in this stage. In contrast, dilution of growing or stationary-phase ino1 mutant in inositol-free medium resulted in the rapid depletion of PI and apolar PIMs. Whereas depletion of these lipids was not associated with loss of viability, subsequent depletion of polar PIMs coincided with loss of major cell wall components and cell viability. Metabolic labeling experiments confirmed that the large pools of PI and apolar PIMs were used to sustain polar PIM and LAM biosynthesis during inositol limitation. They also showed that under non-limiting conditions, PI is catabolized via lyso-PI. These data suggest that large pools of PI and apolar PIMs are not essential for membrane integrity but are required to sustain polar PIM biosynthesis, which is essential for mycobacterial growth.}, keywords = {Inositol, Lipopolysaccharides, Mannose, Mycobacterium smegmatis, Phosphatidylinositols}, issn = {0021-9258}, doi = {10.1074/jbc.M413443200}, author = {Haites, Ruth E and Morita, Yasu S and McConville, Malcolm J and Billman-Jacobe, Helen} } @article {40, title = {Genomic analysis of PIS1 gene expression.}, journal = {Eukaryot Cell}, volume = {4}, year = {2005}, month = {2005 Mar}, pages = {604-14}, abstract = {The Saccharomyces cerevisiae PIS1 gene is essential and required for the final step in the de novo synthesis of phosphatidylinositol. Transcription of the PIS1 gene is uncoupled from the factors that regulate other yeast phospholipid biosynthetic genes. Most of the phospholipid biosynthetic genes are regulated in response to inositol and choline via a regulatory circuit that includes the Ino2p:Ino4p activator complex and the Opi1p repressor. PIS1 is regulated in response to carbon source and anaerobic growth conditions. Both of these regulatory responses are modest, which is not entirely surprising since PIS1 is essential. However, even modest regulation of PIS1 expression has been shown to affect phosphatidylinositol metabolism and to affect cell cycle progression. This prompted the present study, which employed a genomic screen, database mining, and more traditional promoter analysis to identify genes that affect PIS1 expression. A screen of the viable yeast deletion set identified 120 genes that affect expression of a PIS1-lacZ reporter. The gene set included several peroxisomal genes, silencing genes, and transcription factors. Factors suggested by database mining, such as Pho2 and Yfl044c, were also found to affect PIS1-lacZ expression. A PIS1 promoter deletion study identified an upstream regulatory sequence element that was required for carbon source regulation located downstream of three previously defined upstream activation sequence elements. Collectively, these studies demonstrate how a collection of genomic and traditional strategies can be implemented to identify a set of genes that affect the regulation of an essential gene.}, keywords = {Gene Expression Profiling, Gene Expression Regulation, Fungal, Gene Silencing, Genome, Fungal, Glycerol, Oligonucleotide Array Sequence Analysis, Peroxisomes, Phospholipids, Promoter Regions, Genetic, Recombinant Fusion Proteins, Saccharomyces cerevisiae, Transferases (Other Substituted Phosphate Groups)}, issn = {1535-9778}, doi = {10.1128/EC.4.3.604-614.2005}, author = {Gardocki, Mary E and Bakewell, Margaret and Kamath, Deepa and Robinson, Kelly and Borovicka, Kathy and Lopes, John M} } @article {532, title = {Geobacter bemidjiensis sp. nov. and Geobacter psychrophilus sp. nov., two novel Fe(III)-reducing subsurface isolates.}, journal = {Int J Syst Evol Microbiol}, volume = {55}, year = {2005}, month = {2005 Jul}, pages = {1667-74}, abstract = {Fe(III)-reducing isolates were recovered from two aquifers in which Fe(III) reduction is known to be important. Strain Bem(T) was enriched from subsurface sediments collected in Bemidji, MN, USA, near a site where Fe(III) reduction is important in aromatic hydrocarbon degradation. Strains P11, P35(T) and P39 were isolated from the groundwater of an aquifer in Plymouth, MA, USA, in which Fe(III) reduction is important because of long-term inputs of acetate as a highway de-icing agent to the subsurface. All four isolates were Gram-negative, slightly curved rods that grew best in freshwater media. Strains P11, P35(T) and P39 exhibited motility via means of monotrichous flagella. Analysis of the 16S rRNA and nifD genes indicated that all four strains are delta-proteobacteria and members of the Geobacter cluster of the Geobacteraceae. Differences in phenotypic and phylogenetic characteristics indicated that the four isolates represent two novel species within the genus Geobacter. All of the isolates coupled the oxidation of acetate to the reduction of Fe(III) [iron(III) citrate, amorphous iron(III) oxide, iron(III) pyrophosphate and iron(III) nitrilotriacetate]. All four strains utilized ethanol, lactate, malate, pyruvate and succinate as electron donors and malate and fumarate as electron acceptors. Strain Bem(T) grew fastest at 30 degrees C, whereas strains P11, P35(T) and P39 grew equally well at 17, 22 and 30 degrees C. In addition, strains P11, P35(T) and P39 were capable of growth at 4 degrees C. The names Geobacter bemidjiensis sp. nov. (type strain Bem(T)=ATCC BAA-1014(T)=DSM 16622(T)=JCM 12645(T)) and Geobacter psychrophilus sp. nov. (strains P11, P35(T) and P39; type strain P35(T)=ATCC BAA-1013(T)=DSM 16674(T)=JCM 12644(T)) are proposed.}, keywords = {Bacterial Proteins, Bacterial Typing Techniques, Cold Temperature, DNA, Bacterial, DNA, Ribosomal, Ferric Compounds, Fresh Water, Genes, rRNA, Geobacter, Geologic Sediments, Massachusetts, Minnesota, Molecular Sequence Data, Oxidation-Reduction, RNA, Ribosomal, 16S, Species Specificity, Water Supply}, issn = {1466-5026}, doi = {10.1099/ijs.0.63417-0}, author = {Nevin, Kelly P and Holmes, Dawn E and Woodard, Trevor L and Hinlein, Erich S and Ostendorf, David W and Lovley, Derek R} } @article {361, title = {Infection with the insect virus Hz-2v alters mating behavior and pheromone production in female Helicoverpa zea moths.}, journal = {J Insect Sci}, volume = {5}, year = {2005}, month = {2005}, pages = {6}, abstract = {The effect of Hz-2V virus infection on the reproductive physiology and behavior of infected Helicoverpa zea female moths was examined. In the absence of males, infected females exhibited calling behavior and called as often but for shorter periods on average than control females. As expected, control females mated with males for extend periods when they were present and did not call after mating, while virus-infected females made many frequent contacts with males and continued to call even after these contacts. Virus-infected females were found to produce five to seven times more pheromone than control females and attracted twice as many males as did control females in flight tunnel experiments. The ability of Hz-2V to alter the physiology and behavior of infected females observed here may serve to facilitate the transmission of virus in insect populations.}, keywords = {Animals, Female, Infectious Disease Transmission, Vertical, Insect Viruses, Male, Moths, Pheromones, Sexual Behavior, Animal, Time Factors, Vocalization, Animal}, issn = {1536-2442}, author = {Burand, John P and Tan, Weijia and Kim, Woojin and Nojima, Satoshi and Roelofs, Wendell} } @article {820, title = {Localization of RecA in Escherichia coli K-12 using RecA-GFP.}, journal = {Mol Microbiol}, volume = {57}, year = {2005}, month = {2005 Aug}, pages = {1074-85}, abstract = {RecA is important in recombination, DNA repair and repair of replication forks. It functions through the production of a protein-DNA filament. To study the localization of RecA in live Escherichia coli cells, the RecA protein was fused to the green fluorescence protein (GFP). Strains with this gene have recombination/DNA repair activities three- to tenfold below wild type (or about 1000-fold above that of a recA null mutant). RecA-GFP cells have a background of green fluorescence punctuated with up to five foci per cell. Two types of foci have been defined: 4,6-diamidino-2-phenylindole (DAPI)-sensitive foci that are bound to DNA and DAPI-insensitive foci that are DNA-less aggregates/storage structures. In log phase cells, foci were not localized to any particular region. After UV irradiation, the number of foci increased and they localized to the cell centre. This suggested colocalization with the DNA replication factory. recA, recB and recF strains showed phenotypes and distributions of foci consistent with the predicted effects of these mutations.}, keywords = {Chromosomes, Bacterial, DNA Replication, DNA, Bacterial, Escherichia coli, Escherichia coli Proteins, Green Fluorescent Proteins, Mutation, Rec A Recombinases, Recombinant Fusion Proteins, Recombination, Genetic, Ultraviolet Rays}, issn = {0950-382X}, doi = {10.1111/j.1365-2958.2005.04755.x}, author = {Renzette, Nicholas and Gumlaw, Nathan and Nordman, Jared T and Krieger, Marlee and Yeh, Su-Ping and Long, Edward and Centore, Richard and Boonsombat, Ruethairat and Sandler, Steven J} } @article {526, title = {Microbial incorporation of 13C-labeled acetate at the field scale: detection of microbes responsible for reduction of U(VI).}, journal = {Environ Sci Technol}, volume = {39}, year = {2005}, month = {2005 Dec 1}, pages = {9039-48}, abstract = {A field-scale acetate amendment experiment was performed in a contaminated aquifer at Old Rifle, CO to stimulate in situ microbial reduction of U(VI) in groundwater. To evaluate the microorganisms responsible for microbial uranium reduction during the experiment, 13C-labeled acetate was introduced into well bores via bio-traps containing porous activated carbon beads (Bio-Sep). Incorporation of the 13C from labeled acetate into cellular DNA and phospholipid fatty acid (PLFA) biomarkers was analyzed in parallel with geochemical parameters. An enrichment of active sigma-proteobacteria was demonstrated in downgradient monitoring wells: Geobacter dominated in wells closer to the acetate injection gallery, while various sulfate reducers were prominent in different downgradient wells. These results were consistent with the geochemical evidence of Fe(III), U(VI), and SO(4)2- reduction. PLFA profiling of bio-traps suspended in the monitoring wells also showed the incorporation of 13C into bacterial cellular lipids. Community composition of downgradient monitoring wells based on quinone and PLFA profiling was in general agreement with the 13C-DNA result. The direct application of 13C label to biosystems, coupled with DNA and PLFA analysis,}, keywords = {Acetates, Biodegradation, Environmental, Carbon Isotopes, Electrophoresis, Polyacrylamide Gel, Geobacter, Phylogeny, Polymerase Chain Reaction, Proteobacteria, Uranium}, issn = {0013-936X}, author = {Chang, Yun-Juan and Long, Philip E and Geyer, Roland and Peacock, Aaron D and Resch, Charles T and Sublette, Kerry and Pfiffner, Susan and Smithgall, Amanda and Anderson, Robert T and Vrionis, Helen A and Stephen, John R and Dayvault, Richard and Ortiz-Bernad, Irene and Lovley, Derek R and White, David C} } @article {530, title = {Microbiological and geochemical heterogeneity in an in situ uranium bioremediation field site.}, journal = {Appl Environ Microbiol}, volume = {71}, year = {2005}, month = {2005 Oct}, pages = {6308-18}, abstract = {The geochemistry and microbiology of a uranium-contaminated subsurface environment that had undergone two seasons of acetate addition to stimulate microbial U(VI) reduction was examined. There were distinct horizontal and vertical geochemical gradients that could be attributed in large part to the manner in which acetate was distributed in the aquifer, with more reduction of Fe(III) and sulfate occurring at greater depths and closer to the point of acetate injection. Clone libraries of 16S rRNA genes derived from sediments and groundwater indicated an enrichment of sulfate-reducing bacteria in the order Desulfobacterales in sediment and groundwater samples. These samples were collected nearest the injection gallery where microbially reducible Fe(III) oxides were highly depleted, groundwater sulfate concentrations were low, and increases in acid volatile sulfide were observed in the sediment. Further down-gradient, metal-reducing conditions were present as indicated by intermediate Fe(II)/Fe(total) ratios, lower acid volatile sulfide values, and increased abundance of 16S rRNA gene sequences belonging to the dissimilatory Fe(III)- and U(VI)-reducing family Geobacteraceae. Maximal Fe(III) and U(VI) reduction correlated with maximal recovery of Geobacteraceae 16S rRNA gene sequences in both groundwater and sediment; however, the sites at which these maxima occurred were spatially separated within the aquifer. The substantial microbial and geochemical heterogeneity at this site demonstrates that attempts should be made to deliver acetate in a more uniform manner and that closely spaced sampling intervals, horizontally and vertically, in both sediment and groundwater are necessary in order to obtain a more in-depth understanding of microbial processes and the relative contribution of attached and planktonic populations to in situ uranium bioremediation.}, keywords = {Acetates, Biodegradation, Environmental, Deltaproteobacteria, DNA, Bacterial, DNA, Ribosomal, Ferric Compounds, Fresh Water, Geologic Sediments, Phylogeny, Polymerase Chain Reaction, RNA, Ribosomal, 16S, Sulfates, Uranium, Water Pollution}, issn = {0099-2240}, doi = {10.1128/AEM.71.10.6308-6318.2005}, author = {Vrionis, Helen A and Anderson, Robert T and Ortiz-Bernad, Irene and O{\textquoteright}Neill, Kathleen R and Resch, Charles T and Peacock, Aaron D and Dayvault, Richard and White, David C and Long, Philip E and Lovley, Derek R} } @article {534, title = {A novel Geobacteraceae-specific outer membrane protein J (OmpJ) is essential for electron transport to Fe(III) and Mn(IV) oxides in Geobacter sulfurreducens.}, journal = {BMC Microbiol}, volume = {5}, year = {2005}, month = {2005}, pages = {41}, abstract = {BACKGROUND: Metal reduction is thought to take place at or near the bacterial outer membrane and, thus, outer membrane proteins in the model dissimilatory metal-reducing organism Geobacter sulfurreducens are of interest to understand the mechanisms of Fe(III) reduction in the Geobacter species that are the predominant Fe(III) reducers in many environments. Previous studies have implicated periplasmic and outer membrane cytochromes in electron transfer to metals. Here we show that the most abundant outer membrane protein of G. sulfurreducens, OmpJ, is not a cytochrome yet it is required for metal respiration. RESULTS: When outer membrane proteins of G. sulfurreducens were separated via SDS-PAGE, one protein, designated OmpJ (outer membrane protein J), was particularly abundant. The encoding gene, which was identified from mass spectrometry analysis of peptide fragments, is present in other Geobacteraceae, but not in organisms outside this family. The predicted localization and structure of the OmpJ protein suggested that it was a porin. Deletion of the ompJ gene in G. sulfurreducens produced a strain that grew as well as the wild-type strain with fumarate as the electron acceptor but could not grow with metals, such as soluble or insoluble Fe(III) and insoluble Mn(IV) oxide, as the electron acceptor. The heme c content in the mutant strain was ca. 50\% of the wild-type and there was a widespread loss of multiple cytochromes from soluble and membrane fractions. Transmission electron microscopy analyses of mutant cells revealed an unusually enlarged periplasm, which is likely to trigger extracytoplasmic stress response mechanisms leading to the degradation of periplasmic and/or outer membrane proteins, such as cytochromes, required for metal reduction. Thus, the loss of the capacity for extracellular electron transport in the mutant could be due to the missing c-type cytochromes, or some more direct, but as yet unknown, role of OmpJ in metal reduction. CONCLUSION: OmpJ is a putative porin found in the outer membrane of the model metal reducer G. sulfurreducens that is required for respiration of extracellular electron acceptors such as soluble and insoluble metals. The effect of OmpJ in extracellular electron transfer is indirect, as OmpJ is required to keep the integrity of the periplasmic space necessary for proper folding and functioning of periplasmic and outer membrane electron transport components. The exclusive presence of ompJ in members of the Geobacteraceae family as well as its role in metal reduction suggest that the ompJ sequence may be useful in tracking the growth or activity of Geobacteraceae in sedimentary environments.}, keywords = {Amino Acid Sequence, Bacterial Outer Membrane Proteins, Base Sequence, Biological Transport, Deltaproteobacteria, DNA Primers, Ferric Compounds, Gene Deletion, Genome, Bacterial, Geobacter, Manganese Compounds, Molecular Sequence Data, Oxides, Peptide Fragments, Phylogeny, Protein Structure, Secondary, Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization}, issn = {1471-2180}, doi = {10.1186/1471-2180-5-41}, author = {Afkar, Eman and Reguera, Gemma and Schiffer, Marianne and Lovley, Derek R} } @article {536, title = {OmcF, a putative c-Type monoheme outer membrane cytochrome required for the expression of other outer membrane cytochromes in Geobacter sulfurreducens.}, journal = {J Bacteriol}, volume = {187}, year = {2005}, month = {2005 Jul}, pages = {4505-13}, abstract = {Outer membrane cytochromes are often proposed as likely agents for electron transfer to extracellular electron acceptors, such as Fe(III). The omcF gene in the dissimilatory Fe(III)-reducing microorganism Geobacter sulfurreducens is predicted to code for a small outer membrane monoheme c-type cytochrome. An OmcF-deficient strain was constructed, and its ability to reduce and grow on Fe(III) citrate was found to be impaired. Following a prolonged lag phase (150 h), the OmcF-deficient strain developed the ability to grow in Fe(III) citrate medium with doubling times and yields that were ca. 145\% and 70\% of those of the wild type, respectively. Comparison of the c-type cytochrome contents of outer membrane-enriched fractions prepared from wild-type and OmcF-deficient cultures confirmed the outer membrane association of OmcF and revealed multiple changes in the cytochrome content of the OmcF-deficient strain. These changes included loss of expression of two previously characterized outer membrane cytochromes, OmcB and OmcC, and overexpression of a third previously characterized outer membrane cytochrome, OmcS, during growth on Fe(III) citrate. The omcB and omcC transcripts could not be detected in the OmcF-deficient mutant by either reverse transcriptase PCR or Northern blot analyses. Expression of the omcF gene in trans restored both the capacity of the OmcF-deficient mutant to reduce Fe(III) and wild-type levels of omcB and omcC mRNA and protein. Thus, elimination of OmcF may impair Fe(III) reduction by influencing expression of OmcB, which has previously been demonstrated to play a critical role in Fe(III) reduction.}, keywords = {Amino Acid Sequence, Bacterial Outer Membrane Proteins, Cytochromes c, Ferric Compounds, Gene Deletion, Gene Expression Regulation, Bacterial, Geobacter, Molecular Sequence Data, Oxidation-Reduction, Sequence Alignment}, issn = {0021-9193}, doi = {10.1128/JB.187.13.4505-4513.2005}, author = {Kim, Byoung-Chan and Leang, Ching and Ding, Yan-Huai R and Glaven, Richard H and Coppi, Maddalena V and Lovley, Derek R} } @article {527, title = {Outer membrane c-type cytochromes required for Fe(III) and Mn(IV) oxide reduction in Geobacter sulfurreducens.}, journal = {Appl Environ Microbiol}, volume = {71}, year = {2005}, month = {2005 Dec}, pages = {8634-41}, abstract = {The potential role of outer membrane proteins in electron transfer to insoluble Fe(III) oxides by Geobacter sulfurreducens was investigated because this organism is closely related to the Fe(III) oxide-reducing organisms that are predominant in many Fe(III)-reducing environments. Two of the most abundant proteins that were easily sheared from the outer surfaces of intact cells were c-type cytochromes. One, designated OmcS, has a molecular mass of ca. 50 kDa and is predicted to be an outer membrane hexaheme c-type cytochrome. Transcripts for omcS could be detected during growth on Fe(III) oxide, but not on soluble Fe(III) citrate. The omcS mRNA consisted primarily of a monocistronic transcript, and to a lesser extent, a longer transcript that also contained the downstream gene omcT, which is predicted to encode a second hexaheme outer membrane cytochrome with 62.6\% amino acid sequence identity to OmcS. The other abundant c-type cytochrome sheared from the outer surface of G. sulfurreducens, designated OmcE, has a molecular mass of ca. 30 kDa and is predicted to be an outer membrane tetraheme c-type cytochrome. When either omcS or omcE was deleted, G. sulfurreducens could no longer reduce Fe(III) oxide but could still reduce soluble electron acceptors, including Fe(III) citrate. The mutants could reduce Fe(III) in Fe(III) oxide medium only if the Fe(III) chelator, nitrilotriacetic acid, or the electron shuttle, anthraquinone 2,6-disulfonate, was added. Expressing omcS or omcE in trans restored the capacity for Fe(III) oxide reduction. OmcT was not detected among the sheared proteins, and genetic studies indicated that G. sulfurreducens could not reduce Fe(III) oxide when omcT was expressed but OmcS was absent. In contrast, Fe(III) oxide was reduced when omcS was expressed in the absence of OmcT. These results suggest that OmcS and OmcE are involved in electron transfer to Fe(III) oxides in G. sulfurreducens. They also emphasize the importance of evaluating mechanisms for Fe(III) reduction with environmentally relevant Fe(III) oxide, rather than the more commonly utilized Fe(III) citrate, because additional electron transfer components are required for Fe(III) oxide reduction that are not required for Fe(III) citrate reduction.}, keywords = {Amino Acid Sequence, Bacterial Outer Membrane Proteins, Bacterial Proteins, Base Sequence, Cytochromes c, DNA Primers, Ferric Compounds, Geobacter, Kinetics, Manganese Compounds, Molecular Sequence Data, Oxidation-Reduction, Oxides, Peptide Fragments}, issn = {0099-2240}, doi = {10.1128/AEM.71.12.8634-8641.2005}, author = {Mehta, T and Coppi, M V and Childers, S E and Lovley, D R} } @article {39, title = {Phosphatidylinositol biosynthesis: biochemistry and regulation.}, journal = {Biochim Biophys Acta}, volume = {1735}, year = {2005}, month = {2005 Jul 15}, pages = {89-100}, abstract = {Phosphatidylinositol (PI) is a ubiquitous membrane lipid in eukaryotes. It is becoming increasingly obvious that PI and its metabolites play a myriad of very diverse roles in eukaryotic cells. The Saccharomyces cerevisiae PIS1 gene is essential and encodes PI synthase, which is required for the synthesis of PI. Recently, PIS1 expression was found to be regulated in response to carbon source and oxygen availability. It is particularly significant that the promoter elements required for these responses are conserved evolutionarily throughout the Saccharomyces genus. In addition, several genome-wide strategies coupled with more traditional screens suggest that several other factors regulate PIS1 expression. The impact of regulating PIS1 expression on PI synthesis will be discussed along with the possible role(s) that this may have on diseases such as cancer.}, keywords = {Eukaryotic Cells, Gene Expression Regulation, Fungal, Mutation, Phenotype, Phosphatidylinositols, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins, Transferases (Other Substituted Phosphate Groups)}, issn = {0006-3002}, doi = {10.1016/j.bbalip.2005.05.006}, author = {Gardocki, Mary E and Jani, Niketa and Lopes, John M} } @article {698, title = {PIG-V involved in transferring the second mannose in glycosylphosphatidylinositol.}, journal = {J Biol Chem}, volume = {280}, year = {2005}, month = {2005 Mar 11}, pages = {9489-97}, abstract = {Glycosylphosphatidylinositol (GPI) is a glycolipid that anchors many proteins to the eukaryotic cell surface. The biosynthetic pathway of GPI is mediated by sequential additions of sugars and other components to phosphatidylinositol. Four mannoses in the GPI are transferred from dolichol-phosphate-mannose (Dol-P-Man) and are linked through different glycosidic linkages. Therefore, four Dol-P-Man-dependent mannosyltransferases, GPI-MT-I, -MT-II, -MT-III, and -MT-IV for the first, second, third, and fourth mannoses, respectively, are required for generation of GPI. GPI-MT-I (PIG-M), GPI-MT-III (PIG-B), and GPI-MT-IV (SMP3) were previously reported, but GPI-MT-II remains to be identified. Here we report the cloning of PIG-V involved in transferring the second mannose in the GPI anchor. Human PIG-V encodes a 493-amino acid, endoplasmic reticulum (ER) resident protein with eight putative transmembrane regions. Saccharomyces cerevisiae protein encoded in open reading frame YBR004c, which we termed GPI18, has 25\% amino acid identity to human PIG-V. Viability of the yeast gpi18 deletion mutant was restored by human PIG-V cDNA. PIG-V has two functionally important conserved regions facing the ER lumen. Taken together, we suggest that PIG-V is the second mannosyltransferase in GPI anchor biosynthesis.}, keywords = {Amino Acid Sequence, Animals, Cell Line, Tumor, CHO Cells, Cloning, Molecular, Conserved Sequence, Cricetinae, DNA Primers, Glioma, Glycosylphosphatidylinositols, Humans, Mannose, Mannosyltransferases, Molecular Sequence Data, Open Reading Frames, Rats, Restriction Mapping, Saccharomyces cerevisiae, Sequence Alignment, Sequence Homology, Amino Acid}, issn = {0021-9258}, doi = {10.1074/jbc.M413867200}, author = {Kang, Ji Young and Hong, Yeongjin and Ashida, Hisashi and Shishioh, Nobue and Murakami, Yoshiko and Morita, Yasu S and Maeda, Yusuke and Kinoshita, Taroh} } @article {529, title = {Potential for quantifying expression of the Geobacteraceae citrate synthase gene to assess the activity of Geobacteraceae in the subsurface and on current-harvesting electrodes.}, journal = {Appl Environ Microbiol}, volume = {71}, year = {2005}, month = {2005 Nov}, pages = {6870-7}, abstract = {The Geobacteraceae citrate synthase is phylogenetically distinct from those of other prokaryotes and is a key enzyme in the central metabolism of Geobacteraceae. Therefore, the potential for using levels of citrate synthase mRNA to estimate rates of Geobacter metabolism was evaluated in pure culture studies and in four different Geobacteraceae-dominated environments. Quantitative reverse transcription-PCR studies with mRNA extracted from cultures of Geobacter sulfurreducens grown in chemostats with Fe(III) as the electron acceptor or in batch with electrodes as the electron acceptor indicated that transcript levels of the citrate synthase gene, gltA, increased with increased rates of growth/Fe(III) reduction or current production, whereas the expression of the constitutively expressed housekeeping genes recA, rpoD, and proC remained relatively constant. Analysis of mRNA extracted from groundwater collected from a U(VI)-contaminated site undergoing in situ uranium bioremediation revealed a remarkable correspondence between acetate levels in the groundwater and levels of transcripts of gltA. The expression of gltA was also significantly greater in RNA extracted from groundwater beneath a highway runoff recharge pool that was exposed to calcium magnesium acetate in June, when acetate concentrations were high, than in October, when the levels had significantly decreased. It was also possible to detect gltA transcripts on current-harvesting anodes deployed in freshwater sediments. These results suggest that it is possible to monitor the in situ metabolic rate of Geobacteraceae by tracking the expression of the citrate synthase gene.}, keywords = {Acetates, Citrate (si)-Synthase, Deltaproteobacteria, DNA, Ribosomal, Electrodes, Ferric Compounds, Fresh Water, Geobacter, Geologic Sediments, Petroleum, Phylogeny, RNA, Ribosomal, 16S, Uranium, Water Pollutants, Chemical, Water Pollutants, Radioactive}, issn = {0099-2240}, doi = {10.1128/AEM.71.11.6870-6877.2005}, author = {Holmes, Dawn E and Nevin, Kelly P and O{\textquoteright}Neil, Regina A and Ward, Joy E and Adams, Lorrie A and Woodard, Trevor L and Vrionis, Helen A and Lovley, Derek R} } @article {537, title = {Regulation of two highly similar genes, omcB and omcC, in a 10 kb chromosomal duplication in Geobacter sulfurreducens.}, journal = {Microbiology}, volume = {151}, year = {2005}, month = {2005 Jun}, pages = {1761-7}, abstract = {The Fe(III)-reducing micro-organism Geobacter sulfurreducens requires an outer-membrane c-type cytochrome, OmcB, for Fe(III) reduction, but a related cytochrome, OmcC, which is 73 \% identical to OmcB, is not required. The omcB and omcC genes are part of a tandem chromosomal duplication consisting of two repeated clusters of four genes. The 2.7 kb sequences preceding omcB and omcC are identical with the exception of a single base pair change. Studies that combined genetic, Northern blotting and primer extension analyses demonstrated that both omcB and omcC are transcribed as monocistronic and polycistronic (orf1-orf2-omcB/omcC) transcripts. All of the promoters for the various transcripts were found to be located within the 2.7 kb identical region upstream of omcB and omcC. The sequences of the promoter regions for the two monocistronic transcripts are identical and equidistant from the omcB or omcC start codons. The promoters for the two polycistronic transcripts, in contrast, are distinct. One is specific for transcription of orf1-orf2-omcB and the other is associated with transcription of orf1-orf2-omcC. Studies with an RpoS-deficient mutant suggested that transcription from all four promoters is RpoS dependent under one or more growth conditions. Deletion of orfR, a gene immediately upstream of orf1-orf2-omcB that encodes a putative transcriptional regulator, significantly lowered the omcB transcription when Fe(III) was the electron acceptor and partially inhibited Fe(III) reduction. In contrast, levels of omcC transcripts were unaffected in the orfR mutant. These results indicate that omcB and omcC operons represent a rare instance in which duplicated operons, located in tandem on the chromosome, have different transcriptional regulation.}, keywords = {Bacterial Outer Membrane Proteins, Bacterial Proteins, Blotting, Northern, DNA, Bacterial, Gene Deletion, Gene Expression Regulation, Bacterial, Geobacter, Molecular Sequence Data, Operon, Promoter Regions, Genetic, RNA, Bacterial, RNA, Messenger, Sequence Analysis, DNA, Sigma Factor, Transcription, Genetic}, issn = {1350-0872}, doi = {10.1099/mic.0.27870-0}, author = {Leang, Ching and Lovley, Derek R} } @article {528, title = {Remediation and recovery of uranium from contaminated subsurface environments with electrodes.}, journal = {Environ Sci Technol}, volume = {39}, year = {2005}, month = {2005 Nov 15}, pages = {8943-7}, abstract = {Previous studies have demonstrated that Geobacter species can effectively remove uranium from contaminated groundwater by reducing soluble U(VI) to the relatively insoluble U(IV) with organic compounds serving as the electron donor. Studies were conducted to determine whether electrodes might serve as an alternative electron donor for U(VI) reduction by a pure culture of Geobacter sulfurreducens and microorganisms in uranium-contaminated sediments. Electrodes poised at -500 mV (vs a Ag/AgCl reference) rapidly removed U(VI) from solution in the absence of cells. However, when the poise at the electrode was removed, all of the U(VI) returned to solution, demonstrating that the electrode did not reduce U(VI). If G. sulfurreducens was present on the electrode, U(VI) did not return to solution until the electrode was exposed to dissolved oxygen. This suggeststhat G. sulfurreducens on the electrode reduced U(VI) to U(IV) which was stably precipitated until reoxidized in the presence of oxygen. When an electrode was placed in uranium-contaminated subsurface sediments, U(VI) was removed and recovered from groundwater using poised electrodes. Electrodes emplaced in flow-through columns of uranium-contaminated sediments readily removed U(VI) from the groundwater, and 87\% of the uranium that had been removed was recovered from the electrode surface after the electrode was pulled from the sediments. These results suggest that microorganisms can use electrons derived from electrodes to reduce U(VI) and that it may be possible to remove and recover uranium from contaminated groundwater with poised electrodes.}, keywords = {Biodegradation, Environmental, Electrodes, Geobacter, Graphite, Soil Pollutants, Uranium, Water Pollutants, Chemical}, issn = {0013-936X}, author = {Gregory, Kelvin B and Lovley, Derek R} } @article {821, title = {Requirements for replication restart proteins during constitutive stable DNA replication in Escherichia coli K-12.}, journal = {Genetics}, volume = {169}, year = {2005}, month = {2005 Apr}, pages = {1799-806}, abstract = {Constitutive stable DNA replication (cSDR) is a mechanism for replisome loading in Escherichia coli K-12. This occurs in a dnaA-independent fashion in an rnhA mutant. cSDR is dependent on recA, priA, and transcription. In this report, it is shown that dnaA rnhA mutants using cSDR for initiation of their DNA replication additionally require priB, but not priC, for viability. Two subtle priA missense mutations either eliminated the ability to grow using cSDR (priA301 C479Y) or resulted in very small colonies (priA300 K230R). DnaC809, a priA suppressor, failed to allow priA or priB mutants to grow using cSDR to initiate DNA replication. Furthermore, unlike dnaC(+) strains, dnaC809 strains require priC for cSDR. DnaC809,820, a priC-independent suppressor of priA2::kan phenotypes, allowed priA and priC (but not priB) mutants to grow using cSDR to initiate DNA replication. It is also shown that rep and rnhA mutations are synthetically lethal. DnaC809 and dnaC809,820 mutations suppress this lethality. Rep is further shown to be required for cSDR in a dnaC809 strain. A model whereby these different sets of replication restart proteins interact preferentially with substrates associated with either RecA or SSB during replication restart and cSDR, respectively, is proposed.}, keywords = {Bacterial Proteins, DNA, DNA Replication, DNA, Bacterial, DNA-Binding Proteins, Escherichia coli, Escherichia coli Proteins, Gene Expression Regulation, Bacterial, Genome, Bacterial, Macromolecular Substances, Models, Genetic, Mutation, Mutation, Missense, Phenotype, Rec A Recombinases, RNA, SOS Response (Genetics)}, issn = {0016-6731}, doi = {10.1534/genetics.104.036962}, author = {Sandler, Steven J} } @article {751, title = {Shifts in rhizoplane communities of aquatic plants after cadmium exposure.}, journal = {Appl Environ Microbiol}, volume = {71}, year = {2005}, month = {2005 May}, pages = {2484-92}, abstract = {In this study we present the comparative molecular analysis of bacterial communities of the aquatic plant Lemna minor from a contaminated site (RCP) and from a laboratory culture (EPA), as well as each of these with the addition of cadmium. Plants were identified as L. minor by analysis of the rpl16 chloroplast region. Comparative bacterial community studies were based on the analyses of 16S rRNA clone libraries, each containing about 100 clones from the root surfaces of plants. Bacterial communities were compared at three phylogenetic levels of resolution. At the level of bacterial divisions, differences in diversity index scores between treatments, with and without cadmium within the same plant type (EPA or RCP), were small, indicating that cadmium had little effect. When we compared genera within the most dominant group, the beta-proteobacteria, differences between unamended and cadmium-amended libraries were much larger. Bacterial diversity increased upon cadmium addition for both EPA and RCP libraries. Analyses of diversity at the phylotype level showed parallel shifts to more even communities upon cadmium addition; that is, percentage changes in diversity indices due to cadmium addition were the same for either plant type, indicating that contamination history might be independent of disturbance-induced diversity shifts. At finer phylogenetic levels of resolution, the effects of cadmium addition on bacterial communities were very noticeable. This study is a first step in understanding the role of aquatic plant-associated microbial communities in phytoremediation of heavy metals.}, keywords = {Araceae, Bacteria, Base Sequence, Cadmium, Molecular Sequence Data, Phylogeny, Plant Roots, RNA, Ribosomal, 16S}, issn = {0099-2240}, doi = {10.1128/AEM.71.5.2484-2492.2005}, author = {Stout, Lisa M and N{\"u}sslein, Klaus} } @article {750, title = {Simple oligomers as antimicrobial peptide mimics.}, journal = {J Ind Microbiol Biotechnol}, volume = {32}, year = {2005}, month = {2005 Jul}, pages = {296-300}, abstract = {New approaches to antibiotic design are desperately needed. The design of simple oligomers that capture the shape and biological function of natural antimicrobial peptides could prove to be versatile and highly successful. We discuss the use of aromatic backbones to design facially amphiphilic (FA) beta-sheet like structures which are potently antimicrobial. These oligomers capture the physiochemical properties of peptides like the Magainins and Defensins, which fold into specific conformations that are amphiphilic resulting in antimicrobial activity. However, natural peptides are expensive to prepare and difficult to produce on large scale. The design of polymers and oligomers that mimic the complex structures and remarkable biological properties of proteins is an important endeavor and provides attractive alternatives to the difficult synthesis of natural peptides. We therefore have designed a series of FA oligomers that are easy to prepare from inexpensive monomers. They adopt structures very reminiscent of amphiphilic beta-sheets and have significant activity with minimal inhibitory concentrations at 6 h in the low microgram per ml range (muM to nM). They are active against a broad spectrum of bacteria including gram-positive and gram-negative as well as antibiotic resistant strains.}, keywords = {Amides, Anti-Bacterial Agents, Antimicrobial Cationic Peptides, Drug Resistance, Bacterial, Gram-Negative Bacteria, Gram-Positive Bacteria, Inhibitory Concentration 50, Peptides, Protein Conformation}, issn = {1367-5435}, doi = {10.1007/s10295-005-0219-0}, author = {Rennie, Jason and Arnt, Lachelle and Tang, Haizhong and N{\"u}sslein, Klaus and Tew, Gregory N} } @article {822, title = {Allele specific synthetic lethality between priC and dnaAts alleles at the permissive temperature of 30 degrees C in E. coli K-12.}, journal = {BMC Microbiol}, volume = {4}, year = {2004}, month = {2004}, pages = {47}, abstract = {BACKGROUND: DnaA is an essential protein in the regulation and initiation of DNA replication in many bacteria. It forms a protein-DNA complex at oriC to which DnaC loads DnaB. DNA replication forks initiated at oriC by DnaA can collapse on route to the terminus for a variety of reasons. PriA, PriB, PriC, DnaT, Rep and DnaC form multiple pathways to restart repaired replication forks. DnaC809 and dnaC809,820 are suppressors of priA2::kan mutant phenotypes. The former requires PriC and Rep while the latter is independent of them. RnhA339::cat mutations allow DnaA-independent initiation of DNA replication. RESULTS: It is shown herein that a priC303::kan mutation is synthetically lethal with either a dnaA46 or dnaA508 temperature sensitive mutation at the permissive temperature of 30 degrees C. The priC-dnaA lethality is specific for the dnaA allele. The priC303::kan mutant was viable when placed in combination with either dnaA5, dnaA167, dnaA204 or dnaA602. The priC-dnaA508 and priC-dnaA46 lethality could be suppressed by rnhA339::cat. The priC-dnaA508 lethality could be suppressed by a dnaC809,820 mutation, but not dnaC809. Neither of the dnaC mutations could suppress the priC-dnaA46 lethality. CONCLUSIONS: A hitherto unknown function for either DnaA in replication restart or PriC in initiation of DNA replication that occurs in certain dnaA temperature sensitive mutant strains at the permissive temperature of 30 degrees C has been documented. Models considering roles for PriC during initiation of DNA replication and roles for DnaA in replication restart were tested and found not to decisively explain the data. Other roles of dnaA in transcription and nucleoid structure are additionally considered.}, keywords = {Alleles, Bacterial Proteins, DNA Replication, DNA-Binding Proteins, Escherichia coli K12, Escherichia coli Proteins, Gene Expression Regulation, Bacterial, Genes, Lethal, Mutation, Temperature}, issn = {1471-2180}, doi = {10.1186/1471-2180-4-47}, author = {Hinds, Tania and Sandler, Steven J} } @article {716, title = {Caveolin-2 associates with intracellular chlamydial inclusions independently of caveolin-1.}, journal = {BMC Infect Dis}, volume = {4}, year = {2004}, month = {2004 Jul 22}, pages = {23}, abstract = {BACKGROUND: Lipid raft domains form in plasma membranes of eukaryotic cells by the tight packing of glycosphingolipids and cholesterol. Caveolae are invaginated structures that form in lipid raft domains when the protein caveolin-1 is expressed. The Chlamydiaceae are obligate intracellular bacterial pathogens that replicate entirely within inclusions that develop from the phagocytic vacuoles in which they enter. We recently found that host cell caveolin-1 is associated with the intracellular vacuoles and inclusions of some chlamydial strains and species, and that entry of those strains depends on intact lipid raft domains. Caveolin-2 is another member of the caveolin family of proteins that is present in caveolae, but of unknown function. METHODS: We utilized a caveolin-1 negative/caveolin-2 positive FRT cell line and laser confocal immunofluorescence techniques to visualize the colocalization of caveolin-2 with the chlamydial inclusions. RESULTS: We show here that in infected HeLa cells, caveolin-2, as well as caveolin-1, colocalizes with inclusions of C. pneumoniae (Cp), C. caviae (GPIC), and C. trachomatis serovars E, F and K. In addition, caveolin-2 also associates with C. trachomatis serovars A, B and C, although caveolin-1 did not colocalize with these organisms. Moreover, caveolin-2 appears to be specifically, or indirectly, associated with the pathogens at the inclusion membranes. Using caveolin-1 deficient FRT cells, we show that although caveolin-2 normally is not transported out of the Golgi in the absence of caveolin-1, it nevertheless colocalizes with chlamydial inclusions in these cells. However, our results also show that caveolin-2 did not colocalize with UV-irradiated Chlamydia in FRT cells, suggesting that in these caveolin-1 negative cells, pathogen viability and very likely pathogen gene expression are necessary for the acquisition of caveolin-2 from the Golgi. CONCLUSION: Caveolin-2 associates with the chlamydial inclusion independently of caveolin-1. The function of caveolin-2, either in the uninfected cell or in the chlamydial developmental cycle, remains to be elucidated. Nevertheless, this second caveolin protein can now be added to the small number of host proteins that are associated with the inclusions of this obligate intracellular pathogen.}, keywords = {Animals, Blotting, Western, Caveolin 1, Caveolin 2, Caveolins, Cells, Cultured, Chlamydia, Fluorescent Antibody Technique, Golgi Apparatus, Guinea Pigs, HeLa Cells, Humans, Mice, Microscopy, Confocal, Rats, Vacuoles, Virulence}, issn = {1471-2334}, doi = {10.1186/1471-2334-4-23}, author = {Webley, Wilmore C and Norkin, Leonard C and Stuart, Elizabeth S} } @article {857, title = {Cell surface display of the chlamydial glycolipid exoantigen (GLXA) demonstrated by antibody-dependent complement-mediated cytotoxicity.}, journal = {Curr Microbiol}, volume = {49}, year = {2004}, month = {2004 Jul}, pages = {13-21}, abstract = {The chlamydial species are Gram-negative bacterial pathogens critical to human health. Their developmental cycle is associated with the formation and release of the broadly conserved glycolipid exoantigen (GLXA), which has been implicated in the chlamydial elementary body-host cell interaction. This study examines the potential surface display of this glycolipid by chlamydiae-infected cells and the ability of the GLXA they secrete to associate with the plasma membranes of uninfected cells, a prerequisite for exerting influence on them. The sequential incubation of anti-GLXA antibody and complement with Chlamydia trachomatis serovar K or C. pneumoniae AR-39-infected HeLa 229 or macrophage cells resulted in significant cellular cytotoxicity, which preceded the formation of mature elementary bodies. For uninfected cells, co-incubation of GLXA, purified from supernatants of either C. trachomatis or C. pneumoniae-infected HeLa 229 cells, followed by the successive addition of mouse anti-GLXA antibody and complement, yielded similar levels of cellular cytotoxicity. Thus, GLXA indeed is displayed on the surface of infected cells and, therefore, if antibody of appropriate specificity were present, this GLXA could serve to target these infected cells for elimination. Furthermore, released GLXA can associate with uninfected cells and therefore would be positioned to influence their behavior, especially in the context of infection.}, keywords = {Antibody Specificity, Chlamydia, Complement System Proteins, Glycolipids, HeLa Cells, Humans, Polysaccharides, Bacterial}, issn = {0343-8651}, doi = {10.1007/s00284-003-4181-7}, author = {Webley, Wilmore C and Vora, Gary J and Stuart, Elizabeth S} } @article {385, title = {Closing the gaps in kinetoplast DNA network replication.}, journal = {Proc Natl Acad Sci U S A}, volume = {101}, year = {2004}, month = {2004 Mar 30}, pages = {4333-4}, keywords = {Animals, Crithidia fasciculata, DNA Ligases, DNA Replication, DNA, Kinetoplast, DNA, Mitochondrial, Microscopy, Electron, Trypanosoma brucei brucei}, issn = {0027-8424}, doi = {10.1073/pnas.0401400101}, author = {Klingbeil, Michele M and Englund, Paul T} } @article {547, title = {Comparison of 16S rRNA, nifD, recA, gyrB, rpoB and fusA genes within the family Geobacteraceae fam. nov.}, journal = {Int J Syst Evol Microbiol}, volume = {54}, year = {2004}, month = {2004 Sep}, pages = {1591-9}, abstract = {The sequences of five conserved genes, in addition to the 16S rRNA gene, were investigated in 30 members of the Geobacteraceae fam. nov. All members of the Geobacteraceae examined contained nifD, suggesting that they are capable of nitrogen fixation, which may explain their ability to compete effectively in nitrogen-poor subsurface environments undergoing remediation for petroleum or metal contamination. The phylogenies predicted from rpoB, gyrB, fusA, recA and nifD were generally in agreement with the phylogeny predicted from 16S rRNA gene sequences. Furthermore, phylogenetic analysis of concatemers constructed from all five protein-coding genes corresponded closely with the 16S rRNA gene-based phylogeny. This study demonstrated that the Geobacteraceae is a phylogenetically coherent family within the delta-subclass of the Proteobacteria that is composed of three distinct phylogenetic clusters: Geobacter, Desulfuromonas and Desulfuromusa. The sequence data provided here will make it possible to discriminate better between physiologically distinct members of the Geobacteraceae, such as Pelobacter propionicus and Geobacter species, in geobacteraceae-dominated microbial communities and greatly expands the potential to identify geobacteraceae sequences in libraries of environmental genomic DNA.}, keywords = {Bacterial Proteins, Deltaproteobacteria, Desulfuromonas, DNA Gyrase, DNA, Bacterial, DNA, Ribosomal, DNA-Directed RNA Polymerases, Genes, Bacterial, Genes, rRNA, Geobacter, Molecular Sequence Data, Nitrogen Fixation, Peptide Elongation Factor G, Phylogeny, Rec A Recombinases, RNA, Bacterial, RNA, Ribosomal, 16S, Sequence Analysis, DNA}, issn = {1466-5026}, doi = {10.1099/ijs.0.02958-0}, author = {Holmes, Dawn E and Nevin, Kelly P and Lovley, Derek R} } @article {550, title = {Computational prediction of conserved operons and phylogenetic footprinting of transcription regulatory elements in the metal-reducing bacterial family Geobacteraceae.}, journal = {J Theor Biol}, volume = {230}, year = {2004}, month = {2004 Sep 7}, pages = {133-44}, abstract = {Members of the family Geobacteraceae are an important group of microorganisms from the delta subdivision of Proteobacteria that couple the oxidation of organic compounds to metal reduction. In order to uncover transcription regulatory interactions in these organisms, we used computational methods to identify conserved operons and putative cis-regulatory transcription elements. We identified 26 putative operons with gene order and function conserved among two species of Geobacteraceae, Geobacter sulfurreducens and Geobacter metallireducens. Most of these operons were also conserved in Desulfovibrio vulgaris, an additional metal reducing organism from family Desulfovibrionaceae of the delta subdivision of Proteobacteria. The predicted conserved operons were investigated for the presence of transcription factor binding sites by two different methods, (i) comparison of non-coding regions in conserved operons, and (ii) neural network promoter prediction. Predicted motifs were screened to identify most likely transcription factor binding sites and ribosome-binding sites. We provide information on motifs in Geobacteraceae similar to known transcription factor binding sites in Escherichia coli, conserved motifs in other bacterial species, putative palindromic sites, and predicted ribosome-binding sites. These predictions will aid in further elucidation of regulatory networks of gene interactions in Geobacteraceae.}, keywords = {Animals, Computational Biology, Conserved Sequence, DNA Footprinting, Genes, Regulator, Genome, Bacterial, Geobacter, Models, Genetic, Operon, Phylogeny}, issn = {0022-5193}, doi = {10.1016/j.jtbi.2004.04.022}, author = {Yan, Bin and Meth{\'e}, Barbara A and Lovley, Derek R and Krushkal, Julia} } @article {548, title = {Direct correlation between rates of anaerobic respiration and levels of mRNA for key respiratory genes in Geobacter sulfurreducens.}, journal = {Appl Environ Microbiol}, volume = {70}, year = {2004}, month = {2004 Sep}, pages = {5183-9}, abstract = {The predominance of Geobacter species in environments in which Fe(III) reduction is important has suggested that Fe(III) reduction rates might be estimated in Geobacter-dominated environments by assessing in situ activity with molecular techniques. To determine whether mRNA levels of key respiratory genes might be correlated with respiration rates in Geobacter sulfurreducens, studies were conducted with fumarate as the electron acceptor and acetate as the limiting electron donor in anaerobic continuous cultures. Levels of mRNA for a fumarate reductase gene, frdA, quantified by real-time reverse transcription-PCR were directly correlated with fumarate reduction rates. In similar studies with Fe(III) as the electron acceptor, mRNA levels for omcB, a gene for an outer membrane c-type cytochrome involved in Fe(III) reduction, were positively correlated with Fe(III) reduction rates. Levels of mRNA for frdA and omcB were also positively correlated with fumarate and Fe(III) reduction rates, respectively, when growth was limited by the availability of fumarate or Fe(III), but mRNA levels were higher than in acetate-limited cultures. Levels of mRNA for omcC, which encodes a c-type cytochrome highly similar to OmcB but not necessary for Fe(III) reduction, followed patterns different than those of omcB. This agrees with the previous finding that OmcC is not involved in Fe(III) reduction and suggests that changes in mRNA levels of omcB are related to its role in Fe(III) reduction. These results demonstrate that mRNA levels for respiratory genes might be used to estimate in situ Fe(III) reduction rates in Geobacter-dominated environments but suggest that information on environmental conditions and/or the metabolic state of Geobacter species is also required for accurate rate estimates.}, keywords = {Anaerobiosis, Base Sequence, DNA Primers, Geobacter, Kinetics, Oxygen Consumption, Polymerase Chain Reaction, RNA, Messenger}, issn = {0099-2240}, doi = {10.1128/AEM.70.9.5183-5189.2004}, author = {Chin, Kuk-Jeong and Esteve-N{\'u}{\~n}ez, Abraham and Leang, Ching and Lovley, Derek R} } @article {545, title = {Dissimilatory Fe(III) and Mn(IV) reduction.}, journal = {Adv Microb Physiol}, volume = {49}, year = {2004}, month = {2004}, pages = {219-86}, abstract = {Dissimilatory Fe(III) and Mn(IV) reduction has an important influence on the geochemistry of modern environments, and Fe(III)-reducing microorganisms, most notably those in the Geobacteraceae family, can play an important role in the bioremediation of subsurface environments contaminated with organic or metal contaminants. Microorganisms with the capacity to conserve energy from Fe(III) and Mn(IV) reduction are phylogenetically dispersed throughout the Bacteria and Archaea. The ability to oxidize hydrogen with the reduction of Fe(III) is a highly conserved characteristic of hyperthermophilic microorganisms and one Fe(III)-reducing Archaea grows at the highest temperature yet recorded for any organism. Fe(III)- and Mn(IV)-reducing microorganisms have the ability to oxidize a wide variety of organic compounds, often completely to carbon dioxide. Typical alternative electron acceptors for Fe(III) reducers include oxygen, nitrate, U(VI) and electrodes. Unlike other commonly considered electron acceptors, Fe(III) and Mn(IV) oxides, the most prevalent form of Fe(III) and Mn(IV) in most environments, are insoluble. Thus, Fe(III)- and Mn(IV)-reducing microorganisms face the dilemma of how to transfer electrons derived from central metabolism onto an insoluble, extracellular electron acceptor. Although microbiological and geochemical evidence suggests that Fe(III) reduction may have been the first form of microbial respiration, the capacity for Fe(III) reduction appears to have evolved several times as phylogenetically distinct Fe(III) reducers have different mechanisms for Fe(III) reduction. Geobacter species, which are representative of the family of Fe(III) reducers that predominate in a wide diversity of sedimentary environments, require direct contact with Fe(III) oxides in order to reduce them. In contrast, Shewanella and Geothrix species produce chelators that solubilize Fe(III) and release electron-shuttling compounds that transfer electrons from the cell surface to the surface of Fe(III) oxides not in direct contact with the cells. Electron transfer from the inner membrane to the outer membrane in Geobacter and Shewanella species appears to involve an electron transport chain of inner-membrane, periplasmic, and outer-membrane c-type cytochromes, but the cytochromes involved in these processes in the two organisms are different. In addition, Geobacter species specifically express flagella and pili during growth on Fe(III) and Mn(IV) oxides and are chemotactic to Fe(II) and Mn(II), which may lead Geobacter species to the oxides under anoxic conditions. The physiological characteristics of Geobacter species appear to explain why they have consistently been found to be the predominant Fe(III)- and Mn(IV)-reducing microorganisms in a variety of sedimentary environments. In comparison with other respiratory processes, the study of Fe(III) and Mn(IV) reduction is in its infancy, but genome-enabled approaches are rapidly advancing our understanding of this environmentally significant physiology.}, keywords = {Archaea, Biodegradation, Environmental, Ferric Compounds, Geobacter, Manganese, Manganese Compounds, Oxidation-Reduction, Oxides, Shewanella, Soil Microbiology, Soil Pollutants}, issn = {0065-2911}, doi = {10.1016/S0065-2911(04)49005-5}, author = {Lovley, Derek R and Holmes, Dawn E and Nevin, Kelly P} } @article {824, title = {A dnaT mutant with phenotypes similar to those of a priA2::kan mutant in Escherichia coli K-12.}, journal = {Genetics}, volume = {167}, year = {2004}, month = {2004 Jun}, pages = {569-78}, abstract = {The ability to repair damaged replication forks and restart them is important for cell survival. DnaT is essential for replication restart in vitro and yet no definite genetic analysis has been done in Escherichia coli K-12. To begin, dnaT822, an in-frame six-codon (87-92) deletion was constructed. DnaT822 mutants show colony size, cell morphology, inability to properly partition nucleoids, UV sensitivity, and basal SOS expression similar to priA2::kan mutants. DnaT822 priA2::kan double mutants had phenotypes similar to those of the single mutants. DnaT822 and dnaT822 priA2::kan mutant phenotypes were fully suppressed by dnaC809. Previously, a dominant temperature-sensitive lethal mutation, dnaT1, had been isolated in E. coli 15T(-). DnaT1 was found to have a base-pair change relative to the E. coli 15T(-) and E. coli K-12 dnaT genes that led to a single amino acid change: R152C. A plasmid-encoded E. coli K-12 mutant dnaT gene with the R152C amino acid substitution did not display a dominant temperature-sensitive lethal phenotype in a dnaT(+) strain of E. coli K-12. Instead, this mutant dnaT gene was found to complement the E. coli K-12 dnaT822 mutant phenotypes. The significance of these results is discussed in terms of models for replication restart.}, keywords = {Base Sequence, Codon, DNA Primers, DNA Repair, DNA Replication, DNA, Bacterial, DNA-Binding Proteins, Escherichia coli K12, Escherichia coli Proteins, Molecular Sequence Data, Mutation, Plasmids, Polymerase Chain Reaction, Sequence Deletion}, issn = {0016-6731}, doi = {10.1534/genetics.103.025296}, author = {McCool, Jesse D and Ford, Christopher C and Sandler, Steven J} } @article {559, title = {Electron transfer by Desulfobulbus propionicus to Fe(III) and graphite electrodes.}, journal = {Appl Environ Microbiol}, volume = {70}, year = {2004}, month = {2004 Feb}, pages = {1234-7}, abstract = {Desulfobulbus propionicus was able to grow with Fe(III), the humic acids analog anthraquinone-2,6-disulfonate (AQDS), or a graphite electrode as an electron acceptor. These results provide an explanation for the enrichment of Desulfobulbaceae species on the surface of electrodes harvesting electricity from anaerobic marine sediments and further expand the diversity of microorganisms known to have the ability to use both sulfate and Fe(III) as an electron acceptor.}, keywords = {Culture Media, Deltaproteobacteria, Electrodes, Electron Transport, Ferric Compounds, Graphite, Oxidation-Reduction, Pyruvic Acid, Sulfates}, issn = {0099-2240}, author = {Holmes, Dawn E and Bond, Daniel R and Lovley, Derek R} } @article {397, title = {Genetic evidence for pre-recruitment as the mechanism of transcription activation by SoxS of Escherichia coli: the dominance of DNA binding mutations of SoxS.}, journal = {J Mol Biol}, volume = {344}, year = {2004}, month = {2004 Nov 12}, pages = {1-10}, abstract = {SoxS, the direct transcriptional activator of the Escherichia coli superoxide (SoxRS) regulon, displays several unusual characteristics which suggest that it is unlikely to activate transcription by the ususal recruitment mechanism. Thus, agents that generate superoxide endogenously and thereby provoke the defense response elicit the de novo synthesis of SoxS, and with the SoxS binding site being highly degenerate, the number of SoxS binding sites per cell far exceeds the number of SoxS molecules per cell. To account for these distinctive features of the SoxRS system, we proposed "pre-recruitment" as the mechanism by which SoxS activates transcription of the regulon{\textquoteright}s genes. In pre-recruitment, newly synthesized SoxS molecules form binary complexes with RNA polymerase in solution. These complexes provide the information content to allow the 2500 molecules of SoxS per cell to scan the 65,000 SoxS binding sites per cell for the 200 binding sites per cell that reside within SoxS-dependent promoters. As a test of whether SoxS activates transcription by recruitment or pre-recruitment, we determined the dominance relationships of SoxS mutations conferring defective DNA binding. We found that soxS DNA binding mutations are dominant to the wild-type allele, a result consistent with the pre-recruitment hypothesis, but opposite to that expected for an activator that functions by recruitment. Moreover, whereas positive control mutations of activators functioning by recruitment are usually dominant, a soxS positive control mutation was not. Lastly, with the SoxRS system as an example, we discuss the physiological requirement for stringent regulation of transcriptional activators that function by pre-recruitment.}, keywords = {Base Sequence, Binding Sites, DNA, Bacterial, Escherichia coli, Escherichia coli Proteins, Gene Expression Regulation, Bacterial, Genes, Bacterial, Genes, Dominant, Mutation, Regulon, Trans-Activators, Transcriptional Activation}, issn = {0022-2836}, doi = {10.1016/j.jmb.2004.09.007}, author = {Griffith, Kevin L and Wolf, Richard E} } @article {557, title = {Geobacter sulfurreducens can grow with oxygen as a terminal electron acceptor.}, journal = {Appl Environ Microbiol}, volume = {70}, year = {2004}, month = {2004 Apr}, pages = {2525-8}, abstract = {Geobacter sulfurreducens, previously classified as a strict anaerobe, tolerated exposure to atmospheric oxygen for at least 24 h and grew with oxygen as the sole electron acceptor at concentrations of 10\% or less in the headspace. These results help explain how Geobacter species may survive in oxic subsurface environments, being poised to rapidly take advantage of the development of anoxic conditions.}, keywords = {Aerobiosis, Anaerobiosis, Electron Transport, Environmental Microbiology, Geobacter, Oxygen}, issn = {0099-2240}, author = {Lin, W C and Coppi, M V and Lovley, D R} } @article {552, title = {Graphite electrodes as electron donors for anaerobic respiration.}, journal = {Environ Microbiol}, volume = {6}, year = {2004}, month = {2004 Jun}, pages = {596-604}, abstract = {It has been demonstrated previously that Geobacter species can transfer electrons directly to electrodes. In order to determine whether electrodes could serve as electron donors for microbial respiration, enrichment cultures were established from a sediment inoculum with a potentiostat-poised graphite electrode as the sole electron donor and nitrate as the electron acceptor. Nitrate was reduced to nitrite with the consumption of electrical current. The stoichiometry of electron and nitrate consumption and nitrite accumulation were consistent with the electrode serving as the sole electron donor for nitrate reduction. Analysis of 16 rRNA gene sequences demonstrated that the electrodes supplied with current were specifically enriched in microorganisms with sequences most closely related to the sequences of known Geobacter species. A pure culture of Geobacter metallireducens was shown to reduce nitrate to nitrite with the electrode as the sole electron donor with the expected stoichiometry of electron consumption. Cells attached to the electrode appeared to be responsible for the nitrate reduction. Attached cells of Geobacter sulfurreducens reduced fumarate to succinate with the electrode as an electron donor. These results demonstrate for the first time that electrodes may serve as a direct electron donor for anaerobic respiration. This finding has implications for the harvesting of electricity from anaerobic sediments and the bioremediation of oxidized contaminants.}, keywords = {Anaerobiosis, Electrodes, Electrons, Fumarates, Geobacter, Geologic Sediments, Graphite, Kinetics, Microscopy, Electron, Scanning, Nitrates, Phylogeny, RNA, Ribosomal, 16S, Sequence Analysis, DNA}, issn = {1462-2912}, doi = {10.1111/j.1462-2920.2004.00593.x}, author = {Gregory, Kelvin B and Bond, Daniel R and Lovley, Derek R} } @article {1449, title = {High-throughput method for detecting genomic-deletion polymorphisms.}, journal = {J Clin Microbiol}, volume = {42}, year = {2004}, month = {2004 Jul}, pages = {2913-8}, abstract = {DNA microarrays have been successfully used with different microorganisms, including Mycobacterium tuberculosis, to detect genomic deletions relative to a reference strain. However, the cost and complexity of the microarray system are obstacles to its widespread use in large-scale studies. In order to evaluate the extent and role of large sequence polymorphisms (LSPs) or insertion-deletion events in bacterial populations, we developed a technique, termed deligotyping, which hybridizes multiplex-PCR products to membrane-bound, highly specific oligonucleotide probes. The approach has the benefits of being low cost and capable of simultaneously interrogating more than 40 bacterial strains for the presence of 43 genomic regions. The deletions represented on the membrane were selected from previous comparative genomic studies and ongoing microarray experiments. Highly specific probes for these deletions were designed and attached to a membrane for hybridization with strain-derived targets. The targets were generated by multiplex PCR, allowing simultaneous amplifications of 43 different genomic loci in a single reaction. To validate our approach, 100 strains that had been analyzed with a high-density microarray were analyzed. The membrane accurately detected the deletions identified by the microarray approach, with a sensitivity of 99.9\% and a specificity of 98.0\%. The deligotyping technique allows the rapid and reliable screening of large numbers of M. tuberculosis isolates for LSPs. This technique can be used to provide insights into the epidemiology, genomic evolution, and population structure of M. tuberculosis and can be adapted for the study of other organisms.
}, keywords = {DNA Probes, Genome, Bacterial, Mycobacterium tuberculosis, Oligonucleotide Array Sequence Analysis, Polymerase Chain Reaction, Polymorphism, Genetic}, issn = {0095-1137}, doi = {10.1128/JCM.42.7.2913-2918.2004}, author = {Goguet de la Salmoni{\`e}re, Yves-Olivier Luc and Kim, C C and Tsolaki, A G and Pym, A S and Siegrist, M S and Small, Peter M} } @article {363, title = {Horizontal transmission of Hz-2V by virus infected Helicoverpa zea moths.}, journal = {J Invertebr Pathol}, volume = {85}, year = {2004}, month = {2004 Feb}, pages = {128-31}, abstract = {Helicoverpa zea female moths productively infected with Hz-2V have malformed reproductive tissues and are sterile. Virus replication in infected females occurs primarily in the reproductive tissues and culminates with the accumulation of virus-filled vesicles, which form plugs of virus covering the reproductive openings of these insects. The location of this large concentration of virus particles at the terminal abdominal segment of infected females suggests that it may serve as a source of virus that can be transmitted horizontally between moths during mating. In mating experiments it was found that healthy males are attracted to and attempt to mate with infected females, and that these males are able transmit Hz-2V to healthy females during subsequent matings, giving rise to virus infected progeny.}, keywords = {Abdomen, Animals, Entomology, Female, Genitalia, Insect Vectors, Insect Viruses, Male, Moths, Sexual Behavior, Animal, Tissue Distribution}, issn = {0022-2011}, doi = {10.1016/j.jip.2004.01.004}, author = {Burand, John P and Rallis, Christopher P and Tan, Weijia} } @article {555, title = {Identification of an uptake hydrogenase required for hydrogen-dependent reduction of Fe(III) and other electron acceptors by Geobacter sulfurreducens.}, journal = {J Bacteriol}, volume = {186}, year = {2004}, month = {2004 May}, pages = {3022-8}, abstract = {Geobacter sulfurreducens, a representative of the family Geobacteraceae that predominates in Fe(III)-reducing subsurface environments, can grow by coupling the oxidation of hydrogen to the reduction of a variety of electron acceptors, including Fe(III), fumarate, and quinones. An examination of the G. sulfurreducens genome revealed two operons, hya and hyb, which appeared to encode periplasmically oriented respiratory uptake hydrogenases. In order to assess the roles of these two enzymes in hydrogen-dependent growth, Hya- and Hyb-deficient mutants were generated by gene replacement. Hyb was found to be required for hydrogen-dependent reduction of Fe(III), anthraquinone-2,6-disulfonate, and fumarate by resting cell suspensions and to be essential for growth with hydrogen and these three electron acceptors. Hya, in contrast, was not. These findings suggest that Hyb is an essential respiratory hydrogenase in G. sulfurreducens.}, keywords = {Anthraquinones, Ferric Compounds, Fumarates, Geobacter, Hydrogen, Operon, Oxidation-Reduction, Oxidoreductases, Phenotype}, issn = {0021-9193}, author = {Coppi, Maddalena V and O{\textquoteright}Neil, Regina A and Lovley, Derek R} } @article {41, title = {Identification of novel dominant INO2c mutants with an Opi- phenotype.}, journal = {Mol Microbiol}, volume = {52}, year = {2004}, month = {2004 Jun}, pages = {1271-80}, abstract = {The INO2 gene of Saccharomyces cerevisiae is required for derepression of the phospholipid biosynthetic genes in response to inositol depletion. Conversely, the OPI1 gene is required for repression in response to inositol supplementation. Results of an in vitro assay have led to a model in which Opi1p interacts with Ino2p. However, there is no in vivo evidence to support this model. Additionally, most of the previously isolated ino2 mutants offer little insight into this model. Here, we report the isolation of a new class of dominant mutations in the INO2 gene, which yield constitutive expression of a target gene (i.e. an Opi(-) mutant phenotype). Two mutations reside in a region of the Ino2p required for interaction with Opi1p in vitro. Three other mutations are at the amino-terminus in a transcriptional activation domain.}, keywords = {Basic Helix-Loop-Helix Transcription Factors, Gene Expression Regulation, Fungal, Inositol, Mutation, Phenotype, Repressor Proteins, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins, Transcription Factors}, issn = {0950-382X}, doi = {10.1111/j.1365-2958.2004.04069.x}, author = {Gardenour, Kyle R and Levy, Jamie and Lopes, John M} } @article {543, title = {In situ expression of nifD in Geobacteraceae in subsurface sediments.}, journal = {Appl Environ Microbiol}, volume = {70}, year = {2004}, month = {2004 Dec}, pages = {7251-9}, abstract = {In order to determine whether the metabolic state of Geobacteraceae involved in bioremediation of subsurface sediments might be inferred from levels of mRNA for key genes, in situ expression of nifD, a highly conserved gene involved in nitrogen fixation, was investigated. When Geobacter sulfurreducens was grown without a source of fixed nitrogen in chemostats with acetate provided as the limiting electron donor and Fe(III) as the electron acceptor, levels of nifD transcripts were 4 to 5 orders of magnitude higher than in chemostat cultures provided with ammonium. In contrast, the number of transcripts of recA and the 16S rRNA gene were slightly lower in the absence of ammonium. The addition of acetate to organic- and nitrogen-poor subsurface sediments stimulated the growth of Geobacteraceae and Fe(III) reduction, as well as the expression of nifD in Geobacteraceae. Levels of nifD transcripts in Geobacteraceae decreased more than 100-fold within 2 days after the addition of 100 microM ammonium, while levels of recA and total bacterial 16S rRNA in Geobacteraceae remained relatively constant. Ammonium amendments had no effect on rates of Fe(III) reduction in acetate-amended sediments or toluene degradation in petroleum-contaminated sediments, suggesting that other factors, such as the rate that Geobacteraceae could access Fe(III) oxides, limited Fe(III) reduction. These results demonstrate that it is possible to monitor one aspect of the in situ metabolic state of Geobacteraceae species in subsurface sediments via analysis of mRNA levels, which is the first step toward a more global analysis of in situ gene expression related to nutrient status and stress response during bioremediation by Geobacteraceae.}, keywords = {Acetates, Biodegradation, Environmental, Culture Media, DNA, Ribosomal, Fresh Water, Gene Expression Regulation, Bacterial, Geobacter, Geologic Sediments, Nitrogenase, Petroleum, Phylogeny, Polymerase Chain Reaction, Quaternary Ammonium Compounds, Rec A Recombinases, RNA, Ribosomal, 16S, Water Pollutants, Chemical}, issn = {0099-2240}, doi = {10.1128/AEM.70.12.7251-7259.2004}, author = {Holmes, Dawn E and Nevin, Kelly P and Lovley, Derek R} } @article {362, title = {In vivo dose-response of insects to Hz-2V infection.}, journal = {Virol J}, volume = {1}, year = {2004}, month = {2004}, pages = {15}, abstract = {BACKGROUND: Hz-2V infection of female Helicoverpa zea moths is manifested as insects that are either sterile "agonadal" individuals with malformed reproductive tissues or fertile asymptomatic carriers which are capable of transmitting virus on to their progeny. Virus infected progeny arising from eggs laid by asymptomatic carrier females may themselves be either sterile agonadals or asymptomatic carriers. RESULTS: By injecting virus into female moths, a correlation was established between virus doses administered to the females and the levels of resulting asymptomatic and sterile progeny. CONCLUSIONS: The results of these experiments indicate that high virus doses produced a higher level of agonadal progeny and lower doses produced higher levels of asymptomatic carriers.}, keywords = {Animals, Female, Insect Viruses, Male, Moths, Ovum, Virus Diseases}, issn = {1743-422X}, doi = {10.1186/1743-422X-1-15}, author = {Burand, John P and Rallis, Christopher P} } @article {554, title = {Isolation, characterization, and U(VI)-reducing potential of a facultatively anaerobic, acid-resistant Bacterium from Low-pH, nitrate- and U(VI)-contaminated subsurface sediment and description of Salmonella subterranea sp. nov.}, journal = {Appl Environ Microbiol}, volume = {70}, year = {2004}, month = {2004 May}, pages = {2959-65}, abstract = {A facultatively anaerobic, acid-resistant bacterium, designated strain FRCl, was isolated from a low-pH, nitrate- and U(VI)-contaminated subsurface sediment at site FW-024 at the Natural and Accelerated Bioremediation Research Field Research Center in Oak Ridge, Tenn. Strain FRCl was enriched at pH 4.5 in minimal medium with nitrate as the electron acceptor, hydrogen as the electron donor, and acetate as the carbon source. Clones with 16S ribosomal DNA (rDNA) sequences identical to the sequence of strain FRCl were also detected in a U(VI)-reducing enrichment culture derived from the same sediment. Cells of strain FRCl were gram-negative motile regular rods 2.0 to 3.4 micro m long and 0.7 to 0.9 microm in diameter. Strain FRCl was positive for indole production, by the methyl red test, and for ornithine decarboxylase; it was negative by the Voges-Proskauer test (for acetylmethylcarbinol production), for urea hydrolysis, for arginine dihydrolase, for lysine decarboxylase, for phenylalanine deaminase, for H(2)S production, and for gelatin hydrolysis. Strain FRCl was capable of using O(2), NO(3)(-), S(2)O(3)(2-), fumarate, and malate as terminal electron acceptors and of reducing U(VI) in the cell suspension. Analysis of the 16S rDNA sequence of the isolate indicated that this strain was 96.4\% similar to Salmonella bongori and 96.3\% similar to Enterobacter cloacae. Physiological and phylogenetic analyses suggested that strain FRCl belongs to the genus Salmonella and represents a new species, Salmonella subterranea sp. nov.}, keywords = {Anaerobiosis, Culture Media, DNA, Ribosomal, Fresh Water, Geologic Sediments, Hydrogen-Ion Concentration, Molecular Sequence Data, Nitrates, Oxidation-Reduction, Phylogeny, RNA, Ribosomal, 16S, Salmonella, Sequence Analysis, DNA, Uranium, Water Pollution, Chemical}, issn = {0099-2240}, author = {Shelobolina, Evgenya S and Sullivan, Sara A and O{\textquoteright}Neill, Kathleen R and Nevin, Kelly P and Lovley, Derek R} } @article {551, title = {MacA, a diheme c-type cytochrome involved in Fe(III) reduction by Geobacter sulfurreducens.}, journal = {J Bacteriol}, volume = {186}, year = {2004}, month = {2004 Jun}, pages = {4042-5}, abstract = {A 36-kDa diheme c-type cytochrome abundant in Fe(III)-respiring Geobacter sulfurreducens, designated MacA, was more highly expressed during growth with Fe(III) as the electron acceptor than with fumarate. Although MacA has homology to proteins with in vitro peroxidase activity, deletion of macA had no impact on response to oxidative stress. However, the capacity for Fe(III) reduction was greatly diminished, indicating that MacA, which is predicted to be localized in the periplasm, is a key intermediate in electron transfer to Fe(III).}, keywords = {Amino Acid Sequence, Bacterial Proteins, Cytochrome c Group, Deltaproteobacteria, Electron Transport, Ferric Compounds, Gene Deletion, Molecular Sequence Data, Oxidation-Reduction}, issn = {0021-9193}, doi = {10.1128/JB.186.12.4042-4045.2004}, author = {Butler, Jessica E and Kaufmann, Franz and Coppi, Maddalena V and N{\'u}{\~n}ez, Cinthia and Lovley, Derek R} } @article {823, title = {Measurement of SOS expression in individual Escherichia coli K-12 cells using fluorescence microscopy.}, journal = {Mol Microbiol}, volume = {53}, year = {2004}, month = {2004 Sep}, pages = {1343-57}, abstract = {Many recombination, DNA repair and DNA replication mutants have high basal levels of SOS expression as determined by a sulAp-lacZ reporter gene system on a population of cells. Two opposing models to explain how the SOS expression is distributed in these cells are: (i) the {\textquoteright}Uniform Expression Model (UEM){\textquoteright} where expression is evenly distributed in all cells or (ii) the {\textquoteright}Two Population Model (TPM){\textquoteright} where some cells are highly induced while others are not at all. To distinguish between these two models, a method to quantify SOS expression in individual bacterial cells was developed by fusing an SOS promoter (sulAp) to the green fluorescent protein (gfp) reporter gene and inserting it at attlambda on the Escherichia coli chromosome. It is shown that the fluorescence in sulAp-gfp cells is regulated by RecA and LexA. This system was then used to distinguish between the two models for several mutants. The patterns displayed by priA, dnaT, recG, uvrD, dam, ftsK, rnhA, polA and xerC mutants were explained best by the TPM while only lexA (def), lexA3 (ind-) and recA defective mutants were explained best by the UEM. These results are discussed in a context of how the processes of DNA replication and recombination may affect cells in a population differentially.}, keywords = {Adenosine Triphosphatases, DNA Damage, DNA Helicases, DNA Repair, Escherichia coli K12, Escherichia coli Proteins, Gene Expression Regulation, Bacterial, Genes, Reporter, Microscopy, Fluorescence, Models, Genetic, Promoter Regions, Genetic, Recombinant Fusion Proteins, SOS Response (Genetics), Ultraviolet Rays}, issn = {0950-382X}, doi = {10.1111/j.1365-2958.2004.04225.x}, author = {McCool, Jesse D and Long, Edward and Petrosino, Joseph F and Sandler, Hilary A and Rosenberg, Susan M and Sandler, Steven J} } @article {544, title = {Microbial communities associated with electrodes harvesting electricity from a variety of aquatic sediments.}, journal = {Microb Ecol}, volume = {48}, year = {2004}, month = {2004 Aug}, pages = {178-90}, abstract = {The microbial communities associated with electrodes from underwater fuel cells harvesting electricity from five different aquatic sediments were investigated. Three fuel cells were constructed with marine, salt-marsh, or freshwater sediments incubated in the laboratory. Fuel cells were also deployed in the field in salt marsh sediments in New Jersey and estuarine sediments in Oregon, USA. All of the sediments produced comparable amounts of power. Analysis of 16S rRNA gene sequences after 3-7 months of incubation demonstrated that all of the energy-harvesting anodes were highly enriched in microorganisms in the delta-Proteobacteria when compared with control electrodes not connected to a cathode. Geobacteraceae accounted for the majority of delta-Proteobacterial sequences or all of the energy-harvesting anodes, except the one deployed at the Oregon estuarine site. Quantitative PCR analysis of 16S rRNA genes and culturing studies indicated that Geobacteraceae were 100-fold more abundant on the marine-deployed anodes versus controls. Sequences most similar to microorganisms in the family Desulfobulbaceae predominated on the anode deployed in the estuarine sediments, and a significant proportion of the sequences recovered from the freshwater anodes were closely related to the Fe(III)-reducing isolate, Geothrix fermentans. There was also a specific enrichment of microorganisms on energy harvesting cathodes, but the enriched populations varied with the sediment/water source. Thus, future studies designed to help optimize the harvesting of electricity from aquatic sediments or waste organic matter should focus on the electrode interactions of these microorganisms which are most competitive in colonizing anodes and cathodes.}, keywords = {Base Sequence, Biodiversity, Bioelectric Energy Sources, Cloning, Molecular, DNA Primers, Electrodes, Gammaproteobacteria, Geologic Sediments, Molecular Sequence Data, Phylogeny, Polymerase Chain Reaction, Restriction Mapping, RNA, Ribosomal, 16S, Sequence Analysis, DNA}, issn = {0095-3628}, doi = {10.1007/s00248-003-0004-4}, author = {Holmes, D E and Bond, D R and O{\textquoteright}Neil, R A and Reimers, C E and Tender, L R and Lovley, D R} } @article {861, title = {Poly(hydroxyalkanoic acid) Biosynthesis in Ectothiorhodospirashaposhnikovii: Characterization and Reactivity of a Type III PHA Synthase.}, journal = {Biomacromolecules}, volume = {5}, year = {2004}, month = {2004 Jan-Feb}, pages = {40-8}, abstract = {Ectothiorhodospira shaposhnikovii is able to accumulate polyhydroxybutyrate (PHB) photoautotrophically during nitrogen-limited growth. The activity of polyhydroxyalkanoate (PHA) synthase in the cells correlates with PHB accumulation. PHA synthase samples collected during the light period do not show a lag phase during in vitro polymerization. Synthase samples collected in the dark period displays a significant lag phase during in vitro polymerization. The lag phase can be eliminated by reacting the PHA synthase with the monomer, 3-hydroxybutyryl-CoA (3HBCoA). The PHA synthase genes (phaC and phaE) were cloned by screening a genomic library for PHA accumulation in E. coli cells. The PHA synthase expressed in the recombinant E. coli cells was purified to homogeneity. Both sequence analysis and biochemical studies indicated that this PHA synthase consists of two subunits, PhaE and PhaC and, therefore, belongs to the type III PHA synthases. Two major complexes were identified in preparations of purified PHA synthase. The large complex appears to be composed of 12 PhaC subunits and 12 PhaE subunits (dodecamer), whereas the small complex appears to be composed of 6 PhaC and 6 PhaE subunits (hexamer). In dilute aqueous solution, the synthase is predominantly composed of hexamer and has low activity accompanied with a significant lag period at the initial stage of reaction. The percentage of dodecameric complex increases with increasing salt concentration. The dodecameric complex has a greatly increased specific activity for the polymerization of 3HBCoA and a negligible lag period. The results from in vitro polymerizations of 3HBCoA suggest that the PHA synthase from E. shaposhnikovii may catalyze a living polymerization and demonstrate that two PhaC and two PhaE subunits comprise a single catalytic site in the synthase complex.}, keywords = {Acyltransferases, Cloning, Molecular, Ectothiorhodospira shaposhnikovii, Escherichia coli, Hydroxybutyrates, Kinetics, Molecular Weight, Polyesters}, issn = {1525-7797}, doi = {10.1021/bm034171i}, author = {Zhang, Shiming and Kolvek, Steven and Goodwin, Steve and Lenz, Robert W} } @article {546, title = {Potential role of a novel psychrotolerant member of the family Geobacteraceae, Geopsychrobacter electrodiphilus gen. nov., sp. nov., in electricity production by a marine sediment fuel cell.}, journal = {Appl Environ Microbiol}, volume = {70}, year = {2004}, month = {2004 Oct}, pages = {6023-30}, abstract = {Previous studies have shown that members of the family Geobacteraceae that attach to the anodes of sediment fuel cells are directly involved in harvesting electricity by oxidizing organic compounds to carbon dioxide and transferring the electrons to the anode. In order to learn more about this process, microorganisms from the anode surface of a marine sediment fuel cell were enriched and isolated with Fe(III) oxide. Two unique marine isolates were recovered, strains A1(T) and A2. They are gram-negative, nonmotile rods, with abundant c-type cytochromes. Phylogenetic analysis of the 16S rRNA, recA, gyrB, fusA, rpoB, and nifD genes indicated that strains A1(T) and A2 represent a unique phylogenetic cluster within the Geobacteraceae. Both strains were able to grow with an electrode serving as the sole electron acceptor and transferred ca. 90\% of the electrons available in their organic electron donors to the electrodes. These organisms are the first psychrotolerant members of the Geobacteraceae reported thus far and can grow at temperatures between 4 and 30 degrees C, with an optimum temperature of 22 degrees C. Strains A1(T) and A2 can utilize a wide range of traditional electron acceptors, including all forms of soluble and insoluble Fe(III) tested, anthraquinone 2,6-disulfonate, and S(0). In addition to acetate, both strains can utilize a number of other organic acids, amino acids, long-chain fatty acids, and aromatic compounds to support growth with Fe(III) nitrilotriacetic acid as an electron acceptor. The metabolism of these organisms differs in that only strain A1(T) can use acetoin, ethanol, and hydrogen as electron donors, whereas only strain A2 can use lactate, propionate, and butyrate. The name Geopsychrobacter electrodiphilus gen. nov., sp. nov., is proposed for strains A1(T) and A2, with strain A1(T) (ATCC BAA-880(T); DSM 16401(T); JCM 12469) as the type strain. Strains A1(T) and A2 (ATCC BAA-770; JCM 12470) represent the first organisms recovered from anodes that can effectively couple the oxidation of organic compounds to an electrode. Thus, they may serve as important model organisms for further elucidation of the mechanisms of microbe-electrode electron transfer in sediment fuel cells.}, keywords = {Bioelectric Energy Sources, Cytochromes, Deltaproteobacteria, Electron Transport, Genes, Bacterial, Geologic Sediments, Microscopy, Electron, Molecular Sequence Data, Phylogeny, RNA, Bacterial, RNA, Ribosomal, 16S, Temperature}, issn = {0099-2240}, doi = {10.1128/AEM.70.10.6023-6030.2004}, author = {Holmes, Dawn E and Nicoll, Julie S and Bond, Daniel R and Lovley, Derek R} } @article {556, title = {Preferential reduction of FeIII over fumarate by Geobacter sulfurreducens.}, journal = {J Bacteriol}, volume = {186}, year = {2004}, month = {2004 May}, pages = {2897-9}, abstract = {The presence of Fe(III), but not that of Fe(II), resulted in ca. 20-fold-lower levels of mRNA for fumarate reductase, inhibiting fumarate reduction and favoring utilization of fumarate as an electron donor in chemostat cultures of Geobacter sulfurreducens, despite the fact that growth yield with fumarate was 3-fold higher than with Fe(III).}, keywords = {Electrons, Fumarates, Geobacter, Iron, Oxidation-Reduction, Succinate Dehydrogenase}, issn = {0021-9193}, author = {Esteve-N{\'u}{\~n}ez, Abraham and N{\'u}{\~n}ez, Cinthia and Lovley, Derek R} } @article {398, title = {Proteolytic degradation of Escherichia coli transcription activators SoxS and MarA as the mechanism for reversing the induction of the superoxide (SoxRS) and multiple antibiotic resistance (Mar) regulons.}, journal = {Mol Microbiol}, volume = {51}, year = {2004}, month = {2004 Mar}, pages = {1801-16}, abstract = {In Escherichia coli, the SoxRS regulon confers resistance to redox-cycling compounds, and the Mar regulon provides a defence against multiple antibiotics. The response regulators, SoxS and MarA, are synthesized de novo in response to their inducing signals and directly activate transcription of a common set of target genes. Although the mechanisms of transcription activation by SoxS and MarA have been well studied, little is known about how the systems are shut-off once the inducing stress has subsided, except that de novo synthesis of the regulators is known to cease almost immediately. Here, we induced the SoxRS regulon and determined that, upon removal of the inducer, expression of the regulon{\textquoteright}s genes quickly returns to the preinduced level. This rapid shut-off indicates that the system is reset by an active process. We found that SoxS is unstable and infer that SoxS degradation is responsible for the rapid return of the system to the ground state upon removal of the inducing signal. We also found that MarA is unstable and that the instability of both proteins is intrinsic and unregulated. We used null mutations of protease genes to identify the proteases involved in the degradation of SoxS and MarA. Among single protease mutations, only lon mutations increased the half-life of SoxS and MarA. In addition, SoxS appeared to be nearly completely stable in a lon ftsH double mutant. Using hexahistidine tags placed at the respective ends of the activators, we found that access to the amino-terminus is essential for the proteolytic degradation.}, keywords = {Base Sequence, Blotting, Western, DNA Primers, DNA-Binding Proteins, Drug Resistance, Bacterial, Drug Resistance, Multiple, Bacterial, Endopeptidases, Escherichia coli, Escherichia coli Proteins, Genes, Bacterial, Half-Life, Hydrolysis, Kinetics, Mutation, Oxidation-Reduction, Promoter Regions, Genetic, Regulon, Trans-Activators, Transcription, Genetic}, issn = {0950-382X}, author = {Griffith, Kevin L and Shah, Ishita M and Wolf, Richard E} } @article {774, title = {Quorum sensing in Staphylococci is regulated via phosphorylation of three conserved histidine residues.}, journal = {J Biol Chem}, volume = {279}, year = {2004}, month = {2004 Apr 9}, pages = {14665-72}, abstract = {Staphylococcus aureus cause infections by producing toxins, a process regulated by cell-cell communication (quorum sensing) through the histidine-phosphorylation of the target of RNAIII-activating protein (TRAP). We show here that TRAP is highly conserved in staphylococci and contains three completely conserved histidine residues (His-66, His-79, His-154) that are phosphorylated and essential for its activity. This was tested by constructing a TRAP(-) strain with each of the conserved histidine residues changed to alanine by site-directed mutagenesis. All mutants were tested for pathogenesis in vitro (expression of RNAIII and hemolytic activity) and in vivo (murine cellulitis model). Results show that RNAIII is not expressed in the TRAP(-) strain, that it is non hemolytic, and that it does not cause disease in vivo. These pathogenic phenotypes could be rescued in the strain containing the recovered traP, confirming the importance of TRAP in S. aureus pathogenesis. The phosphorylation of TRAP mutated in any of the conserved histidine residues was significantly reduced, and mutants defective in any one of these residues were non-pathogenic in vitro or in vivo, whereas those mutated in a non-conserved histidine residue (His-124) were as pathogenic as the wild type. These results confirm the importance of the three conserved histidine residues in TRAP activity. The phosphorylation pattern, structure, and gene organization of TRAP deviates from signaling molecules known to date, suggesting that TRAP belongs to a novel class of signal transducers.}, keywords = {Amino Acid Sequence, Animals, Blotting, Northern, Cell Communication, DNA, Electrophoresis, Polyacrylamide Gel, Escherichia coli, Histidine, Mice, Molecular Sequence Data, Mutagenesis, Mutagenesis, Site-Directed, Mutation, Phenotype, Phosphorylation, Phylogeny, Plasmids, Protein Structure, Tertiary, RNA, Antisense, RNA, Bacterial, Sequence Homology, Amino Acid, Signal Transduction, Staphylococcus aureus, Time Factors}, issn = {0021-9258}, doi = {10.1074/jbc.M311106200}, author = {Gov, Yael and Borovok, Ilya and Korem, Moshe and Singh, Vineet K and Jayaswal, Radheshyam K and Wilkinson, Brian J and Rich, Stephen M and Balaban, Naomi} } @article {542, title = {Resistance of solid-phase U(VI) to microbial reduction during in situ bioremediation of uranium-contaminated groundwater.}, journal = {Appl Environ Microbiol}, volume = {70}, year = {2004}, month = {2004 Dec}, pages = {7558-60}, abstract = {Speciation of solid-phase uranium in uranium-contaminated subsurface sediments undergoing uranium bioremediation demonstrated that although microbial reduction of soluble U(VI) readily immobilized uranium as U(IV), a substantial portion of the U(VI) in the aquifer was strongly associated with the sediments and was not microbially reducible. These results have important implications for in situ uranium bioremediation strategies.}, keywords = {Acetates, Biodegradation, Environmental, Deltaproteobacteria, Fresh Water, Geologic Sediments, Oxidation-Reduction, Solubility, Uranium, Water Pollutants, Radioactive}, issn = {0099-2240}, doi = {10.1128/AEM.70.12.7558-7560.2004}, author = {Ortiz-Bernad, Irene and Anderson, Robert T and Vrionis, Helen A and Lovley, Derek R} } @article {549, title = {The RpoS sigma factor in the dissimilatory Fe(III)-reducing bacterium Geobacter sulfurreducens.}, journal = {J Bacteriol}, volume = {186}, year = {2004}, month = {2004 Aug}, pages = {5543-6}, abstract = {Geobacter sulfurreducens RpoS sigma factor was shown to contribute to survival in stationary phase and upon oxygen exposure. Furthermore, a mutation in rpoS decreased the rate of reduction of insoluble Fe(III) but not of soluble forms of iron. This study suggests that RpoS plays a role in regulating metabolism of Geobacter under suboptimal conditions in subsurface environments.}, keywords = {Adaptation, Physiological, Bacterial Proteins, Base Sequence, Ferric Compounds, Gene Expression Regulation, Bacterial, Genes, Bacterial, Geobacter, Molecular Sequence Data, Mutation, Oxidation-Reduction, Oxidative Stress, Sigma Factor, Transcription Initiation Site}, issn = {0021-9193}, doi = {10.1128/JB.186.16.5543-5546.2004}, author = {N{\'u}{\~n}ez, Cinthia and Adams, Lorrie and Childers, Susan and Lovley, Derek R} } @article {541, title = {The structure of the core region of the lipopolysaccharide from Geobacter sulfurreducens.}, journal = {Carbohydr Res}, volume = {339}, year = {2004}, month = {2004 Dec 27}, pages = {2901-4}, abstract = {The structure of the core part of the LPS from Geobacter sulfurreducens was analysed. The LPS contained no O-specific polysaccharide (O-side chain) and upon mild hydrolysis gave a core oligosaccharide, which was isolated by gel chromatography. It was studied by chemical methods, NMR and mass spectrometry, and the following structure was proposed. [carbohydrate structure: see text] where Q = 3-O-Me-alpha-L-QuiNAc-(1-->or H (approximately 3:2).}, keywords = {Carbohydrate Sequence, Geobacter, Lipopolysaccharides, Magnetic Resonance Spectroscopy, Molecular Sequence Data, Oligosaccharides}, issn = {0008-6215}, doi = {10.1016/j.carres.2004.10.004}, author = {Vinogradov, Evgeny and Korenevsky, Anton and Lovley, Derek R and Beveridge, Terry J} } @article {752, title = {Tuning the hemolytic and antibacterial activities of amphiphilic polynorbornene derivatives.}, journal = {J Am Chem Soc}, volume = {126}, year = {2004}, month = {2004 Dec 8}, pages = {15870-5}, abstract = {Amphiphilic cationic polynorbornene derivatives, soluble in water, were prepared from modular norbornene monomers, with a wide range of molecular weights (M(n) = 1600-137 500 g/mol) and narrow polydispersities (PDI = 1.1-1.3). The antibacterial activity determined by growth inhibition assays and the hemolytic activity against human red blood cells were measured and compared to determine the selectivity of the polymers for bacterial over mammalian cells. The effects of monomer repeat unit hydrophobicity and polymer molecular weight on antibacterial and hemolytic activities were determined. The hydrophobicity of the repeat unit was observed to have dramatic effects on antibacterial and hemolytic activities. Lipid membrane disruption activities of the polymers was confirmed by measuring polymer-induced dye leakage from large unilamellar vesicles. By tuning the overall hydrophobicity of the polymer through random copolymerizations of modular norbornene derivatives, highly selective, nonhemolytic antibacterial activities were obtained. For appropriate monomer composition, selectivity against bacteria versus human red blood cells was determined to be over 100.}, keywords = {Anti-Bacterial Agents, Bacillus subtilis, Cell Membrane, Cholesterol, Erythrocytes, Escherichia coli, Hemolysis, Humans, Liposomes, Microbial Sensitivity Tests, Molecular Weight, Norbornanes, Polymers, Structure-Activity Relationship}, issn = {0002-7863}, doi = {10.1021/ja045664d}, author = {Ilker, M Firat and N{\"u}sslein, Klaus and Tew, Gregory N and Coughlin, E Bryan} } @article {773, title = {The unpredictable past of Plasmodium vivax revealed in its genome.}, journal = {Proc Natl Acad Sci U S A}, volume = {101}, year = {2004}, month = {2004 Nov 2}, pages = {15547-8}, keywords = {Animals, Biological Evolution, Culicidae, Genetics, Population, Genome, Protozoan, Genomics, Haplorhini, Humans, Malaria, Vivax, Plasmodium vivax}, issn = {0027-8424}, doi = {10.1073/pnas.0405618101}, author = {Rich, Stephen M} } @article {553, title = {Vanadium respiration by Geobacter metallireducens: novel strategy for in situ removal of vanadium from groundwater.}, journal = {Appl Environ Microbiol}, volume = {70}, year = {2004}, month = {2004 May}, pages = {3091-5}, abstract = {Vanadium can be an important contaminant in groundwaters impacted by mining activities. In order to determine if microorganisms of the Geobacteraceae, the predominant dissimilatory metal reducers in many subsurface environments, were capable of reducing vanadium(V), Geobacter metallireducens was inoculated into a medium in which acetate was the electron donor and vanadium(V) was the sole electron acceptor. Reduction of vanadium(V) resulted in the production of vanadium(IV), which subsequently precipitated. Reduction of vanadium(V) was associated with cell growth with a generation time of 15 h. No vanadium(V) was reduced and no precipitate was formed in heat-killed or abiotic controls. Acetate was the most effective of all the electron donors evaluated. When acetate was injected into the subsurface to enhance the growth and activity of Geobacteraceae in an aquifer contaminated with uranium and vanadium, vanadium was removed from the groundwater even more effectively than uranium. These studies demonstrate that G. metallireducens can grow via vanadium(V) respiration and that stimulating the activity of Geobacteraceae, and hence vanadium(V) reduction, can be an effective strategy for in situ immobilization of vanadium in contaminated subsurface environments.}, keywords = {Anaerobiosis, Biodegradation, Environmental, Culture Media, Fresh Water, Geobacter, Geologic Sediments, Mining, Oxidation-Reduction, Vanadium, Water Pollution, Chemical}, issn = {0099-2240}, author = {Ortiz-Bernad, Irene and Anderson, Robert T and Vrionis, Helen A and Lovley, Derek R} } @article {576, title = {Biochemical and genetic characterization of PpcA, a periplasmic c-type cytochrome in Geobacter sulfurreducens.}, journal = {Biochem J}, volume = {369}, year = {2003}, month = {2003 Jan 1}, pages = {153-61}, abstract = {A 9.6 kDa periplasmic c -type cytochrome, designated PpcA, was purified from the Fe(III)-reducing bacterium Geobacter sulfurreducens and characterized. The purified protein is basic (pI 9.5), contains three haems and has an N-terminal amino acid sequence closely related to those of the previously described trihaem c (7) cytochromes of Geobacter metallireducens and Desulfuromonas acetoxidans. The gene encoding PpcA was identified from the G. sulfurreducens genome using the N-terminal sequence, and encodes a protein of 71 amino acids (molecular mass 9.58 kDa) with 49\% identity to the c (7) cytochrome of D. acetoxidans. In order to determine the physiological role of PpcA, a knockout mutant was prepared with a single-step recombination method. Acetate-dependent Fe(III) reduction was significantly inhibited in both growing cultures and cell suspensions of the mutant. When ppcA was expressed in trans, the full capacity for Fe(III) reduction with acetate was restored. The transfer of electrons from acetate to anthraquinone 2,6-disulphonate (AQDS; a humic acid analogue) and to U(VI) was also compromised in the mutant, but acetate-dependent reduction of fumarate was not altered. The rates of reduction of Fe(III), AQDS, U(VI) and fumarate were also the same in the wild type and ppcA mutant when hydrogen was supplied as the electron donor. When taken together with previous studies on other electron transport proteins in G. sulfurreducens, these results suggest that PpcA serves as an intermediary electron carrier from acetate to terminal Fe(III) reductases in the outer membrane, and is also involved in the transfer of electrons from acetate to U(VI) and humics.}, keywords = {Amino Acid Sequence, Base Sequence, Cytochrome c Group, DNA, Bacterial, Molecular Sequence Data, Periplasm, Proteobacteria, Sequence Homology, Amino Acid}, issn = {0264-6021}, doi = {10.1042/BJ20020597}, author = {Lloyd, Jon R and Leang, Ching and Hodges Myerson, Allison L and Coppi, Maddalena V and Cuifo, Stacey and Methe, Barb and Sandler, Steven J and Lovley, Derek R} } @article {561, title = {Biotechnological application of metal-reducing microorganisms.}, journal = {Adv Appl Microbiol}, volume = {53}, year = {2003}, month = {2003}, pages = {85-128}, keywords = {Archaea, Biodegradation, Environmental, Bioreactors, Environmental Microbiology, Geobacter, Gram-Negative Anaerobic Bacteria, Metals, Heavy, Water Pollutants, Chemical}, issn = {0065-2164}, author = {Lloyd, Jonathan R and Lovley, Derek R and Macaskie, Lynne E} } @article {558, title = {Cleaning up with genomics: applying molecular biology to bioremediation.}, journal = {Nat Rev Microbiol}, volume = {1}, year = {2003}, month = {2003 Oct}, pages = {35-44}, abstract = {Bioremediation has the potential to restore contaminated environments inexpensively yet effectively, but a lack of information about the factors controlling the growth and metabolism of microorganisms in polluted environments often limits its implementation. However, rapid advances in the understanding of bioremediation are on the horizon. Researchers now have the ability to culture microorganisms that are important in bioremediation and can evaluate their physiology using a combination of genome-enabled experimental and modelling techniques. In addition, new environmental genomic techniques offer the possibility for similar studies on as-yet-uncultured organisms. Combining models that can predict the activity of microorganisms that are involved in bioremediation with existing geochemical and hydrological models should transform bioremediation from a largely empirical practice into a science.}, keywords = {Bacteria, Bacterial Physiological Phenomena, Biodegradation, Environmental, Ecology, Environmental Microbiology, Fungi, Genetic Vectors, Genome, Bacterial, Genome, Fungal, Genomics, Models, Biological, Molecular Biology}, issn = {1740-1526}, doi = {10.1038/nrmicro731}, author = {Lovley, Derek R} } @article {684, title = {ConSurf: identification of functional regions in proteins by surface-mapping of phylogenetic information.}, journal = {Bioinformatics}, volume = {19}, year = {2003}, month = {2003 Jan}, pages = {163-4}, abstract = {We recently developed algorithmic tools for the identification of functionally important regions in proteins of known three dimensional structure by estimating the degree of conservation of the amino-acid sites among their close sequence homologues. Projecting the conservation grades onto the molecular surface of these proteins reveals patches of highly conserved (or occasionally highly variable) residues that are often of important biological function. We present a new web server, ConSurf, which automates these algorithmic tools. ConSurf may be used for high-throughput characterization of functional regions in proteins. AVAILABILITY: The ConSurf web server is available at:http://consurf.tau.ac.il. SUPPLEMENTARY INFORMATION: A set of examples is available at http://consurf.tau.ac.il under {\textquoteright}GALLERY{\textquoteright}.}, keywords = {Amino Acids, bcl-X Protein, Conserved Sequence, Databases, Protein, Evolution, Molecular, Internet, Phylogeny, Protein Conformation, Proteins, Proto-Oncogene Proteins c-bcl-2, Sequence Alignment, Sequence Analysis, Protein, User-Computer Interface}, issn = {1367-4803}, author = {Glaser, Fabian and Pupko, Tal and Paz, Inbal and Bell, Rachel E and Bechor-Shental, Dalit and Martz, Eric and Ben-Tal, Nir} } @article {776, title = {Cryptosporidium parvum in children with diarrhea in Mulago Hospital, Kampala, Uganda.}, journal = {Am J Trop Med Hyg}, volume = {68}, year = {2003}, month = {2003 Jun}, pages = {710-5}, abstract = {A cross-sectional case-control study (ratio = 3:1) was conducted over a 15-month period to determine the prevalence and consequences of cryptosporidiosis in hospitalized diarrheic children (0-5 years old) at Mulago Hospital in Kampala, Uganda. Cryptosporidium parvum was detected and genotyped among 2,446 children of whom 1,779 (72.7\%) had diarrhea, and 667 (27.3\%) were age- and sex-matched controls. Of the 1,779 children with diarrhea, 532 (29.9\%) had persistent (> 14 days) diarrhea and 1,247 (70.1\%) had acute diarrhea. Overall, 444 (25.0\%) of the 1,779 children with diarrhea had C. parvum, compared with only 57 (8.5\%) of the 667 children without diarrhea (chi2 = 80.2, P < or = 0.0001). Within this group of infected children, 72.8\% were infected with genotype 1, 18.4\% with genotype 2, and 4.1\% with a mixture of both genotypes, and 4.1\% isolates were either unclassified or C. meleagridis. The prevalence was highest during the rainy months of April to June. Of the 532 children with persistent diarrhea, 166 (31.2\%) had C. parvum compared with 278 (22.3\%) of the 1,247 children with acute diarrhea (chi2 = 15.8, P < or = 0.0001). There was a significant association between C. parvum and malnutrition including stunting, being underweight, and wasting. Unfavorable outcome (death or failure to resolve within 14 days) occurred in 139 (72.8\%) of the 191 children with C. parvum, and in only 65.1\% of the 545 without (odds ratio = 1.117, 95\% confidence interval = 1.005-1.243, P = 0.05), Of the 191 children with C. parvum, 24 (12.6\%) died, compared with 34 (6.2\%) of the 545 without C. parvum (P = 0.005). Mortality rates were higher among children with severe dehydration and persistent diarrhea, and in stunted or underweight children infected with C. parvum. Among Ugandan children, cryptosporidiosis, which remains untreatable, is frequently associated with diarrhea and other serious and unfavorable consequences.}, keywords = {Animals, Case-Control Studies, Child, Preschool, Cross-Sectional Studies, Cryptosporidiosis, Cryptosporidium parvum, Diarrhea, Diarrhea, Infantile, DNA, Protozoan, Female, Hospitalization, Humans, Infant, Infant, Newborn, Male, Nutrition Disorders, Polymerase Chain Reaction, Polymorphism, Restriction Fragment Length, Prevalence, Uganda}, issn = {0002-9637}, author = {Tumwine, James K and Kekitiinwa, Addy and Nabukeera, Nicolette and Akiyoshi, Donna E and Rich, Stephen M and Widmer, Giovanni and Feng, Xiaochuan and Tzipori, Saul} } @article {560, title = {Diversity of Geobacteraceae species inhabiting metal-polluted freshwater lake sediments ascertained by 16S rDNA analyses.}, journal = {Microb Ecol}, volume = {46}, year = {2003}, month = {2003 Aug}, pages = {257-69}, abstract = {The abundance, distribution, and phylogenetic diversity of members of the Fe(III)-reducing family Geobacteraceae were studied along a gradient of metal contaminants in Lake Coeur d{\textquoteright}Alene, Idaho. Partial 16S rRNA gene fragments were amplified by PCR using primers directed toward conserved regions of the gene within the family Geobacteraceae. Analysis of amplicons separated by denaturing gradient gel electrophoresis (DGGE) suggested within-site variation was as great as between-site variation. Amplicons were cloned and grouped by RFLP type and DGGE migration distance and representatives were sequenced. Grouping clones with 3\% or less sequence dissimilarity, 15 distinct phylotypes were identified compared to 16 distinct DGGE bands. Only 1 phylotype was recovered from all sites. This clone, B14, is most closely related to Geobacter metallireducens and constituted a greater portion of the pristine community than of the contaminated communities. A second phylotype, Q2, predominated in the contaminated communities and was notably absent from the pristine libraries. Clone Q2 presents a high degree of sequence similarity to two Geobacter spp. previously isolated from this region of Lake Coeur d{\textquoteright}Alene. Six phylotypes were unique to the contaminated sediments, whereas two were found only in the pristine sediments. Indices of diversity (Shannon and Simpson) were consistently higher when calculated with DGGE data than when clone library data were used. Most-probable-number PCR and real-time PCR suggested that the Geobacteraceae phylotypes were spread relatively evenly across all three sites along the gradient. Our data indicate that the Geobacteraceae are diverse and abundant in Lake Coeur d{\textquoteright}Alene sediments, regardless of metals content. These results provide insight into the ability of dissimilatory Fe(III)-reducing bacteria to colonize habitats with elevated metal concentrations, and they have important implications for the management and remediation of metal-contaminated sites.}, keywords = {Cloning, Molecular, Geobacter, Geologic Sediments, Phylogeny, Polymerase Chain Reaction, Population Dynamics, RNA, Ribosomal, 16S, Water Pollutants, Water Supply}, issn = {0095-3628}, author = {Cummings, D E and Snoeyenbos-West, O L and Newby, D T and Niggemyer, A M and Lovley, D R and Achenbach, L A and Rosenzweig, R F} } @article {564, title = {Electricity generation by direct oxidation of glucose in mediatorless microbial fuel cells.}, journal = {Nat Biotechnol}, volume = {21}, year = {2003}, month = {2003 Oct}, pages = {1229-32}, abstract = {Abundant energy, stored primarily in the form of carbohydrates, can be found in waste biomass from agricultural, municipal and industrial sources as well as in dedicated energy crops, such as corn and other grains. Potential strategies for deriving useful forms of energy from carbohydrates include production of ethanol and conversion to hydrogen, but these approaches face technical and economic hurdles. An alternative strategy is direct conversion of sugars to electrical power. Existing transition metal-catalyzed fuel cells cannot be used to generate electric power from carbohydrates. Alternatively, biofuel cells in which whole cells or isolated redox enzymes catalyze the oxidation of the sugar have been developed, but their applicability has been limited by several factors, including (i) the need to add electron-shuttling compounds that mediate electron transfer from the cell to the anode, (ii) incomplete oxidation of the sugars and (iii) lack of long-term stability of the fuel cells. Here we report on a novel microorganism, Rhodoferax ferrireducens, that can oxidize glucose to CO(2) and quantitatively transfer electrons to graphite electrodes without the need for an electron-shuttling mediator. Growth is supported by energy derived from the electron transfer process itself and results in stable, long-term power production.}, keywords = {Bioelectric Energy Sources, Bioreactors, Carbon Dioxide, Comamonadaceae, Electrodes, Energy Transfer, Equipment Design, Equipment Failure Analysis, Glucose, Iron, Oxidation-Reduction}, issn = {1087-0156}, doi = {10.1038/nbt867}, author = {Chaudhuri, Swades K and Lovley, Derek R} } @article {573, title = {Electricity production by Geobacter sulfurreducens attached to electrodes.}, journal = {Appl Environ Microbiol}, volume = {69}, year = {2003}, month = {2003 Mar}, pages = {1548-55}, abstract = {Previous studies have suggested that members of the Geobacteraceae can use electrodes as electron acceptors for anaerobic respiration. In order to better understand this electron transfer process for energy production, Geobacter sulfurreducens was inoculated into chambers in which a graphite electrode served as the sole electron acceptor and acetate or hydrogen was the electron donor. The electron-accepting electrodes were maintained at oxidizing potentials by connecting them to similar electrodes in oxygenated medium (fuel cells) or to potentiostats that poised electrodes at +0.2 V versus an Ag/AgCl reference electrode (poised potential). When a small inoculum of G. sulfurreducens was introduced into electrode-containing chambers, electrical current production was dependent upon oxidation of acetate to carbon dioxide and increased exponentially, indicating for the first time that electrode reduction supported the growth of this organism. When the medium was replaced with an anaerobic buffer lacking nutrients required for growth, acetate-dependent electrical current production was unaffected and cells attached to these electrodes continued to generate electrical current for weeks. This represents the first report of microbial electricity production solely by cells attached to an electrode. Electrode-attached cells completely oxidized acetate to levels below detection (<10 micro M), and hydrogen was metabolized to a threshold of 3 Pa. The rates of electron transfer to electrodes (0.21 to 1.2 micro mol of electrons/mg of protein/min) were similar to those observed for respiration with Fe(III) citrate as the electron acceptor (E(o){\textquoteright} =+0.37 V). The production of current in microbial fuel cell (65 mA/m(2) of electrode surface) or poised-potential (163 to 1,143 mA/m(2)) mode was greater than what has been reported for other microbial systems, even those that employed higher cell densities and electron-shuttling compounds. Since acetate was completely oxidized, the efficiency of conversion of organic electron donor to electricity was significantly higher than in previously described microbial fuel cells. These results suggest that the effectiveness of microbial fuel cells can be increased with organisms such as G. sulfurreducens that can attach to electrodes and remain viable for long periods of time while completely oxidizing organic substrates with quantitative transfer of electrons to an electrode.}, keywords = {Culture Media, Deltaproteobacteria, Electricity, Electrodes, Electron Transport, Microscopy, Electron, Scanning, Oxidation-Reduction}, issn = {0099-2240}, author = {Bond, Daniel R and Lovley, Derek R} } @article {42, title = {Expression of the yeast PIS1 gene requires multiple regulatory elements including a Rox1p binding site.}, journal = {J Biol Chem}, volume = {278}, year = {2003}, month = {2003 Oct 3}, pages = {38646-52}, abstract = {The PIS1 gene is required for de novo synthesis of phosphatidylinositol (PI), an essential phospholipid in Saccharomyces cerevisiae. PIS1 gene expression is unusual because it is uncoupled from the other phospholipid biosynthetic genes, which are regulated in response to inositol and choline. Relatively little is known about regulation of transcription of the PIS1 gene. We reported previously that PIS1 transcription is sensitive to carbon source. To further our understanding of the regulation of PIS1 transcription, we carried out a promoter deletion analysis that identified three regions required for PIS1 gene expression (upstream activating sequence (UAS) elements 1-3). Deletion of either UAS1 or UAS2 resulted in an approximately 45\% reduction in expression, whereas removal of UAS3 yielded an 84\% decrease in expression. A comparison of promoters among several Saccharomyces species shows that these sequences are highly conserved. Curiously, the UAS3 element region (-149 to -138) includes a Rox1p binding site. Rox1p is a repressor of hypoxic genes under aerobic growth conditions. Consistent with this, we have found that expression of a PIS1-cat reporter was repressed under aerobic conditions, and this repression was dependent on both Rox1p and its binding site. Furthermore, PI levels were elevated under anaerobic conditions. This is the first evidence that PI levels are affected by regulation of PIS1 transcription.}, keywords = {Anoxia, Base Sequence, Binding Sites, Carbon, Chloramphenicol O-Acetyltransferase, Choline, Chromatography, Thin Layer, Conserved Sequence, DNA, DNA, Complementary, DNA-Binding Proteins, Gene Deletion, Gene Expression Regulation, Fungal, Genes, Reporter, Inositol, Lipid Metabolism, Models, Biological, Models, Genetic, Molecular Sequence Data, Oxygen, Phospholipids, Plasmids, Promoter Regions, Genetic, Protein Binding, Repressor Proteins, RNA, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins, Transcription, Genetic, Transferases (Other Substituted Phosphate Groups)}, issn = {0021-9258}, doi = {10.1074/jbc.M305251200}, author = {Gardocki, Mary Elizabeth and Lopes, John M} } @article {565, title = {Extending the upper temperature limit for life.}, journal = {Science}, volume = {301}, year = {2003}, month = {2003 Aug 15}, pages = {934}, keywords = {Archaea, Culture Media, Ferric Compounds, Hot Temperature, Oxidation-Reduction, Pacific Ocean, Water Microbiology}, issn = {1095-9203}, doi = {10.1126/science.1086823}, author = {Kashefi, Kazem and Lovley, Derek R} } @article {562, title = {Genome of Geobacter sulfurreducens: metal reduction in subsurface environments.}, journal = {Science}, volume = {302}, year = {2003}, month = {2003 Dec 12}, pages = {1967-9}, abstract = {The complete genome sequence of Geobacter sulfurreducens, a delta-proteobacterium, reveals unsuspected capabilities, including evidence of aerobic metabolism, one-carbon and complex carbon metabolism, motility, and chemotactic behavior. These characteristics, coupled with the possession of many two-component sensors and many c-type cytochromes, reveal an ability to create alternative, redundant, electron transport networks and offer insights into the process of metal ion reduction in subsurface environments. As well as playing roles in the global cycling of metals and carbon, this organism clearly has the potential for use in bioremediation of radioactive metals and in the generation of electricity.}, keywords = {Acetates, Acetyl Coenzyme A, Aerobiosis, Anaerobiosis, Bacterial Proteins, Carbon, Chemotaxis, Chromosomes, Bacterial, Cytochromes c, Electron Transport, Energy Metabolism, Genes, Bacterial, Genes, Regulator, Genome, Bacterial, Geobacter, Hydrogen, Metals, Movement, Open Reading Frames, Oxidation-Reduction, Phylogeny}, issn = {1095-9203}, doi = {10.1126/science.1088727}, author = {Meth{\'e}, B A and Nelson, K E and Eisen, J A and Paulsen, I T and Nelson, W and Heidelberg, J F and Wu, D and Wu, M and Ward, N and Beanan, M J and Dodson, R J and Madupu, R and Brinkac, L M and Daugherty, S C and DeBoy, R T and Durkin, A S and Gwinn, M and Kolonay, J F and Sullivan, S A and Haft, D H and Selengut, J and Davidsen, T M and Zafar, N and White, O and Tran, B and Romero, C and Forberger, H A and Weidman, J and Khouri, H and Feldblyum, T V and Utterback, T R and Van Aken, S E and Lovley, D R and Fraser, C M} } @article {570, title = {Geobacter sulfurreducens has two autoregulated lexA genes whose products do not bind the recA promoter: differing responses of lexA and recA to DNA damage.}, journal = {J Bacteriol}, volume = {185}, year = {2003}, month = {2003 Apr}, pages = {2493-502}, abstract = {The Escherichia coli LexA protein was used as a query sequence in TBLASTN searches to identify the lexA gene of the delta-proteobacterium Geobacter sulfurreducens from its genome sequence. The results of the search indicated that G. sulfurreducens has two independent lexA genes designated lexA1 and lexA2. A copy of a dinB gene homologue, which in E. coli encodes DNA polymerase IV, is present downstream of each lexA gene. Reverse transcription-PCR analyses demonstrated that, in both cases, lexA and dinB constitute a single transcriptional unit. Electrophoretic mobility shift assays with purified LexA1 and LexA2 proteins have shown that both proteins bind the imperfect palindrome GGTTN(2)CN(4)GN(3)ACC found in the promoter region of both lexA1 and lexA2. This sequence is also present upstream of the Geobacter metallireducens lexA gene, indicating that it is the LexA box of this bacterial genus. This palindrome is not found upstream of either the G. sulfurreducens or the G. metallireducens recA genes. Furthermore, DNA damage induces expression of the lexA-dinB transcriptional unit but not that of the recA gene. However, the basal level of recA gene expression is dramatically higher than that of the lexA gene. Likewise, the promoters of the G. sulfurreducens recN, ruvAB, ssb, umuDC, uvrA, and uvrB genes do not contain the LexA box and are not likely to bind to the LexA1 or LexA2 proteins. G. sulfurreducens is the first bacterial species harboring a lexA gene for which a constitutive expression of its recA gene has been described.}, keywords = {Amino Acid Sequence, Bacterial Proteins, Consensus Sequence, DNA Damage, DNA Polymerase beta, DNA Repair, DNA-Binding Proteins, Electrophoretic Mobility Shift Assay, Gene Expression, Genes, Bacterial, Molecular Sequence Data, Promoter Regions, Genetic, Proteobacteria, Rec A Recombinases, Sequence Alignment, Serine Endopeptidases, Transcription, Genetic}, issn = {0021-9193}, author = {Jara, M{\'o}nica and N{\'u}{\~n}ez, Cinthia and Campoy, Susana and Fern{\'a}ndez de Henestrosa, Antonio R and Lovley, Derek R and Barb{\'e}, Jordi} } @article {1215, title = {Identification of 2D-gel proteins: a comparison of MALDI/TOF peptide mass mapping to mu LC-ESI tandem mass spectrometry.}, journal = {J Am Soc Mass Spectrom}, volume = {14}, year = {2003}, month = {2003 Sep}, pages = {957-70}, abstract = {A comparative analysis of protein identification for a total of 162 protein spots separated by two-dimensional gel electrophoresis from two fully sequenced archaea, Methanococcus jannaschii and Pyrococcus furiosus, using MALDI-TOF peptide mass mapping (PMM) and mu LC-MS/MS is presented. 100\% of the gel spots analyzed were successfully matched to the predicted proteins in the two corresponding open reading frame databases by mu LC-MS/MS while 97\% of them were identified by MALDI-TOF PMM. The high success rate from the PMM resulted from sample desalting/concentrating with ZipTip(C18) and optimization of several PMM search parameters including a 25 ppm average mass tolerance and the application of two different protein molecular weight search windows. By using this strategy, low-molecular weight (<23 kDa) proteins could be identified unambiguously with less than 5 peptide matches. Nine percent of spots were identified as containing multiple proteins. By using mu LC-MS/MS, 50\% of the spots analyzed were identified as containing multiple proteins. mu LC-MS/MS demonstrated better protein sequence coverage than MALDI-TOF PMM over the entire mass range of proteins identified. MALDI-TOF and PMM produced unique peptide molecular weight matches that were not identified by mu LC-MS/MS. By incorporating amino acid sequence modifications into database searches, combined sequence coverage obtained from these two complimentary ionization methods exceeded 50\% for approximately 70\% of the 162 spots analyzed. This improved sequence coverage in combination with enzymatic digestions of different specificity is proposed as a method for analysis of post-translational modification from 2D-gel separated proteins.
}, keywords = {Amino Acid Sequence, Chromatography, Liquid, Databases, Protein, Electrophoresis, Gel, Two-Dimensional, Methanococcus, Molecular Sequence Data, Molecular Weight, Peptide Mapping, Proteins, Pyrococcus furiosus, Sensitivity and Specificity, Software, Spectrometry, Mass, Electrospray Ionization, Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization, Trypsin}, issn = {1044-0305}, author = {Lim, Hanjo and Eng, Jimmy and Yates, John R and Tollaksen, Sandra L and Giometti, Carol S and Holden, James F and Adams, Michael W W and Reich, Claudia I and Olsen, Gary J and Hays, Lara G} } @article {715, title = {Lipid rafts, caveolae, caveolin-1, and entry by Chlamydiae into host cells.}, journal = {Exp Cell Res}, volume = {287}, year = {2003}, month = {2003 Jul 1}, pages = {67-78}, abstract = {Obligate intracellular bacterial pathogens of the genus Chlamydia are reported to enter host cells by both clathrin-dependent and clathrin-independent processes. C. trachomatis serovar K recently was shown to enter cells via caveolae-like lipid raft domains. We asked here how widespread raft-mediated entry might be among the Chlamydia. We show that C. pneumoniae, an important cause of respiratory infections in humans that additionally is associated with cardiovascular disease, and C. psittaci, an important pathogen in domestic mammals and birds that also infects humans, each enter host cells via cholesterol-rich lipid raft microdomains. Further, we show that C. trachomatis serovars E and F also use these domains to enter host cells. The involvement of these membrane domains in the entry of these organisms was indicated by the sensitivity of their entry to the raft-disrupting agents Nystatin and filipin, and by their intracellular association with caveolin-1, a 22-kDa protein associated with the formation of caveolae in rafts. In contrast, caveolin-marked lipid raft domains do not mediate entry of C. trachomatis serovars A, 36B, and C, nor of LGV serovar L2 and MoPn. Finally, we show that entry of each of these chlamydial strains is independent of cellular expression of caveolin-1. Thus, entry via the Nystatin and filipin-sensitive pathway is dependent on lipid rafts containing cholesterol, rather than invaginated caveolae per se.}, keywords = {Anti-Bacterial Agents, Caveolae, Caveolin 1, Caveolins, Chlamydia, Chlamydia Infections, Cholesterol, Dose-Response Relationship, Drug, Eukaryotic Cells, Filipin, HeLa Cells, Host-Parasite Interactions, Humans, Membrane Microdomains, Nystatin, Species Specificity}, issn = {0014-4827}, author = {Stuart, Elizabeth S and Webley, Wilmore C and Norkin, Leonard C} } @article {862, title = {Mechanism of the polymerization reaction initiated and catalyzed by the polyhydroxybutyrate synthase of Ralstonia eutropha.}, journal = {Biomacromolecules}, volume = {4}, year = {2003}, month = {2003 May-Jun}, pages = {504-9}, abstract = {Polyhydroxybutyrate (PHB) synthases (polymerases) catalyze the polymerization of the coenzyme A thioester of 3-hydroxybutyrate to PHB. The Ralstonia eutropha PHB synthase purified from recombinant E. coli cells exists in aqueous solution in both monomeric (single subunit) and homodimeric (two subunits) forms in equilibrium. Several lines of evidence suggest that the homodimer is the active form of the synthase. The initial mechanistic model for the polymerization reaction proposed that two different thiol groups form the catalytic site. The cysteine at 319 has been shown to provide one thiol group that is involved in the covalent catalysis, but a second thiol group on the same protein molecule has not yet been identified. It is suggested that cysteines at 319 from each of the two molecules of a homodimer synthase provide two identical thiol groups to jointly form a single catalytic site. To verify this model using the strategy of in vitro reconstitution, heterodimers composed of the wild-type subunit and of the C(319) mutated subunit were constructed and the activities at various ratios of the wild-type subunit to the mutated subunit were measured. The experimental results indicate that the homodimer is the active form of the enzyme, that the heterodimer containing the mutated subunit has no activity, and that a single cysteine is not sufficient for catalysis. Two identical thiol groups from C(319) residues on each subunit of the homodimer are required to form the catalytic site for the initiation and propagation reactions. We further demonstrate that a dimer synthase that has initiated the polymerization reaction (primed synthase) is significantly more stable against dissociation than the unprimed (unreacted) dimer synthase. These two properties explain the nature of lag phenomenon during the in vitro polymerization reaction catalyzed by this enzyme}, keywords = {Catalysis, Cupriavidus necator, Hydroxybutyrates, Polymers}, issn = {1525-7797}, doi = {10.1021/bm020099x}, author = {Zhang, Shiming and Kolvek, Steve and Lenz, Robert W and Goodwin, Steve} } @article {566, title = {Metabolism of organic compounds in anaerobic, hydrothermal sulphate-reducing marine sediments.}, journal = {Environ Microbiol}, volume = {5}, year = {2003}, month = {2003 Jul}, pages = {583-91}, abstract = {Previous studies of hot (>80 degrees C) microbial ecosystems have primarily relied on the study of pure cultures or analysis of 16S rDNA sequences. In order to gain more information on anaerobic metabolism by natural communities in hot environments, sediments were collected from a shallow marine hydrothermal vent system in Baia di Levante, Vulcano, Italy and incubated under strict anaerobic conditions at 90 degrees C. Sulphate reduction was the predominant terminal electron-accepting process in the sediments. The addition of molybdate inhibited sulphate reduction in the sediments and resulted in a linear accumulation of acetate and hydrogen over time. [U-14C]- acetate was completely oxidized to 14CO2, and the addition of molybdate inhibited 14CO2 production by 60\%. [U-14C]-glucose was oxidized to 14CO2, and this was inhibited when molybdate was added. When the pool sizes of short-chain fatty acids were artificially increased, radiolabel from [U-14C]-glucose accumulated in the acetate pool. L-[U-14C]-glutamate, [ring-14C]-benzoate and [U-14C]-palmitate were also anaerobically oxidized to 14CO2 in the sediments, but molybdate had little effect on the oxidation of these compounds. These results demonstrate that natural microbial communities living in a hot, microbial ecosystem can oxidize acetate and a range of other organic electron donors under sulphate-reducing conditions and suggest that acetate is an important extracellular intermediate in the anaerobic degradation of organic matter in hot microbial ecosystems.}, keywords = {Acetates, Anaerobiosis, Bacteria, Anaerobic, Benzoates, Carbon Dioxide, Ecosystem, Geologic Sediments, Glucose, Glutamic Acid, Hot Temperature, Hydrogen, Italy, Molybdenum, Organic Chemicals, Oxidation-Reduction, Palmitates, Seawater, Sulfates, Water Microbiology}, issn = {1462-2912}, author = {Tor, Jason M and Amend, Jan P and Lovley, Derek R} } @article {1216, title = {Microbe-metal interactions in marine hydrothermal environments.}, journal = {Curr Opin Chem Biol}, volume = {7}, year = {2003}, month = {2003 Apr}, pages = {160-5}, abstract = {Marine hydrothermal microorganisms respond rapidly to changes in the concentrations and availability of metals within their environment. Hyperthermophilic archaea appear to possess novel mechanisms for metal detoxification, dissimilatory metal reduction and metal assimilation that may be absent in their mesophilic and bacterial counterparts. For example, tungsten was found in high concentrations in a hydrothermal sulfide deposit where hyperthermophiles were also most abundant, consistent with the unique requirement of these organisms for this element. Furthermore, newly isolated genera of iron-reducing hyperthermophiles expand the scope of carbon cycling in hydrothermal environments. The advent of genome sequences and new molecular techniques will facilitate our further understanding of microbe-mineral interactions in these environments.
}, keywords = {Archaea, Bacteria, Environment, Marine Biology, Metals, Seawater, Temperature}, issn = {1367-5931}, author = {Holden, James F and Adams, Michael W W} } @article {568, title = {Microorganisms associated with uranium bioremediation in a high-salinity subsurface sediment.}, journal = {Appl Environ Microbiol}, volume = {69}, year = {2003}, month = {2003 Jun}, pages = {3672-5}, abstract = {Although stimulation of dissimilatory metal reduction to promote the reductive precipitation of uranium has been shown to successfully remove uranium from some aquifer sediments, the organisms in the family Geobacteraceae that have been found to be associated with metal reduction in previous studies are not known to grow at the high salinities found in some uranium-contaminated groundwaters. Studies with a highly saline uranium-contaminated aquifer sediment demonstrated that the addition of acetate could stimulate the removal of U(VI) from the groundwater. This removal was associated with an enrichment in microorganisms most closely related to Pseudomonas and Desulfosporosinus species.}, keywords = {Acetates, Biodegradation, Environmental, DNA, Ribosomal, Fresh Water, Geologic Sediments, Peptococcaceae, Polymerase Chain Reaction, Pseudomonas, RNA, Ribosomal, 16S, Sodium Chloride, Uranium, Water Pollution}, issn = {0099-2240}, author = {Nevin, Kelly P and Finneran, Kevin T and Lovley, Derek R} } @article {571, title = {OmcB, a c-type polyheme cytochrome, involved in Fe(III) reduction in Geobacter sulfurreducens.}, journal = {J Bacteriol}, volume = {185}, year = {2003}, month = {2003 Apr}, pages = {2096-103}, abstract = {Microorganisms in the family Geobacteraceae are the predominant Fe(III)-reducing microorganisms in a variety of subsurface environments in which Fe(III) reduction is an important process, but little is known about the mechanisms for electron transport to Fe(III) in these organisms. The Geobacter sulfurreducens genome was found to contain a 10-kb chromosomal duplication consisting of two tandem three-gene clusters. The last genes of the two clusters, designated omcB and omcC, encode putative outer membrane polyheme c-type cytochromes which are 79\% identical. The role of the omcB and omcC genes in Fe(III) reduction in G. sulfurreducens was investigated. OmcB and OmcC were both expressed during growth with acetate as the electron donor and either fumarate or Fe(III) as the electron acceptor. OmcB was ca. twofold more abundant under both conditions. Disrupting omcB or omcC by gene replacement had no impact on growth with fumarate. However, the OmcB-deficient mutant was greatly impaired in its ability to reduce Fe(III) both in cell suspensions and under growth conditions. In contrast, the ability of the OmcC-deficient mutant to reduce Fe(III) was similar to that of the wild type. When omcB was reintroduced into the OmcB-deficient mutant, the capacity for Fe(III) reduction was restored in proportion to the level of OmcB production. These results indicate that OmcB, but not OmcC, has a major role in electron transport to Fe(III) and suggest that electron transport to the outer membrane is an important feature in Fe(III) reduction in this organism.}, keywords = {Amino Acid Sequence, Bacterial Outer Membrane Proteins, Bacterial Proteins, Cytochrome c Group, Deltaproteobacteria, Gene Expression Regulation, Bacterial, Iron, Molecular Sequence Data, Multigene Family, Mutation, Oxidation-Reduction, Sequence Homology, Amino Acid}, issn = {0021-9193}, author = {Leang, Ching and Coppi, M V and Lovley, D R} } @article {43, title = {Opi1p, Ume6p and Sin3p control expression from the promoter of the INO2 regulatory gene via a novel regulatory cascade.}, journal = {Mol Microbiol}, volume = {48}, year = {2003}, month = {2003 May}, pages = {823-32}, abstract = {The INO2 gene of Saccharomyces cerevisiae is required for expression of most of the phospholipid biosynthetic genes. INO2 expression is regulated by a complex cascade that includes autoregulation, Opi1p-mediated repression and Ume6p-mediated activation. To screen for mutants with altered INO2 expression directly, we constructed an INO2-HIS3 reporter that provides a plate assay for INO2 promoter activity. This reporter was used to isolate mutants (dim1) that fail to repress expression of the INO2 gene in an otherwise wild-type strain. The dim1 mutants contain mutations in the OPI1 gene. To define further the mechanism for Ume6p regulation of INO2 expression, we isolated suppressors (rum1, 2, 3) of the ume6Delta mutation that overexpress the INO2-HIS3 gene. Two of the rum mutant groups contain mutations in the OPI1 and SIN3 genes showing that opi1 and sin3 mutations are epistatic to the ume6Delta mutation. These results are surprising given that Ume6p, Sin3p and Rpd3p are known to form a complex that represses the expression of a diverse set of yeast genes. This prompted us to examine the effect of sin3Delta and rpd3Delta mutants on INO2-cat expression. Surprisingly, the sin3Delta allele overexpressed INO2-cat, whereas the rpd3Delta mutant had no effect. We also show that the UME6 gene does not affect the expression of an OPI1-cat reporter. This suggests that Ume6p does not regulate INO2 expression indirectly by regulating OPI1 expression.}, keywords = {Basic Helix-Loop-Helix Transcription Factors, DNA-Binding Proteins, Fungal Proteins, Gene Expression Regulation, Fungal, Genes, Regulator, Genes, Reporter, Genetic Complementation Test, Histone Deacetylases, Methyltransferases, Phenotype, Phospholipids, Promoter Regions, Genetic, Repressor Proteins, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins, Transcription Factors}, issn = {0950-382X}, author = {Kaadige, Mohan R and Lopes, John M} } @article {775, title = {Progress in malaria research: the case for phylogenetics.}, journal = {Adv Parasitol}, volume = {54}, year = {2003}, month = {2003}, pages = {255-80}, abstract = {Malaria, from the Italian for "bad air", is a term used to describe a human disease caused by any of four parasites of the genus, Plasmodium. There are in fact over 200 described species of Plasmodium that parasitize reptiles, birds, and mammals, and may or may not cause disease in these various hosts. In this chapter, we highlight important evolutionary studies that have been undertaken to determine the relatedness among these species and their place in the taxonomic hierarchy. We begin by providing an overview of our present understanding of the phylum to which malaria parasites belong--Apicomplexa. The unique characteristics of these parasites reflect both their adaptation to the parasitic life style as well as some vestigial remnants of their pre-parasitic evolutionary past. Phylogenetic analyses provide the means for discerning the means by which these characteristics have come into existence. We next discuss the systematics of the genus Plasmodium. Morphology, genomic structure and content as well as host affiliation of these parasites are all traits that have been used for establishing taxonomic arrangements. Molecular phylogenetics has proven to be an invaluable tool in this regard and so we discuss the current phylogenetic picture of the genus as well as the correspondence among the various datasets (morphology, molecules, and host-preference). Lastly, we present a detailed account of our current understanding of the evolutionary past of the most deadly of the human malaria species--P. falciparum.}, keywords = {Animals, Biomedical Research, Evolution, Molecular, Genome, Malaria, Phylogeny, Plasmodium}, issn = {0065-308X}, author = {Rich, Stephen M and Ayala, Francisco J} } @article {567, title = {Rhodoferax ferrireducens sp. nov., a psychrotolerant, facultatively anaerobic bacterium that oxidizes acetate with the reduction of Fe(III).}, journal = {Int J Syst Evol Microbiol}, volume = {53}, year = {2003}, month = {2003 May}, pages = {669-73}, abstract = {To further investigate the diversity of micro-organisms capable of conserving energy to support growth from dissimilatory Fe(III) reduction, Fe(III)-reducing micro-organisms were enriched and isolated from subsurface sediments collected in Oyster Bay, VA, USA. A novel isolate, designated T118(T), was recovered in a medium with lactate as the sole electron donor and Fe(III) as the sole electron acceptor. Cells of T1 18(T) were Gram-negative, motile, short rods with a single polar flagellum. Strain T1 18(T) grew between pH 6.7 and 7.1, with a temperature range of 4-30 degrees C. The optimal growth temperature was 25 degrees C. Electron donors utilized by strain T1 18(T) with Fe(III) as the sole electron acceptor included acetate, lactate, malate, propionate, pyruvate, succinate and benzoate. None of the compounds tested was fermented. Electron acceptors utilized with either acetate or lactate as the electron donor included Fe(III)-NTA (nitrilotriacetic acid), Mn(IV) oxide, nitrate, fumarate and oxygen. Phylogenetic analysis demonstrated that strain T1 18(T) is most closely related to the genus Rhodoferax. Unlike other species in this genus, strain T1 18(T) is not a phototroph and does not ferment fructose. However, phototrophic genes may be present but not expressed under the experimental conditions tested. No Rhodoferax species have been reported to grow via dissimilatory Fe(III) reduction. Based on these physiological and phylogenetic differences, strain T1 18(T) (=ATCC BAA-621(T) = DSM 15236(T)) is proposed as a novel species, Rhodoferax ferrireducens sp. nov.}, keywords = {Acetates, Anaerobiosis, Bacterial Typing Techniques, Betaproteobacteria, Cold Temperature, Culture Media, DNA, Ribosomal, Electron Transport, Ferric Compounds, Fresh Water, Molecular Sequence Data, Oxidation-Reduction, Phylogeny, RNA, Ribosomal, 16S, Sequence Analysis, DNA}, issn = {1466-5026}, author = {Finneran, Kevin T and Johnsen, Claudia V and Lovley, Derek R} } @article {777, title = {Sequence diversity and evolution of the malaria vaccine candidate merozoite surface protein-1 (MSP-1) of Plasmodium falciparum.}, journal = {Gene}, volume = {304}, year = {2003}, month = {2003 Jan 30}, pages = {65-75}, abstract = {The merozoite surface protein-1 (MSP-1) of the malaria parasite Plasmodium falciparum is a major blood-stage antigen containing highly polymorphic tripeptide repeats in the domain known as block 2 and several non-repetitive domains that are essentially dimorphic. We have analyzed sequence variation in block 2 repeats and in non-repetitive block 17, as well as other polymorphisms within the MSP-1 gene, in clinical isolates of P. falciparum. Repeat haplotypes were defined as unique combinations of repeat motifs within block 2, whereas block 17 haplotypes were defined as unique combinations of single nucleotide replacements in this domain. A new block 17 haplotype, E-TNG-L, was found in one isolate from Vietnam. MSP-1 alleles, defined as unique combinations of haplotypes in blocks 2 and 17 and other polymorphisms within the molecule, were characterized in 60 isolates from hypoendemic Brazil and 37 isolates from mesoendemic Vietnam. Extensive diversity has been created in block 2 and elsewhere in the molecule, while maintaining significant linkage disequilibrium between polymorphisms across the non-telomeric MSP-1 locus separated by a map distance of more than 4 kb, suggesting that low meiotic recombination rates occur in both parasite populations. These results indicate a role for non-homologous recombination, such as strand-slippage mispairing during mitosis and gene conversion, in creating variation in a malarial antigen under strong diversifying selection.}, keywords = {Alleles, Animals, Brazil, DNA, Protozoan, Evolution, Molecular, Genetic Variation, Haplotypes, Linkage Disequilibrium, Malaria Vaccines, Merozoite Surface Protein 1, Molecular Sequence Data, Plasmodium falciparum, Polymorphism, Single Nucleotide, Recombination, Genetic, Sequence Analysis, DNA, Tanzania, Thailand, Vietnam}, issn = {0378-1119}, author = {Ferreira, Marcelo U and Ribeiro, Weber L and Tonon, Angela P and Kawamoto, Fumihiko and Rich, Stephen M} } @article {563, title = {Stimulating the in situ activity of Geobacter species to remove uranium from the groundwater of a uranium-contaminated aquifer.}, journal = {Appl Environ Microbiol}, volume = {69}, year = {2003}, month = {2003 Oct}, pages = {5884-91}, abstract = {The potential for removing uranium from contaminated groundwater by stimulating the in situ activity of dissimilatory metal-reducing microorganisms was evaluated in a uranium-contaminated aquifer located in Rifle, Colo. Acetate (1 to 3 mM) was injected into the subsurface over a 3-month period via an injection gallery composed of 20 injection wells, which was installed upgradient from a series of 15 monitoring wells. U(VI) concentrations decreased in as little as 9 days after acetate injection was initiated, and within 50 days uranium had declined below the prescribed treatment level of 0.18 micro M in some of the monitoring wells. Analysis of 16S ribosomal DNA (rDNA) sequences and phospholipid fatty acid profiles demonstrated that the initial loss of uranium from the groundwater was associated with an enrichment of Geobacter species in the treatment zone. Fe(II) in the groundwater also increased during this period, suggesting that U(VI) reduction was coincident with Fe(III) reduction. As the acetate injection continued over 50 days there was a loss of sulfate from the groundwater and an accumulation of sulfide and the composition of the microbial community changed. Organisms with 16S rDNA sequences most closely related to those of sulfate reducers became predominant, and Geobacter species became a minor component of the community. This apparent switch from Fe(III) reduction to sulfate reduction as the terminal electron accepting process for the oxidation of the injected acetate was associated with an increase in uranium concentration in the groundwater. These results demonstrate that in situ bioremediation of uranium-contaminated groundwater is feasible but suggest that the strategy should be optimized to better maintain long-term activity of Geobacter species.}, keywords = {Acetates, DNA, Ribosomal, Ecosystem, Fatty Acids, Ferric Compounds, Fresh Water, Geobacter, Mining, Oxidation-Reduction, Phospholipids, RNA, Ribosomal, 16S, Sulfates, Uranium, Water Pollution, Chemical}, issn = {0099-2240}, author = {Anderson, Robert T and Vrionis, Helen A and Ortiz-Bernad, Irene and Resch, Charles T and Long, Philip E and Dayvault, Richard and Karp, Ken and Marutzky, Sam and Metzler, Donald R and Peacock, Aaron and White, David C and Lowe, Mary and Lovley, Derek R} } @article {569, title = {Thermophily in the Geobacteraceae: Geothermobacter ehrlichii gen. nov., sp. nov., a novel thermophilic member of the Geobacteraceae from the "Bag City" hydrothermal vent.}, journal = {Appl Environ Microbiol}, volume = {69}, year = {2003}, month = {2003 May}, pages = {2985-93}, abstract = {Little is known about the microbiology of the "Bag City" hydrothermal vent, which is part of a new eruption site on the Juan de Fuca Ridge and which is notable for its accumulation of polysaccharide on the sediment surface. A pure culture, designated strain SS015, was recovered from a vent fluid sample from the Bag City site through serial dilution in liquid medium with malate as the electron donor and Fe(III) oxide as the electron acceptor and then isolation of single colonies on solid Fe(III) oxide medium. The cells were gram-negative rods, about 0.5 micro m by 1.2 to 1.5 micro m, and motile and contained c-type cytochromes. Analysis of the 16S ribosomal DNA (rDNA) sequence of strain SS015 placed it in the family Geobacteraceae in the delta subclass of the Proteobacteria. Unlike previously described members of the Geobacteraceae, which are mesophiles, strain SS015 was a thermophile and grew at temperatures of between 35 and 65 degrees C, with an optimum temperature of 55 degrees C. Like many previously described members of the Geobacteraceae, strain SS015 grew with organic acids as the electron donors and Fe(III) or nitrate as the electron acceptor, with nitrate being reduced to ammonia. Strain SS015 was unique among the Geobacteraceae in its ability to use sugars, starch, or amino acids as electron donors for Fe(III) reduction. Under stress conditions, strain SS015 produced copious quantities of extracellular polysaccharide, providing a model for the microbial production of the polysaccharide accumulation at the Bag City site. The 16S rDNA sequence of strain SS015 was less than 94\% similar to the sequences of previously described members of the Geobacteraceae; this fact, coupled with its unique physiological properties, suggests that strain SS015 represents a new genus in the family Geobacteraceae. The name Geothermobacter ehrlichii gen. nov., sp. nov., is proposed (ATCC BAA-635 and DSM 15274). Although strains of Geobacteraceae are known to be the predominant Fe(III)-reducing microorganisms in a variety of Fe(III)-reducing environments at moderate temperatures, strain SS015 represents the first described thermophilic member of the Geobacteraceae and thus extends the known environmental range of this family to hydrothermal environments.}, keywords = {Base Composition, Cytochromes, Deltaproteobacteria, DNA, Bacterial, DNA, Ribosomal, Drug Resistance, Bacterial, Ecosystem, Electron Transport, Geologic Sediments, Hot Temperature, Hydrogen-Ion Concentration, Iron, Microscopy, Electron, Molecular Sequence Data, Oxidation-Reduction, Pacific Ocean, Phylogeny, Seawater, Sodium Chloride, Species Specificity}, issn = {0099-2240}, author = {Kashefi, Kazem and Holmes, Dawn E and Baross, John A and Lovley, Derek R} } @article {386, title = {Trypanosoma brucei has two distinct mitochondrial DNA polymerase beta enzymes.}, journal = {J Biol Chem}, volume = {278}, year = {2003}, month = {2003 Dec 5}, pages = {49095-101}, abstract = {In higher eukaryotes, DNA polymerase (pol) beta resides in the nucleus and participates primarily in DNA repair. The DNA polymerase beta from the trypanosomatid Crithidia fasciculata, however, was the first mitochondrial enzyme of this type described. Upon searching the nearly completed genome data base of the related parasite Trypanosoma brucei, we discovered genes for two pol beta-like proteins. One is approximately 70\% identical to the C. fasciculata pol beta and is likely the homolog of this enzyme. The other, although approximately 30\% identical within the polymerase region, has unusual structural features including a short C-terminal tail and a long N-terminal extension rich in prolines, alanines, and lysines. Both proteins, when expressed recombinantly, are active as DNA polymerases and deoxyribose phosphate lyases, but their polymerase activity optima differ with respect to pH and KCl and MgCl2 concentrations. Remarkably, green fluorescent protein fusion proteins and immunofluorescence demonstrate that both are mitochondrial, but their locations with respect to the mitochondrial DNA (kinetoplast DNA network) in this organism are strikingly different.}, keywords = {Animals, Base Sequence, Cell Line, DNA Polymerase beta, DNA Primers, Isoenzymes, Mitochondria, Recombinant Proteins, Trypanosoma brucei brucei}, issn = {0021-9258}, doi = {10.1074/jbc.M308565200}, author = {Saxowsky, Tina T and Choudhary, Gunjan and Klingbeil, Michele M and Englund, Paul T} } @article {575, title = {Anaerobic, sulfate-dependent degradation of polycyclic aromatic hydrocarbons in petroleum-contaminated harbor sediment.}, journal = {Environ Sci Technol}, volume = {36}, year = {2002}, month = {2002 Nov 15}, pages = {4811-7}, abstract = {It has previously been demonstrated that [14C]-labeled polycyclic aromatic hydrocarbons (PAHs) can be oxidized to 14CO2 in anoxic, PAH-contaminated, marine harbor sediments in which sulfate reduction is the terminal electron-accepting process. However, it has not previously been determined whether this degradation of [14C]-PAHs accurately reflects the degradation of the in situ pools of contaminant PAHs. In coal tar-contaminated sediments from Boston Harbor, [14C]-naphthalene was readily oxidized to 14CO2, but, after 95 d of incubation under anaerobic conditions, there was no significant decrease in the detectable pool of in situ naphthalene in these sediments. Therefore, to better evaluate the anaerobic biodegradation of the in situ PAH pools, the concentrations of these contaminants were monitored for ca. 1 year during which the sediments were incubated under conditions that mimicked those found in situ. There was loss of all of the PAHs that were monitored (2-5 ring congeners), including high molecular weight PAHs, such as benzo[a]pyrene, that have not previously been shown to be degraded under anaerobic conditions. There was no significant change in the PAH levels in the sediments amended with molybdate to inhibit sulfate-reducing bacteria or in sediments in which all microorganisms had been killed with glutaraldehyde. In some instances, over half of the detectable pools of in situ 2-3 ring PAHs were degraded. In general, the smaller PAHs were degraded more rapidly than the larger PAHs. A distinct exception in the Boston Harbor sediment was naphthalene which was degraded very slowly at a rate comparable to the larger PAHs. In a similar in situ-like study of fuel-contaminated sediments from Liepaja Harbor, Latvia, there was no decline in PAH levels in samples that were sulfate-depleted. However, when the Latvia sediments were supplemented with sufficient sodium sulfate or gypsum to elevate pore water levels of sulfate to approximately 14-25 mM there was a 90\% decline in the naphthalene and a 60\% decline in the 2-methylnaphthalene pool within 90 days. These studies demonstrate for the first time that degradation by anaerobic microorganisms can significantly impact the in situ pools of PAHs in petroleum-contaminated, anoxic, sulfate-reducing harbor sediments and suggest that the self-purification capacity of contaminated harbor sediments is greater than previously considered.}, keywords = {Bacteria, Anaerobic, Biodegradation, Environmental, Environmental Monitoring, Geologic Sediments, Molecular Weight, Petroleum, Polycyclic Hydrocarbons, Aromatic, Ships, Sulfur-Reducing Bacteria, Transportation, Water Pollutants, Chemical}, issn = {0013-936X}, author = {Rothermich, Mary M and Hayes, Lory A and Lovley, Derek R} } @article {572, title = {Analysis of the genetic potential and gene expression of microbial communities involved in the in situ bioremediation of uranium and harvesting electrical energy from organic matter.}, journal = {OMICS}, volume = {6}, year = {2002}, month = {2002}, pages = {331-9}, abstract = {The proposed research will investigate two microbial communities that are of direct relevance to Department of Energy interests. One is the microbial community associated with the in situ bioremediation of uranium-contaminated groundwater. The second is a microbial community that harvests energy from waste organic matter in the form of electricity. These studies will address DOE needs for (1) remediation of metals and radionuclides at DOE sites and (2) the development of cleaner forms of energy and biomass conversion to energy. Our previous studies have demonstrated that the microbial communities involved in uranium bioremediation and energy harvesting are both dominated by microorganisms in the family Geobacteraceae and that the organisms in this family are responsible for uranium bioremediation and electron transfer to electrodes. The initial objectives of this study are to (1) describe the genetic potential of the Geobacteraceae that predominate in the environments of interest; (2) identify conserved patterns of gene expression within the Geobacteraceae family in response to a range of environmental conditions; (3) begin to identify mechanisms controlling the expression of key genes related to survival, growth, and activity in subsurface environments and on electrodes; and (4) use the results from subobjectives 1-3 to develop a conceptual model for predicting gene expression of Geobacteraceae in the environments of interest. This will serve as the basis for a subsequent simulation model of the growth and activity of Geobacteraceae in the subsurface and on electrodes.}, keywords = {Biodegradation, Environmental, Deltaproteobacteria, Electricity, Electrodes, Energy Metabolism, Gene Expression Regulation, Bacterial, Genome, Bacterial, Geologic Sediments, Models, Theoretical, Uranium}, issn = {1536-2310}, doi = {10.1089/153623102321112755}, author = {Lovley, Derek R} } @article {780, title = {Ancient lineages of non-LTR retrotransposons in the primitive eukaryote, Giardia lamblia.}, journal = {Mol Biol Evol}, volume = {19}, year = {2002}, month = {2002 May}, pages = {619-30}, abstract = {Mobile elements that use reverse transcriptase to make new copies of themselves are found in all major lineages of eukaryotes. The non-long terminal repeat (non-LTR) retrotransposons have been suggested to be the oldest of these eukaryotic elements. Phylogenetic analysis of non-LTR elements suggests that they have predominantly undergone vertical transmission, as opposed to the frequent horizontal transmissions found for other mobile elements. One prediction of this vertical model of inheritance is that the oldest lineages of eukaryotes should exclusively harbor the oldest lineages of non-LTR retrotransposons. Here we characterize the non-LTR retrotransposons present in one of the most primitive eukaryotes, the diplomonad Giardia lamblia. Two families of elements were detected in the WB isolate of G. lamblia currently being used for the genome sequencing project. These elements are clearly distinct from all other previously described non-LTR lineages. Phylogenetic analysis indicates that these Genie elements (for Giardia early non-LTR insertion element) are among the oldest known lineages of non-LTR elements consistent with strict vertical descent. Genie elements encode a single open reading frame with a carboxyl terminal endonuclease domain. Genie 1 is site specific, as seven to eight copies are present in a single tandem array of a 771-bp repeat near the telomere of one chromosome. The function of this repeat is not known. One additional, highly divergent, element within the Genie 1 lineage is not located in this tandem array but is near a second telomere. Four different telomere addition sites could be identified within or near the Genie elements on each of these chromosomes. The second lineage of non-LTR elements, Genie 2, is composed of about 10 degenerate copies. Genie 2 elements do not appear to be site specific in their insertion. An unusual aspect of Genie 2 is that all copies contain inverted repeats up to 172 bp in length.}, keywords = {Animals, Base Sequence, DNA, Protozoan, Evolution, Molecular, Giardia lamblia, Long Interspersed Nucleotide Elements, Molecular Sequence Data, Open Reading Frames, Phylogeny, Repetitive Sequences, Nucleic Acid, Sequence Homology, Nucleic Acid, Telomere}, issn = {0737-4038}, author = {Burke, William D and Malik, Harmit S and Rich, Stephen M and Eickbush, Thomas H} } @article {714, title = {Caveolar endocytosis of simian virus 40 is followed by brefeldin A-sensitive transport to the endoplasmic reticulum, where the virus disassembles.}, journal = {J Virol}, volume = {76}, year = {2002}, month = {2002 May}, pages = {5156-66}, abstract = {Simian virus 40 (SV40) enters cells by atypical endocytosis mediated by caveolae that transports the virus to the endoplasmic reticulum (ER) instead of to the endosomal-lysosomal compartment, which is the usual destination for viruses and other cargo that enter by endocytosis. We show here that SV4O is transported to the ER via an intermediate compartment that contains beta-COP, which is best known as a component of the COPI coatamer complexes that are required for the retrograde retrieval pathway from the Golgi to the ER. Additionally, transport of SV40 to the ER, as well as infection, is sensitive to brefeldin A. This drug acts by specifically inhibiting the ARF1 GTPase, which is known to regulate assembly of COPI coat complexes on Golgi cisternae. Moreover, some beta-COP colocalizes with intracellular caveolin-1, which was previously shown to be present on a new organelle (termed the caveosome) that is an intermediate in the transport of SV40 to the ER (L. Pelkmans, J. Kartenbeck, and A. Helenius, Nat. Cell Biol. 3:473-483, 2001). We also show that the internal SV40 capsid proteins VP2 and VP3 become accessible to immunostaining starting at about 5 h. Most of that immunostaining overlays the ER, with some appearing outside of the ER. In contrast, immunostaining with anti-SV40 antisera remains confined to the ER.}, keywords = {Animals, Antiviral Agents, Biological Transport, Brefeldin A, Capsid, Caveolae, Coatomer Protein, Endocytosis, Endoplasmic Reticulum, Simian virus 40, Virus Replication}, issn = {0022-538X}, author = {Norkin, Leonard C and Anderson, Howard A and Wolfrom, Scott A and Oppenheim, Ariella} } @article {399, title = {A comprehensive alanine scanning mutagenesis of the Escherichia coli transcriptional activator SoxS: identifying amino acids important for DNA binding and transcription activation.}, journal = {J Mol Biol}, volume = {322}, year = {2002}, month = {2002 Sep 13}, pages = {237-57}, abstract = {SoxS is the direct transcriptional activator of the superoxide regulon. SoxS recognizes a highly degenerate "soxbox" DNA sequence, and activates transcription from class I and class II promoters. SoxS is the smallest member of the AraC/XylS family of transcription regulators whose hallmark is dual helix-turn-helix (HTH) DNA-binding motifs. Evidence suggests that the N-terminal HTH motif of SoxS interacts with a highly conserved region of the soxbox termed recognition element 1 (RE1), while the C-terminal HTH motif interacts with the less conserved recognition element 2 (RE2). In the work described here, we prepared a complete library of 101 SoxS mutants containing single alanine substitutions of SoxS, and we characterized the mutant proteins in vivo and in vitro. With SoxS being closely related to MarA, we analyzed the effects of the SoxS mutations in the context of the MarA-mar crystal structure and with respect to the NMR study of MarA-DNA complexes in solution. From the properties of the alanine substitutions, we conclude the following. (1) Surface-exposed residues of helix 3 and helix 6, the recognition helices of the dual HTH motifs, are important to DNA binding and transcription activation; however, substitutions of residues predicted from the MarA-mar crystal structure to make contact with the sugar-phosphate backbone are more detrimental to DNA binding than mutations predicted to make base-specific contacts. (2) Substitution of several residues within the recognition helix predicted to make base-specific contacts with RE2 have relatively little effect on DNA-binding, suggesting the possibility of alternative protein-DNA interactions than those inferred from the MarA-mar crystal structure. (3) DNA binding and transcription activation were reduced by substitution of conserved amino acid residues comprising the hydrophobic core, presumably because they disrupt the structural integrity of SoxS. (4) Mutant K30A appears to be a positive control mutant defective in a protein-protein interaction with RNA polymerase that is required for transcription activation at all SoxS-dependent promoters because it binds and bends DNA normally but fails to activate transcription from both classes of promoters. Alanine substitutions of surface-exposed residues H3, K5, D9, S31, and V45 confer a similar phenotype. Since these residues are near K30 on the surface of the protein, the surface formed by the six residues may be used to make protein-protein interactions with RNA polymerase that are required for transcription activation at both class I and class II SoxS-dependent promoters. (5) Mutants F74A, D75A, M78A, D79A and Q85A appear to define a surface required for protein-protein interaction with RNA polymerase specifically at class II promoters because these positive control mutants bind and bend DNA normally but are defective in activation of class II promoters but not class I promoters. These SoxS mutants that bind and bend DNA normally but are defective in transcription activation represent the first positive control mutants with putative defects in protein-protein interactions with RNA polymerase among the SoxS/MarA/Rob subset of the AraC/XylS family of transcription regulators.}, keywords = {Alanine, Amino Acid Substitution, Base Sequence, Binding Sites, DNA, DNA-Binding Proteins, Electrophoretic Mobility Shift Assay, Escherichia coli, Escherichia coli Proteins, Gene Expression Regulation, Bacterial, Genes, Lethal, Hydrophobic and Hydrophilic Interactions, Lac Operon, Models, Molecular, Mutagenesis, Nucleic Acid Conformation, Phenotype, Phosphates, Promoter Regions, Genetic, Protein Binding, Protein Structure, Tertiary, Structure-Activity Relationship, Trans-Activators, Transcriptional Activation}, issn = {0022-2836}, author = {Griffith, Kevin L and Wolf, Richard E} } @article {574, title = {Desulfitobacterium metallireducens sp. nov., an anaerobic bacterium that couples growth to the reduction of metals and humic acids as well as chlorinated compounds.}, journal = {Int J Syst Evol Microbiol}, volume = {52}, year = {2002}, month = {2002 Nov}, pages = {1929-35}, abstract = {Strain 853-15A(T) was enriched and isolated from uranium-contaminated aquifer sediment by its ability to grow under anaerobic conditions via the oxidation of lactate coupled to the reduction of anthraquinone-2,6-disulfonate (AQDS) to anthrahydroquinone-2,6-disulfonate (AHQDS). Lactate was oxidized incompletely to acetate and carbon dioxide according to the reaction CH3CHOHCOO(-)+ 2AQDS+H2O --> CH3COO(-)+ 2AHQDS+CO2. Additional electron donors utilized included formate, ethanol, butanol, butyrate, malate and pyruvate. Lactate also supported growth with Fe(III) citrate, Mn(IV) oxide, humic substances, elemental sulfur, 3-chloro-4-hydroxyphenylacetate, trichloroethylene or tetrachloroethylene serving as the electron acceptor. Growth was not observed with sulfate, sulfite, nitrate or fumarate as the terminal electron acceptor. The temperature optimum for growth was 30 degrees C, but growth was also observed at 20 and 37 degrees C. The pH optimum was approximately 7.0. The 16S rDNA sequence of strain 853-15A(T) suggested that it was most closely related to Desulfitobacterium dehalogenans and closely related to Desulfitobacterium chlororespirans and Desulfitobacterium frappieri. The phylogenetic and physiological properties exhibited by strain 853-15A(T) (= ATCC BAA-636(T)) place it within the genus Desulfitobacterium as the type strain of a novel species, Desulfitobacterium metallireducens sp. nov.}, keywords = {Anaerobiosis, DNA, Bacterial, DNA, Ribosomal, Ecosystem, Electron Transport, Geologic Sediments, Humic Substances, Metals, Microscopy, Electron, Molecular Sequence Data, Oxidation-Reduction, Peptococcaceae, Phenotype, Phylogeny, RNA, Bacterial, RNA, Ribosomal, 16S, Species Specificity}, issn = {1466-5026}, author = {Finneran, Kevin T and Forbush, Heather M and VanPraagh, Catherine V Gaw and Lovley, Derek R} } @article {588, title = {Electrode-reducing microorganisms that harvest energy from marine sediments.}, journal = {Science}, volume = {295}, year = {2002}, month = {2002 Jan 18}, pages = {483-5}, abstract = {Energy in the form of electricity can be harvested from marine sediments by placing a graphite electrode (the anode) in the anoxic zone and connecting it to a graphite cathode in the overlying aerobic water. We report a specific enrichment of microorganisms of the family Geobacteraceae on energy-harvesting anodes, and we show that these microorganisms can conserve energy to support their growth by oxidizing organic compounds with an electrode serving as the sole electron acceptor. This finding not only provides a method for extracting energy from organic matter, but also suggests a strategy for promoting the bioremediation of organic contaminants in subsurface environments.}, keywords = {Aerobiosis, Anaerobiosis, Anthraquinones, Benzoates, Biodegradation, Environmental, Carbon Dioxide, Colony Count, Microbial, Deltaproteobacteria, DNA, Ribosomal, Electricity, Electrodes, Electrons, Energy Metabolism, Geologic Sediments, Oxidation-Reduction, RNA, Ribosomal, 16S, Seawater, Sodium Acetate}, issn = {1095-9203}, doi = {10.1126/science.1066771}, author = {Bond, Daniel R and Holmes, Dawn E and Tender, Leonard M and Lovley, Derek R} } @article {583, title = {Enrichment of members of the family Geobacteraceae associated with stimulation of dissimilatory metal reduction in uranium-contaminated aquifer sediments.}, journal = {Appl Environ Microbiol}, volume = {68}, year = {2002}, month = {2002 May}, pages = {2300-6}, abstract = {Stimulating microbial reduction of soluble U(VI) to insoluble U(IV) shows promise as a strategy for immobilizing uranium in uranium-contaminated subsurface environments. In order to learn more about which microorganisms might be involved in U(VI) reduction in situ, the changes in the microbial community when U(VI) reduction was stimulated with the addition of acetate were monitored in sediments from three different uranium-contaminated sites in the floodplain of the San Juan River in Shiprock, N.Mex. In all three sediments U(VI) reduction was accompanied by concurrent Fe(III) reduction and a dramatic enrichment of microorganisms in the family Geobacteraceae, which are known U(VI)- and Fe(III)-reducing microorganisms. At the point when U(VI) reduction and Fe(III) reduction were nearing completion, Geobacteraceae accounted for ca. 40\% of the 16S ribosomal DNA (rDNA) sequences recovered from the sediments with bacterial PCR primers, whereas Geobacteraceae accounted for fewer than 5\% of the 16S rDNA sequences in control sediments that were not amended with acetate and in which U(VI) and Fe(III) reduction were not stimulated. Between 55 and 65\% of these Geobacteraceae sequences were most similar to sequences from Desulfuromonas species, with the remainder being most closely related to Geobacter species. Quantitative analysis of Geobacteraceae sequences with most-probable-number PCR and TaqMan analyses indicated that the number of Geobacteraceae sequences increased from 2 to 4 orders of magnitude over the course of U(VI) and Fe(III) reduction in the acetate-amended sediments from the three sites. No increase in Geobacteraceae sequences was observed in control sediments. In contrast to the predominance of Geobacteraceae sequences, no sequences related to other known Fe(III)-reducing microorganisms were detected in sediments. These results compare favorably with an increasing number of studies which have demonstrated that Geobacteraceae are important components of the microbial community in a diversity of subsurface environments in which Fe(III) reduction is an important process. The combination of these results with the finding that U(VI) reduction takes place during Fe(III) reduction and prior to sulfate reduction suggests that Geobacteraceae will be responsible for much of the Fe(III) and U(VI) reduction during uranium bioremediation in these sediments.}, keywords = {Gene Library, Geologic Sediments, Iron, Oxidation-Reduction, Proteobacteria, RNA, Ribosomal, 16S, Soil Pollutants, Uranium}, issn = {0099-2240}, author = {Holmes, Dawn E and Finneran, Kevin T and O{\textquoteright}Neil, Regina A and Lovley, Derek R} } @article {400, title = {Evidence for "pre-recruitment" as a new mechanism of transcription activation in Escherichia coli: the large excess of SoxS binding sites per cell relative to the number of SoxS molecules per cell.}, journal = {Biochem Biophys Res Commun}, volume = {291}, year = {2002}, month = {2002 Mar 8}, pages = {979-86}, abstract = {In response to the oxidative stress imposed by redox-cycling compounds like paraquat, Escherichia coli induces the synthesis of SoxS, which then activates the transcription of approximately 100 genes. The DNA binding site for SoxS-dependent transcription activation, the "soxbox," is highly degenerate, suggesting that the genome contains a large number of SoxS binding sites. To estimate the number of soxboxes in the cell, we searched the E. coli genome for SoxS binding sites using as query sequence the previously determined optimal SoxS binding sequence. We found approximately 12,500 sequences that match the optimal binding sequence under the conditions of our search; this agrees with our previous estimate, based on information theory, that a random sequence the size of the E. coli genome contains approximately 13,000 soxboxes. Thus, fast-growing cells with 4-6 genomes per cell have approximately 65,000 soxboxes. This large number of potential SoxS binding sites per cell raises the interesting question of how SoxS distinguishes between the functional soxboxes located within the promoters of target genes and the plethora of equivalent but nonfunctional binding sites scattered throughout the chromosome. To address this question, we treated cells with paraquat and used Western blot analysis to determine the kinetics of SoxS accumulation per cell; we also determined the kinetics of SoxS-activated gene expression. The abundance of SoxS reached a maximum of 2,500 molecules per cell 20 min after induction and gradually declined to approximately 500 molecules per cell over the next 1.5 h. Given that activation of target gene expression began almost immediately and given the large disparity between the number of SoxS molecules per cell, 2,500, and the number of SoxS binding sites per cell, 65,000, we infer that SoxS is not likely to activate transcription by the usual "recruitment" pathway, as this mechanism would require a number of SoxS molecules similar to the number of soxboxes. Instead, we propose that SoxS first interacts in solution with RNA polymerase and then the binary complex scans the chromosome for promoters that contain a soxbox properly positioned and oriented for transcription activation. We name this new pathway "pre-recruitment."}, keywords = {Bacterial Proteins, Binding Sites, Blotting, Western, Cell Division, DNA-Binding Proteins, Escherichia coli, Escherichia coli Proteins, Gene Expression Regulation, Bacterial, Genome, Bacterial, Kinetics, Numerical Analysis, Computer-Assisted, Oxidative Stress, Paraquat, Protein Transport, Trans-Activators, Transcription Factors, Transcriptional Activation}, issn = {0006-291X}, doi = {10.1006/bbrc.2002.6559}, author = {Griffith, Kevin L and Shah, Ishita M and Myers, Todd E and O{\textquoteright}Neill, Michael C and Wolf, Richard E} } @article {782, title = {Experimental evidence for genetic recombination in the opportunistic pathogen Cryptosporidium parvum.}, journal = {Mol Biochem Parasitol}, volume = {119}, year = {2002}, month = {2002 Jan}, pages = {55-62}, abstract = {Cryptosporidium parvum is an intracellular protozoan parasite causing intestinal malabsorption and diarrhea in humans. The infection is usually self-limiting, although persistent cryptosporidosis is observed in immunocompromised and malnourished individuals. As with other Apicomplexa, the life cycle of Cryptosporidium is thought to comprise a sexual phase, during which a motile microgamont fuses with a sessile macrogamont. The four sporozoites found within each oocyst (the infectious form excreted in the feces) are thought to be the product of a meiotic division taking place immediately following fertilization, but the existence of a meiotic cycle in this genus has not been tested experimentally. To substantiate the occurrence of meiotic recombination in this species, we performed a genetic cross between two distinct isolates of C. parvum co-infected in INF-gamma knockout mice. We found that mixed infections produced recombinant progeny characterized by multilocus genotypes comprising alleles inherited from each parental line. This observation represents the first demonstration of sexual recombination in this pathogen. Together with the occurrence of genetically heterogeneous infections, this finding suggests that outcrossing between genotypes may occur in nature. Experimental crosses among Cryptosporidium populations will facilitate mapping of clinically relevant genes, the delineation of Cryptosporidium species, and defining the taxonomical status of C. parvum subtypes and host-specific genotypes.}, keywords = {Animals, Conserved Sequence, Crosses, Genetic, Cryptosporidiosis, Cryptosporidium parvum, Deer, Female, Genotype, Humans, Male, Mice, Mice, Knockout, Microsatellite Repeats, Opportunistic Infections, Polymerase Chain Reaction, Recombination, Genetic, Survival Rate}, issn = {0166-6851}, author = {Feng, Xiaochuan and Rich, Stephen M and Tzipori, Saul and Widmer, Giovanni} } @article {578, title = {Fulvic acid oxidation state detection using fluorescence spectroscopy.}, journal = {Environ Sci Technol}, volume = {36}, year = {2002}, month = {2002 Jul 15}, pages = {3170-5}, abstract = {Humic substances are a heterogeneous class of moderate molecular weight, yellow-colored biomolecules present in all soils, sediments, and natural waters. Although humic substances are generally resistant to microbial degradation under anaerobic conditions, some microorganisms in soils and sediments can use quinone moieties in humic substances as electron acceptors. Laboratory experiments have shown that humic substances can act as electron shuttles in the microbial reduction of ferric iron. Field studies of electron shuttling processes have been constrained by the lack of methods to characterize the oxidation state of quinone moieties in humic substances at natural concentrations. All humic substances have fluorescent properties, and fluorescence spectroscopy can indicate differences in precursor organic source of humic substances. Here we show that the quinone moieties responsible for electron transfer reactions contribute significantly to the fluorescence of humic substances. Further we use fluorescence spectroscopy to elucidate the oxidation state of quinone moieties in humic substances at natural concentrations found in sediment interstitial waters.}, keywords = {Benzopyrans, Environmental Monitoring, Iron, Oxidation-Reduction, Spectrometry, Fluorescence}, issn = {0013-936X}, author = {Klapper, Lisa and McKnight, Diane M and Fulton, J Robin and Blunt-Harris, Elizabeth L and Nevin, Kelly P and Lovley, Derek R and Hatcher, Patrick G} } @article {586, title = {Geobacter metallireducens accesses insoluble Fe(III) oxide by chemotaxis.}, journal = {Nature}, volume = {416}, year = {2002}, month = {2002 Apr 18}, pages = {767-9}, abstract = {Microorganisms that use insoluble Fe(III) oxide as an electron acceptor can have an important function in the carbon and nutrient cycles of aquatic sediments and in the bioremediation of organic and metal contaminants in groundwater. Although Fe(III) oxides are often abundant, Fe(III)-reducing microbes are faced with the problem of how to access effectively an electron acceptor that can not diffuse to the cell. Fe(III)-reducing microorganisms in the genus Shewanella have resolved this problem by releasing soluble quinones that can carry electrons from the cell surface to Fe(III) oxide that is at a distance from the cell. Here we report that another Fe(III)-reducer, Geobacter metallireducens, has an alternative strategy for accessing Fe(III) oxides. Geobacter metallireducens specifically expresses flagella and pili only when grown on insoluble Fe(III) or Mn(IV) oxide, and is chemotactic towards Fe(II) and Mn(II) under these conditions. These results suggest that G. metallireducens senses when soluble electron acceptors are depleted and then synthesizes the appropriate appendages to permit it to search for, and establish contact with, insoluble Fe(III) or Mn(IV) oxide. This approach to the use of an insoluble electron acceptor may explain why Geobacter species predominate over other Fe(III) oxide-reducing microorganisms in a wide variety of sedimentary environments.}, keywords = {Bacterial Proteins, Chemotaxis, Deltaproteobacteria, DNA-Binding Proteins, Ferric Compounds, Ferrous Compounds, Fimbriae Proteins, Fimbriae, Bacterial, Flagella, Manganese Compounds, Movement, Oxidation-Reduction, Oxides, Solubility}, issn = {0028-0836}, doi = {10.1038/416767a}, author = {Childers, Susan E and Ciufo, Stacy and Lovley, Derek R} } @article {581, title = {Geoglobus ahangari gen. nov., sp. nov., a novel hyperthermophilic archaeon capable of oxidizing organic acids and growing autotrophically on hydrogen with Fe(III) serving as the sole electron acceptor.}, journal = {Int J Syst Evol Microbiol}, volume = {52}, year = {2002}, month = {2002 May}, pages = {719-28}, abstract = {A novel, regular to irregular, coccoid-shaped, anaerobic, Fe(III)-reducing microorganism was isolated from the Guaymas Basin hydrothermal system at a depth of 2000 m. Isolation was carried out with a new technique using Fe(III) oxide as the electron acceptor for the recovery of colonies on solid medium. The isolate, designated strain 234T, was strictly anaerobic and exhibited a tumbling motility. The cells had a single flagellum. Strain 234T grew at temperatures between 65 and 90 degrees C, with an optimum at about 88 degrees C. The optimal salt concentration for growth was around 19 g l(-1). The isolate was capable of growth with H2 as the sole electron donor coupled to the reduction of Fe(III) without the need for an organic carbon source. This is the first example of a dissimilatory Fe(III)-reducing micro-organism capable of growing autotrophically on hydrogen. In addition to molecular hydrogen, strain 234T oxidizes pyruvate, acetate, malate, succinate, peptone, formate, fumarate, yeast extract, glycerol, isoleucine, arginine, serine, glutamine, asparagine, stearate, palmitate, valerate, butyrate and propionate with the reduction of Fe(III). This isolate is the first example of a hyperthermophile capable of oxidizing long-chain fatty acids anaerobically. Isolate 234T grew exclusively with Fe(III) as the sole electron acceptor. The G+C content was 58.7 mol\%. Based on detailed analysis of its 16S rDNA sequence, G+C content, distinguishing physiological features and metabolism, strain 234T is proposed to represent a novel genus within the Archaeoglobales. The name proposed for strain 234T is Geoglobus ahangari gen. nov., sp. nov..}, keywords = {Anaerobiosis, Archaea, Bacterial Typing Techniques, DNA, Ribosomal, Electron Transport, Fatty Acids, Ferric Compounds, Hot Temperature, Hydrogen, Molecular Sequence Data, Oxidation-Reduction, Phylogeny, RNA, Ribosomal, 16S, Sequence Analysis, DNA, Water Microbiology}, issn = {1466-5026}, author = {Kashefi, Kazem and Tor, Jason M and Holmes, Dawn E and Gaw Van Praagh, Catherine V and Reysenbach, Anna-Louise and Lovley, Derek R} } @article {580, title = {Harnessing microbially generated power on the seafloor.}, journal = {Nat Biotechnol}, volume = {20}, year = {2002}, month = {2002 Aug}, pages = {821-5}, abstract = {In many marine environments, a voltage gradient exists across the water sediment interface resulting from sedimentary microbial activity. Here we show that a fuel cell consisting of an anode embedded in marine sediment and a cathode in overlying seawater can use this voltage gradient to generate electrical power in situ. Fuel cells of this design generated sustained power in a boat basin carved into a salt marsh near Tuckerton, New Jersey, and in the Yaquina Bay Estuary near Newport, Oregon. Retrieval and analysis of the Tuckerton fuel cell indicates that power generation results from at least two anode reactions: oxidation of sediment sulfide (a by-product of microbial oxidation of sedimentary organic carbon) and oxidation of sedimentary organic carbon catalyzed by microorganisms colonizing the anode. These results demonstrate in real marine environments a new form of power generation that uses an immense, renewable energy reservoir (sedimentary organic carbon) and has near-immediate application.}, keywords = {Bacteria, Bioelectric Energy Sources, Biotechnology, Carbon, Conservation of Energy Resources, DNA, Ribosomal, Electricity, Electrodes, Environmental Microbiology, Geologic Sediments, Molecular Sequence Data, New Jersey, Oceans and Seas, Oregon, Oxidation-Reduction, RNA, Bacterial, RNA, Ribosomal, 16S, Sulfides}, issn = {1087-0156}, doi = {10.1038/nbt716}, author = {Tender, Leonard M and Reimers, Clare E and Stecher, Hilmar A and Holmes, Dawn E and Bond, Daniel R and Lowy, Daniel A and Pilobello, Kanoelani and Fertig, Stephanie J and Lovley, Derek R} } @article {781, title = {Human Stx2-specific monoclonal antibodies prevent systemic complications of Escherichia coli O157:H7 infection.}, journal = {Infect Immun}, volume = {70}, year = {2002}, month = {2002 Feb}, pages = {612-9}, abstract = {Hemolytic-uremic syndrome (HUS) is a serious complication predominantly associated with infection by enterohemorrhagic Escherichia coli (EHEC), such as E. coli O157:H7. EHEC can produce Shiga toxin 1 (Stx1) and/or Shiga toxin 2 (Stx2), both of which are exotoxins comprised of active (A) and binding (B) subunits. In piglets and mice, Stx can induce fatal neurological symptoms. Polyclonal Stx2 antiserum can prevent these effects in piglets infected with the Stx2-producing E. coli O157:H7 strain 86-24. Human monoclonal antibodies (HuMAbs) against Stx2 were developed as potential passive immunotherapeutic reagents for the prevention and/or treatment of HUS. Transgenic mice bearing unrearranged human immunoglobulin (Ig) heavy and kappa light chain loci (HuMAb___Mouse) were immunized with formalin-inactivated Stx2. Thirty-seven stable hybridomas secreting Stx2-specific HuMAbs were isolated: 33 IgG1kappa A-subunit-specific and 3 IgG1kappa and 1 IgG3kappa B-subunit-specific antibodies. Six IgG1kappa A-subunit-specific (1G3, 2F10, 3E9, 4H9, 5A4, and 5C12) and two IgG1kappa B-subunit-specific (5H8 and 6G3) HuMAbs demonstrated neutralization of > 95\% activity of 1 ng of Stx2 in the presence of 0.04 microg of HuMAb in vitro and significant prolongation of survival of mice given 50 microg of HuMAb intraperitoneally (i.p.) and 25 ng of Stx2 intravenously. When administered i.p. to gnotobiotic piglets 6 or 12 h after infection with E. coli O157:H7 strain 86-24, HuMAbs 2F10, 3E9, 5H8, and 5C12 prolonged survival and prevented development of fatal neurological signs and cerebral lesions. The Stx2-neutralizing ability of these HuMAbs could potentially be used clinically to passively protect against HUS development in individuals infected with Stx-producing bacteria, including E. coli O157:H7.}, keywords = {Animals, Antibodies, Bacterial, Antibodies, Monoclonal, Antibody Specificity, Disease Models, Animal, Escherichia coli O157, Female, Germ-Free Life, HeLa Cells, Hemolytic-Uremic Syndrome, Humans, Immunization, Passive, Immunoglobulin G, Immunoglobulin Isotypes, Immunoglobulin kappa-Chains, Mice, Neutralization Tests, Shiga Toxin 2, Swine}, issn = {0019-9567}, author = {Mukherjee, Jean and Chios, Kerry and Fishwild, Dianne and Hudson, Deborah and O{\textquoteright}Donnell, Susan and Rich, Stephen M and Donohue-Rolfe, Arthur and Tzipori, Saul} } @article {589, title = {A hydrogen-based subsurface microbial community dominated by methanogens.}, journal = {Nature}, volume = {415}, year = {2002}, month = {2002 Jan 17}, pages = {312-5}, abstract = {The search for extraterrestrial life may be facilitated if ecosystems can be found on Earth that exist under conditions analogous to those present on other planets or moons. It has been proposed, on the basis of geochemical and thermodynamic considerations, that geologically derived hydrogen might support subsurface microbial communities on Mars and Europa in which methanogens form the base of the ecosystem. Here we describe a unique subsurface microbial community in which hydrogen-consuming, methane-producing Archaea far outnumber the Bacteria. More than 90\% of the 16S ribosomal DNA sequences recovered from hydrothermal waters circulating through deeply buried igneous rocks in Idaho are related to hydrogen-using methanogenic microorganisms. Geochemical characterization indicates that geothermal hydrogen, not organic carbon, is the primary energy source for this methanogen-dominated microbial community. These results demonstrate that hydrogen-based methanogenic communities do occur in Earth{\textquoteright}s subsurface, providing an analogue for possible subsurface microbial ecosystems on other planets.}, keywords = {Bacteria, DNA, Archaeal, DNA, Bacterial, Ecosystem, Euryarchaeota, Exobiology, Hydrogen, Molecular Sequence Data, Phylogeny, RNA, Ribosomal, 16S, Water Microbiology}, issn = {0028-0836}, doi = {10.1038/415312a}, author = {Chapelle, Francis H and O{\textquoteright}Neill, Kathleen and Bradley, Paul M and Meth{\'e}, Barbara A and Ciufo, Stacy A and Knobel, LeRoy L and Lovley, Derek R} } @article {779, title = {Infectivity of a Cryptosporidium parvum isolate of cervine origin for healthy adults and interferon-gamma knockout mice.}, journal = {J Infect Dis}, volume = {185}, year = {2002}, month = {2002 May 1}, pages = {1320-5}, abstract = {The infectivity of a Cryptosporidium parvum isolate of cervine origin (type 2, Moredun) propagated in calves was investigated simultaneously in healthy adult human volunteers and in interferon-gamma knockout (GKO) mice. After exposure to 100-3000 oocysts, 16 volunteers recorded, for a duration of 6 weeks, the number and form of stools that they passed and any symptoms that they experienced. Oocyst excretion was assessed by enzyme-linked immunosorbent assay and direct immunofluorescence assay. Eleven subjects (69\%) became ill, and 8 subjects (50\%) shed oocysts in stool. The median duration of illness was 169 h, and the median number of unformed stools passed was 24. The duration and intensity of symptoms were more severe than were those associated with previously studied isolates. The median infectious dose was estimated to be 300 oocysts for humans and 1 oocyst for the GKO mouse model. The Moredun isolate was more pathogenic than the reference GCH-1 isolate. The GKO mouse model of cryptosporidiosis is useful for discerning isolate-specific differences in pathogenicity.}, keywords = {Adult, Animals, Body Weight, Cryptosporidiosis, Cryptosporidium parvum, Deer, Diarrhea, Disease Susceptibility, Female, Humans, Interferon-gamma, Mice, Mice, Knockout}, issn = {0022-1899}, doi = {10.1086/340132}, author = {Okhuysen, Pablo C and Rich, Stephen M and Chappell, Cynthia L and Grimes, Kevin A and Widmer, Giovanni and Feng, Xiaochuan and Tzipori, Saul} } @article {401, title = {Measuring beta-galactosidase activity in bacteria: cell growth, permeabilization, and enzyme assays in 96-well arrays.}, journal = {Biochem Biophys Res Commun}, volume = {290}, year = {2002}, month = {2002 Jan 11}, pages = {397-402}, abstract = {We describe a high-throughput procedure for measuring beta-galactosidase activity in bacteria. This procedure is unique because all manipulations, including bacterial growth and cell permeabilization, are performed in a 96-well format. Cells are permeabilized by chloroform/SDS treatment directly in the 96-well blocks and then transferred to 96-well microplates for standard colorimetric assay of beta-galactosidase activity as described by Miller [J. H. Miller (1972) Experiments in Molecular Genetics, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY]. Absorbance data are collected with a microplate reader and analyzed using a Microsoft Excel spreadsheet. The beta-galactosidase specific activity values obtained with the high-throughput procedure are identical to those obtained by the traditional single-tube method of Miller. Thus, values obtained with this procedure may be expressed as Miller units and compared directly to Miller units reported in the literature. The 96-well format for permeabilization and assay of enzyme specific activity together with the use of 12-channel and repeater pipettors enables efficient processing of hundreds of samples in an 8-h day.}, keywords = {Bacteria, beta-Galactosidase, Biochemistry, Cell Division, Chloroform, Escherichia coli, Polypropylenes, Sodium Dodecyl Sulfate, Software, Spectrophotometry, Time Factors}, issn = {0006-291X}, doi = {10.1006/bbrc.2001.6152}, author = {Griffith, Kevin L and Wolf, Richard E} } @article {584, title = {Mechanisms for accessing insoluble Fe(III) oxide during dissimilatory Fe(III) reduction by Geothrix fermentans.}, journal = {Appl Environ Microbiol}, volume = {68}, year = {2002}, month = {2002 May}, pages = {2294-9}, abstract = {Mechanisms for Fe(III) oxide reduction were investigated in Geothrix fermentans, a dissimilatory Fe(III)-reducing microorganism found within the Fe(III) reduction zone of subsurface environments. Culture filtrates of G. fermentans stimulated the reduction of poorly crystalline Fe(III) oxide by washed cell suspensions, suggesting that G. fermentans released one or more extracellular compounds that promoted Fe(III) oxide reduction. In order to determine if G. fermentans released electron-shuttling compounds, poorly crystalline Fe(III) oxide was incorporated into microporous alginate beads, which prevented contact between G. fermentans and the Fe(III) oxide. G. fermentans reduced the Fe(III) within the beads, suggesting that one of the compounds that G. fermentans releases is an electron-shuttling compound that can transfer electrons from the cell to Fe(III) oxide that is not in contact with the organism. Analysis of culture filtrates by thin-layer chromatography suggested that the electron shuttle has characteristics similar to those of a water-soluble quinone. Analysis of filtrates by ion chromatography demonstrated that there was as much as 250 microM dissolved Fe(III) in cultures of G. fermentans growing with Fe(III) oxide as the electron acceptor, suggesting that G. fermentans released one or more compounds capable of chelating and solubilizing Fe(III). Solubilizing Fe(III) is another strategy for alleviating the need for contact between cells and Fe(III) oxide for Fe(III) reduction. This is the first demonstration of a microorganism that, in defined medium without added electron shuttles or chelators, can reduce Fe(III) derived from Fe(III) oxide without directly contacting the Fe(III) oxide. These results are in marked contrast to those with Geobacter metallireducens, which does not produce electron shuttles or Fe(III) chelators. These results demonstrate that phylogenetically distinct Fe(III)-reducing microorganisms may use significantly different strategies for Fe(III) reduction. Thus, it is important to know which Fe(III)-reducing microorganisms predominate in a given environment in order to understand the mechanisms for Fe(III) reduction in the environment of interest.}, keywords = {Bacteria, Electrons, Ferric Compounds, Iron Chelating Agents, Oxidation-Reduction, Solubility}, issn = {0099-2240}, author = {Nevin, Kelly P and Lovley, Derek R} } @article {577, title = {Multiple influences of nitrate on uranium solubility during bioremediation of uranium-contaminated subsurface sediments.}, journal = {Environ Microbiol}, volume = {4}, year = {2002}, month = {2002 Sep}, pages = {510-6}, abstract = {Microbiological reduction of soluble U(VI) to insoluble U(IV) has been proposed as a remediation strategy for uranium-contaminated groundwater. Nitrate is a common co-contaminant with uranium. Nitrate inhibited U(VI) reduction in acetate-amended aquifer sediments collected from a uranium-contaminated site in New Mexico. Once nitrate was depleted, both U(VI) and Fe(III) were reduced concurrently. When nitrate was added to sediments in which U(VI) had been reduced, U(VI) reappeared in solution. Parallel studies with the dissimilatory Fe(III)-, U(VI)- and nitrate-reducing microorganism, Geobacter metallireducens, demonstrated that nitrate inhibited reduction of Fe(III) and U(VI) in cell suspensions of cells that had been grown with nitrate as the electron acceptor, but not in Fe(III)-grown cells. Suspensions of nitrate-grown G. metallireducens oxidized Fe(II) and U(IV) with nitrate as the electron acceptor. U(IV) oxidation was accelerated when Fe(II) was also added, presumably due to the Fe(III) being formed abiotically oxidizing U(IV). These studies demonstrate that although the presence of nitrate is not likely to be an impediment to the bioremediation of uranium contamination with microbial U(VI) reduction, it is necessary to reduce nitrate before U(VI) can be reduced. These results also suggest that anaerobic oxidation of U(IV) to U(VI) with nitrate serving as the electron acceptor may provide a novel strategy for solubilizing and extracting microbial U(IV) precipitates from the subsurface.}, keywords = {Ferric Compounds, Ferrous Compounds, Geologic Sediments, Nitrates, Oxidation-Reduction, Proteobacteria, Soil Pollutants, Solubility, Uranium, Water Pollutants, Chemical}, issn = {1462-2912}, author = {Finneran, Kevin T and Housewright, Meghan E and Lovley, Derek R} } @article {388, title = {Multiple mitochondrial DNA polymerases in Trypanosoma brucei.}, journal = {Mol Cell}, volume = {10}, year = {2002}, month = {2002 Jul}, pages = {175-86}, abstract = {Kinetoplast DNA (kDNA), the unusual mitochondrial DNA of Trypanosoma brucei, is a network containing thousands of catenated circles. Database searching for a kDNA replicative polymerase (pol) revealed no mitochondrial pol gamma homolog. Instead, we identified four proteins (TbPOLIA, IB, IC, and ID) related to bacterial pol I. Remarkably, all four localized to the mitochondrion. TbPOLIB and TbPOLIC localized beside the kDNA where replication occurs, and their knockdown by RNA interference caused kDNA network shrinkage. Furthermore, silencing of TbPOLIC caused loss of both minicircles and maxicircles and accumulation of minicircle replication intermediates, consistent with a role in replication. While typical mitochondria contain one DNA polymerase, pol gamma, trypanosome mitochondria contain five such enzymes, including the previously characterized pol beta.}, keywords = {Amino Acid Sequence, Animals, DNA, Kinetoplast, DNA-Directed DNA Polymerase, Mitochondria, Molecular Sequence Data, Phylogeny, Protein Structure, Tertiary, Protein Transport, RNA, Double-Stranded, RNA, Messenger, Sequence Homology, Amino Acid, Trypanosoma brucei brucei}, issn = {1097-2765}, author = {Klingbeil, Michele M and Motyka, Shawn A and Englund, Paul T} } @article {364, title = {Pathology and ultrastructure of Hz-2V infection in the agonadal female corn earworm, Helicoverpa zea.}, journal = {J Invertebr Pathol}, volume = {81}, year = {2002}, month = {2002 Sep}, pages = {33-44}, abstract = {The pathology and ultrastructure of the reproductive tract of Hz-2V-infected female corn earworm moths, Helicoverpa zea, were studied. The identity of malformed reproductive tissues found in virus-infected moths was determined by examining these tissues in moths that were infected with the virus at different life stages. Malformation of reproductive tissues in the progeny of virus-infected female moths was first observed by 3 days post-pupation (dpp), indicating that virus replication had altered the differentiation of these tissues very early on in their development. The ultrastructure of the grossly malformed agonadal reproductive tissues from insects aged 3-10dpp revealed the absence of the cuticular lining found in the oviducts of normal moths, and the proliferation of epithelial cells in these infected oviduct tissues. In addition, large quantities of virus were found aggregated into a large mass in the lumen of the malformed cervix bursa of 10dpp agonadal female pharate adult moths. Prior to eclosion, the virus in the cervix bursa was observed separated into spherical masses, which are thought to exude through the ductus bursa and collect over the vulva, forming a viral "waxy plug" that is likely to play an important role in virus transmission.}, keywords = {Animals, Female, Insect Viruses, Microscopy, Electron, Moths, Virus Diseases}, issn = {0022-2011}, author = {Rallis, Christopher P and Burand, John P} } @article {365, title = {Pathology and ultrastructure of the insect virus, Hz-2V, infecting agonadal male corn earworms, Helicoverpa zea.}, journal = {J Invertebr Pathol}, volume = {80}, year = {2002}, month = {2002 Jun}, pages = {81-9}, abstract = {The pathology of the reproductive tract of Hz-2V-infected agonadal male corn earworm moths, Helicoverpa zea, was studied. The examination of the reproductive tissues of adult agonadal males infected with Hz-2V during different lifestages allowed us to positively correlate the grossly malformed tissues of typical agonadal male moths to the corresponding normal tissues in uninfected males. The reproductive tissues responsible for producing sperm, a pheromonostatic peptide (PSP), and the spermatophore in normal male moths were absent or grossly malformed in the agonadal male moths. Hz-2V was observed replicating in one area of these malformed reproductive tissues in pharate adult males as early as 7 days post-pupation. Interestingly, reproductive tissues essential for initiation of copulation and transfer of reproductive fluids into a female moth during mating appear to be intact and may be functional. These data suggest that agonadal adult males are able to mate with healthy female moths and transfer Hz-2V particles, without fertilizing female moths or altering their sexual receptivity to further mating with other male moths.}, keywords = {Animals, Female, Insect Viruses, Male, Moths, Testis, Virus Diseases}, issn = {0022-2011}, author = {Rallis, Christopher P and Burand, John P} } @article {778, title = {Production and characterization of protective human antibodies against Shiga toxin 1.}, journal = {Infect Immun}, volume = {70}, year = {2002}, month = {2002 Oct}, pages = {5896-9}, abstract = {Hemolytic-uremic syndrome (HUS) is a serious complication which is predominantly associated in children with infection by Shiga toxin-producing Escherichia coli (STEC). By using HuMAb-Mouse (Medarex) animals, human monoclonal antibodies (Hu-MAbs) were developed against Shiga toxin 1 (Stx1) for passive immunotherapy of HUS. Ten stable hybridomas comprised of fully human heavy- and light-chain immunoglobulin elements and secreting Stx1-specific Hu-MAbs (seven immunoglobulin M(kappa)() [IgM(kappa)] elements [one specific for the A subunit and six specific for the B subunit] and three IgG1(kappa) elements specific for subunit B) were isolated. Two IgM(kappa) Hu-MAbs (2D9 and 15G9) and three IgG1(kappa) Hu-MAbs (5A4, 10F4, and 15G2), all specific for subunit B, demonstrated marked neutralization of Stx1 in vitro and significant prolongation of survival in a murine model of Stx1 toxicosis.}, keywords = {Animals, Antibodies, Bacterial, Antibodies, Monoclonal, Child, Escherichia coli, Female, HeLa Cells, Hemolytic-Uremic Syndrome, Humans, Hybridomas, Immunization, Passive, Immunoglobulin G, Immunoglobulin M, Mice, Neutralization Tests, Shiga Toxin 1}, issn = {0019-9567}, author = {Mukherjee, Jean and Chios, Kerry and Fishwild, Dianne and Hudson, Deborah and O{\textquoteright}Donnell, Susan and Rich, Stephen M and Donohue-Rolfe, Arthur and Tzipori, Saul} } @article {685, title = {Protein Explorer: easy yet powerful macromolecular visualization.}, journal = {Trends Biochem Sci}, volume = {27}, year = {2002}, month = {2002 Feb}, pages = {107-9}, abstract = {Protein Explorer (PE, http://www.proteinexplorer.org) enables students, educators and other nonspecialists to visualize macromolecular structures easily. It also offers several advanced capabilities useful to protein structure specialists. Great attention has been given to making PE easy to use. Explanations, color keys and troubleshooting information are displayed automatically. There are also {\textquoteright}Frequently Asked Questions{\textquoteright}, a one-hour {\textquoteright}Quick-Tour{\textquoteright}, an alphabetical {\textquoteright}Help/Index/Glossary{\textquoteright}, and a detailed {\textquoteright}Tutorial{\textquoteright}; all making PE much easier to use than either Chime or RasMol. Moreover, it is much more powerful; in addition to basic macromolecular visualization capabilities common to most similar programs, it offers one-click visualization of interfaces between moieties ({\textquoteright}contacts{\textquoteright}), cation-pi interactions and salt bridges, as well as easy-to-use routines to visualize regions of conservation in three-dimensional protein structures based on multiple sequence alignments.}, keywords = {Computational Biology, DNA, Evolution, Molecular, Macromolecular Substances, Protein Structure, Secondary, Proteins, Sequence Alignment, Software}, issn = {0968-0004}, author = {Martz, Eric} } @article {579, title = {Rapid evolution of redox processes in a petroleum hydrocarbon-contaminated aquifer.}, journal = {Ground Water}, volume = {40}, year = {2002}, month = {2002 Jul-Aug}, pages = {353-60}, abstract = {Ground water chemistry data collected over a six-year period show that the distribution of contaminants and redox processes in a shallow petroleum hydrocarbon-contaminated aquifer has changed rapidly over time. Shortly after a gasoline release occurred in 1990, high concentrations of benzene were present near the contaminant source area. In this contaminated zone, dissolved oxygen in ground water was depleted, and by 1994 Fe(III) reduction and sulfate reduction were the predominant terminal electron accepting processes. Significantly, dissolved methane was below measurable levels in 1994, indicating the absence of significant methanogenesis. By 1996, however, depletion of solid-phase Fe(III)-oxyhydrox ides in aquifer sediments and depletion of dissolved sulfate in ground water resulted in the onset of methanogenesis. Between 1996 and 2000, water-chemistry data indicated that methanogenic metabolism became increasingly prevalent. Molecular analysis of 16S-rDNA extracted from sediments shows the presence of a more diverse methanogenic community inside as opposed to outside the plume core, and is consistent with water-chemistry data indicating a shift toward methanogenesis over time. This rapid evolution of redox processes reflects several factors including the large amounts of contaminants, relatively rapid ground water flow (approximately 0.3 m/day [approximately foot/day]), and low concentrations of microbially reducible Fe(III) oxyhydroxides ( approximately 1 micromol/g) initially present in aquifer sediments. These results illustrate that, under certain hydrologic conditions, redox conditions in petroleum hydrocarbon-contaminated aquifers can change rapidly in time and space, and that the availability of solid-phase Fe(III)-oxyhydroxides affects this rate of change.}, keywords = {Hydrocarbons, Iron, Methane, Oxidation-Reduction, Petroleum, Soil Pollutants, Solubility, Sulfates, Water Pollutants, Water Supply}, issn = {0017-467X}, author = {Chapelle, Francis H and Bradley, Paul M and Lovley, Derek R and O{\textquoteright}Neill, Kathleen and Landmeyer, James E} } @article {585, title = {Reduction of Fe(III) oxide by methanogens in the presence and absence of extracellular quinones.}, journal = {Environ Microbiol}, volume = {4}, year = {2002}, month = {2002 Feb}, pages = {115-24}, abstract = {Five methanogens (Methanosarcina barkeri MS, Methanosphaera cuniculi 1R7, Methanobacterium palustre F, Methanococcus voltaei A3 and Methanolobus vulcani PL-12/M) were investigated for their ability to reduce Fe(III) oxide and the soluble quinone anthraquinone-2,6-disulphonate (AQDS). Two species (M. barkeri and M. voltaei) reduced significant amounts of Fe(III) oxide using hydrogen as the electron donor, and 0.1 mM AQDS greatly accelerated Fe(III) reduction by these organisms. Although Fe(III) appeared to inhibit growth and methanogenesis of some strains, hydrogen partial pressures under donor-limited conditions were much lower (<0.5 Pa) in the presence of Fe(III) than in normal media (1-10 Pa) for all species except for M. vulcani. These results demonstrate that electrons were transferred to Fe(III) by hydrogen-utilizing methanogens even when growth and methanogenesis were inhibited. All species except the obligate methylotroph M. vulcani were able to reduce AQDS when their growth substrates were present as electron donors, and rates were highest when organisms used hydrogen as the electron donor. Purified soil humic acids could also be reduced by the AQDS-reducing methanogens. The ability of methanogens to interact with extracellular quinones, humic acids and Fe(III) oxides raises the possibility that this functional group of organ-isms contributes to Fe(III) and humic acid reduction under certain conditions in the environment and provides an alternative explanation for the inhibition of methanogenesis in some Fe(III)-containing ecosystems.}, keywords = {Anthraquinones, Euryarchaeota, Ferric Compounds, Humic Substances, Oxidation-Reduction, Species Specificity}, issn = {1462-2912}, author = {Bond, Daniel R and Lovley, Derek R} } @article {582, title = {Specific 16S rDNA sequences associated with naphthalene degradation under sulfate-reducing conditions in harbor sediments.}, journal = {Microb Ecol}, volume = {43}, year = {2002}, month = {2002 Jan}, pages = {134-45}, abstract = {Previous studies have demonstrated that naphthalene and other polycyclic aromatic hydrocarbons (PAHs) can be anaerobically oxidized with the reduction of sulfate in PAH-contaminated marine harbor sediments, including those in San Diego Bay. In order to learn more about the microorganisms that might be involved in anaerobic naphthalene degradation, the microorganisms associated with naphthalene degradation in San Diego Bay sediments were evaluated. A dilution-to-extinction enrichment culture strategy, designed to recover the most numerous culturable naphthalene-degrading sulfate reducers, resulted in the enrichment of microorganisms with 16S rDNA sequences in the d-Proteobacteria, which were closely related to a previously described pure culture of a naphthalene-degrading sulfate reducer, NaphS2, isolated from sediments in Germany. A more traditional enrichment culture approach, expected to enrich for the fastest-growing naphthalene-degrading sulfate reducers, yielded 16S rDNA sequences closely related to those found in the dilution-to-extinction enrichments and NaphS2. Analysis of 16S rDNA sequences in sediments from two sites in San Diego Bay that had been adapted for rapid naphthalene degradation by continual amendment with low levels of naphthalene suggested that the microbial community composition in the amended sediments differed from that present in the unamended sediments from the same sites. Most significantly, 6-8\% of the sequences recovered from 100 clones of each of the naphthalene-amended sediments were closely related to the 16S rDNA sequences in the enrichment cultures as well as the sequence of the pure culture, NaphS2. No sequences in this NaphS2 phylotype were recovered from the sediments that were not continually exposed to naphthalene. A PCR primer, which was designed based on these phylotype sequences, was used to amplify additional 16S rDNA sequences belonging to the NaphS2 phylotype from PAH-degrading sediments from Island End River (Boston), MA, and Liepaja Harbor, Latvia. Closely related sequences were also recovered from highly contaminated sediment from Tampa Bay, FL. These results suggest that microorganisms closely related to NaphS2 might be involved in naphthalene degradation in harbor sediments. This finding contrasts with the frequent observation that the environmentally relevant microorganisms cannot be readily recovered in pure culture and suggests that further study of the physiology of NaphS2 may provide insights into factors controlling the rate and extent of naphthalene degradation in marine harbor sediments.}, keywords = {Biodegradation, Environmental, DNA Primers, DNA, Bacterial, Geologic Sediments, Naphthalenes, Oxidation-Reduction, Phylogeny, Polymerase Chain Reaction, RNA, Ribosomal, 16S, Sequence Analysis, DNA, Sulfur-Reducing Bacteria, Water Pollutants, Chemical}, issn = {0095-3628}, doi = {10.1007/s00248-001-1055-z}, author = {Hayes, L A and Lovley, Derek R} } @article {587, title = {Use of Fe(III) as an electron acceptor to recover previously uncultured hyperthermophiles: isolation and characterization of Geothermobacterium ferrireducens gen. nov., sp. nov.}, journal = {Appl Environ Microbiol}, volume = {68}, year = {2002}, month = {2002 Apr}, pages = {1735-42}, abstract = {It has recently been recognized that the ability to use Fe(III) as a terminal electron acceptor is a highly conserved characteristic in hyperthermophilic microorganisms. This suggests that it may be possible to recover as-yet-uncultured hyperthermophiles in pure culture if Fe(III) is used as an electron acceptor. As part of a study of the microbial diversity of the Obsidian Pool area in Yellowstone National Park, Wyo., hot sediment samples were used as the inoculum for enrichment cultures in media containing hydrogen as the sole electron donor and poorly crystalline Fe(III) oxide as the electron acceptor. A pure culture was recovered on solidified, Fe(III) oxide medium. The isolate, designated FW-1a, is a hyperthermophilic anaerobe that grows exclusively by coupling hydrogen oxidation to the reduction of poorly crystalline Fe(III) oxide. Organic carbon is not required for growth. Magnetite is the end product of Fe(III) oxide reduction under the culture conditions evaluated. The cells are rod shaped, about 0.5 microm by 1.0 to 1.2 microm, and motile and have a single flagellum. Strain FW-1a grows at circumneutral pH, at freshwater salinities, and at temperatures of between 65 and 100 degrees C with an optimum of 85 to 90 degrees C. To our knowledge this is the highest temperature optimum of any organism in the Bacteria. Analysis of the 16S ribosomal DNA (rDNA) sequence of strain FW-1a places it within the Bacteria, most closely related to abundant but uncultured microorganisms whose 16S rDNA sequences have been previously recovered from Obsidian Pool and a terrestrial hot spring in Iceland. While previous studies inferred that the uncultured microorganisms with these 16S rDNA sequences were sulfate-reducing organisms, the physiology of the strain FW-1a, which does not reduce sulfate, indicates that these organisms are just as likely to be Fe(III) reducers. These results further demonstrate that Fe(III) may be helpful for recovering as-yet-uncultured microorganisms from hydrothermal environments and illustrate that caution must be used in inferring the physiological characteristics of at least some thermophilic microorganisms solely from 16S rDNA sequences. Based on both its 16S rDNA sequence and physiological characteristics, strain FW-1a represents a new genus among the Bacteria. The name Geothermobacterium ferrireducens gen. nov., sp. nov., is proposed (ATCC BAA-426).}, keywords = {Bacterial Typing Techniques, Culture Media, DNA, Ribosomal, Electron Transport, Ferric Compounds, Fresh Water, Geologic Sediments, Gram-Negative Anaerobic Bacteria, Hot Temperature, Molecular Sequence Data, RNA, Ribosomal, 16S, Sequence Analysis, DNA}, issn = {0099-2240}, author = {Kashefi, Kazem and Holmes, Dawn E and Reysenbach, Anna-Louise and Lovley, Derek R} } @article {603, title = {Acetate oxidation coupled to Fe(iii) reduction in hyperthermophilic microorganisms.}, journal = {Appl Environ Microbiol}, volume = {67}, year = {2001}, month = {2001 Mar}, pages = {1363-5}, abstract = {No hyperthermophilic microorganisms have previously been shown to anaerobically oxidize acetate, the key extracellular intermediate in the anaerobic oxidation of organic matter. Here we report that two hyperthermophiles, Ferroglobus placidus and "Geoglobus ahangari," grow at 85 degrees C by oxidizing acetate to carbon dioxide, with Fe(III) serving as the electron acceptor. These results demonstrate that acetate could potentially be metabolized within the hot microbial ecosystems in which hyperthermophiles predominate, rather than diffusing to cooler environments prior to degradation as has been previously proposed.}, keywords = {Acetates, Anaerobiosis, Archaea, Ferric Compounds, Hot Temperature, Oxidation-Reduction}, issn = {0099-2240}, doi = {10.1128/AEM.67.3.1363-1365.2001}, author = {Tor, J M and Kashefi, K and Lovley, D R} } @article {598, title = {Anaerobic degradation of aromatic compounds coupled to Fe(III) reduction by Ferroglobus placidus.}, journal = {Environ Microbiol}, volume = {3}, year = {2001}, month = {2001 Apr}, pages = {281-7}, abstract = {Aromatic compounds are an important component of the organic matter in some of the anaerobic environments that hyperthermophilic microorganisms inhabit, but the potential for hyperthermophilic microorganisms to metabolize aromatic compounds has not been described previously. In this study, aromatic metabolism was investigated in the hyperthermophile Ferroglobus placidus. F. placidus grew at 85 degrees C in anaerobic medium with a variety of aromatic compounds as the sole electron donor and poorly crystalline Fe(III) oxide as the electron acceptor. Growth coincided with Fe(III) reduction. Aromatic compounds supporting growth included benzoate, phenol, 4-hydroxybenzoate, benzaldehyde, p-hydroxybenzaldehyde and t-cinnamic acid (3-phenyl-2-propenoic acid). These aromatic compounds did not support growth when nitrate was provided as the electron acceptor, even though nitrate supports the growth of this organism with Fe(II) or H2 as the electron donor. The stoichiometry of benzoate and phenol uptake and Fe(III) reduction indicated that F. placidus completely oxidized these aromatic compounds to carbon dioxide, with Fe(III) serving as the sole electron acceptor. This is the first example of an Archaea that can anaerobically oxidize an aromatic compound. These results also demonstrate for the first time that hyperthermophilic microorganisms can anaerobically oxidize aromatic compounds and suggest that hyperthermophiles may metabolize aromatic compounds in hot environments such as the deep hot subsurface and in marine and terrestrial hydrothermal zones in which Fe(III) is available as an electron acceptor.}, keywords = {Anaerobiosis, Archaea, Electron Transport, Ferric Compounds, Hot Temperature, Hydrocarbons, Aromatic, Oxidation-Reduction}, issn = {1462-2912}, author = {Tor, J M and Lovley, D R} } @article {599, title = {Anaerobic degradation of methyl tert-butyl ether (MTBE) and tert-butyl alcohol (TBA).}, journal = {Environ Sci Technol}, volume = {35}, year = {2001}, month = {2001 May 1}, pages = {1785-90}, abstract = {The potential for anaerobic degradation of methyl tert-butyl ether (MTBE) and tert-butyl alcohol (TBA) was investigated in laboratory incubations of sediments from a petroleum-contaminated aquifer and in aquatic sediments. The addition of humic substances (HS) stimulated the anaerobic degradation of MTBE in aquifer sediments in which Fe(III) was available as an electron acceptor. This is attributed to the fact that HS and other extracellular quinones can stimulate the activity of Fe(III)-reducing microorganisms by acting as an electron shuttle between Fe(III)-reducing microorganisms and insoluble Fe(III) oxides. MTBE was not degraded in aquifer sediments without Fe(III) and HS. [14C]-MTBE added to aquatic sediments adapted for anaerobic MTBE degradation was converted to 14CO2 in the presence or absence of HS or the HS analog, anthraquione-2,6-disulfonate. Unamended aquatic sediments produced 14CH4 as well as 14CO2 from [14C]-MTBE. The aquatic sediments also rapidly consumed TBA under anaerobic conditions and converted [14C]-TBA to 14CH4 and 14CO2. An adaptation period of ca. 250-300 days was required prior to the most rapid anaerobic MTBE degradation in both sediment types, whereas TBA was metabolized in the aquatic sediments without a lag. These results demonstrate that, under the appropriate conditions, MTBE and TBA can be degraded in the absence of oxygen. This suggests that it may be possible to design strategies for the anaerobic remediation of MTBE in petroleum-contaminated subsurface environments.}, keywords = {Anaerobiosis, Biodegradation, Environmental, Carbon Dioxide, Geologic Sediments, Methane, Methyl Ethers, tert-Butyl Alcohol, Water Pollutants, Chemical}, issn = {0013-936X}, author = {Finneran, K T and Lovley, D R} } @article {783, title = {Antigen polymorphism in Borrelia hermsii, a clonal pathogenic bacterium.}, journal = {Proc Natl Acad Sci U S A}, volume = {98}, year = {2001}, month = {2001 Dec 18}, pages = {15038-43}, abstract = {The relapsing fever spirochete, Borrelia hermsii, escapes immune selection by alternating expression of surface lipoprotein alleles. The switch results from a duplicative transposition of one of several surface lipoprotein-encoding nucleotide sequences into the singular expression site. These nucleotide sequences constitute a large gene family whose diversity originated, in some cases, before the major divergences of Borrelia species. We have examined the B. hermsii vsp subfamily of alleles, which are carried on linear plasmids within each cell and maintained in several diverse copies as an antigenic archive. Each encodes a distinct serotype-specific protein. We sequenced more than 90\% of the alleles within a single strain-B. hermsii strain HS1. A preponderance of allelic mosaicism suggests that intragenic recombination, coupled with selection imposed by host immune response, has driven diversification of the archived ensemble of vsp alleles. The recombinational diversification of vsp alleles generates change in the associated serotypes of the magnitude (30-40\% amino acid differentiation) necessary for overcoming cross-reactivity of neutralizing antibodies. We conclude that evolution of vsp has occurred by punctuated occurrence of allelic differentiation, rather than by gradual selection of incremental point mutations that do not meet the threshold for antigenic diversity.}, keywords = {Antigens, Bacterial, Base Sequence, Borrelia, DNA, Bacterial, Polymorphism, Genetic, Repetitive Sequences, Nucleic Acid}, issn = {0027-8424}, doi = {10.1073/pnas.071042098}, author = {Rich, S M and Sawyer, S A and Barbour, A G} } @article {597, title = {Application of the 5{\textquoteright} fluorogenic exonuclease assay (TaqMan) for quantitative ribosomal DNA and rRNA analysis in sediments.}, journal = {Appl Environ Microbiol}, volume = {67}, year = {2001}, month = {2001 Jun}, pages = {2781-9}, abstract = {In this study, we report on the development of quantitative PCR and reverse transcriptase PCR assays for the 16S rRNA of Geobacter spp. and identify key issues related to fluorogenic reporter systems for nucleic acid analyses of sediments. The lower detection limit of each assay was 5 to 50 fg of genomic DNA or < or =2 pg of 16S rRNA. TaqMan PCR spectral traces from uncontaminated, amended aquifer sediments were significantly lower (P < 0.0002) than traces for the external standard curve. We also observed a similar, significant decrease in mean quencher emissions for undiluted extracts relative to those for diluted extracts (P < 0.0001). If PCR enumerations were based solely upon the undiluted sample eluant, the TaqMan assay generated an inaccurate result even though the threshold cycle (C(t)) measurements were precise and reproducible in the sediment extracts. Assay accuracy was significantly improved by employing a system of replicate dilutions and replicate analyses for both DNA and rRNA quantitation. Our results clearly demonstrate that fluorescence quenching and autofluorescence can significantly affect TaqMan PCR enumeration accuracy, with subsequent implications for the design and implementation of TaqMan PCR to sediments and related environmental samples.}, keywords = {Deltaproteobacteria, DNA, Ribosomal, Fluorescent Dyes, Geologic Sediments, Polymerase Chain Reaction, Reproducibility of Results, Reverse Transcriptase Polymerase Chain Reaction, RNA, Ribosomal, 16S, Sensitivity and Specificity, Taq Polymerase}, issn = {0099-2240}, doi = {10.1128/AEM.67.6.2781-2789.2001}, author = {Stults, J R and Snoeyenbos-West, O and Methe, B and Lovley, D R and Chandler, D P} } @article {712, title = {Association of caveolin with Chlamydia trachomatis inclusions at early and late stages of infection.}, journal = {Exp Cell Res}, volume = {266}, year = {2001}, month = {2001 Jun 10}, pages = {229-38}, abstract = {The mechanism by which the intracellular bacterial pathogen Chlamydia trachomatis enters eukaryotic cells is poorly understood. There are conflicting reports of entry occurring by clathrin-dependent and clathrin-independent processes. We report here that C. trachomatis serovar K enters HEp-2 and HeLa 229 epithelial cells and J-774A.1 mouse macrophage/monocyte cells via caveolin-containing sphingolipid and cholesterol-enriched raft microdomains in the host cell plasma membranes. First, filipin and nystatin, drugs that specifically disrupt raft function by cholesterol chelation, each impaired entry of C. trachomatis serovar K. In control experiments, filipin did not impair entry of the same organism by an antibody-mediated opsonic process, nor did it impair entry of BSA-coated microspheres. Second, the chlamydia-containing endocytic vesicles specifically reacted with antisera against the caveolae marker protein caveolin. These vesicles are known to become the inclusions in which parasite replication occurs. They avoid fusion with lysosomes and instead traffic to the Golgi region, where they intercept Golgi-derived vesicles that recycle sphingolipids and cholesterol to the plasma membrane. We also report that late-stage C. trachomatis inclusions continue to display high levels of caveolin, which they likely acquire from the exocytic Golgi vesicles. We suggest that the atypical raft-mediated entry process may have important consequences for the host-pathogen interaction well after entry has occurred. These consequences include enabling the chlamydial vesicle to avoid acidification and fusion with lysosomes, to traffic to the Golgi region, and to intercept sphingolipid-containing vesicles from the Golgi.}, keywords = {Animals, Caveolin 1, Caveolins, Cell Line, Chlamydia trachomatis, Epithelium, Filipin, HeLa Cells, Humans, Inclusion Bodies, Macrophages, Membrane Microdomains, Mice, Nystatin, Phagosomes}, issn = {0014-4827}, doi = {10.1006/excr.2001.5202}, author = {Norkin, L C and Wolfrom, S A and Stuart, E S} } @article {591, title = {Bioremediation. Anaerobes to the rescue.}, journal = {Science}, volume = {293}, year = {2001}, month = {2001 Aug 24}, pages = {1444-6}, keywords = {Bacteria, Aerobic, Bacteria, Anaerobic, Biodegradation, Environmental, Euryarchaeota, Ferric Compounds, Genome, Archaeal, Genome, Bacterial, Hydrocarbons, Aromatic, Hydrocarbons, Chlorinated, Metals, Methane, Nitrates, Oxidation-Reduction, Oxygen, Soil Microbiology, Sulfates, Sulfur-Reducing Bacteria, Water Pollutants, Chemical}, issn = {0036-8075}, doi = {10.1126/science.1063294}, author = {Lovley, D R} } @article {713, title = {Caveolae in the uptake and targeting of infectious agents and secreted toxins.}, journal = {Adv Drug Deliv Rev}, volume = {49}, year = {2001}, month = {2001 Jul 28}, pages = {301-15}, abstract = {A variety of microbial pathogens, including viruses, intracellular bacteria, and prions, as well as certain secreted bacterial toxins, can now be added to the list of ligands that enter cells via caveolae or caveolae-like membrane domains. In general, the caveolae-mediated entry pathway results in transport of these microbes and toxins to intracellular destinations that are different from that of cargo entering by other means. As a result, the caveolae-mediated entry pathway can profoundly affect the host cell-pathogen interaction long after entry has occurred. Furthermore, some microbes such as SV40 that enter via cavolae will be valuable as probes to analyze certain poorly understood intracellular trafficking pathways, such as retrograde transport to the ER. Also, viruses that enter via caveolae may have unique potential as gene and drug delivery vectors. In addition, some extracellular microbial pathogens, such as Pneumocystis carinii, may also interact with host cells via caveolae. Finally, caveolae may play a role in host immune defense mechanisms.}, keywords = {Animals, Bacterial Infections, Caveolae, Humans, Toxins, Biological, Virus Diseases}, issn = {0169-409X}, author = {Norkin, L C} } @article {600, title = {Comment on "Abiotic controls on H2 production from basalt-water reactions and implications for aquifer biogeochemistry".}, journal = {Environ Sci Technol}, volume = {35}, year = {2001}, month = {2001 Apr 1}, pages = {1556-9}, keywords = {Hydrogen, Minerals, Organic Chemicals, Oxidation-Reduction, Silicates, Water, Water Microbiology}, issn = {0013-936X}, author = {Anderson, R T and Chapelle, F H and Lovley, D R} } @article {866, title = {Comparative study of the relationship between monomer structure and reactivity for two polyhydroxyalkanoate synthases.}, journal = {Appl Microbiol Biotechnol}, volume = {56}, year = {2001}, month = {2001 Jul}, pages = {131-6}, abstract = {Using organically synthesized hydroxyalkanoate coenzyme A thioesters, the activities of two short-chain polyhydroxalkanoate (PHA) synthases were investigated--Ralstonia eutropha PHA synthase (a type I PHA synthase) and Ectothiorhodospira shaposhnikovii PHA synthase (a type III synthase). The results indicate that the two synthases have similar activities towards most of the monomers tested. 3-Hydroxybutyryl CoA was found to be the most efficient substrate for both synthases. Changes in the side-chain length of the monomers affect monomer reactivity, with shortening of the side-chain length having the more severe effect. Hydrophobicity in the side chain appears to play an important role in the catalytic reaction. The configuration and the position of the hydroxyl group also affect the reactivity of a monomer. Monomers with the [S] configuration can not be recognized by either synthase. Moving the hydroxyl group from the beta carbon to the alpha carbon has a much more severe effect on the reactivity of the monomer than moving the hydroxyl group to the gamma carbon. The results demonstrate that the in vitro system can be used to prepare entirely novel polymers that may not be obtainable from living cells because of metabolic restrictions.}, keywords = {Acyl Coenzyme A, Acyltransferases, Structure-Activity Relationship, Substrate Specificity}, issn = {0175-7598}, author = {Zhang, S and Kamachi, M and Takagi, Y and Lenz, R W and Goodwin, S} } @article {1218, title = {Determination of polydextrose in foods by ion chromatography: collaborative study.}, journal = {J AOAC Int}, volume = {84}, year = {2001}, month = {2001 Mar-Apr}, pages = {472-8}, abstract = {Eight collaborating laboratories assayed 7 blind duplicate pairs of foods for polydextrose content. The 7 test sample pairs ranged from low (2\%) to high (95\%) levels. The following foods were prepared with polydextrose mixed into the other ingredients and then baked, cooked, or otherwise prepared: milk chocolate candy, iced tea, sugar cookie, grape jelly, soft jellied candy, and powdered drink mix. Collaborators received a polydextrose standard to develop a calibration curve. The method determined polydextrose by ion chromatography, after removal of interfering food components (high molecular weight solubles). Repeatability standard deviations (RSDr) ranged from 3.93 to 9.04\%; reproducibility standard deviations (RSDR) ranged from 4.48 to 14.06\%. The average recovery was 94\%.
}, keywords = {Algorithms, Beverages, Cacao, Candy, Chromatography, Ion Exchange, Food Analysis, Glucans, Indicators and Reagents, Reference Standards, Tea, Ultracentrifugation}, issn = {1060-3271}, author = {Craig, S A and Holden, J F and Khaled, M Y} } @article {595, title = {Development of a genetic system for Geobacter sulfurreducens.}, journal = {Appl Environ Microbiol}, volume = {67}, year = {2001}, month = {2001 Jul}, pages = {3180-7}, abstract = {Members of the genus Geobacter are the dominant metal-reducing microorganisms in a variety of anaerobic subsurface environments and have been shown to be involved in the bioremediation of both organic and metal contaminants. To facilitate the study of the physiology of these organisms, a genetic system was developed for Geobacter sulfurreducens. The antibiotic sensitivity of this organism was characterized, and optimal conditions for plating it at high efficiency were established. A protocol for the introduction of foreign DNA into G. sulfurreducens by electroporation was also developed. Two classes of broad-host-range vectors, IncQ and pBBR1, were found to be capable of replication in G. sulfurreducens. In particular, the IncQ plasmid pCD342 was found to be a suitable expression vector for this organism. When the information and novel methods described above were utilized, the nifD gene of G. sulfurreducens was disrupted by the single-step gene replacement method. Insertional mutagenesis of this key gene in the nitrogen fixation pathway impaired the ability of G. sulfurreducens to grow in medium lacking a source of fixed nitrogen. Expression of the nifD gene in trans complemented this phenotype. This paper constitutes the first report of genetic manipulation of a member of the Geobacter genus.}, keywords = {Anti-Bacterial Agents, Bacterial Proteins, Blotting, Southern, Culture Media, Deltaproteobacteria, Electroporation, Fimbriae Proteins, Genetic Complementation Test, Genetic Vectors, Microbial Sensitivity Tests, Mutagenesis, Insertional, Nitrogen Fixation, Plasmids, Transformation, Bacterial}, issn = {0099-2240}, doi = {10.1128/AEM.67.7.3180-3187.2001}, author = {Coppi, M V and Leang, C and Sandler, S J and Lovley, D R} } @article {606, title = {Differences in Fe(III) reduction in the hyperthermophilic archaeon, Pyrobaculum islandicum, versus mesophilic Fe(III)-reducing bacteria.}, journal = {FEMS Microbiol Lett}, volume = {195}, year = {2001}, month = {2001 Feb 20}, pages = {253-8}, abstract = {The discovery that all hyperthermophiles that have been evaluated have the capacity to reduce Fe(III) has raised the question of whether mechanisms for dissimilatory Fe(III) reduction have been conserved throughout microbial evolution. Many studies have suggested that c-type cytochromes are integral components in electron transport to Fe(III) in mesophilic dissimilatory Fe(III)-reducing microorganisms. However, Pyrobaculum islandicum, the hyperthermophile in which Fe(III) reduction has been most intensively studied, did not contain c-type cytochromes. NADPH was a better electron donor for the Fe(III) reductase activity in P. islandicum than NADH. This is the opposite of what has been observed with mesophiles. Thus, if previous models for dissimilatory Fe(III) reduction by mesophilic bacteria are correct, then it is unlikely that a single strategy for electron transport to Fe(III) is present in all dissimilatory Fe(III)-reducing microorganisms.}, keywords = {Cytochrome c Group, Electron Transport, Ferric Compounds, FMN Reductase, NADH, NADPH Oxidoreductases, NADP, Oxidation-Reduction, Temperature, Thermoproteaceae}, issn = {0378-1097}, author = {Childers, S E and Lovley, D R} } @article {1217, title = {Diversity among three novel groups of hyperthermophilic deep-sea Thermococcus species from three sites in the northeastern Pacific Ocean.}, journal = {FEMS Microbiol Ecol}, volume = {36}, year = {2001}, month = {2001 Jun}, pages = {51-60}, abstract = {Eight new strains of deep-sea hyperthermophilic sulfur reducers were isolated from hydrothermal vent fields at 9 degrees 50{\textquoteright}N East Pacific Rise (EPR) and at the Cleft and CoAxial segments along the Juan de Fuca Ridge (JdFR). 16S rRNA gene sequence analysis showed that each strain belongs to the genus Thermococcus. Restriction fragment length polymorphism patterns of the 16S/23S rRNA intergenic spacer region revealed that these isolates fell into three groups: those from the EPR, those from fluid and rock sources on the JdFR, and those isolated from Paralvinella spp. polychaete vent worms from the JdFR. The optimum-temperature specific growth rates and the temperature ranges for growth were significantly higher and broader for those strains isolated from worms relative to those isolated from low-temperature diffuse hydrothermal fluids. Furthermore, the worm-derived isolates generally produced a larger array of proteases and amylases based on zymogram analyses. The zymogram patterns also changed with growth temperature suggesting that these organisms alter their lytic protein suites in response to changes in temperature. This study suggests that there is significant phenotypic diversity in Thermococcus that is not apparent from their highly conserved 16S rRNA nucleotide sequences.
}, issn = {1574-6941}, author = {Holden, J F. and Takai, K and Summit, M and Bolton, S and Zyskowski, J and Baross, J A.} } @article {826, title = {Effects of mutations involving cell division, recombination, and chromosome dimer resolution on a priA2::kan mutant.}, journal = {Proc Natl Acad Sci U S A}, volume = {98}, year = {2001}, month = {2001 Jul 17}, pages = {8203-10}, abstract = {Recombinational repair of replication forks can occur either to a crossover (XO) or noncrossover (non-XO) depending on Holliday junction resolution. Once the fork is repaired by recombination, PriA is important for restarting these forks in Escherichia coli. PriA mutants are Rec(-) and UV sensitive and have poor viability and 10-fold elevated basal levels of SOS expression. PriA sulB mutant cells and their nucleoids were studied by differential interference contrast and fluorescence microscopy of 4{\textquoteright},6-diamidino-2-phenylindole-stained log phase cells. Two populations of cells were seen. Eighty four percent appeared like wild type, and 16\% of the cells were filamented and had poorly partitioned chromosomes (Par(-)). To probe potential mechanisms leading to the two populations of cells, mutations were added to the priA sulB mutant. Mutating sulA or introducing lexA3 decreased, but did not eliminate filamentation or defects in partitioning. Mutating either recA or recB virtually eliminated the Par(-) phenotype. Filamentation in the recB mutant decreased to 3\%, but increased to 28\% in the recA mutant. The ability to resolve and/or branch migrate Holliday junctions also appeared crucial in the priA mutant because removing either recG or ruvC was lethal. Lastly, it was tested whether the ability to resolve chromosome dimers caused by XOs was important in a priA mutant by mutating dif and the C-terminal portion of ftsK. Mutation of dif showed no change in phenotype whereas ftsK1cat was lethal with priA2kan. A model is proposed where the PriA-independent pathway of replication restart functions at forks that have been repaired to non-XOs.}, keywords = {Adenosine Triphosphatases, Bacterial Proteins, Cell Division, Chromosomes, Bacterial, Dimerization, DNA Helicases, Endodeoxyribonucleases, Escherichia coli, Escherichia coli Proteins, Exodeoxyribonuclease V, Exodeoxyribonucleases, Gene Expression, Membrane Proteins, Mutagenesis, Rec A Recombinases, Recombination, Genetic, SOS Response (Genetics)}, issn = {0027-8424}, doi = {10.1073/pnas.121007698}, author = {McCool, J D and Sandler, S J} } @article {601, title = {Geobacter hydrogenophilus, Geobacter chapellei and Geobacter grbiciae, three new, strictly anaerobic, dissimilatory Fe(III)-reducers.}, journal = {Int J Syst Evol Microbiol}, volume = {51}, year = {2001}, month = {2001 Mar}, pages = {581-8}, abstract = {Recent studies on the diversity and ubiquity of Fe(III)-reducing organisms in different environments led to the isolation and identification of four new dissimilatory Fe(III)-reducers (strains H-2T, 172T, TACP-2T and TACP-5). All four isolates are non-motile, Gram-negative, freshwater, mesophilic, strict anaerobes with morphology identical to that of Geobacter metallireducens strain GS-15T. Analysis of the 16S rRNA sequences indicated that the new isolates belong to the genus Geobacter, in the delta-Proteobacteria. Significant differences in phenotypic characteristics, DNA-DNA homology and G+C content indicated that the four isolates represent three new species of the genus. The names Geobacter hydrogenophilus sp. nov. (strain H-2T), Geobacter chapellei sp. nov. (strain 172T) and Geobacter grbiciae sp. nov. (strains TACP-2T and TACP-5) are proposed. Geobacter hydrogenophilus and Geobacter chapellei were isolated from a petroleum-contaminated aquifer and a pristine, deep, subsurface aquifer, respectively. Geobacter grbiciae was isolated from aquatic sediments. All of the isolates can obtain energy for growth by coupling the oxidation of acetate to the reduction of Fe(III). The four isolates also coupled Fe(III) reduction to the oxidation of other simple, volatile fatty acids. In addition, Geobacter hydrogenophilus and Geobacter grbiciae were able to oxidize aromatic compounds such as benzoate, whilst Geobacter grbiciae was also able to use the monoaromatic hydrocarbon toluene.}, keywords = {Base Composition, Deltaproteobacteria, Ferric Compounds, Gram-Negative Anaerobic Straight, Curved, and Helical Rods, Hydrocarbons, Iron, Molecular Sequence Data, Nucleic Acid Hybridization, Oxidation-Reduction, RNA, Ribosomal, 16S}, issn = {1466-5026}, author = {Coates, J D and Bhupathiraju, V K and Achenbach, L A and Mclnerney, M J and Lovley, D R} } @article {1211, title = {Identification of membrane proteins in the hyperthermophilic archaeon pyrococcus furiosus using proteomics and prediction programs.}, journal = {Comp Funct Genomics}, volume = {2}, year = {2001}, month = {2001}, pages = {275-88}, abstract = {Cell-free extracts from the hyperthermophilic archaeon Pyrococcus furiosus were separated into membrane and cytoplasmic fractions and each was analyzed by 2D-gel electrophoresis. A total of 66 proteins were identified, 32 in the membrane fraction and 34 in the cytoplasmic fraction. Six prediction programs were used to predict the subcellular locations of these proteins. Three were based on signal-peptides (SignalP, TargetP, and SOSUISignal) and three on transmembrane-spanning alpha-helices (TSEG, SOSUI, and PRED-TMR2). A consensus of the six programs predicted that 23 of the 32 proteins (72\%) from the membrane fraction should be in the membrane and that all of the proteins from the cytoplasmic fraction should be in the cytoplasm. Two membrane-associated proteins predicted to be cytoplasmic by the programs are also predicted to consist primarily of transmembrane-spanning beta-sheets using porin protein models, suggesting that they are, in fact, membrane components. An ATPase subunit homolog found in the membrane fraction, although predicted to be cytoplasmic, is most likely complexed with other ATPase subunits in the membrane fraction. An additional three proteins predicted to be cytoplasmic but found in the membrane fraction, may be cytoplasmic contaminants. These include a chaperone homolog that may have attached to denatured membrane proteins during cell fractionation. Omitting these three proteins would boost the membrane-protein predictability of the models to near 80\%. A consensus prediction using all six programs for all 2242 ORFs in the P. furiosus genome estimates that 24\% of the ORF products are found in the membrane. However, this is likely to be a minimum value due to the programs{\textquoteright} inability to recognize certain membrane-related proteins, such as subunits associated with membrane complexes and porin-type proteins.
}, issn = {1531-6912}, doi = {10.1002/cfg.110}, author = {Holden, J F and Poole Ii, F L and Tollaksen, S L and Giometti, C S and Lim, H and Yates Iii, J R and Adams, M W} } @article {592, title = {Isolation and characterization of a soluble NADPH-dependent Fe(III) reductase from Geobacter sulfurreducens.}, journal = {J Bacteriol}, volume = {183}, year = {2001}, month = {2001 Aug}, pages = {4468-76}, abstract = {NADPH is an intermediate in the oxidation of organic compounds coupled to Fe(III) reduction in Geobacter species, but Fe(III) reduction with NADPH as the electron donor has not been studied in these organisms. Crude extracts of Geobacter sulfurreducens catalyzed the NADPH-dependent reduction of Fe(III)-nitrilotriacetic acid (NTA). The responsible enzyme, which was recovered in the soluble protein fraction, was purified to apparent homogeneity in a four-step procedure. Its specific activity for Fe(III) reduction was 65 micromol. min(-1). mg(-1). The soluble Fe(III) reductase was specific for NADPH and did not utilize NADH as an electron donor. Although the enzyme reduced several forms of Fe(III), Fe(III)-NTA was the preferred electron acceptor. The protein possessed methyl viologen:NADP(+) oxidoreductase activity and catalyzed the reduction of NADP(+) with reduced methyl viologen as electron donor at a rate of 385 U/mg. The enzyme consisted of two subunits with molecular masses of 87 and 78 kDa and had a native molecular mass of 320 kDa, as determined by gel filtration. The purified enzyme contained 28.9 mol of Fe, 17.4 mol of acid-labile sulfur, and 0.7 mol of flavin adenine dinucleotide per mol of protein. The genes encoding the two subunits were identified in the complete sequence of the G. sulfurreducens genome from the N-terminal amino acid sequences derived from the subunits of the purified protein. The sequences of the two subunits had about 30\% amino acid identity to the respective subunits of the formate dehydrogenase from Moorella thermoacetica, but the soluble Fe(III) reductase did not possess formate dehydrogenase activity. This soluble Fe(III) reductase differs significantly from previously characterized dissimilatory and assimilatory Fe(III) reductases in its molecular composition and cofactor content.}, keywords = {Amino Acid Sequence, Bacterial Proteins, Deltaproteobacteria, Ferric Compounds, FMN Reductase, Molecular Sequence Data, NADH, NADPH Oxidoreductases, NADP, Nitrilotriacetic Acid, Sequence Analysis, Protein, Sequence Homology, Amino Acid, Solubility}, issn = {0021-9193}, doi = {10.1128/JB.183.15.4468-4476.2001}, author = {Kaufmann, F and Lovley, D R} } @article {590, title = {Isolation, characterization and gene sequence analysis of a membrane-associated 89 kDa Fe(III) reducing cytochrome c from Geobacter sulfurreducens.}, journal = {Biochem J}, volume = {359}, year = {2001}, month = {2001 Oct 1}, pages = {147-52}, abstract = {Geobacter sulfurreducens is capable of anaerobic respiration with Fe(III) as a terminal electron acceptor via a membrane-bound Fe(III) reductase activity associated with a large molecular mass cytochrome c. This cytochrome was purified by detergent extraction of the membrane fraction, Q-Sepharose ion-exchange chromatography, preparative electrophoresis, and MonoQ ion-exchange chromatography. Spectrophotometric analysis of the purified cytochrome reveals a c-type haem, with no evidence of haem a, haem b or sirohaem. The cytochrome has an M(r) of 89000 as determined by denaturing PAGE, and has an isoelectric point of 5.2 as determined by analytical isoelectric focusing. Dithionite-reduced cytochrome can donate electrons to Fe(III)-nitrilotriacetic acid and synthetic ferrihydrite, thus demonstrating that the cytochrome has redox and thermodynamic properties required for reduction of Fe(III). Analysis using cyclic voltammetry confirmed that the reduced cytochrome can catalytically transfer electrons to ferrihydrite, further demonstrating its ability to be an electron transport mediator in anaerobic Fe(III) respiration. Sequence analysis of a cloned chromosomal DNA fragment revealed a 2307 bp open reading frame (ferA) encoding a 768 amino acid protein corresponding to the 89 kDa cytochrome. The deduced amino acid sequence (FerA) translated from the open reading frame contained 12 putative haem-binding motifs, as well as a hydrophobic N-terminal membrane anchor sequence, a lipid-attachment site and an ATP/GTP-binding site. FerA displayed 20\% or less identity with amino acid sequences of other known cytochromes, although it does share some features with characterized polyhaem cytochromes c.}, keywords = {Amino Acid Sequence, Bacterial Proteins, Chromatography, Ion Exchange, Cloning, Molecular, Cytochrome c Group, Deltaproteobacteria, DNA Primers, Electron Transport, Electrophoresis, Polyacrylamide Gel, Ferric Compounds, Gene Expression Regulation, Bacterial, Molecular Sequence Data, Molecular Weight, Nitrilotriacetic Acid, Oxidation-Reduction, Polymerase Chain Reaction, Sequence Homology, Amino Acid}, issn = {0264-6021}, author = {Magnuson, T S and Isoyama, N and Hodges-Myerson, A L and Davidson, G and Maroney, M J and Geesey, G G and Lovley, D R} } @article {1219, title = {Key role for sulfur in peptide metabolism and in regulation of three hydrogenases in the hyperthermophilic archaeon Pyrococcus furiosus.}, journal = {J Bacteriol}, volume = {183}, year = {2001}, month = {2001 Jan}, pages = {716-24}, abstract = {The hyperthermophilic archaeon Pyrococcus furiosus grows optimally at 100 degrees C by the fermentation of peptides and carbohydrates. Growth of the organism was examined in media containing either maltose, peptides (hydrolyzed casein), or both as the carbon source(s), each with and without elemental sulfur (S(0)). Growth rates were highest on media containing peptides and S(0), with or without maltose. Growth did not occur on the peptide medium without S(0). S(0) had no effect on growth rates in the maltose medium in the absence of peptides. Phenylacetate production rates (from phenylalanine fermentation) from cells grown in the peptide medium containing S(0) with or without maltose were the same, suggesting that S(0) is required for peptide utilization. The activities of 14 of 21 enzymes involved in or related to the fermentation pathways of P. furiosus were shown to be regulated under the five different growth conditions studied. The presence of S(0) in the growth media resulted in decreases in specific activities of two cytoplasmic hydrogenases (I and II) and of a membrane-bound hydrogenase, each by an order of magnitude. The primary S(0)-reducing enzyme in this organism and the mechanism of the S(0) dependence of peptide metabolism are not known. This study provides the first evidence for a highly regulated fermentation-based metabolism in P. furiosus and a significant regulatory role for elemental sulfur or its metabolites.
}, keywords = {Culture Media, Cytoplasm, Gene Expression Regulation, Archaeal, Gene Expression Regulation, Enzymologic, Glycolysis, Hydrogenase, Membrane Proteins, Oxidation-Reduction, Peptides, Pyrococcus furiosus, Sulfur}, issn = {0021-9193}, doi = {10.1128/JB.183.2.716-724.2001}, author = {Adams, M W and Holden, J F and Menon, A L and Schut, G J and Grunden, A M and Hou, C and Hutchins, A M and Jenney, F E and Kim, C and Ma, K and Pan, G and Roy, R and Sapra, R and Story, S V and Verhagen, M F} } @article {785, title = {Lone star tick-infecting borreliae are most closely related to the agent of bovine borreliosis.}, journal = {J Clin Microbiol}, volume = {39}, year = {2001}, month = {2001 Feb}, pages = {494-7}, abstract = {Although Borrelia theileri, the agent of bovine borreliosis, was described at the turn of the century (in 1903), its relationship with borreliae causing Lyme disease or relapsing fever remains undescribed. We tested the previously published hypothesis that spirochetes infecting Lone Star ticks (Amblyomma americanum) may comprise B. theileri by analyzing the 16S ribosomal DNAs (rDNAs) and flagellin genes of these spirochetes. B. theileri, the Amblyomma agent, and B. miyamotoi formed a natural group or clade distinct from but most closely related to that of the relapsing fever spirochetes. B. theileri and the Amblyomma agent were 97 and 98\% similar at the nucleotide level within the analyzed portions of the 16S rDNA and the flagellin gene respectively, suggesting a recent divergence. The agent of bovine borreliosis might be explored as a surrogate antigen for the as-yet-uncultivatable Amblyomma agent in studies designed to explore the etiology of a Lyme disease-like infection associated with Lone Star ticks.}, keywords = {Animals, Borrelia, Borrelia Infections, Cattle, Cattle Diseases, Flagellin, Humans, Lyme Disease, Phylogeny, Relapsing Fever, RNA, Bacterial, RNA, Ribosomal, 16S, Species Specificity, Tick-Borne Diseases, Ticks}, issn = {0095-1137}, doi = {10.1128/JCM.39.2.494-497.2001}, author = {Rich, S M and Armstrong, P M and Smith, R D and Telford, S R} } @article {596, title = {Microbial detoxification of metals and radionuclides.}, journal = {Curr Opin Biotechnol}, volume = {12}, year = {2001}, month = {2001 Jun}, pages = {248-53}, abstract = {Microorganisms have important roles in the biogeochemical cycling of toxic metals and radionuclides. Recent advances have been made in understanding metal-microbe interactions and new applications of these processes to the detoxification of metal and radionuclide contamination have been developed.}, keywords = {Bacteria, Biodegradation, Environmental, Biotechnology, Environmental Pollution, Genetic Engineering, Geologic Sediments, Metals, Heavy, Radioisotopes}, issn = {0958-1669}, author = {Lloyd, J R and Lovley, D R} } @article {1199, title = {Phosphoenolpyruvate synthetase from the hyperthermophilic archaeon Pyrococcus furiosus.}, journal = {J Bacteriol}, volume = {183}, year = {2001}, month = {2001 Jan}, pages = {709-15}, abstract = {Phosphoenolpyruvate synthetase (PpsA) was purified from the hyperthermophilic archaeon Pyrococcus furiosus. This enzyme catalyzes the conversion of pyruvate and ATP to phosphoenolpyruvate (PEP), AMP, and phosphate and is thought to function in gluconeogenesis. PpsA has a subunit molecular mass of 92 kDa and contains one calcium and one phosphorus atom per subunit. The active form has a molecular mass of 690+/-20 kDa and is assumed to be octomeric, while approximately 30\% of the protein is purified as a large ( approximately 1.6 MDa) complex that is not active. The apparent K(m) values and catalytic efficiencies for the substrates pyruvate and ATP (at 80 degrees C, pH 8.4) were 0.11 mM and 1.43 x 10(4) mM(-1). s(-1) and 0.39 mM and 3.40 x 10(3) mM(-1) x s(-1), respectively. Maximal activity was measured at pH 9.0 (at 80 degrees C) and at 90 degrees C (at pH 8.4). The enzyme also catalyzed the reverse reaction, but the catalytic efficiency with PEP was very low [k(cat)/K(m) = 32 (mM. s(-1)]. In contrast to several other nucleotide-dependent enzymes from P. furiosus, PpsA has an absolute specificity for ATP as the phosphate-donating substrate. This is the first PpsA from a nonmethanogenic archaeon to be biochemically characterized. Its kinetic properties are consistent with a role in gluconeogenesis, although its relatively high cellular concentration ( approximately 5\% of the cytoplasmic protein) suggests an additional function possibly related to energy spilling. It is not known whether interconversion between the smaller, active and larger, inactive forms of the enzyme has any functional role.
}, keywords = {Adenosine Monophosphate, Adenosine Triphosphate, Gluconeogenesis, Hydrogen-Ion Concentration, Phosphates, Phosphoenolpyruvate, Phosphotransferases (Paired Acceptors), Pyrococcus furiosus, Pyruvic Acid, Substrate Specificity}, issn = {0021-9193}, doi = {10.1128/JB.183.2.709-715.2001}, author = {Hutchins, A M and Holden, J F and Adams, M W} } @article {825, title = {PriA mutations that affect PriA-PriC function during replication restart.}, journal = {Mol Microbiol}, volume = {41}, year = {2001}, month = {2001 Aug}, pages = {697-704}, abstract = {In Escherichia coli, repair and restart of collapsed replication forks is thought to be essential for cell growth. The replication restart proteins, PriA, PriB, PriC, DnaB, DnaC, DnaG, DnaT and Rep, form redundant pathways that recognize repaired replication forks and restart them. Recognition, modulation of specific DNA structures and loading of the replicative helicase by the replication restart proteins, is likely to be important for replication restart. It has been hypothesized that PriB and PriC function with PriA in genetically separate and redundant PriA-PriB and PriA-PriC pathways. In this study, the del(priB)302 or priC303:kan mutations were used to isolate the PriA-PriB and PriA-PriC pathways genetically so that the effects of three priA missense mutations, priA300 (K230R), priA301 (C479Y) and priA306 (L557P), on these pathways could be assessed. In a wild-type background, the three priA mutations had little, if any, effect on the phenotypes of UV resistance, basal levels of SOS expression and cell viability. In the priB mutant, priA300 and priA301 caused dramatic negative changes in the three phenotypes listed above (and others), whereas the third priA mutant allele, priA306, showed very little negative effect. In the priC mutant, all three priA mutations behaved similarly, producing little, if any, changes in phenotypes. We conclude that priA300 and priA301 mostly affect the PriA-PriC pathway and do so more than priA306. We suggest that PriA{\textquoteright}s helicase activity is important for the PriA-PriC pathway of replication restart.}, keywords = {Bacterial Proteins, Bacteriophage mu, Cell Division, DNA Replication, DNA-Binding Proteins, Escherichia coli, Escherichia coli Proteins, Genes, Lethal, Genotype, Models, Biological, Mutation, Missense, Phenotype, Replication Protein A, SOS Response (Genetics), Ultraviolet Rays}, issn = {0950-382X}, author = {Sandler, S J and McCool, J D and Do, T T and Johansen, R U} } @article {594, title = {Reductive precipitation of gold by dissimilatory Fe(III)-reducing bacteria and archaea.}, journal = {Appl Environ Microbiol}, volume = {67}, year = {2001}, month = {2001 Jul}, pages = {3275-9}, abstract = {Studies with a diversity of hyperthermophilic and mesophilic dissimilatory Fe(III)-reducing Bacteria and Archaea demonstrated that some of these organisms are capable of precipitating gold by reducing Au(III) to Au(0) with hydrogen as the electron donor. These studies suggest that models for the formation of gold deposits in both hydrothermal and cooler environments should consider the possibility that dissimilatory metal-reducing microorganisms can reductively precipitate gold from solution.}, keywords = {Archaea, Bacteria, Chemical Precipitation, Ferric Compounds, Gold, Oxidation-Reduction}, issn = {0099-2240}, doi = {10.1128/AEM.67.7.3275-3279.2001}, author = {Kashefi, K and Tor, J M and Nevin, K P and Lovley, D R} } @article {389, title = {Replication of kinetoplast DNA: an update for the new millennium.}, journal = {Int J Parasitol}, volume = {31}, year = {2001}, month = {2001 May 1}, pages = {453-8}, abstract = {In this review we will describe the replication of kinetoplast DNA, a subject that our lab has studied for many years. Our knowledge of kinetoplast DNA replication has depended mostly upon the investigation of the biochemical properties and intramitochondrial localisation of replication proteins and enzymes as well as a study of the structure and dynamics of kinetoplast DNA replication intermediates. We will first review the properties of the characterised kinetoplast DNA replication proteins and then describe our current model for kinetoplast DNA replication.}, keywords = {Animals, Crithidia fasciculata, DNA Replication, DNA, Kinetoplast, Forecasting}, issn = {0020-7519}, author = {Morris, J C and Drew, M E and Klingbeil, M M and Motyka, S A and Saxowsky, T T and Wang, Z and Englund, P T} } @article {366, title = {Replication of the gonad-specific virus Hz-2V in Ld652Y cells mimics replication in vivo.}, journal = {J Invertebr Pathol}, volume = {77}, year = {2001}, month = {2001 Jan}, pages = {44-50}, abstract = {A newly discovered, nonoccluded insect virus, known as gonad-specific virus or Hz-2V, was found to replicate differently in two insect cell lines derived from ovarian tissues (Tn-368 cells from Trichoplusia ni and Ld652Y from Lymantria dispar). Differences between these two cell lines were observed in virus plaque forming ability, rate of viral DNA replication, time course of infectious virus production, and the mechanism of virus release from infected cells. Replication of Hz-2V in Ld652Y cells was more productive and more closely resembled in vivo virus replication.}, keywords = {Animals, Baculoviridae, Cell Line, DNA Replication, Lepidoptera, Molecular Mimicry, Time Factors, Virus Cultivation, Virus Replication}, issn = {0022-2011}, doi = {10.1006/jipa.2000.4990}, author = {Lu, H and Burand, J P} } @article {402, title = {Systematic mutagenesis of the DNA binding sites for SoxS in the Escherichia coli zwf and fpr promoters: identifying nucleotides required for DNA binding and transcription activation.}, journal = {Mol Microbiol}, volume = {40}, year = {2001}, month = {2001 Jun}, pages = {1141-54}, abstract = {SoxS is the direct transcriptional activator of at least 15 genes of the Escherichia coli superoxide regulon. SoxS is small (107 amino acids), binds DNA as a monomer and recognizes a highly degenerate DNA binding site, termed {\textquoteright}soxbox{\textquoteright}. Like other members of the AraC/XylS family, SoxS has two putative helix-turn-helix (HTH) DNA-binding motifs, and it has been proposed that each HTH motif recognizes a highly conserved recognition element of the soxbox. To determine which nucleotides are important for SoxS binding, we conducted a systematic mutagenesis of the DNA binding sites for SoxS in the zwf and fpr promoters and determined the effect of the soxbox mutations on SoxS DNA binding and transcription activation in vivo by measuring beta-galactosidase activity in strains with fusions to lacZ. We found that the sequences GCAC and CAAA, termed recognition elements 1 and 2 (RE 1 and RE 2), respectively, are critical for SoxS binding, as mutations within these elements severely hinder or eliminate SoxS-dependent transcription activation; substitutions within RE 2 (CAAA), however, are tolerated better than changes within RE 1 (GCAC). Although substitutions at the seven positions separating the two REs had only a modest effect on SoxS binding, AT basepairs were favoured within this {\textquoteright}spacer{\textquoteright} region, presumably because, by facilitating DNA bending, they help bring the two recognition elements into proper juxtaposition. We also found that the {\textquoteright}invariant A{\textquoteright} present at position 1 of 14/15 functional soxboxes identified thus far is important for SoxS binding, as a change to any other nucleotide at this position reduced SoxS-dependent transcription by approximately 50\%. In addition, positions surrounding the REs seem to show a context effect, in that certain substitutions there have little or no effect when the RE has the optimal binding sequence, but produce a pronounced effect when the RE has a suboptimal sequence. We propose that these nucleotides play an important role in effecting differential expression from the various promoters. Lastly, we used gel retardation assays to show that alterations in transcription activation in vivo are caused by effects on DNA binding. Based on this exhaustive mutagenesis, we propose the following optimal sequence for SoxS binding: AnVGCACWWWnKRHCAAAHn (n = A, C, G, T; V = A, C, G; W = A, T; K = G, T; R = A, G; H = A, C, T).}, keywords = {Amino Acid Sequence, Bacterial Proteins, Base Sequence, Binding Sites, Carrier Proteins, DNA, Bacterial, Electrophoresis, Escherichia coli, Escherichia coli Proteins, Molecular Sequence Data, Mutagenesis, Promoter Regions, Genetic, Trans-Activators, Transcription Factors, Transcriptional Activation}, issn = {0950-382X}, author = {Griffith, K L and Wolf, R E} } @article {602, title = {Trichlorobacter thiogenes should be renamed as a Geobacter species.}, journal = {Appl Environ Microbiol}, volume = {67}, year = {2001}, month = {2001 Feb}, pages = {1020-2}, keywords = {Deltaproteobacteria, DNA, Ribosomal, Phylogeny, RNA, Ribosomal, 16S, Terminology as Topic}, issn = {0099-2240}, author = {Snoeyenbos-West, O and Van Praagh, C G and Lovley, D R} } @article {387, title = {Unlocking the secrets of trypanosome kinetoplast DNA network replication.}, journal = {Protist}, volume = {152}, year = {2001}, month = {2001 Dec}, pages = {255-62}, keywords = {Animals, DNA Replication, DNA, Kinetoplast, Protozoan Proteins, Trypanosoma brucei brucei}, issn = {1434-4610}, author = {Klingbeil, M M and Drew, M E and Liu, Y and Morris, J C and Motyka, S A and Saxowsky, T T and Wang, Z and Englund, P T} } @article {593, title = {Anaerobic benzene degradation.}, journal = {Biodegradation}, volume = {11}, year = {2000}, month = {2000}, pages = {107-16}, abstract = {Although many studies have indicated that benzene persists under anaerobic conditions in petroleum-contaminated environments, it has recently been documented that benzene can be anaerobically oxidized with most commonly considered electron acceptors for anaerobic respiration. These include: Fe(III), sulfate, nitrate, and possibly humic substances. Benzene can also be converted to methane and carbon dioxide under methanogenic conditions. There is evidence that benzene can be degraded under in situ conditions in petroleum-contaminated aquifers in which either Fe(III) reduction or methane production is the predominant terminal electron-accepting process. Furthermore, evidence from laboratory studies suggests that benzene may be anaerobically degraded in petroleum-contaminated marine sediments under sulfate-reducing conditions. Laboratory studies have suggested that within the Fe(III) reduction zone of petroleum-contaminated aquifers, benzene degradation can be stimulated with the addition of synthetic chelators which make Fe(III) more available for microbial reduction. The addition of humic substances and other compounds that contain quinone moieties can also stimulate anaerobic benzene degradation in laboratory incubations of Fe(III)-reducing aquifer sediments by providing an electron shuttle between Fe(III)-reducing microorganisms and insoluble Fe(III) oxides. Anaerobic benzene degradation in aquifer sediments can be stimulated with the addition of sulfate, but in some instances an inoculum of benzene-oxidizing, sulfate-reducing microorganisms must also be added. In a field trial, sulfate addition to the methanogenic zone of a petroleum-contaminated aquifer stimulated the growth and activity of sulfate-reducing microorganisms and enhanced benzene removal. Molecular phylogenetic studies have provided indications of what microorganisms might be involved in anaerobic benzene degradation in aquifers. The major factor limiting further understanding of anaerobic benzene degradation is the lack of a pure culture of an organism capable of anaerobic benzene degradation.}, keywords = {Bacteria, Anaerobic, Benzene, Biodegradation, Environmental, Oxidation-Reduction}, issn = {0923-9820}, author = {Lovley, D R} } @article {789, title = {Animal propagation and genomic survey of a genotype 1 isolate of Cryptosporidium parvum.}, journal = {Mol Biochem Parasitol}, volume = {108}, year = {2000}, month = {2000 May}, pages = {187-97}, abstract = {Human cryptosporidiosis is attributed to two major Cryptosporidium parvum genotypes of which type 1 appears to be the predominant. Most laboratory investigations however are performed using genotype 2 isolates, the only type which readily infects laboratory animals. So far type 1 has only been identified in humans and primates. A type 1 isolate, obtained from an individual with HIV and cryptosporidiosis, was successfully adapted to propagate in gnotobiotic piglets. Genotypic characterization of oocyst DNA from this isolate using multiple restriction fragment length polymorphisms, a genotype-specific PCR marker, and direct sequence analysis of two polymorphic loci confirmed that this isolate, designated NEMC1, is indeed type 1. No changes in the genetic profile were identified during multiple passages in piglets. In contrast, the time period between infection and onset of fecal oocyst shedding, an indicator of adaptation, decreased with increasing number of passages. Consistent with other type 1 isolates, NEMC1 failed to infect mice. A preliminary survey of the NEMC1 genome covering approximately 2\% of the genome and encompassing 200 kb of unique sequence showed an average similarity of approximately 95\% between type 1 and 2 sequences. Twenty-four percent of the NEMC1 sequences were homologous to previously determined genotype 2 C. parvum sequences. To our knowledge, this is the first successful serial propagation of genotype 1 in animals, which should facilitate characterization of the unique features of this human pathogen.}, keywords = {Animals, Cryptosporidiosis, Cryptosporidium parvum, Genome, Protozoan, Genotype, Germ-Free Life, Humans, Mice, Mice, Knockout, Molecular Sequence Data, Polymerase Chain Reaction, Polymorphism, Restriction Fragment Length, Sequence Analysis, DNA, Swine}, issn = {0166-6851}, author = {Widmer, G and Akiyoshi, D and Buckholt, M A and Feng, X and Rich, S M and Deary, K M and Bowman, C A and Xu, P and Wang, Y and Wang, X and Buck, G A and Tzipori, S} } @article {613, title = {Characterization of a membrane-bound NADH-dependent Fe(3+) reductase from the dissimilatory Fe(3+)-reducing bacterium Geobacter sulfurreducens.}, journal = {FEMS Microbiol Lett}, volume = {185}, year = {2000}, month = {2000 Apr 15}, pages = {205-11}, abstract = {Geobacter sulfurreducens produces a single, membrane-associated Fe(3+) reductase activity when grown on fumarate or Fe(3+). The activity was initially isolated by solubilization of membranes with the non-ionic detergent dodecyl-beta-D-maltoside, and partially purified by a combination of ion exchange chromatography and preparative non-denaturing gel electrophoresis. Molecular mass of the reductase, as determined by gel filtration chromatography, was approximately 300 kDa. Cofactor analysis of the purified reductase demonstrates that it contains a hemoprotein and flavin adenine dinucleotide. Kinetic and inhibitor studies show that the reductase is specific for NADH as electron donor, and confirm that the reductase enzymatically reduces Fe(3+). The cytochrome associated with the complex undergoes a reoxidation upon addition of Fe(3+) compounds, indicating an ability to pass reducing equivalents to Fe(3+). This is the first description of a purified NADH-dependent Fe(3+) reductase from a microorganism capable of coupling Fe(3+) reduction to growth.}, keywords = {Cell Membrane, Deltaproteobacteria, Ferric Compounds, FMN Reductase, NADH, NADPH Oxidoreductases, Oxidation-Reduction}, issn = {0378-1097}, author = {Magnuson, T S and Hodges-Myerson, A L and Lovley, D R} } @article {868, title = {Characterization, seasonal occurrence, and diel fluctuation of poly(hydroxyalkanoate) in photosynthetic microbial mats.}, journal = {Appl Environ Microbiol}, volume = {66}, year = {2000}, month = {2000 Oct}, pages = {4279-91}, abstract = {In situ poly(hydroxyalkanoate) (PHA) levels and repeating-unit compositions were examined in stratified photosynthetic microbial mats from Great Sippewissett Salt Marsh, Mass., and Ebro Delta, Spain. Unlike what has been observed in pure cultures of phototrophic bacteria, the prevalence of hydroxyvalerate (HV) repeating units relative to hydroxybutyrate (HB) repeating units was striking. In the cyanobacteria-dominated green material of Sippewissett mats, the mole percent ratio of repeating units was generally 1HB:1HV. In the purple sulfur bacteria-dominated pink material the relationship was typically 1HB:2HV. In Sippewissett mats, PHA contributed about 0.5 to 1\% of the organic carbon in the green layer and up to 6\% in the pink layer. In Ebro Delta mats, PHA of approximately 1HB:2HV-repeating-unit distribution contributed about 2\% of the organic carbon of the composite photosynthetic layers (the green and pink layers were not separated). Great Sippewissett Salt Marsh mats were utilized for more extensive investigation of seasonal, diel, and exogenous carbon effects. When the total PHA content was normalized to organic carbon, there was little seasonal variation in PHA levels. However, routine daily variation was evident at all sites and seasons. In every case, PHA levels increased during the night and decreased during the day. This phenomenon was conspicuous in the pink layer, where PHA levels doubled overnight. The daytime declines could be inhibited by artificial shading. Addition of exogenous acetate, lactate, and propionate induced two- to fivefold increases in the total PHA levels when applied in the daylight but had no effect when applied at night. The distinct diel pattern of in situ PHA accumulation at night appears to be related, in some phototrophs, to routine dark energy metabolism and is not influenced by the availability of organic nutrients.}, keywords = {Alkanes, Biofilms, Hydroxybutyrates, Massachusetts, Photosynthesis, Seasons, Spain, Valerates, Water, Water Microbiology}, issn = {0099-2240}, author = {Rothermich, M M and Guerrero, R and Lenz, R W and Goodwin, S} } @article {45, title = {Combinatorial regulation of phospholipid biosynthetic gene expression by the UME6, SIN3 and RPD3 genes.}, journal = {Nucleic Acids Res}, volume = {28}, year = {2000}, month = {2000 Aug 15}, pages = {3160-7}, abstract = {The Ume6p-Sin3p-Rpd3p complex negatively regulates expression of genes containing a Ume6p binding site. However, these regulatory proteins also function independently to regulate gene expression both negatively and positively. The model system for this combinatorial regulation is the yeast phospholipid biosynthetic pathway. Sin3p negatively regulates the INO1, CHO1, CHO2 and OPI3 genes while Ume6p negatively regulates the INO1 gene and positively regulates the other genes. We have suggested that the positive regulation results from indirect effects on expression of the INO2 transcriptional activator gene. Here, we demonstrate that the effect of Ume6p on INO2 gene expression is also indirect. We also show that Rpd3p is a negative regulator of phospholipid biosynthetic gene expression. The ability of Ume6p, Sin3p and Rpd3p to differentially regulate expression of the phospholipid biosynthetic genes affects phospholipid composition. A sin3 mutant strain lacks detectable levels of phosphatidylethanolamine and elevated levels of phosphatidylcholine (PC) and a rpd3 mutant strain has reduced levels of PC. These alterations in membrane composition suggest that there may exist additional differences in regulation of phospholipid biosynthetic gene expression and that membrane compositions may be coordinated with other biological processes regulated by Ume6p, Sin3p and Rpd3p.}, keywords = {Basic Helix-Loop-Helix Transcription Factors, DNA-Binding Proteins, Fungal Proteins, Gene Expression Regulation, Fungal, Helix-Loop-Helix Motifs, Histone Deacetylases, Kinetics, Phospholipids, Repressor Proteins, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins, Transcription Factors, Transcription, Genetic}, issn = {1362-4962}, author = {Elkhaimi, M and Kaadige, M R and Kamath, D and Jackson, J C and Biliran, H and Lopes, J M} } @article {1198, title = {Determination of polydextrose as dietary fiber in foods.}, journal = {J AOAC Int}, volume = {83}, year = {2000}, month = {2000 Jul-Aug}, pages = {1006-12}, abstract = {Polydextrose (Litesse) provides physiological effects consistent with dietary fiber. However, AOAC methods for measuring total dietary fiber (TDF) in foods include an ethanol precipitation step in which polydextrose and similar carbohydrates are discarded and therefore not quantitated. This study describes a method developed to quantitate polydextrose in foods. The new method includes water extraction, centrifugal ultrafiltration, multienzyme hydrolysis, and anion exchange chromatography with electrochemical detection. Six foods were prepared with 4 levels of polydextrose to test the ruggedness of the method. Internal validation demonstrated the ruggedness of the method with recoveries ranging from 83 to 104\% with an average of 95\% (n = 24) and relative standard deviation of recoveries ranging from 0.7 to 13\% with an average of 3.3\% (n = 24). The value is added to that obtained for dietary fiber content of foods using the AOAC methods, to determine the TDF content of the food.
}, keywords = {Animals, Anions, Bacterial Proteins, Beverages, Cacao, Candy, Chromatography, Ion Exchange, Dietary Fiber, Ethanol, Food Analysis, Glucan 1,4-alpha-Glucosidase, Glucans, Glycoside Hydrolases, Humans, Hydrolysis, Isoamylase, Tea, Ultrafiltration}, issn = {1060-3271}, author = {Craig, S A and Holden, J F and Khaled, M Y and Craig, S A and Holden, J F and Khaled, M Y} } @article {607, title = {Direct and Fe(II)-mediated reduction of technetium by Fe(III)-reducing bacteria.}, journal = {Appl Environ Microbiol}, volume = {66}, year = {2000}, month = {2000 Sep}, pages = {3743-9}, abstract = {The dissimilatory Fe(III)-reducing bacterium Geobacter sulfurreducens reduced and precipitated Tc(VII) by two mechanisms. Washed cell suspensions coupled the oxidation of hydrogen to enzymatic reduction of Tc(VII) to Tc(IV), leading to the precipitation of TcO(2) at the periphery of the cell. An indirect, Fe(II)-mediated mechanism was also identified. Acetate, although not utilized efficiently as an electron donor for direct cell-mediated reduction of technetium, supported the reduction of Fe(III), and the Fe(II) formed was able to transfer electrons abiotically to Tc(VII). Tc(VII) reduction was comparatively inefficient via this indirect mechanism when soluble Fe(III) citrate was supplied to the cultures but was enhanced in the presence of solid Fe(III) oxide. The rate of Tc(VII) reduction was optimal, however, when Fe(III) oxide reduction was stimulated by the addition of the humic analog and electron shuttle anthaquinone-2,6-disulfonate, leading to the rapid formation of the Fe(II)-bearing mineral magnetite. Under these conditions, Tc(VII) was reduced and precipitated abiotically on the nanocrystals of biogenic magnetite as TcO(2) and was removed from solution to concentrations below the limit of detection by scintillation counting. Cultures of Fe(III)-reducing bacteria enriched from radionuclide-contaminated sediment using Fe(III) oxide as an electron acceptor in the presence of 25 microM Tc(VII) contained a single Geobacter sp. detected by 16S ribosomal DNA analysis and were also able to reduce and precipitate the radionuclide via biogenic magnetite. Fe(III) reduction was stimulated in aquifer material, resulting in the formation of Fe(II)-containing minerals that were able to reduce and precipitate Tc(VII). These results suggest that Fe(III)-reducing bacteria may play an important role in immobilizing technetium in sediments via direct and indirect mechanisms.}, keywords = {Culture Media, Deltaproteobacteria, Ferric Compounds, Ferrous Compounds, Fresh Water, Oxidation-Reduction, Technetium}, issn = {0099-2240}, author = {Lloyd, J R and Sole, V A and Van Praagh, C V and Lovley, D R} } @article {610, title = {Enrichment of Geobacter Species in Response to Stimulation of Fe(III) Reduction in Sandy Aquifer Sediments.}, journal = {Microb Ecol}, volume = {39}, year = {2000}, month = {2000 Feb}, pages = {153-167}, abstract = {Engineered stimulation of Fe(III) has been proposed as a strategy to enhance the immobilization of radioactive and toxic metals in metal-contaminated subsurface environments. Therefore, laboratory and field studies were conducted to determine which microbial populations would respond to stimulation of Fe(III) reduction in the sediments of sandy aquifers. In laboratory studies, the addition of either various organic electron donors or electron shuttle compounds stimulated Fe(III) reduction and resulted in Geobacter sequences becoming important constituents of the Bacterial 16S rDNA sequences that could be detected with PCR amplification and denaturing gradient gel electrophoresis (DGGE). Quantification of Geobacteraceae sequences with a PCR most-probable-number technique indicated that the extent to which numbers of Geobacter increased was related to the degree of stimulation of Fe(III) reduction. Geothrix species were also enriched in some instances, but were orders of magnitude less numerous than Geobacter species. Shewanella species were not detected, even when organic compounds known to be electron donors for Shewanella species were used to stimulate Fe(III) reduction in the sediments. Geobacter species were also enriched in two field experiments in which Fe(III) reduction was stimulated with the addition of benzoate or aromatic hydrocarbons. The apparent growth of Geobacter species concurrent with increased Fe(III) reduction suggests that Geobacter species were responsible for much of the Fe(III) reduction in all of the stimulation approaches evaluated in three geographically distinct aquifers. Therefore, strategies for subsurface remediation that involve enhancing the activity of indigenous Fe(III)-reducing populations in aquifers should consider the physiological properties of Geobacter species in their treatment design.}, issn = {1432-184X}, author = {Snoeyenbos-West, O L and Nevin, K P and Anderson, R T and Lovley, D R} } @article {863, title = {Enzymatic hydrolysis of oligomeric models of poly-3-hydroxybutyrate.}, journal = {Biomacromolecules}, volume = {1}, year = {2000}, month = {2000 Winter}, pages = {577-83}, abstract = {The mechanism of the enzymatic degradation of poly([R]-3-hydroxybutyrate) (PHB) was investigated by using well-defined model substrates, including both linear and cyclic [R]-3-hydroxybutyrate (3HB) and [R]-3-hydroxyvalerate (3HV) oligomers, with two different PHB depolymerases. The linear and cyclic oligomers containing from 2 to 10 repeating units were hydrolyzed in solutions of the depolymerase isolated from Aspergillus fumigatus and Alcaligenes faecalis, and the rates of hydrolysis and types of products formed were characterized. Both of the depolymerases catalyzed the hydrolysis of the cyclic oligomers (macrolides) which contained more than three 3HB and 3HV repeating units. The degradation reactions of the linear and cyclic 3HB oligomers with the A. fumigatus depolymerase gave similar ratios of monomer-to-dimer products, but PHB itself formed mostly monomer on hydrolysis, indicating that the enzymatic hydrolysis reactions occurred by different mechanisms for these different types of substrates. The results of this study conclusively show that at least the endo mode of polymer hydrolysis occurs with the two enzymes studied, while the A. fumigatus depolymerase was found to utilize both endo and exo modes of hydrolysis to efficiently degrade PHB and 3HB oligomers.}, keywords = {3-Hydroxybutyric Acid, Alcaligenes, Aspergillus fumigatus, Carboxylic Ester Hydrolases, Escherichia coli, Hydrolysis, Kinetics, Macrolides, Models, Chemical, Polymers}, issn = {1525-7797}, author = {Scherer, T M and Fuller, R C and Goodwin, S and Lenz, R W} } @article {787, title = {Extensive polymorphism in Cryptosporidium parvum identified by multilocus microsatellite analysis.}, journal = {Appl Environ Microbiol}, volume = {66}, year = {2000}, month = {2000 Aug}, pages = {3344-9}, abstract = {Restriction fragment length polymorphism and DNA sequence analysis discern two main types of Cryptosporidium parvum. We present a survey of length polymorphism at several microsatellite loci for type 1 and type 2 isolates. A total of 14 microsatellite loci were identified from C. parvum DNA sequences deposited in public databases. All repeats were mono-, di-, and trinucleotide repeats of A, AT, and AAT, reflecting the high AT content of the C. parvum genome. Several of these loci showed significant length polymorphism, with as many as seven alleles identified for a single locus. Differences between alleles ranged from 1 to 27 bp. Karyotype analysis using probes flanking three microsatellites localized each marker to an individual chromosomal band, suggesting that these markers are single copy. In a sample of 19 isolates for which at least three microsatellites were typed, a majority of isolates displayed a unique multilocus fingerprint. Microsatellite analysis of isolates passaged between different host species identified genotypic changes consistent with changes in parasite populations.}, keywords = {Animals, Base Sequence, Cattle, Cryptosporidiosis, Cryptosporidium parvum, DNA, Protozoan, Humans, Karyotyping, Mice, Mice, Knockout, Microsatellite Repeats, Molecular Sequence Data, Polymerase Chain Reaction, Polymorphism, Genetic, Sequence Analysis, DNA}, issn = {0099-2240}, author = {Feng, X and Rich, S M and Akiyoshi, D and Tumwine, J K and Kekitiinwa, A and Nabukeera, N and Tzipori, S and Widmer, G} } @article {784, title = {Genetic variation and the recent worldwide expansion of Plasmodium falciparum.}, journal = {Gene}, volume = {261}, year = {2000}, month = {2000 Dec 30}, pages = {161-70}, abstract = {Plasmodium falciparum, the agent of human malignant malaria, diverged from Plasmodium reichenowi, the chimpanzee parasite, about the time the human and chimpanzee lineages diverged from each other. The absence of synonymous nucleotide variation at ten loci indicates that the world populations of P. falciparum derive most recently from one single strain, or {\textquoteright}cenancestor,{\textquoteright} which lived a few thousand years ago. Antigenic genes of P. falciparum (such as Csp, Msp-1, and Msp-2) exhibit numerous polymorphisms that have been estimated to be millions of years old. We have discovered in these antigenic genes short repetitive sequences that distort the alignment of alleles and account for the apparent old age of the polymorphisms. The processes of intragenic recombination that generate the repeats occur at rates about 10(-3) to 10(-2), several orders of magnitude greater than the typical mutational process of nucleotide substitutions. We conclude that the antigenic polymorphisms of P. falciparum are consistent with a recent expansion of the world populations of the parasite from a cenancestor that lived in tropical Africa a few thousand years ago.}, keywords = {Animals, Antigens, Protozoan, Base Sequence, Genetic Variation, Merozoite Surface Protein 1, Molecular Sequence Data, Phylogeny, Plasmodium, Plasmodium falciparum, Polymorphism, Genetic, Protozoan Proteins, Repetitive Sequences, Nucleic Acid, RNA, Ribosomal, Sequence Homology, Nucleic Acid, Species Specificity}, issn = {0378-1119}, author = {Ayala, F J and Rich, S M} } @article {612, title = {Hexadecane decay by methanogenesis.}, journal = {Nature}, volume = {404}, year = {2000}, month = {2000 Apr 13}, pages = {722-3}, keywords = {Alkanes, Anaerobiosis, Bacteria, Biodegradation, Environmental, Geologic Sediments, Methane, Petroleum}, issn = {0028-0836}, doi = {10.1038/35008145}, author = {Anderson, R T and Lovley, D R} } @article {828, title = {The importance of repairing stalled replication forks.}, journal = {Nature}, volume = {404}, year = {2000}, month = {2000 Mar 2}, pages = {37-41}, abstract = {The bacterial SOS response to unusual levels of DNA damage has been recognized and studied for several decades. Pathways for re-establishing inactivated replication forks under normal growth conditions have received far less attention. In bacteria growing aerobically in the absence of SOS-inducing conditions, many replication forks encounter DNA damage, leading to inactivation. The pathways for fork reactivation involve the homologous recombination systems, are nonmutagenic, and integrate almost every aspect of DNA metabolism. On a frequency-of-use basis, these pathways represent the main function of bacterial DNA recombination systems, as well as the main function of a number of other enzymatic systems that are associated with replication and site-specific recombination.}, keywords = {Bacteria, Bacterial Proteins, Chromosomes, Bacterial, DNA Replication, DNA, Bacterial, Escherichia coli, Recombination, Genetic, Replication Origin, SOS Response (Genetics)}, issn = {0028-0836}, doi = {10.1038/35003501}, author = {Cox, M M and Goodman, M F and Kreuzer, K N and Sherratt, D J and Sandler, S J and Marians, K J} } @article {864, title = {In vitro polymerization and copolymerization of 3-hydroxypropionyl-CoA with the PHB synthase from Ralstonia eutropha.}, journal = {Biomacromolecules}, volume = {1}, year = {2000}, month = {2000 Fall}, pages = {433-9}, abstract = {The poly(3-hydroxybutyrate) (PHB) synthase of Ralstonia eutropha, which was produced by a recombinant strain of Escherichia coli and purified in one step with a methyl-HIC column to a purity of more than 90\%, was used to polymerize 3-hydroxypropionyl-CoA (3HPCoA) and to copolymerize 3HPCoA with 3-hydroxybutyryl-CoA (3HBCoA). A Km of 189 microM and a kcat of 10 s-1 were determined for the activity of the enzyme in the polymerization reaction of 3HPCoA based on the assumption that the dimer form of PHB synthase was the active form. Free coenzyme A was found to be a very effective competitive inhibitor for the polymerization of 3HPCoA with a Ki of 85 microM. The maximum degree of conversion of 3HPCoA to polymer was less than 40\%. In the simultaneous copolymerization reactions of these two monomers, both the turnover number for the copolymerization reaction and the maximum degree of conversion of 3HPCoA and 3HBCoA to copolymers increased with an increase in the amount of 3HBCoA in the monomer mixture. However, the maximum conversion of 3HPCoA to copolymer was always less than 35\%, regardless of the ratio of 3HPCoA to 3HBCoA. Block copolymers were obtained by the sequential copolymerization of the two monomers and these copolymers had a much narrower molecular weight distribution than those obtained by the simultaneous copolymerization for the same molar ratio of 3HPCoA to 3HBCoA.}, keywords = {Acyltransferases, Coenzyme A, Escherichia coli, Kinetics, Magnetic Resonance Spectroscopy, Molecular Weight}, issn = {1525-7797}, author = {Song, J J and Zhang, S and Lenz, R W and Goodwin, S} } @article {865, title = {Kinetic and mechanistic characterization of the polyhydroxybutyrate synthase from Ralstonia eutropha.}, journal = {Biomacromolecules}, volume = {1}, year = {2000}, month = {2000 Summer}, pages = {244-51}, abstract = {Purified Ralstonia eutropha polyhydroxybutyrate (PHB) synthase from recombinant cells can exist as monomer and dimer. The polymerization reaction catalyzed by this enzyme displays a lag phase, which causes difficulties for kinetic and mechanistic characterization of the enzymatic polymerization reaction. In this study, we developed a method to eliminate the lag phase of PHB synthase by physical means, i.e., adding multihydroxyl compounds to the enzyme solution. This method allows us to recognize the nature of the lag phase as a physical rather than a chemical process. With such lag-phase-free-enzyme, the kinetic properties of the enzyme were investigated. The results indicate that 3-hydroxybutyryl-CoA (3HBCoA) is the optimal substrate for the enzyme. A slower catalytic rate and lower binding ability account for a lower reactivity of 3-hydroxyvaleryl-CoA (3HVCoA) compared to that of 3HBCoA. The change of hydroxyl group from the beta to the gamma position causes dramatic decreases in the binding ability of 4-hydroxybutyryl-CoA (4HBCoA). By using a dilution strategy and size exclusion chromatographic technique, the active form of the enzyme was identified to be the dimeric form. The number of catalytic sites in the dimeric form of the enzyme was examined by comparing the molecular weight of polyhydroxybutyrate as a function of substrate-to-enzyme ratio. The results suggest that the dimeric enzyme has only one catalytic site. A revised model of polymerization reaction catalyzed by R. eutropha PHB synthase is described.}, keywords = {Acyltransferases, Catalysis, Chromatography, Gel, Coenzyme A, Culture Media, Hydrogen-Ion Concentration, Kinetics, Molecular Weight, Proteobacteria}, issn = {1525-7797}, author = {Zhang, S and Yasuo, T and Lenz, R W and Goodwin, S} } @article {611, title = {Lack of production of electron-shuttling compounds or solubilization of Fe(III) during reduction of insoluble Fe(III) oxide by Geobacter metallireducens.}, journal = {Appl Environ Microbiol}, volume = {66}, year = {2000}, month = {2000 May}, pages = {2248-51}, abstract = {Studies with the dissimilatory Fe(III)-reducing microorganism Geobacter metallireducens demonstrated that the common technique of separating Fe(III)-reducing microorganisms and Fe(III) oxides with semipermeable membranes in order to determine whether the Fe(III) reducers release electron-shuttling compounds and/or Fe(III) chelators is invalid. This raised doubts about the mechanisms for Fe(III) oxide reduction by this organism. However, several experimental approaches indicated that G. metallireducens does not release electron-shuttling compounds and does not significantly solubilize Fe(III) during Fe(III) oxide reduction. These results suggest that G. metallireducens directly reduces insoluble Fe(III) oxide.}, keywords = {Deltaproteobacteria, Electron Transport, Ferric Compounds, Iron, Kinetics, Oxidation-Reduction, Solubility}, issn = {0099-2240}, author = {Nevin, K P and Lovley, D R} } @article {867, title = {Methane production and release from two New England peatlands.}, journal = {Int Microbiol}, volume = {3}, year = {2000}, month = {2000 Jun}, pages = {89-95}, abstract = {The rate of methane production and release to the atmosphere was determined for two New England peat bogs. Methane production rates from peat sediments, which were measured down to depths of 150 cm, ranged from 1 to 15 micromoles per liter per hour. The highest rates of methane production occurred at depths of 60-100 cm. Methane release from these same peats was quantified from various habitats on the bog using gas collection chambers. The chambers enclose a two-liter volume and cover an area of 0.02 m2. Methane accumulation in the chambers was measured for periods of up to 18 days. Methane release was related to pH and habitat zone. The lowest rates of methane release were from those portions of the bogs that had pH values below 5.0. Peak methane release occurred during or immediately after ice melt in both wetlands with release rates as high as 34 mmoles/m2/d. The overall estimate of yearly release of methane from these bog systems is 2,900 and 14,900 moles per year for Arcadia and Hawley Bogs respectively. Both of these bogs have pH environments close to the lowest limit for methanogenesis, and small differences in pH values can have a large impact on both the rate of methane production and the rate of methane release to the atmosphere.}, keywords = {Anaerobiosis, Euryarchaeota, Hydrogen-Ion Concentration, Methane, New England, Soil, Soil Microbiology}, issn = {1139-6709}, author = {Duval, B and Goodwin, S} } @article {609, title = {Microbes with a mettle for bioremediation.}, journal = {Nat Biotechnol}, volume = {18}, year = {2000}, month = {2000 Jun}, pages = {600-1}, keywords = {Bacterial Adhesion, Cadmium, Cupriavidus necator, Industrial Waste, Metallothionein, Metals, Heavy, Soil Microbiology, Soil Pollutants}, issn = {1087-0156}, doi = {10.1038/76433}, author = {Lovley, D R and Lloyd, J R} } @article {827, title = {Multiple genetic pathways for restarting DNA replication forks in Escherichia coli K-12.}, journal = {Genetics}, volume = {155}, year = {2000}, month = {2000 Jun}, pages = {487-97}, abstract = {In Escherichia coli, the primosome assembly proteins, PriA, PriB, PriC, DnaT, DnaC, DnaB, and DnaG, are thought to help to restart DNA replication forks at recombinational intermediates. Redundant functions between priB and priC and synthetic lethality between priA2::kan and rep3 mutations raise the possibility that there may be multiple pathways for restarting replication forks in vivo. Herein, it is shown that priA2::kan causes synthetic lethality when placed in combination with either Deltarep::kan or priC303:kan. These determinations were made using a nonselective P1 transduction-based viability assay. Two different priA2::kan suppressors (both dnaC alleles) were tested for their ability to rescue the priA-priC and priA-rep double mutant lethality. Only dnaC809,820 (and not dnaC809) could rescue the lethality in each case. Additionally, it was shown that the absence of the 3{\textquoteright}-5{\textquoteright} helicase activity of both PriA and Rep is not the critical missing function that causes the synthetic lethality in the rep-priA double mutant. One model proposes that replication restart at recombinational intermediates occurs by both PriA-dependent and PriA-independent pathways. The PriA-dependent pathways require at least priA and priB or priC, and the PriA-independent pathway requires at least priC and rep. It is further hypothesized that the dnaC809 suppression of priA2::kan requires priC and rep, whereas dnaC809,820 suppression of priA2::kan does not.}, keywords = {Base Sequence, DNA Primers, DNA Replication, Escherichia coli, Genes, Lethal, Mutation, Polymerase Chain Reaction}, issn = {0016-6731}, author = {Sandler, S J} } @article {604, title = {N2-dependent growth and nitrogenase activity in the metal-metabolizing bacteria, Geobacter and Magnetospirillum species.}, journal = {Environ Microbiol}, volume = {2}, year = {2000}, month = {2000 Jun}, pages = {266-73}, abstract = {Cells of Geobacter metallireducens, Magnetospirillum strain AMB-1, Magnetospirillum magnetotacticum and Magnetospirillum gryphiswaldense showed N2-dependent growth, the first anaerobically with Fe(III) as the electron acceptor, and the latter three species microaerobically in semi-solid oxygen gradient cultures. Cells of the Magnetospirillum species grown with N2 under microaerobic conditions were magnetotactic and therefore produced magnetosomes. Cells of Geobacter metallireducens reduced acetylene to ethylene (11.5+/-5.9 nmol C2H4 produced min(-1) mg(-1) cell protein) while growing with Fe(III) as the electron acceptor in anaerobic growth medium lacking a fixed nitrogen source. Cells of the Magnetospirillum species, grown in a semi-solid oxygen gradient medium, also reduced acetylene at comparable rates. Uncut chromosomal and fragments from endonuclease-digested chromosomal DNA from these species, as well as Geobacter sulphurreducens organisms, hybridized with a nifHDK probe from Rhodospirillum rubrum, indicating the presence of these nitrogenase structural genes in these organisms. The evidence presented here shows that members of the metal-metabolizing genera, Geobacter and Magnetospirillum, fix atmospheric dinitrogen.}, keywords = {Acetylene, DNA, Bacterial, Ethylenes, Genes, Bacterial, Iron, Nitrogen Fixation, Nitrogenase, Oxidation-Reduction, Oxidoreductases, Proteobacteria, Rhodospirillaceae}, issn = {1462-2912}, author = {Bazylinski, D A and Dean, A J and Sch{\"u}ler, D and Phillips, E J and Lovley, D R} } @article {44, title = {A network of yeast basic helix-loop-helix interactions.}, journal = {Nucleic Acids Res}, volume = {28}, year = {2000}, month = {2000 Nov 15}, pages = {4460-6}, abstract = {The Ino4 protein belongs to the basic helix-loop-helix (bHLH) family of proteins. It is known to form a dimer with Ino2p, which regulates phospholipid biosynthetic genes. Mammalian bHLH proteins have been shown to form multiple dimer combinations. However, this flexibility in dimerization had not been documented for yeast bHLH proteins. Using the yeast two-hybrid assay and a biochemical assay we show that Ino4p dimerizes with the Pho4p, Rtg1p, Rtg3p and Sgc1p bHLH proteins. Screening a yeast cDNA library identified three additional proteins that interact with Ino4p: Bck2p, YLR422W and YNR064C. The interaction with Bck2p prompted us to examine if any of the Bck2p-associated functions affect expression of phospholipid biosynthetic genes. We found that hyperosmotic growth conditions altered the growth phase regulation of a phospholipid biosynthetic gene, CHO1. There are two recent reports of initial whole genome yeast two-hybrid interactions. Interestingly, one of these reports identified five proteins that interact with Ino4p: Ino2p, Hcs1p, Apl2p, YMR317W and YNL279W. Ino2p is the only protein in common with the data presented here. Our finding that Ino4p interacts with five bHLH proteins suggests that Ino4p is likely to be a central player in the coordination of multiple biological processes.}, keywords = {DNA-Binding Proteins, Fungal Proteins, Gene Expression Regulation, Fungal, Helix-Loop-Helix Motifs, Lac Operon, Methyltransferases, Phosphatidylethanolamine N-Methyltransferase, Plasmids, Protein Binding, Recombinant Fusion Proteins, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins, Trans-Activators, Transcription Factors, Transcriptional Activation, Two-Hybrid System Techniques}, issn = {1362-4962}, author = {Robinson, K A and Koepke, J I and Kharodawala, M and Lopes, J M} } @article {608, title = {Novel forms of anaerobic respiration of environmental relevance.}, journal = {Curr Opin Microbiol}, volume = {3}, year = {2000}, month = {2000 Jun}, pages = {252-6}, abstract = {Novel forms of anaerobic respiration continue to be discovered. Many of these are environmentally significant as they have important impacts on the fate of organic carbon and the cycling of many inorganic compounds. Furthermore, anaerobic respiration is becoming increasing recognized as a strategy for the remediation of organic and metal contaminants in the subsurface.}, keywords = {Anaerobiosis, Bacteria, Anaerobic, Environment, Industrial Microbiology, Industrial Waste}, issn = {1369-5274}, author = {Lovley, D R and Coates, J D} } @article {786, title = {The origin of antigenic diversity in Plasmodium falciparum.}, journal = {Parasitol Today}, volume = {16}, year = {2000}, month = {2000 Sep}, pages = {390-6}, abstract = {Most studies of genetic variability of Plasmodium falciparum have focused on protein antigens and the genes that encode them. The consensus is that populations exhibit high levels of genetic polymorphism, most notably the genes encoding surface proteins of the merozoite (Msp1, Msp2) and the sporozoite (Csp). The age and derivation of this variation is a subject that warrants further careful consideration, as discussed here by Stephen Rich, Marcelo Ferreira and Francisco Ayala.}, keywords = {Amino Acid Sequence, Animals, Antigens, Protozoan, Base Sequence, Genes, Protozoan, Genetic Variation, Merozoite Surface Protein 1, Molecular Sequence Data, Plasmodium falciparum, Protozoan Proteins, Sequence Homology}, issn = {0169-4758}, author = {Rich, S M and Ferreira, M U and Ayala, F J} } @article {788, title = {Population structure and recent evolution of Plasmodium falciparum.}, journal = {Proc Natl Acad Sci U S A}, volume = {97}, year = {2000}, month = {2000 Jun 20}, pages = {6994-7001}, abstract = {Plasmodium falciparum is the agent of malignant malaria, one of mankind{\textquoteright}s most severe maladies. The parasite exhibits antigenic polymorphisms that have been postulated to be ancient. We have proposed that the extant world populations of P. falciparum have derived from one single parasite, a cenancestor, within the last 5, 000-50,000 years. This inference derives from the virtual or complete absence of synonymous nucleotide polymorphisms at genes not involved in immune or drug responses. Seeking to conciliate this claim with extensive antigenic polymorphism, we first note that allele substitutions or polymorphisms can arise very rapidly, even in a single generation, in large populations subject to strong natural selection. Second, new alleles can arise not only by single-nucleotide mutations, but also by duplication/deletion of short simple-repeat DNA sequences, a process several orders of magnitude faster than single-nucleotide mutation. We analyze three antigenic genes known to be extremely polymorphic: Csp, Msp-1, and Msp-2. We identify regions consisting of tandem or proximally repetitive short DNA sequences, including some previously unnoticed. We conclude that the antigenic polymorphisms are consistent with the recent origin of the world populations of P. falciparum inferred from the analysis of nonantigenic genes.}, keywords = {Amino Acid Sequence, Animals, Base Sequence, Biological Evolution, Genes, Protozoan, Genetics, Population, Molecular Sequence Data, Plasmodium falciparum}, issn = {0027-8424}, author = {Rich, S M and Ayala, F J} } @article {46, title = {The promoter of the yeast INO4 regulatory gene: a model of the simplest yeast promoter.}, journal = {J Bacteriol}, volume = {182}, year = {2000}, month = {2000 May}, pages = {2746-52}, abstract = {In Saccharomyces cerevisiae, the phospholipid biosynthetic genes are transcriptionally regulated in response to inositol and choline. This regulation requires the transcriptional activator proteins Ino4p and Ino2p, which form a heterodimer that binds to the UAS(INO) element. We have previously shown that the promoters of the INO4 and INO2 genes are among the weakest promoters characterized in yeast. Because little is known about the promoters of weakly expressed yeast genes, we report here the analysis of the constitutive INO4 promoter. Promoter deletion constructs scanning 1,000 bp upstream of the INO4 gene identified a small region (-58 to -46) that is absolutely required for expression. S1 nuclease mapping shows that this region contains the transcription start sites for the INO4 gene. An additional element (-114 to -86) modestly enhances INO4 promoter activity (fivefold). Thus, the region required for INO4 transcription is limited to 68 bp. These studies also found that INO4 gene expression is not autoregulated by Ino2p and Ino4p, despite the presence of a putative UAS(INO) element in the INO4 promoter. We further report that the INO4 steady-state transcript levels and Ino4p levels are regulated twofold in response to inositol and choline, suggesting a posttranscriptional mechanism of regulation.}, keywords = {Basic Helix-Loop-Helix Transcription Factors, DNA-Binding Proteins, Fungal Proteins, Gene Expression Regulation, Fungal, Promoter Regions, Genetic, Repressor Proteins, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins, Trans-Activators, Transcription Factors}, issn = {0021-9193}, author = {Robinson, K A and Lopes, J M} } @article {614, title = {Reduction of Fe(III), Mn(IV), and toxic metals at 100 degrees C by Pyrobaculum islandicum.}, journal = {Appl Environ Microbiol}, volume = {66}, year = {2000}, month = {2000 Mar}, pages = {1050-6}, abstract = {It has recently been noted that a diversity of hyperthermophilic microorganisms have the ability to reduce Fe(III) with hydrogen as the electron donor, but the reduction of Fe(III) or other metals by these organisms has not been previously examined in detail. When Pyrobaculum islandicum was grown at 100 degrees C in a medium with hydrogen as the electron donor and Fe(III)-citrate as the electron acceptor, the increase in cell numbers of P. islandicum per mole of Fe(III) reduced was found to be ca. 10-fold higher than previously reported. Poorly crystalline Fe(III) oxide could also serve as the electron acceptor for growth on hydrogen. The stoichiometry of hydrogen uptake and Fe(III) oxide reduction was consistent with the oxidation of 1 mol of hydrogen resulting in the reduction of 2 mol of Fe(III). The poorly crystalline Fe(III) oxide was reduced to extracellular magnetite. P. islandicum could not effectively reduce the crystalline Fe(III) oxide minerals goethite and hematite. In addition to using hydrogen as an electron donor for Fe(III) reduction, P. islandicum grew via Fe(III) reduction in media in which peptone and yeast extract served as potential electron donors. The closely related species P. aerophilum grew via Fe(III) reduction in a similar complex medium. Cell suspensions of P. islandicum reduced the following metals with hydrogen as the electron donor: U(VI), Tc(VII), Cr(VI), Co(III), and Mn(IV). The reduction of these metals was dependent upon the presence of cells and hydrogen. The metalloids arsenate and selenate were not reduced. U(VI) was reduced to the insoluble U(IV) mineral uraninite, which was extracellular. Tc(VII) was reduced to insoluble Tc(IV) or Tc(V). Cr(VI) was reduced to the less toxic, less soluble Cr(III). Co(III) was reduced to Co(II). Mn(IV) was reduced to Mn(II) with the formation of manganese carbonate. These results demonstrate that biological reduction may contribute to the speciation of metals in hydrothermal environments and could account for such phenomena as magnetite accumulation and the formation of uranium deposits at ca. 100 degrees C. Reduction of toxic metals with hyperthermophilic microorganisms or their enzymes might be applied to the remediation of metal-contaminated waters or waste streams.}, keywords = {Chromium, Cobalt, Energy Metabolism, Ferric Compounds, Geological Phenomena, Geology, Iron, Manganese, Metals, Heavy, Oxidation-Reduction, Technetium, Thermoproteaceae, Uranium, Water Pollutants, Chemical}, issn = {0099-2240}, author = {Kashefi, K and Lovley, D R} } @article {829, title = {Role of PriA in replication fork reactivation in Escherichia coli.}, journal = {J Bacteriol}, volume = {182}, year = {2000}, month = {2000 Jan}, pages = {9-13}, keywords = {Bacterial Proteins, DNA Primase, DNA Repair, DNA Replication, DNA, Bacterial, DNA-Binding Proteins, Escherichia coli, Escherichia coli Proteins, Models, Genetic, Replication Protein A}, issn = {0021-9193}, author = {Sandler, S J and Marians, K J} } @article {832, title = {Diversity of radA genes from cultured and uncultured archaea: comparative analysis of putative RadA proteins and their use as a phylogenetic marker.}, journal = {J Bacteriol}, volume = {181}, year = {1999}, month = {1999 Feb}, pages = {907-15}, abstract = {Archaea-specific radA primers were used with PCR to amplify fragments of radA genes from 11 cultivated archaeal species and one marine sponge tissue sample that contained essentially an archaeal monoculture. The amino acid sequences encoded by the PCR fragments, three RadA protein sequences previously published (21), and two new complete RadA sequences were aligned with representative bacterial RecA proteins and eucaryal Rad51 and Dmc1 proteins. The alignment supported the existence of four insertions and one deletion in the archaeal and eucaryal sequences relative to the bacterial sequences. The sizes of three of the insertions were found to have taxonomic and phylogenetic significance. Comparative analysis of the RadA sequences, omitting amino acids in the insertions and deletions, shows a cladal distribution of species which mimics to a large extent that obtained by a similar analysis of archaeal 16S rRNA sequences. The PCR technique also was used to amplify fragments of 15 radA genes from uncultured natural sources. Phylogenetic analysis of the amino acid sequences encoded by these fragments reveals several clades with affinity, sometimes only distant, to the putative RadA proteins of several species of Crenarcheota. The two most deeply branching archaeal radA genes found had some amino acid deletion and insertion patterns characteristic of bacterial recA genes. Possible explanations are discussed. Finally, signature codons are presented to distinguish among RecA protein family members.}, keywords = {Amino Acid Sequence, Archaea, Archaeal Proteins, Bacteria, Bacterial Proteins, Cloning, Molecular, DNA Primers, DNA Repair, DNA-Binding Proteins, Evolution, Molecular, Humans, Molecular Sequence Data, Phylogeny, Polymerase Chain Reaction, Recombinant Proteins, RNA, Ribosomal, 16S, Saccharomyces cerevisiae, Sequence Alignment}, issn = {0021-9193}, author = {Sandler, S J and Hugenholtz, P and Schleper, C and DeLong, E F and Pace, N R and Clark, A J} } @article {830, title = {dnaC mutations suppress defects in DNA replication- and recombination-associated functions in priB and priC double mutants in Escherichia coli K-12.}, journal = {Mol Microbiol}, volume = {34}, year = {1999}, month = {1999 Oct}, pages = {91-101}, abstract = {PriA, PriB and PriC were originally discovered as proteins essential for the PhiX174 in vitro DNA replication system. Recent studies have shown that PriA mutants are poorly viable, have high basal levels of SOS expression (SOSH), are recombination deficient (Rec-), sensitive to UV irradiation (UVS) and sensitive to rich media. These data suggest that priA{\textquoteright}s role may be more complex than previously thought and may involve both DNA replication and homologous recombination. Based on the PhiX174 system, mutations in priB and priC should cause phenotypes like those seen in priA2:kan mutants. To test this, mutations in priB and priC were constructed. We found that, contrary to the PhiX174 model, del(priB)302 and priC303:kan mutants have almost wild-type phenotypes. Most unexpectedly, we then found that the priBC double mutant had very poor viability and/or a slow growth rate (even less than a priA2:kan mutant). This suggests that priB and priC have a redundant and important role in Escherichia coli. The priA2:kan suppressor, dnaC809, partially suppressed the poor viability/slow growth phenotype of the priBC double mutant. The resulting triple mutant (priBC dnaC809 ) had small colony size, recombination deficiency and levels of SOS expression similar to a priA2:kan mutant. The priBC dnaC809 mutant, however, was moderately UVR and had good viability, unlike a priA2:kan mutant. Additional mutations in the triple mutant were selected to suppress the slow growth phenotype. One suppressor restored all phenotypes tested to nearly wild-type levels. This mutation was identified as dnaC820 (K178N) [mapping just downstream of dnaC809 (E176G)]. Experiments suggest that dnaC820 makes dnaC809 suppression of priA and or priBC mutants priB and or priC independent. A model is proposed for the roles of these proteins in terms of restarting collapsed replication forks from recombinational intermediates.}, keywords = {Bacterial Proteins, Bacteriophage mu, Cell Division, DNA Replication, DNA, Bacterial, DNA-Binding Proteins, Escherichia coli, Escherichia coli Proteins, Mutation, Phenotype, Recombination, Genetic, Replication Protein A, SOS Response (Genetics), Substrate Specificity, Suppression, Genetic}, issn = {0950-382X}, author = {Sandler, S J and Marians, K J and Zavitz, K H and Coutu, J and Parent, M A and Clark, A J} } @article {790, title = {Evolution of Plasmodium and the recent origin of the world populations of Plasmodium falciparum.}, journal = {Parassitologia}, volume = {41}, year = {1999}, month = {1999 Sep}, pages = {55-68}, abstract = {We have investigated the evolution of Plasmodium parasites by analyzing DNA sequences of several genes. We reach the following conclusions: (1) The four human parasites, P. falciparum, P. malariae, P. ovale, and P. vivax are very remotely related to each other, so that their evolutionary divergence predates the origin of the hominids; several of these parasites became associated with the human lineage by lateral transfer from other hosts. (2) P. falciparum diverged from P. reichenowi about 8 million years ago, consistently with the time of divergence of the human lineage from the apes; a parsimonious inference is that falciparum has been associated with humans since the origin of the hominids. (3) P. malariae is genetically indistinguishable from P. brasilianum, a parasite of New World monkeys; and, similarly. (4) P. vivax is genetically indistinguishable from the New World monkey parasite P. simium. We infer in each of these two cases a very recent lateral transfer between the human and monkey hosts, and explore alternative hypotheses about the direction of the transfer. We have also investigated the population structure of P. falciparum by analyzing 10 genes and conclude that the extant world populations of this parasite have evolved from a single strain within the last several thousand years. The extensive polymorphisms observed in the highly repetitive central region of the Csp gene, as well as the apparently very divergent two classes of alleles at the Msa-1 gene, are consistent with this conclusion.}, keywords = {Amino Acid Sequence, Animals, Base Sequence, DNA, Protozoan, Evolution, Molecular, Hominidae, Humans, Merozoite Surface Protein 1, Molecular Sequence Data, Plasmodium, Plasmodium falciparum, Polymorphism, Genetic, Sequence Alignment}, issn = {0048-2951}, author = {Ayala, F J and Escalante, A A and Rich, S M} } @article {869, title = {Extracellular degradation of medium chain length poly(beta-hydroxyalkanoates) by Comamonas sp.}, journal = {Int J Biol Macromol}, volume = {25}, year = {1999}, month = {1999 Jun-Jul}, pages = {135-43}, abstract = {The PHA-degrading isolate, strain P37C, was enriched from residential compost for its ability to hydrolyze the medium chain length PHA, poly(beta-hydroxyoctanoate) (PHO). It was subsequently found to grow on a wide range of PHAs, including both short chain length and medium chain length PHAs. The isolate was identified as belonging to the genus Comamonas. Strain P37C formed clear zones on poly(beta-hydroxybutyrate) (PHB), (PHO) and poly(beta-hydroxyphenylvalerate) (PHPV) overlay plates. PHA clear zone tubes were prepared using seven different kinds of PHAs, ranging from PHB with four-carbon repeating units, to poly(beta-hydroxyoctanoate-co-beta-hydroxyundecanoate) (PHOU) with 8- and 11-carbon repeating units. There was a direct correlation between PHA side chain length and rate of hydrolysis of the PHAs. A series of PHOUs containing varying percentages of unsaturated bonds were used to make a series of epoxidized PHOUs (PHOEs) with varying percentages of epoxy functions. Results of clear zone tube assays showed that these functionalized PHAs were all biodegradable by strain P37C, and there was no apparent correlation between rate of biodegradation and the proportion of functional groups in the PHAs. Biodegradability of these PHAs was verified using respirometry and enzyme assays. Cell-free supernatants containing activity toward PHAs were prepared, and strain P37C was shown to synthesize at least two distinct PHA depolymerases for the hydrolysis of SCL and MCL PHAs.}, keywords = {Carboxylic Ester Hydrolases, Gram-Negative Aerobic Rods and Cocci, Kinetics, Polyesters, Structure-Activity Relationship, Substrate Specificity}, issn = {0141-8130}, author = {Quinteros, R and Goodwin, S and Lenz, R W and Park, W H} } @article {870, title = {Extracellular polymerization of 3-hydroxyalkanoate monomers with the polymerase of Alcaligenes eutrophus.}, journal = {Int J Biol Macromol}, volume = {25}, year = {1999}, month = {1999 Jun-Jul}, pages = {55-60}, abstract = {Previous investigations on the role of the polymerase in the synthesis of poly-3-hydroxybutyrate (PHB) are reviewed, and the results from earlier in vitro studies on the activity and selectivity of the polymerase of Alcaligenes eutrophus are discussed. In the present study the effect of glycerol on stabilizing the polymerase after purification and on eliminating the lag phase in in vitro polymerization reactions of 3-hydroxybutyl CoA (HBCoA), and 3-hydroxyvaleryl CoA (HVCoA) are described. K(M) values were determined for the activity of the polymerase with both HBCoA and HVCoA, and the rates of propagation for both monomers were estimated. With a racemic mixture of HBCoA, the enzyme polymerized only the [R] monomer.}, keywords = {Acyl Coenzyme A, Acyltransferases, Alcaligenes, Hydroxybutyrates, Kinetics, Polyesters, Substrate Specificity}, issn = {0141-8130}, author = {Lenz, R W and Farcet, C and Dijkstra, P J and Goodwin, S and Zhang, S} } @article {710, title = {Extracellular simian virus 40 transmits a signal that promotes virus enclosure within caveolae.}, journal = {Exp Cell Res}, volume = {246}, year = {1999}, month = {1999 Jan 10}, pages = {83-90}, abstract = {It was reported earlier that entry of simian virus 40 (SV40) into cells is promoted by a signal transmitted by the virus from the cell surface and that SV40 enters cells through caveolae. It is shown here that bound SV40 begins to partition into a caveolae-enriched Triton X-100-insoluble membrane fraction at 30 min postadsorption. Maximal levels of SV40 were seen in that fraction at 1 h. The sterol-binding agent nystatin, which selectively disrupts the cholesterol-enriched caveolae-containing membrane microdomain, selectively blocked the SV40-induced signal. This implies that the SV40 signal is transmitted from that membrane microdomain. The tyrosine kinase inhibitor genistein, which was earlier shown to block the SV40-induced signal and infectious entry, did not block the partitioning of SV40 into the detergent-insoluble membrane fraction. This shows that the signal is not required for the translocation of SV40 to the detergent-insoluble membrane and is consistent with the finding that the signal is likely transmitted from that membrane microdomain. However, electron microscopy of the Triton X-100-insoluble membrane fraction showed that genistein caused SV40 particles to accumulate at the annuli or mouths of the caveolae. In contrast, most SV40 particles were found enclosed within caveolae in parallel samples from untreated control cells. Together, these results imply that SV40 initially binds to flat detergent-soluble membrane. The virus then translocates to a caveolae-containing detergent-insoluble membrane microdomain. From the flat portion of that membrane microdomain the virus induces a signal which promotes its entry into caveolae.}, keywords = {Animals, Blotting, Northern, Blotting, Western, Caveolin 1, Caveolins, Cell Line, Cell Membrane, Cercopithecus aethiops, Cholesterol, Endocytosis, Genes, myc, Genistein, Kidney, Membrane Proteins, Microscopy, Electron, Nystatin, Octoxynol, Signal Transduction, Simian virus 40, Solubility}, issn = {0014-4827}, doi = {10.1006/excr.1998.4301}, author = {Chen, Y and Norkin, L C} } @article {616, title = {Geothrix fermentans gen. nov., sp. nov., a novel Fe(III)-reducing bacterium from a hydrocarbon-contaminated aquifer.}, journal = {Int J Syst Bacteriol}, volume = {49 Pt 4}, year = {1999}, month = {1999 Oct}, pages = {1615-22}, abstract = {In an attempt to understand better the micro-organisms involved in anaerobic degradation of aromatic hydrocarbons in the Fe(III)-reducing zone of petroleum-contaminated aquifers, Fe(III)-reducing micro-organisms were isolated from contaminated aquifer material that had been adapted for rapid oxidation of toluene coupled to Fe(III) reduction. One of these organisms, strain H-5T, was enriched and isolated on acetate/Fe(III) medium. Strain H-5T is a Gram-negative strict anaerobe that grows with various simple organic acids such as acetate, propionate, lactate and fumarate as alternative electron donors with Fe(III) as the electron acceptor. In addition, strain H-5T also oxidizes long-chain fatty acids such as palmitate with Fe(III) as the sole electron acceptor. Strain H-5T can also grow by fermentation of citrate or fumarate in the absence of an alternative electron acceptor. The primary end-products of citrate fermentation are acetate and succinate. In addition to various forms of soluble and insoluble Fe(III), strain H-5T grows with nitrate, Mn(IV), fumarate and the humic acid analogue 2,6-anthraquinone disulfonate as alternative electron acceptors. As with other organisms that can oxidize organic compounds completely with the reduction of Fe(III), cell suspensions of strain H-5T have absorbance maxima indicative of a c-type cytochrome(s). It is proposed that strain H-5T represents a novel genus in the Holophaga-Acidobacterium phylum and that it should be named Geothrix fermentans sp. nov., gen. nov.}, keywords = {Biodegradation, Environmental, DNA, Bacterial, DNA, Ribosomal, Ferric Compounds, Geologic Sediments, Gram-Negative Anaerobic Bacteria, Molecular Sequence Data, Oxidation-Reduction, Petroleum, Phylogeny, RNA, Ribosomal, 16S, Water Microbiology, Water Pollutants, Chemical, Water Supply}, issn = {0020-7713}, author = {Coates, J D and Ellis, D J and Gaw, C V and Lovley, D R} } @article {605, title = {Humics as an electron donor for anaerobic respiration.}, journal = {Environ Microbiol}, volume = {1}, year = {1999}, month = {1999 Feb}, pages = {89-98}, abstract = {The possibility that microorganisms might use reduced humic substances (humics) as an electron donor for the reduction of electron acceptors with a more positive redox potential was investigated. All of the Fe(III)- and humics-reducing microorganisms evaluated were capable of oxidizing reduced humics and/or the reduced humics analogue anthrahydroquinone-2,6,-disulphonate (AHODS), with nitrate and/or fumarate as the electron acceptor. These included Geobacter metallireducens, Geobacter sulphurreducens, Geothrix fermentans, Shewanella alga, Wolinella succinogenes and {\textquoteright}S. barnesii{\textquoteright}. Several of the humics-oxidizing microorganisms grew in medium with AHQDS as the sole electron donor and fumarate as the electron acceptor. Even though it does not reduce Fe(III) or humics, Paracoccus denitrificans could use AHQDS and reduced humics as electron donors for denitrification. However, another denitrifier, Pseudomonas denitrificans, could not. AHODS could also serve as an electron donor for selenate and arsenate reduction by W. succinogenes. Electron spin resonance studies demonstrated that humics oxidation was associated with the oxidation of hydroquinone moieties in the humics. Studies with G. metallireducens and W. succinogenes demonstrated that the anthraquinone-2,6-disulphonate (AQDS)/AHQDS redox couple mediated an interspecies electron transfer between the two organisms. These results suggest that, as microbially reduced humics enter less reduced zones of soils and sediments, the reduced humics may serve as electron donors for microbial reduction of several environmentally significant electron acceptors.}, keywords = {Aerobiosis, Anaerobiosis, Anthraquinones, Arsenates, Bacteria, Electron Transport, Fumarates, Humic Substances, Selenium Compounds, Tumor Cells, Cultured}, issn = {1462-2912}, author = {Lovley, D R and Fraga, J L and Coates, J D and Blunt-Harris, E L} } @article {403, title = {Interdependence of the position and orientation of SoxS binding sites in the transcriptional activation of the class I subset of Escherichia coli superoxide-inducible promoters.}, journal = {Mol Microbiol}, volume = {34}, year = {1999}, month = {1999 Nov}, pages = {414-30}, abstract = {SoxS is the direct transcriptional activator of the member genes of the Escherichia coli superoxide regulon. At class I SoxS-dependent promoters, e.g. zwf and fpr, whose SoxS binding sites ({\textquoteright}soxbox{\textquoteright}) lie upstream of the -35 region of the promoter, activation requires the C-terminal domain of the RNA polymerase alpha-subunit, while at class II SoxS-dependent promoters, e.g. fumC and micF, whose binding sites overlap the -35 region, activation is independent of the alpha-CTD. To determine whether SoxS activation of its class I promoters shows the same helical phase-dependent spacing requirement as class I promoters activated by catabolite gene activator protein, we increased the 7 bp distance between the 20 bp zwf soxbox and the zwf -35 promoter hexamer by 5 bp and 11 bp, and we decreased the 15 bp distance between the 20 bp fpr soxbox and the fpr -35 promoter hexamer by the same amounts. In both cases, displacement of the binding site by a half or full turn of the DNA helix prevented transcriptional activation. With constructs containing the binding site of one gene fused to the promoter of the other, we demonstrated that the positional requirements are a function of the specific binding site, not the promoter. Supposing that opposite orientation of the SoxS binding site at the two promoters might account for the positional requirements, we placed the zwf and fpr soxboxes in the reverse orientation at the various positions upstream of the promoters and determined the effect of orientation on transcription activation. We found that reversing the orientation of the zwf binding site converts its positional requirement to that of the fpr binding site in its normal orientation, and vice versa. Analysis by molecular information theory of DNA sequences known to bind SoxS in vitro is consistent with the opposite orientation of the zwf and fpr soxboxes.}, keywords = {Bacterial Proteins, Base Sequence, Binding Sites, Escherichia coli, Escherichia coli Proteins, Molecular Sequence Data, Plasmids, Promoter Regions, Genetic, Sequence Analysis, DNA, Superoxides, Trans-Activators, Transcription Factors, Transcription, Genetic, Transcriptional Activation}, issn = {0950-382X}, author = {Wood, T I and Griffith, K L and Fawcett, W P and Jair, K W and Schneider, T D and Wolf, R E} } @article {619, title = {Microbial communities associated with anaerobic benzene degradation in a petroleum-contaminated aquifer.}, journal = {Appl Environ Microbiol}, volume = {65}, year = {1999}, month = {1999 Jul}, pages = {3056-63}, abstract = {Microbial community composition associated with benzene oxidation under in situ Fe(III)-reducing conditions in a petroleum-contaminated aquifer located in Bemidji, Minn., was investigated. Community structure associated with benzene degradation was compared to sediment communities that did not anaerobically oxidize benzene which were obtained from two adjacent Fe(III)-reducing sites and from methanogenic and uncontaminated zones. Denaturing gradient gel electrophoresis of 16S rDNA sequences amplified with bacterial or Geobacteraceae-specific primers indicated significant differences in the composition of the microbial communities at the different sites. Most notable was a selective enrichment of microorganisms in the Geobacter cluster seen in the benzene-degrading sediments. This finding was in accordance with phospholipid fatty acid analysis and most-probable-number-PCR enumeration, which indicated that members of the family Geobacteraceae were more numerous in these sediments. A benzene-oxidizing Fe(III)-reducing enrichment culture was established from benzene-degrading sediments and contained an organism closely related to the uncultivated Geobacter spp. This genus contains the only known organisms that can oxidize aromatic compounds with the reduction of Fe(III). Sequences closely related to the Fe(III) reducer Geothrix fermentans and the aerobe Variovorax paradoxus were also amplified from the benzene-degrading enrichment and were present in the benzene-degrading sediments. However, neither G. fermentans nor V. paradoxus is known to oxidize aromatic compounds with the reduction of Fe(III), and there was no apparent enrichment of these organisms in the benzene-degrading sediments. These results suggest that Geobacter spp. play an important role in the anaerobic oxidation of benzene in the Bemidji aquifer and that molecular community analysis may be a powerful tool for predicting a site{\textquoteright}s capacity for anaerobic benzene degradation.}, keywords = {Anaerobiosis, Benzene, Biodegradation, Environmental, Culture Media, DNA, Bacterial, DNA, Ribosomal, Fresh Water, Geologic Sediments, Gram-Negative Anaerobic Bacteria, Molecular Sequence Data, Oxidation-Reduction, Petroleum, Phylogeny, Polymerase Chain Reaction, RNA, Ribosomal, 16S, Sequence Analysis, DNA, Water Pollutants}, issn = {0099-2240}, author = {Rooney-Varga, J N and Anderson, R T and Fraga, J L and Ringelberg, D and Lovley, D R} } @article {615, title = {The periplasmic 9.6-kilodalton c-type cytochrome of Geobacter sulfurreducens is not an electron shuttle to Fe(III).}, journal = {J Bacteriol}, volume = {181}, year = {1999}, month = {1999 Dec}, pages = {7647-9}, abstract = {Geobacter sulfurreducens contains a 9.6-kDa c-type cytochrome that was previously proposed to serve as an extracellular electron shuttle to insoluble Fe(III) oxides. However, when the cytochrome was added to washed-cell suspensions of G. sulfurreducens it did not enhance Fe(III) oxide reduction, whereas similar concentrations of the known electron shuttle, anthraquinone-2,6-disulfonate, greatly stimulated Fe(III) oxide reduction. Furthermore, analysis of the extracellular c-type cytochromes in cultures of G. sulfurreducens demonstrated that the dominant c-type cytochrome was not the 9.6-kDa cytochrome, but rather a 41-kDa cytochrome. These results and other considerations suggest that the 9.6-kDa cytochrome is not an important extracellular electron shuttle to Fe(III) oxides.}, keywords = {Cytochrome c Group, Electron Transport, Electrophoresis, Polyacrylamide Gel, Ferric Compounds, Gram-Negative Anaerobic Bacteria, Molecular Weight, Periplasm}, issn = {0021-9193}, author = {Lloyd, J R and Blunt-Harris, E L and Lovley, D R} } @article {831, title = {Replication fork assembly at recombination intermediates is required for bacterial growth.}, journal = {Proc Natl Acad Sci U S A}, volume = {96}, year = {1999}, month = {1999 Mar 30}, pages = {3552-5}, abstract = {PriA, a 3{\textquoteright} --> 5{\textquoteright} DNA helicase, directs assembly of a primosome on some bacteriophage and plasmid DNAs. Primosomes are multienzyme replication machines that contribute both the DNA-unwinding and Okazaki fragment-priming functions at the replication fork. The role of PriA in chromosomal replication is unclear. The phenotypes of priA null mutations suggest that the protein participates in replication restart at recombination intermediates. We show here that PriA promotes replication fork assembly at a D loop, an intermediate formed during initiation of homologous recombination. We also show that DnaC810, encoded by a naturally arising intergenic suppressor allele of the priA2::kan mutation, bypasses the need for PriA during replication fork assembly at D loops in vitro. These findings underscore the essentiality of replication fork restart at recombination intermediates under normal growth conditions in bacteria.}, keywords = {Bacteria, Bacteriophage phi X 174, Base Sequence, DNA Polymerase III, DNA Replication, DNA-Binding Proteins, Escherichia coli, Molecular Sequence Data, Oligodeoxyribonucleotides, Open Reading Frames, Recombination, Genetic, Replication Protein A, Templates, Genetic}, issn = {0027-8424}, author = {Liu, J and Xu, L and Sandler, S J and Marians, K J} } @article {791, title = {Reply}, journal = {Parasitol Today}, volume = {15}, year = {1999}, month = {1999 Jan}, pages = {39-40}, issn = {0169-4758}, author = {Rich, S M and Ayala, F J} } @article {617, title = {Role of humic-bound iron as an electron transfer agent in dissimilatory Fe(III) reduction.}, journal = {Appl Environ Microbiol}, volume = {65}, year = {1999}, month = {1999 Sep}, pages = {4252-4}, abstract = {The dissimilatory Fe(III) reducer Geobacter metallireducens reduced Fe(III) bound in humic substances, but the concentrations of Fe(III) in a wide range of highly purified humic substances were too low to account for a significant portion of the electron-accepting capacities of the humic substances. Furthermore, once reduced, the iron in humic substances could not transfer electrons to Fe(III) oxide. These results suggest that other electron-accepting moieties in humic substances, such as quinones, are the important electron-accepting and shuttling agents under Fe(III)-reducing conditions.}, keywords = {Deltaproteobacteria, Electron Transport, Ferric Compounds, Humic Substances, Iron, Oxidation-Reduction}, issn = {0099-2240}, author = {Lovley, D R and Blunt-Harris, E L} } @article {711, title = {Simian virus 40 infection via MHC class I molecules and caveolae.}, journal = {Immunol Rev}, volume = {168}, year = {1999}, month = {1999 Apr}, pages = {13-22}, abstract = {MHC class I molecules are a necessary component of the cell surface receptor for simian virus 40 (SV40). After binding to class I molecules, SV40 enters cells via a unique endocytic pathway that involves caveolae, rather than clathrin-coated pits. This pathway is dependent on a transmembrane signal that SV40 transmits from the cell surface. Furthermore, it delivers SV40 to the endoplasmic reticulum, rather than to the endosomal/lysosomal compartment, which is the usual target for endocytic traffic. The glycosphingolipid and cholesterol-enriched plasma membrane domains that contain caveolae are also enriched for class I molecules, relative to whole plasma membrane. Nevertheless, although class I molecules bind SV40, they do not enter with SV40, nor do they enter spontaneously into uninfected SV40 host cells. Instead, they are shed from the cell surface by the activity of a metalloprotease. These results imply the existence of a putative secondary receptor for SV40 that might mediate SV40 entry. It is not yet clear whether class I molecules are active in transmitting the SV40 signal. Monoclonal antibodies against class I molecules also induce a signal in the SV40 host cells. However, the antibody-induced signal is mediated by mitogen-activated protein kinase (MAP kinase), whereas the SV40 signal is independent of MAP kinase.}, keywords = {Animals, Cell Membrane, Histocompatibility Antigens Class I, Humans, Papillomavirus Infections, Receptors, Virus, Simian virus 40, Tumor Virus Infections}, issn = {0105-2896}, author = {Norkin, L C} } @article {753, title = {Soil bacterial community shift correlated with change from forest to pasture vegetation in a tropical soil.}, journal = {Appl Environ Microbiol}, volume = {65}, year = {1999}, month = {1999 Aug}, pages = {3622-6}, abstract = {The change in vegetative cover of a Hawaiian soil from forest to pasture led to significant changes in the composition of the soil bacterial community. DNAs were extracted from both soil habitats and compared for the abundance of guanine-plus-cytosine (G+C) content, by analysis of abundance of phylotypes of small-subunit ribosomal DNA (SSU rDNA) amplified from fractions with 63 and 35\% G+C contents, and by phylogenetic analysis of the dominant rDNA clones in the 63\% G+C content fraction. All three methods showed differences between the forest and pasture habitats, providing evidence that vegetation had a strong influence on microbial community composition at three levels of taxon resolution. The forest soil DNA had a peak in G+C content of 61\%, while the DNA of the pasture soil had a peak in G+C content of 67\%. None of the dominant phylotypes found in the forest soil were detected in the pasture soil. For the 63\% G+C fraction SSU rDNA sequence analysis of the three most dominant members revealed that their phyla changed from Fibrobacter and Syntrophomonas assemblages in the forest soil to Burkholderia and Rhizobium-Agrobacterium assemblages in the pasture soil.}, keywords = {Agriculture, Bacteria, Base Composition, DNA, Bacterial, Ecosystem, Molecular Sequence Data, Soil Microbiology, Trees, Tropical Climate}, issn = {0099-2240}, author = {N{\"u}sslein, K and Tiedje, J M} } @article {618, title = {Sulfurospirillum barnesii sp. nov. and Sulfurospirillum arsenophilum sp. nov., new members of the Sulfurospirillum clade of the epsilon Proteobacteria.}, journal = {Int J Syst Bacteriol}, volume = {49 Pt 3}, year = {1999}, month = {1999 Jul}, pages = {1177-80}, abstract = {Two strains of dissimilatory arsenate-reducing vibrio-shaped bacteria are assigned to the genus Sulfurospirillum. These two new species, Sulfurospirillum barnesii strain SES-3T and Sulfurospirillum arsenophilum strain MIT-13T, in addition to Sulfurospirillum sp. SM-5, two strains of Sulfurospirillum deleyianum, and Sulfurospirillum arcachonense, form a distinct clade within the epsilon subclass of the Proteobacteria based on 16S rRNA analysis.}, keywords = {Arsenates, Bacterial Typing Techniques, DNA, Ribosomal, Genes, rRNA, Gram-Negative Bacteria, Molecular Sequence Data, Oxidation-Reduction, Phylogeny, RNA, Ribosomal, 16S, Selenium Compounds, Sequence Analysis, DNA}, issn = {0020-7713}, author = {Stolz, J F and Ellis, D J and Blum, J S and Ahmann, D and Lovley, D R and Oremland, R S} } @article {626, title = {Anaerobic benzene degradation in petroleum-contaminated aquifer sediments after inoculation with a benzene-oxidizing enrichment.}, journal = {Appl Environ Microbiol}, volume = {64}, year = {1998}, month = {1998 Feb}, pages = {775-8}, abstract = {Sediments from the sulfate-reduction zone of a petroleum-contaminated aquifer, in which benzene persisted, were inoculated with a benzene-oxidizing, sulfate-reducing enrichment from aquatic sediments. Benzene was degraded, with apparent growth of the benzene-degrading population over time. These results suggest that the lack of benzene degradation in the sulfate-reduction zones of some aquifers may result from the failure of the appropriate benzene-degrading sulfate reducers to colonize the aquifers rather than from environmental conditions that are adverse for anaerobic benzene degradation.}, keywords = {Anaerobiosis, Benzene, Biodegradation, Environmental, Fresh Water, Geologic Sediments, Iron, Oxidation-Reduction, Petroleum, Sulfates}, issn = {0099-2240}, author = {Weiner, JM and Lovley, D R} } @article {516, title = {Carbohydrate oxidation coupled to Fe(III) reduction, a novel form of anaerobic metabolism.}, journal = {Anaerobe}, volume = {4}, year = {1998}, month = {1998 Dec}, pages = {277-82}, abstract = {An isolate, designated GC-29, that could incompletely oxidize glucose to acetate and carbon dioxide with Fe(III) serving as the electron acceptor was recovered from freshwater sediments of the Potomac River, Maryland. This metabolism yielded energy to support cell growth. Strain GC-29 is a facultatively anaerobic, gram-negative motile rod which, in addition to glucose, also used sucrose, lactate, pyruvate, yeast extract, casamino acids or H2 as alternative electron donors for Fe(III) reduction. Stain GC-29 could reduce NO3(-), Mn(IV), U(VI), fumarate, malate, S2O3(2-), and colloidal S0 as well as the humics analog, 2,6-anthraquinone disulfonate. Analysis of the almost complete 16S rRNA sequence indicated that strain GC-29 belongs in the Shewanella genus in the epsilon subdivision of the Proteobacteria. The name Shewanella saccharophilia is proposed. Shewanella saccharophilia differs from previously described fermentative microorganisms that metabolize glucose with the reduction of Fe(III) because it transfers significantly more electron equivalents to Fe(III); acetate and carbon dioxide are the only products of glucose metabolism; energy is conserved from Fe(III) reduction; and glucose is not metabolized in the absence of Fe(III). The metabolism of organisms like S. saccharophilia may account for the fact that glucose is metabolized primarily to acetate and carbon dioxide in a variety of sediments in which Fe(III) reduction is the terminal electron accepting process.}, issn = {1075-9964}, doi = {10.1006/anae.1998.0172}, author = {Coates, J D and Councell, T and Ellis, D J and Lovley, D R} } @article {754, title = {Characterization of the dominant and rare members of a young Hawaiian soil bacterial community with small-subunit ribosomal DNA amplified from DNA fractionated on the basis of its guanine and cytosine composition.}, journal = {Appl Environ Microbiol}, volume = {64}, year = {1998}, month = {1998 Apr}, pages = {1283-9}, abstract = {The small-subunit ribosomal DNA (rDNA) diversity was found to be very high in a Hawaiian soil community that might be expected to have lower diversity than the communities in continental soils because the Hawaiian soil is geographically isolated and only 200 years old, is subjected to a constant climate, and harbors low plant diversity. Since an underlying community structure could not be revealed by analyzing the total eubacterial rDNA, we first fractionated the DNA on the basis of guanine-plus-cytosine (G + C) content by using bisbenzimidazole and equilibrium centrifugation and then analyzed the bacterial rDNA amplified from a fraction with a high biomass (63\% G + C fraction) and a fraction with a low biomass (35\% G + C fraction). The rDNA clone libraries were screened by amplified rDNA restriction analysis to determine phylotype distribution. The dominant biomass reflected by the 63\% G + C fraction contained several dominant phylotypes, while the community members that were less successful (35\% G + C fraction) did not show dominance but there was a very high diversity of phylotypes. Nucleotide sequence analysis revealed taxa belonging to the groups expected for the G + C contents used. The dominant phylotypes in the 63\% G + C fraction were members of the Pseudomonas, Rhizobium-Agrobacterium, and Rhodospirillum assemblages, while all of the clones sequenced from the 35\% G + C fraction were affiliated with several Clostridium assemblages. The two-step rDNA analysis used here uncovered more diversity than can be detected by direct rDNA analysis of total community DNA. The G + C separation step is also a way to detect some of the less dominant organisms in a community.}, keywords = {Base Composition, Clostridium, DNA, Bacterial, DNA, Ribosomal, Ecosystem, Genetic Variation, Hawaii, Phylogeny, Pseudomonas, Rhizobium, Rhodospirillum, Soil Microbiology}, issn = {0099-2240}, author = {N{\"u}sslein, K and Tiedje, J M} } @article {621, title = {Evidence against hydrogen-based microbial ecosystems in basalt aquifers}, journal = {Science}, volume = {281}, year = {1998}, month = {1998 Aug 14}, pages = {976-7}, abstract = {It has been proposed that hydrogen produced from basalt-ground-water interactions may serve as an energy source that supports the existence of microorganisms in the deep subsurface on Earth and possibly on other planets. However, experiments demonstrated that hydrogen is not produced from basalt at an environmentally relevant, alkaline pH. Small amounts of hydrogen were produced at a lower pH in laboratory incubations, but even this hydrogen production was transitory. Furthermore, geochemical considerations suggest that previously reported rates of hydrogen production cannot be sustained over geologically significant time frames. These findings indicate that hydrogen production from basalt-ground-water interactions may not support microbial metabolism in the subsurface.}, issn = {1095-9203}, author = {Anderson, R T and Chapelle, F H and Lovley, D R} } @article {623, title = {Growth of geobacter sulfurreducens with acetate in syntrophic cooperation with hydrogen-oxidizing anaerobic partners}, journal = {Appl Environ Microbiol}, volume = {64}, year = {1998}, month = {1998 Jun}, pages = {2232-6}, abstract = {Pure cultures of Geobacter sulfurreducens and other Fe(III)-reducing bacteria accumulated hydrogen to partial pressures of 5 to 70 Pa with acetate, butyrate, benzoate, ethanol, lactate, or glucose as the electron donor if electron release to an acceptor was limiting. G. sulfurreducens coupled acetate oxidation with electron transfer to an anaerobic partner bacterium in the absence of ferric iron or other electron acceptors. Cocultures of G. sulfurreducens and Wolinella succinogenes with nitrate as the electron acceptor degraded acetate efficiently and grew with doubling times of 6 to 8 h. The hydrogen partial pressures in these acetate-degrading cocultures were considerably lower, in the range of 0.02 to 0.04 Pa. From these values and the concentrations of the other reactants, it was calculated that in this cooperation the free energy change available to G. sulfurreducens should be about -53 kJ per mol of acetate oxidized, assuming complete conversion of acetate to CO2 and H2. However, growth yields (18.5 g of dry mass per mol of acetate for the coculture, about 14 g for G. sulfurreducens) indicated considerably higher energy gains. These yield data, measurement of hydrogen production rates, and calculation of the diffusive hydrogen flux indicated that electron transfer in these cocultures may not proceed exclusively via interspecies hydrogen transfer but may also proceed through an alternative carrier system with higher redox potential, e.g., a c-type cytochrome that was found to be excreted by G. sulfurreducens into the culture fluid. Syntrophic acetate degradation was also possible with G. sulfurreducens and Desulfovibrio desulfuricans CSN but only with nitrate as electron acceptor. These cultures produced cell yields of 4.5 g of dry mass per mol of acetate, to which both partners contributed at about equal rates. These results demonstrate that some Fe(III)-reducing bacteria can oxidize organic compounds under Fe(III) limitation with the production of hydrogen, and they provide the first example of rapid acetate oxidation via interspecies electron transfer at moderate temperature.}, issn = {1098-5336}, author = {Cord-Ruwisch, R and Lovley, D R and Schink, B} } @article {622, title = {Humic acids as electron acceptors for anaerobic microbial oxidation of vinyl chloride and dichloroethene.}, journal = {Appl Environ Microbiol}, volume = {64}, year = {1998}, month = {1998 Aug}, pages = {3102-5}, abstract = {Anaerobic oxidation of [1,2-14C]vinyl chloride and [1, 2-14C]dichloroethene to 14CO2 under humic acid-reducing conditions was demonstrated. The results indicate that waterborne contaminants can be oxidized by using humic acid compounds as electron acceptors and suggest that natural aquatic systems have a much larger capacity for contaminant oxidation than previously thought.}, keywords = {Aerobiosis, Anaerobiosis, Bacteria, Biodegradation, Environmental, Geologic Sediments, Humic Substances, Oxidation-Reduction, Vinyl Chloride, Water Microbiology, Water Pollutants}, issn = {0099-2240}, author = {Bradley, P M and Chapelle, F H and Lovley, D R} } @article {793, title = {Malaria{\textquoteright}s Eve: evidence of a recent population bottleneck throughout the world populations of Plasmodium falciparum.}, journal = {Proc Natl Acad Sci U S A}, volume = {95}, year = {1998}, month = {1998 Apr 14}, pages = {4425-30}, abstract = {We have analyzed DNA sequences from world-wide geographic strains of Plasmodium falciparum and found a complete absence of synonymous DNA polymorphism at 10 gene loci. We hypothesize that all extant world populations of the parasite have recently derived (within several thousand years) from a single ancestral strain. The upper limit of the 95\% confidence interval for the time when this most recent common ancestor lived is between 24,500 and 57,500 years ago (depending on different estimates of the nucleotide substitution rate); the actual time is likely to be much more recent. The recent origin of the P. falciparum populations could have resulted from either a demographic sweep (P. falciparum has only recently spread throughout the world from a small geographically confined population) or a selective sweep (one strain favored by natural selection has recently replaced all others). The selective sweep hypothesis requires that populations of P. falciparum be effectively clonal, despite the obligate sexual stage of the parasite life cycle. A demographic sweep that started several thousand years ago is consistent with worldwide climatic changes ensuing the last glaciation, increased anthropophilia of the mosquito vectors, and the spread of agriculture. P. falciparum may have rapidly spread from its African tropical origins to the tropical and subtropical regions of the world only within the last 6,000 years. The recent origin of the world-wide P. falciparum populations may account for its virulence, as the most malignant of human malarial parasites.}, keywords = {Africa, Animals, Asia, Climate, Demography, DNA, Protozoan, Evolution, Molecular, Genes, Protozoan, Humans, Life Cycle Stages, Malaria, Malaria, Falciparum, Netherlands, Phylogeny, Plasmodium, Plasmodium falciparum, Polymorphism, Genetic, South America, Tetrahydrofolate Dehydrogenase, Time}, issn = {0027-8424}, author = {Rich, S M and Licht, M C and Hudson, R R and Ayala, F J} } @article {709, title = {MHC class I molecules are enriched in caveolae but do not enter with simian virus 40.}, journal = {J Gen Virol}, volume = {79 ( Pt 6)}, year = {1998}, month = {1998 Jun}, pages = {1469-77}, abstract = {Simian virus 40 (SV40) binds to MHC class I molecules anywhere on the cell surface and then enters through caveolae. The fate of class I molecules after SV40 binding is not known. Sensitivity of 125I-surface-labelled class I molecules to papain cleavage was used to distinguish internalized class I molecules from class I molecules remaining at the cell surface. Whereas the caveolae-enriched membrane microdomain was found to also be enriched for class I molecules, no internalized papain-resistant 125I-surface-labelled class I molecules could be detected at any time in either control cells or in cells preadsorbed with saturating amounts of SV40. Instead, 125I-surface-labelled class I molecules, as well as preadsorbed 125I-labelled anti-class I antibodies, accumulated in the medium, coincident with the turnover of class I molecules at the cell surface. The class I heavy chains that accumulated in the medium were truncated and their release was specifically prevented by the metalloprotease inhibitor 1,10-phenanthroline. Thus, whereas class I molecules mediate SV40 binding, they do not appear to mediate SV40 entry.}, keywords = {Animals, Caveolin 1, Caveolins, Cell Line, Cell Membrane, Cercopithecus aethiops, Histocompatibility Antigens Class I, Humans, Membrane Proteins, Simian virus 40}, issn = {0022-1317}, author = {Anderson, H A and Chen, Y and Norkin, L C} } @article {620, title = {Microbiological evidence for Fe(III) reduction on early Earth.}, journal = {Nature}, volume = {395}, year = {1998}, month = {1998 Sep 3}, pages = {65-7}, abstract = {It is generally considered that sulphur reduction was one of the earliest forms of microbial respiration, because the known microorganisms that are most closely related to the last common ancestor of modern life are primarily anaerobic, sulphur-reducing hyperthermophiles. However, geochemical evidence indicates that Fe(III) is more likely than sulphur to have been the first external electron acceptor of global significance in microbial metabolism. Here we show that Archaea and Bacteria that are most closely related to the last common ancestor can reduce Fe(III) to Fe(II) and conserve energy to support growth from this respiration. Surprisingly, even Thermotoga maritima, previously considered to have only a fermentative metabolism, could grow as a respiratory organism when Fe(III) was provided as an electron acceptor. These results provide microbiological evidence that Fe(III) reduction could have been an important process on early Earth and suggest that microorganisms might contribute to Fe(III) reduction in modern hot biospheres. Furthermore, our discovery that hyperthermophiles that had previously been thought to require sulphur for cultivation can instead be grown without the production of toxic and corrosive sulphide, should aid biochemical investigations of these poorly understood organisms.}, keywords = {Earth (Planet), Electron Transport, Environmental Microbiology, Ferric Compounds, Gram-Negative Anaerobic Bacteria, Oxidation-Reduction, Thermoproteaceae}, issn = {0028-0836}, doi = {10.1038/25720}, author = {Vargas, M and Kashefi, K and Blunt-Harris, E L and Lovley, D R} } @article {833, title = {RadA protein is an archaeal RecA protein homolog that catalyzes DNA strand exchange.}, journal = {Genes Dev}, volume = {12}, year = {1998}, month = {1998 May 1}, pages = {1248-53}, abstract = {With the discovery that the Saccharomyces cerevisiae Rad51 protein is both structurally and functionally similar to the Escherichia coli RecA protein, the RecA paradigm for homologous recombination was extended to the Eucarya. The ubiquitous presence of RecA and Rad51 protein homologs raises the question of whether this archetypal protein exists within the third domain of life, the Archaea. Here we present the isolation of a Rad51/RecA protein homolog from the archaeon Sulfolobus solfataricus, and show that this protein, RadA, possesses the characteristics of a DNA strand exchange protein: The RadA protein is a DNA-dependent ATPase, forms a nucleoprotein filament on DNA, and catalyzes DNA pairing and strand exchange.}, keywords = {Adenosine Triphosphatases, Archaeal Proteins, Bacterial Proteins, DNA Helicases, DNA, Archaeal, DNA-Binding Proteins, Escherichia coli, Escherichia coli Proteins, Genes, Archaeal, Nucleoproteins, Rad51 Recombinase, Rec A Recombinases, Recombinant Proteins, Recombination, Genetic, Sulfolobus}, issn = {0890-9369}, author = {Seitz, E M and Brockman, J P and Sandler, S J and Clark, A J and Kowalczykowski, S C} } @article {624, title = {Rapid Benzene Degradation in Methanogenic Sediments from a Petroleum-Contaminated Aquifer}, journal = {Appl Environ Microbiol}, volume = {64}, year = {1998}, month = {1998 May 1}, pages = {1937-9}, abstract = {In methanogenic sediments from a petroleum-contaminated aquifer, [14C]benzene was converted to 14CH4 and 14CO2 without an apparent lag. Phenol, acetate, and propionate were intermediates in benzene mineralization. These results suggest that alternative electron acceptors need not be available for there to be significant natural attenuation of benzene in some petroleum-contaminated aquifers.}, issn = {1098-5336}, author = {Weiner, JM and Lovley, D R} } @article {792, title = {The recent origin of allelic variation in antigenic determinants of Plasmodium falciparum.}, journal = {Genetics}, volume = {150}, year = {1998}, month = {1998 Sep}, pages = {515-7}, keywords = {Alleles, Animals, Epitopes, Genetic Variation, Merozoite Surface Protein 1, Plasmodium falciparum, Protozoan Proteins}, issn = {0016-6731}, author = {Rich, S M and Ayala, F J} } @article {625, title = {Recovery of humic-reducing bacteria from a diversity of environments.}, journal = {Appl Environ Microbiol}, volume = {64}, year = {1998}, month = {1998 Apr}, pages = {1504-9}, abstract = {To evaluate which microorganisms might be responsible for microbial reduction of humic substances in sedimentary environments, humic-reducing bacteria were isolated from a variety of sediment types. These included lake sediments, pristine and contaminated wetland sediments, and marine sediments. In each of the sediment types, all of the humic reducers recovered with acetate as the electron donor and the humic substance analog, 2,6-anthraquinone disulfonate (AQDS), as the electron acceptor were members of the family Geobacteraceae. This was true whether the AQDS-reducing bacteria were enriched prior to isolation on solid media or were recovered from the highest positive dilutions of sediments in liquid media. All of the isolates tested not only conserved energy to support growth from acetate oxidation coupled to AQDS reduction but also could oxidize acetate with highly purified soil humic acids as the sole electron acceptor. All of the isolates tested were also able to grow with Fe(III) serving as the sole electron acceptor. This is consistent with previous studies that have suggested that the capacity for Fe(III) reduction is a common feature of all members of the Geobacteraceae. These studies demonstrate that the potential for microbial humic substance reduction can be found in a wide variety of sediment types and suggest that Geobacteraceae species might be important humic-reducing organisms in sediments.}, keywords = {Acetic Acid, Anthraquinones, Base Sequence, DNA Primers, Electron Transport, Fresh Water, Gram-Negative Anaerobic Bacteria, Humic Substances, Iron, Molecular Sequence Data, Phylogeny, Polymerase Chain Reaction, RNA, Bacterial, RNA, Ribosomal, 16S, Seawater, Sulfur-Reducing Bacteria, Water Microbiology}, issn = {0099-2240}, author = {Coates, J D and Ellis, D J and Blunt-Harris, E L and Gaw, C V and Roden, E E and Lovley, D R} } @article {390, title = {Altered phosphorylation/inactivation of a novel pyruvate dehydrogenase in adult Ascaris suum muscle.}, journal = {Mol Biochem Parasitol}, volume = {90}, year = {1997}, month = {1997 Dec 1}, pages = {323-6}, keywords = {Animals, Ascaris suum, Cattle, Kidney, Muscles, Phosphorylation, Protein Kinases, Protein-Serine-Threonine Kinases, Pyruvate Dehydrogenase (Lipoamide), Pyruvate Dehydrogenase Complex}, issn = {0166-6851}, author = {Klingbeil, M M and Walker, D J and Huang, Y J and Komuniecki, R} } @article {628, title = {Anaerobic degradation of polycyclic aromatic hydrocarbons and alkanes in petroleum-contaminated marine harbor sediments.}, journal = {Appl Environ Microbiol}, volume = {63}, year = {1997}, month = {1997 Sep}, pages = {3589-93}, abstract = {Although polycyclic aromatic hydrocarbons (PAHs) have usually been found to persist under strict anaerobic conditions, in a previous study an unusual site was found in San Diego Bay in which two PAHs, naphthalene and phenanthrene, were oxidized to carbon dioxide under sulfate-reducing conditions. Further investigations with these sediments revealed that methylnaphthalene, fluorene, and fluoranthene were also anaerobically oxidized to carbon dioxide in these sediments, while pyrene and benzo[a]pyrene were not. Studies with naphthalene indicated that PAH oxidation was sulfate dependent. Incubating the sediments with additional naphthalene for 1 month resulted in a significant increase in the oxidation of [14C]naphthalene. In sediments from a less heavily contaminated site in San diego Bay where PAHs were not readily degraded, naphthalene degradation could be stimulated through inoculation with active PAH-degrading sediments from the most heavily contaminated site. Sediments from the less heavily contaminated site that had been adapted for rapid anaerobic degradation of high concentrations of benzene did not oxidize naphthalene, suggesting that the benzene- and naphthalene-degrading populations were different. When fuels containing complex mixtures of alkanes were added to sediments from the two sites, there was significant degradation in the alkanes. [14C]hexadecane was also anaerobically oxidized to 14CO2 in these sediments. Molybdate, a specific inhibitor of sulfate reduction, inhibited hexadecane oxidation. These results demonstrate that a wide variety of hydrocarbon contaminants can be degraded under sulfate-reducing conditions in hydrocarbon-contaminated sediments, and they suggest that it may be possible to use sulfate reduction rather than aerobic respiration as a treatment strategy for hydrocarbon-contaminated dredged sediments.}, keywords = {Alkanes, Anaerobiosis, Biodegradation, Environmental, California, Naphthalenes, Oxidation-Reduction, Petroleum, Polycyclic Hydrocarbons, Aromatic, Sulfates, Water Microbiology, Water Pollutants, Chemical}, issn = {0099-2240}, author = {Coates, J D and Woodward, J and Allen, J and Philp, P and Lovley, D R} } @article {629, title = {Bioremediation of metal contamination.}, journal = {Curr Opin Biotechnol}, volume = {8}, year = {1997}, month = {1997 Jun}, pages = {285-9}, abstract = {Recent studies have demonstrated that microbes might be used to remediate metal contamination by removing metals from contaminated water or waste streams, sequestering metals in soils and sediments or solubilizing metals to aid in their extraction. This is primarily accomplished either by biosorption of metals or enzymatically catalyzed changes in the metal redox state. Bioremediation of metals is still primarily a research problem with little large-scale application of this technology.}, keywords = {Adsorption, Bacteria, Biomass, Biotechnology, Environmental Pollutants, Metals, Oxidation-Reduction}, issn = {0958-1669}, author = {Lovley, D R and Coates, J D} } @article {627, title = {Dissimilatory arsenate and sulfate reduction in Desulfotomaculum auripigmentum sp. nov.}, journal = {Arch Microbiol}, volume = {168}, year = {1997}, month = {1997 Nov}, pages = {380-8}, abstract = {A newly discovered arsenate-reducing bacterium, strain OREX-4, differed significantly from strains MIT-13 and SES-3, the previously described arsenate-reducing isolates, which grew on nitrate but not on sulfate. In contrast, strain OREX-4 did not respire nitrate but grew on lactate, with either arsenate or sulfate serving as the electron acceptor, and even preferred arsenate. Both arsenate and sulfate reduction were inhibited by molybdate. Strain OREX-4, a gram-positive bacterium with a hexagonal S-layer on its cell wall, metabolized compounds commonly used by sulfate reducers. Scorodite (FeAsO42. H2O) an arsenate-containing mineral, provided micromolar concentrations of arsenate that supported cell growth. Physiologically and phylogenetically, strain OREX-4 was far-removed from strains MIT-13 and SES-3: strain OREX-4 grew on different electron donors and electron acceptors, and fell within the gram-positive group of the Bacteria, whereas MIT-13 and SES-3 fell together in the epsilon-subdivision of the Proteobacteria. Together, these results suggest that organisms spread among diverse bacterial phyla can use arsenate as a terminal electron acceptor, and that dissimilatory arsenate reduction might occur in the sulfidogenic zone at arsenate concentrations of environmental interest. 16S rRNA sequence analysis indicated that strain OREX-4 is a new species of the genus Desulfotomaculum, and accordingly, the name Desulfotomaculum auripigmentum is proposed.}, keywords = {Arsenates, Bacteria, Anaerobic, Biotransformation, Geologic Sediments, Gram-Positive Endospore-Forming Rods, Molecular Sequence Data, Oxidation-Reduction, Phylogeny, RNA, Ribosomal, 16S, Substrate Specificity, Sulfates, Sulfides, Sulfur-Reducing Bacteria, Water Microbiology}, issn = {0302-8933}, author = {Newman, D K and Kennedy, E K and Coates, J D and Ahmann, D and Ellis, D J and Lovley, D R and Morel, F M} } @article {834, title = {Evolutionary comparisons of RecA-like proteins across all major kingdoms of living organisms.}, journal = {J Mol Evol}, volume = {44}, year = {1997}, month = {1997 May}, pages = {528-41}, abstract = {Protein sequences with similarities to Escherichia coli RecA were compared across the major kingdoms of eubacteria, archaebacteria, and eukaryotes. The archaeal sequences branch monophyletically and are most closely related to the eukaryotic paralogous Rad51 and Dmc1 groups. A multiple alignment of the sequences suggests a modular structure of RecA-like proteins consisting of distinct segments, some of which are conserved only within subgroups of sequences. The eukaryotic and archaeal sequences share an N-terminal domain which may play a role in interactions with other factors and nucleic acids. Several positions in the alignment blocks are highly conserved within the eubacteria as one group and within the eukaryotes and archaebacteria as a second group, but compared between the groups these positions display nonconservative amino acid substitutions. Conservation within the RecA-like core domain identifies possible key residues involved in ATP-induced conformational changes. We propose that RecA-like proteins derive evolutionarily from an assortment of independent domains and that the functional homologs of RecA in noneubacteria comprise an array of RecA-like proteins acting in series or cooperatively.}, keywords = {Amino Acid Sequence, Animals, Archaeal Proteins, Bacteria, Bacterial Proteins, Cell Cycle Proteins, Consensus Sequence, Conserved Sequence, DNA-Binding Proteins, Escherichia coli Proteins, Evolution, Molecular, Humans, Molecular Sequence Data, Phylogeny, Rad51 Recombinase, Rec A Recombinases, Sequence Alignment}, issn = {0022-2844}, author = {Brendel, V and Brocchieri, L and Sandler, S J and Clark, A J and Karlin, S} } @article {795, title = {Heterogeneity of the internal transcribed spacer (ITS-2) region within individual deer ticks.}, journal = {Insect Mol Biol}, volume = {6}, year = {1997}, month = {1997 May}, pages = {123-9}, abstract = {To determine whether nuclear rDNA sequences provide a useful means for assessing the structure of populations of Ixodes ticks, we compared variability among copies of an internal transcribed spacer (ITS-2) sequence within individual ticks to the variability between ticks. At least 4\% of the nucleotides comprising this sequence vary among the copies present within individual ticks. ITS-2 diversity in each of two ticks is nearly half as great as that reported between ticks from geographically disparate populations. Because individual ticks retain ancestral polymorphism, ITS-2 variation does not accurately reflect descent relationships among these ticks. Sequencing single copies of PCR-amplified ITS-2 therefore does not permit assessment of the phylogenetic relationships among the I. ricinus-like ticks in eastern North America. We recommend caution in future analyses, and emphasize the importance of procedures designed to ensure that the many paralogous copies of the rDNA cistron have been sufficiently homogenized by concerted evolutionary processes. Such precautionary measures will make certain that phylogenetic trees based on these gene sequences reflect the phyletic relatedness of the biological species.}, keywords = {Animals, Base Sequence, Deer, DNA, DNA, Ribosomal, Genetic Heterogeneity, Ixodes, Molecular Sequence Data}, issn = {0962-1075}, author = {Rich, S M and Rosenthal, B M and Telford, S R and Spielman, A and Hartl, D L and Ayala, F J} } @article {794, title = {Plasmodium falciparum antigenic diversity: evidence of clonal population structure.}, journal = {Proc Natl Acad Sci U S A}, volume = {94}, year = {1997}, month = {1997 Nov 25}, pages = {13040-5}, abstract = {Plasmodium falciparum, the agent of malignant malaria, is one of mankind{\textquoteright}s most severe scourges. Efforts to develop preventive vaccines or remedial drugs are handicapped by the parasite{\textquoteright}s rapid evolution of drug resistance and protective antigens. We examine 25 DNA sequences of the gene coding for the highly polymorphic antigenic circumsporozoite protein. We observe total absence of silent nucleotide variation in the two nonrepeated regions of the gene. We propose that this absence reflects a recent origin (within several thousand years) of the world populations of P. falciparum from a single individual; the amino acid polymorphisms observed in these nonrepeat regions would result from strong natural selection. Analysis of these polymorphisms indicates that: (i) the incidence of recombination events does not increase with nucleotide distance; (ii) the strength of linkage disequilibrium between nucleotides is also independent of distance; and (iii) haplotypes in the two nonrepeat regions are correlated with one another, but not with the central repeat region they span. We propose two hypotheses: (i) variation in the highly polymorphic central repeat region arises by mitotic intragenic recombination, and (ii) the population structure of P. falciparum is clonal--a state of affairs that persists in spite of the necessary stage of physiological sexuality that the parasite must sustain in the mosquito vector to complete its life cycle.}, keywords = {Amino Acid Sequence, Animals, Antigens, Protozoan, Molecular Sequence Data, Phylogeny, Plasmodium falciparum, Polymorphism, Genetic, Protozoan Proteins, Recombination, Genetic, Sequence Homology, Amino Acid}, issn = {0027-8424}, author = {Rich, S M and Hudson, R R and Ayala, F J} } @article {708, title = {Bound simian virus 40 translocates to caveolin-enriched membrane domains, and its entry is inhibited by drugs that selectively disrupt caveolae.}, journal = {Mol Biol Cell}, volume = {7}, year = {1996}, month = {1996 Nov}, pages = {1825-34}, abstract = {Simian virus 40 (SV40) entry leading to infection occurred only after the virus was at the cell surface for 1.5 to 2 h. SV40 infectious entry was not sensitive to cytosol acidification, a treatment that blocks endocytosis via clathrin-coated vesicles. Instead, SV40 infectious entry was blocked by treating cells with the phorbol ester PMA or nystatin, which selectively disrupts caveolae. In control experiments, transferrin internalization was sensitive to cytosol acidification but was not sensitive to PMA or nystatin. Also, absorbed transferrin entered cells within minutes. Finally, bound SV40 translocated to caveolin-enriched membrane complexes isolated by a Triton X-100 insolubility protocol. Treatment with nystatin did not impair SV40 binding but did block the partitioning of virus into the caveolin-enriched complexes.}, keywords = {Animals, Antigens, Polyomavirus Transforming, Caveolin 1, Caveolins, Cell Line, Cell Membrane, Coated Vesicles, Endocytosis, Membrane Proteins, Nystatin, Organelles, Simian virus 40, Tetradecanoylphorbol Acetate}, issn = {1059-1524}, author = {Anderson, H A and Chen, Y and Norkin, L C} } @article {836, title = {Differential suppression of priA2::kan phenotypes in Escherichia coli K-12 by mutations in priA, lexA, and dnaC.}, journal = {Genetics}, volume = {143}, year = {1996}, month = {1996 May}, pages = {5-13}, abstract = {First identified as an essential component of the phi X174 in vitro DNA replication system, PriA has ATPase, helicase, translocase, and primosome-assembly activities. priA1::kan strains of Escherichia coli are sensitive to UV irradiation, deficient in homologous recombination following transduction, and filamentous. priA2::kan strains have eightfold higher levels of uninduced SOS expression than wild type. We show that (1) priA1::kan strains have eightfold higher levels of uninduced SOS expression, (2) priA2::kan strains are UVS and Rec-, (3) lexA3 suppresses the high basal levels of SOS expression of a priA2::kan strain, and (4) plasmid-encoded priA300 (K230R), a mutant allele retaining only the primosome-assembly activity of priA+, restores both UVR and Rec+ phenotypes to a priA2::kan strain. Finally, we have isolated 17 independent UVR Rec+ revertants of priA2::kan strains that carry extragenic suppressors. All 17 map in the C-terminal half of the dnaC gene. DnaC loads the DnaB helicase onto DNA as a prelude for primosome assembly and DNA replication. We conclude that priA{\textquoteright}s primosome-assembly activity is essential for DNA repair and recombination and that the dnaC suppressor mutations allow these processes to occur in the absence of priA.}, keywords = {Bacterial Proteins, Bacteriophage phi X 174, beta-Galactosidase, Chromosomes, Bacterial, DNA Helicases, DNA Replication, DNA-Binding Proteins, Dose-Response Relationship, Radiation, Escherichia coli, Escherichia coli Proteins, Genes, Bacterial, Genetic Markers, Mutagenesis, Phenotype, Recombination, Genetic, Replication Protein A, Repressor Proteins, Serine Endopeptidases, Suppression, Genetic, Transduction, Genetic, Ultraviolet Rays}, issn = {0016-6731}, author = {Sandler, S J and Samra, H S and Clark, A J} } @article {707, title = {Extracellular simian virus 40 induces an ERK/MAP kinase-independent signalling pathway that activates primary response genes and promotes virus entry.}, journal = {J Gen Virol}, volume = {77 ( Pt 9)}, year = {1996}, month = {1996 Sep}, pages = {2173-82}, abstract = {Simian virus 40 (SV40) binding to growth-arrested cells activated an intracellular signalling pathway that induced the up-regulation of the primary response genes c-myc, c-jun and c-sis within 30 min and of JE within 90 min. The up-regulation of the primary response genes occurred in the presence of cycloheximide and when UV-inactivated SV40 was adsorbed to cells. SV40 binding did not activate Raf or mitogen-activated protein kinase (MAP/ERK1), or mobilize intracellular Ca2+. The SV40-induced up-regulation of c-myc and c-jun was blocked by the tyrosine kinase inhibitor, genistein, and by the protein kinase C (PKC) inhibitor, calphostin C, but not by expression of the MAP kinase-specific phosphatase, MKP-1. These results suggest that the SV40-induced signalling pathway includes the activities of a tyrosine kinase and a Ca(2+)-independent isoform of PKC, but not of Raf or MAP kinase. Finally, SV40 infectious entry into cells was specifically and reversibly blocked by genistein.}, keywords = {Animals, Calcium, Calcium-Calmodulin-Dependent Protein Kinases, Cell Cycle Proteins, Cell Line, Cercopithecus aethiops, Dual Specificity Phosphatase 1, Enzyme Inhibitors, Gene Expression Regulation, Viral, Genistein, Humans, Immediate-Early Proteins, Isoflavones, Mitogen-Activated Protein Kinase 3, Mitogen-Activated Protein Kinases, Naphthalenes, Phosphoprotein Phosphatases, Platelet-Derived Growth Factor, Protein Kinase C, Protein Phosphatase 1, Protein Tyrosine Phosphatases, Protein-Serine-Threonine Kinases, Proto-Oncogene Proteins, Proto-Oncogene Proteins c-jun, Proto-Oncogene Proteins c-myc, Proto-Oncogene Proteins c-raf, Proto-Oncogene Proteins c-sis, Signal Transduction, Simian virus 40, Up-Regulation}, issn = {0022-1317}, author = {Dangoria, N S and Breau, W C and Anderson, H A and Cishek, D M and Norkin, L C} } @article {630, title = {Geovibrio ferrireducens, a phylogenetically distinct dissimilatory Fe(III)-reducing bacterium.}, journal = {Arch Microbiol}, volume = {165}, year = {1996}, month = {1996 Jun}, pages = {370-6}, abstract = {A new, phylogenetically distinct, dissimilatory, Fe(III)-reducing bacterium was isolated from surface sediment of a hydrocarbon-contaminated ditch. The isolate, designated strain PAL-1, was an obligately anaerobic, non-fermentative, motile, gram-negative vibrio. PAL-1 grew in a defined medium with acetate as electron donor and ferric pyrophosphate, ferric oxyhydroxide, ferric citrate, Co(III)-EDTA, or elemental sulfur as sole electron acceptor. PAL-1 also used proline, hydrogen, lactate, propionate, succinate, fumarate, pyruvate, or yeast extract as electron donors for Fe(III) reduction. It is the first bacterium known to couple the oxidation of an amino acid to Fe(III) reduction. PAl-1 did not reduce oxygen, Mn(IV), U(VI), Cr(VI), nitrate, sulfate, sulfite, or thiosulfate with acetate as the electron donor. Cell suspensions of PAL-1 exhibited dithionite-reduced minus air-oxidized difference spectra that were characteristic of c-type cytochromes. Analysis of the 16S rRNA gene sequence of PAL-1 showed that the strain is not related to any of the described metal-reducing bacteria in the Proteobacteria and, together with Flexistipes sinusarabici, forms a separate line of descent within the Bacteria. Phenotypically and phylogenetically, strain PAl-1 differs from all other described bacteria, and represents the type strain of a new genus and species, Geovibrio ferrireducens.}, keywords = {Bacteria, Base Composition, Iron, Phylogeny}, issn = {0302-8933}, author = {Caccavo, F and Coates, J D and Rossello-Mora, R A and Ludwig, W and Schleifer, K H and Lovley, D R and McInerney, M J} } @article {631, title = {Hydrogen-based microbial ecosystems in the Earth.}, journal = {Science}, volume = {272}, year = {1996}, month = {1996 May 10}, pages = {896b}, issn = {0036-8075}, doi = {10.1126/science.272.5263.896b}, author = {Lovley, D R and Chapelle, F H} } @article {391, title = {Identification of a novel dihydrolipoyl dehydrogenase-binding protein in the pyruvate dehydrogenase complex of the anaerobic parasitic nematode, Ascaris suum.}, journal = {J Biol Chem}, volume = {271}, year = {1996}, month = {1996 Mar 8}, pages = {5451-7}, abstract = {A novel dihydrolipoyl dehydrogenase-binding protein (E3BP) which lacks an amino-terminal lipoyl domain, p45, has been identified in the pyruvate dehydrogenase complex (PDC) of the adult parasitic nematode, Ascaris suum. Sequence at the amino terminus of p45 exhibited significant similarity with internal E3-binding domains of dihydrolipoyl transacetylase (E2) and E3BP. Dissociation and resolution of a pyruvate dehydrogenase-depleted adult A. suum PDC in guanidine hydrochloride resulted in two E3-depleted E2 core preparations which were either enriched or substantially depleted of p45. Following reconstitution, the p45-enriched E2 core exhibited enhanced E3 binding, whereas, the p45-depleted E2 core exhibited dramatically reduced E3 binding. Reconstitution of either the bovine kidney or A. suum PDCs with the A. suum E3 suggested that the ascarid E3 was more sensitive to NADH inhibition when bound to the bovine kidney core. The expression of p45 was developmentally regulated and p45 was most abundant in anaerobic muscle. In contrast, E3s isolated from anaerobic muscle or aerobic second-stage larvae were identical. These results suggest that during the transition to anaerobic metabolism, E3 remains unchanged, but it appears that a novel E3BP, p45, is expressed which may help to maintain the activity of the PDC in the face of the elevated intramitochondrial NADH/NAD+ ratios associated with anaerobiosis.}, keywords = {Amino Acid Sequence, Anaerobiosis, Animals, Ascaris suum, Binding Sites, Carrier Proteins, Dihydrolipoamide Dehydrogenase, Electrophoresis, Polyacrylamide Gel, Flavin-Adenine Dinucleotide, Helminth Proteins, Kinetics, Larva, Molecular Sequence Data, NAD, Oxidation-Reduction, Pyruvate Dehydrogenase Complex, Sequence Homology, Amino Acid}, issn = {0021-9258}, author = {Klingbeil, M M and Walker, D J and Arnette, R and Sidawy, E and Hayton, K and Komuniecki, P R and Komuniecki, R} } @article {633, title = {Isolation of Geobacter species from diverse sedimentary environments.}, journal = {Appl Environ Microbiol}, volume = {62}, year = {1996}, month = {1996 May}, pages = {1531-6}, abstract = {In an attempt to better understand the microorganisms responsible for Fe(III) reduction in sedimentary environments, Fe(III)-reducing microorganisms were enriched for and isolated from freshwater aquatic sediments, a pristine deep aquifer, and a petroleum-contaminated shallow aquifer. Enrichments were initiated with acetate or toluene as the electron donor and Fe(III) as the electron acceptor. Isolations were made with acetate or benzoate. Five new strains which could obtain energy for growth by dissimilatory Fe(III) reduction were isolated. All five isolates are gram-negative strict anaerobes which grow with acetate as the electron donor and Fe(III) as the electron acceptor. Analysis of the 16S rRNA sequence of the isolated organisms demonstrated that they all belonged to the genus Geobacter in the delta subdivision of the Proteobacteria. Unlike the type strain, Geobacter metallireducens, three of the five isolates could use H2 as an electron donor for Fe(III) reduction. The deep subsurface isolate is the first Fe(III) reducer shown to completely oxidize lactate to carbon dioxide, while one of the freshwater sediment isolates is only the second Fe(III) reducer known that can oxidize toluene. The isolation of these organisms demonstrates that Geobacter species are widely distributed in a diversity of sedimentary environments in which Fe(III) reduction is an important process.}, keywords = {Base Sequence, Ferric Compounds, Gram-Negative Anaerobic Bacteria, Molecular Sequence Data, Phylogeny, RNA, Bacterial, RNA, Ribosomal, 16S, Water Microbiology}, issn = {0099-2240}, author = {Coates, J D and Phillips, E J and Lonergan, D J and Jenter, H and Lovley, D R} } @article {632, title = {Microbial degradation of hydrochlorofluorocarbons (CHCl2F and CHCl2CF3) in soils and sediments.}, journal = {Appl Environ Microbiol}, volume = {62}, year = {1996}, month = {1996 May}, pages = {1818-21}, abstract = {The ability of microorganisms to degrade trace levels of the hydrochlorofluorocarbons HCFC-21 and HCFC-123 was investigated. Methanotroph-linked oxidation of HCFC-21 was observed in aerobic soils, and anaerobic degradation of HCFC-21 occurred in freshwater and salt marsh sediments. Microbial degradation of HCFC-123 was observed in anoxic freshwater and salt marsh sediments, and the recovery of 1,1,1-trifluoro-2-chloroethane indicated the involvement of reductive dechlorination. No degradation of HCFC-123 was observed in aerobic soils. In some experiments, HCFCs were degraded at low (parts per billion) concentrations, raising the possibility that bacteria in nature remove HCFCs from the atmosphere.}, keywords = {Biodegradation, Environmental, Chlorofluorocarbons, Soil Microbiology}, issn = {0099-2240}, author = {Oremland, R S and Lonergan, D J and Culbertson, C W and Lovley, D R} } @article {796, title = {Mosaic structure of plasmids from natural populations of Escherichia coli.}, journal = {Genetics}, volume = {143}, year = {1996}, month = {1996 Jul}, pages = {1091-100}, abstract = {The distribution of plasmids related to the fertility factor F was examined in the ECOR reference collection of Escherichia coli. Probes specific for four F-related genes were isolated and used to survey the collection by DNA hybridization. To estimate the genetic diversity of genes in F-like plasmids, DNA sequences were obtained for four plasmid genes. The phylogenetic relationships among the plasmids in the ECOR strains is very different from that of the strains themselves. This finding supports the view that plasmid transfer has been frequent within and between the major groups of ECOR. Furthermore, the sequences indicate that recombination between genes in plasmids takes place at a considerably higher frequency than that observed for chromosomal genes. The plasmid genes, and by inference the plasmids themselves, are mosaic in structure with different regions acquired from different sources. Comparison of gene sequences from a variety of naturally occurring plasmids suggested a plausible donor of some of the recombinant regions as well as implicating a chi site in the mechanism of genetic exchange. The relatively high rate of recombination in F-plasmid genes suggests that conjugational gene transfer may play a greater role in bacterial population structure than previously appreciated.}, keywords = {Bacterial Proteins, Base Sequence, DNA Helicases, DNA, Bacterial, DNA-Binding Proteins, Escherichia coli, Escherichia coli Proteins, Evolution, Molecular, Membrane Proteins, Molecular Sequence Data, Mosaicism, Nucleic Acid Hybridization, Phylogeny, Plasmids, Polymorphism, Genetic, Proteins, Recombination, Genetic, Repressor Proteins, RNA-Binding Proteins, Trans-Activators}, issn = {0016-6731}, author = {Boyd, E F and Hill, C W and Rich, S M and Hartl, D L} } @article {706, title = {Multiple stages of virus-receptor interactions as shown by simian virus 40.}, journal = {Adv Exp Med Biol}, volume = {408}, year = {1996}, month = {1996}, pages = {159-67}, keywords = {Animals, Cell Membrane, Histocompatibility Antigens Class I, Humans, Receptors, Virus, Signal Transduction, Simian virus 40}, issn = {0065-2598}, author = {Norkin, L C and Anderson, H A} } @article {797, title = {A new Borrelia infecting Lone Star ticks.}, journal = {Lancet}, volume = {347}, year = {1996}, month = {1996 Jan 6}, pages = {67-8}, keywords = {Animals, Base Sequence, Borrelia, Borrelia Infections, Deer, Disease Vectors, DNA Primers, Maryland, Molecular Sequence Data, Ticks}, issn = {0140-6736}, author = {Armstrong, P M and Rich, S M and Smith, R D and Hartl, D L and Spielman, A and Telford, S R} } @article {837, title = {Overlapping functions for recF and priA in cell viability and UV-inducible SOS expression are distinguished by dnaC809 in Escherichia coli K-12.}, journal = {Mol Microbiol}, volume = {19}, year = {1996}, month = {1996 Feb}, pages = {871-80}, abstract = {The recF and priA genes have roles in DNA repair and homologous recombination. Mutations in these genes also cause decreases in cell viability and alterations in UV-inducible sulAp-lacZ (SOS) expression. To find out if the two genes are in the same or different pathways for viability and SOS expression, the phenotypes of the double mutant strains were studied. The recF priA double mutant showed a lower viability and SOS expression level than either of the single mutants. In the case of cell viability, recF missense mutations decreased viability of a priA2::kan strain two to five-fold whereas recF null priA2::kan double mutants were not viable at all. dnaC809, a mutation that suppresses the UV-sensitive (UVs and Rec- phenotypes of priA2::kan, restored cell viability, but not UV-inducible SOS expression, to a priA recF strain. Since recF is epistatic with recO and recR (recOR) for UV resistance, recOR mutations were also tested with priA2::kan. No overlap was found between recOR and priA for viability and SOS expression. It is concluded that priA and recF have two different overlapping functions in viability and SOS expression that are distinguishable by the effects of dnaC809. The role of recF in a priA2::kan strain in cell viability is a new function for recF and unlike recF{\textquoteright}s other roles in DNA repair and recombination, is independent of recOR. A new role for priA in UV-inducible SOS expression in a recF mutant is also defined.}, keywords = {Bacterial Proteins, DNA Repair, DNA, Bacterial, DNA-Binding Proteins, Escherichia coli, Escherichia coli Proteins, Genes, Bacterial, Lac Operon, Mutation, Recombinant Fusion Proteins, Recombination, Genetic, Replication Protein A, SOS Response (Genetics), Ultraviolet Rays}, issn = {0950-382X}, author = {Sandler, S J} } @article {1197, title = {Overview of hyperthermophiles and their heat-shock proteins.}, journal = {Adv Protein Chem}, volume = {48}, year = {1996}, month = {1996}, pages = {1-34}, keywords = {Archaea, Bacterial Proteins, Heat-Shock Proteins, Hot Temperature, Phylogeny}, issn = {0065-3233}, author = {Baross, J A and Holden, J F} } @article {635, title = {Oxidation of Polycyclic Aromatic Hydrocarbons under Sulfate-Reducing Conditions.}, journal = {Appl Environ Microbiol}, volume = {62}, year = {1996}, month = {1996 Mar}, pages = {1099-101}, abstract = {[(sup14)C]naphthalene and phenanthrene were oxidized to (sup14)CO(inf2) without a detectable lag under strict anaerobic conditions in sediments from San Diego Bay, San Diego, Calif., that were heavily contaminated with polycyclic aromatic hydrocarbons (PAHs) but not in less contaminated sediments. Sulfate reduction was necessary for PAH oxidation. These results suggest that the self-purification capacity of PAH-contaminated sulfate-reducing environments may be greater than previously recognized.}, issn = {0099-2240}, author = {Coates, J D and Anderson, R T and Lovley, D R} } @article {634, title = {Phylogenetic analysis of dissimilatory Fe(III)-reducing bacteria.}, journal = {J Bacteriol}, volume = {178}, year = {1996}, month = {1996 Apr}, pages = {2402-8}, abstract = {Evolutionary relationships among strictly anaerobic dissimilatory Fe(III)-reducing bacteria obtained from a diversity of sedimentary environments were examined by phylogenetic analysis of 16S rRNA gene sequences. Members of the genera Geobacter, Desulfuromonas, Pelobacter, and Desulfuromusa formed a monophyletic group within the delta subdivision of the class Proteobacteria. On the basis of their common ancestry and the shared ability to reduce Fe(III) and/or S0, we propose that this group be considered a single family, Geobacteraceae. Bootstrap analysis, characteristic nucleotides, and higher-order secondary structures support the division of Geobacteraceae into two subgroups, designated the Geobacter and Desulfuromonas clusters. The genus Desulfuromusa and Pelobacter acidigallici make up a distinct branch within the Desulfuromonas cluster. Several members of the family Geobacteraceae, none of which reduce sulfate, were found to contain the target sequences of probes that have been previously used to define the distribution of sulfate-reducing bacteria and sulfate-reducing bacterium-like microorganisms. The recent isolations of Fe(III)-reducing microorganisms distributed throughout the domain Bacteria suggest that development of 16S rRNA probes that would specifically target all Fe(III) reducers may not be feasible. However, all of the evidence suggests that if a 16S rRNA sequence falls within the family Geobacteraceae, then the organism has the capacity for Fe(III) reduction. The suggestion, based on geological evidence, that Fe(III) reduction was the first globally significant process for oxidizing organic matter back to carbon dioxide is consistent with the finding that acetate-oxidizing Fe(III) reducers are phylogenetically diverse.}, keywords = {Bacteria, Anaerobic, Base Sequence, DNA, Ribosomal, Ferric Compounds, Iron, Molecular Sequence Data, Oxidation-Reduction, Phylogeny, RNA, Ribosomal, 16S, Sequence Analysis, DNA, Sequence Homology, Nucleic Acid}, issn = {0021-9193}, author = {Lonergan, D J and Jenter, H L and Coates, J D and Phillips, E J and Schmidt, T M and Lovley, D R} } @article {636, title = {Rapid Anaerobic Benzene Oxidation with a Variety of Chelated Fe(III) Forms.}, journal = {Appl Environ Microbiol}, volume = {62}, year = {1996}, month = {1996 Jan}, pages = {288-91}, abstract = {Fe(III) chelated to such compounds as EDTA, N-methyliminodiacetic acid, ethanol diglycine, humic acids, and phosphates stimulated benzene oxidation coupled to Fe(III) reduction in anaerobic sediments from a petroleum-contaminated aquifer as effectively as or more effectively than nitrilotriacetic acid did in a previously demonstrated stimulation experiment. These results indicate that many forms of chelated Fe(III) might be applicable to aquifer remediation.}, issn = {0099-2240}, author = {Lovley, D R and Woodward, J C and Chapelle, F H} } @article {835, title = {recA-like genes from three archaean species with putative protein products similar to Rad51 and Dmc1 proteins of the yeast Saccharomyces cerevisiae.}, journal = {Nucleic Acids Res}, volume = {24}, year = {1996}, month = {1996 Jun 1}, pages = {2125-32}, abstract = {The process of homologous recombination has been documented in bacterial and eucaryotic organisms. The Escherichia coli RecA and Saccharomyces cerevisiae Rad51 proteins are the archetypal members of two related families of proteins that play a central role in this process. Using the PCR process primed by degenerate oligonucleotides designed to encode regions of the proteins showing the greatest degree of identity, we examined DNA from three organisms of a third phylogenetically divergent group, Archaea, for sequences encoding proteins similar to RecA and Rad51. The archaeans examined were a hyperthermophilic acidophile, Sulfolobus sofataricus (Sso); a halophile, Haloferax volcanii (Hvo); and a hyperthermophilic piezophilic methanogen, Methanococcus jannaschii (Mja). The PCR generated DNA was used to clone a larger genomic DNA fragment containing an open reading frame (orf), that we refer to as the radA gene, for each of the three archaeans. As shown by amino acid sequence alignments, percent amino acid identities and phylogenetic analysis, the putative proteins encoded by all three are related to each other and to both the RecA and Rad51 families of proteins. The putative RadA proteins are more similar to the Rad51 family (approximately 40\% identity at the amino acid level) than to the RecA family (approximately 20\%). Conserved sequence motifs, putative tertiary structures and phylogenetic analysis implied by the alignment are discussed. The 5{\textquoteright} ends of mRNA transcripts to the Sso radA were mapped. The levels of radA mRNA do not increase after treatment with UV irradiation as do recA and RAD51 transcripts in E.coli and S.cerevisiae. Hence it is likely that radA in this organism is a constitutively expressed gene and we discuss possible implications of the lack of UV-inducibility.}, keywords = {Amino Acid Sequence, Archaea, Base Sequence, Cell Cycle Proteins, DNA-Binding Proteins, Escherichia coli, Halobacteriaceae, Methanococcus, Models, Molecular, Molecular Sequence Data, Phylogeny, Rad51 Recombinase, Rec A Recombinases, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins, Sequence Alignment, Sulfolobus}, issn = {0305-1048}, author = {Sandler, S J and Satin, L H and Samra, H S and Clark, A J} } @article {640, title = {Benzene oxidation coupled to sulfate reduction.}, journal = {Appl Environ Microbiol}, volume = {61}, year = {1995}, month = {1995 Mar}, pages = {953-8}, abstract = {Highly reduced sediments from San Diego Bay, Calif., that were incubated under strictly anaerobic conditions metabolized benzene within 55 days when they were exposed initially to 1 (mu)M benzene. The rate of benzene metabolism increased as benzene was added back to the benzene-adapted sediments. When a [(sup14)C]benzene tracer was included with the benzene added to benzene-adapted sediments, 92\% of the added radioactivity was recovered as (sup14)CO(inf2). Molybdate, an inhibitor of sulfate reduction, inhibited benzene uptake and production of (sup14)CO(inf2) from [(sup14)C]benzene. Benzene metabolism stopped when the sediments became sulfate depleted, and benzene uptake resumed when sulfate was added again. The stoichiometry of benzene uptake and sulfate reduction was consistent with the hypothesis that sulfate was the principal electron acceptor for benzene oxidation. Isotope trapping experiments performed with [(sup14)C]benzene revealed that there was no production of such potential extracellular intermediates of benzene oxidation as phenol, benzoate, p-hydroxybenzoate, cyclohexane, catechol, and acetate. The results demonstrate that benzene can be oxidized in the absence of O(inf2), with sulfate serving as the electron acceptor, and suggest that some sulfate reducers are capable of completely oxidizing benzene to carbon dioxide without the production of extracellular intermediates. Although anaerobic benzene oxidation coupled to chelated Fe(III) has been documented previously, the study reported here provides the first example of a natural sediment compound that can serve as an electron acceptor for anaerobic benzene oxidation.}, issn = {0099-2240}, author = {Lovley, D R and Coates, J D and Woodward, J C and Phillips, E} } @article {641, title = {Bioremediation of organic and metal contaminants with dissimilatory metal reduction.}, journal = {J Ind Microbiol}, volume = {14}, year = {1995}, month = {1995 Feb}, pages = {85-93}, abstract = {Dissimilatory metal reduction has the potential to be a helpful mechanism for both intrinsic and engineered bioremediation of contaminated environments. Dissimilatory Fe(III) reduction is an important intrinsic process for removing organic contaminants from aquifers contaminated with petroleum or landfill leachate. Stimulation of microbial Fe(III) reduction can enhance the degradation of organic contaminants in ground water. Dissimilatory reduction of uranium, selenium, chromium, technetium, and possibly other metals, can convert soluble metal species to insoluble forms that can readily be removed from contaminated waters or waste streams. Reduction of mercury can volatilize mercury from waters and soils. Despite its potential, there has as yet been limited applied research into the use of dissimilatory metal reduction as a bioremediation tool.}, keywords = {Bacteria, Biodegradation, Environmental, Chromium, Hydrocarbons, Iron, Mercury, Metals, Oxidation-Reduction, Selenium, Uranium}, issn = {0169-4146}, author = {Lovley, D R} } @article {637, title = {Desulfuromonas palmitatis sp. nov., a marine dissimilatory Fe(III) reducer that can oxidize long-chain fatty acids.}, journal = {Arch Microbiol}, volume = {164}, year = {1995}, month = {1995 Dec}, pages = {406-13}, abstract = {Studies on the microorganisms living in hydrocarbon-contaminated sediments in San Diego Bay, California led to the isolation of a novel Fe(III)-reducing microorganism. This organism, designated strain SDBY1, was an obligately anaerobic, non-motile, non-flagellated, gram-negative rod. Strain SDBY1 conserves energy to support growth from the oxidation of acetate, lactate, succinate, fumarate, laurate, palmitate, or stearate. H2 was also oxidized with the reduction of Fe(III), but growth with H2 as the sole electron donor was not observed. In addition to various forms of soluble and insoluble Fe(III), strain SDBY1 also coupled growth to the reduction of fumarate, Mn(IV), or S0. Air-oxidized minus dithionite-reduced difference spectra exhibited peaks at 552.8, 523.6, and 422.8 nm, indicative of c-type cytochrome(s). Strain SDBY1 shares physiological characteristics with organisms in the genera Geobacter, Pelobacter, and Desulfuromonas. Detailed analysis of the 16S rRNA sequence indicated that strain SDBY1 should be placed in the genus Desulfuromonas. The new species name Desulfuromonas palmitatis is proposed. D. palmitatis is only the second marine organism found (after D. acetoxidans) to oxidize multicarbon organic compounds completely to carbon dioxide with Fe(III) as an electron acceptor and provides the first pure culture model for the oxidation of long-chain fatty acids coupled to Fe(III) reduction.}, keywords = {Electron Transport, Fatty Acids, Gram-Negative Anaerobic Bacteria, Iron, Microscopy, Electron, Scanning, Oxidation-Reduction, Phylogeny, RNA, Bacterial, RNA, Ribosomal, 16S, Water Microbiology}, issn = {0302-8933}, author = {Coates, J D and Lonergan, D J and Philips, E J and Jenter, H and Lovley, D R} } @article {798, title = {Discriminating between Ixodes ticks by means of mitochondrial DNA sequences.}, journal = {Mol Phylogenet Evol}, volume = {4}, year = {1995}, month = {1995 Dec}, pages = {361-5}, abstract = {Ticks of the genus Ixodes have recently assumed prominence because they frequently serve as vectors of important zoonoses, including Lyme disease and babesiosis. The morphological characteristics that have been used in their identification often are ambiguous and are useful solely at a particular stage of development. Here we report the DNA sequence of the mitochondrially encoded 16S rRNA gene of nine different Ixodes ticks and an outgroup from another genus, Dermacentor. The sequences readily discriminate between these ticks. Samples of I. dammini from the northeastern and upper midwestern United States differ from southeastern I. scapularis at about 2\% of the nucleotides. This difference is about half that separating other members of the I. ricinus group of species, but exceeds typical levels of intraspecific variation. Two major clades exist within the I. ricinus complex. One includes I. cookei, I. hexagonus, and I. angustus. The other includes I. persulcatus, I. pacificus, I. muris, I. ricinus, I. scapularis, and I. dammini. We conclude that mtDNA sequences are useful for unravelling the systematics of these important vectors of human disease.}, keywords = {Animals, Arachnid Vectors, Base Sequence, Dermacentor, DNA Primers, DNA, Mitochondrial, Genes, Humans, Ixodes, Molecular Sequence Data, Phylogeny, Polymerase Chain Reaction, RNA, Ribosomal, 16S, Species Specificity}, issn = {1055-7903}, doi = {10.1006/mpev.1995.1033}, author = {Caporale, D A and Rich, S M and Spielman, A and Telford, S R and Kocher, T D} } @article {799, title = {Distribution of the Ixodes ricinus-like ticks of eastern North America.}, journal = {Proc Natl Acad Sci U S A}, volume = {92}, year = {1995}, month = {1995 Jul 3}, pages = {6284-8}, abstract = {We analyzed the geographic distribution of the Ixodes ricinus-like ticks in eastern North America by comparing the mitochondrial 16S rDNA sequences of specimens sampled directly from the field during the 1990s. Two distinct lineages are evident. The southern clade includes ticks from the southeastern and middle-eastern regions of the United States. The range of the northern clade, which appears to have been restricted to the northeastern region until the mid-1900s, now extends throughout the northeastern and middle-eastern regions. These phyletic units correspond to northern and southern taxa that have previously been assigned specific status as Ixodes dammini and Ixodes scapularis, respectively. The expanding range of I. dammini appears to drive the present outbreaks of zoonotic disease in eastern North America that include Lyme disease and human babesiosis.}, keywords = {Animals, Base Sequence, Demography, DNA Primers, DNA, Mitochondrial, DNA, Ribosomal, Geography, Mitochondria, Molecular Sequence Data, Phylogeny, Polymerase Chain Reaction, Population, RNA, Ribosomal, 16S, Ticks, United States}, issn = {0027-8424}, author = {Rich, S M and Caporale, D A and Telford, S R and Kocher, T D and Hartl, D L and Spielman, A} } @article {639, title = {Fe(III) and S0 reduction by Pelobacter carbinolicus.}, journal = {Appl Environ Microbiol}, volume = {61}, year = {1995}, month = {1995 Jun}, pages = {2132-8}, abstract = {There is a close phylogenetic relationship between Pelobacter species and members of the genera Desulfuromonas and Geobacter, and yet there has been a perplexing lack of physiological similarities. Pelobacter species have been considered to have a fermentative metabolism. In contrast, Desulfuromonas and Geobacter species have a respiratory metabolism with Fe(III) serving as the common terminal electron acceptor in all species. However, the ability of Pelobacter species to reduce Fe(III) had not been previously evaluated. When a culture of Pelobacter carbinolicus that had grown by fermentation of 2,3-butanediol was inoculated into the same medium supplemented with Fe(III), the Fe(III) was reduced. There was less accumulation of ethanol and more production of acetate in the presence of Fe(III). P. carbinolicus grew with ethanol as the sole electron donor and Fe(III) as the sole electron acceptor. Ethanol was metabolized to acetate. Growth was also possible on Fe(III) with the oxidation of propanol to propionate or butanol to butyrate if acetate was provided as a carbon source. P. carbinolicus appears capable of conserving energy to support growth from Fe(III) respiration as it also grew with H2 or formate as the electron donor and Fe(III) as the electron acceptor. Once adapted to Fe(III) reduction, P. carbinolicus could also grow on ethanol or H2 with S0 as the electron acceptor. P. carbinolicus did not contain detectable concentrations of the c-type cytochromes that previous studies have suggested are involved in electron transport to Fe(III) in other organisms that conserve energy to support growth from Fe(III) reduction.(ABSTRACT TRUNCATED AT 250 WORDS)}, keywords = {Bacteria, Anaerobic, Base Sequence, Butylene Glycols, Culture Media, Ethanol, Ferrous Compounds, Hydrogen, Molecular Sequence Data, Oxidation-Reduction, Phylogeny, RNA, Ribosomal, 16S, Sulfur}, issn = {0099-2240}, author = {Lovley, D R and Phillips, E J and Lonergan, D J and Widman, P K} } @article {638, title = {Growth of Strain SES-3 with Arsenate and Other Diverse Electron Acceptors.}, journal = {Appl Environ Microbiol}, volume = {61}, year = {1995}, month = {1995 Oct}, pages = {3556-61}, abstract = {The selenate-respiring bacterial strain SES-3 was able to use a variety of inorganic electron acceptors to sustain growth. SES-3 grew with the reduction of arsenate to arsenite, Fe(III) to Fe(II), or thiosulfate to sulfide. It also grew in medium in which elemental sulfur, Mn(IV), nitrite, trimethylamine N-oxide, or fumarate was provided as an electron acceptor. Growth on oxygen was microaerophilic. There was no growth with arsenite or chromate. Washed suspensions of cells grown on selenate or nitrate had a constitutive ability to reduce arsenate but were unable to reduce arsenite. These results suggest that strain SES-3 may occupy a niche as an environmental opportunist by being able to take advantage of a diversity of electron acceptors.}, issn = {0099-2240}, author = {Laverman, A M and Blum, J S and Schaefer, J K and Phillips, E and Lovley, D R and Oremland, R S} } @article {838, title = {recO and recR mutations delay induction of the SOS response in Escherichia coli.}, journal = {Mol Gen Genet}, volume = {246}, year = {1995}, month = {1995 Jan 20}, pages = {254-8}, abstract = {RecF, RecO and RecR, three of the important proteins of the RecF pathway of recombination, are also needed for repair of DNA damage due to UV irradiation. recF mutants are not proficient in cleaving LexA repressor in vivo following DNA damage: therefore they show a delay of induction of the SOS response. In this communication, by measuring the in vivo levels of LexA repressor using anti-LexA antibodies, we show that recO and recR mutant strains are also not proficient in LexA cleavage reactions. In addition, we show that recO and recR mutations delay induction of beta-galactosidase activity expressed from a lexA-regulated promoter following exposure of cells to UV, thus further supporting the idea that recF, recO and recR gene products are needed for induction of the SOS response.}, keywords = {Bacterial Proteins, beta-Galactosidase, DNA-Binding Proteins, Escherichia coli, Escherichia coli Proteins, Gene Expression Regulation, Bacterial, Mutation, Promoter Regions, Genetic, Recombinant Fusion Proteins, Repressor Proteins, Serine Endopeptidases, SOS Response (Genetics), Ultraviolet Rays}, issn = {0026-8925}, author = {Hegde, S and Sandler, S J and Clark, A J and Madiraju, M V} } @article {705, title = {Relationship between expression of epidermal growth factor and simian virus 40 T antigen in a line of transgenic mice.}, journal = {Transgenic Res}, volume = {4}, year = {1995}, month = {1995 Sep}, pages = {306-14}, abstract = {The pattern of expression of the simian virus 40 (SV40) T antigen gene and resultant dysplasia were re-examined in a line of transgenic mice in which the T antigen gene was under the control of the SV40 early promoter. We found that T antigen expression in the kidney, and resulting dysplastic lesions, occurred exclusively in the distal convoluted tubules and the ascending limbs of Henle. Epidermal growth factor (EGF) expression in the kidney of normal mice was similarly immunolocalized. The correlation between high EGF immunoreactivity in normal mouse tissues and T antigen expression in the transgenic counterpart was also seen in the choroid plexus epithelium and in the submandibular glands of male mice. T antigen was not found in the submandibular gland of transgenic females. Similarly, EGF was only rarely detected in the normal female submandibular gland. In contrast to the correlation between T antigen expression in the transgenic mice and EGF expression in the corresponding tissues of the normal mice, within the dysplastic lesions of the transgenic mice EGF expression was severely diminished. Adenocarcinomas of the male submandibular gland from another line of transgenic mice that expresses the Int-1 transgene, showed similarly reduced levels of immunostaining for EGF. Thus, reduced expression of EGF might be a general feature of dysplasia and tumorigenesis in those tissues that normally express EGF.}, keywords = {Adenocarcinoma, Animals, Antigens, Viral, Tumor, Cattle, Choroid Plexus, Cricetinae, Epidermal Growth Factor, Female, Gene Expression Regulation, Neoplastic, Kidney, Kidney Tubules, Distal, Male, Mice, Mice, Transgenic, Proto-Oncogene Proteins, Simian virus 40, Submandibular Gland, Submandibular Gland Neoplasms, Wnt Proteins, Wnt1 Protein, Zebrafish Proteins}, issn = {0962-8819}, author = {Lafond, R E and Giammalvo, J T and Norkin, L C} } @article {704, title = {Virus receptors: implications for pathogenesis and the design of antiviral agents.}, journal = {Clin Microbiol Rev}, volume = {8}, year = {1995}, month = {1995 Apr}, pages = {293-315}, abstract = {A virus initiates infection by attaching to its specific receptor on the surface of a susceptible host cell. This prepares the way for the virus to enter the cell. Consequently, the expression of the receptor on specific cells and tissues of the host is a major determinant of the route of entry of the virus into the host and of the patterns of virus spread and pathogenesis in the host. This review emphasizes the virus-receptor interactions of human immunodeficiency virus, the rhinoviruses, the herpesviruses, and the coronaviruses. These interactions are often found to be complex and dynamic, involving multiple sites or factors on both the virus and the host cell. Also, the receptor may play an important role in virus entry per se in addition to its role in virus binding. In the cases of human immunodeficiency virus and the rhinoviruses, ingenious approaches to therapeutic strategies based on inhibiting virus attachment and entry are under development and in clinical trials.}, keywords = {Antigens, CD4, Antiviral Agents, Coronavirus, Drug Design, Herpesviridae, HIV, Humans, Intercellular Adhesion Molecule-1, Models, Molecular, Picornaviridae, Receptors, Virus, Retroviridae}, issn = {0893-8512}, author = {Norkin, L C} } @article {642, title = {Geobacter sulfurreducens sp. nov., a hydrogen- and acetate-oxidizing dissimilatory metal-reducing microorganism.}, journal = {Appl Environ Microbiol}, volume = {60}, year = {1994}, month = {1994 Oct}, pages = {3752-9}, abstract = {A dissimilatory metal- and sulfur-reducing microorganism was isolated from surface sediments of a hydrocarbon-contaminated ditch in Norman, Okla. The isolate, which was designated strain PCA, was an obligately anaerobic, nonfermentative nonmotile, gram-negative rod. PCA grew in a defined medium with acetate as an electron donor and ferric PPi, ferric oxyhydroxide, ferric citrate, elemental sulfur, Co(III)-EDTA, fumarate, or malate as the sole electron acceptor. PCA also coupled the oxidation of hydrogen to the reduction of Fe(III) but did not reduce Fe(III) with sulfur, glucose, lactate, fumarate, propionate, butyrate, isobutyrate, isovalerate, succinate, yeast extract, phenol, benzoate, ethanol, propanol, or butanol as an electron donor. PCA did not reduce oxygen, Mn(IV), U(VI), nitrate, sulfate, sulfite, or thiosulfate with acetate as the electron donor. Cell suspensions of PCA exhibited dithionite-reduced minus air-oxidized difference spectra which were characteristic of c-type cytochromes. Phylogenetic analysis of the 16S rRNA sequence placed PCA in the delta subgroup of the proteobacteria. Its closest known relative is Geobacter metallireducens. The ability to utilize either hydrogen or acetate as the sole electron donor for Fe(III) reduction makes strain PCA a unique addition to the relatively small group of respiratory metal-reducing microorganisms available in pure culture. A new species name, Geobacter sulfurreducens, is proposed.}, keywords = {Acetic Acid, Acetic Acids, Base Sequence, DNA Primers, DNA, Bacterial, DNA, Ribosomal, Electron Transport, Gram-Negative Anaerobic Bacteria, Hydrogen, Metals, Microscopy, Electron, Molecular Sequence Data, Oxidation-Reduction, Phylogeny, RNA, Bacterial, RNA, Ribosomal, 16S, Soil Microbiology}, issn = {0099-2240}, author = {Caccavo, F and Lonergan, D J and Lovley, D R and Davis, M and Stolz, J F and McInerney, M J} } @article {842, title = {Homologous genetic recombination: the pieces begin to fall into place.}, journal = {Crit Rev Microbiol}, volume = {20}, year = {1994}, month = {1994}, pages = {125-42}, abstract = {One of the authors (AJC) acknowledges with gratitude the important role Fernando Bastarrachea played in the author{\textquoteright}s discovery that E. coli could carry out homologous genetic recombination by multiple pathways. This in turn led to the discovery of several genes, including recF, recO, and recR, whose role in recombination would not otherwise have been detected. Subsequent genetic and biochemical studies have led to a general formulation in which there are multiple nucleolytic ways to achieve a presynaptic intermediate bound to RecA protein. Postsynaptic events in the general formulation occur by means of multiple branch migration enzymes to form Holliday DNA structures and a specific nuclease to cleave them. The general formulation is built on synapsis catalyzed by RecA protein. A second RecA-independent synapsis catalyzed by RecT (and RecE?) protein is now under study and a third type independent of both RecA and RecT has apparently been discovered. How these will affect the general formulation remains to be seen. Some proteins, most prominently RecF, RecO, and RecR, have no role in the general formulation. The hypothesis is presented that these proteins act as a switch between replication and recombination by helping to convert replication to recombination intermediates. Universality of the general formulation is supported by the widespread occurrence of recA, recB, recC, and recD genes among bacteria. Recent discovery of recA-like genes in several eukaryotes further supports its universality. We have contributed additional support by sequencing a recA-like gene from an archaeal species, thus making it plausible that the mechanism of synapsis worked out for E. coli RecA protein will hold for all three organismal domains. The boundaries of the puzzle of homologous genetic recombination therefore seem complete and the pieces to the complex picture they encompass are falling into place.}, keywords = {DNA Replication, Escherichia coli, Genes, Bacterial, Rec A Recombinases, Recombination, Genetic}, issn = {1040-841X}, doi = {10.3109/10408419409113552}, author = {Clark, A J and Sandler, S J} } @article {840, title = {Mutational analysis of sequences in the recF gene of Escherichia coli K-12 that affect expression.}, journal = {J Bacteriol}, volume = {176}, year = {1994}, month = {1994 Jul}, pages = {4011-6}, abstract = {The level of translation of recF-lacZ fusions is reduced 20-fold by nucleotides 49 to 146 of recF. In this region of recF, we found a previously described ribosome-interactive sequence called epsilon and a hexapyrimidine tract located just upstream of the epsilon sequence. Mutational studies indicate that the hexapyrimidine sequence is involved in at least some of the reduced translation. When the hexapyrimidine sequence is mutant, mutating epsilon increases the level of translation maximally. We ruled out the possibility that ribosome frameshifting explains most of the effect of these two sequences on expression and suspect that multiple mechanisms may be responsible. In a separate report, we show that mutations in the hexapyrimidine tract and epsilon increase expression of the full-sized recF gene.}, keywords = {Bacterial Proteins, Base Sequence, DNA Mutational Analysis, DNA-Binding Proteins, Escherichia coli, Escherichia coli Proteins, Gene Expression Regulation, Bacterial, Genes, Bacterial, Half-Life, Lac Operon, Molecular Sequence Data, Protein Biosynthesis, Recombinant Fusion Proteins, Regulatory Sequences, Nucleic Acid, Ribosomes}, issn = {0021-9193}, author = {Sandler, S J and Clark, A J} } @article {645, title = {Novel processes for anaerobic sulfate production from elemental sulfur by sulfate-reducing bacteria.}, journal = {Appl Environ Microbiol}, volume = {60}, year = {1994}, month = {1994 Jul}, pages = {2394-9}, abstract = {Sulfate reducers and related organisms which had previously been found to reduce Fe(III) with H(2) or organic electron donors oxidized S to sulfate when Mn(IV) was provided as an electron acceptor. Organisms catalyzing this reaction in washed cell suspensions included Desulfovibrio desulfuricans, Desulfomicrobium baculatum, Desulfobacterium autotrophicum, Desulfuromonas acetoxidans, and Geobacter metallireducens. These organisms produced little or no sulfate from S with Fe(III) as a potential electron acceptor or in the absence of an electron acceptor. In detailed studies with Desulfovibrio desulfuricans, the stoichiometry of sulfate and Mn(II) production was consistent with the reaction S + 3 MnO(2) + 4H-->SO(4) + 3Mn(II) + 2H(2)O. None of the organisms evaluated could be grown with S as the sole electron donor and Mn(IV) as the electron acceptor. In contrast to the other sulfate reducers evaluated, Desulfobulbus propionicus produced sulfate from S in the absence of an electron acceptor and Fe(III) oxide stimulated sulfate production. Sulfide also accumulated in the absence of Mn(IV) or Fe(III). The stoichiometry of sulfate and sulfide production indicated that Desulfobulbus propionicus disproportionates S as follows: 4S + 4H(2)O-->SO(4) + 3HS + 5 H. Growth of Desulfobulbus propionicus with S as the electron donor and Fe(III) as a sulfide sink and/or electron acceptor was very slow. The S oxidation coupled to Mn(IV) reduction described here provides a potential explanation for the Mn(IV)-dependent sulfate production that previous studies have observed in anoxic marine sediments. Desulfobulbus propionicus is the first example of a pure culture known to disproportionate S.}, issn = {0099-2240}, author = {Lovley, D R and Phillips, E J} } @article {841, title = {RecOR suppression of recF mutant phenotypes in Escherichia coli K-12.}, journal = {J Bacteriol}, volume = {176}, year = {1994}, month = {1994 Jun}, pages = {3661-72}, abstract = {The recF, recO, and recR genes form the recFOR epistasis group for DNA repair. recF mutants are sensitive to UV irradiation and fail to properly induce the SOS response. Using plasmid derivatives that overexpress combinations of the recO+ and recR+ genes, we tested the hypothesis that high-level expression of recO+ and recR+ (recOR) in vivo will indirectly suppress the recF mutant phenotypes mentioned above. We found that overexpression of just recR+ from the plasmid will partially suppress both phenotypes. Expression of the chromosomal recO+ gene is essential for the recR+ suppression. Hence we call this RecOR suppression of recF mutant phenotypes. RecOR suppression of SOS induction is more efficient with recO+ expression from a plasmid than with recO+ expression from the chromosome. This is not true for RecOR suppression of UV sensitivity (the two are equal). Comparison of RecOR suppression with the suppression caused by recA801 and recA803 shows that RecOR suppression of UV sensitivity is more effective than recA803 suppression and that RecOR suppression of UV sensitivity, like recA801 suppression, requires recJ+. We present a model that explains the data and proposes a function for the recFOR epistasis group in the induction of the SOS response and recombinational DNA repair.}, keywords = {Bacterial Proteins, Base Sequence, DNA Damage, DNA Repair, DNA-Binding Proteins, Epistasis, Genetic, Escherichia coli, Escherichia coli Proteins, Models, Genetic, Molecular Sequence Data, Phenotype, Plasmids, SOS Response (Genetics), Suppression, Genetic, Ultraviolet Rays}, issn = {0021-9193}, author = {Sandler, S J and Clark, A J} } @article {646, title = {Reduction of Chromate by Desulfovibrio vulgaris and Its c(3) Cytochrome.}, journal = {Appl Environ Microbiol}, volume = {60}, year = {1994}, month = {1994 Feb}, pages = {726-8}, abstract = {Washed cell suspensions of Desulfovibrio vulgaris rapidly reduced Cr(VI) to Cr(III) with H(2) as the electron donor. The c(3) cytochrome from this organism functioned as a Cr(VI) reductase. D. vulgaris may have advantages over previously described Cr(VI) reducers for the bioremediation of Cr(VI)-contaminated waters.}, issn = {0099-2240}, author = {Lovley, D R and Phillips, E J} } @article {643, title = {Stimulated anoxic biodegradation of aromatic hydrocarbons using Fe(III) ligands.}, journal = {Nature}, volume = {370}, year = {1994}, month = {1994 Jul 14}, pages = {128-31}, abstract = {Contamination of ground waters with water-soluble aromatic hydrocarbons, common components of petroleum pollution, often produces anoxic conditions under which microbial degradation of the aromatics is slow. Oxygen is often added to contaminated ground water to stimulate biodegradation, but this can be technically difficult and expensive. Insoluble Fe(III) oxides, which are generally abundant in shallow aquifers, are alternative potential oxidants, but are difficult for microorganisms to access. Here we report that adding organic ligands that bind to Fe(III) dramatically increases its bioavailability, and that in the presence of these ligands, rates of degradation of aromatic hydrocarbons in anoxic aquifer sediments are comparable to those in oxic sediments. We find that even benzene, which is notoriously refractory in the absence of oxygen, can be rapidly degraded. Our results suggest that increasing the bioavailability of Fe(III) by adding suitable ligands provides a potential alternative to oxygen addition for the bioremediation of petroleum-contaminated aquifers.}, keywords = {Benzene, Biodegradation, Environmental, Ferric Compounds, Hydrocarbons, Ligands, Methane, Nitrilotriacetic Acid, Oxidation-Reduction, Toluene, Water Pollutants, Chemical}, issn = {0028-0836}, doi = {10.1038/370128a0}, author = {Lovley, D R and Woodward, J C and Chapelle, F H} } @article {839, title = {Studies on the mechanism of reduction of UV-inducible sulAp expression by recF overexpression in Escherichia coli K-12.}, journal = {Mol Gen Genet}, volume = {245}, year = {1994}, month = {1994 Dec 15}, pages = {741-9}, abstract = {UV-inducible sulAp expression, an indicator of the SOS response, is reduced by recF+ overexpression in vivo. Different DNA-damaging agents and amounts of RecO and RecR were tested for their effects on this phenotype. It was found that recF+ overexpression reduced sulAp expression after DNA damage by mitomycin C or nalidixic acid, recO+ and recR+ overexpression partially suppressed the reduction of UV-induced sulAp expression caused by recF+ overexpression. The requirement for ATP binding to RecF to produce the phenotype was tested by genetically altering the putative phosphate binding cleft of recF in a way that should prevent the mutant recF protein from binding ATP. It was found that a change of lysine to glutamine at codon 36 results in a mutant recF protein (RecF4115) that is unable to reduce UV-inducible sulAp expression when overproduced. It is inferred from these results that recF overexpression may reduce UV-inducible sulAp expression by a mechanism that is sensitive to the ability of RecF to bind ATP and to the levels of RecO and RecR (RecOR) in the cell, but not to the type of DNA damage per se. Models are explored that can explain how recF+ overexpression reduces UV induction of sulAp and how RecOR overproduction might suppress this phenotype.}, keywords = {Bacterial Proteins, Base Sequence, DNA Primers, DNA-Binding Proteins, Escherichia coli, Escherichia coli Proteins, Gene Expression Regulation, Bacterial, Genes, Bacterial, Molecular Sequence Data, Mutagenesis, Site-Directed, Restriction Mapping, RNA, Messenger, SOS Response (Genetics), Structure-Activity Relationship, Ultraviolet Rays}, issn = {0026-8925}, author = {Sandler, S J} } @article {644, title = {Use of dissolved h2 concentrations to determine distribution of microbially catalyzed redox reactions in anoxic groundwater.}, journal = {Environ Sci Technol}, volume = {28}, year = {1994}, month = {1994 Jul 1}, pages = {1205-10}, issn = {0013-936X}, doi = {10.1021/es00056a005}, author = {Lovley, D R and Chapelle, F H and Woodward, J C} } @article {649, title = {Anaerobes into heavy metal: Dissimilatory metal reduction in anoxic environments.}, journal = {Trends Ecol Evol}, volume = {8}, year = {1993}, month = {1993 Jun}, pages = {213-7}, abstract = {Within the last decade, a novel form of microbial metabolism of major environmental significance has been elucidated. In this process, known as dissimilatory metal reduction, specialized microorganisms, living in anoxic aquatic sediments and ground water, oxidize organic compounds to carbon dioxide with metals serving as the oxidant. Recent studies have demonstrated that this metabolism explains a number of important geochemical phenomena in ancient and modern sedimentary environments, affecting not only the cycling of metals but also the fate of organic matter. Furthermore, this metabolism may have practical application in remediation of environments contaminated with toxic metals and/or organics.}, issn = {0169-5347}, doi = {10.1016/0169-5347(93)90102-U}, author = {Lovley, D R} } @article {392, title = {Characterization of cDNA clones for the 2-methyl branched-chain enoyl-CoA reductase. An enzyme involved in branched-chain fatty acid synthesis in anaerobic mitochondria of the parasitic nematode Ascaris suum.}, journal = {J Biol Chem}, volume = {268}, year = {1993}, month = {1993 Oct 25}, pages = {22391-6}, abstract = {The 2-methyl branched-chain enoyl-CoA reductase plays a pivotal role in the reversal of beta-oxidation operating in anaerobic mitochondria of the parasitic nematode Ascaris suum. An affinity-purified polyclonal anti-serum against the reductase was used to screen a cDNA library constructed in lambda gt11 with poly(A)+ RNA from adult A. suum muscle. A 1.2-kilobase partial cDNA clone was isolated, subcloned, and sequenced in both directions. Additional sequence at the 5{\textquoteright} end of the mRNA was determined by the RACE (rapid amplification of cDNA ends) procedure. Nucleotide sequence analysis of the cDNAs revealed the 22-nucleotide trans-spliced leader sequence characteristic of many nematode mRNAs, an open reading frame of 1236 nucleotides and a 3{\textquoteright}-untranslated sequence of 109 nucleotides including a short poly(A) tail 14 nucleotides from a polyadenylation signal (AATAAA). The open reading frame encoded a 396-amino acid sequence (M(r) 43,046) including a 16-amino acid leader peptide. Two-dimensional gel electrophoresis of the purified reductase yielded multiple spots with two distinct but overlapping amino-terminal amino acid sequences. Both sequences overlapped with the sequence predicted from the mRNA, and one of the sequences was identical to the predicted sequence. Comparison of the ascarid sequence with that of mammalian acyl-CoA dehydrogenases revealed a high degree of sequence identity, suggesting that these enzymes may have evolved from a common ancestral gene even though the ascarid enzyme functions as a reductase, not as a dehydrogenase. Immunoblotting of A. suum larval stages and adult tissues with antisera that cross-reacted with each of the spots separated on two-dimensional gels suggested that the reductase was only found in adult muscle. Northern blotting using the partial cDNA revealed a hybridization band of about 1.5 kilobases and also suggested that the enzyme was tissue-specific and developmentally regulated in agreement with the results of the immunoblotting.}, keywords = {Amino Acid Sequence, Anaerobiosis, Animals, Ascaris suum, Base Sequence, Cloning, Molecular, Consensus Sequence, DNA, DNA Primers, DNA, Complementary, Fatty Acid Desaturases, Gene Library, Humans, Mitochondria, Molecular Sequence Data, Oligonucleotides, Antisense, Oxidoreductases, Oxidoreductases Acting on CH-CH Group Donors, Poly A, RNA, RNA, Messenger, Sequence Homology, Amino Acid}, issn = {0021-9258}, author = {Duran, E and Komuniecki, R W and Komuniecki, P R and Wheelock, M J and Klingbeil, M M and Ma, Y C and Johnson, K R} } @article {801, title = {Competence of Peromyscus maniculatus (Rodentia: Cricetidae) as a reservoir host for Borrelia burgdorferi (Spirochaetares: Spirochaetaceae) in the wild.}, journal = {J Med Entomol}, volume = {30}, year = {1993}, month = {1993 May}, pages = {614-8}, abstract = {Although capable of maintaining and transmitting Borrelia burgdorferi Johnson, Schmidt, Hyde, Steigerwalt \& Brenner, the causative spirochete of Lyme disease, in the laboratory, the specific ability of deer mice, Peromyscus maniculatus Le Conte, to support this zoonosis has not been established. Demonstration that P. maniculatus is a competent reservoir host in the wild would indicate that the spread of Lyme disease is not limited to the range of the primary reservoir host, P. leucopus Rafinesque. Isle au Haut, an offshore Maine island upon which the vector tick Ixodes dammini Spielman, Clifford, Piesman \& Corwin has become established, supports an isolated population of mice that are exclusively P. maniculatus. We examined the reservoir competence of this species by comparing infection rates of B. burgdorferi among juvenile ticks removed from livetrapped mice on this island with those removed from P. leucopus obtained at a mainland site endemic for Lyme disease. Equivalent rates of infection among engorged larval ticks, survival of infection through the larval-nymphal molt, and the isolation of B. burgdorferi from mice at both sites attest to the reservoir competence of P. maniculatus.}, keywords = {Animals, Borrelia burgdorferi Group, Host-Parasite Interactions, Lyme Disease, Peromyscus, Ticks}, issn = {0022-2585}, author = {Rand, P W and Lacombe, E H and Smith, R P and Rich, S M and Kilpatrick, C W and Dragoni, C A and Caporale, D} } @article {648, title = {Composition of Non-Microbially Reducible Fe(III) in Aquatic Sediments.}, journal = {Appl Environ Microbiol}, volume = {59}, year = {1993}, month = {1993 Aug}, pages = {2727-9}, abstract = {The production of small quantities of Fe(II) during the initial phase of microbial Fe(III) reduction greatly increased the amount of Fe(III) that could be extracted from freshwater sediments with oxalate. This finding and other evidence suggest that the oxalate-extractable Fe(III) that is unavailable for microbial reduction in anoxic sediments is not in the form of mixed Fe(III)-Fe(II) forms, as was previously suggested, but rather is in the form of highly crystalline Fe(III) oxides.}, issn = {0099-2240}, author = {Phillips, E J and Lovley, D R and Roden, E E} } @article {650, title = {Dissimilatory Fe(III) Reduction by the Marine Microorganism Desulfuromonas acetoxidans.}, journal = {Appl Environ Microbiol}, volume = {59}, year = {1993}, month = {1993 Mar}, pages = {734-42}, abstract = {The ability of the marine microorganism Desulfuromonas acetoxidans to reduce Fe(III) was investigated because of its close phylogenetic relationship with the freshwater dissimilatory Fe(III) reducer Geobacter metallireducens. Washed cell suspensions of the type strain of D. acetoxidans reduced soluble Fe(III)-citrate and Fe(III) complexed with nitriloacetic acid. The c-type cytochrome(s) of D. acetoxidans was oxidized by Fe(III)-citrate and Mn(IV)-oxalate, as well as by two electron acceptors known to support growth, colloidal sulfur and malate. D. acetoxidans grew in defined anoxic, bicarbonate-buffered medium with acetate as the sole electron donor and poorly crystalline Fe(III) or Mn(IV) as the sole electron acceptor. Magnetite (Fe(3)O(4)) and siderite (FeCO(3)) were the major end products of Fe(III) reduction, whereas rhodochrosite (MnCO(3)) was the end product of Mn(IV) reduction. Ethanol, propanol, pyruvate, and butanol also served as electron donors for Fe(III) reduction. In contrast to D. acetoxidans, G. metallireducens could only grow in freshwater medium and it did not conserve energy to support growth from colloidal S reduction. D. acetoxidans is the first marine microorganism shown to conserve energy to support growth by coupling the complete oxidation of organic compounds to the reduction of Fe(III) or Mn(IV). Thus, D. acetoxidans provides a model enzymatic mechanism for Fe(III) or Mn(IV) oxidation of organic compounds in marine and estuarine sediments. These findings demonstrate that 16S rRNA phylogenetic analyses can suggest previously unrecognized metabolic capabilities of microorganisms.}, issn = {0099-2240}, author = {Roden, E E and Lovley, D R} } @article {652, title = {Dissimilatory metal reduction.}, journal = {Annu Rev Microbiol}, volume = {47}, year = {1993}, month = {1993}, pages = {263-90}, abstract = {Microorganisms can enzymatically reduce a variety of metals in metabolic processes that are not related to metal assimilation. Some microorganisms can conserve energy to support growth by coupling the oxidation of simple organic acids and alcohols, H2, or aromatic compounds to the reduction of Fe(III) or Mn(IV). This dissimilatory Fe(III) and Mn(IV) reduction influences the organic as well as the inorganic geochemistry of anaerobic aquatic sediments and ground water. Microorganisms that use U(VI) as a terminal electron acceptor play an important role in uranium geochemistry and may be a useful tool for removing uranium from contaminated environments. Se(VI) serves as a terminal electron acceptor to support anaerobic growth of some microorganisms. Reduction of Se(VI) to Se(O) is an important mechanism for the precipitation of selenium from contaminated waters. Enzymatic reduction of Cr(VI) to the less mobile and less toxic Cr(III), and reduction of soluble Hg(II) to volatile Hg(O) may affect the fate of these compounds in the environment and might be used as a remediation strategy. Microorganisms can also enzymatically reduce other metals such as technetium, vanadium, molybdenum, gold, silver, and copper, but reduction of these metals has not been studied extensively.}, keywords = {Bacteria, Metals, Oxidation-Reduction}, issn = {0066-4227}, doi = {10.1146/annurev.mi.47.100193.001403}, author = {Lovley, D R} } @article {1196, title = {Enhanced thermotolerance and temperature-induced changes in protein composition in the hyperthermophilic archaeon ES4.}, journal = {J Bacteriol}, volume = {175}, year = {1993}, month = {1993 May}, pages = {2839-43}, abstract = {The hyperthermophilic archaeon ES4, a heterotrophic sulfur reducer isolated from a deep-sea hydrothermal vent, is capable of protecting itself from thermal stress at temperatures above its optimum for growth. The thermotolerance of ES4 was determined by exposing log-phase cells to various lethal high temperatures. When ES4 was shifted from 95 to 102 degrees C, it displayed recovery from an exponential rate of death, followed by transient thermotolerance. When ES4 was shifted directly from 95 to either 105 or 108 degrees C, only exponential death occurred. However, a shift from 95 to 105 degrees C with an intermediate incubation at 102 degrees C also gave ES4 transient thermotolerance to 105 degrees C. The protein composition of ES4 was examined at temperatures ranging from 75 to 102 degrees C by one-dimensional electrophoresis. Two proteins with molecular masses of approximately 90 and 150 kDa significantly decreased in abundance with increasing growth temperature, while a 98-kDa protein, present at very low levels at normal growth temperatures (76 to 99 degrees C), was more abundant at higher temperatures. The enhanced tolerance to hyperthermal conditions after a mild hyperthermal exposure and the increased abundance of the 98-kDa protein at above-optimal temperatures imply that ES4 is capable of a heat shock-like response previously unseen in hyperthermophilic archaea.
}, keywords = {Archaea, Bacterial Proteins, Blotting, Western, Electrophoresis, Polyacrylamide Gel, Hot Temperature}, issn = {0021-9193}, author = {Holden, J F and Baross, J A} } @article {651, title = {Geobacter metallireducens gen. nov. sp. nov., a microorganism capable of coupling the complete oxidation of organic compounds to the reduction of iron and other metals.}, journal = {Arch Microbiol}, volume = {159}, year = {1993}, month = {1993}, pages = {336-44}, abstract = {The gram-negative metal-reducing microorganism, previously known as strain GS-15, was further characterized. This strict anaerobe oxidizes several short-chain fatty acids, alcohols, and monoaromatic compounds with Fe(III) as the sole electron acceptor. Furthermore, acetate is also oxidized with the reduction of Mn(IV), U(VI), and nitrate. In whole cell suspensions, the c-type cytochrome(s) of this organism was oxidized by physiological electron acceptors and also by gold, silver, mercury, and chromate. Menaquinone was recovered in concentrations comparable to those previously found in gram-negative sulfate reducers. Profiles of the phospholipid ester-linked fatty acids indicated that both the anaerobic desaturase and the branched pathways for fatty acid biosynthesis were operative. The organism contained three lipopolysaccharide hydroxy fatty acids which have not been previously reported in microorganisms, but have been observed in anaerobic freshwater sediments. The 16S rRNA sequence indicated that this organism belongs in the delta proteobacteria. Its closest known relative is Desulfuromonas acetoxidans. The name Geobacter metallireducens is proposed.}, keywords = {Cytochrome c Group, Fatty Acids, Gram-Negative Anaerobic Bacteria, Iron, Oxidation-Reduction, Phylogeny, RNA, Ribosomal, 16S}, issn = {0302-8933}, author = {Lovley, D R and Giovannoni, S J and White, D C and Champine, J E and Phillips, E J and Gorby, Y A and Goodwin, S} } @article {800, title = {Norway rats as reservoir hosts for Lyme disease spirochetes on Monhegan Island, Maine.}, journal = {J Infect Dis}, volume = {168}, year = {1993}, month = {1993 Sep}, pages = {687-91}, abstract = {To determine whether the agent of Lyme disease, Borrelia burgdorferi, may be maintained in the absence of its usual white-footed mouse reservoir host, Ixodes dammini ticks from an island where mice are absent were examined. Prevalence of spirochetal infection was described for ticks removed from mammals, birds, and vegetation on Monhegan Island, Maine. Forty percent of adult I. dammini removed from vegetation were infected. Norway rats were heavily infested with ticks, and > 60\% of such ticks contained spirochetes. Other hosts were less frequently infested by ticks, and few such ticks were infected by spirochetes. The prevalence of antibody to B. burgdorferi was 23\% in dogs and cats; 4\% of island residents had Lyme disease. Thus, rats maintain Lyme disease spirochetes on Monhegan Island, and there may be transmission of this agent by I. dammini to island residents and their pets.}, keywords = {Animals, Animals, Domestic, Antibodies, Bacterial, Borrelia burgdorferi Group, Cats, Disease Reservoirs, Disease Vectors, Dogs, Geography, Humans, Incidence, Lyme Disease, Maine, Rats, Ticks}, issn = {0022-1899}, author = {Smith, R P and Rand, P W and Lacombe, E H and Telford, S R and Rich, S M and Piesman, J and Spielman, A} } @article {647, title = {Reduction of uranium by cytochrome c3 of Desulfovibrio vulgaris.}, journal = {Appl Environ Microbiol}, volume = {59}, year = {1993}, month = {1993 Nov}, pages = {3572-6}, abstract = {The mechanism for U(VI) reduction by Desulfovibrio vulgaris (Hildenborough) was investigated. The H2-dependent U(VI) reductase activity in the soluble fraction of the cells was lost when the soluble fraction was passed over a cationic exchange column which extracted cytochrome c3. Addition of cytochrome c3 back to the soluble fraction that had been passed over the cationic exchange column restored the U(VI)-reducing capacity. Reduced cytochrome c3 was oxidized by U(VI), as was a c-type cytochrome(s) in whole-cell suspensions. When cytochrome c3 was combined with hydrogenase, its physiological electron donor, U(VI) was reduced in the presence of H2. Hydrogenase alone could not reduce U(VI). Rapid U(VI) reduction was followed by a subsequent slow precipitation of the U(IV) mineral uraninite. Cytochrome c3 reduced U(VI) in a uranium-contaminated surface water and groundwater. Cytochrome c3 provides the first enzyme model for the reduction and biomineralization of uranium in sedimentary environments. Furthermore, the finding that cytochrome c3 can catalyze the reductive precipitation of uranium may aid in the development of fixed-enzyme reactors and/or organisms with enhanced U(VI)-reducing capacity for the bioremediation of uranium-contaminated waters and waste streams.}, keywords = {Biotransformation, Chemical Precipitation, Cytochrome c Group, Desulfovibrio vulgaris, Oxidation-Reduction, Uranium, Water Pollutants, Radioactive}, issn = {0099-2240}, author = {Lovley, D R and Widman, P K and Woodward, J C and Phillips, E J} } @article {843, title = {Use of high and low level overexpression plasmids to test mutant alleles of the recF gene of Escherichia coli K-12 for partial activity.}, journal = {Genetics}, volume = {135}, year = {1993}, month = {1993 Nov}, pages = {643-54}, abstract = {We showed that sufficient overexpression of the wild-type recF gene interfered with three normal cell functions: (1) UV induction of transcription from the LexA-protein-repressed sulA promoter, (2) UV resistance and (3) cell viability at 42 degrees. To show this, we altered a low-level overexpressing recF+ plasmid with a set of structurally neutral mutations that increased the rate of expression of recF. The resulting high-level overexpressing plasmid interfered with UV induction of the sulA promoter, as did the low-level overexpressing plasmid. It also reduced UV resistance more than its low level progenitor and decreased viability at 42 degrees, an effect not seen with the low-level plasmid. We used the high-level plasmid to test four recF structural mutations for residual activity. The structural alleles consisted of an insertion mutation, two single amino acid substitution mutations and a double amino acid substitution mutation. On the Escherichia coli chromosome the three substitution mutations acted similarly to a recF deletion in reducing UV resistance in a recB21 recC22 sbcB15 sbcC201 genetic background. By this test, therefore, all three appeared to be null alleles. Measurements of conjugational recombination revealed, however, that the three substitution mutations may have residual activity. On the high-level overexpressing plasmid all three substitution mutations definitely showed partial activity. By contrast, the insertion mutation on the high-level overexpressing plasmid showed no partial activity and can be considered a true null mutation. One of the substitutions, recF143, showed a property attributable to a leaky mutation. Another substitution, recF4101, may block selectively two of the three interference phenotypes, thus allowing us to infer a mechanism for them.}, keywords = {Alleles, Bacterial Proteins, Base Sequence, Chromosomes, Bacterial, DNA Repair, DNA, Bacterial, DNA-Binding Proteins, Escherichia coli, Escherichia coli Proteins, Gene Expression, Genes, Bacterial, Molecular Sequence Data, Mutagenesis, Site-Directed, Mutation, Phenotype, Plasmids, Radiation Tolerance, Recombination, Genetic, Ultraviolet Rays}, issn = {0016-6731}, author = {Sandler, S J and Clark, A J} } @article {654, title = {Acetate oxidation by dissimilatory Fe(III) reducers.}, journal = {Appl Environ Microbiol}, volume = {58}, year = {1992}, month = {1992 Sep}, pages = {3205-8}, keywords = {Acetates, Ferric Compounds, Gram-Negative Bacteria, Oxidation-Reduction}, issn = {0099-2240}, author = {Lovley, D R and Phillips, E J and Caccavo, F} } @article {703, title = {Class I major histocompatibility proteins are an essential component of the simian virus 40 receptor.}, journal = {J Virol}, volume = {66}, year = {1992}, month = {1992 Apr}, pages = {2037-45}, abstract = {The class I molecules encoded by the major histocompatibility complex (MHC) present endogenously synthesized antigenic peptide fragments to cytotoxic T lymphocytes. We show here that these proteins are an essential component of the cell surface receptor for simian virus 40 (SV40). First, SV40 binding to cells can be blocked by two monoclonal antibodies against class I human lymphocyte antigen (HLA) proteins but not by monoclonal antibodies specific for other cell surface proteins. Second, SV40 does not bind to cells of two different human lymphoblastoid cell lines which do not express surface class I MHC proteins because of genetic defects in the beta 2-microglobulin gene in one line and in the HLA complex in the other. Transfection of these cell lines with cloned genes for beta 2-microglobulin and HLA-B8, respectively, restored expression of their surface class I MHC proteins and resulted in concomitant SV40 binding. Finally, SV40 binds to purified HLA proteins in vitro and selectively binds to class I MHC proteins in a cell surface extract.}, keywords = {Animals, Binding, Competitive, Blotting, Western, Cell Line, Flow Cytometry, Histocompatibility Antigens Class I, Humans, Precipitin Tests, Receptors, Virus, Simian virus 40, Transfection}, issn = {0022-538X}, author = {Breau, W C and Atwood, W J and Norkin, L C} } @article {755, title = {Expression and transfer of engineered catabolic pathways harbored by Pseudomonas spp. introduced into activated sludge microcosms.}, journal = {Appl Environ Microbiol}, volume = {58}, year = {1992}, month = {1992 Oct}, pages = {3380-6}, abstract = {Two genetically engineered microorganisms (GEMs), Pseudomonas sp. strain B13 FR1(pFRC20P) (FR120) and Pseudomonas putida KT2440(pWWO-EB62) (EB62), were introduced into activated sludge microcosms that had the level of aeration, nutrient makeup, and microbial community structure of activated sludge reactors. FR120 contains an experimentally assembled ortho cleavage route for simultaneous degradation of 3-chlorobenzoate (3CB) and 4-methyl benzoate (4MB); EB62 contains a derivative TOL plasmid-encoded degradative pathway for toluene experimentally evolved so that it additionally processes 4-ethyl benzoate (4EB). Experiments assessed survival of the GEMs, their ability to degrade target substrates, and lateral transfer of plasmid-encoded recombinant DNA. GEMs added at initial densities of 10(6) to 10(7) bacteria per ml of activated sludge declined to stable population densities of 10(4) to 10(5) bacteria per ml. FR120 degraded combinations of 3CB and 4MB (1 mM each) following 3 days of adaptation in the microcosms. Indigenous microorganisms required an 8-day adaptation period before degradation of 4MB was observed; 3CB was degraded only after the concentration of 4MB was much reduced. The indigenous microbial community was killed when both compounds were present at concentrations of 4.0 mM. However, in parallel microcosms containing FR120, the microbial community maintained a normal density of viable cells. Indigenous microbes readily degraded 4EB (2 mM), and EB62 did not significantly increase the observed rate of degradation. In filter matings, transfer of pFRC20P, which specifies mobilization but not transfer functions, from FR120 to P. putida UWC1 was not detectable (< 10(-7) transconjugants per donor cell).(ABSTRACT TRUNCATED AT 250 WORDS)}, keywords = {Benzoates, Biodegradation, Environmental, Gene Expression Regulation, Bacterial, Genetic Engineering, Industrial Waste, Pseudomonas, Transfection, Waste Disposal, Fluid}, issn = {0099-2240}, author = {N{\"u}sslein, K and Maris, D and Timmis, K and Dwyer, D F} } @article {653, title = {A Hydrogen-Oxidizing, Fe(III)-Reducing Microorganism from the Great Bay Estuary, New Hampshire.}, journal = {Appl Environ Microbiol}, volume = {58}, year = {1992}, month = {1992 Oct}, pages = {3211-6}, abstract = {A dissimilatory Fe(III)- and Mn(IV)-reducing bacterium was isolated from bottom sediments of the Great Bay estuary, New Hampshire. The isolate was a facultatively anaerobic gram-negative rod which did not appear to fit into any previously described genus. It was temporarily designated strain BrY. BrY grew anaerobically in a defined medium with hydrogen or lactate as the electron donor and Fe(III) as the electron acceptor. BrY required citrate, fumarate, or malate as a carbon source for growth on H(2) and Fe(III). With Fe(III) as the sole electron acceptor, BrY metabolized hydrogen to a minimum threshold at least 60-fold lower than the threshold reported for pure cultures of sulfate reducers. This finding supports the hypothesis that when Fe(III) is available, Fe(III) reducers can outcompete sulfate reducers for electron donors. Lactate was incompletely oxidized to acetate and carbon dioxide with Fe(III) as the electron acceptor. Lactate oxidation was also coupled to the reduction of Mn(IV), U(VI), fumarate, thiosulfate, or trimethylamine n-oxide under anaerobic conditions. BrY provides a model for how enzymatic metal reduction by respiratory metal-reducing microorganisms has the potential to contribute to the mobilization of iron and trace metals and to the immobilization of uranium in sediments of Great Bay Estuary.}, issn = {0099-2240}, author = {Caccavo, F and Blakemore, R P and Lovley, D R} } @article {871, title = {Influence of mass transfer limitations on determination of the half saturation constant for hydrogen uptake in a mixed-culture CH(4)-producing enrichment.}, journal = {Biotechnol Bioeng}, volume = {40}, year = {1992}, month = {1992 Oct 5}, pages = {768-76}, abstract = {There is strong evidence in the literature supporting the existence of significant mass transfer limitations on the kinetics of exogenous H(2) consumption by methanogens. The half saturation constant for H (2) uptake by a mixed-culture, CH(4) producing enrichment was measured using an experimental protocol that avoided internal mass transfer limitations. The value obtained was two orders of magnitude smaller than any other previously reported. A mathematical model for acetogenic syntrophic associations was developed to check the capacity of H(2) as electron transporter between syntrophic partners. It was found that H(2) diffusion could account for the rate of transport of electrons between the syntrophic microorganisms and that formate is not a necessary intermediate. The possibility that formate may be an intermediate in this system was not ruled out. A Monod-type kinetic equation was modified to include the observed H(2) threshold effect. This modified equation was used to predict the CH(4)-production rate in a batch-fed digester. The results show that the external and internal H(2) pools are kinetically coupled.}, issn = {0006-3592}, doi = {10.1002/bit.260400704}, author = {Giraldo-Gomez, E and Goodwin, S and Switzenbaum, M S} } @article {655, title = {Reduction of uranium by Desulfovibrio desulfuricans.}, journal = {Appl Environ Microbiol}, volume = {58}, year = {1992}, month = {1992 Mar}, pages = {850-6}, abstract = {The possibility that sulfate-reducing microorganisms contribute to U(VI) reduction in sedimentary environments was investigated. U(VI) was reduced to U(IV) when washed cells of sulfate-grown Desulfovibrio desulfuricans were suspended in a bicarbonate buffer with lactate or H2 as the electron donor. There was no U(VI) reduction in the absence of an electron donor or when the cells were killed by heat prior to the incubation. The rates of U(VI) reduction were comparable to those in respiratory Fe(III)-reducing microorganisms. Azide or prior exposure of the cells to air did not affect the ability of D. desulfuricans to reduce U(VI). Attempts to grow D. desulfuricans with U(VI) as the electron acceptor were unsuccessful. U(VI) reduction resulted in the extracellular precipitation of the U(IV) mineral uraninite. The presence of sulfate had no effect on the rate of U(VI) reduction. Sulfate and U(VI) were reduced simultaneously. Enzymatic reduction of U(VI) by D. desulfuricans was much faster than nonenzymatic reduction of U(VI) by sulfide, even when cells of D. desulfuricans were added to provide a potential catalytic surface for the nonenzymatic reaction. The results indicate that enzymatic U(VI) reduction by sulfate-reducing microorganisms may be responsible for the accumulation of U(IV) in sulfidogenic environments. Furthermore, since the reduction of U(VI) to U(IV) precipitates uranium from solution, D. desulfuricans might be a useful organism for recovering uranium from contaminated waters and waste streams.}, keywords = {Biodegradation, Environmental, Desulfovibrio, Kinetics, Oxidation-Reduction, Sulfates, Uranium, X-Ray Diffraction}, issn = {0099-2240}, author = {Lovley, D R and Phillips, E J} } @article {844, title = {Sequence and complementation analysis of recF genes from Escherichia coli, Salmonella typhimurium, Pseudomonas putida and Bacillus subtilis: evidence for an essential phosphate binding loop.}, journal = {Nucleic Acids Res}, volume = {20}, year = {1992}, month = {1992 Feb 25}, pages = {839-45}, abstract = {We have compared the recF genes from Escherichia coli K-12, Salmonella typhimurium, Pseudomonas putida, and Bacillus subtilis at the DNA and amino acid sequence levels. To do this we determined the complete nucleotide sequence of the recF gene from Salmonella typhimurium and we completed the nucleotide sequence of recF gene from Pseudomonas putida begun by Fujita et al. (1). We found that the RecF proteins encoded by these two genes contain respectively 92\% and 38\% amino acid identity with the E. coli RecF protein. Additionally, we have found that the S. typhimurium and P. putida recF genes will complement an E. coli recF mutant, but the recF gene from Bacillus subtilis [showing about 20\% identity with E. coli (2)] will not. Amino acid sequence alignment of the four proteins identified four highly conserved regions. Two of these regions are part of a putative phosphate binding loop. In one region (position 36), we changed the lysine codon (which is essential for ATPase, GTPase and kinase activity in other proteins having this phosphate binding loop) to an arginine codon. We then tested this mutation (recF4101) on a multicopy plasmid for its ability to complement a recF chromosomal mutation and on the E. coli chromosome for its effect on sensitivity to UV irradiation. The strain with recF4101 on its chromosome is as sensitive as a null recF mutant strain. The strain with the plasmid-borne mutant allele is however more UV resistant than the null mutant strain. We conclude that lysine-36 and possibly a phosphate binding loop is essential for full recF activity. Lastly we made two chimeric recF genes by exchanging the amino terminal 48 amino acids of the S. typhimurium and E. coli recF genes. Both chimeras could complement E. coli chromosomal recF mutations.}, keywords = {Amino Acid Sequence, Bacillus subtilis, Bacterial Proteins, Base Sequence, Binding Sites, DNA, Bacterial, DNA-Binding Proteins, Escherichia coli, Escherichia coli Proteins, Genes, Bacterial, Genetic Complementation Test, Molecular Sequence Data, Nucleic Acid Conformation, Phosphates, Pseudomonas putida, Salmonella typhimurium, Sequence Alignment}, issn = {0305-1048}, author = {Sandler, S J and Chackerian, B and Li, J T and Clark, A J} } @article {872, title = {Acetate catabolism in the dissimilatory iron-reducing isolate GS-15.}, journal = {J Bacteriol}, volume = {173}, year = {1991}, month = {1991 Apr}, pages = {2704-6}, abstract = {Acetate-grown GS-15 whole-cell suspensions were disrupted with detergent and assayed for enzymes associated with acetate catabolism. Carbon monoxide dehydrogenase and formate dehydrogenase were not observed in GS-15. Catabolic levels of acetokinase and phosphotransacetylase were observed. Enzyme activities of the citric acid cycle, i.e., isocitrate dehydrogenase, 2-oxoglutarate sythase, succinate dehydrogenase, fumarase, and malate dehydrogenase, were observed.}, keywords = {Acetate Kinase, Acetates, Acetyl Coenzyme A, Bacteria, Carbon Monoxide, Citric Acid Cycle, Electron Transport, Euryarchaeota, Fumarate Hydratase, Isocitrate Dehydrogenase, Ketone Oxidoreductases, Malate Dehydrogenase, Phosphate Acetyltransferase, Pseudomonas aeruginosa, Succinate Dehydrogenase}, issn = {0021-9193}, author = {Champine, J E and Goodwin, S} } @article {656, title = {Dissimilatory Fe(III) and Mn(IV) reduction.}, journal = {Microbiol Rev}, volume = {55}, year = {1991}, month = {1991 Jun}, pages = {259-87}, abstract = {The oxidation of organic matter coupled to the reduction of Fe(III) or Mn(IV) is one of the most important biogeochemical reactions in aquatic sediments, soils, and groundwater. This process, which may have been the first globally significant mechanism for the oxidation of organic matter to carbon dioxide, plays an important role in the oxidation of natural and contaminant organic compounds in a variety of environments and contributes to other phenomena of widespread significance such as the release of metals and nutrients into water supplies, the magnetization of sediments, and the corrosion of metal. Until recently, much of the Fe(III) and Mn(IV) reduction in sedimentary environments was considered to be the result of nonenzymatic processes. However, microorganisms which can effectively couple the oxidation of organic compounds to the reduction of Fe(III) or Mn(IV) have recently been discovered. With Fe(III) or Mn(IV) as the sole electron acceptor, these organisms can completely oxidize fatty acids, hydrogen, or a variety of monoaromatic compounds. This metabolism provides energy to support growth. Sugars and amino acids can be completely oxidized by the cooperative activity of fermentative microorganisms and hydrogen- and fatty-acid-oxidizing Fe(III) and Mn(IV) reducers. This provides a microbial mechanism for the oxidation of the complex assemblage of sedimentary organic matter in Fe(III)- or Mn(IV)-reducing environments. The available evidence indicates that this enzymatic reduction of Fe(III) or Mn(IV) accounts for most of the oxidation of organic matter coupled to reduction of Fe(III) and Mn(IV) in sedimentary environments. Little is known about the diversity and ecology of the microorganisms responsible for Fe(III) and Mn(IV) reduction, and only preliminary studies have been conducted on the physiology and biochemistry of this process.}, keywords = {Bacteria, Electron Transport, Ferric Compounds, Fungi, Geological Phenomena, Geology, Manganese, Oxidation-Reduction, Soil Microbiology, Water Microbiology}, issn = {0146-0749}, author = {Lovley, D R} } @article {657, title = {Electron Transport in the Dissimilatory Iron Reducer, GS-15.}, journal = {Appl Environ Microbiol}, volume = {57}, year = {1991}, month = {1991 Mar}, pages = {867-70}, abstract = {Mechanisms for electron transport to Fe(III) were investigated in GS-15, a novel anaerobic microorganism which can obtain energy for growth by coupling the complete oxidation of organic acids or aromatic compounds to the reduction of Fe(III) to Fe(II). The results indicate that Fe(III) reduction proceeds through a type b cytochrome and a membrane-bound Fe(III) reductase which is distinct from the nitrate reductase.}, issn = {0099-2240}, author = {Gorby, Y A and Lovley, D R} } @article {659, title = {Anaerobic Oxidation of Toluene, Phenol, and p-Cresol by the Dissimilatory Iron-Reducing Organism, GS-15.}, journal = {Appl Environ Microbiol}, volume = {56}, year = {1990}, month = {1990 Jun}, pages = {1858-64}, abstract = {The dissimilatory Fe(III) reducer, GS-15, is the first microorganism known to couple the oxidation of aromatic compounds to the reduction of Fe(III) and the first example of a pure culture of any kind known to anaerobically oxidize an aromatic hydrocarbon, toluene. In this study, the metabolism of toluene, phenol, and p-cresol by GS-15 was investigated in more detail. GS-15 grew in an anaerobic medium with toluene as the sole electron donor and Fe(III) oxide as the electron acceptor. Growth coincided with Fe(III) reduction. [ring-C]toluene was oxidized to CO(2), and the stoichiometry of CO(2) production and Fe(III) reduction indicated that GS-15 completely oxidized toluene to carbon dioxide with Fe(III) as the electron acceptor. Magnetite was the primary iron end product during toluene oxidation. Phenol and p-cresol were also completely oxidized to carbon dioxide with Fe(III) as the sole electron acceptor, and GS-15 could obtain energy to support growth by oxidizing either of these compounds as the sole electron donor. p-Hydroxybenzoate was a transitory extracellular intermediate of phenol and p-cresol metabolism but not of toluene metabolism. GS-15 oxidized potential aromatic intermediates in the oxidation of toluene (benzylalcohol and benzaldehyde) and p-cresol (p-hydroxybenzylalcohol and p-hydroxybenzaldehyde). The metabolism described here provides a model for how aromatic hydrocarbons and phenols may be oxidized with the reduction of Fe(III) in contaminated aquifers and petroleum-containing sediments.}, issn = {0099-2240}, author = {Lovley, D R and Lonergan, D J} } @article {845, title = {Factors affecting expression of the recF gene of Escherichia coli K-12.}, journal = {Gene}, volume = {86}, year = {1990}, month = {1990 Jan 31}, pages = {35-43}, abstract = {This report describes four factors which affect expression of the recF gene from strong upstream lambda promoters under temperature-sensitive cIAt2-encoded repressor control. The first factor was the long mRNA leader sequence consisting of the Escherichia coli dnaN gene and 95\% of the dnaA gene and lambda bet, N (double amber) and 40\% of the exo gene. When most of this DNA was deleted, RecF became detectable in maxicells. The second factor was the vector, pBEU28, a runaway replication plasmid. When we substituted pUC118 for pBEU28, RecF became detectable in whole cells by the Coomassie blue staining technique. The third factor was the efficiency of initiation of translation. We used site-directed mutagenesis to change the mRNA leader, ribosome-binding site and the 3 bp before and after the translational start codon. Monitoring the effect of these mutational changes by translational fusion to lacZ, we discovered that the efficiency of initiation of translation was increased 30-fold. Only an estimated two- or threefold increase in accumulated levels of RecF occurred, however. This led us to discover the fourth factor, namely sequences in the recF gene itself. These sequences reduce expression of the recF-lacZ fusion genes 100-fold. The sequences responsible for this decrease in expression occur in four regions in the N-terminal half of recF. Expression is reduced by some sequences at the transcriptional level and by others at the translational level.}, keywords = {Bacterial Proteins, Base Sequence, Cloning, Molecular, Escherichia coli, Gene Expression Regulation, Bacterial, Genes, Bacterial, Molecular Sequence Data, Peptide Chain Initiation, Translational, Recombinant Fusion Proteins, Recombination, Genetic, Transcription, Genetic}, issn = {0378-1119}, author = {Sandler, S J and Clark, A J} } @article {658, title = {Rates of microbial metabolism in deep coastal plain aquifers.}, journal = {Appl Environ Microbiol}, volume = {56}, year = {1990}, month = {1990 Jun}, pages = {1865-74}, abstract = {Rates of microbial metabolism in deep anaerobic aquifers of the Atlantic coastal plain of South Carolina were investigated by both microbiological and geochemical techniques. Rates of [2-C]acetate and [U-C]glucose oxidation as well as geochemical evidence indicated that metabolic rates were faster in the sandy sediments composing the aquifers than in the clayey sediments of the confining layers. In the sandy aquifer sediments, estimates of the rates of CO(2) production (millimoles of CO(2) per liter per year) based on the oxidation of [2-C] acetate were 9.4 x 10 to 2.4 x 10 for the Black Creek aquifer, 1.1 x 10 for the Middendorf aquifer, and <7 x 10 for the Cape Fear aquifer. These estimates were at least 2 orders of magnitude lower than previously published estimates that were based on the accumulation of CO(2) in laboratory incubations of similar deep subsurface sediments. In contrast, geochemical modeling of groundwater chemistry changes along aquifer flowpaths gave rate estimates that ranged from 10 to 10 mmol of CO(2) per liter per year. The age of these sediments (ca. 80 million years) and their organic carbon content suggest that average rates of CO(2) production could have been no more than 10 mmol per liter per year. Thus, laboratory incubations may greatly overestimate the in situ rates of microbial metabolism in deep subsurface environments. This has important implications for the use of laboratory incubations in attempts to estimate biorestoration capacities of deep aquifers. The rate estimates from geochemical modeling indicate that deep aquifers are among the most oligotrophic aquatic environments in which there is ongoing microbial metabolism.}, issn = {0099-2240}, author = {Chapelle, F H and Lovley, D R} } @article {661, title = {Hydrogen and Formate Oxidation Coupled to Dissimilatory Reduction of Iron or Manganese by Alteromonas putrefaciens.}, journal = {Appl Environ Microbiol}, volume = {55}, year = {1989}, month = {1989 Mar}, pages = {700-6}, abstract = {The ability of Alteromonas putrefaciens to obtain energy for growth by coupling the oxidation of various electron donors to dissimilatory Fe(III) or Mn(IV) reduction was investigated. A. putrefaciens grew with hydrogen, formate, lactate, or pyruvate as the sole electron donor and Fe(III) as the sole electron acceptor. Lactate and pyruvate were oxidized to acetate, which was not metabolized further. With Fe(III) as the electron acceptor, A. putrefaciens had a high affinity for hydrogen and formate and metabolized hydrogen at partial pressures that were 25-fold lower than those of hydrogen that can be metabolized by pure cultures of sulfate reducers or methanogens. The electron donors for Fe(III) reduction also supported Mn(IV) reduction. The electron donors for Fe(III) and Mn(IV) reduction and the inability of A. putrefaciens to completely oxidize multicarbon substrates to carbon dioxide distinguish A. putrefaciens from GS-15, the only other organism that is known to obtain energy for growth by coupling the oxidation of organic compounds to the reduction of Fe(III) or Mn(IV). The ability of A. putrefaciens to reduce large quantities of Fe(III) and to grow in a defined medium distinguishes it from a Pseudomonas sp., which is the only other known hydrogen-oxidizing, Fe(III)-reducing microorganism. Furthermore, A. putrefaciens is the first organism that is known to grow with hydrogen as the electron donor and Mn(IV) as the electron acceptor and is the first organism that is known to couple the oxidation of formate to the reduction of Fe(III) or Mn(IV). Thus, A. putrefaciens provides a much needed microbial model for key reactions in the oxidation of sediment organic matter coupled to Fe(III) and Mn(IV) reduction.}, issn = {0099-2240}, author = {Lovley, D R and Phillips, E J and Lonergan, D J} } @article {660, title = {Requirement for a Microbial Consortium To Completely Oxidize Glucose in Fe(III)-Reducing Sediments.}, journal = {Appl Environ Microbiol}, volume = {55}, year = {1989}, month = {1989 Dec}, pages = {3234-6}, abstract = {In various sediments in which Fe(III) reduction was the terminal electron-accepting process, [C]glucose was fermented to C-fatty acids in a manner similar to that observed in methanogenic sediments. These results are consistent with the hypothesis that, in Fe(III)-reducing sediments, fermentable substrates are oxidized to carbon dioxide by the combined activity of fermentative bacteria and fatty acid-oxidizing, Fe(III)-reducing bacteria.}, issn = {0099-2240}, author = {Lovley, D R and Phillips, E J} } @article {662, title = {Novel mode of microbial energy metabolism: organic carbon oxidation coupled to dissimilatory reduction of iron or manganese.}, journal = {Appl Environ Microbiol}, volume = {54}, year = {1988}, month = {1988 Jun}, pages = {1472-80}, abstract = {A dissimilatory Fe(III)- and Mn(IV)-reducing microorganism was isolated from freshwater sediments of the Potomac River, Maryland. The isolate, designated GS-15, grew in defined anaerobic medium with acetate as the sole electron donor and Fe(III), Mn(IV), or nitrate as the sole electron acceptor. GS-15 oxidized acetate to carbon dioxide with the concomitant reduction of amorphic Fe(III) oxide to magnetite (Fe(3)O(4)). When Fe(III) citrate replaced amorphic Fe(III) oxide as the electron acceptor, GS-15 grew faster and reduced all of the added Fe(III) to Fe(II). GS-15 reduced a natural amorphic Fe(III) oxide but did not significantly reduce highly crystalline Fe(III) forms. Fe(III) was reduced optimally at pH 6.7 to 7 and at 30 to 35 degrees C. Ethanol, butyrate, and propionate could also serve as electron donors for Fe(III) reduction. A variety of other organic compounds and hydrogen could not. MnO(2) was completely reduced to Mn(II), which precipitated as rhodochrosite (MnCO(3)). Nitrate was reduced to ammonia. Oxygen could not serve as an electron acceptor, and it inhibited growth with the other electron acceptors. This is the first demonstration that microorganisms can completely oxidize organic compounds with Fe(III) or Mn(IV) as the sole electron acceptor and that oxidation of organic matter coupled to dissimilatory Fe(III) or Mn(IV) reduction can yield energy for microbial growth. GS-15 provides a model for how enzymatically catalyzed reactions can be quantitatively significant mechanisms for the reduction of iron and manganese in anaerobic environments.}, issn = {0099-2240}, author = {Lovley, D R and Phillips, E J} } @article {663, title = {Competitive mechanisms for inhibition of sulfate reduction and methane production in the zone of ferric iron reduction in sediments.}, journal = {Appl Environ Microbiol}, volume = {53}, year = {1987}, month = {1987 Nov}, pages = {2636-41}, abstract = {Mechanisms for inhibition of sulfate reduction and methane production in the zone of Fe(III) reduction in sediments were investigated. Addition of amorphic iron(III) oxyhydroxide to sediments in which sulfate reduction was the predominant terminal electron-accepting process inhibited sulfate reduction 86 to 100\%. The decrease in electron flow to sulfate reduction was accompanied by a corresponding increase in electron flow to Fe(III) reduction. In a similar manner, Fe(III) additions also inhibited methane production in sulfate-depleted sediments. The inhibition of sulfate reduction and methane production was the result of substrate limitation, because the sediments retained the potential for sulfate reduction and methane production in the presence of excess hydrogen and acetate. Sediments in which Fe(III) reduction was the predominant terminal electron-accepting process had much lower concentrations of hydrogen and acetate than sediments in which sulfate reduction or methane production was the predominant terminal process. The low concentrations of hydrogen and acetate in the Fe(III)-reducing sediments were the result of metabolism by Fe(III)-reducing organisms of hydrogen and acetate at concentrations lower than sulfate reducers or methanogens could metabolize them. The results indicate that when Fe(III) is in a form that Fe(III)-reducing organisms can readily reduce, Fe(III)-reducing organisms can inhibit sulfate reduction and methane production by outcompeting sulfate reducers and methanogens for electron donors.}, issn = {0099-2240}, author = {Lovley, D R and Phillips, E J} } @article {664, title = {Rapid assay for microbially reducible ferric iron in aquatic sediments.}, journal = {Appl Environ Microbiol}, volume = {53}, year = {1987}, month = {1987 Jul}, pages = {1536-40}, abstract = {The availability of ferric iron for microbial reduction as directly determined by the activity of iron-reducing organisms was compared with its availability as determined by a newly developed chemical assay for microbially reducible iron. The chemical assay was based on the reduction of poorly crystalline ferric iron by hydroxylamine under acidic conditions. There was a strong correlation between the extent to which hydroxylamine could reduce various synthetic ferric iron forms and the susceptibility of the iron to microbial reduction in an enrichment culture of iron-reducing organisms. When sediments that contained hydroxylamine-reducible ferric iron were incubated under anaerobic conditions, ferrous iron accumulated as the concentration of hydroxylamine-reducible ferric iron declined over time. Ferrous iron production stopped as soon as the hydroxylamine-reducible ferric iron was depleted. In anaerobic incubations of reduced sediments that did not contain hydroxylamine-reducible ferric iron, there was no microbial iron reduction, even though the sediments contained high concentrations of oxalate-extractable ferric iron. A correspondence between the presence of hydroxylamine-reducible ferric iron and the extent of ferric iron reduction in anaerobic incubations was observed in sediments from an aquifer and in fresh- and brackish-water sediments from the Potomac River estuary. The assay is a significant improvement over previously described procedures for the determination of hydroxylamine-reducible ferric iron because it provides a correction for the high concentrations of solid ferrous iron which may also be extracted from sediments with acid. This is a rapid, simple technique to determine whether ferric iron is available for microbial reduction.}, issn = {0099-2240}, author = {Lovley, D R and Phillips, E J} } @article {665, title = {Availability of ferric iron for microbial reduction in bottom sediments of the freshwater tidal potomac river.}, journal = {Appl Environ Microbiol}, volume = {52}, year = {1986}, month = {1986 Oct}, pages = {751-7}, abstract = {The distribution of Fe(III), its availability for microbial reduction, and factors controlling Fe(III) availability were investigated in sediments from a freshwater site in the Potomac River Estuary. Fe(III) reduction in sediments incubated under anaerobic conditions and depth profiles of oxalate-extractable Fe(III) indicated that Fe(III) reduction was limited to depths of 4 cm or less, with the most intense Fe(III) reduction in the top 1 cm. In incubations of the upper 4 cm of the sediments, Fe(III) reduction was as important as methane production as a pathway for anaerobic electron flow because of the high rates of Fe(III) reduction in the 0- to 0.5-cm interval. Most of the oxalate-extractable Fe(III) in the sediments was not reduced and persisted to a depth of at least 20 cm. The incomplete reduction was not the result of a lack of suitable electron donors. The oxalate-extractable Fe(III) that was preserved in the sediments was considered to be in a form other than amorphous Fe(III) oxyhydroxide, since synthetic amorphous Fe(III) oxyhydroxide, amorphous Fe(III) oxyhydroxide adsorbed onto clay, and amorphous Fe(III) oxyhydroxide saturated with adsorbed phosphate or fulvic acids were all readily reduced. Fe(3)O(4) and the mixed Fe(III)-Fe(II) compound(s) that were produced during the reduction of amorphous Fe(III) oxyhydroxide in an enrichment culture were oxalate extractable but were not reduced, suggesting that mixed Fe(III)-Fe(II) compounds might account for the persistence of oxalate-extractable Fe(III) in the sediments. The availability of microbially reducible Fe(III) in surficial sediments demonstrates that microbial Fe(III) reduction can be important to organic matter decomposition and iron geochemistry. However, the overall extent of microbial Fe(III) reduction is governed by the inability of microorganisms to reduce most of the Fe(III) in the sediment.}, issn = {0099-2240}, author = {Lovley, D R and Phillips, E J} } @article {666, title = {Organic matter mineralization with reduction of ferric iron in anaerobic sediments.}, journal = {Appl Environ Microbiol}, volume = {51}, year = {1986}, month = {1986 Apr}, pages = {683-9}, abstract = {The potential for ferric iron reduction with fermentable substrates, fermentation products, and complex organic matter as electron donors was investigated with sediments from freshwater and brackish water sites in the Potomac River Estuary. In enrichments with glucose and hematite, iron reduction was a minor pathway for electron flow, and fermentation products accumulated. The substitution of amorphous ferric oxyhydroxide for hematite in glucose enrichments increased iron reduction 50-fold because the fermentation products could also be metabolized with concomitant iron reduction. Acetate, hydrogen, propionate, butyrate, ethanol, methanol, and trimethylamine stimulated the reduction of amorphous ferric oxyhydroxide in enrichments inoculated with sediments but not in uninoculated or heat-killed controls. The addition of ferric iron inhibited methane production in sediments. The degree of inhibition of methane production by various forms of ferric iron was related to the effectiveness of these ferric compounds as electron acceptors for the metabolism of acetate. The addition of acetate or hydrogen relieved the inhibition of methane production by ferric iron. The decrease of electron equivalents proceeding to methane in sediments supplemented with amorphous ferric oxyhydroxides was compensated for by a corresponding increase of electron equivalents in ferrous iron. These results indicate that iron reduction can outcompete methanogenic food chains for sediment organic matter. Thus, when amorphous ferric oxyhydroxides are available in anaerobic sediments, the transfer of electrons from organic matter to ferric iron can be a major pathway for organic matter decomposition.}, issn = {0099-2240}, author = {Lovley, D R and Phillips, E J} } @article {667, title = {Minimum threshold for hydrogen metabolism in methanogenic bacteria.}, journal = {Appl Environ Microbiol}, volume = {49}, year = {1985}, month = {1985 Jun}, pages = {1530-1}, abstract = {Methanogenic isolates did not consume hydrogen below partial pressures of 6.5 Pa. Thus, in contrast to a previous report, results from pure-culture studies do not invalidate the threshold model for methane production from hydrogen in sediments.}, issn = {0099-2240}, author = {Lovley, D R} } @article {668, title = {Production and Consumption of H(2) during Growth of Methanosarcina spp. on Acetate.}, journal = {Appl Environ Microbiol}, volume = {49}, year = {1985}, month = {1985 Jan}, pages = {247-9}, abstract = {Methanosarcina sp. strain TM-1 and Methanosarcina acetivorans produced and consumed H(2) to maintain H(2) partial pressures of 16 to 92 Pa in closed cultures during growth on acetate. Strain TM-1 produced H(2) continuously when H(2) was continuously removed from the culture. The potential physiological significance of H(2) in acetate metabolism to methane is discussed.}, issn = {0099-2240}, author = {Lovley, D R and Ferry, J G} } @article {847, title = {Genes of the RecE and RecF pathways of conjugational recombination in Escherichia coli.}, journal = {Cold Spring Harb Symp Quant Biol}, volume = {49}, year = {1984}, month = {1984}, pages = {453-62}, keywords = {Chromosome Mapping, Conjugation, Genetic, Escherichia coli, Genes, Bacterial, Mutation, Protein Biosynthesis, Recombination, Genetic}, issn = {0091-7451}, author = {Clark, A J and Sandler, S J and Willis, D K and Chu, C C and Blanar, M A and Lovett, S T} } @article {669, title = {Identification of methyl coenzyme M as an intermediate in methanogenesis from acetate in Methanosarcina spp.}, journal = {J Bacteriol}, volume = {160}, year = {1984}, month = {1984 Nov}, pages = {521-5}, abstract = {The transfer of the methyl group of acetate to coenzyme M (2-mercaptoethanesulfonic acid; HS-CoM) during the metabolism of acetate to methane was investigated in cultures of Methanosarcina strain TM-1. The organism metabolized CD3COO- to 83\% CD3H and 17\% CD2H2 and produced no CDH3 or CH4. The isotopic composition of coenzyme M in cells grown on CD3COO- was analyzed with a novel gas chromatography-mass spectrometry technique. The cells contained CD3-D-CoM and CD2H-S-CoM) in a proportion similar to that of CD3H to CD2H2. These results, in conjunction with a report (J.K. Nelson and J.G. Ferry, J. Bacteriol. 160:526-532, 1984) that extracts of acetate-grown strain TM-1 contain high levels of CH3-S-CoM methylreductase, indicate that CH3-S-CoM is an intermediate in the metabolism of acetate to methane in this organism.}, keywords = {Acetates, Deuterium, Euryarchaeota, Gas Chromatography-Mass Spectrometry, Mercaptoethanol, Mesna, Methane, Methylation}, issn = {0021-9193}, author = {Lovley, D R and White, R H and Ferry, J G} } @article {846, title = {Molecular analysis of the recF gene of Escherichia coli.}, journal = {Proc Natl Acad Sci U S A}, volume = {81}, year = {1984}, month = {1984 Aug}, pages = {4622-6}, abstract = {We analyzed the nucleotide sequence of a 1.325-kilobase region of wild-type Escherichia coli containing a functional recF gene and six Tn3 mutations that inactivate recF. The analysis shows a potentially translatable reading frame of 1071 nucleotides, which is interrupted by all six insertions. A protein of 40.5 kilodaltons would result from translation of the open reading frame, and a radioactive band of protein of an apparent molecular weight of approximately 40 kilodaltons was seen by the maxicell method using a recF+ plasmid. Putative truncated peptides were seen when two recF::Tn3 mutant plasmids were used. Differential expression of dnaN and recF from a common promoter was noted. recF332::Tn3 was transferred to the chromosome where, in hemizygous condition, it produced UV sensitivity indistinguishable from that produced by two presumed recF point mutations.}, keywords = {Amino Acid Sequence, Bacterial Proteins, Base Sequence, DNA Repair, DNA Replication, Escherichia coli, Genes, Bacterial, Molecular Weight, Mutation, Plasmids, Recombination, Genetic}, issn = {0027-8424}, author = {Blanar, M A and Sandler, S J and Armengod, M E and Ream, L W and Clark, A J} } @article {670, title = {Rapidly growing rumen methanogenic organism that synthesizes coenzyme M and has a high affinity for formate.}, journal = {Appl Environ Microbiol}, volume = {48}, year = {1984}, month = {1984 Jul}, pages = {81-7}, abstract = {Methanogenic bacteria with a coccobacillus morphology similar to Methanobrevibacter ruminantium were isolated from the bovine rumen. One isolate, 10-16B, represented a previously undescribed rumen population that, unlike M. ruminantium, synthesized coenzyme M, grew rapidly (mu = 0.24 h-1) on H2-CO2 in a complex medium, had simple nutritional requirements, and metabolized formate at reported rumen concentrations. H2 was metabolized to partial pressures 10-fold lower than those reported for the rumen. After H2 starvation for 26 h, strain 10-16B rapidly resumed growth when H2 was made available. The minimum concentrations of acetate (6 mM) and ammonia (less than 7 mM) that were required for optimal growth were lower than the reported acetate and ammonia concentrations in the rumen. Isoleucine and leucine stimulated growth, but only at concentrations (greater than 50 microM) higher than those reported for the rumen. Another coccobacillary methanogenic organism that synthesized coenzyme M was isolated from a different animal as were organisms that required an exogenous supply of coenzyme M. In general, methanogenic bacteria that required an exogenous supply of coenzyme M had lower maximum growth rates and more complex nutritional requirements than organisms that synthesized the cofactor. The ability of all isolates to metabolize formate below the detection limit of 10 microM indicated that, in contrast to previous reports, methanogenic bacteria have the potential to directly metabolize formate in the rumen. This study demonstrated that there are physiologically diverse populations of coccobacillary methanogenic bacteria in the rumen that can interact competitively and cooperatively.}, keywords = {Animals, Cattle, Culture Media, Enzyme Induction, Euryarchaeota, Formates, Hydrogen, Male, Mercaptoethanol, Mesna, Rumen, Substrate Specificity}, issn = {0099-2240}, author = {Lovley, D R and Greening, R C and Ferry, J G} } @article {671, title = {Metabolism of acetate, methanol, and methylated amines in intertidal sediments of lowes cove, maine.}, journal = {Appl Environ Microbiol}, volume = {45}, year = {1983}, month = {1983 Jun}, pages = {1848-53}, abstract = {The fates and the rates of metabolism of acetate, trimethylamine, methylamine, and methanol were examined to determine the significance of these compounds as in situ methane precursors in surface sediments of an intertidal zone in Maine. Concentrations of these potential methane precursors were generally <3 muM, with the exception of sediments containing fragments of the seaweed Ascophyllum nodosum, in which acetate was 96 muM. [2-C]acetate turnover in all samples was rapid (turnover time <2 h), with CO(2) as the primary product. [C]trimethylamine and methylamine turnover times were slower (>8 h) and were characterized by formation of both CH(4) and CO(2). Ratios of CH(4)/CO(2) from [C]trimethylamine and methylamine in uninhibited sediments indicated that a significant fraction of these substrates were catabolized via a non-methanogenic process. Data from inhibition experiments involving sodium molybdate and 2-bromoethanesulfonic acid supported this interpretation. [C]methanol was oxidized relatively slowly compared with the other substrates and was catabolized mainly to CO(2). Results from experiments with molybdate and 2-bromoethanesulfonic acid suggested that methanol was oxidized primarily through sulfate reduction. In Lowes Cove sediments, trimethylamine accounted for 35.1 to 61.1\% of total methane production.}, issn = {0099-2240}, author = {King, G M and Klug, M J and Lovley, D R} } @article {672, title = {Methanogenesis from methanol and methylamines and acetogenesis from hydrogen and carbon dioxide in the sediments of a eutrophic lake.}, journal = {Appl Environ Microbiol}, volume = {45}, year = {1983}, month = {1983 Apr}, pages = {1310-5}, abstract = {C-tracer techniques were used to examine the metabolism of methanol and methylamines and acetogenesis from hydrogen and carbon dioxide in sediments from the profundal and littoral zones of eutrophic Wintergreen Lake, Michigan. Methanogens were primarily responsible for the metabolism of methanol, monomethylamine, and trimethylamine and maintained the pool size of these substrates below 10 muM in both sediment types. Methanol and methylamines were the precursors for less than 5 and 1\%, respectively, of the total methane produced. Methanol and methylamines continued to be metabolized to methane when the sulfate concentration in the sediment was increased to 20 mM. Less than 2\% of the total acetate production was derived from carbon dioxide reduction. Hydrogen consumption by hydrogen-oxidizing acetogens was 5\% or less of the total hydrogen uptake by acetogens and methanogens. These results, in conjunction with previous studies, emphasize that acetate and hydrogen are the major methane precursors and that methanogens are the predominant hydrogen consumers in the sediments of this eutrophic lake.}, issn = {0099-2240}, author = {Lovley, D R and Klug, M J} } @article {673, title = {Sulfate reducers can outcompete methanogens at freshwater sulfate concentrations.}, journal = {Appl Environ Microbiol}, volume = {45}, year = {1983}, month = {1983 Jan}, pages = {187-92}, abstract = {Acetate and hydrogen metabolism by sulfate reducers and methanogens in the profundal sediments of an oligotrophic lake were examined. Inhibition of sulfate reduction with molybdate stimulated methane production from both hydrogen and acetate. Molybdate did not stimulate methane production in sediments that were preincubated to deplete the sulfate pool. Sulfate reduction accounted for 30 to 81\% of the total of terminal metabolism proceeding through sulfate reduction and methane production in Eckman grab samples of surface sediments. The ability of sulfate reducers to effectively compete with methanogens for acetate was related to the sulfate reducers{\textquoteright} lower half-saturation constant for acetate metabolism at in situ sulfate concentrations. Processes other than sulfate reduction and methanogenesis consumed hydrogen at elevated hydrogen partial pressures and prevented a kinetic analysis of hydrogen uptake by sulfate reducers and methanogens. The demonstration that sulfate reducers can successfully compete with methanogens for hydrogen and acetate in sediments at in situ sulfate concentrations of 60 to 105 muM extends the known range of sediment habitats in which sulfate reduction can be a dominant terminal process.}, issn = {0099-2240}, author = {Lovley, D R and Klug, M J} } @article {675, title = {Intermediary metabolism of organic matter in the sediments of a eutrophic lake.}, journal = {Appl Environ Microbiol}, volume = {43}, year = {1982}, month = {1982 Mar}, pages = {552-60}, abstract = {The rates, products, and controls of the metabolism of fermentation intermediates in the sediments of a eutrophic lake were examined. C-fatty acids were directly injected into sediment subcores for turnover rate measurements. The highest rates of acetate turnover were in surface sediments (0- to 2-cm depth). Methane was the dominant product of acetate metabolism at all depths. Simultaneous measurements of acetate, propionate, and lactate turnover in surface sediments gave turnover rates of 159, 20, and 3 muM/h, respectively. [2-C]propionate and [U-C]lactate were metabolized to [C]acetate, CO(2), and CH(4). [C]formate was completely converted to CO(2) in less than 1 min. Inhibition of methanogenesis with chloroform resulted in an immediate accumulation of volatile fatty acids and hydrogen. Hydrogen inhibited the metabolism of C(3)-C(5) volatile fatty acids. The rates of fatty acid production were estimated from the rates of fatty acid accumulation in the presence of chloroform or hydrogen. The mean molar rates of production were acetate, 82\%; propionate, 13\%; butyrates, 2\%; and valerates, 3\%. A working model for carbon and electron flow is presented which illustrates that fermentation and methanogenesis are the predominate steps in carbon flow and that there is a close interaction between fermentative bacteria, acetogenic hydrogen-producing bacteria, and methanogens.}, issn = {0099-2240}, author = {Lovley, D R and Klug, M J} } @article {674, title = {Kinetic analysis of competition between sulfate reducers and methanogens for hydrogen in sediments.}, journal = {Appl Environ Microbiol}, volume = {43}, year = {1982}, month = {1982 Jun}, pages = {1373-9}, abstract = {The competition between sulfate-reducing and methanogenic bacteria for hydrogen was investigated in eutrophic lake sediments that contained low in situ sulfate concentrations and in sulfate-amended sediments. Sulfate reduction and methane production coexisted in situ in lake surface sediments (0 to 2 cm), but methane production was the dominant terminal process. Addition of 10 to 20 mM sulfate to sediments resulted in a decrease in the hydrogen partial pressure and a concomitant inhibition of methane production over time. Molybdate inhibition of sulfate reduction in sulfate-amended sediments was followed by an increase in the hydrogen partial pressure and the methane production rate to values comparable to those in sediments not amended with sulfate. The sulfate reducer population had a half-saturation constant for hydrogen uptake of 141 pascals versus 597 pascals for the methanogen population. Thus, when sulfate was not limiting, the lower half-saturation constant of sulfate reducers enabled them to inhibit methane production by lowering the hydrogen partial pressure below levels that methanogens could effectively utilize. However, methanogens coexisted with sulfate reducers in the presence of sulfate, and the outcome of competition at any time was a function of the rate of hydrogen production, the relative population sizes, and sulfate availability.}, issn = {0099-2240}, author = {Lovley, D R and Dwyer, D F and Klug, M J} } @article {1739, title = {Bacterial metabolism of resorcinylic compounds: purification and properties of orcinol hydroxylase and resorcinol hydroxylase from Pseudomonas putida ORC.}, journal = {Eur J Biochem}, volume = {61}, year = {1976}, month = {1976 Jan 2}, pages = {259-69}, abstract = {The hydroxylase activities observed in extracts of Pseudomonas putida ORC after growth on orcinol and resorcinol as sole source of carbon have been purified to homogeneity. Both enzymes were shown to be flavoproteins and to contain approximately 1 mol of FAD for each polypeptide chain, S20,W values for each enzyme are 4.1 +/- 0.1 and are independent of the presence of their aromatic substrates. Molecular weight determinations under native (approximately 68000) and denaturing (approximately 70000) conditions indicated that they are monomeric. The visible absorption spectra identical but the circular dichroic spectra of the two proteins can be distinguished. Although each protein catalyzes the NAD(P)H and O2-dependent hydroxylation of both orcinol and resorcinol, the efficiency of the transformations of the substrates by the two enzymes is radically different; furthermore resorcinol hydroxylase is much more versatile in the aromatic compounds it can utilize as substrates and effectors. Other properties of the enzymes which clearly establish their own identity include their serological characteristics and amino acid composition; the latter property is particularly evident when the quantities of valine and alanine residues are compared. The synthesis of each enzyme is also under different regulatory constraints, being controlled by the substrate used for growth.
}, keywords = {Amino Acids, Circular Dichroism, Flavin-Adenine Dinucleotide, Mixed Function Oxygenases, Molecular Weight, NAD, NADP, Oxidation-Reduction, Protein Conformation, Pseudomonas, Resorcinols, Spectrophotometry, Spectrophotometry, Ultraviolet}, issn = {0014-2956}, author = {Ohta, Y and Ribbons, D W} } @article {1707, title = {Citrate synthase.}, journal = {Curr Top Cell Regul}, volume = {10}, year = {1976}, month = {1976}, pages = {161-204}, keywords = {Adenosine Diphosphate, Adenosine Monophosphate, Adenosine Triphosphate, Amino Acids, Animals, Bacteria, Binding Sites, Cations, Divalent, Cations, Monovalent, Citrate (si)-Synthase, Dithionitrobenzoic Acid, Ketoglutaric Acids, Kinetics, Molecular Weight, NAD, NADP, Oxo-Acid-Lyases, Protein Binding, Species Specificity}, issn = {0070-2137}, author = {Weitzman, P D and Danson, M J} } @article {1731, title = {Disposition of (15,16-3H)naltrexone in the central nervous system of the rat.}, journal = {Drug Metab Dispos}, volume = {4}, year = {1976}, month = {1976 May-Jun}, pages = {276-80}, abstract = {After injection of (15,16-3H)naltrexone (10 mg/kg s.c.) in male Wistar rats, peak concentrations of drug occurred in brain and plasma within 0.5 hr. Levels of naltrexone were sustained in brain between 2 and 24 hr and were barely detectable at 48 hr. Significant amounts of metabolities were present in brain and plasma at longer time periods. The t1/2 of naltrexone in brain and plasma were approximately 8.0 and 11.4 hr. respectively. The brain/plasma ratios of naltrexone at earlier times (0.5-1 hr) were higher than those at later times. The binding of naltrexone in vitro with rat plasma proteins in concentrations of 1-10 mug/ml ranged between 41 and 59\% 6beta-Naltrexol was present in very small amounts in brain but not in plasma. In addition to 7,8-dihydro-14-hydroxynormophinone and 7,8-dihydro-14-hydroxynormophine, tentative evidence was obtained for three other metabolites of naltrexone in brain. These metabolites were also present in plasma in addition to free and conjugated naltrexone and its N-dealkylated metabolites.
}, keywords = {Animals, Blood-Brain Barrier, Brain, Chromatography, Thin Layer, Male, Naloxone, Naltrexone, Rats}, issn = {0090-9556}, author = {Misra, A L and Bloch, R and Vardy, J and Mul{\'e}, S J and Verebely, K} } @article {1715, title = {Mechanistic studies of glutamine synthetase from Escherichia coli: kinetic evidence for two reaction intermediates in biosynthetic reaction.}, journal = {Proc Natl Acad Sci U S A}, volume = {73}, year = {1976}, month = {1976 Feb}, pages = {476-80}, abstract = {Fast reaction techniques were used to study the kinetics of protein fluorescence intensity changes that are associated with the reactions of unadenylylated Escherichia coli glutamine synthetase [L-glutamate: ammonia ligase (ADP-forming), EC 6.3.1.2] with its substrates. It was established that the synthesis of glutamine occurs by a stepwise mechanism. During the catalytic process two fluorometrically distinct intermediates were observed. Both forward and reverse rate constants which lead to the formation and consumption of these intermediates were evaluated. The catalytic rate constant, kc, which was calculated from these rate constants agrees well with the values of kc which were determined by direct measurement of the overall biosynthetic activities by means of stopped-flow technique or the steady-state assay method.
}, keywords = {Enzyme Activation, Escherichia coli, Glutamate-Ammonia Ligase, Kinetics, Magnesium}, issn = {0027-8424}, author = {Rhee, S G and Chock, P B} } @article {1734, title = {The specificity of heterophil antibodies in patients and healthy donors with no or minimal signs of infectious mononucleosis.}, journal = {Blood}, volume = {47}, year = {1976}, month = {1976 Jan}, pages = {91-8}, abstract = {Over several years sera were collected from 14 heterophil-positive students or patients who did not fulfill minimal hematologic criteria for infectious mononucleosis (I.M.) The specificity of these heterophil reactions for I.M. was investigated by determining antibodies to Epstein-Barr virus-determined antigens, i.e., to viral capsid antigens (VCA), early antigens (EA), and EBV-associated nuclear antigens (EBNA). On the basis of detectable anti-EA and/or the early absence and late emergence of anti-EBNA, four of these 14 individuals showed evidence of a current or very recent primary Epstein-Barr virus infection. The other ten patients showed antibody patterns indicative of Epstein-Barr virus infections in the past, and no firm conclusions could be drawn with regard to the specificity of their heterophil reactions. It was assumed, however, that some represented atypical clinical forms of EBV infection and that timing of specimen collection was a factor in explaining the paucity of Downey cells. In three patients, the absorbed heterophil-positive reactions persisted with little change in titer for at least 22 mo and thus might represent false-positive tests.
}, keywords = {Adolescent, Adult, Agglutinins, Antibodies, Heterophile, Antigens, Aspartate Aminotransferases, Capsid, Female, gamma-Glutamyltransferase, Herpesvirus 4, Human, Humans, Infectious Mononucleosis, Male, Serologic Tests}, issn = {0006-4971}, author = {Horwitz, C A and Henle, W and Henle, G and Polesky, H and Wexler, H and Ward, P} } @article {1735, title = {[Spectral properties of porcine plasminogen: study of the acidic transition (author{\textquoteright}s transl)].}, journal = {Eur J Biochem}, volume = {63}, year = {1976}, month = {1976 Apr 1}, pages = {553-62}, abstract = {The acidic transition of porcine plasminogen, prepared by affinity chromatography, was studied by non-destructive methods. These methods are based on the analysis of the behaviour of the tryptophyls under various conditions. The perturbation of the absorption and emission spectra by pH or temperature and the dynamic quenching of the intrinsic fluorescence are used to obtain information on structural changes which affect the environment of these residues. It is shown that by decreasing pH the fluorescence emission spectra are shifted toward the long wavelengths, with a broadening of the fluorescence band. The same effect can be obtained at constant pH by heating the protein solution. In order to analyze these phenomena, it is assumed that the fluorescence intensities at 355 nm and 328 nm reflect the proportion of the tryptophans which are exposed to the solvent, and buried, respectively. The plot of the ratio of the fluorescence intensities at these wavelengths versus pH or temperature leads to a titration curve showing an unmasking of tryptophans. The proportion of exposed tryptophans is measured by the dynamic fluorescence quenching technique and the data analyzed according to Lehrer. The plot of the fraction of exposed tryptophyls versus pH also shows the unmasking of these chromophores. Thermal perturbation of a solution of plaminogen at neutral pH induces a difference absorption spectrum whose amplitudes at the maxima are proportional to the number of exposed aromatic residues. The comparison with a solution of fully denatured plasminogen in 6 M guanidium chloride, where all the tryptophyls are exposed, shows that the percentage of exposure is equal to 59\%. This number is significantly higher than the percentage found by the fluorescence quenching technique (20\%), indicating that some tryptophyls are located in crevices, exposed to the solvent but not to the iodide. At acidic pH the absorption difference spectra induced by thermal perturbation are not classical, since they show an inversion and a new band between 300 nm and 305 nm. This band is mentioned in the literature as a minor band of tryptophan which appears when this chromophore is located in an asymmetric environment. On plotting the maximum amplitude of these spectra obtained at acidic pH versus temperature, we obtain a curve indicating that two types of antagonistic interactions are involved in the perturbation of the chromophores spectra. The spectrophotometric titration of plasminogen gives classical absorption difference spectra. By plotting the maximum amplitude at 292 nm versus pH, we obtain a titration curve with an apparent pK of 2.9 units. This pK is acidic which respect to the pK value of a normal carboxyl. This low value can be due to a positively charged group in the neighbourhood of a carboxyl, which interacts with one or more chromophores. When the carboxyl becomes protonated, this positively charged group is free and available to perturb the environment of some chromophores...
}, keywords = {Animals, Binding Sites, Guanidines, Hydrogen-Ion Concentration, Plasminogen, Protein Binding, Protein Conformation, Spectrometry, Fluorescence, Spectrophotometry, Spectrophotometry, Ultraviolet, Swine, Temperature}, issn = {0014-2956}, author = {Rodier, F} } @article {1716, title = {Stereochemical studies of adrenergic drugs. Optically active derivatives of imidazolines.}, journal = {J Med Chem}, volume = {19}, year = {1976}, month = {1976 Dec}, pages = {1382-4}, abstract = {The synthesis of (R)-(+)-4-methyl-2-(1-naphthylmethyl)imidazoline hydrochloride (2) and (S)-(-)-4-methyl-2-(1-naphthylmethyl)imidazoline hydrochloride (3) is presented. The synthesis involves the preparation of (R)-(+)- and (S)-(-)-1,2-diaminopropane dihydrochloride and then allowing the appropriate diaminopropane to react with ethyl 1-naphthyliminoacetate hydrochloride in the presence of triethylamine. The parent compound, naphazoline, is a potent alpha-adrenoreceptor agonist (-log ED50 = 7.22), whereas the methylated derivatives, 2 and 3, were moderately potent antagonists (pA2 = 5.6 and 5.8, respectively) of the alpha-adrenoreceptor. Compounds 2 and 3 also produced blockade of the response to histamine on the rabbit aorta, but at concentrations approximately 20 times higher than necessary to produce equal blockade of the alpha-adrenoreceptor.
}, keywords = {Animals, Aorta, Histamine H1 Antagonists, Imidazoles, In Vitro Techniques, Muscle, Smooth, Naphazoline, Phenylephrine, Rabbits, Receptors, Adrenergic, Receptors, Adrenergic, alpha, Stereoisomerism}, issn = {0022-2623}, author = {Miller, D D and Hsu, F L and Ruffolo, R R and Patil, P N} } @article {1740, title = {[Acid and alkaline denaturation of superoxide dismutase].}, journal = {Biofizika}, volume = {20}, year = {1975}, month = {1975 Sep-Oct}, pages = {783-7}, abstract = {Optical and ESR spectra of erythrocyte superoxide dismutase denaturated with acid and alkali are described. Sharp changes in activity and spectra were found. "Residual" activity of alkaline denaturated protein was higher than of acidic denaturated sample. It is suggested that covalent bonding copper-nitrogen is essential for superoxide dismutase activity of the protein or synthetic copper complexes.
}, keywords = {Animals, Cattle, Erythrocytes, Hydrogen-Ion Concentration, In Vitro Techniques, Protein Denaturation, Superoxide Dismutase}, issn = {0006-3029}, author = {Simonian, M A and Nalbandian, R M} } @article {1721, title = {Acid-base balance in amphibian gastric mucosa.}, journal = {Am J Physiol}, volume = {229}, year = {1975}, month = {1975 Sep}, pages = {721-30}, abstract = {It has been established that H+ secretion can be maintained in frog stomach in the absence of exogenous CO2 by using a nutrient bathing fluid containing 25 mM H2PO4 (pH approximately equal to 4.5) or by lowering the pH of a nonbuffered nutrient solution to about 3.0-3.6. Exogenous CO2 in the presence of these nutrient solutions uniformly caused a marked decrease in H+ secretion, PD, adn short-circuit current (Isc) and an increase in transmucosal resistance (R). Elevation of nutrient [k+] to 83 mM reduced R significantly but transiently without change in H+ when nutrient pH less than 5.0, whereas R returned to base line and H+ increased when nutrient pH greater than 5.0. Acidification of the nutrient medium in the presence of exogenous CO2 results in inhibition of the secretory pump, probably by decreasing intracellular pH, and also interferes with conductance at the nutrient membrane. Removal of exogenous CO2 from standard bicarbonate nutrient solution reduced by 50\% the H+, PD, and Isc without change in R; K+-free nutrient solutions reverse these changes in Isc and PD but not in H+. The dropping PD and rising R induced by K+-free nutrient solutions in 5\% CO2 - 95\% O2 are returned toward normal by 100\% O2. Our findings support an important role for exogenous CO2 in maintaining normal acid-base balance in frog mucosa by acting as an acidifying agent.
}, keywords = {Acid-Base Equilibrium, Animals, Anura, Bicarbonates, Carbon Dioxide, Electrophysiology, Gastric Mucosa, Hydrogen-Ion Concentration, Phosphates, Potassium, Rana catesbeiana, Solutions}, issn = {0002-9513}, author = {Silen, W and Machen, T E and Forte, J G} } @article {1736, title = {Atomic models for the polypeptide backbones of myohemerythrin and hemerythrin.}, journal = {Biochem Biophys Res Commun}, volume = {66}, year = {1975}, month = {1975 Oct 27}, pages = {1349-56}, keywords = {Animals, Cnidaria, Computers, Hemerythrin, Metalloproteins, Models, Molecular, Muscle Proteins, Protein Conformation, Species Specificity}, issn = {0006-291X}, author = {Hendrickson, W A and Ward, K B} } @article {1738, title = {Bile acids. XLVII. 12alpha-Hydroxylation of precursors of allo bile acids by rabbit liver microsomes.}, journal = {Biochim Biophys Acta}, volume = {409}, year = {1975}, month = {1975 Nov 21}, pages = {249-57}, abstract = {Rabbit liver microsomal preparations fortified with 0.1 mM NADPH effectively promote hydroxylation of [3beta-3H]- or [24-14C]allochenodeoxycholic acid or [5alpha,6alpha-3H2]5alpha-cholestane-3alpha,7alpha-diol to their respective 12alpha-hydroxyl derivatives in yields of about 25 or 65\% in 60 min. Minor amounts of other products are formed from the diol. The requirements for activity of rabbit liver microsomal 12alpha-hydroxylase resemble those of rat liver microsomes. Of a number of enzyme inhibitors studied only p-chloromercuribenzoate demonstrated a marked ability to inhibit the reaction with either tritiated substrate. There was no difference in the quantity of product produced from the tritiated acid or the 14C-labeled acid. No clear sex difference was found in activity of the enzyme, nor was an appreciable difference noted in activity of the enzyme between mature and immature animals.
}, keywords = {Aging, Animals, Bile Acids and Salts, Female, Kinetics, Male, Microsomes, Liver, Mixed Function Oxygenases, NADP, Oxidation-Reduction, Rabbits, Sex Factors}, issn = {0006-3002}, author = {Ali, S S and Elliott, W H} } @article {1711, title = {Comparison between procaine and isocarboxazid metabolism in vitro by a liver microsomal amidase-esterase.}, journal = {Biochem Pharmacol}, volume = {24}, year = {1975}, month = {1975 Aug 15}, pages = {1517-21}, keywords = {Amidohydrolases, Animals, Esterases, Hydrogen-Ion Concentration, In Vitro Techniques, Isocarboxazid, Kinetics, Male, Metals, Microsomes, Liver, Phospholipids, Procaine, Proteins, Rats, Subcellular Fractions, Temperature}, issn = {0006-2952}, author = {Moroi, K and Sato, T} } @article {1725, title = {A comparison of the substrate specificities of endo-beta-N-acetylglucosaminidases from Streptomyces griseus and Diplococcus Pneumoniae.}, journal = {Biochem Biophys Res Commun}, volume = {67}, year = {1975}, month = {1975 Nov 3}, pages = {455-62}, keywords = {Acetylglucosaminidase, Fucose, Hexosaminidases, Kinetics, Oligosaccharides, Species Specificity, Streptococcus pneumoniae, Streptomyces griseus, Structure-Activity Relationship}, issn = {0006-291X}, author = {Tarentino, A L and Maley, F} } @article {1710, title = {Delineation of the intimate details of the backbone conformation of pyridine nucleotide coenzymes in aqueous solution.}, journal = {Biochem Biophys Res Commun}, volume = {66}, year = {1975}, month = {1975 Oct 27}, pages = {1173-9}, keywords = {Fourier Analysis, Magnetic Resonance Spectroscopy, Models, Molecular, Molecular Conformation, NAD, NADP, Structure-Activity Relationship, Temperature}, issn = {0006-291X}, author = {Bose, K S and Sarma, R H} } @article {1706, title = {Digitoxin metabolism by rat liver microsomes.}, journal = {Biochem Pharmacol}, volume = {24}, year = {1975}, month = {1975 Sep 1}, pages = {1639-41}, keywords = {Animals, Chromatography, Thin Layer, Digitoxigenin, Digitoxin, Hydroxylation, In Vitro Techniques, Male, Microsomes, Liver, NADP, Rats, Time Factors}, issn = {0006-2952}, author = {Schmoldt, A and Benthe, H F and Haberland, G} } @article {1723, title = {Ecological observation of the 137Cs-contamination in beef of animals from the southern-Bavarian area.}, journal = {Environ Qual Saf}, volume = {4}, year = {1975}, month = {1975}, pages = {24-36}, abstract = {Certain climatic and edaphic conformations in the Bavarian sub-alpine mountains and in the Alps favor above all the development of a land utilization system and farm structures similar to those in the northern part of Scandinavia. In 1963/64, the years of the highest environmental contamination up to the present, we established in 600 beef samples from the round or shoulder of male and female cattle (mainly Highland cattle) close connections between the 137Cs-contamination of green crop and the long lastnig yearly precipitation quantities, as well as certain relations between the 137Cs-contamination of meat and differences in the feeding and keeping of the animals. During summer-seasons (April-October), beef of cattle from pastures with heavy rainfall (Alps) was contaiminated by 137Cs up to 15 times more than that of confined animals. Hereby the rate of 137Cs-contamination in the meat of grazing cattle was nearly proportional to the quantities of precipitation. When confined cattle were fed on pastures in autumn after harvesting for 2 to 3 weeks, a quick increase of 137Cs-contamination of the meat was caused within this time up to values which in this district were otherwise only observed in grazing cattle. The lower 137Cs-content in meat of cattle housed during the summer season is due to the more varied fodder, which is at that time less contaminated than green crop. During the winter season (November to March), the highest contaminations in the meat of confined (Bohemian Forest) or grazing cattle (Alps) was measured when the animals in these districts were almost exclusively fed with fodder from the own farmground or with leafy silage. The highest contamination was almost regularly noticed in January, February and March, as generally during these months the highly contaminated first cut hay is fed. Here the meat was often even more contaminated than that of grazing cattle. After the quick decrease of 137Cs in fallout noticed in the years 1964 and 1965, in 1965/66 a dependance in the 137Cs-contamination of beef on the methods of keeping and feeding could still be observed in only the extreme cases (Alps, Bavarian- and Bohemian Forest); though in general, meat of animals from districts with heavy rainfall was slightly more contaminated than meat of animals from regions with less precipitation.
}, keywords = {Animals, Cattle, Cesium Radioisotopes, Ecology, Food Contamination, Radioactive, Fungi, Germany, West, Hydrogen-Ion Concentration, Meat, Plants, Radioactive Pollutants, Soil, Soil Microbiology, Time Factors, Trees, Weather}, issn = {0300-824X}, author = {Kreuzer, W} } @article {1729, title = {Effect of chloroquine on cultured fibroblasts: release of lysosomal hydrolases and inhibition of their uptake.}, journal = {Biochem Biophys Res Commun}, volume = {66}, year = {1975}, month = {1975 Oct 27}, pages = {1338-43}, keywords = {Biological Transport, Cells, Cultured, Cerebroside-Sulfatase, Chloroquine, Dextrans, Fibroblasts, Glucuronidase, Humans, Leukodystrophy, Metachromatic, Lysosomes, Pinocytosis, Skin, Sulfatases}, issn = {0006-291X}, author = {Wiesmann, U N and DiDonato, S and Herschkowitz, N N} } @article {1703, title = {Effect of etafenone on total and regional myocardial blood flow.}, journal = {Arzneimittelforschung}, volume = {25}, year = {1975}, month = {1975 Sep}, pages = {1400-3}, abstract = {The distribution of blood flow to the subendocardial, medium and subepicardial layers of the left ventricular free wall was studied in anaesthetized dogs under normoxic (A), hypoxic (B) conditions and under pharmacologically induced (etafenone) coronary vasodilation (C). Regional myocardial blood flow was determined by means of the particle distribution method. In normoxia a transmural gradient of flow was observed, with the subendocardial layers receiving a significantly higher flow rate compared with the subepicardial layers. In hypoxia induced vasodilation this transmural gradient of flow was persistent. In contrast a marked redistribution of regional flow was observed under pharmacologically induced vasodilation. The transmural gradient decreased. In contrast to some findings these experiments demonstrate that a considerable vasodilatory capacity exists in all layers of the myocardium and can be utilized by drugs. The differences observed for the intramural distribution pattern of flow under hypoxia and drug induced vasodilation support the hypothesis that this pattern reflects corresponding gradients of regional myocardial metabolism.
}, keywords = {Animals, Anoxia, Blood Pressure, Carbon Dioxide, Cardiac Output, Coronary Circulation, Coronary Vessels, Dogs, Heart Rate, Heart Septum, Heart Ventricles, Hydrogen-Ion Concentration, Oxygen, Propiophenones, Vasodilator Agents}, issn = {0004-4172}, author = {Flohr, H and Breull, W} } @article {1702, title = {Effect of sodium nitrate loading on electrolyte transport by the renal tubule.}, journal = {Am J Physiol}, volume = {229}, year = {1975}, month = {1975 Sep}, pages = {746-53}, abstract = {Effects of sodium nitrate were compared with sodium chloride loading on transport of electrolytes by the nephron. Maximal levels of free water clearance/clomerular filtration rate (CH2O/GFR) averaged 8.4\% with nitrate loading and 14.4\% with saline loading. Since ethacrynic acid and chlorothiazide exert their major natriuretic effect in the distal nephron, the increment in Na ad Cl reabsorbed beyond the proximal tubule. The administration of these agents resulted in an increase in fractional sodium excretion (CNa/GFR) of 21.1\%, urinary sodium excretion (UNaV) of 1,126 mueq/min, and urinary chloride excretion (UClV) of 848 mueq/min during nitrate loading compared with an increase in CNa/GFR of 37.6\%, UNaV of 2,362 mueq/min, and UClV of 2,397 mueq/min during saline loading. The smaller diuretic-induced increment in Na and Cl excretion in the nitrate studies suggests, as do the hydrated studies, that less Cl and Na are reabsorbed in the distal nephron during nitrate than saline loading. At every level of UNaV, fractional bicarbonate reabsorption was higher, urine pH was lower, and urinary potassium excretion (UKV) was higher in the nitrate studies. Thus, compared with saline loading, sodium nitrate decreases chloride and sodium reabsorption in the distal nephron. The higher hydrogen and potassium secretion in the nitrate studies may be consequent to the decreased ability of the distal nephron to reabsorb chloride.
}, keywords = {Animals, Bicarbonates, Biological Transport, Chlorides, Chlorothiazide, Diuresis, Dogs, Electrolytes, Ethacrynic Acid, Glomerular Filtration Rate, Hydrogen-Ion Concentration, Kidney Tubules, Nitrates, Potassium, Sodium, Sodium Chloride, Urine}, issn = {0002-9513}, author = {Kahn, T and Bosch, J and Levitt, M F and Goldstein, M H} } @article {1712, title = {Evaluation of amplitude and frequency changes of miniature potentials with a poor signal-to-noise ratio.}, journal = {Comp Biochem Physiol C}, volume = {52}, year = {1975}, month = {1975 Oct 1}, pages = {11-6}, keywords = {Animals, Gallamine Triethiodide, Grasshoppers, Muscle Contraction, Potentiometry, Sodium Chloride, Strychnine, Sucrose}, issn = {0306-4492}, author = {Huijbregts, L A and Schreurs, M} } @article {1701, title = {Formate assay in body fluids: application in methanol poisoning.}, journal = {Biochem Med}, volume = {13}, year = {1975}, month = {1975 Jun}, pages = {117-26}, keywords = {Aldehyde Oxidoreductases, Animals, Body Fluids, Carbon Dioxide, Formates, Haplorhini, Humans, Hydrogen-Ion Concentration, Kinetics, Methanol, Methods, Pseudomonas}, issn = {0006-2944}, author = {Makar, A B and McMartin, K E and Palese, M and Tephly, T R} } @article {1741, title = {Hydrogen peroxide and iron: a microbial cellulolytic system?}, journal = {Biotechnol Bioeng Symp}, year = {1975}, month = {1975}, pages = {151-9}, keywords = {Cellulase, Cellulose, Glycoside Hydrolases, Hydrogen Peroxide, Hydrogen-Ion Concentration, Iron, Kinetics, Mitosporic Fungi, Structure-Activity Relationship, Trichoderma}, issn = {0572-6565}, author = {Koenigs, J W} } @article {1722, title = {Influence of a new virostatic compound on the induction of enzymes in rat liver.}, journal = {Arzneimittelforschung}, volume = {25}, year = {1975}, month = {1975 Sep}, pages = {1426-9}, abstract = {The virostatic compound N,N-diethyl-4-[2-(2-oxo-3-tetradecyl-1-imidazolidinyl)-ethyl]-1-piperazinecarboxamide-hydrochloride (5531) was analyzed as to its effect on the induction of tryptophan-pyrrolase and tyrosineaminotransferase in rat liver. 1. The basic activity of the enzymes was not influenced by the substance either in normal or in adrenalectomized animals. 2. The induction of the enzymes by cortisone increased in the presence of the compound whereas the substrate induction remained unchanged. 3. The induction of tyrosine-aminotransferase by dexamethasonephosphate in tissue culture is inhibited if the dose of compound 5531 is higher than 5 mug/ml.
}, keywords = {Adrenalectomy, Animals, Antiviral Agents, Cortisone, Culture Techniques, Dose-Response Relationship, Drug, Enzyme Induction, Female, Imidazoles, Liver, Piperazines, Rats, Time Factors, Tryptophan Oxygenase, Tyrosine, Tyrosine Transaminase}, issn = {0004-4172}, author = {Kr{\"o}ger, H and Donner, I and Skiello, G} } @article {1720, title = {[Influence of beta-blocking agents on experimental atherosclerosis of cocks (author{\textquoteright}s transl)].}, journal = {Cas Lek Cesk}, volume = {114}, year = {1975}, month = {1975 Oct 24}, pages = {1320-5}, keywords = {Adrenergic beta-Antagonists, Animals, Arteriosclerosis, Chickens, Cholesterol, Dietary, Disease Models, Animal, Male}, issn = {0008-7335}, author = {Reinis, Z and Lojda, Z and Heyrovsk{\'y}, A and Hor{\'a}kov{\'a}, D and Reisenauer, R} } @article {1727, title = {Inhibition of aldehyde reductase isoenzymes in human and rat brain.}, journal = {Biochem Pharmacol}, volume = {24}, year = {1975}, month = {1975 Oct 15}, pages = {1865-9}, keywords = {Aldehyde Oxidoreductases, Animals, Anticonvulsants, Barbiturates, Brain, Glutethimide, Humans, Hydantoins, In Vitro Techniques, Isoenzymes, Kinetics, NAD, NADP, Rats, Succinimides}, issn = {0006-2952}, author = {Ris, M M and Deitrich, R A and Von Wartburg, J P} } @article {1714, title = {[Intermediate plateaux in kinetics of the reaction catalyzed by biodegradative L-threonine dehydratase from Escherichia coli].}, journal = {Biokhimiia}, volume = {40}, year = {1975}, month = {1975 May-Jun}, pages = {645-51}, abstract = {It has been shown that for the reaction catalyzed by "biodegradative" L-threonine dehydratase from E. coli strains K-12 and 980 in 0.5 M phosphate-carbonate buffer, pH 8.4 and pH 9.5, the plots of initial reaction rate (v) versus the initial substrate concentration ([S]0 are characterized by several inflection points, i. e. an intermediate plateau. The plot of v versus the allosteric activator (AMP) concentration have very complicated shapes: there are several inflection points, and also the maximum at L-threonine concentration equal to 3-10(2) and 5-10(-2) M. High AMP concentrations inhibit the enzyme at high substrate concentrations. The reduced glutathion dose not influence the enzyme and does not alter the activating effect of AMP. On the basis of the data obtained it is proposed that the substrate and AMP shift the equilibrium between multiple oligomeric enzyme forms differing in catalytic activity and kinetic manifestations of allosteric interactions between the active and allosteric AMP-binding sites towards polymerization. Thus, the functioning the enzyme under study is discussed in the frames of the model of dissociating regulatory enzymes with multiple intermediate oligomeric forms.
}, keywords = {Adenosine Monophosphate, Allosteric Regulation, Allosteric Site, Binding Sites, Escherichia coli, Hydro-Lyases, Hydrogen-Ion Concentration, Kinetics, Protein Binding, Threonine Dehydratase}, issn = {0320-9725}, author = {Sinelnikova, E M and Dvoretskova, T V and Kagan, Z S} } @article {1742, title = {Lorazepam in sexual disorders.}, journal = {Br J Clin Pract}, volume = {29}, year = {1975}, month = {1975 Jul}, pages = {175-6}, keywords = {Adult, Anti-Anxiety Agents, Anxiety, Clinical Trials as Topic, Female, Humans, Lorazepam, Male, Placebos, Psychotherapy, Sexual Dysfunction, Physiological}, issn = {0007-0947}, author = {Maneksha, S and Harry, T V} } @article {1733, title = {Lysosomal hydrolases of the epidermis. 2. Ester hydrolases.}, journal = {Br J Dermatol}, volume = {93}, year = {1975}, month = {1975 Oct}, pages = {391-8}, abstract = {Five distinct ester hydrolases (EC 3-1) have been characterized in guinea-pig epidermis. These are carboxylic esterase, acid phosphatase, pyrophosphatase, and arylsulphatase A and B. Their properties are consistent with those of lysosomal enzymes.
}, keywords = {Acid Phosphatase, Animals, Carboxylic Ester Hydrolases, Cerebroside-Sulfatase, Chondro-4-Sulfatase, Guinea Pigs, Hydrogen-Ion Concentration, Hydrolases, Kinetics, Lysosomes, Pyrophosphatases, Skin}, issn = {0007-0963}, author = {Mier, P D and van den Hurk, J J} } @article {1724, title = {Lysosomal hydrolases of the epidermis. I. Glycosidases.}, journal = {Br J Dermatol}, volume = {93}, year = {1975}, month = {1975 Jul}, pages = {1-10}, abstract = {Seven distinct glycosidases (EC 3.2) have been characterized in guinea-pig epidermis. Their properties indicate them to be of lysosomal origin. The {\textquoteright}profile{\textquoteright} of the epidermal glycosidases is significantly different from that reported for whole skin, the activities of beta-galactosidase and beta-acetylglucosaminidase being very high and those of the remaining enzymes relatively low in epidermis.
}, keywords = {Acetylglucosaminidase, Animals, Galactosidases, Glycoside Hydrolases, Guinea Pigs, Hydrogen-Ion Concentration, Kinetics, Lysosomes, Skin}, issn = {0007-0963}, author = {Mier, P D and van den Hurk, J J} } @article {1704, title = {Maturation of the adrenal medulla--IV. Effects of morphine.}, journal = {Biochem Pharmacol}, volume = {24}, year = {1975}, month = {1975 Aug 15}, pages = {1469-74}, keywords = {Adrenal Medulla, Aging, Animals, Animals, Newborn, Body Weight, Catecholamines, Dopamine beta-Hydroxylase, Epinephrine, Female, Humans, In Vitro Techniques, Maternal-Fetal Exchange, Metaraminol, Morphine, Morphine Dependence, Pregnancy, Rats, Tyrosine 3-Monooxygenase}, issn = {0006-2952}, author = {Anderson, T R and Slotkin, T A} } @article {1730, title = {Medical approach to the hypotensive patient and the patient in shock.}, journal = {Heart Lung}, volume = {4}, year = {1975}, month = {1975 Jul-Aug}, pages = {577-87}, keywords = {Adrenergic alpha-Antagonists, Anti-Bacterial Agents, Assisted Circulation, Autonomic Agents, Central Venous Pressure, Electrocardiography, Humans, Hypotension, Infusions, Parenteral, Mannitol, Plasma Substitutes, Shock, Shock, Cardiogenic, Shock, Septic, Steroids}, issn = {0147-9563}, author = {Jahre, J A and Grace, W J and Greenbaum, D M and Sarg, M J} } @article {1719, title = {Metal substitutions incarbonic anhydrase: a halide ion probe study.}, journal = {Biochem Biophys Res Commun}, volume = {66}, year = {1975}, month = {1975 Oct 27}, pages = {1281-6}, keywords = {Animals, Binding Sites, Cadmium, Carbonic Anhydrases, Cattle, Humans, Hydrogen-Ion Concentration, Magnetic Resonance Spectroscopy, Mercury, Protein Binding, Protein Conformation, Zinc}, issn = {0006-291X}, author = {Smith, R J and Bryant, R G} } @article {1713, title = {The relative importance of monoamine oxidase and catechol-O-methyl transferase on the physiologic response to administered norepinephrine in the turkey.}, journal = {Comp Biochem Physiol C}, volume = {52}, year = {1975}, month = {1975 Oct 1}, pages = {35-9}, keywords = {Animals, Blood Pressure, Catechol O-Methyltransferase, Chickens, Epinephrine, Heart, Kidney, Kinetics, Liver, Monoamine Oxidase, Myocardium, Norepinephrine, Organ Specificity, Rats, Species Specificity, Turkeys}, issn = {0306-4492}, author = {El Halawani, M E and Waibel, P E} } @article {1732, title = {Relaxation as a factor in semantic desensitization.}, journal = {J Consult Clin Psychol}, volume = {43}, year = {1975}, month = {1975 Dec}, pages = {925}, keywords = {Behavior Therapy, Desensitization, Psychologic, Female, Follow-Up Studies, Humans, Phobic Disorders, Relaxation Therapy, Semantics}, issn = {0022-006X}, author = {Bechtel, J E and McNamara, J R} } @article {1726, title = {Studies of oxygen binding energy to hemoglobin molecule.}, journal = {Biochem Biophys Res Commun}, volume = {66}, year = {1975}, month = {1975 Oct 27}, pages = {1424-31}, keywords = {Binding Sites, Cobalt, Hemoglobins, Humans, Hydrogen-Ion Concentration, Iron, Ligands, Mathematics, Oxygen, Oxyhemoglobins, Protein Binding, Spectrum Analysis}, issn = {0006-291X}, author = {Chow, Y W and Pietranico, R and Mukerji, A} } @article {1717, title = {[Studies on the action of an anticholinergic agent in combination with a tranquilizer on gastric juice secretion in man].}, journal = {Arzneimittelforschung}, volume = {25}, year = {1975}, month = {1975 Sep}, pages = {1460-3}, abstract = {A double-blind study with intra-individual comparisons was carried out to investigate the effects of 15 mg of (8r)-3alpha-hydroxy-8-isopropyl-1alphaH-tropanium bromide(+/-)-tropate (Sch 1000), 15 mg Sch 1000 + 10 mg oxazepam, 10 mg oxazepam and placebo with oral administration in randomized sequence on gastric juice volume, amount of acid, concentration and pH values in 12 healthy volunteers. The secretion parameters were measured during a 1-h basal period and a 2-h stimulation period. The gastric juice was obtained in 15 min portions via stomach tube. Stimulation was effected by 1 mug/kg/h pentagastrin via drip infusion. The Friedman test was used for the comparative statistical evaluation, and individual comparisons were carried out by means of the Wilcoxon test (pair-differences rank). The results show that Sch 1000 and Sch 1000 + oxazepam were equal in effect on basal and stimulated secretion volume. As compared with placebo, it was not possible to establish an effect on secretion volume for oxazepam alone. Sch 1000 and Sch 1000 + oxazepam were found to be equipotent in reducing the amount of basal acid, while oxazepam reduced this quantity only during the first 30 min of basal secretion. None of the three active preparations was capable of inhibiting the stimulated acid, although both Sch 1000 preparations produced a clear trend towards lowered mean values. During the basal secretion period, all three test preparations had an inhibiting action on acid concentration, but none of them had a significant effect during the stimulation period. The pH value was savely increased only by Sch 1000 and Sch 1000 + oxazepam, and this even only during the basal period. The results are discussed.
}, keywords = {Adult, Atropine Derivatives, Clinical Trials as Topic, Depression, Chemical, Drug Combinations, Female, Gastric Juice, Humans, Hydrogen-Ion Concentration, Ipratropium, Male, Middle Aged, Oxazepam, Pentagastrin, Time Factors}, issn = {0004-4172}, author = {Scherberger, R R and Kaess, H and Br{\"u}ckner, S} } @article {1705, title = {Treatment of renal hypertension.}, journal = {Clin Nephrol}, volume = {4}, year = {1975}, month = {1975 Nov}, pages = {194-7}, abstract = {There are different types of renal hypertension: hypertension due to parenchymal renal disease, renovascular hypertension, hypertension due to urological disease, hypertension of endstage renal disease. Treatment has to consider-above all-the possibility of specific, medical or surgical procedures that may cause the underlying condition. If the underlying disease is not amenable to specific therapy, symptomatic medical treatment to lower blood pressure is indicated: besides control of sodium-intake and body weight antihypertensive drugs are generally indicated. We use them, alone or in combination, in the following line of order: diuretics, beta-adrenergic blockers, dihydralazine, reserpine, clonidine, alpha-methyldopa, guanethidine.
}, keywords = {Adrenergic beta-Antagonists, Antihypertensive Agents, Diuretics, Humans, Hypertension, Renal, Kidney Diseases, Kidney Failure, Chronic, Middle Aged, Nephrectomy, Urologic Diseases}, issn = {0301-0430}, author = {Losse, H} } @article {1709, title = {[Psychoneurological problems of human adaptation to changed daily schedules].}, journal = {Kosm Biol Med}, volume = {7}, year = {1973}, month = {1973 Jul-Aug}, pages = {69-75}, keywords = {Adaptation, Psychological, Circadian Rhythm, Humans, Personality, Space Flight}, issn = {0023-4192}, author = {Kuznetsov, O N and Litsov, A N} }