Sulfate-reducing bacteria methylate mercury at variable rates in pure culture and in marine sediments

Differences in methylmercury (CH(3)Hg) production normalized to the sulfate reduction rate (SRR) in various species of sulfate-reducing bacteria (SRB) were quantified in pure cultures and in marine sediment slurries in order to determine if SRB strains which differ phylogenetically methylate mercury (Hg) at similar rates. Cultures representing five genera of the SRB (Desulfovibrio desulfuricans, Desulfobulbus propionicus, Desulfococcus multivorans, Desulfobacter sp. strain BG-8, and Desulfobacterium sp. strain BG-33) were grown in a strictly anoxic, minimal medium that received a dose of inorganic Hg 120 h after inoculation. The mercury methylation rates (MMR) normalized per cell were up to 3 orders of magnitude higher in pure cultures of members of SRB groups capable of acetate utilization (e.g., the family Desulfobacteriaceae) than in pure cultures of members of groups that are not able to use acetate (e.g., the family Desulfovibrionaceae). Little or no Hg methylation was observed in cultures of Desulfobacterium or Desulfovibrio strains in the absence of sulfate, indicating that Hg methylation was coupled to respiration in these strains. Mercury methylation, sulfate reduction, and the identities of sulfate-reducing bacteria in marine sediment slurries were also studied. Sulfate-reducing consortia were identified by using group-specific oligonucleotide probes that targeted the 16S rRNA molecule. Acetate-amended slurries, which were dominated by members of the Desulfobacterium and Desulfobacter groups, exhibited a pronounced ability to methylate Hg when the MMR were normalized to the SRR, while lactate-amended and control slurries had normalized MMR that were not statistically different. Collectively, the results of pure-culture and amended-sediment experiments suggest that members of the family Desulfobacteriaceae have a greater potential to methylate Hg than members of the family Desulfovibrionaceae have when the MMR are normalized to the SRR. Hg methylation potential may be related to genetic composition and/or carbon metabolism in the SRB. Furthermore, we found that in marine sediments that are rich in organic matter and dissolved sulfide rapid CH(3)Hg accumulation is coupled to rapid sulfate reduction. The observations described above have broad implications for understanding the control of CH(3)Hg formation and for developing remediation strategies for Hg-contaminated sediments.     

A highly selective direct method of detecting sulphate-reducing bacteria in crude oil

AIMS: The aim of this work was to develop a highly selective method of detecting sulphate-reducing bacteria (SRB) in crude oil. METHODS: A pair of PCR primers was designed based on an alignment of the nucleotide sequence of the 16S rRNA genes from the Desulfovibrionaceae family. DNA extraction from crude oil was performed by the method using zirconia beads and a stool kit. RESULTS: The PCR specifically detected Desulfovibrio and Desulfomicrobium in a sediment sample. When nucleic acids extracted directly from crude oil were used for the PCR, 16S rRNA genes of Desulfovibrio and Thermodesulforhabdus norvegicus were detected. IMPACT OF STUDY: A simple direct method for detection of the SRB in crude oil using PCR was established.     

Temperature and nutrient induced responses of Lake Fryxell sulfate-reducing prokaryotes and description of Desulfovibrio lacusfryxellense, sp. nov., a pervasive, cold-active, sulfate-reducing bacterium from Lake Fryxell, Antarctica

The effects of temperature and carbon substrate availability on the stimulation of sulfate reduction by indigenous populations of sulfate-reducing prokaryotes (SRP) in permanently ice-covered Lake Fryxell, Antarctica were investigated. Psychrophilic and halotolerant, lactate-degrading SRP showed significant metabolic activity throughout all sampled depths of the water column, suggesting that such organisms, possibly of marine origin, may be key contributors to carbon and sulfur cycling in Lake Fryxell. Planktonic and benthic strains of lactate-oxidizing sulfate-reducing bacteria (SRB) were isolated from samples of various depths of the anoxic water column and from surficial sediments. Phylogenetic analyses of 16S rRNA gene sequences placed the Fryxell sulfate-reducer (FSR) strains within the Deltaproteobacteria and showed them to be most closely related to the Arctic marine species of SRB Desulfovibrio frigidus and Desulfovibrio ferrireducens. Based on phylogenetic and phenotypic differences between the Antarctic FSR strains and related species of the genus Desulfovibrio, strain FSRs^T (=DSM 23315^T =ATCC BAA-2083^T) is proposed as the type strain of a novel species of cold-active SRB, Desulfovibrio lacusfryxellense, sp. nov.     

Diversity of bacteria and Archaea in sulphate-reducing enrichment cultures inoculated from serial dilution of Zostera noltii rhizosphere samples

We have analysed the diversity of culturable sulphate-reducing bacteria (SRB) in Zostera noltii colonized sediments from Bassin d'Arcachon (France). Four organic substrates have been tested as well as the combination of H2 and CO2 to select for lithotrophic SRB. All energy sources were supplied in parallel cultures that were amended with yeast extract plus NH4+ and prepared without a source of combined nitrogen, the latter to select for diazotrophic SRB. The 10 different enrichment media were inoculated from serial dilution of rhizosphere samples. The highest dilution cultures yielding positive growth (i.e. 10-7) were studied by molecular techniques (16S rDNA clone libraries, RISA and ARDRA). Lactate as a single organic substrate in combination with a source of combined nitrogen resulted in selection of members of the Desulfovibrionaceae. Surprisingly, when lactate was added without a source of combined nitrogen, Desulfobacteriaceae were selected. A strong influence of the presence or absence of combined nitrogen was also observed for the substrates sucrose and fructose. Whereas the liquid culture growing on sucrose and NH4+ systematically yielded 16S rDNA clones related to an environmental unidentified green sulphur bacterium (OPS185), on plates we were able to isolate a SRB related to Desulfovibrio dechloracetivorans, which likely represents a non-described species. Under diazotrophic conditions, sucrose selected for SRB clones related to the cluster formed by Desulfovibrio zosterae, Desulfovibrio salexigens and Desulfovibrio bastinii. The corresponding isolate obtained on plates showed only low sequence similarity with this closest neighbour (93.8%), and we suggest that it also represents a non-described species. Surprisingly, a 16S rDNA sequence corresponding to an archaeon, i.e. a non-extremophile Crenoarchaeota, was retrieved from several of the SRB enrichment cultures even after subsequent transfers.     

Isolation and characterization of sulphate-reducing bacteria <Emphasis Type="Italic">Desulfovibrio vulgaris</Emphasis> from Vajreshwari thermal springs in Maharashtra,India

Sulphate-reducing bacteria (SRB) in the thermal springs of Vajreshwari were investigated with combined microbiological and molecular approaches. A sulphate-reducing bacteria medium containing lactate was used for enrichment and isolation, which yielded Gram negative, rod shaped, anaerobic, non-spore forming and motile bacteria capable of reducing sulphate to sulphide. These grew at temperatures ranging from 25 to 55 °C and could use pyruvate, lactate and ethanol as electron donors. Desulfoviridin was detected in all the isolates. The partial 16S rRNA and dissimilatory sulphite reductase (DSR) gene sequences of five representative isolates revealed that the strains belonged to the sulphur reducing bacterial species Desulfovibrio vulgaris.     

Overestimation of the abundance of sulfate-reducing bacteria in human feces by quantitative PCR targeting the Desulfovibrio 16S rRNA gene

The dominant genus of sulfate-reducing bacteria (SRB) in humans is Desulfovibrio, and quantitative PCR (QPCR) targeting the 16S rRNA gene is often used in assays. We show that the 16S rRNA gene assay overestimated SRB abundance in feces from 24 adults compared to QPCR assays using primers targeting two genes involved in SRB energy metabolism.     

Phylogenetic Identification and Substrate Uptake Patterns of Sulfate-Reducing Bacteria Inhabiting an Oxic-Anoxic Sewer Biofilm Determined by Combining Microautoradiography and Fluorescent In Situ Hybridization

We simultaneously determined the phylogenetic identification and substrate uptake patterns of sulfate-reducing bacteria (SRB) inhabiting a sewer biofilm with oxygen, nitrate, or sulfate as an electron acceptor by combining microautoradiography and fluorescent in situ hybridization (MAR-FISH) with family- and genus-specific 16S rRNA probes. The MAR-FISH analysis revealed that Desulfobulbus hybridized with probe 660 was a dominant SRB subgroup in this sewer biofilm, accounting for 23% of the total SRB. Approximately 9 and 27% of Desulfobulbus cells detected with probe 660 could take up [¹⁴C]propionate with oxygen and nitrate, respectively, as an electron acceptor, which might explain the high abundance of this species in various oxic environments. Furthermore, more than 40% of Desulfobulbus cells incorporated acetate under anoxic conditions. SRB were also numerically important members of H₂-utilizing and ¹⁴CO₂-fixing microbial populations in this sewer biofilm, accounting for roughly 42% of total H₂-utilizing bacteria hybridized with probe EUB338. A comparative 16S ribosomal DNA analysis revealed that two SRB populations, related to the Desulfomicrobium hypogeium and the Desulfovibrio desulfuricans MB lineages, were found to be important H₂ utilizers in this biofilm. The substrate uptake characteristics of different phylogenetic SRB subgroups were compared with the characteristics described to date. These results provide further insight into the correlation between the 16S rRNA phylogenetic diversity and the physiological diversity of SRB populations inhabiting sewer biofilms.     

Quantification of Gram-negative sulphate-reducing bacteria in rice field soil by 16S rRNA gene-targeted real-time PCR

For the quantification of Gram-negative sulphate reducers in rice fields, 11 real-time PCR assays were established targeting 16S rRNA genes combined with SybrGreen detection. Three of these assays were specific for the "main" groups, i.e. the Desulfovibrionaceae, the Desulfobacteraceae and Desulfobulbus sp., whereas eight assays were developed for subgroups within the first two main groups. The detection limits of the assays were between 2 x 10(5) and 4 x 10(3) targets g(-1) (wet weight) or less than 0.02% of the eubacterial 16S rDNA targets in bulk soil, rhizosphere soil and rice root DNA extracts. Analysis of soil spiked with defined cell numbers of sulphate-reducing bacteria showed good correlation of measured target numbers to amended cells. In rice field bulk and rhizosphere soil, the Desulfobacteraceae were the predominant main group with target numbers of 6.4 x 10(7) (+/-1.0 x 10(7)) and 7.5 x 10(7) (+/-1.7 x 10(7)), respectively. Within this group the Desulforhabdus/Synthrophobacter assemblage and Desulfobacterium sp. were predominant. At the rice roots, the three main groups were abundant in similar numbers (approx. 1.0 x 10(8)) indicating that the relative abundance of the Desulfovibrionaceae and also of Desulfobulbus sp. was increased, relatively to the Desulfobacteraceae. Within the Desulfovibrionaceae the subgroup was predominant that was detected by assay DSV-II. This assay detects many from rice field soil isolated Desulfovibrio-strains and molecular retrieved sequences. Therefore these organisms that were already detected in the rice field environment by isolation and by molecular techniques are indeed best adapted to the conditions provided by the rice roots.     

Biogeochemical cycling of methylmercury at Barn Island Salt Marsh,Stonington, CT,USA

Monomethylmercury (MMHg) is toxic, and is the primary form of Hg thatbioaccumulates in the food web. An understanding of its distribution,production, and transport is needed. Prior investigations indicate thatmethylation is mediated by sulfate-reducing bacteria, yet limited in highsulfate environments. High rates of microbial respiration and strong oxygengradients are found in salt marshes. It is hypothesized that significant in situ methylation takes place in the redox transition zone of sulfate rich( 28 mM) salt marsh sediment. Results from a water column surveyof Barn Island Salt Marsh in October 1996 showed that ca. 61pmol m^-2 d^-1 of dissolved MMHg were discharged toadjacent coastal waters, while 16 pmol m^-2 d^-1 ofparticulate MMHg were entrained in the marsh, implying an in situsource. In-sediment MMHg production rates were determined by²⁰³Hg radiotracer studies. At the surface, methylation rates variedover both long (i.e., 100's m; 11–1120 pmol m^-2 d^-1) andshort (i.e., 10 cm; 11–108 pmol m^-2 d^-1) spatial scales. Methylation rate profiles from both low and high MMHg production sitesexhibited an exponential decrease below the redox transition zone. Porewater was collected with multi-chambered in situ dialysis (30 kDa)samplers [Peepers] and analyzed for MMHg. Temporal differences in porewater MMHg accumulation (i.e., May > September > November)were found. Results from May showed a significant gradient at thesediment water interface. The transport out of the sediments estimated byFick's Law (ca. 390 pmol MMHg m-2 d^-1) suggeststhat MMHg entered the marsh water by diffusion. This workdemonstrates the potential for elevated in situ Hg methylation in highsulfate environments.     

Population Dynamics of a Single-Stage Sulfidogenic Bioreactor Treating Synthetic Zinc-Containing Waste Streams

Waste streams from industrial processes such as metal smelting or mining contain high concentrations of sulfate and metals with low pH. Dissimilatory sulfate reduction carried out by sulfate-reducing bacteria (SRB) at low pH can combine sulfate reduction with metal-sulfide precipitation and thus open possibilities for selective metal recovery. This study investigates the microbial diversity and population changes of a single-stage sulfidogenic gas-lift bioreactor treating synthetic zinc-rich waste water at pH 5.5 by denaturing gradient gel electrophoresis of 16S rRNA gene fragments and quantitative polymerase chain reaction. The results indicate the presence of a diverse range of phylogenetic groups with the predominant microbial populations belonging to the Desulfovibrionaceae from δ-Proteobacteria. Desulfovibrio desulfuricans-like populations were the most abundant among the SRB during the three stable phases of varying sulfide and zinc concentrations and increased from 13% to 54% of the total bacterial populations over time. The second largest group was Desulfovibrio marrakechensis-like SRB that increased from 1% to about 10% with decreasing sulfide concentrations. Desulfovibrio aminophilus-like populations were the only SRB to decrease in numbers with decreasing sulfide concentrations. However, their population was <1% of the total bacterial population in the reactor at all analyzed time points. The number of dissimilatory sulfate reductase (DsrA) gene copies per number of SRB cells decreased from 3.5 to 2 DsrA copies when the sulfide concentration was reduced, suggesting that the cells' sulfate-reducing capacity was also lowered. This study has identified the species present in a single-stage sulfidogenic bioreactor treating zinc-rich wastewater at low pH and provides insights into the microbial ecology of this biotechnological process.     

Genetic Diversity among Thermophilic Bacteria Isolated from Geothermal Sites by Using Two PCR Typing Methods

Microbial communities thriving at two hot springs, Hammam Pharaon (Pharaoh's Bath) and Oyoun Mossa (Moses springs), in Egypt was studied by cultural and molecular methods. Thirteen morphologically distinct strains of facultative anaerobic thermophilic bacterial isolates have been characterized and identified using phenotypic and genotypic characters including RAPD-PCR, ERIC-PCR typing, plasmid analysis and 16S rRNA sequencing. All isolates produced plasmid DNA with various sizes ranging from 0.7 kb to a larger plasmid 7.2 kb. The bacterial strains could tolerate a temperature range between 45 to 85°C and a pH between 4–11. Also, sulphate-reducing bacteria (SRB) in the thermal springs were investigated with combined biochemical and molecular approaches. A sulphate-reducing bacteria medium containing lactate was used for enrichment and isolation, which yielded Gram negative, rod shaped, anaerobic, non-spore-forming and motile bacteria capable of reducing sulphate to sulphide. These grew at temperatures ranging from 30 to 50°C and could use pyruvate, lactate and ethanol as electron donors. The dissimilatory sulphite reductase (DSR) gene sequences of eleven representative isolates revealed that the strains belonged to the sulphur reducing bacterial species Desulfovibrio vulgaris. 16S rRNA gene partial sequence results indicated the presence of novel or existing species of Bacillus (one species), Anoxybacillus (four species) and Geobacillus (eight species). In this study phenotypic and genotypic diversity were applied for the first time to differentiate thermophilic bacteria of such geothermal sites in Sinai, Egypt.     

Depth profile of sulfate-reducing bacterial ribosomal RNA and mercury methylation in an estuarine sediment

Abstract: The community structure of complex anaerobic microbial communities has been difficult to elucidate because of an inability to cultivate most of the contributing populations. In this study, the distribution of sulfate-reducing bacteria (SRB) in anaerobic sediments was determined using oligonucleotide probes complementary to the 16S ribosomal RNAs of major phylogenetic groups. Sediment cores were collected from Santa Rosa Sound in northwest Florida, and sectioned by depth into 1 to 2 cm fractions. Nucleic acids were extracted from each fraction and hybridized with the SRB-specific ribosomal RNA probes. SRB ribosomal RNAs accounted for almost 5% of the microbial community ribosomal RNA pool in the 3–4 cm depth fraction and were dominated by Desulfovibrionaceae ribosomal RNA. The SRB ribosomal RNA peak coincided with mercury methylation, an activity attributed to SRB. Profiles of the ribosomal RNAs indicate that SRB populations in sediments are stratified by depth.     

Mercury Methylation Independent of the Acetyl-Coenzyme A Pathway in Sulfate-Reducing Bacteria

Sulfate-reducing bacteria (SRB) in anoxic waters and sediments are the major producers of methylmercury in aquatic systems. Although a considerable amount of work has addressed the environmental factors that control methylmercury formation and the conditions that control bioavailability of inorganic mercury to SRB, little work has been undertaken analyzing the biochemical mechanism of methylmercury production. The acetyl-coenzyme A (CoA) pathway has been implicated as being key to mercury methylation in one SRB strain, Desulfovibrio desulfuricans LS, but this result has not been extended to other SRB species. To probe whether the acetyl-CoA pathway is the controlling biochemical process for methylmercury production in SRB, five incomplete-oxidizing SRB strains and two Desulfobacter strains that do not use the acetyl-CoA pathway for major carbon metabolism were assayed for methylmercury formation and acetyl-CoA pathway enzyme activities. Three of the SRB strains were also incubated with chloroform to inhibit the acetyl-CoA pathway. So far, all species that have been found to have acetyl-CoA activity are complete oxidizers that require the acetyl-CoA pathway for basic metabolism, as well as methylate mercury. Chloroform inhibits Hg methylation in these species either by blocking the methylating enzyme or by indirect effects on metabolism and growth. However, we have identified four incomplete-oxidizing strains that clearly do not utilize the acetyl-CoA pathway either for metabolism or mercury methylation (as confirmed by the absence of chloroform inhibition). Hg methylation is thus independent of the acetyl-CoA pathway and may not require vitamin B₁₂ in some and perhaps many incomplete-oxidizing SRB strains.     

Diversity of sulfate-reducing bacteria from an extreme hypersaline sediment, Great Salt Lake (Utah)

The diversity of sulfate-reducing bacteria (SRB) inhabiting the extreme hypersaline sediment (270 g L(-1) NaCl) of the northern arm of Great Salt Lake was studied by integrating cultivation and genotypic identification approaches involving PCR-based retrieval of 16S rRNA and dsrAB genes, the latter encoding major subunits of dissimilatory (bi) sulfite reductase. The majority (85%) of dsrAB sequences retrieved directly from the sediment formed a lineage of high (micro) diversity affiliated with the genus Desulfohalobium, while others represented novel lineages within the families Desulfohalobiaceae and Desulfobacteraceae or among Gram-positive SRB. Using the same sediment, SRB enrichment cultures were established in parallel at 100 and at 190 g L(-1) NaCl using different electron donors. After 5-6 transfers, dsrAB and 16S rRNA gene-based profiling of these enrichment cultures recovered a SRB community composition congruent with the cultivation-independent profiling of the sediment. Pure culture representatives of the predominant Desulfohalobium-related lineage and of one of the Desulfobacteraceae-affilated lineages were successfully obtained. The growth performance of these isolates and of the enrichment cultures suggests that the sediment SRB community of the northern arm of Great Salt Lake consists of moderate halophiles, which are salt-stressed at the in situ salinity of 27%.     

Characterization of bacterial community associated to biofilms of corroded oil pipelines from the southeast of Mexico

Microbial communities associated to biofilms promote corrosion of oil pipelines. The community structure of bacteria in the biofilm formed in oil pipelines is the basic knowledge to understand the complexity and mechanisms of metal corrosion. To assess bacterial diversity, biofilm samples were obtained from X52 steel coupons corroded after 40 days of exposure to normal operation and flow conditions. The biofilm samples were directly used to extract metagenomic DNA, which was used as template to amplify 16S ribosomal gene by PCR. The PCR products of 16S ribosomal gene were also employed as template for sulfate-reducing bacteria (SRB) specific nested-PCR and both PCR products were utilized for the construction of gene libraries. The V3 region of the 16S rRNA gene was also amplified to analyse the bacterial diversity by analysis of denaturing gradient gel electrophoresis (DGGE). Ribosomal library and DGGE profiles exhibited limited bacterial diversity, basically including Citrobacter spp., Enterobacter spp. and Halanaerobium spp. while Desulfovibrio alaskensis and a novel clade within the genus Desulfonatronovibrio were detected from the nested PCR library. The biofilm samples were also taken for the isolation of SRB. Desulfovibrio alaskensis and Desulfovibrio capillatus, as well as some strains related to Citrobacter were isolated. SRB consists in a very small proportion of the community and Desulfovibrio spp. were the relatively abundant groups among the SRB. This is the first study directly exploring bacterial diversity in corrosive biofilms associated to steel pipelines subjected to normal operation conditions.     

Diversity of Sulfate-Reducing Bacteria Inhabiting the Rhizosphere of Phragmites australis in Lake Velencei (Hungary) Revealed by a Combined Cultivation-based and Molecular approach

The community structure of sulfate-reducing bacteria (SRB) associated with reed (Phragmites australis) rhizosphere in Lake Velencei (Hungary) was investigated by using cultivation-based and molecular methods. The cultivation methods were restricted to recover lactate-utilizing species with the exclusion of Desulfobacter and some Desulfobacterium species presumably not being dominant members of the examined community. The most-probable-number (MPN) estimations of lactate-utilizing SRB showed that the cell counts in reed rhizosphere were at least one order of magnitude higher than that in the bulk sediment. The number of endospores was low compared to the total SRB counts. From the highest positive dilution of MPN series, 47 strains were isolated and grouped by restriction fragment length polymorphism (RFLP) analysis of the amplified 16S ribosomal RNA (rRNA) and dsrAB (dissimilatory sulfite reductase) genes. Contrary to the physiological diversity of the isolates, the combined results of RFLP analysis revealed higher diversity at species as well as at subspecies level. Based on the partial 16S rRNA sequences, the representative strains were closely affiliated with the genera Desulfovibrio and Desulfotomaculum. The partial dsrAB sequences of the clones, recovered after isolation and PCR amplification of the community DNA, were related to hitherto uncultured species of the genera Desulfovibrio and Desulfobulbus. Nevertheless, the representative of the second largest clone group was shown to be closely affiliated with the sequenced dsrAB gene of a strain isolated from the same environment and identified as Desulfovibrio alcoholivorans. Another clone sequence was closely related to a possible novel species also isolated within the scope of this work.     

Effects of the antimicrobial tylosin on the microbial community structure of an anaerobic sequencing batch reactor

The effects of the antimicrobial tylosin on a methanogenic microbial community were studied in a glucose‐fed laboratory‐scale anaerobic sequencing batch reactor (ASBR) exposed to stepwise increases of tylosin (0, 1.67, and 167 mg/L). The microbial community structure was determined using quantitative fluorescence in situ hybridization (FISH) and phylogenetic analyses of bacterial 16S ribosomal RNA (rRNA) gene clone libraries of biomass samples. During the periods without tylosin addition and with an influent tylosin concentration of 1.67 mg/L, 16S rRNA gene sequences related to Syntrophobacter were detected and the relative abundance of Methanosaeta species was high. During the highest tylosin dose of 167 mg/L, 16S rRNA gene sequences related to Syntrophobacter species were not detected and the relative abundance of Methanosaeta decreased considerably. Throughout the experimental period, Propionibacteriaceae and high GC Gram‐positive bacteria were present, based on 16S rRNA gene sequences and FISH analyses, respectively. The accumulation of propionate and subsequent reactor failure after long‐term exposure to tylosin are attributed to the direct inhibition of propionate‐oxidizing syntrophic bacteria closely related to Syntrophobacter and the indirect inhibition of Methanosaeta by high propionate concentrations and low pH. Biotechnol. Bioeng. 2011;108: 296–305. © 2010 Wiley Periodicals, Inc.     

High overall diversity and dominance of microdiverse relationships in salt marsh sulphate-reducing bacteria

The biogeochemistry of North Atlantic salt marshes is characterized by the interplay between the marsh grass Spartina and sulphate-reducing bacteria (SRB), which mineralize the diverse carbon substrates provided by the plants. It was hypothesized that SRB populations display high diversity within the sediment as a result of the rich spatial and chemical structuring provided by Spartina roots. A 2000-member 16S rRNA gene library, prepared with delta-proteobacterial SRB-selective primers, was analysed for diversity patterns and phylogenetic relationships. Sequence clustering detected 348 16S rRNA sequence types (ribotypes) related to delta-proteobacterial SRB, and it was estimated that a total of 623 ribotypes were present in the library. Similarity clustering showed that approximately 46% of these sequences fell into groups with < 1% divergence; thus, microheterogeneity accounts for a large portion of the observable genetic diversity. Phylogenetic comparison revealed that sequences most frequently recovered were associated with the Desulfobacteriaceae and Desulfobulbaceae families. Sequences from the Desulfovibrionaceae family were also observed, but were infrequent. Over 80% of the delta-proteobacterial ribotypes clustered with cultured representatives of Desulfosarcina, Desulfococcus and Desulfobacterium genera, suggesting that complete oxidizers with high substrate versatility dominate. The large-scale approach demonstrates the co-existence of numerous SRB-like sequences and reveals an unexpected amount of microdiversity.     

Diversity and origin of Desulfovibrio species: phylogenetic definition of a family

The different nutritional properties of several Desulfovibrio desulfuricans strains suggest that either the strains are misclassified or there is a high degree of phenotypic diversity within the genus Desulfovibrio. The results of partial 16S rRNA and 23S rRNA sequence determinations demonstrated that Desulfovibrio desulfuricans ATCC 27774 and "Desulfovibrio multispirans" are closely related to the type strain (strain Essex 6) and that strains ATCC 7757, Norway 4, and El Agheila Z are not. Therefore, these latter three strains of Desulfovibrio desulfuricans are apparently misclassified. A comparative analysis of nearly complete 16S rRNA sequences in which we used a least-squares analysis method for evolutionary distances, an unweighted pair group method, a signature analysis method, and maximum parsimony was undertaken to further investigate the phylogeny of Desulfovibrio species. The species analyzed were resolved into two branches with origins deep within the delta subdivision of the purple photosynthetic bacteria. One branch contained five deep lineages, which were represented by (i) Desulfovibrio salexigens and Desulfovibrio desulfuricans El Agheila Z; (ii) Desulfovibrio africanus; (iii) Desulfovibrio desulfuricans ATCC 27774, Desulfomonas pigra, and Desulfovibrio vulgaris; (iv) Desulfovibrio gigas; and (v) Desulfomicrobium baculatus (Desulfovibrio baculatus) and Desulfovibrio desulfuricans Norway 4. A correlation between 16S rRNA sequence similarity and percentage of DNA relatedness showed that these five deep lineages are related at levels below the minimum genus level suggested by Johnson (in Bergey's Manual of Systematic Bacteriology, vol. 1, 1984). We propose that this branch should be grouped into a single family, the Desulfovibrionaceae. The other branch includes other genera of sulfate-reducing bacteria (e.g., Desulfobacter and Desulfococcus) and contains Desulfovibrio sapovorans and Desulfovibrio baarsii as separate, distantly related lineages.     

Activity and structure of the sulfate-reducing bacterial community in the sediments of the southern part of Lake Baikal

The rates of sulfate reduction (SR) and the diversity of sulfate-reducing bacteria (SRB) were studied in the sediments of the Posol’skaya Banka elevation in the southern part of Lake Baikal. SR rates varied from 1.2 to 1641 nmol/(dm³ day), with high rates (>600 nmol/(dm³ day)) observed at both deep-water stations and in subsurface silts. Integral SR rates calculated for the uppermost 50 cm of the sediments were higher for gas-saturated and gas hydrate-bearing sediments than in those with low methane content. Enrichment cultures were obtained in Widdel medium for freshwater SRB. Analysis of the 16S rRNA gene fragments from clone libraries obtained from the enrichments revealed the presence of SRB belonged to the genus Desulfosporosinus, with D. lacus as the most closely related member (capable of sulfate, sulfite, and thiosulfate reduction), as well as members of the order Clostridiales.