Microbial reduction of structural iron in interstratified illite-smectite minerals by a sulfate-reducing bacterium

Clay minerals are ubiquitous in soils, sediments, and sedimentary rocks and could coexist with sulfate‐reducing bacteria (SRB) in anoxic environments, however, the interactions of clay minerals and SRB are not well understood. The objective of this study was to understand the reduction rate and capacity of structural Fe(III) in dioctahedral clay minerals by a mesophilic SRB, Desulfovibrio vulgaris and the potential role in catalyzing smectite illitization. Bioreduction experiments were performed in batch systems, where four different clay minerals (nontronite NAu‐2, mixed‐layer illite‐smectite RAr‐1 and ISCz‐1, and illite IMt‐1) were exposed to D. vulgaris in a non‐growth medium with and without anthraquinone‐2,6‐disulfonate (AQDS) and sulfate. Our results demonstrated that D. vulgaris was able to reduce structural Fe(III) in these clay minerals, and AQDS enhanced the reduction rate and extent. In the presence of AQDS, sulfate had little effect on Fe(III) bioreduction. In the absence of AQDS, sulfate increased the reduction rate and capacity, suggesting that sulfide produced during sulfate reduction reacted with the phyllosilicate Fe(III). The extent of bioreduction of structural Fe(III) in the clay minerals was positively correlated with the percentage of smectite and mineral surface area of these minerals. X‐ray diffraction, and scanning and transmission electron microscopy results confirmed formation of illite after bioreduction. These data collectively showed that D. vulgaris could promote smectite illitization through reduction of structural Fe(III) in clay minerals.     

Hydrogen metabolism in Shewanella oneidensis MR-1

Shewanella oneidensis MR-1 is a facultative sediment microorganism which uses diverse compounds, such as oxygen and fumarate, as well as insoluble Fe(III) and Mn(IV) as electron acceptors. The electron donor spectrum is more limited and includes metabolic end products of primary fermenting bacteria, such as lactate, formate, and hydrogen. While the utilization of hydrogen as an electron donor has been described previously, we report here the formation of hydrogen from pyruvate under anaerobic, stationary-phase conditions in the absence of an external electron acceptor. Genes for the two S. oneidensis MR-1 hydrogenases, hydA, encoding a periplasmic [Fe-Fe] hydrogenase, and hyaB, encoding a periplasmic [Ni-Fe] hydrogenase, were found to be expressed only under anaerobic conditions during early exponential growth and into stationary-phase growth. Analyses of DeltahydA, DeltahyaB, and DeltahydA DeltahyaB in-frame-deletion mutants indicated that HydA functions primarily as a hydrogen-forming hydrogenase while HyaB has a bifunctional role and represents the dominant hydrogenase activity under the experimental conditions tested. Based on results from physiological and genetic experiments, we propose that hydrogen is formed from pyruvate by multiple parallel pathways, one pathway involving formate as an intermediate, pyruvate-formate lyase, and formate-hydrogen lyase, comprised of HydA hydrogenase and formate dehydrogenase, and a formate-independent pathway involving pyruvate dehydrogenase. A reverse electron transport chain is potentially involved in a formate-hydrogen lyase-independent pathway. While pyruvate does not support a fermentative mode of growth in this microorganism, pyruvate, in the absence of an electron acceptor, increased cell viability in anaerobic, stationary-phase cultures, suggesting a role in the survival of S. oneidensis MR-1 under stationary-phase conditions.     

Growth of Thermophilic and Hyperthermophilic Fe(III)-Reducing Microorganisms on a Ferruginous Smectite as the Sole Electron Acceptor

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.     

Morphological and metabolic responses to starvation by the dissimilatory metal-reducing bacterium Shewanella alga BrY.

The response of the dissimilatory metal-reducing bacterium Shewanella alga BrY to carbon and nitrogen starvation was examined. Starvation resulted in a gradual decrease in the mean cell volume from 0.48 to 0.2 micron 3 and a dramatic decrease in Fe(III) reductase activity. Growth of starved cultures was initiated with O2, ferric oxyhydroxide, Co(III)-EDTA, or Fe(III)-bearing subsurface materials as the sole electron acceptor. Microbially reduced subsurface materials reduced CrO(4)2-. Starvation of dissimilatory metal-reducing bacteria may provide a means of delivering this metabolism to contaminated subsurface environments for in situ bioremediation.     

Involvement of cyclic AMP (cAMP) and cAMP receptor protein in anaerobic respiration of Shewanella oneidensis

Shewanella oneidensis is a metal reducer that can use several terminal electron acceptors for anaerobic respiration, including fumarate, nitrate, dimethyl sulfoxide (DMSO), trimethylamine N-oxide (TMAO), nitrite, and insoluble iron and manganese oxides. Two S. oneidensis mutants, SR-558 and SR-559, with Tn5 insertions in crp, were isolated and analyzed. Both mutants were deficient in Fe(III) and Mn(IV) reduction. They were also deficient in anaerobic growth with, and reduction of, nitrate, fumarate, and DMSO. Although nitrite reductase activity was not affected by the crp mutation, the mutants failed to grow with nitrite as a terminal electron acceptor. This growth deficiency may be due to the observed loss of cytochromes c in the mutants. In contrast, TMAO reduction and growth were not affected by loss of cyclic AMP (cAMP) receptor protein (CRP). Fumarate and Fe(III) reductase activities were induced in rich medium by the addition of cAMP to aerobically growing wild-type S. oneidensis. These results indicate that CRP and cAMP play a role in the regulation of anaerobic respiration, in addition to their known roles in catabolite repression and carbon source utilization in other bacteria.     

Chemotactic responses to metals and anaerobic electron acceptors in Shewanella oneidensis MR-1

Although a previous study indicated that the dissimilatory metal-reducing bacterium Shewanella oneidensis MR-1 lacks chemotactic responses to metals that can be used as anaerobic electron acceptors, new results show that this bacterium responds to both Mn(III) and Fe(III). Cells were also shown to respond to another unusual electron acceptor, the humic acid analog anthraquinone-2,6-disulfonate. These results indicate that S. oneidensis is capable of moving towards a number of unusual anaerobic electron acceptors, including some that would normally be insoluble in the environment. Additionally, S. oneidensis was shown to migrate in gradients of several divalent cations under anaerobic conditions. Although responses to the reduced forms of redox-active metals, such as Mn(II) and Fe(II), might indicate that S. oneidensis uses gradients of these metals to locate the insoluble electron acceptors Mn(III/IV) and Fe(III) for dissimilatory purposes, responses to non-redox-active metals, such as Zn(II), suggest that movement towards divalent cations might serve other, potentially assimilatory, purposes.     

Plutonium(IV) reduction by the metal-reducing bacteria Geobacter metallireducens GS15 and Shewanella oneidensis MR1

The bacterial reduction of actinides has been suggested as a possible remedial strategy for actinide-contaminated environments, and the bacterial reduction of Pu(VI/V) has the potential to produce highly insoluble Pu(IV) solid phases. However, the behavior of plutonium with regard to bacterial reduction is more complex than for other actinides because it is possible for Pu(IV) to be further reduced to Pu(III), which is relatively more soluble than Pu(IV). This work investigates the ability of the metal-reducing bacteria Geobacter metallireducens GS15 and Shewanella oneidensis MR1 to enzymatically reduce freshly precipitated amorphous Pu(IV) (OH)(4) [Pu(IV)(OH)(4(am))] and soluble Pu(IV)(EDTA). In cell suspensions without added complexing ligands, minor Pu(III) production was observed in cultures containing S. oneidensis, but little or no Pu(III) production was observed in cultures containing G. metallireducens. In the presence of EDTA, most of the Pu(IV)(OH)(4(am)) present was reduced to Pu(III) and remained soluble in cell suspensions of both S. oneidensis and G. metallireducens. When soluble Pu(IV)(EDTA) was provided as the terminal electron acceptor, cell suspensions of both S. oneidensis and G. metallireducens rapidly reduced Pu(IV)(EDTA) to Pu(III)(EDTA) with nearly complete reduction within 20 to 40 min, depending on the initial concentration. Neither bacterium was able to use Pu(IV) (in any of the forms used) as a terminal electron acceptor to support growth. These results have significant implications for the potential remediation of plutonium and suggest that strongly reducing environments where complexing ligands are present may produce soluble forms of reduced Pu species.     

Anaerobic Biooxidation of Fe(II) by Dechlorosoma suillum

Anaerobic microbial oxidation of Fe(II) was only recently discovered and very little is known about this metabolism. We recently demonstrated that several dissimilatory perchlorate-reducing bacteria could utilize Fe(II) as an electron donor under anaerobic conditions. Here we report on a more in-depth analysis of Fe(II) oxidation by one of these organisms, Dechlorosoma suillum. Similarly to most known nitrate-dependent Fe(II) oxidizers, D. suillum did not grow heterotrophically or lithoautotrophically by anaerobic Fe(II) oxidation. In the absence of a suitable organic carbon source, cells rapidly lysed even though nitrate-dependent Fe(II) oxidation was still occurring. The coupling of Fe(II) oxidation to a particular electron acceptor was dependent on the growth conditions of cells of D. suillum. As such, anaerobically grown cultures of D. suillum did not mediate Fe(II) oxidation with oxygen as the electron acceptor, while conversely, aerobically grown cultures did not mediate Fe(II) oxidation with nitrate as the electron acceptor. Anaerobic washed cell suspensions of D. suillum rapidly produced an orange/brown precipitate which X-ray diffraction analysis identified as amorphous ferric oxyhydroxide or ferrihydrite. This is similar to all other identified nitrate-dependent Fe(II) oxidizers but is in contrast to what is observed for growth cultures of D. suillum, which produced a mixed-valence Fe(II)-Fe(III) precipitate known as green rust. D. suillum rapidly oxidized the Fe(II) content of natural sediments. Although the form of ferrous iron in these sediments is unknown, it is probably a component of an insoluble mineral, as previous studies indicated that soluble Fe(II) is a relatively minor form of the total Fe(II) content of anoxic environments. The results of this study further enhance our knowledge of a poorly understood form of microbial metabolism and indicate that anaerobic Fe(II) oxidation by D. suillum is significantly different from previously described forms of nitrate-dependent microbial Fe(II) oxidation.     

Melanin production and use as a soluble electron shuttle for Fe(III) oxide reduction and as a terminal electron acceptor by Shewanella algae BrY

Dissimilatory metal-reducing bacteria (DMRB) utilize numerous compounds as terminal electron acceptors, including insoluble iron oxides. The mechanism(s) of insoluble-mineral reduction by DMRB is not well understood. Here we report that extracellular melanin is produced by Shewanella algae BrY. The extracted melanin served as the sole terminal electron acceptor. Upon reduction the reduced, soluble melanin reduced insoluble hydrous ferric oxide in the absence of bacteria, thus demonstrating that melanin produced by S. algae BrY is a soluble Fe(III)-reducing compound. In the presence of bacteria, melanin acted as an electron conduit to Fe(III) minerals and increased Fe(III) mineral reduction rates. Growth of S. algae BrY occurred in anaerobic minimal medium supplemented with melanin extracted from previously grown aerobic cultures of S. algae BrY. Melanin produced by S. algae BrY imparts increased versatility to this organism as a soluble Fe(III) reductant, an electron conduit for iron mineral reduction, and a sole terminal electron acceptor that supports growth.     

Use of Ferric and Ferrous Iron Containing Minerals for Respiration by Desulfitobacterium frappieri

The potential of Desulfitobacterium frappieri strain G2, which was isolated from subsurface smectite bedding, to participate in iron redox reactions was investigated. Strain G2 can use poorly crystalline Fe(III) oxide, soluble forms of Fe(III) and Fe(III) in the structure of phyllosilicate minerals as electron acceptors. It can also oxidize Fe(II)-NTA or Fe(II) in the structure of phyllosilicate minerals with nitrate as the electron acceptor. These results suggest for the first time that strains of Desulfitobacterium frappieri may play an important role in iron cycling in sedimentary environments.     

Dissimilatory iron reduction in Escherichia coli: identification of CymA of Shewanella oneidensis and NapC of E. coli as ferric reductases

Over geological time scales, microbial reduction of chelated Fe(III) or Fe(III) minerals has profoundly affected today's composition of our bio- and geosphere. However, the electron transfer reactions that are specific and defining for dissimilatory iron(III)-reducing (DIR) bacteria are not well understood. Using a synthetic biology approach involving the reconstruction of the putative electron transport chain of the DIR bacterium Shewanella oneidensis MR-1 in Escherichia coli , we showed that expression of cymA was necessary and sufficient to convert E. coli into a DIR bacterium. In intact cells, the Fe(III)-reducing activity was limited to Fe(III) NTA as electron acceptor. In vitro biochemical analysis indicated that CymA, which is a cytoplasmic membrane-associated tetrahaem c -type cytochrome, carries reductase activity towards Fe(III) NTA, Fe(III) citrate, as well as to AQDS, a humic acid analogue. The in vitro specific activities of Fe(III) citrate reductase and AQDS reductase of E. coli spheroplasts were 10× and 30× higher, respectively, relative to the specific rates observed in intact cells, suggesting that access of chelated and insoluble forms of Fe(III) and AQDS is restricted in whole cells. Interestingly, the E. coli CymA orthologue NapC also carried ferric reductase activity. Our data support the argument that the biochemical mechanism of Fe(III) reduction per se was not the key innovation leading to environmental relevant DIR bacteria. Rather, the evolution of an extension of the electron transfer pathway from the Fe(III) reductase CymA to the cell surface via a system of periplasmic and outer membrane cytochrome proteins enabled access to diffusion-impaired electron acceptors.     

Specific bonds between an iron oxide surface and outer membrane cytochromes MtrC and OmcA from Shewanella oneidensis MR-1

Shewanella oneidensis MR-1 is purported to express outer membrane cytochromes (e.g., MtrC and OmcA) that transfer electrons directly to Fe(III) in a mineral during anaerobic respiration. A prerequisite for this type of reaction would be the formation of a stable bond between a cytochrome and an iron oxide surface. Atomic force microscopy (AFM) was used to detect whether a specific bond forms between a hematite (Fe(2)O(3)) thin film, created with oxygen plasma-assisted molecular beam epitaxy, and recombinant MtrC or OmcA molecules coupled to gold substrates. Force spectra displayed a unique force signature indicative of a specific bond between each cytochrome and the hematite surface. The strength of the OmcA-hematite bond was approximately twice that of the MtrC-hematite bond, but direct binding to hematite was twice as favorable for MtrC. Reversible folding/unfolding reactions were observed for mechanically denatured MtrC molecules bound to hematite. The force measurements for the hematite-cytochrome pairs were compared to spectra collected for an iron oxide and S. oneidensis under anaerobic conditions. There is a strong correlation between the whole-cell and pure-protein force spectra, suggesting that the unique binding attributes of each cytochrome complement one another and allow both MtrC and OmcA to play a prominent role in the transfer of electrons to Fe(III) in minerals. Finally, by comparing the magnitudes of binding force for the whole-cell versus pure-protein data, we were able to estimate that a single bacterium of S. oneidensis (2 by 0.5 microm) expresses approximately 10(4) cytochromes on its outer surface.     

Melanin Production and Use as a Soluble Electron Shuttle for Fe(III) Oxide Reduction and as a Terminal Electron Acceptor by Shewanella algae BrY

Dissimilatory metal-reducing bacteria (DMRB) utilize numerous compounds as terminal electron acceptors, including insoluble iron oxides. The mechanism(s) of insoluble-mineral reduction by DMRB is not well understood. Here we report that extracellular melanin is produced by Shewanella algae BrY. The extracted melanin served as the sole terminal electron acceptor. Upon reduction the reduced, soluble melanin reduced insoluble hydrous ferric oxide in the absence of bacteria, thus demonstrating that melanin produced by S. algae BrY is a soluble Fe(III)-reducing compound. In the presence of bacteria, melanin acted as an electron conduit to Fe(III) minerals and increased Fe(III) mineral reduction rates. Growth of S. algae BrY occurred in anaerobic minimal medium supplemented with melanin extracted from previously grown aerobic cultures of S. algae BrY. Melanin produced by S. algae BrY imparts increased versatility to this organism as a soluble Fe(III) reductant, an electron conduit for iron mineral reduction, and a sole terminal electron acceptor that supports growth.     

Morphology of biogenic iron oxides records microbial physiology and environmental conditions: toward interpreting iron microfossils

Despite the abundance of Fe and its significance in Earth history, there are no established robust biosignatures for Fe(II)‐oxidizing micro‐organisms. This limits our ability to piece together the history of Fe biogeochemical cycling and, in particular, to determine whether Fe(II)‐oxidizers played a role in depositing ancient iron formations. A promising candidate for Fe(II)‐oxidizer biosignatures is the distinctive morphology and texture of extracellular Fe(III)‐oxyhydroxide stalks produced by mat‐forming microaerophilic Fe(II)‐oxidizing micro‐organisms. To establish the stalk morphology as a biosignature, morphologic parameters must be quantified and linked to the microaerophilic Fe(II)‐oxidizing metabolism and environmental conditions. Toward this end, we studied an extant model organism, the marine stalk‐forming Fe(II)‐oxidizing bacterium, Mariprofundus ferrooxydans PV‐1. We grew cultures in flat glass microslide chambers, with FeS substrate, creating opposing oxygen/Fe(II) concentration gradients. We used solid‐state voltammetric microelectrodes to measure chemical gradients in situ while using light microscopy to image microbial growth, motility, and mineral formation. In low‐oxygen (2.7–28 μm ) zones of redox gradients, the bacteria converge into a narrow (100 μm–1 mm) growth band. As cells oxidize Fe(II), they deposit Fe(III)‐oxyhydroxide stalks in this band; the stalks orient directionally, elongating toward higher oxygen concentrations. M. ferrooxydans stalks display a narrow range of widths and uniquely biogenic branching patterns, which result from cell division. Together with filament composition, these features (width, branching, and directional orientation) form a physical record unique to microaerophilic Fe(II)‐oxidizer physiology; therefore, stalk morphology is a biosignature, as well as an indicator of local oxygen concentration at the time of formation. Observations of filamentous Fe(III)‐oxyhydroxide microfossils from a ~170 Ma marine Fe‐Si hydrothermal deposit show that these morphological characteristics can be preserved in the microfossil record. This study demonstrates the potential of morphological biosignatures to reveal microbiology and environmental chemistry associated with geologic iron formation depositional processes.     

Transcriptional analysis of Shewanella oneidensis MR-1 with an electrode compared to Fe(III)citrate or oxygen as terminal electron acceptor

Shewanella oneidensis is a target of extensive research in the fields of bioelectrochemical systems and bioremediation because of its versatile metabolic capabilities, especially with regard to respiration with extracellular electron acceptors. The physiological activity of S. oneidensis to respire at electrodes is of great interest, but the growth conditions in thin-layer biofilms make physiological analyses experimentally challenging. Here, we took a global approach to evaluate physiological activity with an electrode as terminal electron acceptor for the generation of electric current. We performed expression analysis with DNA microarrays to compare the overall gene expression with an electrode to that with soluble iron(III) or oxygen as the electron acceptor and applied new hierarchical model-based statistics for the differential expression analysis. We confirmed the differential expression of many genes that have previously been reported to be involved in electrode respiration, such as the entire mtr operon. We also formulate hypotheses on other possible gene involvements in electrode respiration, for example, a role of ScyA in inter-protein electron transfer and a regulatory role of the cbb3-type cytochrome c oxidase under anaerobic conditions. Further, we hypothesize that electrode respiration imposes a significant stress on S. oneidensis, resulting in higher energetic costs for electrode respiration than for soluble iron(III) respiration, which fosters a higher metabolic turnover to cover energy needs. Our hypotheses now require experimental verification, but this expression analysis provides a fundamental platform for further studies into the molecular mechanisms of S. oneidensis electron transfer and the physiologically special situation of growth on a poised-potential surface.     

Comparative analysis of membranous proteomics of Shewanella decolorationis S12 grown with azo compound or Fe (III) citrate as sole terminal electron acceptor

Shewanella decolorationis S12 is capable of carrying out anaerobic respiration using azo dyes and Fe (III) citrate as electron acceptors. In the present study, proteomic techniques including two-dimensional gel electrophoresis (2-DE) and matrix-assisted laser desorption/ionization time-of-flight/time-of-flight mass spectrometry were used to analyze the similarity and the dissimilarity of the membrane proteins isolated from strain S12 cells grown in amaranth or Fe (III) citrate with defined inorganic salt medium. The cells of strain S12 grown under a saturated dissolved oxygen condition served as controls. This is the first work that made the comparative analysis of cell membranous proteomics of strain S12 grown with azo compound or Fe (III) citrate as a sole terminal electron acceptor. The results showed that most of the membrane proteins of strain S12 under azo respiration are similar to those under Fe (III) respiration, but dissimilar from those of oxygen-grown cells. FdnH and FrdB were expressed specifically in azo respiration. NqrA-2, DctP, and hypothetical protein SO_4719 showed relative overexpression in azo respiration compared with Fe (III) respiration. OmpA family protein SO_3545 was detected to be specific to Fe (III) respiration. Furthermore, ArgF, SdhA, and HoxK were expressed markedly in both amaranth- and Fe (III) citrate-grown cultures compared with oxygen-grown cultures.     

Importance of clay size minerals for Fe(III) respiration in a petroleum-contaminated aquifer

The availability of Fe(III)‐bearing minerals for dissimilatory Fe(III) reduction was evaluated in sediments from a petroleum‐contaminated sandy aquifer near Bemidji, Minnesota (USA). First, the sediments from a contaminated area of the aquifer, in which Fe(III) reduction was the predominant terminal electron accepting process, were compared with sediments from a nearby, uncontaminated site. Data from 0.5 m HCl extraction of different size fractions of the sediments revealed that the clay size fraction contributed a significant portion of the ‘bio‐available’ Fe(III) in the background sediment and was the most depleted in ‘bio‐available’ Fe(III) in the iron‐reducing sediment. Analytical transmission electron microscopy (TEM) revealed the disappearance of thermodynamically unstable Fe(III) and Mn(IV) hydroxides (ferrihydrite and Fe vernadite), as well as a decrease in the abundance of goethite and lepidocrocite in the clay size fraction from the contaminated sediment. TEM observations and X‐ray diffraction examination did not provide strong evidence of Fe(III)‐reduction‐related changes within another potential source of ‘bio‐available’ Fe(III) in the clay size fraction – ferruginous phyllosilicates. However, further testing in the laboratory with sediments from the methanogenic portion of the aquifer that were depleted in microbially reducible Fe(III) revealed the potential for microbial reduction of Fe(III) associated with phyllosilicates. Addition of a clay size fraction from the uncontaminated sediment, as well as Fe(III)‐coated kaolin and ferruginous nontronite SWa‐1, as sources of poorly crystalline Fe(III) hydroxides and structural iron of phyllosilicates respectively, lowered steady‐state hydrogen concentrations consistent with a stimulation of Fe(III) reduction in laboratory incubations of methanogenic sediments. There was no change in hydrogen concentration when non‐ferruginous clays or no minerals were added. This demonstrated that Fe(III)‐bearing clay size minerals were essential for microbial Fe(III) reduction and suggested that both potential sources of ‘bio‐available’ Fe(III) in the clay size fraction, poorly crystalline Fe(III) hydroxides and structural Fe(III) of phyllosilicates, were important sources of electron acceptor for indigenous iron‐reducing microorganisms in this aquifer.     

Response to oxygen limitation in bacteria of the genus sulfobacillus

For cultures of moderately thermophilic chemolithotrophic bacteria Sulfobacillus sibiricus N1 and SSO, S. thermosulfidooxidans subsp. asporogenes 41, and the thermotolerant strain S. thermotolerans Kr1 grown under forced aeration and in a high medium layer without aeration, growth characteristics, substrate consumption, and exometabolite formation were compared. Sulfobacilli grown under oxygen limitation exhibited greater generation time, longer growth period, cell yield decreased by from 40 to 85% (depending on the strain), suppressed cell respiration ( demonstrated for S. sibiricus N1 ), accumulation of exometabolites (acetate and propionate) in the medium, and emergence of resting forms. For strains N1, SSO, and Kr1, oscillations of Fe(II) and Fe(III) content in the medium were revealed. For S. sibiricus N1 and S. thermotolerans Kr1, grown under hypoxia (0.07% O₂ in the gas phase), coupling of substrate oxidation with Fe(III) reduction was revealed, as well as utilization of Fe(III) as an electron acceptor alternative to oxygen. The role of labile energy and constructive metabolism for survival of sulfobacilli under diverse conditions is discussed.     

Repeated anaerobic microbial redox cycling of iron

Some nitrate- and Fe(III)-reducing microorganisms are capable of oxidizing Fe(II) with nitrate as the electron acceptor. This enzymatic pathway may facilitate the development of anaerobic microbial communities that take advantage of the energy available during Fe-N redox oscillations. We examined this phenomenon in synthetic Fe(III) oxide (nanocrystalline goethite) suspensions inoculated with microflora from freshwater river floodplain sediments. Nitrate and acetate were added at alternate intervals in order to induce repeated cycles of microbial Fe(III) reduction and nitrate-dependent Fe(II) oxidation. Addition of nitrate to reduced, acetate-depleted suspensions resulted in rapid Fe(II) oxidation and accumulation of ammonium. High-resolution transmission electron microscopic analysis of material from Fe redox cycling reactors showed amorphous coatings on the goethite nanocrystals that were not observed in reactors operated under strictly nitrate- or Fe(III)-reducing conditions. Microbial communities associated with N and Fe redox metabolism were assessed using a combination of most-probable-number enumerations and 16S rRNA gene analysis. The nitrate-reducing and Fe(III)-reducing cultures were dominated by denitrifying Betaproteobacteria (e.g., Dechloromonas) and Fe(III)-reducing Deltaproteobacteria (Geobacter), respectively; these same taxa were dominant in the Fe cycling cultures. The combined chemical and microbiological data suggest that both Geobacter and various Betaproteobacteria participated in nitrate-dependent Fe(II) oxidation in the cycling cultures. Microbially driven Fe-N redox cycling may have important consequences for both the fate of N and the abundance and reactivity of Fe(III) oxides in sediments.     

Toxic effects of chromium(VI) on anaerobic and aerobic growth of Shewanella oneidensis MR-1

Cr(VI) was added to early- and mid-log-phase Shewanella oneidensis (S. oneidensis) MR-1 cultures to study the physiological state-dependent toxicity of Cr(VI). Cr(VI) reduction and culture growth were measured during and after Cr(VI) reduction. Inhibition of growth was observed when Cr(VI) was added to cultures of MR-1 growing aerobically or anaerobically with fumarate as the terminal electron acceptor. Under anaerobic conditions, there was immediate cessation of growth upon addition of Cr(VI) in early- and mid-log-phase cultures. However, once Cr(VI) was reduced below detection limits (0.002 mM), the cultures resumed growth with normal cell yield values observed. In contrast to anaerobic MR-1 cultures, addition of Cr(VI) to aerobically growing cultures resulted in a gradual decrease of the growth rate. In addition, under aerobic conditions, lower cell yields were also observed with Cr(VI)-treated cultures when compared to cultures that were not exposed to Cr(VI). Differences in response to Cr(VI) between aerobically and anaerobically growing cultures indicate that Cr(VI) toxicity in MR-1 is dependent on the physiological growth condition of the culture. Cr(VI) reduction has been previously studied in Shewanella spp., and it has been proposed that Shewanella spp. may be used in Cr(VI) bioremediation systems. Studies of Shewanella spp. provide valuable information on the microbial physiology of dissimilatory metal reducing bacteria; however, our study indicates that S. oneidensis MR-1 is highly susceptible to growth inhibition by Cr(VI) toxicity, even at low concentrations [0.015 mM Cr(VI)].