@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 {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 {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 {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 {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 {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 {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 {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 {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 {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 {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 {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 {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 {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 {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 {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 {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 {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 (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 {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 {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 {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 {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 {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 {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} }