Metal-reducing Bacterium Shewanella Oneidensis MR-1 Research Articles

Single molecule tracking of bacterial cell surface cytochromes reveals dynamics that impact long-distance electron transport

Using a series of multiheme cytochromes, the metal-reducing bacterium Shewanella oneidensis MR-1 can perform extracellular electron transfer (EET) to respire redox-active surfaces, including minerals and electrodes outside the cell. While the role of multiheme cytochromes in transporting electrons across the cell wall is well established, these cytochromes were also recently found to facilitate long-distance (micrometer-scale) redox conduction along outer membranes and across multiple cells bridging electrodes. Recent studies proposed that long-distance conduction arises from the interplay of electron hopping and cytochrome diffusion, which allows collisions and electron exchange between cytochromes along membranes. However, the diffusive dynamics of the multiheme cytochromes have never been observed or quantified in vivo, making it difficult to assess their hypothesized contribution to the collision-exchange mechanism. Here, we use quantum dot labeling, total internal reflection fluorescence microscopy, and single-particle tracking to quantify the lateral diffusive dynamics of the outer membrane-associated decaheme cytochromes MtrC and OmcA, two key components of EET in S. oneidensis. We observe confined diffusion behavior for both quantum dot-labeled MtrC and OmcA along cell surfaces (diffusion coefficients DMtrC = 0.0192 ± 0.0018 µm2/s, DOmcA = 0.0125 ± 0.0024 µm2/s) and the membrane extensions thought to function as bacterial nanowires. We find that these dynamics can trace a path for electron transport via overlap of cytochrome trajectories, consistent with the long-distance conduction mechanism. The measured dynamics inform kinetic Monte Carlo simulations that combine direct electron hopping and redox molecule diffusion, revealing significant electron transport rates along cells and membrane nanowires.

Electrical properties of outer membrane extensions from Shewanella oneidensis MR-1.

Shewanella oneidensis MR-1 is a metal-reducing bacterium that is able to exchange electrons with solid-phase minerals outside the cell. These bacterial cells can produce outer membrane extensions (OMEs) that are tens of nanometers wide and several microns long. The capability of these OMEs to transport electrons is currently under investigation. Tubular chemically fixed OMEs from S. oneidensis have shown good dc conducting properties when measured in an air environment. However, no direct demonstration of the conductivity of the more common bubble-like OMEs has been provided yet, due to the inherent difficulties in measuring it. In the present work, we measured the electrical properties of bubble-like OMEs in a dry air environment by Scanning Dielectric Microscopy (SDM) in force detection mode. We found that at the frequency of the measurements (∼2 kHz), OMEs show an insulating behavior, with an equivalent homogeneous dielectric constant εOME = 3.7 ± 0.7 and no dephasing between the applied ac voltage and the measured ac electric force. The dielectric constant measured for the OMEs is comparable to that obtained for insulating supramolecular protein structures (εprotein = 3-4), pointing towards a rich protein composition of the OMEs, probably coming from the periplasm. Based on the detection sensitivity of the measuring instrument, the upper limit for the ac longitudinal conductivity of bubble-like OMEs in a dry air environment has been set to σOME,ac < 10-5 S m-1, a value several orders of magnitude smaller than the dc conductivity measured in tubular chemically fixed OMEs. The lack of conductivity of bubble-like OMEs can be attributed to the relatively large separation between cytochromes in these larger OMEs and to the suppression of cytochrome mobility due to the dry environmental conditions.

Degradation of 2, 2′, 4, 4′-Tetrabrominated diphenyl ether (BDE-47) via the Fenton reaction driven by the dissimilatory metal-reducing bacterium Shewanella oneidensis MR-1

A microbially facilitated approach was developed to degrade 2, 2′, 4, 4′-tetrabrominated diphenyl ether (BDE-47). This approach consisted of biological production of Fe(II) and H2O2 by the dissimilatory metal-reducing bacterium Shewanella oneidensis MR-1 during the repetitive anoxic/oxic cycles and abiotic production of hydroxyl radical (HO●) with the biologically produced Fe(II) and H2O2 via Fenton reaction. Under the condition tested, BDE-47 did not inhibit the growth of S. oneidensis MR-1. Water soluble Fe(III)-citrate and the solid minerals ferrihydrite [Fe(III)2O3•0.5H2O] and goethite [Fe(III)OOH] were tested in this study. Under anoxic condition, the amounts of Fe(II) produced by S. oneidensis MR-1 varied among the Fe(III)s tested, which decreased in the order of Fe(III)-citrate > ferrihydrite > goethite. Under subsequent oxic condition, H2O2 was produced via O2 reduction by S. oneidensis MR-1. The amounts of H2O2 detected also varied, which decreased in the order of the reactions with Fe(III)-citrate > goethite > ferrihydrite. S. oneidensis MR-1 maintained its ability to produce Fe(II) and H2O2 for up to seven anoxic/oxic cycles. At each end of anoxic/oxic cycle, HO● was detected. The amount of HO● produced decreased in the order of the reactions with ferrihydrite > goethite > Fe(III)-citrate, which was opposite to that of H2O2 detected. Compared to the controls without HO●, the amounts of BDE-47 in the reactions with HO● decreased. The more HO● in the reaction, the less amount of BDE-47 detected. Furthermore, no BDE-47 degradation was observed when HO● was scavenged or ferrihydrite was either omitted or replaced by nitrate. Finally, identification of degradation products, such as hydroxylated BDE-47 and trisBDE, dibromophenol and monobromophenol, suggested that OH-addition and Br-substitution by HO● were the main mechanisms for degrading BDE-47. Collectively, all these results demonstrated for the first time that the Fenton reaction driven by S. oneidensis MR-1 degraded BDE-47 effectively.

Biotic dissolution of autunite under anaerobic conditions: effect of bicarbonates and Shewanella oneidensis MR1 microbial activity.

Uranium is a contaminant of major concern across the US Department of Energy complex that served a leading role in nuclear weapon fabrication for half a century. In an effort to decrease the concentration of soluble uranium, tripolyphosphate injections were identified as a feasible remediation strategy for sequestering uranium in situ in contaminated groundwater at the Hanford Site. The introduction of sodium tripolyphosphate into uranium-bearing porous media results in the formation of uranyl phosphate minerals (autunite) of general formula {X1-2[(UO2)(PO4)]2-1·nH2O}, where X is a monovalent or divalent cation. The stability of the uranyl phosphate minerals is a critical factor that determines the long-term effectiveness of this remediation strategy that can be affected by biogeochemical factors such as the presence of bicarbonates and bacterial activity. The objective of this research was to investigate the effect of bicarbonate ions present in the aqueous phase on Ca-autunite dissolution under anaerobic conditions, as well as the role of metal-reducing facultative bacterium Shewanella oneidensis MR1. The concentration of total uranium determined in the aqueous phase was in direct correlation to the concentration of bicarbonate present in the solution, and the release of Ca, U and P into the aqueous phase was non-stoichiometric. Experiments revealed the absence of an extensive biofilm on autunite surface, while thermodynamic modeling predicted the presence of secondary minerals, which were identified through microscopy. In conclusion, the dissolution of autunite under the conditions studied is susceptible to bicarbonate concentration, as well as microbial presence.

Spin-Dependent Electron Transport through Bacterial Cell Surface Multiheme Electron Conduits.

Multiheme cytochromes, located on the bacterial cell surface, function as long-distance (>10 nm) electron conduits linking intracellular reactions to external surfaces. This extracellular electron transfer process, which allows microorganisms to gain energy by respiring solid redox-active minerals, also facilitates the wiring of cells to electrodes. While recent studies have suggested that a chiral induced spin selectivity effect is linked to efficient electron transmission through biomolecules, this phenomenon has not been investigated in extracellular electron conduits. Using magnetic conductive probe atomic force microscopy, Hall voltage measurements, and spin-dependent electrochemistry of the decaheme cytochromes MtrF and OmcA from the metal-reducing bacterium Shewanella oneidensis MR-1, we show that electron transport through these extracellular conduits is spin-selective. Our study has implications for understanding how spin-dependent interactions and magnetic fields may control electron transport across biotic-abiotic interfaces in both natural and biotechnological systems.

Surface-Induced Formation and Redox-Dependent Staining of Outer Membrane Extensions in Shewanella oneidensis MR-1

The metal-reducing bacterium Shewanella oneidensis MR-1 produces extensions of its outer membrane (OM) and periplasm that contain cytochromes responsible for extracellular electron transfer (EET) to external redox-active surfaces, including minerals and electrodes. While the role of multi-heme cytochromes in transporting electrons across the cell wall is well established, their distribution along S. oneidensis OM extensions is also thought to allow lateral electron transport along these filaments. These proposed bacterial nanowires, which can be several times the cell length, would thereby extend EET to more distant electron acceptors. However, it is still unclear why these extensions form, and to what extent they contribute to respiration in living cells. Here, we investigate physical contributors to their formation using in vivo fluorescence microscopy. While previous studies focused on the display of S. oneidensis outer membrane extensions (OMEs) as a response to oxygen limitation, we find that cell-to-surface contact is sufficient to trigger the production of OMEs, including some that reach >100 µm in length, irrespective of medium composition, agitation, or aeration. To visualize the extent of heme redox centers along OMEs, and help distinguish these structures from other extracellular filaments, we also performed histochemical redox-dependent staining with transmission electron microscopy on wild type and cytochrome-deficient strains. We demonstrate that redox-active components are limited to OMEs and not present on other extracellular appendages, such as pili and flagella. We also observed that the loss of 8 functional periplasmic and outer membrane cytochromes significantly decreased both the frequency and intensity of redox-dependent staining found widespread on OMEs. These results will improve our understanding of the environmental conditions that influence the formation of S. oneidensis OMEs, as well as the distribution and functionality of EET components along extracellular appendages.

Shewanella oneidensis MR-1 Utilizes both Sodium- and Proton-Pumping NADH Dehydrogenases during Aerobic Growth.

ABSTRACTShewanella oneidensis MR-1 is a metal-reducing bacterium with the ability to utilize many different terminal electron acceptors, including oxygen and solid-metal oxides. Both metal oxide reduction and aerobic respiration have been studied extensively in this organism. However, electron transport chain processes upstream of the terminal oxidoreductases have been relatively understudied in this organism, especially electron transfer from NADH to respiratory quinones. Genome annotation indicates that S. oneidensis MR-1 encodes four NADH dehydrogenases, a proton-translocating dehydrogenase (Nuo), two sodium ion-translocating dehydrogenases (Nqr1 and Nqr2), and an “uncoupling” dehydrogenase (Ndh), but none of these complexes have been studied. Therefore, we conducted a study specifically focused on the effects of individual NADH dehydrogenase knockouts in S. oneidensis MR-1. We observed that two of the single-mutant strains, the ΔnuoN and ΔnqrF1 mutants, exhibited significant growth defects compared with the wild type. However, the defects were minor and only apparent under certain growth conditions. Further testing of the ΔnuoN ΔnqrF1 double-mutant strain yielded no growth in minimal medium under oxic conditions, indicating that Nuo and Nqr1 have overlapping functions, but at least one is necessary for aerobic growth. Coutilization of proton- and sodium ion-dependent energetics has important implications for the growth of this organism in environments with varied pH and salinity, including microbial electrochemical systems.IMPORTANCE Bacteria utilize a wide variety of metabolic pathways that allow them to take advantage of different energy sources, and to do so with varied efficiency. The efficiency of a metabolic process determines the growth yield of an organism, or the amount of biomass it produces per amount of substrate consumed. This parameter has important implications in biotechnology and wastewater treatment, where low growth yields are often preferred to minimize the production of microbial biomass. In this study, we investigated respiratory pathways containing NADH dehydrogenases with varied efficiency (i.e., the number of ions translocated per NADH oxidized) in the metal-reducing bacterium Shewanella oneidensis MR-1. We observed that two different respiratory pathways are used concurrently, and at least one pathway must be functional for growth under oxic conditions.

A mechanistic study of TiO2 nanoparticle toxicity on Shewanella oneidensis MR-1 with UV-containing simulated solar irradiation: Bacterial growth, riboflavin secretion, and gene expression

Toxicity of nanomaterials to ecological systems has recently emerged as an important field of research, and thus, many researchers are exploring the mechanisms of how nanoparticles impact organisms. Herein, we probe the mechanisms of bacteria-nanoparticle interaction by investigating how TiO2 nanoparticles impact a model organism, the metal-reducing bacterium Shewanella oneidensis MR-1. In addition to examining the effect of TiO2 exposure, the effect of synergistic simulated solar irradiation containing UV was explored in this study, as TiO2 nanoparticles are known photocatalysts. The data reveal that TiO2 nanoparticles cause an inhibition of S. oneidensis growth at high dosage without compromising cell viability, yet co-exposure of nanoparticles and illumination does not increase the adverse effects on bacterial growth relative to TiO2 alone. Measurements of intracellular reactive oxygen species and riboflavin secretion, on the same nanoparticle-exposed bacteria, reveal that TiO2 nanoparticles have no effect on these cell functions, but application of UV-containing illumination with TiO2 nanoparticles has an impact on the level of riboflavin outside bacterial cells. Finally, gene expression studies were employed to explore how cells respond to TiO2 nanoparticles and illumination, and these results were correlated with cell growth and cell function assessment. Together these data suggest a minimal impact of TiO2 NPs and simulated solar irradiation containing UV on S. oneidensis MR-1, and the minimal impact could be accounted for by the nutrient-rich medium used in this work. These measurements demonstrate a comprehensive scheme combining various analytical tools to enable a mechanistic understanding of nanoparticle-cell interactions and to evaluate the potential adverse effects of nanoparticles beyond viability/growth considerations.

Microbial reduction of Cr(VI) in the presence of chromate conversion coating constituents

ABSTRACTThe reduction of Cr(VI) by the metal-reducing bacterium Shewanella oneidensis MR-1 was evaluated, to determine the potential for exploiting Cr(VI) bioreduction as a means of treating chromate conversion coating (CCC) waste streams. Inclusion of Cr(VI) at concentrations ≥1 mM inhibited aerobic growth of S. oneidensis, but that organism was able to reduce Cr(VI) at a concentration of up to 1 mM under anaerobic, nongrowth conditions. S. oneidensis reduced Cr(VI) in the presence of common CCC constituents, with the exception of ferricyanide, when these CCC constituents were included at concentrations typical of CCC waste streams. Ferricyanide inhibited neither aerobic growth nor metabolism under aerobic, nitrate- or iron-reducing conditions, suggesting that the ferricyanide-depended inhibition of Cr(VI) reduction is not due to broad metabolic inhibition, but is specific to Cr(VI) reduction. Results indicate that under some conditions, the activities of metal-reducing bacteria, such as S. oneidensis, could be exploited for the removal of Cr(VI) from CCC waste streams under appropriate conditions.

Electron Cryo-Tomography of Nanowires in Shewanella Oneidensis MR-1

The dissimilatory metal-reducing bacterium Shewanella oneidensis MR-1 is known to make ‘nanowires,’ conductive cell appendages used in electron transport. Novel live fluorescence imaging techniques recently clarified that the conductive filaments in S. oneidensis are not pili, as previously thought, but rather extensions of the outer membrane. However, their fine structure and mechanism remained unclear. Here, we report the first high-resolution in vivo images of nanowires in frozen-hydrated cells in a near-native state by electron cryo-tomography (ECT). We first developed a method for in vivo production of nanowires in cells grown on EM grids. Using correlative light microscopy and ECT, we then identified and imaged nanowires at high resolution and in three dimensions. Interestingly, we find that nanowires are composed of outer membrane vesicles (OMVs) strung together to form a chain. The size of the vesicles and the length and thickness of the vesicle chain are highly variable. However, even apparently smooth filaments such as those described previously are in fact, at higher resolution, revealed to be chains of OMVs. This work will help us investigate the electron transport mechanism along Shewanella nanowires. Additionally, this work has implications in understanding how OMVs, ubiquitous in Gram-negative bacteria, can assemble into chains.

Multiheme Cytochromes and the Bacterial Nanowires of Shewanella oneidensis MR-1: Regulation, Structure, and Extracellular Electron Transport Mechanisms

Dissimilatory metal-reducing bacteria can extract free energy from their environment by performing electron transfer to solid-phase minerals outside the cell. This extracellular electron transport (EET) has important implications in global elemental cycles as well as renewable energy technologies. Among the pathways for EET, bacterial nanowires have received significant attention in the past decade due to their unique ability to mediate long-range electron transport to electron acceptors microns away from the cell surface. Here we report a comprehensive characterization of the composition, structure, and regulatory network that underlies bacterial nanowires from the metal-reducing bacterium Shewanella oneidensis MR-1. Using fluorescent and atomic force techniques, we find that the Shewanella nanowires are extensions of the outer membrane and periplasm that contain multiple multiheme cytochromes. The localization of decaheme cytochromes MtrC and OmcA supports a multistep redox hopping mechanism, allowing long-range electron transport along a membrane network of heme cofactors that line the nanowires. The electron flux resulting from such a mechanism strongly depends on the cytochrome density and topology. Using correlated electron cryo-tomography and in vivo fluorescent microscopy, we are gaining new insight into the localization patterns of cytochromes along nanowires as well as the morphology and the formation mechanism of these structures. Finally, we report our progress on understanding the underlying regulatory network, by testing targeted mutations and analyzing the transcriptome of Shewanella chemostat cultures as they encounter electron acceptor limitation and form nanowires. The transcriptional response includes an increase in the expression of multiheme cytochromes, heme synthesis enzymes, and cytochrome maturation proteins. Our findings on the regulation, ultrastructure and electron transport mechanism help shape a biophysical understanding of these redox-functionalized membrane and vesicular extensions as a microbial strategy for electron transport and energy distribution.

Photocatalytic properties of zinc sulfide nanocrystals biofabricated by metal-reducing bacterium Shewanella oneidensis MR-1

Accumulation and utilization of heavy metals from wastewater by biological treatment system has aroused great interest. In the present study, a metal-reducing bacterium Shewanella oneidensis MR-1 was used to explore the biofabrication of ZnS nanocrystals from the artificial wastewater. The biogenic H2S produced via the reduction of thiosulfate precipitated the Zn(II) as sulfide extracellularly. Characterization by X-ray diffraction (XRD), high-resolution transmission electron microscopy (HRTEM), and field emission scanning electron microscope (FESEM) confirmed the precipitates as ZnS nanocrystals. The biogenic ZnS nanocrystals appeared spherical in shape with an average diameter of 5nm and mainly aggregated in the medium and cell surface of S. oneidensis MR-1. UV–vis DRS spectra showed ZnS nanoparticles appeared a strong absorption below 360nm. Thus, the photocatalytic activity of ZnS was evaluated by the photodegradation of rhodamine B (RhB) under UV irradiation. The biogenic ZnS nanocrystals showed a high level of photodegradation efficiency to RhB coupled with a significant blue-shift of maximum adsorption peak. A detailed analysis indicated the photogenerated holes, rather than hydroxyl radicals, contributed to the photocatalytic decolorization of RhB. This approach of coupling biosynthesis of nanoparticles with heavy metal removal may offer a potential avenue for efficient bioremediation of heavy metal wastewater.

Bacterial Nanowires of Shewanella Oneidensis MR-1 are Outer Membrane and Periplasmic Extensions of the Extracellular Electron Transport Components

Bacterial nanowires offer a pathway for extracellular electron transfer (EET) by linking the respiratory chain of bacteria to external surfaces, including oxidized metals in the environment and engineered electrodes in renewable energy devices. Specifically, nanowires of the model metal-reducing bacterium Shewanella oneidensis MR-1 were previously shown to be conductive under non-physiological conditions. Despite the global, environmental, and technological consequences of bacterial nanowire-mediated EET, the composition, electron transport mechanism, and physiological relevance of these appendages remain unclear. The nanowires of S. oneidensis MR-1 were previously thought, but never shown, to be bacterial pili. In addition, the transport mechanism through bacterial nanowires has been the subject of intense debate, with “metallic-like” band transport and multistep redox hopping between multiheme cytochromes as the two proposed mechanisms. Here we report the first in vivo observations of the formation and respiratory impact of nanowires in S. oneidensis MR-1. Using live fluorescence measurements and quantitative gene expression analysis, we demonstrate that S. oneidensis MR-1 nanowires are extensions of the outer membrane and periplasm, rather than pilin-based structures. We show, through immunolabeling, that multiheme cytochromes localize to nanowires, in turn supporting the multistep redox hopping model as the transport mechanism. Furthermore, these bacterial nanowires are associated with outer membrane vesicles, structures ubiquitous in Gram-negative bacteria, and occasionally appear as membrane vesicle chains that transition to smoother filaments. Redox-functionalized membrane and vesicular extensions may represent a general microbial strategy for electron transport and energy distribution.

Accumulation of amorphous Cr(III)-Te(IV) nanoparticles on the surface of Shewanella oneidensis MR-1 through reduction of Cr(VI).

Industrial effluents constitute a major source of metal pollution of aquatic bodies. Moreover, due to their environmental persistence, toxic metal pollution is of special concern. Microbial reduction is considered a promising strategy for toxic metal removal among the several methods available for metal remediation. Here, we describe the coremediation of toxic Cr(VI) and Te(IV) by the dissimilatory metal reducing bacterium-Shewanella oneidensis MR-1. In the presence of both Cr(VI) and Te(IV), S. oneidensis MR-1 reduced Cr(VI) to the less toxic Cr(III) form, but not Te(IV) to Te(0). The reduced Cr(III) ions complexed rapidly with Te(IV) ions and were precipitated from the cell cultures. Electron microscopic analyses revealed that the Cr-Te complexed nanoparticles localized on the bacterial outer membranes. K-edge X-ray absorption spectrometric analyses demonstrated that Cr(III) produced by S. oneidensis MR-1 was rapidly complexed with Te(IV) ions, followed by formation of amorphous Cr(III)-Te(IV) nanoparticles on the cell surface. Our results could be applied for the simultaneous sequestration and detoxification of both Cr(VI) and Te(IV) as well as for the preparation of nanomaterials through environmental friendly processes.

Conservation of transcription start sites within genes across a bacterial genus.

ABSTRACTTranscription start sites (TSSs) lying inside annotated genes, on the same or opposite strand, have been observed in diverse bacteria, but the function of these unexpected transcripts is unclear. Here, we use the metal-reducing bacterium Shewanella oneidensis MR-1 and its relatives to study the evolutionary conservation of unexpected TSSs. Using high-resolution tiling microarrays and 5′-end RNA sequencing, we identified 2,531 TSSs in S. oneidensis MR-1, of which 18% were located inside coding sequences (CDSs). Comparative transcriptome analysis with seven additional Shewanella species revealed that the majority (76%) of the TSSs within the upstream regions of annotated genes (gTSSs) were conserved. Thirty percent of the TSSs that were inside genes and on the sense strand (iTSSs) were also conserved. Sequence analysis around these iTSSs showed conserved promoter motifs, suggesting that many iTSS are under purifying selection. Furthermore, conserved iTSSs are enriched for regulatory motifs, suggesting that they are regulated, and they tend to eliminate polar effects, which confirms that they are functional. In contrast, the transcription of antisense TSSs located inside CDSs (aTSSs) was significantly less likely to be conserved (22%). However, aTSSs whose transcription was conserved often have conserved promoter motifs and drive the expression of nearby genes. Overall, our findings demonstrate that some internal TSSs are conserved and drive protein expression despite their unusual locations, but the majority are not conserved and may reflect noisy initiation of transcription rather than a biological function.

Bacterial synthesis of silver sulfide nanoparticles

The metal-reducing bacterium Shewanella oneidensis MR-1 has been employed to obtain Ag2S nanoparticles from an aqueous solution of AgNO3 and Na2S2O3 at an ordinary temperature and pressure. The nanoparticles vary in size within 2–16 nm, and the fraction 6 to 12 nm in size constitutes about 70%. The maximum yield of nanoparticles in silver equivalent is 53%. Being visualized by transmission electron microscopy, the particles look like spheres with average diameters varying from 7 ± 2 nm to 9 ± 2 nm. The elemental composition of synthesized nanoparticles has been analyzed by energy-dispersive X-ray spectroscopy, and the estimated silver to sulfur atomic ratio is 2: 1. The presence of living bacterial cells is mandatory for the formation of Ag2S nanoparticles in the aqueous salt solution. Changes in the reaction conditions (reagent concentrations, temperature, and cell-incubation time in the reaction mixture) influence the yield of nanoparticles dramatically, but have little influence on their size.

Single-cell imaging and spectroscopic analyses of Cr(VI) reduction on the surface of bacterial cells.

We investigate the single-cell reduction of toxic Cr(VI) by the dissimilatory metal-reducing bacterium Shewanella oneidensis MR-1 (MR-1), an important bioremediation process, using Raman spectroscopy and scanning electron microscopy (SEM) combined with energy-dispersive X-ray spectroscopy (EDX). Our experiments indicate that the toxic, highly soluble Cr(VI) can be efficiently reduced to less toxic, nonsoluble Cr(2)O(3) nanoparticles by MR-1. Cr(2)O(3) is observed to emerge as nanoparticles adsorbed on the cell surface and its chemical nature is identified by EDX imaging and Raman spectroscopy. Co-localization of Cr(2)O(3) and cytochromes by EDX imaging and Raman spectroscopy suggests a terminal reductase role for MR-1 surface-exposed cytochromes MtrC and OmcA. Our experiments revealed that the cooperation of surface proteins OmcA and MtrC makes the reduction reaction most efficient, and the sequence of the reducing reactivity of MR-1 is wild type > single mutant ΔmtrC or mutant ΔomcA > double mutant (ΔomcA-ΔmtrC). Moreover, our results also suggest that direct microbial Cr(VI) reduction and Fe(II) (hematite)-mediated Cr(VI) reduction mechanisms may coexist in the reduction processes.

Mtr extracellular electron-transfer pathways in Fe(III)-reducing or Fe(II)-oxidizing bacteria: a genomic perspective

Originally discovered in the dissimilatory metal-reducing bacterium Shewanella oneidensis MR-1 (MR-1), key components of the Mtr (i.e. metal-reducing) pathway exist in all strains of metal-reducing Shewanella characterized. The protein components identified to date for the Mtr pathway of MR-1 include four multihaem c-Cyts (c-type cytochromes), CymA, MtrA, MtrC and OmcA, and a porin-like outer membrane protein MtrB. They are strategically positioned along the width of the MR-1 cell envelope to mediate electron transfer from the quinone/quinol pool in the inner membrane to Fe(III)-containing minerals external to the bacterial cells. A survey of microbial genomes has identified homologues of the Mtr pathway in other dissimilatory Fe(III)-reducing bacteria, including Aeromonas hydrophila, Ferrimonas balearica and Rhodoferax ferrireducens, and in the Fe(II)-oxidizing bacteria Dechloromonas aromatica RCB, Gallionella capsiferriformans ES-2 and Sideroxydans lithotrophicus ES-1. The apparent widespread distribution of Mtr pathways in both Fe(III)-reducing and Fe(II)-oxidizing bacteria suggests a bidirectional electron transfer role, and emphasizes the importance of this type of extracellular electron-transfer pathway in microbial redox transformation of iron. The organizational and electron-transfer characteristics of the Mtr pathways may be shared by other pathways used by micro-organisms for exchanging electrons with their extracellular environments.

Identification of diverse carbon utilization pathways in Shewanella oneidensis MR-1 via expression profiling.

To identify pathways of carbon utilization in the metal-reducing marine bacterium Shewanella oneidensis MR-1, we assayed the expression of cells grown with various carbon sources using a high-density oligonucleotide Affymetrix microarray. Our expression profiles reveal genes and regulatory mechanisms which govern the sensing, import, and utilization of the nucleoside inosine, the chitin monomer N-acetylglucosamine, and a casein-derived mixture of amino acids. Our analysis suggests a prominent role for the pentose-phosphate and Entner-Doudoroff pathways in energy metabolism, and regulatory coupling between carbon catabolism and electron acceptor pathways. In sum, these results indicate that S. oneidensis possesses a broader capacity for carbon utilization than previously reported, a view with implications for optimizing its role in microbial fuel cell and bioremediative applications.

Physical constraints on charge transport through bacterial nanowires.

Extracellular appendages of the dissimilatory metal-reducing bacterium Shewanella oneidensis MR-1 were recently shown to sustain currents of 10(10) electrons per second over distances of 0.5 microns [El-Naggar et al., Proc. Natl. Acad. Sci. U. S. A., 2010, 107, 18127]. However, the identity of the charge localizing sites and their organization along the "nanowire" remain unknown. We use theory to predict redox cofactor separation distances that would permit charge flow at rates of 10(10) electrons per second over 0.5 microns for voltage biases of < or = IV, using a steady-state analysis governed by a non-adiabatic electron transport mechanism. We find the observed currents necessitate a multi-step hopping transport mechanism, with charge localizing sites separated by less than 1 nm and reorganization energies that rival the lowest known in biology.