Sulfurospirillum Barnesii Research Articles

Microbes-Assisted Arsenate Reduction Activity in Bangladesh Drinking Water Wells as Revealed by Enrichment Cultivation

Arsenate-reducing microorganisms may mobilize arsenic into groundwater. Phylogenetic DNA sequence information cannot exclusively be used to infer arsenate reduction, as the spotted homologies may not translate to biochemical activities. We here develop a cultivation-based metagenomic strategy for inferring actual activities: we complement a cultivation-independent microbial community survey covering 22 arsenic-contaminated drinking water wells in Bangladesh, with the characterization of enrichments of anaerobic arsenate reducers. All investigated samples revealed microbial arsenate reduction activity. The enriched microbial communities were phylogenetically diverse within and between the enrichments. The enrichment also unveiled species: the arsenate-reducing enrichments held 16S rRNA gene sequences closest to Dechloromonas sp., Azonexus fungiphilus, Youngiibacter fragile, Methanogenic prokaryote and Fusibacter sp., in addition to arsenate-reducing Sulfurospirillum strains NP4, b10 and dissimilatory iron-reducing Geobacter species. The enriched arsenate reductase gene (arrA) sequences were closest to sequences in known arsenate reducing Sulfurospirillum barnesii. This study bridges the knowledge gap between cultivation-independent and cultivation-dependent analyses. It clarifies the mechanism for arsenic mobilization and redistribution in rural drinking water wells, as a balance between bioremediation and harmful activities. The identified, potentially dissimilatory arsenate and iron-reducing microbes, constitute a risk for arsenic bioremediation. Appropriate metagenomic and dynamic analyses should help design and maintain safer, effective bioremediation.

Biosynthesis, Synthesis, and Activities of Barnesin A, a NRPS-PKS Hybrid Produced by an Anaerobic Epsilonproteobacterium.

Despite the wealth of physiological knowledge and plentiful genomes available, only few natural products of anaerobic bacteria have been identified until today and even less have been linked to their biosynthetic gene cluster. Here, we analyzed a unique NRPS-PKS hybrid gene cluster from an anaerobic Epsilonproteobacterium ( Sulfurospirillum barnesii). Phylogenetic analysis of key biosynthetic genes, gene expression studies, and comparative metabolomics resulted in the identification of the first anoxically biosynthesized NRPS-PKS hybrid metabolite: a lipo-dipeptide with a vinylogous side chain, called barnesin A. The absolute structure was verified by a modular total synthesis, and barnesin and derivatives were found to have antimicrobial activity, as well as selective and nanomolar inhibitory activity, against pharmacological important cysteine proteases, such as cathepsin B.

Oxygen kinetic isotope effects in selenate during microbial reduction

The redox cycling of selenium oxyanions, elemental selenium, and selenides within water resources has implications for Se bioavailability and ecotoxicity. Dual stable isotope analysis of Se and O may provide important information for interpreting environmental Se transformations. Stable Se isotope systematics within the Se redox cycle has been well characterized within the literature, however concomitant oxygen isotope composition requires additional investigation. This study reports the O isotope fractionation of selenate (SeO42−) during microbial reduction by the dissimilatory Se-reducing bacterium Sulfurospirillum barnesii SES-3. Microbial reduction experiments were conducted under various conditions in order to investigate the range of 18O enrichment factors (εO) in selenate. The reduction of selenate to selenite coupled to the oxidation of lactate to acetate resulted in an 18O kinetic isotope effect with εO values 1.5–5.8‰. Greater 18O enrichment was observed with increasing pH, but no correlation was found between εO and reduction rates, lactate availability, or cell density. εO values from biotic reduction by S. barnesii here are significantly less than those observed for abiotic reduction with Fe(II)-rich minerals reported in the literature, and this difference could be explained by a diffusion limitation during enzymatic reduction. Our results expand the isotope systematics of the selenium redox cycle and suggest εO has potential usefulness as indicators for in situ selenate reduction.

이화형비산염환원균의 특성

Although, microbial arsenic mobilization by dissimilatory arsenate-reducing bacteria (DARB) and the practical use to the removal technology of arsenic from contaminated soil are expected, most previous research mainly has been focused on the geochemical circulation of arsenic. Therefore, in this review we summarized the previously reported DARB to grasp the characteristic for bioremediation of arsenic. Evidence of microbial growth on arsenate is presented based on isolate analyses, after which a summary of the physiology of the following arsenaterespiring bacteria is provided: Chrysiogenes arsenatis strain BAL-1 T , Sulfurospirillum barnesii, Desulfotomaculum strain Ben-RB, Desulfotomaculum auripigmentum strains OREX-4, GFAJ-1, Bacillus sp., Desulfitobacterium hafniense DCB-2 T , strain SES-3, Citrobacter sp. (TSA-1 and NC-1), Sulfurospirillum arsenophilum sp. nov., Shewanella sp., Chrysiogenes arsenatis BAL-l T , Deferribacter desulfuricans. Among the DARB, Citrobacter sp. NC-1 is superior to other dissimilatory arsenate-reducing bacteria with respect to arsenate reduction, particularly at high concentrations as high as 60 mM. A gram-negative anaerobic bacterium, Citrobacter sp. NC-1, which was isolated from arsenic contaminated soil, can grow on glucose as an electron donor and arsenate as an electron acceptor. Strain NC-1 rapidly reduced arsenate at 5 mM to arsenite with concomitant cell growth, indicating that arsenate can act as the terminal electron acceptor for anaerobic respiration (dissimilatory arsenate reduction). To characterize the reductase systems in strain NC-1, arsenate and nitrate reduction activities were investigated with washed-cell suspensions and crude cell extracts from cells grown on arsenate or nitrate. These reductase activities were induced individually by the two electron acceptors. Tungstate, which is a typical inhibitory antagonist of molybdenum containing dissimilatory reductases, strongly inhibited the reduction of arsenate and nitrate in anaerobic growth cultures. These results suggest that strain NC-1 catalyzes the reduction of arsenate and nitrate by distinct terminal reductases containing a molybdenum cofactor. This may be advantageous during bioremediation processes where both contaminants are present. Moreover, a brief explanation of arsenic extraction from a model soil artificially contaminated with As (V) using a novel DARB (Citrobacter sp. NC-1) is given in this article. We conclude with a discussion of the importance of microbial arsenate reduction in the environment. The successful application and use of DARB should facilitate the effective bioremediation of arsenic contaminated sites.

Localization of the Dissimilatory Arsenate Reductase in <I>Sulfurospirillum Barnesi</I> Strain SeS-3

Problem statement: Sulfurospirillim barnesii strain SeS-3 is one of the recently known bacterial isolates that can obtain energy to support growth by respiring the toxic oxyanions of arsenic and selenium under anaerobic conditions. Approach: The ultimate goal of this investigation is to localize the active sites for the reduction of arsenate arsenite as well as selenate selenite in S. barnesii strain SeS-3. Results: The ability of the type strain Sulfurospirillium barnesii strain SES-3 (ATCC 700032) to reduce selenate and arsenate were tested using cell grown anaerobically with 20 mM lactate as the carbon source and either 10 mM selenate or 5 mM arsenate as the terminal electron acceptors were harvested after the turbidity reached 0.3, 0.4 absorption units at 600 nm, cell density 4gm wet cells. The results of this study showed that the mobilization of the toxic oxyanions of arsenic and selenium by Sulfurospirillum barnesii strain SES-3 is linked to the membrane. The enzyme is specific for the reduction of arsenate, selenate, selenite, nitrite, thiosulfate and phosphate. Whereas, no specificity was detected for arsenite nitrate, fumarate, when they served as the final electron acceptor. Conclusion/Recommendations: This study concluded that the mechanism of arsenate reduction by S.barnesii strain SeS-3 is connected into the membrane. The environmental significance of this bacterium and its impact to the bioremediation potential in the underground water and sedimentary environment is also discussed.

Microbial Reduction of Chromate in the Presence of Nitrate by Three Nitrate Respiring Organisms

A major challenge for the bioremediation of toxic metals is the co-occurrence of nitrate, as it can inhibit metal transformation. Geobacter metallireducens, Desulfovibrio desulfuricans, and Sulfurospirillum barnesii are three soil bacteria that can reduce chromate [Cr(VI)] and nitrate, and may be beneficial for developing bioremediation strategies. All three organisms respire through dissimilatory nitrate reduction to ammonia (DNRA), employing different nitrate reductases but similar nitrite reductase (Nrf). G. metallireducens reduces nitrate to nitrite via the membrane bound nitrate reductase (Nar), while S. barnesii and D. desulfuricans strain 27774 have slightly different forms of periplasmic nitrate reductase (Nap). We investigated the effect of DNRA growth in the presence of Cr(VI) in these three organisms and the ability of each to reduce Cr(VI) to Cr(III), and found that each organisms responded differently. Growth of G. metallireducens on nitrate was completely inhibited by Cr(VI). Cultures of D. desulfuricans on nitrate media was initially delayed (48 h) in the presence of Cr(VI), but ultimately reached comparable cell yields to the non-treated control. This prolonged lag phase accompanied the transformation of Cr(VI) to Cr(III). Viable G. metallireducens cells could reduce Cr(VI), whereas Cr(VI) reduction by D. desulfuricans during growth, was mediated by a filterable and heat stable extracellular metabolite. S. barnesii growth on nitrate was not affected by Cr(VI), and Cr(VI) was reduced to Cr(III). However, Cr(VI) reduction activity in S. barnesii, was detected in both the cell free spent medium and cells, indicating both extracellular and cell associated mechanisms. Taken together, these results have demonstrated that Cr(VI) affects DNRA in the three organisms differently, and that each have a unique mechanism for Cr(VI) reduction.

Shedding Light on Selenium Biomineralization: Proteins Associated with Bionanominerals

Selenium-reducing microorganisms produce elemental selenium nanoparticles with particular physicochemical properties due to an associated organic fraction. This study identified high-affinity proteins associated with such bionanominerals and with nonbiogenic elemental selenium. Proteins with an anticipated functional role in selenium reduction, such as a metalloid reductase, were found to be associated with nanoparticles formed by one selenium respirer, Sulfurospirillum barnesii.

Bioaugmentation of UASB reactors with immobilized Sulfurospirillum barnesii for simultaneous selenate and nitrate removal

Whole-cell immobilization of selenate-respiring Sulfurospirillum barnesii in polyacrylamide gels was investigated to allow the treatment of selenate contaminated (790 µg Se × L−1) synthetic wastewater with a high molar excess of nitrate (1,500 times) and sulfate (200 times). Gel-immobilized S. barnesii cells were used to inoculate a mesophilic (30°C) bioreactor fed with lactate as electron donor at an organic loading rate of 5 g chemical oxygen demand (COD) × L−1 day−1. Selenate was reduced efficiently (>97%) in the nitrate and sulfate fed bioreactor, and a minimal effluent concentration of 39 µg Se × L−1 was obtained. Scanning electron microscopy with energy dispersive X-ray (SEM–EDX) analysis revealed spherical bioprecipitates of ≤2 µm diameter mostly on the gel surface, consisting of selenium with a minor contribution of sulfur. To validate the bioaugmentation success under microbial competition, gel cubes with immobilized S. barnesii cells were added to an Upflow Anaerobic Sludge Bed (UASB) reactor, resulting in earlier selenate (24 hydraulic retention times (HRTs)) and sulfate (44 HRTs) removal and higher nitrate/nitrite removal efficiencies compared to a non-bioaugmented control reactor. S. barnesii was efficiently immobilized inside the UASB bioreactors as the selenate-reducing activity was maintained during long-term operation (58 days), and molecular analysis showed that S. barnesii was present in both the sludge bed and the effluent. This demonstrates that gel immobilization of specialized bacterial strains can supersede wash-out and out-competition of newly introduced strains in continuous bioaugmented systems. Eventually, proliferation of a selenium-respiring specialist occurred in the non-bioaugmented control reactor, resulting in simultaneous nitrate and selenate removal during a later phase of operation.Electronic supplementary materialThe online version of this article (doi:10.1007/s00253-009-1915-x) contains supplementary material, which is available to authorized users.

Contrasting Effects of Dissimilatory Iron(III) and Arsenic(V) Reduction on Arsenic Retention and Transport

Reduction of arsenate As(V) and As-bearing Fe (hydr)- oxides have been proposed as dominant pathways of As release within soils and aquifers. Here we examine As elution from columns loaded with ferrihydrite-coated sand presorbed with As(V) or As(III) at circumneutral pH upon Fe and/or As reduction; biotic stimulated reduction is then compared to abiotic elution. Columns were inoculated with Shewanella putrefaciens strain CN-32 or Sulfurospirillum barnesii strain SES-3, organisms capable of As (V) and Fe (III) reduction, or Bacillus benzoevorans strain HT-1, an organism capable of As(V) but not Fe(III) reduction. On the basis of equal surface coverages, As(III) elution from abiotic columns exceeded As(V) elution by a factor of 2; thus, As(III) is more readily released from ferrihydrite under the imposed reaction conditions. Biologically mediated Asreduction induced by B. benzoevorans enhances the release of total As relative to As (V) under abiotic conditions. However, under Fe reducing conditions invoked by either S. barnesii or S. putrefaciens, approximately three times more As (V or III) was retained within column solids relative to the abiotic experiments, despite appreciable decreases in surface area due to biotransformation of solid phases. Enhanced As sequestration upon ferrihydrite reduction is consistent with adsorption or incorporation of As into biotransformed solids. Our observations indicate that As retention and release from Fe (hydr)oxide(s) is controlled by complex pathways of Fe biotransformation and that reductive dissolution of As-bearing ferrihydrite can promote As sequestration rather than desorption under conditions examined here.

Anaerobic reduction and oxidation of quinone moieties and the reduction of oxidized metals by halorespiring and related organisms

Halorespiring microorganisms have been detected in soils that were not polluted with chlorinated compounds. In this study, we describe alternative electron acceptor utilization by some halorespiring bacteria and phylogenetically related bacteria. It appears that oxidized metals like selenate, arsenate and manganese are rather common electron acceptors for halorespiring species of Desulfitobacterium and Sulfurospirillum and related bacteria. All tested microorganisms are able to reduce anthraquinone-2,6-disulfonate (AQDS) and four tested organisms ( Desulfitobacterium hafniense DP7, Sulfurospirillum barnesii, Sulfurospirillum deleyianum and Sulfurospirillum arsenophilum) are able to oxidize reduced anthrahydroquinone-2,6,-disulfonate (AH 2QDS) as well. The characteristic to reduce oxidized metals, and to reduce and oxidize quinone moieties coupled to energy conservation is a likely explanation for the presence of halorespiring microorganisms in unpolluted soils.

Structural and spectral features of selenium nanospheres produced by Se-respiring bacteria.

Certain anaerobic bacteria respire toxic selenium oxyanions and in doing so produce extracellular accumulations of elemental selenium [Se(0)]. We examined three physiologically and phylogenetically diverse species of selenate- and selenite-respiring bacteria, Sulfurospirillum barnesii, Bacillus selenitireducens, and Selenihalanaerobacter shriftii, for the occurrence of this phenomenon. When grown with selenium oxyanions as the electron acceptor, all of these organisms formed extracellular granules consisting of stable, uniform nanospheres (diameter, approximately 300 nm) of Se(0) having monoclinic crystalline structures. Intracellular packets of Se(0) were also noted. The number of intracellular Se(0) packets could be reduced by first growing cells with nitrate as the electron acceptor and then adding selenite ions to washed suspensions of the nitrate-grown cells. This resulted in the formation of primarily extracellular Se nanospheres. After harvesting and cleansing of cellular debris, we observed large differences in the optical properties (UV-visible absorption and Raman spectra) of purified extracellular nanospheres produced in this manner by the three different bacterial species. The spectral properties in turn differed substantially from those of amorphous Se(0) formed by chemical oxidation of H(2)Se and of black, vitreous Se(0) formed chemically by reduction of selenite with ascorbate. The microbial synthesis of Se(0) nanospheres results in unique, complex, compacted nanostructural arrangements of Se atoms. These arrangements probably reflect a diversity of enzymes involved in the dissimilatory reduction that are subtly different in different microbes. Remarkably, these conditions cannot be achieved by current methods of chemical synthesis.

Reduction of Elemental Selenium to Selenide: Experiments with Anoxic Sediments and Bacteria that Respire Se-Oxyanions

A selenite-respiring bacterium, Bacillus selenitireducens, produced significant levels of Se(-II) (as aqueous HSe−) when supplied with Se(0). B. selenitireducens was also able to reduce selenite [Se(IV)] through Se(0) to Se(-II). Reduction of Se(0) by B. selenitireducens was more rapid in cells grown on colloidal sulfur [S(0)] or Se(IV) as their electron acceptor than for cell lines grown on fumarate. In contrast, three cultures of selenate-respiring bacteria, Sulfurospirillum barnesii, B. arsenicoselenatis, and Selenihalanaerobacter shriftii either were unable to reduce Se(0) to Se(-II) or had only a very limited capacity to achieve this reduction. Biological reduction of Se(0) to Se(-II) was observed during incubation of estuarine sediment slurries, while no such activity was noted in formalin-killed controls. The majority of the Se(-II) produced was found in the sediments as a solid precipitate of FeSe, rather than in solution as HSe−. These results demonstrate that certain anaerobic bacteria have the capacity to reduce Se(0) to Se(-II), providing a possible biological explanation for the occurrence of the selenide species in some sedimentary rocks.

Fractionation of selenium isotopes during bacterial respiratory reduction of selenium oxyanions

Reduction of selenium oxyanions by microorganisms is an important process in the biogeochemical cycling of selenium. Numerous bacteria can reduce Se oxyanions, which are used as electron acceptors during the oxidation of organic matter in anoxic environments. In this study, we used a double spike ( 82Se and 74Se) thermal ionization mass spectrometry technique to quantify the isotopic fractionation achieved by three different species of anaerobic bacteria capable of accomplishing growth by respiratory reduction of selenate [SeO 4 2− or Se(VI)] or selenite [SeO 3 2− or Se(IV)] to Se(IV) or elemental selenium [Se(0)] coupled with the oxidation of lactate. Isotopic discrimination in these closed system experiments was evaluated by Rayleigh fractionation equations and numerical models. Growing cultures of Bacillus selenitireducens, a haloalkaliphile capable of growth using Se(IV) as an electron acceptor, induced a 80Se/ 76Se fractionation of −8.0 ± 0.4‰ (instantaneous ϵ value) during reduction of Se(IV) to Se(0). With Bacillus arsenicoselenatis, a haloalkaliphile capable of growth using Se(VI) as an electron acceptor, fractionations of −5.0 ± 0.5‰ and −6.0 ± 1.0‰ were observed for reduction of Se(VI) to Se(IV) and reduction of Se(IV) to Se(0), respectively. In growing cultures of Sulfurospirillum barnesii, a freshwater species capable of growth using Se(VI), fractionation was small initially, but near the end of the log growth phase, it increased to −4.0 ± 1.0‰ and −8.4 ± 0.4‰ for reduction of Se(VI) to Se(IV) and reduction of Se(IV) to Se(0), respectively. Washed cell suspensions of S. barnesii induced fractionations of −1.1 ± 0.4‰ during Se(VI) reduction, and −9.1 ± 0.5% for Se(IV) reduction, with some evidence for smaller values (e.g., −1.7‰) in the earliest-formed Se(0) results. These results demonstrate that dissimilatory reduction of selenate or selenite induces significant isotopic fractionation, and suggest that significant Se isotope ratio variation will be found in nature.

Mobilization of Arsenite by Dissimilatory Reduction of Adsorbed Arsenate

Sulfurospirillum barnesii is capable of anaerobic growth using ferric iron or arsenate as electron acceptors. Cell suspensions of S. barnesii were able to reduce arsenate to arsenite when the former oxyanion was dissolved in solution, or when it was adsorbed onto the surface of ferrihydrite, a common soil mineral, by a variety of mechanisms (e.g., coprecipitation, presorption). Reduction of Fe(III) in ferrihydrite to soluble Fe(II) also occurred, but dissolution of ferrihydrite was not required in order for adsorbed arsenate reduction to be achieved. This was illustrated by bacterial reduction of arsenate coprecipitated with aluminum hydroxide, a mineral that does not undergo reductive dissolution. The rate of arsenate reduction was influenced by the method in which arsenate became associated with the mineral phases and may have been strongly coupled with arsenate desorption rates. The extent of release of arsenite into solution was governed by adsorption of arsenite onto the ferrihydrite or alumina phase...

Sulfurospirillum barnesii sp. nov. and Sulfurospirillum arsenophilum sp. nov., new members of the Sulfurospirillum clade of the epsilon Proteobacteria.

Two strains of dissimilatory arsenate-reducing vibrio-shaped bacteria are assigned to the genus Sulfurospirillum. These two new species, Sulfurospirillum barnesii strain SES-3T and Sulfurospirillum arsenophilum strain MIT-13T, in addition to Sulfurospirillum sp. SM-5, two strains of Sulfurospirillum deleyianum, and Sulfurospirillum arcachonense, form a distinct clade within the epsilon subclass of the Proteobacteria based on 16S rRNA analysis.