Ecology of the bacteria of the sulphur cycle with special reference to anoxic-oxic interface environments

H2S is produced as a main end-product of anaerobic mineralization in anoxic, sulphate-rich environments by a diverse population of sulphate-reducing bacteria. The sulphate reducers can carry out an almost complete oxidation of detrital organic matter to CO2. The H2S consequently becomes an important electron carrier from the anoxic to the oxic world. Thiobacilli and other colourless sulphur bacteria have the potential to oxidize the H2S at the oxic-anoxic interface in sediments or stratified waters, but their role is still poorly understood. A comparison of sulphide oxidation processes in the chemoclines of the Black Sea, the Solar Lake and in A beggiatoa mat indicated that depth scales and retention times of coexisting O2 and H2S regulate the bacterial involvement in the sulphide oxidation. The H2S specialists, Beggiatoa and Thiovulum, are optimally adapted to compete with the autocatalytic oxidation of H2S by O2. Microelectrode measurements show retention times of O2-H2S in the bacterial mats or veils of less than 1 s. In photic chemoclines of stratified waters or sulfureta, the phototrophic sulphur bacteria or cyanobacteria interact with the sulphide oxidation at the O2-H2S interface. Short cycles between H2S and intermediate oxidation products, So or S2O2 3-, are created. The bacteria of the sulfuretum are highly adapted to the diurnal rhythm of light, O2 and H2S.     

Correlation of viable cell counts,metabolic activity of sulphur-oxidizing bacteria and chemical parameters of marine sediments

Viable counts of aerobic and anaerobic chemotrophic sulphur-oxidizers as well as phototrophic sulphur bacteria were determined in sediment samples taken from two different areas along the Baltic Sea shore which were known to regularly develop sulphidic conditions. Depth profiles of bacterial cell counts were correlated with concentration profiles of chloride, sulphate, sulphide, nitrate and phosphate in the pore water of these sediments and with potential activities of nitrate reduction, thiosulphate transformation and sulphate formation. The data revealed a complex multilayered structure within the sediments. Sulphide was released into the water from sediments of both sampling areas, but it was found that light and the availability of oxygen significantly reduced this amount. In the highly reduced sediment at Hiddensee, the highest numbers of phototrophic and chemotrophic sulphur-oxidizers were found near the sediment surface. Therefore, it was concluded that the combined action of both groups of bacteria most efficiently oxidizes reduced sulphur compounds in the top layers of the sediments. Nitrate may replace oxygen as final electron acceptor and will support oxidation of sulphide, in particular when oxygen and light are limiting.     

The influence of sulphate on substrate utilization in a thermophilic sewage sludge digestor

Summary Bacterial sulphate reduction and the interaction between sulphate reduction and methane production was studied in an unadapted and sulphate-adapted thermophilic anaerobic sludge digestor. Addition of sulphate to a concentration of 5 mm (100 times the background level) did not influence gas production or volatile fatty acid concentration compared to the control digestor. When sulphate reduction was not limited by the sulphate concentration, the sulphate-adapted digestor had a sulphate reduction rate of 910 mol l^–1 day compared with 17 mol l^–1 day in the control digestor. The results indicate that the potential for sulphate reduction is low in a thermophilic sewage sludge digestor receiving a low sulphate concentration. Counts of sulphate-reducing bacteria and methanogens showed that sulphate-reducing bacteria were found only in significant numbers in the sulphate-adapted digestor and only with H₂/CO₂ as substrate. Only low numbers of acetate-utilizing sulphate-reducing bacteria were found in both digestors. When using radio-labelled acetate, the relative percentage of 2-labelled acetate converted to CO₂ was two to four times higher in the sulphate-adapted digestor compared to the control digestor. These results suggest that oxidation of acetate seems to play a larger role in the sulphate-adapted digestor.Offprint requests to: B. K. Ahring     

Effect of some sulphur compounds on soil microflora of spruce rhizosphere

The effect of a long-term application of sulphite, thiosulphate and sodium sulphate on the soil microflora and spruce seedlings was investigated in a pot experiment. Sulphur compounds decreased the concentration of bacteria, including thiobacilli, increased the concentration of microscopic fungi and sulphate-reducing bacteria; they inhibited respiration, nitrification and oxidation of thiosulphate, stimulated ammonification and oxidation of elemental sulphur. In certain cases the spruce rhizosphere exhibited just the opposite effect. In the rhizosphere the sulphate-reducing bacteria were suppressed together with thiobacilli, whose unit oxidative activity increased substantially. Growth of seedlings was inhibited by sulphite and stimulated by thiosulphate and sulphate. Sulphite, the effects of which were similar to those of sulphur dioxide immissions, was the most effective compound. In regions influenced by immissions the soil is apparently intoxicated by the absorbed sulphite.     

Long-term investigations on the changes of the MPN values of bacterial communities participating in the sulphur cycle in Lake Velencei,Hungary

Lake Velencei is a shallow lake with a fairly variable water quality due to the effect of the earlier basin reconstruction and external factors, such as the weather and the organic matter loading from the catchment areas. For studying the changes in sediment of Lake Velencei, MPN technique, based bacteriological investigations, as well as measurements of water chemical parameters were performed. Microbial communities of aerobic thiosulphate-oxidising, anaerobic phototrophic, and sulphate-reducing bacteria taking part in the sulphur cycle were monitored between 1993 and 2001. In the western part of the lake the bacteriological results and the chemical parameters verified the presence of sulphuretum. In the first low water period of the studies the quantity of the bacteria contributing to the sulphur cycle was low in the sediment. After the increase of the water level the distribution of these microbes became somewhat homogeneous in the sediment. However, the different sediment regions might be characterised with dissimilar MPN values of the studied bacterial communities.

     

Thiosulphate oxidation in the phototrophic sulphur bacterium Allochromatium vinosum

Two different pathways for thiosulphate oxidation are present in the purple sulphur bacterium Allochromatium vinosum: oxidation to tetrathionate and complete oxidation to sulphate with obligatory formation of sulphur globules as intermediates. The tetrathionate:sulphate ratio is strongly pH-dependent with tetrathionate formation being preferred under acidic conditions. Thiosulphate dehydrogenase, a constitutively expressed monomeric 30 kDa c-type cytochrome with a pH optimum at pH 4.2 catalyses tetrathionate formation. A periplasmic thiosulphate-oxidizing multienzyme complex (Sox) has been described to be responsible for formation of sulphate from thiosulphate in chemotrophic and phototrophic sulphur oxidizers that do not form sulphur deposits. In the sulphur-storing A. vinosum we identified five sox genes in two independent loci (soxBXA and soxYZ). For SoxA a thiosulphate-dependent induction of expression, above a low constitutive level, was observed. Three sox-encoded proteins were purified: the heterodimeric c-type cytochrome SoxXA, the monomeric SoxB and the heterodimeric SoxYZ. Gene inactivation and complementation experiments proved these proteins to be indispensable for thiosulphate oxidation to sulphate. The intermediary formation of sulphur globules in A. vinosum appears to be related to the lack of soxCD genes, the products of which are proposed to oxidize SoxY-bound sulphane sulphur. In their absence the latter is instead transferred to growing sulphur globules.     

Incorporation of [methyl-3H]thymidine by obligate and facultative anaerobic bacteria when grown under defined culture conditions

Abstract Incorporation of [ methyl -³H]thymidine into bacterial DNA was determined for a range of axenic anaerobic bacterial cultures: fermentative heterotrophs, sulphate-reducing bacteria, purple sulphur bacteria, acetogens and methanogens. Anaerobically growing Bacillus sp. and the obligate aerobe Thiobacillus ferrooxidans were also investigated. Actively growing cultures of sulphate-reducing bacteria belonging to the genera Desulfovibrio, Desulfotomaculum, Desulfobacter, Desulfobotulus and Desulfobulbus , purple sulphur bacteria ( Chromatium vinosum OP2 and Thiocapsa roseopersicina OP1), methanogens ( Methanococcus GS16 and Methanosarcina barkeri ) and an acetogen ( Acetobacterium woodii ) did not incorporate [ methyl -³H]thymidine into DNA. The only obligate anaerobes in which thymidine incorporation into DNA could be unequivocally demonstrated were members of the genus Clostridium . Anaerobically growing Bacillus sp. also incorporated thymidine. These data demonstrate that pure culture representatives of major groups of anaerobic bacteria involved in the terminal oxidation of organic carbon and anoxygenic phototrophs within sediments are unable to incorporate [ methyl -³H]thymidine into DNA, although some obligate and facultative anaerobes can. Variability in thymidine incorporation amongst pure culture isolates indicates that unless existing techniques can be calibrated to take this into consideration then productivity estimates in both aerobic and anaerobic environments may be greatly underestimated using the [ methyl -³H]thymidine technique.     

Dimethyl sulphide and methanethiol formation in microbial mats: potential pathways for biogenic signatures

Mechanisms of dimethyl sulphide (DMS) and methanethiol (MT) production and consumption were determined in moderately hypersaline mats, Guerrero Negro, Mexico. Biological pathways regulated the net flux of DMS and MT as revealed by increases in flux resulting from decreased salinity, increased temperature and the removal of oxygen. Dimethylsulphoniopropionate (DMSP) was not present in these microbial mats and DMS and MT are probably formed by the reaction of photosynthetically produced low-molecular weight organic carbon and biogenic hydrogen sulphide derived from sulphate reduction. These observations provide an alternative to the notion that DMSP or S-containing amino acids are the dominant precursors of DMS in intertidal sediment systems. The major sink for DMS in the microbial mats was biological consumption, whereas photochemical oxidation to dimethylsulphoxide was the major sink for DMS in the overlying water column. Diel flux measurements demonstrated that significantly more DMS is released from the system during the night than during the day. The major consumers of DMS in the presence of oxygen were monooxygenase-utilizing bacteria, whereas under anoxic conditions, DMS was predominantly consumed by sulphate-reducing bacteria and methanethiol was consumed by methanogenic bacteria. Aerobic and anaerobic consumption rates of DMS were nearly identical. Mass balance estimates suggest that the consumption in the water column is likely to be smaller than net the flux from the mats. Volatile organic sulphur compounds are thus indicators of high rates of carbon fixation and sulphate reduction in these laminated sediment ecosystems, and atmospheric sulphur can be generated as a biogenic signature of the microbial mat community.     

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.     

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.     

Characteristics of a new species of budding purple bacteria containing bacteriochlorophyll b]

A culture of phototrophic purple bacteria containing bacteriochlorophyll b has been isolated. The bacterium forms a sessile bud and, along with producing motile swarmer cells, it produces immobile oval cells surrounded with a slime capsule. It grows on media with simple organic substances in the presence of yeast extract or vitamins, under anaerobic conditions in the light or under microaerophilic conditions in the darkness. Besides organic substances, the culture assimilates hydrogen sulphide and thiosulphate as electron donors; it is incapable of assimilative sulphate reduction. In the light, the bacterium oxidizes thiosulphate to sulphates, without accumulation of molecular sulphur. The bacterium is classed as a new species of the genus Rhodopseudomonas--Rhodopseudomonas sulfoviridis nov. sp.     

Metabolism of organic compounds in anaerobic,hydrothermal sulphate-reducing marine sediments

Previous studies of hot (>80 degrees C) microbial ecosystems have primarily relied on the study of pure cultures or analysis of 16S rDNA sequences. In order to gain more information on anaerobic metabolism by natural communities in hot environments, sediments were collected from a shallow marine hydrothermal vent system in Baia di Levante, Vulcano, Italy and incubated under strict anaerobic conditions at 90 degrees C. Sulphate reduction was the predominant terminal electron-accepting process in the sediments. The addition of molybdate inhibited sulphate reduction in the sediments and resulted in a linear accumulation of acetate and hydrogen over time. [U-14C]- acetate was completely oxidized to 14CO2, and the addition of molybdate inhibited 14CO2 production by 60%. [U-14C]-glucose was oxidized to 14CO2, and this was inhibited when molybdate was added. When the pool sizes of short-chain fatty acids were artificially increased, radiolabel from [U-14C]-glucose accumulated in the acetate pool. L-[U-14C]-glutamate, [ring-14C]-benzoate and [U-14C]-palmitate were also anaerobically oxidized to 14CO2 in the sediments, but molybdate had little effect on the oxidation of these compounds. These results demonstrate that natural microbial communities living in a hot, microbial ecosystem can oxidize acetate and a range of other organic electron donors under sulphate-reducing conditions and suggest that acetate is an important extracellular intermediate in the anaerobic degradation of organic matter in hot microbial ecosystems.     

Microbiological Reduction of Sulphates in Salty Environments and Mineralogical Characterization of the Transformation Products

This research was focused on the selection, growth and identification of SRB from soils that were subjected to long-term activity of brine, and an evaluation of mineral phases formed during the biodegradation of organic compounds and sulphate reduction. Isolated communities of anaerobic microorganisms were incubated on Postgate C medium with lactate and/or ethanol as the sole carbon source and were adapted for growth at 4% NaCl. Active reduction of sulphates with simultaneous biodegradation of organic compounds was observed in all cultures. The largest reduction of sulphates was noted in cultures with lactate as the sole carbon source; it reached 1438 mg/L, which corresponds to a 43% reduction of sulphates introduced to the medium. SRB activity in the biodegradation of organic compounds varied between 20 and 80% and depended on the level of salinity of the environment in which the SRB communities were isolated, and on the electron donor applied. The presence of biotransformation products in the post-culture deposits in the form of elemental sulphur reflects the activity of the communities. Additionally, the influence of selected communities on the salinity index was analyzed. Active SRB communities decreased the salinity of the environment by as much as 50%. Sulphate-reducing bacteria are an important group of anaerobic microorganisms, especially considering their participation in such geological processes as mineral precipitation and mineralization of organic matter in extreme environmental conditions, including high salinity.     

Metabolic interactions between anaerobic bacteria in methanogenic environments

In methanogenic environments organic matter is degraded by associations of fermenting, acetogenic and methanogenic bacteria. Hydrogen and formate consumption, and to some extent also acetate consumption, by methanogens affects the metabolism of the other bacteria. Product formation of fermenting bacteria is shifted to more oxidized products, while acetogenic bacteria are only able to metabolize compounds when methanogens consume hydrogen and formate efficiently. These types of metabolic interaction between anaerobic bacteria is due to the fact that the oxidation of NADH and FADH₂ coupled to proton or bicarbonate reduction is thermodynamically only feasible at low hydrogen and formate concentrations. Syntrophic relationships which depend on interspecies hydrogen or formate transfer were described for the degradation of e.g. fatty acids, amino acids and aromatic compounds.     

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.     

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.     

Sulphate reduction and sulphur cycling in lake sediments: a review

1. The concentration of sulphate is low in lakes and sulphur cycling has often been neglected in studies of organic matter diagenesis in lake sediments. The cycling of sulphur is, however, both spatially and temporally dynamic and strongly influences many biogeochemical reactions in sediments, such as the binding of phosphorus. This review examines the control of sulphate reduction and sulphur cycling in sediments of lakes with different trophic status. 2. The factors that control the rate of sulphate reduction have not been identified with certainty in the various environments because many factors are involved, e.g. oxygen and sulphate concentrations, temperature and organic matter availability. 3. Sulphate reduction is less significant under oligotrophic conditions, where mineralization is dominated by oxic decomposition. The supply of organic matter may not be sufficient to support sulphate reduction in the anoxic parts of sediments and, also, sulphate availability may control the rate as the concentration is generally low in oligotrophic lakes. 4. There is a potential for significant sulphate reduction in eutrophic lakes, as both the availability of organic matter and sulphate concentration are often higher than in oligotrophic lakes. Sulphate is rapidly depleted with sediment depth, however, and methanogenesis is generally the most important process in overall carbon mineralization. Sulphate reduction is generally low in acidic lakes because of low sulphate availability and reduced microbial activity. 5. It is still unclear which of the forms of sulphur deposits are the most important and under which conditions burial occurs. Sulphur deposition is controlled by the rate of sulphate reduction and reoxidation. Reoxidation of sulphides occurs rapidly through several pathways, both under oxic and anoxic conditions. Only a few studies have been able to examine the importance of reoxidation, but it is hypothesized that most of the reoxidation takes place under anoxic conditions and that disproportionation is often involved. The presence of sulphide oxidizing bacteria, benthic fauna and rooted macrophytes may substantially enhance oxic reoxidation. Deposition of sulphur is generally higher in eutrophic than in oligotrophic lakes because of a number of factors: a higher rate of sulphate reduction, enhanced sedimentation of organic sulphur and less reoxidation as a result of reduced penetration of oxygen into the sediments, a lack of faunal activity and rooted macrophytes.     

Sulphate reduction in oxic and sub-oxic North-East Atlantic sediments

Abstract Oxic and sub-oxic N.-E. Atlantic sediments were examined for sulphate-reducing activity. Oxygen and/or nitrate reduction are probably the dominant mineralisation processes in the abyssal plain sediment studied. A low rate of sulphate reduction (0.1 nmol SO²⁻₄/ml/day) was recorded in the surface 5 cm of the continental slope sediment, together with the presence of a range of sulphate-reducing bacteria (SRB). A higher activity of sulphate reduction (2.2 nmol SO²⁻₄/ml/day) occurred in the continental shelf sediment which led to a small decrease in pore water sulphate and an increase in titration alkalinity. This sediment contained approx. 10²–10³ acetate, lactate and propionate oxidising SRB/ml. No low- M _r organic acids were detected in these sediments. However, amendment with 75 μM acetate stimulated sulphate-reducing activity in the shelf sediment.