Anthraquinone-2,6-disulfonate Research Articles

Effects of quinoid redox mediators on the activity of anammox biomass

This study first explored the relationship between the activity of anammox biomass/key enzymes and quinoid redox mediators, which were anthraquinone-2,6-disulfonate (AQDS), 2-hydroxy-1,4-napthoquinone (LAW) and anthraquinone-2-carboxylic acid (AQC). Experimental results demonstrated that the total nitrogen removal performance showed a downward trend with all three redox mediators (RMs) dosage increasing. For instance, when the AQC addition increased to 0.8 mM, the TN removal rate sharply reduced to 17.2 mg-N/gVSS/h, only about 20% of the control. This phenomenon might be caused by microbial poisoning with the extracellular RMs additions. Nevertheless, the crude hydrazine dehydrogenase, nitrite reductase, and nitrate reductase activities were enhanced with RMs addition, about 0.6-3 folds compared to the control experiments without RMs addition. The RMs was inferred to play the role as ubiquinol/ubiquinone (Q/QH2) during the anammox process. Furthermore, the specific ladderane membrane structure could block the contacting between RMs and the key enzymes inside anammoxosome. This might be the main reason for the contrary effects of RMs on anammox biomass and the key enzymes.

Effects of redox mediators on azo dye decolorization by Shewanella algae under saline conditions

Azo dye decolorization by Shewanella algae (SAL) in the presence of high concentrations of NaCl and different quinones or humic acids was investigated to reveal the effects of redox mediator under saline conditions. Growth of SAL and the other two marine Shewanella strains coupled to anthraquinone-2,6-disulfonate (AQDS) reduction was observed in a wide range of NaCl concentrations (0-7%). AQDS showed the best enhancing effects, whereas some other quinones demonstrated poorer stimulating or even inhibiting effects on acid red 27 (AR27) decolorization. Different humic acids could also enhance the decolorization. The correlation between specific AQDS-mediated reduction rate and initial AR27 concentration could be described with Michaelis-Menten kinetics (Km=0.2 mM and Vmax=9.3 μmol mg cell(-1) h(-1)). AQDS reduction by SAL was determined to be the rate-limiting step of mediated reduction. Mediated decolorization products of AR27 were determined to be less phytotoxic aromatic amines.

Effective and characteristics of anthraquinone-2,6-disulfonate (AQDS) on denitrification by Paracoccus versutus sp.GW1

The effects of anthraquinone-2,6-disulfonate (AQDS) on microbial populations and the biocatalysing characteristics of AQDS on denitrifying strain GW1 are discussed. The results showed that microbial population abundances were 30, 6, 12 and 18% during the different periods (1st, 5th, 10th and 20th days). Strain GW1 utilized AQDS as an electron acceptor in the respiration chain and reduced AQDS to hydroquinone (QH2). The nitrate removal efficiency of strain GW1 was increased 1.14–1.63 fold with 0.04–0.32 mmol/L AQDS. A linear correlation was found between the nitrate removal reaction constant (where VSS is volatile suspended solids) and AQDS concentration (CAQDS), which was k=19.332 CAQDS+11.115(R2=0.9749). The stabilized oxidation–reduction potential (ORP) values with AQDS were lower 22–92 mv than the control during the denitrification process. The concentration of nitrite sequentially accumulated up to around 250 mg/L when nitrate have removed completely. The results suggested that AQDS as redox mediator was capable of biocatalysing the nitrate and nitrite removal rate.

Extracellular Quinones Affecting Methane Production and Methanogenic Community in Paddy Soil

This study investigated the change of CH4 production and methanogenic community in response to the presence of humic substances (humics) analogue, anthraquinone-2,6-disulfonate (AQDS). Anaerobic experiments used a Chinese paddy soil, and three concentration levels of 0.5, 5, and 20mM AQDS were conducted. Results suggested that the effect of AQDS on methanogenesis was time-dependent and concentration-dependent. Twenty millimolars of AQDS was toxic for methanogenic activity almost for the entire experimental period. Slight inhibition of methanogenesis by AQDS respiration in the 0.5- and 5-mM AQDS-supplemented treatments occurred within the early period, while CH4 accumulated throughout the later period was approximately five and ten times greater than that of the controls without AQDS, respectively. AQDS reduction coupling to acetate oxidization enriched Geobacter species, and the mcrA-targeted T-RFLP profiles revealed significant increase of Methanosarcina at the expense of Methanobacterium in the 0.5- and 5-mM AQDS treatments. The enriched syntrophic association between Geobacter and Methanosarcina was deduced to be an effective methanogenic pathway for converting acetate to CH4 via direct interspecies electron transfer. This study implied the ecological importance of syntrophic interaction between methanogens and microorganisms enriched by anaerobic respiration of non-methanogenic terminal electron acceptors in paddy soils.

Natranaerobaculum magadiense gen. nov., sp. nov., an anaerobic, alkalithermophilic bacterium from soda lake sediment

An obligately alkaliphilic, anaerobic, thermo- and halotolerant, spore-forming bacterium was isolated from sediments of soda lake Magadi (Kenya) and designated strain Z-1001(T). Cells of strain Z-1001(T) were straight, Gram-positive rods, slowly motile. Strain Z-1001(T) was found to be an obligate anaerobe. It grew within a pH range from 7.5 to 10.7 with an optimum at 9.25-9.5 (at 40 °C), a temperature range from 20 to 57 °C with an optimum at 45-50 °C, and a NaCl concentration range from 0 to 1.55 M with an optimum at 1.2-1.4 M. Peptides, such as meat and yeast extracts, peptone and tryptone, were fermented by Z-1001(T). Carbohydrates did not support growth. With yeast extract as an electron donor, strain Z-1001(T) reduced S(2)O(3)(2-), NO(-)(3), AsO(3-)(4), Fe(III) citrate and anthraquinone-2,6-disulfonate (AQDS) as electron acceptors. The isolate was able to grow oligotrophically with a very small amount of yeast extract: 0.03 g l(-1). The main fatty acids were C16 : 0, C16 : 1ω7c, C18 : 0 and C18 : 1ω9. The DNA G+C content of the isolate was 35.6 mol%. 16S rRNA gene sequence analysis showed that strain Z-1001(T) is a member of family Natranaerobiaceae, clustering with the type strain of Natranaerobius thermophilus (95.8-96.0 % sequence similarity). On the basis of physiological and phylogenetic data it is proposed that strain Z-1001(T) ( = DSM 24923(T) = VKM B-2666(T)) represents a novel genus and species, Natranaerobaculum magadiense gen. nov., sp. nov.

Extracellular Electron Transfer to Fe(III) Oxides by the Hyperthermophilic Archaeon Geoglobus ahangari via a Direct Contact Mechanism

The microbial reduction of Fe(III) plays an important role in the geochemistry of hydrothermal systems, yet it is poorly understood at the mechanistic level. Here we show that the obligate Fe(III)-reducing archaeon Geoglobus ahangari uses a direct-contact mechanism for the reduction of Fe(III) oxides to magnetite at 85°C. Alleviating the need to directly contact the mineral with the addition of a chelator or the electron shuttle anthraquinone-2,6-disulfonate (AQDS) stimulated Fe(III) reduction. In contrast, entrapment of the oxides within alginate beads to prevent cell contact with the electron acceptor prevented Fe(III) reduction and cell growth unless AQDS was provided. Furthermore, filtered culture supernatant fluids had no effect on Fe(III) reduction, ruling out the secretion of an endogenous mediator too large to permeate the alginate beads. Consistent with a direct contact mechanism, electron micrographs showed cells in intimate association with the Fe(III) mineral particles, which once dissolved revealed abundant curled appendages. The cells also produced several heme-containing proteins. Some of them were detected among proteins sheared from the cell's outer surface and were required for the reduction of insoluble Fe(III) oxides but not for the reduction of the soluble electron acceptor Fe(III) citrate. The results thus support a mechanism in which the cells directly attach and transfer electrons to the Fe(III) oxides using redox-active proteins exposed on the cell surface. This strategy confers on G. ahangari a competitive advantage for accessing and reducing Fe(III) oxides under the extreme physical and chemical conditions of hot ecosystems.

Accelerated removal of Sudan dye by Shewanella oneidensis MR-1 in the presence of quinones and humic acids

Although there have been many studies on bacterial removal of soluble azo dyes, much less information is available for biological treatment of water-insoluble azo dyes. The few bacterial species capable of removing Sudan dye generally require a long time to remove low concentrations of insoluble dye particles. The present work examined the efficient removal of Sudan I by Shewanella oneidensis MR-1 in the presence of redox mediator. It was found that the microbially reduced anthraquinone-2,6-disulfonate (AQDS) could abiotically reduce Sudan I, indicating the feasibility of microbially-mediated reduction. The addition of 100 μM AQDS and other different quinone compounds led to 4.3-54.7 % increase in removal efficiencies in 22 h. However, adding 5-hydroxy-1,4-naphthoquinone into the system inhibited Sudan I removal. The presence of 10, 50 and 100 μM AQDS stimulated the removal efficiency in 10 h from 26.4 to 42.8, 54.9 and 64.0 %, respectively. The presence of 300 μM AQDS resulted in an eightfold increase in initial removal rate from 0.19 to 1.52 mg h⁻¹ g⁻¹ cell biomass. A linear relationship was observed between the initial removal rates and AQDS concentrations (0-100 μM). Comparison of Michaelis-Menten kinetic constants revealed the advantage of AQDS-mediated removal over direct reduction. Different species of humic acid could also stimulate the removal of Sudan I. Scanning electronic microscopy analysis confirmed the accelerated removal performance in the presence of AQDS. These results provide a potential method for the efficient removal of insoluble Sudan dye.

Azo dye decolorization assisted by chemical and biogenic sulfide

The effectiveness of chemical and biogenic sulfide in decolorizing three sulfonated azo dyes and the robustness of a sulfate-reducing process for simultaneous decolorization and sulfate removal were evaluated. The results demonstrated that decolorization of azo dyes assisted by chemical sulfide and anthraquinone-2,6-disulfonate (AQDS) was effective. In the absence of AQDS, biogenic sulfide was more efficient than chemical sulfide for decolorizing the azo dyes. The performance of sulfate-reducing bacteria in attached-growth sequencing batch reactors suggested the absence of competition between the studied azo dyes and the sulfate-reducing process for the reducing equivalents. Additionally, the presence of chemical reduction by-products had an almost negligible effect on the sulfate removal rate, which was nearly constant (94%) after azo dye injection.

Microbial formation of palladium nanoparticles by Geobacter sulfurreducens for chromate reduction

Geobacter sulfurreducens was studied for the reduction of Pd(II) and production of Pd(0) nanoparticles capable of reducing Cr(VI). Transmission electronic microscopy, energy dispersive X-ray and X-ray diffraction analyses revealed that the nanoscale Pd(0) particles formed were associated with the cell surface and located inside the periplasm. The increase of cell dry weight (CDW):Pd ratio and addition of anthraquinone-2,6-disulfonate (AQDS) not only stimulated Pd(II) reduction, but also resulted in increase of nanoparticle number, decrease of particle diameter and improvement of Cr(VI) reduction efficiency. The relationship between reduction rate and initial Cr(VI) concentration (150–750μM) followed Michaelis–Menten kinetics (Vmax=3.6μmolh−1mg bio-Pd−1 and Km=891.3μM). These findings indicated the potential of using G. sulfurreducens cells for reclamation of palladium, formation of Pd(0) nanoparticles and efficient treatment of Cr(VI) pollution.

Microbial reduction of Fe(III) in smectite minerals by thermophilic methanogen Methanothermobacter thermautotrophicus

Clay minerals and thermophilic methanogens can co-exist in hot anoxic environments, including the continental subsurface, geysers, terrestrial hot springs, and deep-sea hydrothermal vent systems. However, it is unclear whether thermophilic methanogens are able to reduce structural Fe(III) in clay minerals. In this study, the ability of a thermophilic methanogen Methanothermobacter thermautotrophicus to reduce structural Fe(III) in iron-rich and iron-poor smectites, (nontronite NAu-2 and Wyoming montmorillonite SWy-2) and the relationship between iron reduction and methanogenesis were investigated. M. thermautotrophicus reduced Fe(III) in nontronite NAu-2 and montmorillonite SWy-2 with H2/CO2 as substrate. The extent of bioreduction was 27% for nontronite and 13–15% for montmorillonite. Anthraquinone-2,6-disulfonate (AQDS) did not enhance the extent of bioreduction, but accelerated the rate. When methanogenesis was inhibited via addition of 2-bromoethane sulfonate (BES), the extent of bioreduction decreased to 16% for NAu-2 and 9% for SWy-2. These data suggest that Fe(III) bioreduction and methanogenesis were mutually beneficial. The likely mechanism was that Fe(III) bioreduction lowered the reduction potential of the system so that methanogenesis became favorable, and methanogenesis in turn stimulated the growth of the methanogen, which enhanced Fe(III) bioreduction. NAu-2 was partly dissolved and high charge smectite and biogenic silica formed as a result of bioreduction.

Reduction of palladium and production of nano-catalyst by Geobacter sulfurreducens

The present study is the first report on the ability of Geobacter sulfurreducens PCA to reduce Pd(II) and produce Pd(0) nano-catalyst, using acetate as electron donor at neutral pH (7.0 ± 0.1) and 30 °C. The microbial production of Pd(0) nanoparticles (NPs) was greatly enhanced by the presence of the redox mediator, anthraquinone-2,6-disulfonate (AQDS) when compared with controls lacking AQDS and cell-free controls. A cell dry weight (CDW) concentration of 800 mg/L provided a larger surface area for Pd(0) NPs deposition than a CDW concentration of 400 mg/L. Sample analysis by transmission electron microscopy revealed the formation of extracellular Pd(0) NPs ranging from 5 to 15 nm and X-ray diffraction confirmed the Pd(0) nature of the nano-catalyst produced. The present findings open the possibility for a new alternative to synthesize Pd(0) nano-catalyst and the potential application for microbial metal recovery from metal-containing waste streams.

Hexavalent chromium reduction by Cellulomonas sp. strain ES6: the influence of carbon source, iron minerals, and electron shuttling compounds

The reduction of hexavalent chromium, Cr(VI), to trivalent chromium, Cr(III), can be an important aspect of remediation processes at contaminated sites. Cellulomonas species are found at several Cr(VI) contaminated and uncontaminated locations at the Department of Energy site in Hanford, Washington. Members of this genus have demonstrated the ability to effectively reduce Cr(VI) to Cr(III) fermentatively and therefore play a potential role in Cr(VI) remediation at this site. Batch studies were conducted with Cellulomonas sp. strain ES6 to assess the influence of various carbon sources, iron minerals, and electron shuttling compounds on Cr(VI) reduction rates as these chemical species are likely to be present in, or added to, the environment during in situ bioremediation. Results indicated that the type of carbon source as well as the type of electron shuttle present influenced Cr(VI) reduction rates. Molasses stimulated Cr(VI) reduction more effectively than pure sucrose, presumably due to presence of more easily utilizable sugars, electron shuttling compounds or compounds with direct Cr(VI) reduction capabilities. Cr(VI) reduction rates increased with increasing concentration of anthraquinone-2,6-disulfonate (AQDS) regardless of the carbon source. The presence of iron minerals and their concentrations did not significantly influence Cr(VI) reduction rates. However, strain ES6 or AQDS could directly reduce surface-associated Fe(III) to Fe(II), which was capable of reducing Cr(VI) at a near instantaneous rate. These results suggest the rate limiting step in these systems was the transfer of electrons from strain ES6 to the intermediate or terminal electron acceptor whether that was Cr(VI), Fe(III), or AQDS.

Redox Reactions of Reduced Flavin Mononucleotide (FMN), Riboflavin (RBF), and Anthraquinone-2,6-disulfonate (AQDS) with Ferrihydrite and Lepidocrocite

Flavins are secreted by the dissimilatory iron-reducing bacterium Shewanella and can function as endogenous electron transfer mediators. To assess the potential importance of flavins in Fe(III) bioreduction, we investigated the redox reaction kinetics of reduced flavin mononucleotide (i.e., FMNH(2)) and reduced riboflavin (i.e., RBFH(2)) with ferrihydrite and lepidocrocite. The organic reductants rapidly reduced and dissolved ferrihydrite and lepidocrocite in the pH range 4-8. The rate constant k for 2-line ferrihydrite reductive dissolution by FMNH(2) was 87.5 ± 3.5 M(-1)·s(-1) at pH 7.0 in batch reactors, and k was similar for RBFH(2). For lepidocrocite, k was 500 ± 61 M(-1)·s(-1) for FMNH(2) and 236 ± 22 M(-1)·s(-1) for RBFH(2). The surface area normalized initial reaction rates (r(a)) were between 0.08 and 77 μmol·m(-2)·s(-1) for various conditions in stopped-flow experiments. Initial rates (r(o)) were first-order with respect to iron(III) oxide concentration, and r(a) increased with decreasing pH. Poorly crystalline 2-line ferrihydrite yielded the highest r(a), followed by more crystalline 6-line ferrihydrite and crystalline lepidocrocite. Compared to a previous whole-cell study with Shewanella oneidensis strain MR-1, our findings suggest that the reduction of electron transfer mediators by the Mtr (i.e., metal-reducing) pathway coupled to lactate oxidation is rate limiting, rather than heterogeneous electron transfer to the iron(III) oxide.

Anaerobic Benzene Oxidation by Geobacter Species

The abundance of Geobacter species in contaminated aquifers in which benzene is anaerobically degraded has led to the suggestion that some Geobacter species might be capable of anaerobic benzene degradation, but this has never been documented. A strain of Geobacter, designated strain Ben, was isolated from sediments from the Fe(III)-reducing zone of a petroleum-contaminated aquifer in which there was significant capacity for anaerobic benzene oxidation. Strain Ben grew in a medium with benzene as the sole electron donor and Fe(III) oxide as the sole electron acceptor. Furthermore, additional evaluation of Geobacter metallireducens demonstrated that it could also grow in benzene-Fe(III) medium. In both strain Ben and G. metallireducens the stoichiometry of benzene metabolism and Fe(III) reduction was consistent with the oxidation of benzene to carbon dioxide with Fe(III) serving as the sole electron acceptor. With benzene as the electron donor, and Fe(III) oxide (strain Ben) or Fe(III) citrate (G. metallireducens) as the electron acceptor, the cell yields of strain Ben and G. metallireducens were 3.2 × 10(9) and 8.4 × 10(9) cells/mmol of Fe(III) reduced, respectively. Strain Ben also oxidized benzene with anthraquinone-2,6-disulfonate (AQDS) as the sole electron acceptor with cell yields of 5.9 × 10(9) cells/mmol of AQDS reduced. Strain Ben serves as model organism for the study of anaerobic benzene metabolism in petroleum-contaminated aquifers, and G. metallireducens is the first anaerobic benzene-degrading organism that can be genetically manipulated.

Fe(III) Reduction and U(VI) Immobilization by Paenibacillus sp. Strain 300A, Isolated from Hanford 300A Subsurface Sediments

A facultative iron-reducing [Fe(III)-reducing] Paenibacillus sp. strain was isolated from Hanford 300A subsurface sediment biofilms that was capable of reducing soluble Fe(III) complexes [Fe(III)-nitrilotriacetic acid and Fe(III)-citrate] but unable to reduce poorly crystalline ferrihydrite (Fh). However, Paenibacillus sp. 300A was capable of reducing Fh in the presence of low concentrations (2 μM) of either of the electron transfer mediators (ETMs) flavin mononucleotide (FMN) or anthraquinone-2,6-disulfonate (AQDS). Maximum initial Fh reduction rates were observed at catalytic concentrations (<10 μM) of either FMN or AQDS. Higher FMN concentrations inhibited Fh reduction, while increased AQDS concentrations did not. We also found that Paenibacillus sp. 300A could reduce Fh in the presence of natural ETMs from Hanford 300A subsurface sediments. In the absence of ETMs, Paenibacillus sp. 300A was capable of immobilizing U(VI) through both reduction and adsorption. The relative contributions of adsorption and microbial reduction to U(VI) removal from the aqueous phase were ∼7:3 in PIPES [piperazine-N,N'-bis(2-ethanesulfonic acid)] and ∼1:4 in bicarbonate buffer. Our study demonstrated that Paenibacillus sp. 300A catalyzes Fe(III) reduction and U(VI) immobilization and that these reactions benefit from externally added or naturally existing ETMs in 300A subsurface sediments.

Reduction of Cr(VI) by Escherichia coli BL21 in the presence of redox mediators

Anthraquinone-2,7-disulfonate (2,7-AQDS), anthraquinone-1-sulfonate (α-AQS), anthraquinone-2-sulfonate (AQS), anthraquinone-2,6-disulfonate (AQDS) and anthraquinone-1,5-disulfonate (1,5-AQDS) were selected as redox mediators for Cr(VI) reduction by Escherichia coli BL21. In the presence of 0.8mmoll−1 AQS, the Cr(VI) reduction efficiency was 98.5% in 7.5h, whereas it was only 21–34% in the absence of a mediator or in the presence of α-AQS, 1,5-AQDS, AQDS, 2,7-AQDS. A linear correlation, k=0.768CAQS+1.5531 (R2=0.9935), was found for the reaction constant, k (mg Cr(VI) g−1 dry cell weight h−1) and the AQS concentration (CAQS). The Arrhenius equation described the Cr(VI) reduction in the tested temperature range, and the pre-exponential factor A was 13.249mg Cr(VI) g−1 dry cell weight h−1,and the activation energy Ea was 28.01kJmol−1. Glucose was the best carbon sources, and the optimum pH was 6.0. The Cr(VI) reduction rate obtained with AQS is a significant improvement over low conventional anaerobic reduction rates.

Kinetics during the redox biotransformation of pollutants mediated by immobilized and soluble humic acids

The aim of this study was to elucidate the kinetic constraints during the redox biotransformation of the azo dye, Reactive Red 2 (RR2), and carbon tetrachloride (CT) mediated by soluble humic acids (HAs) and immobilized humic acids (HAi), as well as by the quinoid model compounds, anthraquinone-2,6-disulfonate (AQDS) and 1,2-naphthoquinone-4-sulfonate (NQS). The microbial reduction of both HAs and HAi by anaerobic granular sludge (AGS) was the rate-limiting step during decolorization of RR2 since the reduction of RR2 by reduced HAi proceeded at more than three orders of magnitute faster than the electron-transferring rate observed during the microbial reduction of HAi by AGS. Similarly, the reduction of RR2 by reduced AQDS proceeded 1.6- and 1.9-fold faster than the microbial reduction of AQDS by AGS when this redox mediator (RM) was supplied in soluble and immobilized form, respectively. In contrast, the reduction of NQS by AGS occurred 1.6- and 19.2-fold faster than the chemical reduction of RR2 by reduced NQS when this RM was supplied in soluble and immobilized form, respectively. The microbial reduction of HAs and HAi by a humus-reducing consortium proceeded 1,400- and 790-fold faster than the transfer of electrons from reduced HAs and HAi, respectively, to achieve the reductive dechlorination of CT to chloroform. Overall, the present study provides elucidation on the rate-limiting steps involved in the redox biotransformation of priority pollutants mediated by both HAs and HAi and offers technical suggestions to overcome the kinetic restrictions identified in the redox reactions evaluated.

Biostimulation of Indigenous Microbial Communities for Anaerobic Transformation of Pentachlorophenol in Paddy Soils of Southern China

This study explored biostimulation mechanisms with an electron donor and a shuttle for accelerating pentachlorophenol (PCP) transformation in iron-rich soils. The results indicated that indigenous microbial communities are important for PCP transformation in soils. Biostimulation of indigenous microbial communities by the addition of lactate and anthraquinone-2,6-disulfonate (AQDS) led to the enhanced rates of PCP dechlorination by the dechlorinating- and iron-reducing bacteria in soils. The electrochemical studies using cyclic voltammograms and microbial current measurements confirmed the high reduction potential and the large amount of electrons generated under biostimulation conditions, which were responsible for the higher rates of PCP transformation. After biostimulation treatments by the additions of lactate and/or AQDS during PCP dechlorination processes, microbial community analysis by the terminal restriction fragment length polymorphism (T-RFLP) method showed the abundance terminal restricted fragments (T-RFs), an indicator of bacterial abundance, which represents the dechlorinating- and iron-reducing bacteria, suggesting their critical roles in PCP dechlorination in soils.

Biotic and Abiotic Reduction and Solubilization of Pu(IV)O2•xH2O(am) as Affected by Anthraquinone-2,6-disulfonate (AQDS) and Ethylenediaminetetraacetate (EDTA)

This study measured reductive solubilization of plutonium(IV) hydrous oxide (Pu(IV)O(2)·xH(2)O((am))) with hydrogen (H(2)) as electron donor, in the presence or absence of dissimilatory metal-reducing bacteria (DMRB), anthraquinone-2,6-disulfonate (AQDS), and ethylenediaminetetraacetate (EDTA). In PIPES buffer at pH 7 with excess H(2), Shewanella oneidensis and Geobacter sulfurreducens both solubilized <0.001% of 0.5 mM Pu(IV)O(2)·xH(2)O((am)) over 8 days, with or without AQDS. However, Pu((aq)) increased by an order of magnitude in some treatments, and increases in solubility were associated with production of Pu(III)((aq)). The solid phase of these treatments contained Pu(III)(OH)(3(am)), with more in the DMRB treatments compared with abiotic controls. In the presence of EDTA and AQDS, PuO(2)·xH(2)O((am)) was completely solubilized by S. oneidensis and G. sulfurreducens in ∼24 h. Without AQDS, bioreductive solubilization was slower (∼22 days) and less extensive (∼83-94%). In the absence of DMRB, EDTA facilitated reductive solubilization of 89% (without AQDS) to 98% (with AQDS) of the added PuO(2)·xH(2)O((am)) over 418 days. An in vitro assay demonstrated electron transfer to PuO(2)·xH(2)O((am)) from the S. oneidensis outer-membrane c-type cytochrome MtrC. Our results (1) suggest that PuO(2)·xH(2)O((am)) reductive solubilization may be important in reducing environments, especially in the presence of complexing ligands and electron shuttles, (2) highlight the environmental importance of polynuclear, colloidal Pu, (3) provide additional evidence that Pu(III)-EDTA is a more likely mobile form of Pu than Pu(IV)-EDTA, and (4) provide another example of outer-membrane cytochromes and electron-shuttling compounds facilitating bioreduction of insoluble electron acceptors in geologic environments.

Surface multiheme c -type cytochromes from Thermincola potens and implications for respiratory metal reduction by Gram-positive bacteria

Almost nothing is known about the mechanisms of dissimilatory metal reduction by Gram-positive bacteria, although they may be the dominant species in some environments. Thermincola potens strain JR was isolated from the anode of a microbial fuel cell inoculated with anaerobic digester sludge and operated at 55 °C. Preliminary characterization revealed that T. potens coupled acetate oxidation to the reduction of hydrous ferric oxides (HFO) or anthraquinone-2,6-disulfonate (AQDS), an analog of the redox active components of humic substances. The genome of T. potens was recently sequenced, and the abundance of multiheme c-type cytochromes (MHCs) is unusual for a Gram-positive bacterium. We present evidence from trypsin-shaving LC-MS/MS experiments and surface-enhanced Raman spectroscopy (SERS) that indicates the expression of a number of MHCs during T. potens growth on either HFO or AQDS, and that several MHCs are localized to the cell wall or cell surface. Furthermore, one of the MHCs can be extracted from cells with low pH or denaturants, suggesting a loose association with the cell wall or cell surface. Electron microscopy does not reveal an S-layer, and the precipitation of silver metal on the cell surface is inhibited by cyanide, supporting the involvement of surface-localized redox-active heme proteins in dissimilatory metal reduction. These results provide unique direct evidence for cell wall-associated cytochromes and support MHC involvement in conducting electrons across the cell envelope of a Gram-positive bacterium.