The low-temperature germinating spores of the thermophilic Desulfofundulus contribute to an extremely high sulfate reduction in burning coal seams.

Burning coal seams, characterized by massive carbon monoxide (CO) emissions, the presence of secondary sulfates, and high temperatures, represent suitable environments for thermophilic sulfate reduction. The diversity and activity of dissimilatory sulfate reducers in these environments remain unexplored. In this study, using metagenomic approaches, in situ activity measurements with a radioactive tracer, and cultivation we have shown that members of the genus Desulfofundulus are responsible for the extremely high sulfate reduction rate (SRR) in burning lignite seams in the Altai Mountains. The maximum SRR reached 564 ± 21.9 nmol S cm-3 day-1 at 60°C and was of the same order of magnitude for both thermophilic (60°C) and mesophilic (23°C) incubations. The 16S rRNA profiles and the search for dsr gene sequences in the metagenome revealed members of the genus Desulfofundulus as the main sulfate reducers. The thermophilic Desulfofundulus sp. strain Al36 isolated in pure culture, did not grow at temperatures below 50°C, but produced spores that germinated into metabolically active cells at 20 and 15°C. Vegetative cells germinating from spores produced up to 0.738 ± 0.026 mM H2S at 20°C and up to 0.629 ± 0.007 mM H2S at 15°C when CO was used as the sole electron donor. The Al36 strain maintains significant production of H2S from sulfate over a wide temperature range from 15°C to 65°C, which is important in variable temperature biotopes such as lignite burning seams. Burning coal seams producing CO are ubiquitous throughout the world, and biogenic H2S may represent an overlooked significant flux to the atmosphere. The thermophilic spore outgrowth and their metabolic activity at temperatures below the growth minimum may be important for other spore-forming bacteria of environmental, industrial and clinical importance.

Biogeochemical Activity of Methane-Related Microbial Communities in Bottom Sediments of Cold Seeps of the Laptev Sea.

Bottom sediments at methane discharge sites of the Laptev Sea shelf were investigated. The rates of microbial methanogenesis and methane oxidation were measured, and the communities responsible for these processes were analyzed. Methane content in the sediments varied from 0.9 to 37 µmol CH4 dm-3. Methane carbon isotopic composition (δ13C-CH4) varied from -98.9 to -77.6‱, indicating its biogenic origin. The rates of hydrogenotrophic methanogenesis were low (0.4-5.0 nmol dm-3 day-1). Methane oxidation rates varied from 0.4 to 1.2 µmol dm-3 day-1 at the seep stations. Four lineages of anaerobic methanotrophic archaea (ANME) (1, 2a-2b, 2c, and 3) were found in the deeper sediments at the seep stations along with sulfate-reducing Desulfobacteriota. The ANME-2a-2b clade was predominant among ANME. Aerobic ammonium-oxidizing Crenarchaeota (family Nitrosopumilaceae) predominated in the upper sediments along with heterotrophic Actinobacteriota and Bacteroidota, and mehtanotrophs of the classes Alphaproteobacteria (Methyloceanibacter) and Gammaproteobacteria (families Methylophilaceae and Methylomonadaceae). Members of the genera Sulfurovum and Sulfurimonas occurred in the sediments of the seep stations. Mehtanotrophs of the classes Alphaproteobacteria (Methyloceanibacter) and Gammaproteobacteria (families Methylophilaceae and Methylomonadaceae) occurred in the sediments of all stations. The microbial community composition was similar to that of methane seep sediments from geographically remote areas of the global ocean.

Active Sulfate-Reducing Bacterial Community in the Camel Gut.

The diversity and activity of sulfate-reducing bacteria (SRB) in the camel gut remains largely unexplored. An abundant SRB community has been previously revealed in the feces of Bactrian camels (Camelus bactrianus). This study aims to combine the 16S rRNA gene profiling, sulfate reduction rate (SRR) measurement with a radioactive tracer, and targeted cultivation to shed light on SRB activity in the camel gut. Fresh feces of 55 domestic Bactrian camels grazing freely on semi-arid mountain pastures in the Kosh-Agach district of the Russian Altai area were analyzed. Feces were sampled in early winter at an ambient temperature of -15 °C, which prevented possible contamination. SRR values measured with a radioactive tracer in feces were relatively high and ranged from 0.018 to 0.168 nmol S cm-3 day-1. The 16S rRNA gene profiles revealed the presence of Gram-negative Desulfovibrionaceae and spore-forming Desulfotomaculaceae. Targeted isolation allowed us to obtain four pure culture isolates belonging to Desulfovibrio and Desulforamulus. An active SRB community may affect the iron and copper availability in the camel intestine due to metal ions precipitation in the form of sparingly soluble sulfides. The copper-iron sulfide, chalcopyrite (CuFeS2), was detected by X-ray diffraction in 36 out of 55 analyzed camel feces. In semi-arid areas, gypsum, like other evaporite sulfates, can be used as a solid-phase electron acceptor for sulfate reduction in the camel gastrointestinal tract.

Coexistence of Psychrophilic, Mesophilic, and Thermophilic Sulfate-Reducing Bacteria in a Deep Subsurface Aquifer Associated with Coal-Bed Methane Production.

The microbial community of subsurface environments remains understudied due to limited access to deep strata and aquifers. Coal-bed methane (CBM) production is associated with a large number of wells pumping water out of coal seams. CBM wells provide access to deep biotopes associated with coal-bed water. Temperature is one of the key constraints for the distribution and activity of subsurface microorganisms, including sulfate-reducing prokaryotes (SRP). The 16S rRNA gene amplicon sequencing coupled with in situ sulfate reduction rate (SRR) measurements with a radioactive tracer and cultivation at various temperatures revealed that the SRP community of the coal bed water of the Kuzbass coal basin is characterized by an overlapping mesophilic-psychrophilic boundary. The genus Desulfovibrio comprised a significant share of the SRP community. The D. psychrotolerans strain 1203, which has a growth optimum below 20 °C, dominated the cultivated SRP. SRR in coal bed water varied from 0.154 ± 0.07 to 2.04 ± 0.048 nmol S cm-3 day-1. Despite the ambient water temperature of ~ 10-20 °C, an active thermophilic SRP community occurred in the fracture water, which reduced sulfate with the rate of 0.159 ± 0.023 to 0.198 ± 0.007 nmol S cm-3 day-1 at 55 °C. A novel moderately thermophilic "Desulforudis audaxviator"-clade SRP has been isolated in pure culture from the coal-bed water.

A Novel View of the Diversity of Anoxygenic Phototrophic Bacteria Inhabiting the Chemocline of Meromictic Karst Lakes.

The rates of oxygenic and anoxygenic photosynthesis, the microorganisms responsible for these processes, and the hydrochemical characteristics of the sulfide-containing karst lakes, Black Kichier and Big Kichier (Mari El Republic), were investigated. In these lakes, a plate of anoxygenic phototrophic bacteria (APB) is formed at the upper boundary of sulfide occurrence in the water. The phototrophic community of the chemocline zone was analyzed using a combination of high-throughput sequencing of the 16S rRNA gene fragments and light and electron microscopic techniques. Green-colored Chlorobium clathratiforme were absolutely predominant in both lakes. The minor components included green sulfur bacteria (GSB) Chlorobium spp., symbiotic consortia Chlorochromatium magnum and Pelochromatium roseum, purple sulfur bacteria (PSB) Chromatium okenii, and unidentified phylotypes of the family Chromatiaceae, as well as members of the Chloroflexota: Chloronema sp. and Oscillochloris sp. Based on the results of the molecular analysis, the taxonomic status of Ancalochloris perfilievii and other prosthecate GSB, as well as of the PSB Thiopedia rosea, which were visually revealed in the studied freshwater lakes, is discussed.

Phylogenetic diversity in sulphate-reducing bacterial communities from oxidised and reduced bottom sediments of the Barents Sea.

In the bottom sediments from a number of the Barents Sea sites, including coastal areas of the Novaya Zemlya, Franz Josef Land, and Svalbard archipelagos, sulphate reduction rates were measured and the phylogenetic composition of sulphate-reducing bacterial (SRB) communities was analysed for the first time. Molecular genetic analysis of the sequences of the 16S rRNA and dsrB genes (the latter encodes the ß-subunit of dissimilatory (bi)sulphite reductase) revealed significant differences in the composition of bacterial communities in different sampling stations and sediment horizons of the Barents Sea depending on the physicochemical conditions. The major bacteria involved in reduction of sulphur compounds in Arctic marine bottom sediments belonged to Desulfobulbaceae, Desulfobacteraceae, Desulfovibrionaceae, Desulfuromonadaceae, and Desulfarculaceae families, as well as to uncultured clades SAR324 and Sva0485. Desulfobulbaceae and Desulfuromonadaceae predominated in the oxidised (Eh = 154-226 mV) upper layers of the sediments (up to 9% and 5.9% from all reads of the 16S rRNA gene sequences in the sample, correspondingly), while in deeper, more reduced layers (Eh = -210 to -105 mV) the share of Desulfobacteraceae in the SRB community was also significant (up to 5%). The highest relative abundance of members of Desulfarculaceae family (3.1%) was revealed in reduced layers of sandy-clayey sediments from the Barents Sea area affected by currents of transformed (mixed, with changed physicochemical characteristics) Atlantic waters.

Microbial Communities Involved in Methane, Sulfur, and Nitrogen Cycling in the Sediments of the Barents Sea.

A combination of physicochemical and radiotracer analysis, high-throughput sequencing of the 16S rRNA, and particulate methane monooxygenase subunit A (pmoA) genes was used to link a microbial community profile with methane, sulfur, and nitrogen cycling processes. The objects of study were surface sediments sampled at five stations in the northern part of the Barents Sea. The methane content in the upper layers (0-5 cm) ranged from 0.2 to 2.4 µM and increased with depth (16-19 cm) to 9.5 µM. The rate of methane oxidation in the oxic upper layers varied from 2 to 23 nmol CH4 L-1 day-1 and decreased to 0.3 nmol L-1 day-1 in the anoxic zone at a depth of 16-19 cm. Sulfate reduction rates were much higher, from 0.3 to 2.8 µmol L-1 day-1. In the surface sediments, ammonia-oxidizing Nitrosopumilaceae were abundant; the subsequent oxidation of nitrite to nitrate can be carried out by Nitrospira sp. Aerobic methane oxidation could be performed by uncultured deep-sea cluster 3 of gamma-proteobacterial methanotrophs. Undetectable low levels of methanogenesis were consistent with a near complete absence of methanogens. Anaerobic methane oxidation in the deeper sediments was likely performed by ANME-2a-2b and ANME-2c archaea in consortium with sulfate-reducing Desulfobacterota. Sulfide can be oxidized by nitrate-reducing Sulfurovum sp. Thus, the sulfur cycle was linked with the anaerobic oxidation of methane and the nitrogen cycle, which included the oxidation of ammonium to nitrate in the oxic zone and denitrification coupled to the oxidation of sulfide in the deeper sediments. Methane concentrations and rates of microbial biogeochemical processes in sediments in the northern part of the Barents Sea were noticeably higher than in oligotrophic areas of the Arctic Ocean, indicating that an increase in methane concentration significantly activates microbial processes.

Microbial sulfate reduction by Desulfovibrio is an important source of hydrogen sulfide from a large swine finishing facility.

There is still a lack of understanding of H2S formation in agricultural waste, which leads to poor odour prevention and control. Microbial sulfate reduction is a major process contributing to sulfide formation in natural and technogenic environments with high sulfate and low oxygen concentration. Agricultural waste can be considered a low-sulfate system with no obvious input of oxidised sulfur compounds. The purpose of this study was to characterise a microbial community participating in H2S production and estimate the microbial sulfate reduction rate (SRR) in manure slurry from a large-scale swine finishing facility in Western Siberia. In a series of manure slurry microcosms, we identified bacterial consortia by 16S rRNA gene profiling and metagenomic analysis and revealed that sulfate-reducing Desulfovibrio were key players responsible for H2S production. The SRR measured with radioactive sulfate in manure slurry was high and comprised 7.25 nmol S cm-3 day-1. Gypsum may be used as a solid-phase electron acceptor for sulfate reduction. Another plausible source of sulfate is a swine diet, which often contains supplements in the form of sulfates, including lysine sulfate. Low-sulfur diet, manure treatment with iron salts, and avoiding gypsum bedding are possible ways to mitigate H2S emissions from swine manure.

Microbial Processes and Microbial Communities in the Water Column of the Polar Meromictic Lake Bol'shie Khruslomeny at the White Sea Coast.

Microbiological, molecular ecological, biogeochemical, and isotope geochemical research was carried out at the polar Lake Bol'shie Khruslomeny at the coast of the Kandalaksha Bay, White Sea in March and September 2017. The uppermost mixolimnion was oxic, with low salinity (3-5%). The lower chemocline layer was brown-green colored, with very high content of particulate organic matter (up to 11.8 mg C L-1). The lowermost monimolimnion had marine salinity (22-24%) and very high concentrations of sulfide (up to 18 mmol L-1) and CH4 (up to 1.8 mmol L-1). In the chemocline, total microbial abundance and the rate of anoxygenic photosynthesis were 8.8 × 106 cells mL-1 and 34.4 µmol C L-1 day-1, respectively. Both in March and September, sulfate reduction rate increased with depth, peaking (up to 0.6-1.1 µmol S L-1 day-1) in the lower chemocline. Methane oxidation rates in the chemocline were up to 85 and 180 nmol CH4 L-1 day-1 in March and September, respectively; stimulation of this process by light was observed in September. The percentages of cyanobacteria and methanotrophs in the layer where light-induced methane oxidation occurred were similar, ∼2.5% of the microbial community. Light did not stimulate methane oxidation in deeper layers. The carbon isotope composition of particulate organic matter (δ13C-Corg), dissolved carbonates (δ13C-DIC), and methane (δ13C- CH4) indicated high microbial activity in the chemocline. Analysis of the 16S rRNA gene sequences revealed predominance of Cyanobium cyanobacteria (order Synechococcales) in the mixolimnion. Green sulfur bacteria Chlorobium phaeovibrioides capable of anoxygenic photosynthesis constituted ∼20% of the chemocline community both in March and in September. Methyloprofundus gammaptoteobacteria (family Methylomonaceae) were present in the upper chemocline, where active methane oxidation occurred. During winter, cyanobacteria were less abundant in the chemocline, while methanotrophs occurred in higher horizons, including the under-ice layer. Chemolithotrophic gammaproteobacteria of the genus Thiomicrorhabdus, oxidizing reduced sulfur compounds at low oxygen concentrations, were revealed in the chemocline in March. Both in March and September archaea constituted up to 50% of all microorganisms in the hypolimnion. The percentage of putative methanogens in the archaeal community was low, and they occurred mainly in near-bottom horizons.

Microbial diversity and autotrophic activity in Kamchatka hot springs.

Microbial communities of Kamchatka Peninsula terrestrial hot springs were studied using molecular, radioisotopic and cultural approaches. Analysis of 16S rRNA gene fragments performed by means of high-throughput sequencing revealed that aerobic autotrophic sulfur-oxidizing bacteria of the genus Sulfurihydrogenibium (phylum Aquificae) dominated in a majority of streamers. Another widely distributed and abundant group was that of anaerobic bacteria of the genus Caldimicrobium (phylum Thermodesulfobacteria). Archaea of the genus Vulcanisaeta were abundant in a high-temperature, slightly acidic hot spring, where they were accompanied by numerous Nanoarchaeota, while the domination of uncultured Thermoplasmataceae A10 was characteristic for moderately thermophilic acidic habitats. The highest rates of inorganic carbon assimilation determined by the in situ incubation of samples in the presence of 14C-labeled bicarbonate were found in oxygen-dependent streamers; in two sediment samples taken from the hottest springs this process, though much weaker, was found to be not dependent on oxygen. The isolation of anaerobic lithoautotrophic prokaryotes from Kamchatka hot springs revealed a wide distribution of the ability for sulfur disproportionation, a new lithoautotrophic process capable to fuel autonomous anaerobic ecosystems.

Microbial processes of the carbon and sulfur cycles in an ice-covered, iron-rich meromictic lake Svetloe (Arkhangelsk region, Russia).

Biogeochemical, isotope geochemical and microbiological investigation of Lake Svetloe (White Sea basin), a meromictic freshwater was carried out in April 2014, when ice thickness was ∼0.5 m, and the ice-covered water column contained oxygen to 23 m depth. Below, the anoxic water column contained ferrous iron (up to 240 µµM), manganese (60 µM), sulfide (up to 2 µM) and dissolved methane (960 µM). The highest abundance of microbial cells revealed by epifluorescence microscopy was found in the chemocline (redox zone) at 23-24.5 m. Oxygenic photosynthesis exhibited two peaks: the major one (0.43 µmol C L-1  day-1 ) below the ice and the minor one in the chemocline zone, where cyanobacteria related to Synechococcus rubescens were detected. The maximum of anoxygenic photosynthesis (0.69 µmol C L-1  day-1 ) at the oxic/anoxic interface, for which green sulfur bacteria Chlorobium phaeoclathratiforme were probably responsible, exceeded the value for oxygenic photosynthesis. Bacterial sulfate reduction peaked (1.5 µmol S L-1  day-1 ) below the chemocline zone. The rates of methane oxidation were as high as 1.8 µmol CH4  L-1  day-1 at the oxi/anoxic interface and much lower in the oxic zone. Small phycoerythrin-containing Synechococcus-related cyanobacteria were probably involved in accumulation of metal oxides in the redox zone.

Microbial life in Bourlyashchy, the hottest thermal pool of Uzon Caldera, Kamchatka.

Bourlyashchy is the largest and hottest pool in the Uzon Caldera, located in the territory of Kronotsky Nature Reserve, Kamchatka, Russia, with sediment surface temperatures at the margins ranging from 86 to 97 °C, and pH from 6.0 to 7.0. The microbial communities of the pool water and sediments were studied comprehensively from 2005 to 2014. Radioisotopic tracer studies revealed the processes of inorganic carbon assimilation, sulfate reduction, lithotrophic methanogenesis and potentially very active process of acetate oxidation to CO2. The total number of microbial cells in water was different in different years ranging from 5.2 to 7.0 × 10(6); in sediments, it changed from year to year between 6.3 × 10(6) and 1.75 × 10(8), increasing with a decrease in temperature. FISH with Archaea- and Bacteria-specific probes showed that the share of Bacteria differed with year, changing from 34 to 71%. According to 16S rRNA gene pyrosequencing data, lithoautotrophs (Aquificales and Thermoproteales) predominated in water samples, while in sediments they shared the niche with organotrophic Crenarchaeota, Korarchaeota, and bacteria of the genus Caldimicrobium (phylum Thermodesulfobacteria). The majority of organisms in water belonged to cultivated orders of prokaryotes; the only large uncultured group was that representing a novel order in class Thermoprotei. In sediments, unclassified Aquificeae comprised a significant part of the bacterial population. Thus, we showed that the hottest of the terrestrial hot pools studied contains numerous and active microbial populations where Bacteria represent a significant part of the microbial community, and planktonic and sediment populations differ in both composition and function.

Anaerobic transformation of carbon monoxide by microbial communities of Kamchatka hot springs.

Carbon monoxide (CO) is one of the common gaseous compounds found in hot volcanic environments. It is known to serve as the growth substrate for a number of thermophilic prokaryotes, both aerobic and anaerobic. The goal of this work was to study the process of anaerobic transformation of CO by microbial communities inhabiting natural thermal environments: hot springs of Uzon Caldera, Kamchatka. The anaerobic microbial community of Treshchinny Spring (80°C, pH 6.5) was found to exhibit two peaks of affinity for CO (K (S1) = 54 nM and K (S2) = 1 µM). The actual rate of anaerobic CO transformation by the microbial community of this spring, calculated after obtaining the concentration dependence curve and extrapolated to the natural concentration of CO dissolved in the hot spring water (20 nM), was found to be 120 µmol l(-1) of sediment day(-1). In all the hot springs studied, more than 90% of the carbon of (14)CO upon anaerobic incubation was recovered as (14)CO(2). From 1 to 5% of (14)CO was transformed to volatile fatty acids (VFA). The number of microorganisms capable of anaerobic CO oxidation determined by dilution-to-extinction method reached 10(6) cells ml(-1) of sediment. CO-transforming anaerobic thermophilic microorganisms isolated from the springs under study exhibited hydrogenogenic type of CO oxidation and belonged to the bacterial genera Carboxydocella and Dictyoglomus. These data suggest a significant role of hydrogenogenic carboxydotrophic prokaryotes in anaerobic CO transformation in Uzon Caldera hot springs.

Activity and distribution of thermophilic prokaryotes in hydrothermal fluid, sulfidic structures, and sheaths of alvinellids (East Pacific Rise, 13°N).

Processes of inorganic carbon assimilation, methanogenesis, sulfate reduction, and acetate oxidation to CO(2) occurring in samples from the East Pacific Rise at 13°N were traced, using radioisotopically labeled substrates, at temperatures ranging from 65 to 100°C. Molecular hydrogen stimulated lithotrophic methanogenesis and sulfate reduction but inhibited inorganic carbon assimilation. Active mineralization of acetate was observed in an organic-rich Alvinella-associated system at 80°C. Members of the Thermococcales were the most numerous hyperthermophilic archaea in these samples, their density achieving 10(8) cells per cm(3), while the numbers of cultured hydrogen-utilizing thermophilic lithotrophs were several orders of magnitude lower.

Sulfidogenesis under extremely haloalkaline conditions in soda lakes of Kulunda Steppe (Altai, Russia).

Sulfidogenic activity (SA) in anoxic sediments of several soda lakes with variable salinity in south Kulunda Steppe (Altai, Russia) has been investigated. The study included in situ measurements of sulfate reduction rates and laboratory experiments with sediment slurries in which sulfate, thiosulfate or elemental sulfur were used as electron acceptors. Despite the extreme conditions (high salt concentrations and high pH), the SA values were relatively high (ranging from 0.02 to 1.20 micromol HS(-) cm(-3) h(-1)), and only hampered under salt-saturated conditions. The highest SA was observed with elemental sulfur, followed by thiosulfate, while the lowest SA was determined in the presence of sulfate. Of all the electron donors tested, the addition of formate resulted in the highest SA with all three sulfur electron acceptors. Surprisingly, hydrogen as an electron donor had very little effect. Acetate was utilized as an electron donor only under sulfur-reducing conditions. Indigenous populations of sulfidogens in soda lake sediments showed an obligately alkaliphilic pH response of SA, showing a pattern that corresponded well to the in situ pH conditions. Sulfate reduction was much more susceptible to salt inhibition than thiosulfate and sulfur reduction. Microbiological investigations indicated that sulfate-reducing bacteria belonging to the orders Desulfovibrionales and Desulfobacterales could very likely be responsible for the SA with sulfate and thiosulfate as electron acceptors at moderate salt concentrations. Sulfur reduction at moderate salinity was carried out by a specialized group of haloalkaliphilic sulfur-reducing bacteria that utilize volatile fatty acids. In saturated soda brine, extremely natronophilic representatives of the order Halanaerobiales were responsible for the sulfur-dependent respiration.