Microbial sulphate reduction at a low pH

It is now well established that microbial sulphate-reduction can proceed in environments with a pH<5. This review summarizes existing reports on sulphate reduction at low pH and discusses possible pH effects on sulphate-reducing bacteria. Microbial sulphate reduction has been observed in acidic lakes, wetlands, mesocosms, acidic sulphate soils and bioreactors. Possible inhibitory factors include the metabolites H(2)S and organic acids, which can be toxic depending on pH. Metal sulphide precipitation and competition with other bacteria, namely iron-reducing bacteria, can inhibit sulphate reduction. Theoretical considerations show that normal sulphate reduction rates are too low to maintain a neutral micro niche in an acidic environment. The first acidotolerant sulphate-reducing bacteria have been isolated recently.     

Molecular analysis of a sulphate-reducing consortium used to treat metal-containing effluents

A sulphate-reducing consortium used in a bioprocess to remove toxic metals from solution as insoluble sulphides, was characterised using molecular (PCR-based) and traditional culturing techniques. After prolonged cultivation under anoxic biofilm-forming conditions, the mixed culture contained a low diversity of sulphate-reducing bacteria, dominated by one strain closely related to Desulfomicrobium norvegicum, identified by three independent PCR-based analyses. The genetic targets used were the 16S rRNA gene, the 16S-23S rRNA gene intergenic spacer region and the disulfite reductase (dsr) gene, which is conserved amongst all known sulphate-reducing bacteria. This organism was also isolated by conventional anaerobic techniques, confirming its presence in the mixed culture. A surprising diversity of other non-sulphate-reducing facultative and obligate anaerobes were detected, supporting a model of the symbiotic/commensal nature of carbon and energy fluxes in such a mixed culture while suggesting the physiological capacity for a wide range of biotransformations by this stable microbial consortium.     

Distribution of sulphur-cycle bacteria in the Ems-Dollard estuary

Summary A quantitative study of the sulphur cycle in the tidal flat-sediments of the Eems-Dollard estuary was started by determining the distribution of two physiologically different groups of bacteria: sulphate-reducing and sulphide-oxidizing bacteria. Viable counts of these bacterial groups were determined by most probable number techniques.The highest numbers of aerobic sulphide-oxidizing bacteria were found in the upper 2 cm of the sediment. A rapid decrease was observed with increasing depth.The anaerobic sulphate-reducing bacteria showed different distribution-patterns with depth. Frequently high numbers of these bacteria were found above the redox-discontinuity-layer. This may be attributed to the presence of anaerobic micro-pockets in this largely aerobic top-layer of the sediment.The horizontal distribution of the sulphide-oxidizing bacteria appeared to be highly correlated with sediment parameters such as organic carbon and clay content of the sediment. The sulphate-reducing bacteria showed only a small linear correlation with these parameters.By means of polyfactor-analysis mathematical models were made with bacterial numbers as the dependent variables and with some environmental parameters as independent variables. The parameters used in this models could explain the variance of the viable counts for approximately 70%. The clay content of the sediment and the number of sulphate reducing bacteria appeared to determine to a large extent the variance in numbers of sulphide-oxidizing bacteria. There are indications that a great deal of the sulphide-oxidizing bacteria might be mixotrophic.For the explanation of the variance in numbers of sulphate-reducing bacteria the most important parameters were the clay content of the sediment, the number of aerobic heterotrophic bacteria and temperature (or season). Therefore the numbers of these organisms were varying throughout the year. It is assumed that the heterotrophic bacteria supply the sulphate-reducing bacteria with organic substrates.     

Aluminum and sulphate removal by a highly Al-resistant dissimilatory sulphate-reducing bacteria community

A highly Al-resistant dissimilatory sulphate-reducing bacteria community was isolated from sludge of the wetland of Urgeiri?a mine (community W). This community showed excellent sulphate removal at the presence of Al3?. After 27 days of incubation, 73, 86 and 81% of sulphate was removed in the presence of 0.48, 0.90 and 1.30 mM of Al3?, respectively. Moreover, Al3? was simultaneously removed: 55, 85 and 78% of metal was removed in the presence of 0.48, 0.90 and 1.30 mM of Al3?, respectively. The dissociation of aluminium-lactate soluble complexes due to lactate consumption by dissimilatory sulphate-reducing bacteria can be responsible for aluminum removal, which probably precipitates as insoluble aluminium hydroxide. Phylogenetic analysis of 16S rRNA gene showed that this community was mainly composed by bacteria closely related to Desulfovibrio desulfuricans. However, bacteria affiliated to Proteus and Ralstonia were also present in the community.     

Biological sulphate reduction using food industry wastes as carbon sources

Biological treatment with dissimilatory sulphate-reducing bacteria has been considered the most promising alternative for decontamination of sulphate rich effluents. These wastewaters are usually deficient in electron donors and require their external addition to achieve complete sulphate reduction. The aim of the present study was to investigate the possibility of using food industry wastes (a waste from the wine industry and cheese whey) as carbon sources for dissimilatory sulphate-reducing bacteria. The results show that these wastes can be efficiently used by these bacteria provided that calcite tailing is present as a neutralizing and buffer material. A 95 and 50 % sulphate reduction was achieved within 20 days of experiment by a consortium of dissimilatory sulphate-reducing bacteria grown on media containing waste from the wine industry or cheese whey respectively. Identification of the dissimilatory sulphate-reducing bacteria community using the dsr gene revealed the presence of the species Desulfovibrio fructosovorans, Desulfovibrio aminophilus and Desulfovibrio desulfuricans. The findings of the present study emphasise the potential of using wastes from the wine industry as carbon source for dissimilatory sulphate-reducing bacteria, combined with calcite tailing, in the development of cost effective and environmentally friendly bioremediation processes.     

The effect of pressure and temperature on sulphate-reducing bacteria and the action of biocides in oilfield water injection systems

The effects of elevated pressures and temperatures on the growth, morphology and metabolic activity of sulphate-reducing bacteria, isolated from the North Sea, are described. Pressure/temperature profiles, growth curves and sulphate reduction rates are presented for several isolates. The maximum pressure and temperature that supported growth were 65 000 KPa and 45°C respectively. The results are discussed in connection with water injection into oil-bearing reservoirs where there is a concern that generation of hydrogen sulphide by sulphate-reducing bacteria may lead to increased hydrogen sulphide levels (souring) in oil and gas, and to corrosion problems in production facilities. The bacteriostatic effects of a number of commercial biocides were enhanced at elevated hydrostatic pressures and temperatures.     

Anaerobic degradation of naphthalene by a pure culture of a novel type of marine sulphate-reducing bacterium

Incubation of marine sediment in anoxic, sulphate-rich medium in the presence of naphthalene resulted in the enrichment of sulphate-reducing bacteria. Pure cultures with short, oval cells (1.3 by 1.3–1.9 μm) were isolated that grew with naphthalene as the only organic carbon source and electron donor for sulphate reduction to sulphide. One strain, NaphS2, was characterized. It affiliated with completely oxidizing sulphate-reducing bacteria of the δ-subclass of the Proteobacteria, as revealed by 16S rRNA sequence analysis. 2-Naphthoate, benzoate, pyruvate and acetate were used in addition to naphthalene. Quantification of substrate consumption, sulphide formation and formed cell mass revealed that naphthalene was completely oxidized with sulphate as the electron acceptor.     

Immunofluorescence of sulphate-reducing bacteria

An indirect fluorescent antibody technique was used as a method of rapidly assessing and identifying sulphate-reducing bacteria. Five specific antisera and one polyvalent serum were raised and tested against 44 strains of the genera Desulfovibrio and Desulfotomaculum along with 4 control organisms. Immunofluorescence was found to be mainly strain specific with the sulphate-reducing bacteria although weak fluorescence was seen both within and between recognised groups. A polyvalent antiserum was successfully used to detect sulphate-reducing bacteria. No interference from 4 control organisms was found.     

Related assemblages of sulphate-reducing bacteria associated with ultradeep gold mines of South Africa and deep basalt aquifers of Washington State

We characterized the diversity of sulphate-reducing bacteria (SRB) associated with South African gold mine boreholes and deep aquifer systems in Washington State, USA. Sterile cartridges filled with crushed country rock were installed on two hydrologically isolated and chemically distinct sites at depths of 3.2 and 2.7 km below the land surface (kmbls) to allow development of biofilms. Enrichments of sulphate-reducing chemolithotrophic (H2) and organotrophic (lactate) bacteria were established from each site under both meso- and thermophilic conditions. Dissimilatory sulphite reductase (Dsr) and 16S ribosomal RNA (rRNA) genes amplified from DNA extracted from the cartridges were most closely related to the Gram-positive species Desulfotomaculum thermosapovorans and Desulfotomaculum geothermicum, or affiliated with a novel deeply branching clade. The dsr sequences recovered from the Washington State deep aquifer systems affiliated closely with the South African sequences, suggesting that Gram-positive sulphate-reducing bacteria are widely distributed in the deep subsurface.     

Transfer of broad host-range plasmids to sulphate-reducing bacteria

Abstract 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 Sm^R marker was expressed in both sulphate-reducing bacteria strains while the Su^R 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.     

A rapid method for determination of viable sulphate-reducing bacteria in human faeces

Adenosine phosphosulphate reductase (APS reductase) (E.C.1.8.99.2), viable counts of sulphate-reducing bacteria and rates of sulphate reduction were determined in 20 human samples of faeces. The activity of APS reductase, in contrast to sulphate reduction rates, correlated well with viable counts ( r = 0.987) and may therefore be used rapidly to quantify sulphate-reducing bacteria in human gut contents.     

Translation termination factor aRF1 from the archaeon Methanococcus jannaschii is active with eukaryotic ribosomes

A bacterioferritin was recently isolated from the anaerobic sulphate-reducing bacterium Desulfovibrio desulfuricans ATCC 27774 [Romão et al. (2000) Biochemistry 39, 6841–6849]. Although its properties are in general similar to those of the other bacterioferritins, it contains a haem quite distinct from the haem B, found in bacterioferritins from aerobic organisms. Using visible and NMR spectroscopies, as well as mass spectrometry analysis, the haem is now unambiguously identified as iron-coproporphyrin III, the first example of such a prosthetic group in a biological system. This unexpected finding is discussed in the framework of haem biosynthetic pathways in anaerobes and particularly in sulphate-reducing bacteria.     

Iron-coproporphyrin III is a natural cofactor in bacterioferritin from the anaerobic bacterium Desulfovibrio desulfuricans

A bacterioferritin was recently isolated from the anaerobic sulphate-reducing bacterium Desulfovibrio desulfuricans ATCC 27774 [Romão et al. (2000) Biochemistry 39, 6841–6849]. Although its properties are in general similar to those of the other bacterioferritins, it contains a haem quite distinct from the haem B, found in bacterioferritins from aerobic organisms. Using visible and NMR spectroscopies, as well as mass spectrometry analysis, the haem is now unambiguously identified as iron-coproporphyrin III, the first example of such a prosthetic group in a biological system. This unexpected finding is discussed in the framework of haem biosynthetic pathways in anaerobes and particularly in sulphate-reducing bacteria.     

Characterization of sulphate-reducing bacterial populations within marine and estuarine sediments with different rates of sulphate reduction

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 (<2 mM) and this had a significant effect on propionate and butyrate metabolism, as well as on methane production. These results demonstrate that although the sulphate-reducing bacterial population at each site could metabolise identical substrates, the types of sulphate-reducing bacteria involved and their sulphate thresholds were characteristically different.     

Effect of temperature on sulphate reduction, growth rate and growth yield in five psychrophilic sulphate-reducing bacteria from Arctic sediments

Five psychrophilic sulphate-reducing bacteria (strains ASv26, LSv21, PSv29, LSv54 and LSv514) isolated from Arctic sediments were examined for their adaptation to permanently low temperatures. All strains grew at −1.8°C, the freezing point of sea water, but their optimum temperature for growth ( T _opt) were 7°C (PSv29), 10°C (ASv26, LSv54) and 18°C (LSv21, LSv514). Although T _opt was considerably above the in situ temperatures of their habitats (−1.7°C and 2.6°C), relative growth rates were still high at 0°C, accounting for 25–41% of those at T _opt. Short-term incubations of exponentially growing cultures showed that the highest sulphate reduction rates occurred 2–9°C above T _opt. In contrast to growth and sulphate reduction rates, growth yields of strains ASv26, LSv54 and PSv29 were almost constant between −1.8°C and T _opt. For strains LSv21 and LSv514, however, growth yields were highest at the lowest temperatures, around 0°C. The results indicate that psychrophilic sulphate-reducing bacteria are specially adapted to permanently low temperatures by high relative growth rates and high growth yields at in situ conditions.     

Treatment of acid mine drainage by sulphate-reducing bacteria using permeable reactive barriers: A review from laboratory to full-scale experiments

Acid mine drainage in-situbioremediation has in the last decades drawnthe attention in the field of environmentalbiotechnology. The most recent treatmenttechnique are the permeable reactive barriersusing sulphate-reducing bacteria. This viewdescribes the basis of many of the currentapproaches to use sulphate-reducing bacteria inacid mine drainage treatment, from laboratoryto full-scale realisations, and the limitationsencountered when applied to full scaleapplications.     

Metal removal by sulphate-reducing bacteria from natural and constructed wetlands

The use of wetlands is a promising technology to treat acid mine drainage, yet there is little understanding of the fundamental biological processes involved. They are considered to centre on the complex anaerobic ecology within sediments and involve the removal of metals by sulphate-reducing bacteria (SRB). These bacteria generate hydrogen sulphide and cause precipitation of metals from solution as the insoluble metal sulphide. Sulphate-reducing bacteria have been isolated from natural and constructed wetlands receiving acid mine drainage. Sulphide production by isolates and removal of the metals iron, manganese and zinc were measured, as well as utilization of a range of carbon sources. Marked ecological differences between the wetlands were reflected in population composition of SRB enrichments, and these consortia displayed significant differences in sulphide generation and rates of metal removal from solution. Rates of metal removal did not correlate with sulphide generation in all cultures, suggesting the involvement of other biological mechanisms of metal removal. Differences in substrate utilization have highlighted the need for further investigation of carbon flow and potential carbon sources within constructed wetlands.     

The mechanisms of inhibition of Desulfovibrio and Desulfotomaculum species by selenate and molybdate

The effects of two specific inhibitors of sulphate-reducing bacteria, selenate and molybdate, have been studied by measurement of sulphate uptake and by measurement of the effects of these compounds on growth of the bacteria on a variety of media. The results obtained with selenate are consistent with the view that it acts as a competitive inhibitor of sulphate transport. The results obtained with molybdate show that there is more than one site of action and suggest that the primary effect is also on sulphate transport.     

Marble stone processing powder residue as chemical adjuvant for the biologic treatment of acid mine drainage

Marble stone powder (calcite tailing) is a residual material resulting from the cutting and polishing of marble stone. Its use as adjuvant for the biologic treatment of acid mine drainage is studied. The performance of a combined chemical/biologic packed bed reactor containing a sulphate-reducing bacteria inoculum and calcite tailing as neutralising agent was compared with the individual neutralisation and biologic steps. Unlike the individual steps, the combined column is efficient in terms of metals and sulphate removal and acidity neutralisation, producing treated water suitable for irrigation. This work emphasizes on the key role played by waste calcite tailing in the inlet of the reactor. By thus adjusting pH to values adequate for sulphate-reducing bacteria water suitable for irrigation can be obtained in a single equipment with low energy demand. This goal would be impossible without the neutralisation action.