Potential role of a novel psychrotolerant member of the family Geobacteraceae, Geopsychrobacter electrodiphilus gen. nov., sp. nov., in electricity production by a marine sediment fuel cell

Previous studies have shown that members of the family Geobacteraceae that attach to the anodes of sediment fuel cells are directly involved in harvesting electricity by oxidizing organic compounds to carbon dioxide and transferring the electrons to the anode. In order to learn more about this process, microorganisms from the anode surface of a marine sediment fuel cell were enriched and isolated with Fe(III) oxide. Two unique marine isolates were recovered, strains A1(T) and A2. They are gram-negative, nonmotile rods, with abundant c-type cytochromes. Phylogenetic analysis of the 16S rRNA, recA, gyrB, fusA, rpoB, and nifD genes indicated that strains A1(T) and A2 represent a unique phylogenetic cluster within the Geobacteraceae. Both strains were able to grow with an electrode serving as the sole electron acceptor and transferred ca. 90% of the electrons available in their organic electron donors to the electrodes. These organisms are the first psychrotolerant members of the Geobacteraceae reported thus far and can grow at temperatures between 4 and 30 degrees C, with an optimum temperature of 22 degrees C. Strains A1(T) and A2 can utilize a wide range of traditional electron acceptors, including all forms of soluble and insoluble Fe(III) tested, anthraquinone 2,6-disulfonate, and S(0). In addition to acetate, both strains can utilize a number of other organic acids, amino acids, long-chain fatty acids, and aromatic compounds to support growth with Fe(III) nitrilotriacetic acid as an electron acceptor. The metabolism of these organisms differs in that only strain A1(T) can use acetoin, ethanol, and hydrogen as electron donors, whereas only strain A2 can use lactate, propionate, and butyrate. The name Geopsychrobacter electrodiphilus gen. nov., sp. nov., is proposed for strains A1(T) and A2, with strain A1(T) (ATCC BAA-880(T); DSM 16401(T); JCM 12469) as the type strain. Strains A1(T) and A2 (ATCC BAA-770; JCM 12470) represent the first organisms recovered from anodes that can effectively couple the oxidation of organic compounds to an electrode. Thus, they may serve as important model organisms for further elucidation of the mechanisms of microbe-electrode electron transfer in sediment fuel cells.     

Enrichment of members of the family Geobacteraceae associated with stimulation of dissimilatory metal reduction in uranium-contaminated aquifer sediments

Stimulating microbial reduction of soluble U(VI) to insoluble U(IV) shows promise as a strategy for immobilizing uranium in uranium-contaminated subsurface environments. In order to learn more about which microorganisms might be involved in U(VI) reduction in situ, the changes in the microbial community when U(VI) reduction was stimulated with the addition of acetate were monitored in sediments from three different uranium-contaminated sites in the floodplain of the San Juan River in Shiprock, N.Mex. In all three sediments U(VI) reduction was accompanied by concurrent Fe(III) reduction and a dramatic enrichment of microorganisms in the family Geobacteraceae, which are known U(VI)- and Fe(III)-reducing microorganisms. At the point when U(VI) reduction and Fe(III) reduction were nearing completion, Geobacteraceae accounted for ca. 40% of the 16S ribosomal DNA (rDNA) sequences recovered from the sediments with bacterial PCR primers, whereas Geobacteraceae accounted for fewer than 5% of the 16S rDNA sequences in control sediments that were not amended with acetate and in which U(VI) and Fe(III) reduction were not stimulated. Between 55 and 65% of these Geobacteraceae sequences were most similar to sequences from Desulfuromonas species, with the remainder being most closely related to Geobacter species. Quantitative analysis of Geobacteraceae sequences with most-probable-number PCR and TaqMan analyses indicated that the number of Geobacteraceae sequences increased from 2 to 4 orders of magnitude over the course of U(VI) and Fe(III) reduction in the acetate-amended sediments from the three sites. No increase in Geobacteraceae sequences was observed in control sediments. In contrast to the predominance of Geobacteraceae sequences, no sequences related to other known Fe(III)-reducing microorganisms were detected in sediments. These results compare favorably with an increasing number of studies which have demonstrated that Geobacteraceae are important components of the microbial community in a diversity of subsurface environments in which Fe(III) reduction is an important process. The combination of these results with the finding that U(VI) reduction takes place during Fe(III) reduction and prior to sulfate reduction suggests that Geobacteraceae will be responsible for much of the Fe(III) and U(VI) reduction during uranium bioremediation in these sediments.     

Geoalkalibacter ferrihydriticus gen. nov., sp. nov., the first alkaliphilic representative of the family Geobacteraceae, isolated from a soda lake

Investigation of iron reduction in bottom sediments of alkaline soda lakes resulted in the isolation of a new obligately anaerobic iron-reducing bacterium, strain Z-0531, from Lake Khadyn (Tuva Republic, Russia) sediment samples. The cells of strain Z-0531 are short (1.0-1.5 by 0.3-0.5 microm), motile, non-spore-forming, gram-negative rods. The isolate is an obligate alkaliphile, developing in the pH range of 7.8-10.0, with an optimum at pH 8.6. It does not require NaCl but grows at NaCl concentrations of 0-50 g/1l. It can oxidize acetate with such electron acceptors as amorphous Fe(llI) hydroxide (AFH), EDTA-Fe(III), anthraquinone-2,6-disulfonate (quinone), Mn(IV), and S(0). On media with EDTA-Fe(III), the isolate can oxidize, apart from acetate, ethanol, pyruvate, oxalate, arginine, tartrate, lactate, propionate, and serine. H2 is not utilized. The reduced products formed during growth with AFH are siderite or magnetite, depending on the growth conditions. The isolate is incapable of fermenting sugars, peptides, and amino acids. Yeast extract or vitamins are required as growth factors. The organism is capable of dinitrogen fixation and harbors the nifH gene. The DNA G+C content is 55.3 mol %. 16S rRNA analysis places strain Z-0531 into the family Geobacteraceae. Its closest relative (93% similarity) is Desulfuromonas palmitatis. Based on phenotypic distinctions and phylogenetic position, it is proposed that strain Z-0531 be assigned to the new genus and species Geoalkalibacter ferrihydriticus gen. nov., sp. nov.     

Use of Fe(III) as an electron acceptor to recover previously uncultured hyperthermophiles: isolation and characterization of Geothermobacterium ferrireducens gen. nov., sp. nov

It has recently been recognized that the ability to use Fe(III) as a terminal electron acceptor is a highly conserved characteristic in hyperthermophilic microorganisms. This suggests that it may be possible to recover as-yet-uncultured hyperthermophiles in pure culture if Fe(III) is used as an electron acceptor. As part of a study of the microbial diversity of the Obsidian Pool area in Yellowstone National Park, Wyo., hot sediment samples were used as the inoculum for enrichment cultures in media containing hydrogen as the sole electron donor and poorly crystalline Fe(III) oxide as the electron acceptor. A pure culture was recovered on solidified, Fe(III) oxide medium. The isolate, designated FW-1a, is a hyperthermophilic anaerobe that grows exclusively by coupling hydrogen oxidation to the reduction of poorly crystalline Fe(III) oxide. Organic carbon is not required for growth. Magnetite is the end product of Fe(III) oxide reduction under the culture conditions evaluated. The cells are rod shaped, about 0.5 microm by 1.0 to 1.2 microm, and motile and have a single flagellum. Strain FW-1a grows at circumneutral pH, at freshwater salinities, and at temperatures of between 65 and 100 degrees C with an optimum of 85 to 90 degrees C. To our knowledge this is the highest temperature optimum of any organism in the BACTERIA: Analysis of the 16S ribosomal DNA (rDNA) sequence of strain FW-1a places it within the Bacteria, most closely related to abundant but uncultured microorganisms whose 16S rDNA sequences have been previously recovered from Obsidian Pool and a terrestrial hot spring in Iceland. While previous studies inferred that the uncultured microorganisms with these 16S rDNA sequences were sulfate-reducing organisms, the physiology of the strain FW-1a, which does not reduce sulfate, indicates that these organisms are just as likely to be Fe(III) reducers. These results further demonstrate that Fe(III) may be helpful for recovering as-yet-uncultured microorganisms from hydrothermal environments and illustrate that caution must be used in inferring the physiological characteristics of at least some thermophilic microorganisms solely from 16S rDNA sequences. Based on both its 16S rDNA sequence and physiological characteristics, strain FW-1a represents a new genus among the Bacteria. The name Geothermobacterium ferrireducens gen. nov., sp. nov., is proposed (ATCC BAA-426).     

Potential Role of a Novel Psychrotolerant Member of the Family Geobacteraceae, Geopsychrobacter electrodiphilus gen. nov., sp. nov., in Electricity Production by a Marine Sediment Fuel Cell

Previous studies have shown that members of the family Geobacteraceae that attach to the anodes of sediment fuel cells are directly involved in harvesting electricity by oxidizing organic compounds to carbon dioxide and transferring the electrons to the anode. In order to learn more about this process, microorganisms from the anode surface of a marine sediment fuel cell were enriched and isolated with Fe(III) oxide. Two unique marine isolates were recovered, strains A1^T and A2. They are gram-negative, nonmotile rods, with abundant c-type cytochromes. Phylogenetic analysis of the 16S rRNA, recA, gyrB, fusA, rpoB, and nifD genes indicated that strains A1^T and A2 represent a unique phylogenetic cluster within the Geobacteraceae. Both strains were able to grow with an electrode serving as the sole electron acceptor and transferred ca. 90% of the electrons available in their organic electron donors to the electrodes. These organisms are the first psychrotolerant members of the Geobacteraceae reported thus far and can grow at temperatures between 4 and 30°C, with an optimum temperature of 22°C. Strains A1^T and A2 can utilize a wide range of traditional electron acceptors, including all forms of soluble and insoluble Fe(III) tested, anthraquinone 2,6-disulfonate, and S⁰. In addition to acetate, both strains can utilize a number of other organic acids, amino acids, long-chain fatty acids, and aromatic compounds to support growth with Fe(III) nitrilotriacetic acid as an electron acceptor. The metabolism of these organisms differs in that only strain A1^T can use acetoin, ethanol, and hydrogen as electron donors, whereas only strain A2 can use lactate, propionate, and butyrate. The name Geopsychrobacter electrodiphilus gen. nov., sp. nov., is proposed for strains A1^T and A2, with strain A1^T (ATCC BAA-880^T; DSM 16401^T; JCM 12469) as the type strain. Strains A1^T and A2 (ATCC BAA-770; JCM 12470) represent the first organisms recovered from anodes that can effectively couple the oxidation of organic compounds to an electrode. Thus, they may serve as important model organisms for further elucidation of the mechanisms of microbe-electrode electron transfer in sediment fuel cells.     

Use of Fe(III) as an Electron Acceptor To Recover Previously Uncultured Hyperthermophiles: Isolation and Characterization of Geothermobacterium ferrireducens gen. nov., sp. nov.

It has recently been recognized that the ability to use Fe(III) as a terminal electron acceptor is a highly conserved characteristic in hyperthermophilic microorganisms. This suggests that it may be possible to recover as-yet-uncultured hyperthermophiles in pure culture if Fe(III) is used as an electron acceptor. As part of a study of the microbial diversity of the Obsidian Pool area in Yellowstone National Park, Wyo., hot sediment samples were used as the inoculum for enrichment cultures in media containing hydrogen as the sole electron donor and poorly crystalline Fe(III) oxide as the electron acceptor. A pure culture was recovered on solidified, Fe(III) oxide medium. The isolate, designated FW-1a, is a hyperthermophilic anaerobe that grows exclusively by coupling hydrogen oxidation to the reduction of poorly crystalline Fe(III) oxide. Organic carbon is not required for growth. Magnetite is the end product of Fe(III) oxide reduction under the culture conditions evaluated. The cells are rod shaped, about 0.5 μm by 1.0 to 1.2 μm, and motile and have a single flagellum. Strain FW-1a grows at circumneutral pH, at freshwater salinities, and at temperatures of between 65 and 100°C with an optimum of 85 to 90°C. To our knowledge this is the highest temperature optimum of any organism in the Bacteria. Analysis of the 16S ribosomal DNA (rDNA) sequence of strain FW-1a places it within the Bacteria, most closely related to abundant but uncultured microorganisms whose 16S rDNA sequences have been previously recovered from Obsidian Pool and a terrestrial hot spring in Iceland. While previous studies inferred that the uncultured microorganisms with these 16S rDNA sequences were sulfate-reducing organisms, the physiology of the strain FW-1a, which does not reduce sulfate, indicates that these organisms are just as likely to be Fe(III) reducers. These results further demonstrate that Fe(III) may be helpful for recovering as-yet-uncultured microorganisms from hydrothermal environments and illustrate that caution must be used in inferring the physiological characteristics of at least some thermophilic microorganisms solely from 16S rDNA sequences. Based on both its 16S rDNA sequence and physiological characteristics, strain FW-1a represents a new genus among the Bacteria. The name Geothermobacterium ferrireducens gen. nov., sp. nov., is proposed (ATCC BAA-426).     

Geomesophilobacter sediminis gen. nov., sp. nov., Geomonas propionica sp. nov. and Geomonas anaerohicana sp. nov., three novel members in the family Geobacterecace isolated from river sediment and paddy soil

Bacteria in the family Geobacteraceae have been proven to fill important niches in a diversity of anaerobic environments and global biogeochemical processes. Here, three bacterial strains in this family, designated Red875^T, Red259^T, and Red421^T were isolated from river sediment and paddy soils in Japan. All of them are Gram-staining-negative, strictly anaerobic, motile, flagellum-harboring cells that form red colonies on agar plates and are capable of utilizing Fe(III)-NTA, Fe(III) citrate, ferrihydrite, MnO₂, fumarate, and nitrate as electron acceptors with acetate, propionate, pyruvate, and glucose as electron donors. Phylogenetic analysis based on the 16S rRNA gene and 92 concatenated core proteins sequences revealed that strains Red259^T and Red421^T clustered with the type strains of Geomonas species, whereas strain Red875^T formed an independent lineage within the family Geobacteraceae. Genome comparison based on  average nucleotide identity (ANI) and digital DNA–DNA hybridization (dDDH) values clearly distinguished these three strains from other Geobacteraceae members, with lower values than the thresholds for species delineation. Moreover, strain Red875^T also shared low average amino acid identity (AAI) and percentage of conserved proteins (POCP) values with the type species of the family Geobacteraceae. Based on these physiological, chemotaxonomic, and phylogenetic distinctions, we propose that strain Red875^T (=NBRC 114290^T = MCCC 1K04407^T) represents a novel genus in the family Geobacteraceae, namely, Geomesophilobacter sediminis gen. nov., sp. nov., and strains Red259^T (=NBRC 114288^T = MCCC 1K05016^T) and Red421^T (=NBRC 114289^T = MCCC 1K06216^T) represent two novel independent species in the genus Geomonas, namely, Geomonas propionica sp. nov. and Geomonas anaerohicana sp. nov., respectively.     

Geoalkalibacter ferrihydriticus gen. nov. sp. nov., the first alkaliphilic representative of the family Geobacteraceae, isolated from a soda lake

Investigation of iron reduction in bottom sediments of alkaline soda lakes resulted in the isolation of a new obligately anaerobic iron-reducing bacterium, strain Z-0531, from Lake Khadyn (Tuva, Russia) sediment samples. The cells of strain Z-0531 are short (1.0–1.5 by 0.3–0.5 μm), motile, non-spore-forming, gram-negative rods. The isolate is an obligate alkaliphile, developing in the pH range of 7.8–10.0, with an optimum at pH 8.6. It does not require NaCl but grows at NaCl concentrations of 0–50 g/l. It can oxidize acetate with such electron acceptors as amorphous Fe(III) hydroxide (AFH), EDTA-Fe(III), anthraquinone-2,6-disulfonate (quinone), Mn(IV), and S⁰. On medium with EDTA-Fe(III), the isolate can oxidize, apart from acetate, ethanol, pyruvate, oxalate, arginine, tartrate, lactate, propionate, and serine. H₂ is not utilized. The reduced products formed during growth with AFH are siderite or magnetite, depending on the growth conditions. The isolate is incapable of fermenting sugars, peptides, and amino acids. Yeast extract or vitamins are required as growth factors. The organism is capable of dinitrogen fixation and harbors the nifH gene. The DNA G+C content is 55.3 mol %. 16S rRNA analysis places strain Z-0531 into the family Geobacteraceae. Its closest relative (93% similarity) is Desulfuromonas palmitatis. Based on phenotypic distinctions and phylogenetic position, it is proposed that this strain be assigned to the new genus and species Geoalkalibacter ferrihydriticus gen. nov., sp. nov. (Z-0531^T-DSMZ-17813-VKMB-2401). Original Russian Text ? D.G. Zavarzina, T.V. Kolganova, E.S. Boulygina, N.A. Kostrikina, T.P. Tourova, G.A. Zavarzin, 2006, published in Mikrobiologiya, 2006, Vol. 75, No. 6, pp. 775–785.     

Enumeration and characterization of iron(III)-reducing microbial communities from acidic subsurface sediments contaminated with uranium(VI)

Iron(III)-reducing bacteria have been demonstrated to rapidly catalyze the reduction and immobilization of uranium(VI) from contaminated subsurface sediments. Thus, these organisms may aid in the development of bioremediation strategies for uranium contamination, which is prevalent in acidic subsurface sediments at U.S. government facilities. Iron(III)-reducing enrichment cultures were initiated from pristine and contaminated (high in uranium, nitrate; low pH) subsurface sediments at pH 7 and pH 4 to 5. Enumeration of Fe(III)-reducing bacteria yielded cell counts of up to 240 cells ml(-1) for the contaminated and background sediments at both pHs with a range of different carbon sources (glycerol, acetate, lactate, and glucose). In enrichments where nitrate contamination was removed from the sediment by washing, MPN counts of Fe(III)-reducing bacteria increased substantially. Sediments of lower pH typically yielded lower counts of Fe(III)-reducing bacteria in lactate- and acetate-amended enrichments, but higher counts were observed when glucose was used as an electron donor in acidic enrichments. Phylogenetic analysis of 16S rRNA gene sequences extracted from the highest positive MPN dilutions revealed that the predominant members of Fe(III)-reducing consortia from background sediments were closely related to members of the Geobacteraceae family, whereas a recently characterized Fe(III) reducer (Anaeromyxobacter sp.) and organisms not previously shown to reduce Fe(III) (Paenibacillus and Brevibacillus spp.) predominated in the Fe(III)-reducing consortia of contaminated sediments. Analysis of enrichment cultures by terminal restriction fragment length polymorphism (T-RFLP) strongly supported the cloning and sequencing results. Dominant members of the Fe(III)-reducing consortia were observed to be stable over several enrichment culture transfers by T-RFLP in conjunction with measurements of Fe(III) reduction activity and carbon substrate utilization. Enrichment cultures from contaminated sites were also shown to rapidly reduce millimolar amounts of U(VI) in comparison to killed controls. With DNA extracted directly from subsurface sediments, quantitative analysis of 16S rRNA gene sequences with MPN-PCR indicated that Geobacteraceae sequences were more abundant in pristine compared to contaminated environments,whereas Anaeromyxobacter sequences were more abundant in contaminated sediments. Thus, results from a combination of cultivation-based and cultivation-independent approaches indicate that the abundance/community composition of Fe(III)-reducing consortia in subsurface sediments is dependent upon geochemical parameters (pH, nitrate concentration) and that microorganisms capable of producing spores (gram positive) or spore-like bodies (Anaeromyxobacter) were representative of acidic subsurface environments.     

Characterization of citrate synthase from Geobacter sulfurreducens and evidence for a family of citrate synthases similar to those of eukaryotes throughout the Geobacteraceae

Members of the family Geobacteraceae are commonly the predominant Fe(III)-reducing microorganisms in sedimentary environments, as well as on the surface of energy-harvesting electrodes, and are able to effectively couple the oxidation of acetate to the reduction of external electron acceptors. Citrate synthase activity of these organisms is of interest due to its key role in acetate metabolism. Prior sequencing of the genome of Geobacter sulfurreducens revealed a putative citrate synthase sequence related to the citrate synthases of eukaryotes. All citrate synthase activity in G. sulfurreducens could be resolved to a single 49-kDa protein via affinity chromatography. The enzyme was successfully expressed at high levels in Escherichia coli with similar properties as the native enzyme, and kinetic parameters were comparable to related citrate synthases (kcat= 8.3 s(-1); Km= 14.1 and 4.3 microM for acetyl coenzyme A and oxaloacetate, respectively). The enzyme was dimeric and was slightly inhibited by ATP (Ki= 1.9 mM for acetyl coenzyme A), which is a known inhibitor for many eukaryotic, dimeric citrate synthases. NADH, an allosteric inhibitor of prokaryotic hexameric citrate synthases, did not affect enzyme activity. Unlike most prokaryotic dimeric citrate synthases, the enzyme did not have any methylcitrate synthase activity. A unique feature of the enzyme, in contrast to citrate synthases from both eukaryotes and prokaryotes, was a lack of stimulation by K+ ions. Similar citrate synthase sequences were detected in a diversity of other Geobacteraceae members. This first characterization of a eukaryotic-like citrate synthase from a prokaryote provides new insight into acetate metabolism in Geobacteraceae members and suggests a molecular target for tracking the presence and activity of these organisms in the environment.     

Growth of iron(III)-reducing bacteria on clay minerals as the sole electron acceptor and comparison of growth yields on a variety of oxidized iron forms

Smectite clay minerals are abundant in soils and sediments worldwide and are typically rich in Fe. While recent investigations have shown that the structural Fe(III) bound in clay minerals is reduced by microorganisms, previous studies have not tested growth with clay minerals as the sole electron acceptor. Here we have demonstrated that a pure culture of Shewanella oneidensis strain MR-1 as well as enrichment cultures of Fe(III)-reducing bacteria from rice paddy soil and subsurface sediments are capable of conserving energy for growth with the structural Fe(III) bound in smectite clay as the sole electron acceptor. Pure cultures of S. oneidensis were used for more detailed growth rate and yield experiments on various solid- and soluble-phase electron acceptors [smectite, Fe(III) oxyhydroxide FeOOH, Fe(III) citrate, and oxygen] in the same minimal medium. Growth was assessed as direct cell counts or as an increase in cell carbon (measured as particulate organic carbon). Cell counts showed that similar growth of S. oneidensis (10(8) cells ml(-1)) occurred with smectitic Fe(III) and on other Fe forms [amorphous Fe(III) oxyhydroxide, and Fe citrate] or oxygen as the electron acceptor. In contrast, cell yields of S. oneidensis measured as the increase in cell carbon were similar on all Fe forms tested while yields on oxygen were five times higher, in agreement with thermodynamic predictions. Over a range of particle loadings (0.5 to 4 g liter(-1)), the increase in cell number was highly correlated to the amount of structural Fe in smectite reduced. From phylogenetic analysis of the complete 16S rRNA gene sequences, a predominance of clones retrieved from the clay mineral-reducing enrichment cultures were most closely related to the low-G+C gram-positive members of the Bacteria (Clostridium and Desulfitobacterium) and the delta-Proteobacteria (members of the Geobacteraceae). Results indicate that growth with smectitic Fe(III) is similar in magnitude to that with Fe(III) oxide minerals and is dependent upon the mineral surface area available. Iron(III) bound in clay minerals should be considered an important electron acceptor supporting the growth of bacteria in soils or sedimentary environments.     

Ferrovibrio denitrificans gen. nov., sp. nov., a novel neutrophilic facultative anaerobic Fe(II)-oxidizing bacterium

A neutrophilic Fe(II)-oxidizing bacterium was isolated from the redox zone of a low-salinity spring in Krasnodar krai (Russia), at the FeS-Fe(OH)(3) interface deposited at the sediment surface. The cells of strain Sp-1 were short, thin motile vibrioids with one polar flagellum dividing by binary fission. The optimal values and ranges for pH and temperature were pH 6.2 (5.5-8) and 35?°C (5-45?°C), respectively. The organism was a facultative anaerobe. Strain Sp-1 was capable of organotrophic, lithoheterotrophic and mixotrophic growth with Fe(II) as an electron donor. The denitrification chain was 'disrupted'. Oxidation of Fe(II) was coupled to reduction of NO3 - to NO2 - or of N(2) O to N(2) , as well as under microaerobic conditions, with O(2) as an electron acceptor. The DNA G+C content was 64.2?mol%. According to the results of phylogenetic analysis, the strain was 10.6-12% remote from the closest relatives, members of the genera Sneathiella, Inquilinus, Oceanibaculum and Phaeospirillum within the Alphaproteobacteria. Based on its morphological, physiological and taxonomic characteristics, together with the results of phylogenetic analysis, strain Sp-1 is described as a member of a new genus Ferrovibrio gen. nov., with the type species Ferrovibrio denitrificans sp. nov. and the type strain Sp-1(T) (=?LMG 25817(T) =?VKM B-2673(T) ).     

Phylogenetic and physiological diversity of dissimilatory ferric iron reducers in sediments of the polluted Scheldt estuary, Northwest Europe

The potential for dissimilatory ferric iron [Fe(III)] reduction in intertidal sediments of the polluted Scheldt estuary, Northwest Europe, was assessed by combining field-based geochemical measurements with laboratory experiments on the associated microbiology. Microbial communities at a freshwater and brackish location were characterized by culture-independent 16S rRNA gene analysis, as well as enrichments, strain isolation and physiological screening. Dilution-to-extinction batch enrichments using a variety of Fe(III) sources were performed. The dilution factor of the inoculum in the enrichments had a more determining effect on the Fe(III)-reducing microbial community structure than the Fe(III) source. Well-known Fe(III) reducers, including members of the family Geobacteraceae and the genus Shewanella, constituted only a small fraction (< or = 1%) of the in situ microbial community. Instead, facultative anaerobic Ralstonia and strictly anaerobic, spore-forming Clostridium species dominated Fe(III) reduction. These species were able to utilize a variety of electron acceptors. This flexibility may help the organisms to survive in the dynamic estuarine environment. The high diversity and abundance of culturable Fe(III) reducers (4.6 x 10(5) and 2.4 x 10(4) cells g(-1) sediment at the freshwater and brackish site respectively), plus the high concentrations of chemically reducible solid-phase Fe(III) at the sites, implied a high potential for dissimilatory Fe(III) reduction in the estuarine sediments. Pore water chemical data further supported in situ dissimilatory Fe(III) reduction.     

Ferribacterium limneticum, gen. nov., sp. nov., an Fe(III)-reducing microorganism isolated from mining-impacted freshwater lake sediments

A dissimilatory Fe(III)-reducing bacterium was isolated from mining-impacted lake sediments and designated strain CdA-1. The strain was isolated from a 4-month enrichment culture with acetate and Fe(III)-oxyhydroxide. Strain CdA-1 is a motile, obligately anaerobic rod, capable of coupling the oxidation of acetate and other organic acids to the reduction of ferric iron. Fe(III) reduction was not observed using methanol, ethanol, isopropanol, propionate, succinate, fumarate, H₂, citrate, glucose, or phenol as potential electron donors. With acetate as an electron donor, strain CdA-1 also grew by reducing nitrate or fumarate. Growth was not observed with acetate as electron donor and O₂, sulfoxyanions, nitrite, trimethylamine N-oxide, Mn(IV), As(V), or Se(VI) as potential terminal electron acceptors. Comparative 16 S rRNA gene sequence analyses show strain CdA-1 to be most closely related (93.6% sequence similarity) to Rhodocyclus tenuis. However, R. tenuis did not grow heterotrophically by Fe(III) reduction, nor did strain CdA-1 grow photrophically. We propose that strain CdA-1 represents a new genus and species, Ferribacterium limneticum. Strain CdA-1 represents the first dissimilatory Fe(III) reducer in the β subclass of Proteobacteria, as well as the first Fe(III) reducer isolated from mine wastes. Received: 14 July 1998 / Accepted: 14 December 1998     

Microbial communities associated with anaerobic benzene degradation in a petroleum-contaminated aquifer.

Microbial community composition associated with benzene oxidation under in situ Fe(III)-reducing conditions in a petroleum-contaminated aquifer located in Bemidji, Minn., was investigated. Community structure associated with benzene degradation was compared to sediment communities that did not anaerobically oxidize benzene which were obtained from two adjacent Fe(III)-reducing sites and from methanogenic and uncontaminated zones. Denaturing gradient gel electrophoresis of 16S rDNA sequences amplified with bacterial or Geobacteraceae-specific primers indicated significant differences in the composition of the microbial communities at the different sites. Most notable was a selective enrichment of microorganisms in the Geobacter cluster seen in the benzene-degrading sediments. This finding was in accordance with phospholipid fatty acid analysis and most-probable-number-PCR enumeration, which indicated that members of the family Geobacteraceae were more numerous in these sediments. A benzene-oxidizing Fe(III)-reducing enrichment culture was established from benzene-degrading sediments and contained an organism closely related to the uncultivated Geobacter spp. This genus contains the only known organisms that can oxidize aromatic compounds with the reduction of Fe(III). Sequences closely related to the Fe(III) reducer Geothrix fermentans and the aerobe Variovorax paradoxus were also amplified from the benzene-degrading enrichment and were present in the benzene-degrading sediments. However, neither G. fermentans nor V. paradoxus is known to oxidize aromatic compounds with the reduction of Fe(III), and there was no apparent enrichment of these organisms in the benzene-degrading sediments. These results suggest that Geobacter spp. play an important role in the anaerobic oxidation of benzene in the Bemidji aquifer and that molecular community analysis may be a powerful tool for predicting a site's capacity for anaerobic benzene degradation.     

Thermococcus indicus sp. nov., a Fe(III)-reducing hyperthermophilic archaeon isolated from the Onnuri Vent Field of the Central Indian Ocean ridge

A strictly anaerobic, dissimilatory Fe(III)-reducing hyperthermophilic archaeon, designated as strain IOH1T, was isolated from a new deep-sea hydrothermal vent (Onnuri Vent Field) area in the Central Indian Ocean ridge. Strain IOH1T showed > 99% 16S rRNA gene sequence similarity with Thermococcus celericrescens TS2T (99.4%) and T. siculi DSM 12349T (99.2%). Additional three species T. barossii SHCK-94T (99.0%), T. celer Vu13T (98.8%), and T. piezophilus (98.6%) showed > 98.6% of 16S rRNA gene sequence similarity, however, the maximum OrthoANI value is 89.8% for the genome of T. celericrescens TS2T. Strain IOH1T cells are coccoid, 1.2–1.8 μm in diameter, and motile by flagella. Growth was at 70–82°C (optimum 80°C), pH 5.4–8.0 (optimum pH 6.0) with 2–4% (optimum 3%) NaCl. Growth of strain IOH1T was enhanced by starch, pyruvate, D(+)-maltose and maltodextrin as a carbon sources, and elemental sulfur as an electron acceptor; clearly different from those of related species T. celecrescens DSM 17994T and T. siculi DSM 12349T. Strain IOH1T, T. celercrescence DSM 17994T, and T. siculi DSM 12349T reduced soluble Fe(III)-citrate present in the medium, whereas the amount of total cellular proteins increased with the concomitant accumulation of Fe(II). We determined a circular chromosome of 2,234 kb with an extra-chromosomal archaeal plasmid, pTI1, of 7.7 kb and predicted 2,425 genes. The DNA G + C content was 54.9 mol%. Based on physiological properties, phylogenetic, and genome analysis, we proposed that strain IOH1T (= KCTC 15844T = JCM 39077T) is assigned to a new species in the genus Thermococcus and named Thermococcus indicus sp. nov.     

Salinicola socius gen. nov., sp. nov., a moderately halophilic bacterium from a naphthalene-utilizing microbial association

A chemoorganotrophic, moderately halophilic bacterium (strain SMB35) has been isolated from a naphthalene-utilizing microbial community obtained from salt mines (Perm region of Russia). Strain SMB35 grows in a wide salinity range, 0.5 to 30% (wt/vol) NaCl. Cells are gram-negative rods motile by means of a single polar flagellum. The predominant fatty acids are 16:1omega7, 16:0, 18:1omega7, and 19 cy. The major lipoquinone is an unsaturated ubiquinone with nine isoprene units (Q-9). The DNA G+C content is 63.0 mol%. The 16S rDNA-based phylogenetic analysis has shown that strain SMB35 formed a separate clade in the cluster of the family Halomonadaceae. The 16S rDNA sequence similarity of the isolate to the members of the family is in the range from 90.6% to 95.1%. The phylogenetic and phenotypic differences from Halomonas elongata (the type species of the genus) and from other members of the family suggest that the isolate represents a novel genus and species, for which the name Salinicola socius gen. nov., sp. nov. is proposed. The type strain is SMB35(T) (=VKM B-2397(T)).