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Growth media have been developed to facilitate the enrichment and isolation of acidophilic and acid-tolerant sulfate-reducing bacteria (aSRB) from environmental and industrial samples, and to allow their cultivation in vitro The main features of the 'standard' solid and liquid devised media are as follows: (i) use of glycerol rather than an aliphatic acid as electron donor; (ii) inclusion of stoichiometric concentrations of zinc ions to both buffer pH and to convert potentially harmful hydrogen sulphide produced by the aSRB to insoluble zinc sulphide; (iii) inclusion of Acidocella aromatica (an heterotrophic acidophile that does not metabolize glycerol or yeast extract) in the gel underlayer of double layered (overlay) solid media, to remove acetic acid produced by aSRB that incompletely oxidize glycerol and also aliphatic acids (mostly pyruvic) released by acid hydrolysis of the gelling agent used (agarose). Colonies of aSRB are readily distinguished from those of other anaerobes due to their deposition and accumulation of metal sulphide precipitates. Data presented illustrate the effectiveness of the overlay solid media described for isolating aSRB from acidic anaerobic sediments and low pH sulfidogenic bioreactors.

The fates of the two different sulfur atoms of the thiosulfate molecule during anaerobic disproportionation by the sulfate-reducing bacterium Desulfovibrio desulfuricans were followed by isotope mass spectrometry. During disproportionation, 32S-thiosulfate was preferentially metabolized, and the residual thiosulfate became enriched in 34S. The sulfate formed was isotopically heavier than the inner sulfur of the consumed thiosulfate. Vice versa, the sulfide formed was isotopically lighter than the outer sulfur of the consumed thiosulfate. These results indicate that thiosulfate is cleaved to intermediates that undergo further disproportionation to sulfate and sulfide in a second step. These intermediates are probably elemental sulfur and sulfite. It is concluded that disproportionation of thiosulfate, sulfite and elemental sulfur includes a combined pathway.

Sequence data had indicated that cyanobacteria might possess a bidirectional hydrogenase with properties similar to the soluble enzymes from Alcaligenes eutrophus, Nocardia opaca and Desulfovibrio fructosovorans. The present study shows that extracts from the cyanobacterium Anacystis nidulans catalyse NAD(P)H-dependent H2 evolution with low but significant activity and uptake of the gas with NAD(P)+ as the electron acceptor. NAD+ is the preferred electron acceptor and NADH the preferred donor compared to NADP+ and NADPH, respectively. Activity levels of this NAD(P)+dependent, bidirectional hydrogenase are too low to support chemoautotrophic growth in A. nidulans.

Twenty-five isolates of dissimilatory sulfate-reducing bacteria were clustered based on similarity analysis of their phospholipid ester-linked fatty acids (PLFA). Of these, 22 showed that phylogenetic relationships based on the sequence similarity of their 16S rRNA directly paralleled the PLFA relationships. Desulfobacter latus and Desulfobacter curvatus grouped with the other Desulfobacter spp. by 16S rRNA comparison but not with the PLFA analysis as they contained significantly more monoenoic PLFA than the others. Similarly, Desulfovibrio africanus clustered with the Desulfovibrio spp. by 16S rRNA but not with them when analyzed by PLFA patterns because of higher monoenoic PLFA content. Otherwise, clustering obtained with either analysis was essentially congruent. The relationships defined by PLFA patterns appeared robust to shifts in nutrients and terminal electron acceptors. Additional analyses utilizing the lipopolysaccharide-lipid A hydroxy fatty acid patterns appeared not to shift the relationships based on PLFA significantly except when completely absent, as in Gram-positive bacteria. Phylogenetic relationships between isolates defined by 16S rRNA sequence divergence represent a selection clearly different from the multi-enzyme activities responsible for the PLFA patterns. Determination of bacterial relationships based on different selective pressures for various cellular components provides more clues to evolutionary history leading to a more rational nomenclature.

A mesophilic, dehalogenating, sulfate-reducing diculture was isolated from an anaerobic lake sediment. One strain of the diculture is proposed to be an endospore-forming Desulfotomaculum species, the second strain was a vibrioid, motile and non-sporeforming species which is tentatively assigned to the genus Desulfovibrio. The diculture was able to mineralize 4- and 2-fluorobenzoate both isomers being incompletely oxidized with the release of acetate, which was subsequently used by both sulfate-reducing strains. Other electron donors used for growth included benzoate, 3- and 4-hydroxybenzoate, protocatechuate, catechol, phenol, 2,5-dimethoxyphenol, fatty acids up to C8, malate and pyruvate. The culture obtained from a freshwater habitat grew optimally at NaCl concentrations of 0.3-0.5 g l-1, 33-37 degrees C, and pH 7.4. Our experiments showed that certain fluorinated aromatic hydrocarbons could serve as sole sources of carbon and energy for sulfate-reducing bacteria.

Viable counts of sulphate-reducing bacteria, able to use a range of different growth substrates were determined in sediments from two Sea Lochs (Etive and Eil) and an estuarine site (Tay), in Scotland. The composition of the sulphate-reducing bacterial population, in terms of substrate utilization, broadly corresponded to the in situ substrates for sulphate reduction and concentration of substrates at each site. Addition of acetate, lactate, propionate, butyrate, hydrogen and glutamate/serine (20 mM) to replicate slurries from each site resulted in stimulation of the corresponding population of sulphate-reducing bacteria and the in situ rates of sulphate reduction. The metabolism of the added substrates and changes in bacterial phospholipid fatty acids (PLFA) were quantified. With the exception of acetate and hydrogen, added substrates were incompletely oxidised, producing a mixture of further substrates, which predominantly were sequentially oxidised, and resulted in the stimulation of a mixed population of sulphate-reducing bacteria. There were significant changes in the PLFA of slurries with added substrate compared to controls. Acetate was completely removed at all sites and the small increase in even chain PLFA together with the absence of stimulation of any other biomarker, indicated that acetate was oxidised by sulphate-reducing bacteria distinctly different from those using other substrates. A biomarker for Desulfobacter, 10 Methyl 16:0, was not stimulated in any of the acetate slurries or in slurries where acetate was produced. Biomarkers for the propionate utilizing Desulfobulbus sp (17:1w6, 15:1w6) were always stimulated in propionate slurries and also in lactate slurries, where partial lactate fermentation produced propionate and acetate. In lactate and glutamate / serine slurries from the Tay estuary and lactate and hydrogen slurries from Loch Etive the biomarker for Desulfovibrio sp (i17:1w7) as well as those for Desulfobulbus were stimulated. This provides direct evidence for the significance of Desulfovibrio sp. within sediment slurries and demonstrates the competitive interaction between members of this genus and Desulfobulbus sp. for lactate, hydrogen and amino acid metabolism. At the estuarine site, sulphate reduction was limited at higher sulphate concentrations (about 3.5 mM) than the Sea Loch sites (

During fermentation in the human large intestine, terminal oxidative processes may involve the activities of dissimilatory sulphate-reducing bacteria (SRB). Approximately 50% of healthy individuals harbour significant populations of SRB in faeces. In mixed culture, growth of SRB in vitro was modulated by sulphate availability, with sulphated polysaccharides such as mucin, chondroitin sulphate and carrageenan causing increased growth rates and sulphide production when compared with starch, pectin and arabino-galactan. Rates of H2S production were higher among SRB isolated from patients with ulccrative colitis in contrast to those present in healthy volunteers. The majority (up to 92%) of SRB in faecal samples belonged to the genus Desulfovibrio. In vitro studies demonstrated that compared to isolates from healthy subjects. Desulfovibrio desulfuricans from colitic individuals were better able to adapt to high dilution rates, which may be associated with the disease. These findings indicate that the metabolic capabilities of SRB isolated from the human large intestine are not uniform and may respond to the type of substrate available in the gut as well as the rate of passage of digesta.

In a mineral medium containing sulfate as terminal electron acceptor, the sulfate-reducing bacterium Desulfovibrio alcoholovorans oxidized stoichiometrically 1 mol glycerol to 1 mol acetate and 1 mol 1,3-propanediol to 1 mol acetate with the concomitant reduction of 0.75 and 1 mol sulfate, respectively; 1 mol 1,2-propanediol was degraded to 0.8 mol acetate and 0.1 mol proprionate, with the reduction of approximately 1 mol sulfate. The maximum specific growth rates (μmax in h−1) were 0.22, 0.086 and 0.09 with glycerol, 1,3-propanediol and 1,2-propanediol, respectively. The growth yields were 12.7 g, 11.1 g and 7.2 g dry weight/mol 1,3-propanediol, glycerol and 1,2-propanediol degraded, respectively. The growth yields and maximum specific growth rates of the H2-transferring associations were also calculated. In the absense of sulfate, all these reduced substrates were degraded to acids and methane when D. alcoholovorans was cocultured with Methanospirillum hungatei. Changes in the metabolic pathway were observed in the degradation of 1,2- and 1,3-propanediol. The metabolic efficiency of D. alcoholovorans to degrade glycerol, 1.2- and 1,3-propanediol is discussed.

Microhabitats and survival of sulfate-reducing bacteria (SRB) in an oxic surface sediment of a seawater lake were examined. The size of fractionation of the sediment suspension showed that most of SRB were associated with sediment particles larger than 10 μm. The D values (time in h required to destroy 90% of the initial viable population) for SRB in the whole sediment suspension and for SRB i n the μ m and the μ m fractions were, respectively, 23.7, 10 and 4 when the SRB were exposed to air. Survival of the FeS-associated Desulfovibrio desulfuricans ( D value, 9.3) was higher than that of the free-living ones ( D value, 1.8). These results show that particle-associated SRB are more protected against oxygen than free-living ones in oxic sediments.

Free-living cells produced colonies more rapidly than particle-associated ones when a pure culture of Desulfovibrio desulfuricans containing sediment particles was inoculated on an agar medium. The time required for the appearance of the first colony, tr of the first order reation model [4] was 53 h for free-living cells and 112 h for particle-associated ones. The value of tr may be determined from the growth rate and sulfate reduction activity because the activity of free-living cells was much higher than that of aggregating ones.

Faecal microbial communities in South African Windsnyer-type indigenous pigs (SAWIPs) and Large White × Landrace (LW × LR) crosses were investigated using high-throughput sequencing of the 16S rDNA genes. The faecal microbial communities in LW × LR crosses and SAWIPs fed control (CON) and high maize cob (HMC) diets were evaluated through parallel sequencing of 16S rDNA genes. Butrivibrio, Faecalibacterium and Desulfovibrio, although present in LW × LR pigs, were absent from the SAWIP microbial community. Bacteroides, Succiniclasticum, Peptococcus and Akkermansia were found in SAWIPs but not in LW × LR crosses. The ratios of Bacteroidia to Clostridia on the CON and HMC diets were similar (0.37 versus 0.39) in SAWIPs but different (0.24 versus 0.1) in LW × LR crosses. The faecal microbial profiles determined were different between the LW × LR and SAWIP breeds but not between pigs fed the CON and HMC diets. The composition of faecal bacterial communities in SAWIPs was determined for the first time. The differences in microbial communities detected may explain the enhanced ability of SAWIPs to digest fibrous diets compared with the LW × LR crosses.

Fifteen (soil and intestinal) strains of Desulfovibrio desulfuricans species were typed by PCR method with the use of primers specific for repetitive extragenic palindromic (REP) and enterobacterial repetitive intergenic consensus (ERIC) sequences. As a result, characteristic DNA fingerprints for the strains were obtained. Moreover, the genetic profiles were found to be useful for typing and distinguishing the strains of D. desulfuricans. According to cluster analysis, PCR with primers complementary to the sequences REP appeared to be slightly more discriminatory than PCR with ERIC primers for the investigated strains. Distinct fingerprint patterns of two isolates derived from the same patient pointed to the different origin of both strains.

The genes encoding the desulforubidin type of dissimilatory sulfite reductase (Dsr) from the sulfate-reducing bacteria Desulfobacter vibrioformis and Desulfobulbus rhabdoformis were cloned and sequenced. Similar to the genes for dissimilatory sulfite reductase from the genera Archaeoglobus, Desulfovibrio and Desulfotomaculum the dsr genes were found to form an operon, dsrABD, where dsrA and dsrB encode the structural subunits, alpha and beta, of Dsr, respectively. dsrD encodes a conserved unknown protein apparently restricted to sulfate-reducing species. In Desulfobacter vibrioformis a fourth gene, designated dsrN, was found downstream of dsrD forming a contiguous operon, dsrABDN. DsrN showed significant sequence homology to cobyrinic a,c-diamide synthase, which is involved in the biosynthesis of vitamin B12. A function for DsrN in amidation of siroheme is likely. Analysis of the dsrAB-encoded proteins confirmed that the high conservation observed for other types of dissimilatory sulfite reductase is also found in desulforubidin. The use of Dsr sequences in unravelling the phylogeny of sulfate-reducing bacteria is discussed.

Methane hydrates represent an enormous carbon and energy source in many low temperature deep marine sediments. However, little information is available concerning the nature of the microbial communities associated with these structures. Here, we describe a phylogenetic analysis based on ribosomal DNA (rDNA) sequences obtained from sediment and fluid samples present in a region of gas hydrate formation in shallow sediments within the Cascadia margin in and around Ocean Drilling Program (ODP) Site 892B. Our studies detected diverse sulfur-utilizing microbes, methanogens, methanotrophs, and non-thermophilic members of the kingdom Crenarchaeota. This is the first culture-independent phylogenetic analysis of a gas hydrate habitat.

The genomes of the phylogenetically and physiologically unique bacteria Desulfurella acetivorans DSM 5264T and D. multipotens DSM 8415T were characterized and compared by pulsed field gel electrophoresis (PFGE). Macrorestriction patterns made of large PFGE separated DNA fragments were generated by digesting the genomic DNAs of both strains with the rare cutting restriction endonucleases ApaI, AscI, EagI, RsrII, SacII, SalI as well as with the intron encoded endonuclease I-CeuI. The sum of calculated fragment sizes from digests of the first six enzymes yielded estimates for the chromosome sizes of D. acetivorans with a mean of 1939.0 +/- 26.0 kb and for D. multipotens with a mean of 1864.0 +/- 23.0 kb. Within the patterns obtained from EagI and RsrII cleavages the apparent differences could be attributed to DNA insertion or deletion and to point mutation. The single, circular chromosomes of the two strains contain two copies of 23S rRNA genes each. Different extrachromosomal elements were detected in both strains.

A new mesophilic, thermotolerant sulfate-reducing bacterium, was isolated from the flowing bore waters of a deep aquifer, the Great Artesian Basin, Australia. The strain, designated isolate AB16910a, is a curved rod and resembled members of the genus Desulfovibrio. However, the isolate can be differentiated from other members of the Desulfovibrio species because of the high G+C content of 69 +/- 0.25% the 16S rRNA sequence data and other physiological characteristics. The name Desulfovibrio longreachii is proposed for the new isolate.

Downstream of the genes for the structural alpha and beta subunits of the periplasmic Desulfovibrio vulgaris (Hildenborough) hydrogenase a DNA fragment was detected with sequence homology to these genes. This fragment was cloned in Escherichia coli and the nucleotide sequence was determined. A gene was detected on the fragment with coding capacity for a 65.8 kDa polypeptide, hyd gamma. The central part of hyd gamma has an unusually high degree of homology with the alpha subunit and the C-terminal part has similarity with the beta subunit. These results strongly suggest that the three genes for hyd gamma and the alpha and beta subunits derive from one common ancestor gene. We succeeded in the identification of the translational product of this gene in E. coli, but were unable to determine the function of hyd gamma after expression in E. coli.

In cell suspensions of the marine sulfate-reducing bacterium Desulfovibrio 20020 (DSM 3099) permeabilized with formaldehyde or Triton X-100, sulfite-dependent NADH oxidation activities of 0.05 μmol · min −1 · mg −1 protein were detected. NADH oxidation coupled to APS, thiosulfate and fumarate reduction was also demonstrated. All the activities were subject to inhibition by HOQNO and antimycin A. The rate of NADH oxidation coupled to the reduction of sulfite was extremely low in cell-free extracts. The physiological function and possible mechanism of the NADH oxidation coupled to the reduction of various electron acceptors are discussed.

A multi-stage bi-directional chemostat system has been developed in which solutes but not cells are allowed to diffuse between the individual growth chambers which are separated by 0.2 micron pore size polyvinyledene difluoride membranes. The experimental system enables the generation of physico-chemical gradients which, together with the spatial separation of the individual microbial processes, provides a useful laboratory model to study microbial interactions. This paper describes the construction of a multi-stage diffusion chemostat and its application in studying carbon flow in anaerobic estuarine sediments. Populations of Clostridium butyricum, Desulfovibrio desulfuricans and Chromatium vinosum were grown in the compound diffusion chemostat at a dilution rate of 0.03 h−1 at 25°C, and the effects of inorganic nitrogen source and availability on carbon flow and individual cell populations were determined. C. butyricum and D. desulfuricans both used NO−3 as an e− acceptor with an increase in cell numbers. Under these growth conditions, free S2− concentrations were lower, resulting in more stable cell populations than in comparable cultures grown on NH+4 as nitrogen source.

The oxidation of protoporphyrinogen by membranes from Desulfovibrio gigas with nitrite as the electron acceptor proceeds at the ratio of 1 mol protoporphyrin produced for each mol of nitrite reduced. Coupled to the protoporphyrinogen-nitrite reaction is the esterification of ortho-phosphate with a P/2e− value of 0.92. Pentachlorophenol, 3 × 10−5 M, and carbonyl cyanide m-chlorophenyl hydrazone, 3 × 10−6 M, serve as uncouplers of phosphorylation while rotenone, 9 × 10−6 M, and hydroxyquinoline-N-oxide, 0.1 × 10−6 M, inhibit electron flow from protoporphyrinogen oxidase to nitrite reductase.

An unusual sulphate-reducing bacterium was isolated from Lake Fryxell, Antarctica. Designated strain FESu, it illustrates the difficulty in assigning some isolates of Desulfovibrio spp. to a described species. FESu was characterized by its relatively low mol% G + C (45%), its rod-shaped morphology, inability to grow on lactate, pyruvate, malate or choline in the absence of sulphate, and by its lack of desulfoviridin.

Permeabilized cells of Desulfovibrio baarsii catalyzed an isotopic exchange between 14CO and the carboxyl group of acetyl-CoA and an isotopic exchange between 14CO2 and the carboxyl group of acetyl-CoA. Cyanide, which inactivated the carbon monoxide dehydrogenase activity present in the cells, inhibited the 14CO2/acetyl-CoA rather than the 14CO/acetyl-CoA exchange reaction.

The reduction of tetrazolium salts by the sulfate-reducing bacteria, Desulfovibrio desulfuricans and Desulfotomaculum orientis, was examined. D. desulfuricans and D. orientis reduced triphenyltetrazolium chloride (TTC) and 2-(p-iodophenyl)-3-(p-nitrophenyl)-5-phenyltetrazolium chloride (INT) forming intracellular formazan deposits. The reduction rate of INT was higher than that of TTC. INT reduction was not inhibited by the addition of sulfate or molybdate, and sulfate uptake was inhibited by the addition of both INT and molybdate. The ratio of intracellular formazan forming cells to acridine orange direct counts in both strains decreased with culture age and starvation time.

The broad-host-range, IncQ, plasmid R300B (Sm, Su) has been stably transferred to two strains of sulphate-reducing bacteria (Desulfovibrio sp. 8301 and Desulfovibrio desulfuricans 8312), using the IncP1 transfer system of the helper plasmid pRK2013 and cocultivation of sulphate-reducing bacteria with facultative anaerobes in media provided with sulphate and nitrate ions as electron acceptors. R300B was transferred at a frequency of 10−2 to 1 per acceptor cell. The SmR marker was expressed in both sulphate-reducing bacteria strains while the SuR was expressed only in strain 8301. R300B can also be transferred back to E. coli strains provided with IncP1 plasmids taking advantage of the retrotransfer ability of these plasmids. This occurs at a frequency up to 10−4 by recipient E. coli cell.

In order to validate unusual fatty acids as biomarkers for sulphate-reducing bacteria, selective conditions were arranged for the enrichment of a marine glutamate-fermenting bacterium which made hydrogen and acetate available for oxidation via the respiration of sulphate. Under these conditions the complete oxidation of glutamate via sulphate reduction accounted for 84% of the available electron equivalents. Fatty acid biomarkers for hydrogen-oxidizing Desulfovibrio sp. (iso 17:1w7c and branched monoenoics) and for acetate-oxidizing Desulfobacter (10 methyl 16:0) were detected in the enrichment. These biomarkers were demonstrated in pure cultures of Desulfovibrio sp. and Desulfobacter sp. obtained from the enrichment. The predominant glutamate-fermenting bacterium isolated from the consortium contained no branched ester-linked phospholipid fatty acids, and produced acetate and hydrogen. With energy limitation the enriched consortium produced increased amounts of extracellular polysaccharide and the endogenous storage lipid poly-beta-hydroxybutyrate as detected with Fourier transform/infra-red (FT-IR) spectroscopy.

Genomic DNA from 13 diazotrophic strains of Desulfovibrio showed homology to structural nif DNA from Klebsiella pneumoniae; DNA from 3 non-diazotrophic strains did not. The nif DNA is chromosomal in 10 strains, but is carried on 130-MDa plasmids in 3 strains of Desulfovibrio vulgaris.

Lactate is the most commonly employed energy substrate for the growth of sulfate-reducing bacteria [1], except for some of the recently discovered new types [2]. Both isomeric forms of lactate are utilized and metabolized to acetate and CO 2. Very little is known about the first enzyme involved in the oxidation of lactate. An N A D + independent membrane-bound L(+) lactate hydrogenase (LDH) activity has been reported in extracts of Desulfovibrio gigas but further details are not available [3]. D(--) L D H was recently detected in Desulfovibrio desulfuricans [4] and Desulfovibrio vulgaris [5]. This enzyme was partially purified and retained its activity without special precautions. Both D(--) LDHs were membranebound, NAD(P)+independent and could be assayed with 2,6-dichlorophenolindophenol (DCPIP) as an artificial electron acceptor. The identity of the in vivo elec'tron acceptors of the LDHs remains to be established with certainty; the D(--) L D H of D. vulgaris could be coupled to cytochrome c553 which is also thought to be the acceptor of the formate dehydrogenase [5]. In this paper we report the presence of high activities of a membrane bound NAD(P) + independent L( + ) L D H in a D. desulfuricans strain. This enzyme was fairly stable when kept anaerobically but extremely unstable in the presence of oxygen. Data on the conditions required for reactivation of oxygen-inactivated enzyme activity in extracts are given. Only very low activities of D(--) L D H were detected despite the fact that our strain grows well oh both L (+ ) and D ) lactate.

The metabolism of methanol by acidogenic bacteria (Butyribacterium methylotrophicum, Sporomusa ovata and Acetobacterium woodii) was studied in pure culture and in defined mixed cultures with sulfate-reducing bacteria (Desulfovibrio vulgaris) or methanogenic bacteria (Methanobrevibacter arboriphilus strain AZ). In the mixed cultures, less acids (acetate and/or butyrate) were formed per unit methanol converted than in pure cultures. In these mixed cultures, a significant production of sulfide or methane was observed despite the inability of the sulfate reducer and the methanogen to use methanol as an energy substrate. These results are explained in terms of interspecies hydrogen transfer between the acidogens (converting part of the methanol to 1 CO2 and 3 H2) and the Desulfovibrio or Methanobrevibacter species. The bioenergetic aspects of this process and its ecological implications are discussed.

Three types of hydrogenases have been isolated from the sulfate-reducing bacteria of the genus Desulfovibrio. They differ in their subunit and metal compositions, physico-chemical characteristics, amino acid sequences, immunological reactivities, gene structures and their catalytic properties. Broadly, the hydrogenases can be considered as 'iron only' hydrogenases and nickel-containing hydrogenases. The iron-sulfur-containing hydrogenase ([Fe] hydrogenase) contains two ferredoxin-type (4Fe-4S) clusters and an atypical iron-sulfur center believed to be involved in the activation of H2. The [Fe] hydrogenase has the highest specific activity in the evolution and consumption of hydrogen and in the proton-deuterium exchange reaction and this enzyme is the most sensitive to CO and NO2-. It is not present in all species of Desulfovibrio. The nickel-(iron-sulfur)-containing hydrogenases [( NiFe] hydrogenases) possess two (4Fe-4S) centers and one (3Fe-xS) cluster in addition to nickel and have been found in all species of Desulfovibrio so far investigated. The redox active nickel is ligated by at least two cysteinyl thiolate residues and the [NiFe] hydrogenases are particularly resistant to inhibitors such as CO and NO2-. The genes encoding the large and small subunits of a periplasmic and a membrane-bound species of the [NiFe] hydrogenase have been cloned in Escherichia (E.) coli and sequenced. Their derived amino acid sequences exhibit a high degree of homology (70%); however, they show no obvious metal-binding sites or homology with the derived amino acid sequence of the [Fe] hydrogenase. The third class is represented by the nickel-(iron-sulfur)-selenium-containing hydrogenases [( NiFe-Se] hydrogenases) which contain nickel and selenium in equimolecular amounts plus (4Fe-4S) centers and are only found in some species of Desulfovibrio. The genes encoding the large and small subunits of the periplasmic hydrogenase from Desulfovibrio (D.) baculatus (DSM 1743) have been cloned in E. coli and sequenced. The derived amino acid sequence exhibits homology (40%) with the sequence of the [NiFe] hydrogenase and the carboxy-terminus of the gene for the large subunit contains a codon (TGA) for selenocysteine in a position homologous to a codon (TGC) for cysteine in the large subunit of the [NiFe] hydrogenase. EXAFS and EPR studies with the 77Se-enriched D. baculatus hydrogenase indicate that selenium is a ligand to nickel and suggest that the redox active nickel is ligated by at least two cysteinyl thiolate and one selenocysteine selenolate residues.(ABSTRACT TRUNCATED AT 400 WORDS)

Summary Two marine strains of Desulfovibrio were able to use alanine, serine and glycine as good growth substrates; aspartate, cysteine, threonine and branched-chain amino acids supported only slow growth.

Data are presented that all known periplasmic redox proteins from the sulfate reducing bacteria included in the genus, Desulfovibrio have aminoterminal (N-terminal) amino-acid sequences commonly found in other Gram-negative bacteria and are indicative of recognition sites for signal peptides. In contrast, none of the cytoplasmic redox proteins exhibited these unique N-terminal amino-acid sequences. It is proposed that the N-terminal amino-acid residues of a given protein can be used as an indicator of its cellular localization within the bacterial cell.

During mesophilic methanogenesis of sugars, lactate and ethanol are commonly found as intermediates produced by hydrolytic and fermentative bacteria [1]. Under anaerobiosis, various bacterial species utilize ethanol as sole energy source for their growth concurrently with volatile fatty acid production. According to this production, several patterns of fermentation were distinguished inc!uding (a) the production of butyric, valeric and caproic acids which is characteristic of Clostridium kluyveri. This micro-organism utilizes ethanol and condenses it with short volatile fatty acids (C 2 to C4) into acids with two more carbons [2]; (b) the ethanol oxidation to acetate with concurrent reduction of CO 2 to methane which was first described by Barker [3]. The responsible microorganism, Methanobaeillus omelianskii , has later been shown to be a syntrophic association of two strains [4]: the 'S organism' which oxidizes ethanol to acetate and reduces protons to H 2, and a H 2utilizing methanogenic bacter ium Methanobacterium strain Moll , i.e. Methanobacterium bryantii. Growth of the acetogenic bacterium was inhibited by low partial pressure of hydrogen and occurred only when an H 2 utilizer was present. In low sulfate media, hydrogenase-forming strains of Desulfovibrio or Desulfotomaculum performed the same type of syntrophic association with methanogens [5]. In this report, we provide the first documentation for the fermentation of ethanol into propionate and acetate and show that the responsible anaerobe requires a growth factor produced by a H2-oxidizing species. These findings were presented in preliminary form at the 2nd International Symposium on Anaerobic Digestion (poster session, Travemunde, Germany, Sept. 81).

The Desulfovibrio gigas nifH gene has been cloned and sequenced. It consists of an open-reading frame of 822 base pairs encoding a 274 amino acid polypeptide. A potential ntrA-dependent promotor sequence is present. The gene lacks an upstream activator sequence homologous to those often found in nif genes subject to activation by nifA.