Antiquity and evolutionary status of bacterial sulfate reduction: Sulfur isotope evidence

The presently available sedimentary sulfur isotope record for the Precambrian seems to allow the following conclusions: (1) In the Early Archaean, sedimentary ^{3 4} patterns attributable to bacteriogenic sulfate reduction are generally absent. In particular, the ^{3 4} spread observed in the Isua banded iron formation (3.7×10⁹ yr) is extremely narrow and coincides completely with the respective spreads yielded by contemporaneous rocks of assumed mantle derivation. Incipient minor differentiation of the isotope patterns notably of Archaean sulfates may be accounted for by photosynthetic sulfur bacteria rather than by sulfate reducers. (2) Isotopic evidence of dissimilatory sulfate reduction is first observed in the upper Archaean of the Aldan Shield, Siberia (3.0×10⁹ yr) and in the Michipicoten and Woman River banded iron formations of Canada (2.75×10⁹ yr). This narrows down the possible time of appearance of sulfate respirers to the interval 2.8–3.1×10⁹ yr. (3) Various lines of evidence indicate that photosynthesis is older than sulfate respiration, the SO ₄ ^2– utilized by the first sulfate reducers deriving most probably from oxidation of reduced sulfur compounds by photosynthetic sulfur bacteria. Sulfate respiration must, in turn, have antedated oxygen respiration as O₂-respiring multicellular eucaryotes appear late in the Precambrian. (4) With the bulk of sulfate in the Archaean oceans probably produced by photosynthetic sulfur bacteria, the accumulation of SO ₄ ^2– in the ancient seas must have preceded the buildup of appreciable steady state levels of free oxygen. Hence, the occurrence of sulfate evaporites in Archaean sediments does not necessarily provide testimony of oxidation weathering on the ancient continents and, consequently, of the existence of an atmospheric oxygen reservoir.Paper presented at the Fourth College Park Colloquium on Chemical Evolution, Limits of Life, October 18–20, 1978.     

Occurrence of Purple Sulfur Bacteria in a Sewage Treatment Lagoon

The ecology of purple sulfur bacteria in a sewage oxidation lagoon was investigated. Chemical changes in the lagoon were investigated by monitoring biochemical oxygen demand (BOD(5)), sulfide, sulfate, phosphate, total carbohydrates, volatile acids, alkalinity, and pH. Lagoon water temperatures were observed daily. Microbial ecological relationships were deduced by enumerating coliforms, total bacteria other than anaerobes [Tryptone Glucose Extract (TGE) agar], methane formers such as Methanobacterium formicicum, sulfate reducers, purple sulfur bacteria, and algae. Finally, two strains of purple sulfur bacteria were characterized. Two populations, purple sulfur bacteria and total bacteria (TGE agar), reached maximal concentrations in the warmest part of the 1967 summer. Purple sulfur bacteria reached maximal numbers as concentrations of sulfide and volatile acids were depleted, whereas carbohydrates and alkalinity remained unchanged. Low sulfate levels, which were not limiting for sulfate reducers, may be attributable to storage of sulfur within purple sulfur bacteria. No biological, chemical, or physical agent was linked to the removal of coliforms. The increase of algae in the late summer of 1967 may have been related to the low organic content of the lagoon during this period. Although lagoon pH (7.7 to 8.2) was favorable for purple sulfur bacterial growth, temperatures and sulfides were not optimal in the lagoon for these organisms. Chromatium vinosum and Thiocapsa floridana (the predominant lagoon purple sulfur organism in 1967 and 1968) utilized certain carbohydrates, amino acids, volatile acids, and Krebs cycle intermediates. Also purple sulfur bacteria lowered BOD levels as demonstrated by the growth of T. floridana in sterilized sewage.     

Sulfate-reducing bacteria in rice field soil and on rice roots.

Rice plants that were grown in flooded rice soil microcosms were examined for their ability to exhibit sulfate reducing activity. Washed excised rice roots showed sulfate reduction potential when incubated in anaerobic medium indicating the presence of sulfate-reducing bacteria. Rice plants, that were incubated in a double-chamber (phylloshpere and rhizosphere separated), showed potential sulfate reduction rates in the anoxic rhizosphere compartment. These rates decreased when oxygen was allowed to penetrate through the aerenchyma system of the plants into the anoxic root compartment, indicating that sulfate reducers on the roots were partially inhibited by oxygen or that sulfate was regenerated by oxidation of reduced S-compounds. The potential activity of sulfate reducers on rice roots was consistent with MPN enumerations showing that H2-utilizing sulfate-reducing bacteria were present in high numbers on the rhizoplane (4.1 x 10(7) g-1 root fresh weight) and in the adjacent rhizosperic soil (2.5 x 10(7) g-1 soil dry weight). Acetate-oxidizing sulfate reducers, on the other hand, showed highest numbers in the unplanted bulk soil (1.9 x 10(6) g-1 soil dry weight). Two sulfate reducing bacteria were isolated from the highest dilutions of the MPN series and were characterized physiologically and phylogenetically. Strain F1-7b which was isolated from the rhizoplane with H2 as electron donor was related to subgroup II of the family Desulfovibrionaceae. Strain EZ-2C2, isolated from the rhizoplane on acetate, grouped together with Desulforhabdus sp. and Syntrophobacter wolinii. Other strains of sulfate-reducing bacteria originated from bulk soil of rice soil microcosms and were isolated using different electron donors. From these isolates, strains R-AcA1, R-IbutA1, R-PimA1 and R-AcetonA170 were Gram-positive bacteria which were affiliated with the genus Desulfotomaculum. The other isolates were members of subgroup II of the Desulfovibrionaceae (R-SucA1 and R-LacA1), were related to Desulforhabdus sp. (strain BKA11), Desulfobulbus (R-PropA1), or culstered between Desulfobotulus sapovorans and Desulfosarcina variabilis (R-ButA1 and R-CaprA1).     

Isotope effects associated with the anaerobic oxidation of sulfite and thiosulfate by the photosynthetic bacterium, Chromatium vinosum

Abstract The purple photosynthetic bacterium Chromatium vinosum , strain D, catalyzes several oxidations of reduced sulfur compounds under anaerobic conditions in the light: e.g., sulfide → sulfur → sulfate, sulfite → sulfate, and thiosulfate → sulfur + sulfate. Here it is shown that no sulfur isotope effect is associated with the last of these processes; isotopic compositions of the sulfur and sulfate produced can differ, however, if the sulfane and sulfonate positions within the thiosulfate have different isotopic compositions. In the second process, an observed change from an inverse to a normal isotope effect during oxidation of sulfite may indicate the operation of 2 enzymatic pathways. In contrast to heterotrophic anaerobic reduction of oxidized sulfur compounds, anaerobic oxidations of inorganic sulfur compounds by photosynthetic bacteria are characterized by relatively small isotope effects.     

Rates of sulfate reduction and thiosulfate consumption in a marine microbial mat

Abstract The sulfur cycle in a microbial mat was studied by determining viable counts of sulfate-reducing bacteria, chemolithoautotrophic sulfur bacteria and anoxygenic phototrophic bacteria. All three functional groups of sulfur bacteria revealed a maximum population density in the uppermost 5 mm of the mat: 1.1 × 10⁸ cells of sulfate reducers cm⁻³ sediment, 2.0 × 10⁹ cells of chemolithoautotrophs cm⁻³ sediment, and 4.0 × 10⁷ cells of anoxygenic phototrophs cm⁻³ sediment. Bacterial dynamics were studied by sulfate reduction rate measurements, both under anoxic conditions (dark incubation) and oxic conditions (incubation in the light), and determination of the vertical distribution of the potential rate of thiosulfate consumption under oxic conditions. Sulfate reduction rates in the top 5 mm of the sediment were 566 nmol cm⁻³ d⁻¹ in the absence of oxygen, and 123 nmol cm⁻³ d⁻¹ in the presence of oxygen. In the latter case, the maximum rate was found in the 5–10-mm depth horizon (361 nmol cm⁻³ d⁻¹). Biological consumption of amended thiosulfate was rapid and decreased with depth, while in the presence of molybdate, thiosulfate consumption decreased to 10–30% of the original rate.     

Ecological observations on Heterotrophic,methane oxidizing and sulfate reducing bacteria in Pond

Numbers of heterotrophic, methane oxidizing and sulfate reducing bacteria were counted in Lake Vechten. A dynamic distribution pattern was found in the stratified lake. A maximum of heterotrophs (numbers of 10⁹ bact./l) occured in the deepest part of the lake in spring and in the metalimnion during summer-stratification. These bacteria use nearly all available oxygen in the hypolimnion. It was found that the concentration of available organic material and the oxygen tension caused the numbers of heterotrophs in the metalimnion to be high.The maximal numbers of methane oxidizers (numbers of 5.10⁵ bact./l) were found at a depth of maximal methane concentration: the de-oxygenated hypolimnion. Preliminary evidence indicated that these organisms were facultative methane oxidizers and must be regarded as micro-aerophyllics. By oxidizing methane they removed the residual oxygen under the metalimnion.The sulfate reducing bacteria could be observed in the hypolimnion only. Decreased SO _inf4 ^sup–2 concentration and increased numbers, of bacterai were found in the bottom water. An association between the methane oxidizers and the sulfate reducers could be deduced. It was assumed that favourable redox requirements for obligate anaerobic sulfate reducers were the results of the activities of the methane oxidizing bacteria.The dynamic distribution equilibrium of the investigated groups of bacteria was disturbed by the autumn turn-over. The heterotrophic and methane oxidizing bacteria decreased in number at that period and were equally distributed, no sulfate reducers could be detected in the free water of Lake Vechten.     

Diversity of sulfur isotope fractionations by sulfate-reducing prokaryotes

Batch culture experiments were performed with 32 different sulfate-reducing prokaryotes to explore the diversity in sulfur isotope fractionation during dissimilatory sulfate reduction by pure cultures. The selected strains reflect the phylogenetic and physiologic diversity of presently known sulfate reducers and cover a broad range of natural marine and freshwater habitats. Experimental conditions were designed to achieve optimum growth conditions with respect to electron donors, salinity, temperature, and pH. Under these optimized conditions, experimental fractionation factors ranged from 2.0 to 42.0 per thousand. Salinity, incubation temperature, pH, and phylogeny had no systematic effect on the sulfur isotope fractionation. There was no correlation between isotope fractionation and sulfate reduction rate. The type of dissimilatory bisulfite reductase also had no effect on fractionation. Sulfate reducers that oxidized the carbon source completely to CO2 showed greater fractionations than sulfate reducers that released acetate as the final product of carbon oxidation. Different metabolic pathways and variable regulation of sulfate transport across the cell membrane all potentially affect isotope fractionation. Previous models that explained fractionation only in terms of sulfate reduction rates appear to be oversimplified. The species-specific physiology of each sulfate reducer thus needs to be taken into account to understand the regulation of sulfur isotope fractionation during dissimilatory sulfate reduction.     

Immunological cross-reactivities of adenosine-5'-phosphosulfate reductases from sulfate-reducing and sulfide-oxidizing bacteria

Crude extracts from 14 species of sulfate-reducing bacteria comprising the genera Desulfovibrio, Desulfotomaculum, Desulfobulbus, and Desulfosarcina and from three species of sulfide-oxidizing bacteria were tested in an enzyme-linked immunosorbent assay with polyclonal antisera to adenosine 5'-phosphosulfate reductase from Desulfovibrio desulfuricans G100A. The results showed that extracts from Desulfovibrio species were all highly cross-reactive, whereas extracts from the other sulfate-reducing genera showed significantly less cross-reaction. An exception was Desulfotomaculum orientis, which responded more like Desulfovibrio species than the other Desulfotomaculum strains tested. Extracts from colorless or photosynthetic sulfur bacteria were either unreactive or exhibited very low levels of reactivity with the antibodies to the enzyme from sulfate reducers. These results were confirmed by using partially purified enzymes from sulfate reducers and the most cross-reactive sulfide oxidizer, Thiobacillus denitrificans. Two types of monoclonal antibodies to adenosine 5'-phosphosulfate reductase were also isolated. One type reacted more variably with the enzymes of the sulfate reducers and poorly with the Thiobacillus enzyme, whereas the second reacted strongly with Desulfovibrio, Desulfotomaculum orientis, and Thiobacillus enzymes.     

Diversity of Sulfur Isotope Fractionations by Sulfate-Reducing Prokaryotes

Batch culture experiments were performed with 32 different sulfate-reducing prokaryotes to explore the diversity in sulfur isotope fractionation during dissimilatory sulfate reduction by pure cultures. The selected strains reflect the phylogenetic and physiologic diversity of presently known sulfate reducers and cover a broad range of natural marine and freshwater habitats. Experimental conditions were designed to achieve optimum growth conditions with respect to electron donors, salinity, temperature, and pH. Under these optimized conditions, experimental fractionation factors ranged from 2.0 to 42.0‰. Salinity, incubation temperature, pH, and phylogeny had no systematic effect on the sulfur isotope fractionation. There was no correlation between isotope fractionation and sulfate reduction rate. The type of dissimilatory bisulfite reductase also had no effect on fractionation. Sulfate reducers that oxidized the carbon source completely to CO₂ showed greater fractionations than sulfate reducers that released acetate as the final product of carbon oxidation. Different metabolic pathways and variable regulation of sulfate transport across the cell membrane all potentially affect isotope fractionation. Previous models that explained fractionation only in terms of sulfate reduction rates appear to be oversimplified. The species-specific physiology of each sulfate reducer thus needs to be taken into account to understand the regulation of sulfur isotope fractionation during dissimilatory sulfate reduction.     

Iron isotope fractionation during microbial dissimilatory iron oxide reduction in simulated Archaean seawater

The largest Fe isotope excursion yet measured in marine sedimentary rocks occurs in shales, carbonates, and banded iron formations of Neoarchaean and Paleoproterozoic age. The results of field and laboratory studies suggest a potential role for microbial dissimilatory iron reduction (DIR) in producing this excursion. However, most experimental studies of Fe isotope fractionation during DIR have been conducted in simple geochemical systems, using pure Fe(III) oxide substrates that are not direct analogues to phases likely to have been present in Precambrian marine environments. In this study, Fe isotope fractionation was investigated during microbial reduction of an amorphous Fe(III) oxide-silica coprecipitate in anoxic, high-silica, low-sulphate artificial Archaean seawater at 30 °C to determine if such conditions alter the extent of reduction or isotopic fractionations relative to those observed in simple systems. The Fe(III)-Si coprecipitate was highly reducible (c. 80% reduction) in the presence of excess acetate. The coprecipitate did not undergo phase conversion (e.g. to green rust, magnetite or siderite) during reduction. Iron isotope fractionations suggest that rapid and near-complete isotope exchange took place among all Fe(II) and Fe(III) components, in contrast to previous work on goethite and hematite, where exchange was limited to the outer few atom layers of the substrate. Large quantities of low-δ(56)Fe Fe(II) (aqueous and solid phase) were produced during reduction of the Fe(III)-Si coprecipitate. These findings shed new light on DIR as a mechanism for producing Fe isotope variations observed in Neoarchaean and Paleoproterozoic marine sedimentary rocks.     

In vitro and in vivo sulfate reduction in the gut contents of the termite Mastotermes darwiniensis and the rose-chafer Pachnoda marginata

Sulfate-reducing bacteria (SRB) from termites have been assigned to the genus Desulfovibrio. Desulfovibrio intestinalis lives in the gut of the Australian termite Mastotermes darwiniensis. For the first time we were able to enrich and identify a sulfate-reducing bacterium from the gut of the rose-chafer Pachnoda marginata, which showed the highest 16S rDNA sequence identity (93%) to Desulfovibrio intestinalis and Desulfovibrio strain STL1. Compared to Mastotermes darwiniensis (1x10(7) cells of SRB per ml gut contents), sulfate-reducing bacteria occurred in higher numbers in the gut contents of Pachnoda marginata reaching cell titers of up to 2x10(8) cells per ml gut contents. In vitro sulfate reduction rates were determined with SRB from the gut contents of the termite Mastotermes darwiniensis and the beetle Pachnoda marginata. Due to the higher cell titer, the sulfate reduction rate of Pachnoda marginata was 10(4) nmolxh-1xml-1 and therefore, 21 times higher than that of Mastotermes darwiniensis. In addition, we detected in vivo sulfate reduction in Mastotermes darwiniensis, which indicates that sulfate reducers play an active role in the sulfur metabolism in the termite gut.     

Microbiological investigation of methane- and hydrocarbon-discharging mud volcanoes in the Carpathian Mountains, Romania

Paclele Mici is a terrestrial mud volcano field located in the Carpathian Mountains (Romania), where thermal alteration of sedimentary organic compounds leads to methane, higher hydrocarbons and other petroleum compounds that are continuously released into the environment. The hydrocarbons represent potential substrates for microorganisms. We studied lipid biomarkers, stable isotope ratios, the effect of substrate (methane, other organic compounds) addition and 16S rRNA genes to gain insights into the hitherto unknown microbial community at this site. Quantitative real-time polymerase chain reaction analysis demonstrated that bacteria were much more abundant than archaea. Phylogenetic analyses of 16S rDNA clone sequences indicated the presence of bacterial and archaeal lineages generally associated with the methane cycle (methanogens, aerobic and anaerobic methanotrophs), the sulfur cycle (sulfate reducers), and groups linked to the anaerobic degradation of alkanes or aromatic hydrocarbons. The presence of sulfate reducers, methanogens and methanotrophs in this habitat was also confirmed by concurrent surveys of lipid biomarkers and their isotopic signatures. Incubation experiments with several common and complex substrates revealed the potential of the indigenous microbial community for sulfate reduction, methanogenesis and aerobic methanotrophy. Additionally, consistently to the detection of methane-oxidizing archaea (ANME) and 13C-depleted archaeal lipids, a weak but significant activity of anaerobic methane oxidation was measured by radiotracer techniques and in vitro. This survey is the first to report the presence and activity of ANME in a terrestrial environment.     

Citric-acid cycle, 50 years on. Modifications and an alternative pathway in anaerobic bacteria

Many anaerobic bacteria can completely oxidize organic matter to CO2 with either sulfur, sulfate, or protons as electron acceptor. The sulfur-reducing bacteria and one genus of sulfate reducers use a modified citric-acid cycle with a novel anaplerotic sequence as pathway of terminal respiration. All other anaerobes use an alternative pathway, in which carbon monoxide dehydrogenase is a key enzyme and in which acetyl-CoA is cleaved into two C1 units at the oxidation level of CH3OH and CO. Thus almost 50 years after the discovery of the citric acid cycle by Hans Krebs in 1937, a second pathway for acetyl-CoA oxidation was found.     

Sulfur and Oxygen Isotope Fractionation During Bacterial Sulfur Disproportionation Under Anaerobic Haloalkaline Conditions

Sulfur and oxygen isotope fractionation of elemental sulfur disproportionation at anaerobic haloalkaline conditions was evaluated for the first time. Isotope enrichment factors of the strains Desulfurivibrio alkaliphilus and Dethiobacter alkaliphilus growing at pH 9 or 10 were ?0.9‰ to ?1‰ for sulfide (³⁴?), +3.6‰ to +4.7‰ for sulfate (³⁴?), and +3.5‰ to +7.7‰ for oxygen in sulfate (¹⁸?). These values are significantly smaller compared to previously published values of sulfur disproportionators at neutral pH. We propose that this discrepancy is caused by masking effects due to preferential formation of polysulfides at high pH leading to accelerated internal sulfur turnover rates, but cannot rule out distinct isotope effects due to specific enzymatic disproportionation reactions under haloalkaline conditions. The results imply that the microbial sulfur cycle in haloalkaline environments is characterized by specific stable sulfur and oxygen isotope patterns.     

‘Follow the Water’: Hydrogeochemical Constraints on Microbial Investigations 2.4 km Below Surface at the Kidd Creek Deep Fluid and Deep Life Observatory

Abstract

Microbiological and geochemical data are presented to characterize the hydrogeochemistry and to investigate extant microbial life in fracture waters 2.4?km below surface, at the Kidd Creek Observatory in Canada. Previous studies identified the world’s oldest groundwaters with mean residence times on the order of millions to billions of years trapped in fractures in Precambrian host rock here. In this study, major ion chemistry, δ¹⁸O and δ²H isotopic signatures and dissolved gases in the fracture waters are shown to be distinct from potential contamination end-members, demonstrating the fracture waters are not impacted by waters used in mining operations. A previous work on sulfur isotope signatures suggested a longstanding indigenous population of sulfate-reducing bacteria in these highly reducing fluids and sufficient sulfate to support microbial activity. Here, we report the first evidence for extant visible and cultivable microbial life at this location. Anaerobic metabolisms were investigated using the Most Probable Number (MPN) method. The fracture fluids contained extant cells at low biomass density (～10³–10⁴ cells/mL) and showed a strong response from autotrophic sulfate-reducers and alkane-oxidizing sulfate reducers. These lines of evidence provide the interpretational framework (chemical, hydrogeologic, and microbiologic) essential to the on-going genomic and metagenomic investigations at the Kidd Creek Observatory – the world’s most longstanding location for investigation of subsurface fluids and deep life at such profound depth.     

High bacterial diversity in permanently cold marine sediments.

A 16S ribosomal DNA (rDNA) clone library from permanently cold marine sediments was established. Screening 353 clones by dot blot hybridization with group-specific oligonucleotide probes suggested a predominance of sequences related to bacteria of the sulfur cycle (43.4% potential sulfate reducers). Within this fraction, the major cluster (19.0%) was affiliated with Desulfotalea sp. and other closely related psychrophilic sulfate reducers isolated from the same habitat. The cloned sequences showed between 93 and 100% similarity to these bacteria. Two additional groups were frequently encountered: 13% of the clones were related to Desulfuromonas palmitatis, and a second group was affiliated with Myxobacteria spp. and Bdellovibrio spp. Many clones (18.1%) belonged to the gamma subclass of the class Proteobacteria and were closest to symbiotic or free-living sulfur oxidizers. Probe target groups were further characterized by amplified rDNA restriction analysis to determine diversity within the groups and within the clone library. Rarefaction analysis suggested that the total diversity assessed by 16S rDNA analysis was very high in these permanently cold sediments and was only partially revealed by screening of 353 clones.     

Devonian and modern relatives of the Precambrian Eosphaera: possible significance for the early eukaryotes

The Middle Precambrian problematical microorganism Eosphaera Barghoorn and Eosphaera- like structures known from Early and Middle Precambrian banded iron formations have been compared with the recently discovered Devonian volvocacean alga Eovolvox Kaźmierczak and some modern colonial Volvocales. The volvocacean interpretation of Eosphaera implies that algal eukaryotes (green phytoflagellates) werc already prewmt in the earth biosphere before at least 1.9 b. y. ago (Gunflint Iron Formaticn), and probably before 2.7 b. y. ago (Soudan Iron Formation). The type of metabolism and thc oxygen requirements of modern colonial Volvocales indicate that Eosphaera was most probably a photoorgano-trophic (mixotrophic) organism able to live in the extremely oxygen-deficicnt or anoxy-genous Early Precambrian environment. As an oxygen-releasing photosynthesizer, Eosphaera could have played a considerable role in the production of free oxygen during the Precambrian. The abundance of Eospkaera- like ferriferous structures in the iron microbands of many banded iron formations implies active participation of these organisns in the formation of Precambrian sedimentary iron ores. The exclusively fresh-water habitat of extant volvocacean algae suggests that the Procambrian environments inhabited by Eosphaera were non-marine.     

Oxygen isotopic composition of sulfate in deep sea pore fluid: evidence for rapid sulfur cycling

We present new data of oxygen isotopes in marine sulfate (δ¹⁸O_SO4) in pore fluid profiles through organic‐rich deep‐sea sediments from 11 ODP sites around the world. In almost all sites studied sulfate is depleted with depth, through both organic matter oxidation and anaerobic methane oxidation. The δ¹⁸O_SO4 increases rapidly near the top of the sediments, from seawater values of 9 to maxima between 22 and 25, and remains isotopically heavy and constant at these values with depth. The δ¹⁸O_SO4 in these pore fluid profiles is decoupled from variations in sulfur isotopes measured on the same sulfate samples (δ³⁴S_SO4); the δ³⁴S_SO4 increases continuously with depth and exhibits a shallower isotopic increase. This isotopic decoupling between the δ³⁴S_SO4 and the δ¹⁸O_SO4 is hard to reconcile with the traditional understanding of bacterial sulfate reduction in sediments. Our data support the idea that sulfate or sulfite and water isotopically exchange during sulfate reduction and that some of the isotopically altered sulfur pool returns to the environment. We calculate that the rapid increase in the δ¹⁸O_SO4 in the upper part of these sediments requires rates of this oxygen isotope exchange that are several orders of magnitude higher than the rates of net sulfate reduction calculated from the sulfate concentration profiles and supported by the δ³⁴S_SO4. We suggest several mechanisms by which this may occur, including ‘net‐zero’ sulfur cycling, as well as further experiments through which we can test and resolve these processes.     

Assimilatory reduction of sulfate and sulfite by methanogenic bacteria.

A variety of sulfur-containing compounds were investigated for use as medium reductants and sulfur sources for growth of four methanogenic bacteria. Sulfide (1 to 2 mM) served all methanogens investigated well. Methanococcus thermolithotrophicus and Methanobacterium thermoautotrophicum Marburg and delta H grew well with S0, SO3(2-), or thiosulfate as the sole sulfur source. Only Methanococcus thermolithotrophicus was able to grow with SO4(2-) as the sole sulfur source. 2-Mercaptoethanol at 20 mM was greatly inhibitory to growth of Methanococcus thermolithotrophicus on SO4(2-) or SO2(2-) and Methanobacterium thermoautotrophicum Marburg on SO3(2-) but not to growth of strain delta H on SO3(2-). Sulfite was metabolized during growth by Methanococcus thermolithotrophicus. Sulfide was produced in cultures of Methanococcus thermolithotrophicus growing on SO4(2-), SO3(2-), thiosulfate, and S0. Methanobacterium thermoautotrophicum Marburg was successfully grown in a 10-liter fermentor with S0, SO3(2-), or thiosulfate as the sole sulfur source.     

Immunological Cross-Reactivities of Adenosine-5′-Phosphosulfate Reductases from Sulfate-Reducing and Sulfide-Oxidizing Bacteria

Crude extracts from 14 species of sulfate-reducing bacteria comprising the genera Desulfovibrio, Desulfotomaculum, Desulfobulbus, and Desulfosarcina and from three species of sulfide-oxidizing bacteria were tested in an enzyme-linked immunosorbent assay with polyclonal antisera to adenosine 5′-phosphosulfate reductase from Desulfovibrio desulfuricans G100A. The results showed that extracts from Desulfovibrio species were all highly cross-reactive, whereas extracts from the other sulfate-reducing genera showed significantly less cross-reaction. An exception was Desulfotomaculum orientis, which responded more like Desulfovibrio species than the other Desulfotomaculum strains tested. Extracts from colorless or photosynthetic sulfur bacteria were either unreactive or exhibited very low levels of reactivity with the antibodies to the enzyme from sulfate reducers. These results were confirmed by using partially purified enzymes from sulfate reducers and the most cross-reactive sulfide oxidizer, Thiobacillus denitrificans. Two types of monoclonal antibodies to adenosine 5′-phosphosulfate reductase were also isolated. One type reacted more variably with the enzymes of the sulfate reducers and poorly with the Thiobacillus enzyme, whereas the second reacted strongly with Desulfovibrio, Desulfotomaculum orientis, and Thiobacillus enzymes.