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The review deals with the unique microbial group responsible for anaerobic ammonium oxidation with nitrite (anammox), and with the role of this process in development of the biotechnology for removal of nitrogen compounds from wastewater. The history of the study of this process is briefly related. Up-to date knowledge on the intracellular organization, energy metabolism, growth stoichiometry, and physiology of anammox bacteria is described, and the main methods for cultivation of these microorganisms are characterized. Special attention is paid to the problems associated with practical application of anammox bacteria, which result from their extremely slow growth, the absence of pure cultures, and the interaction with other microbial groups.

Hydroasoustic techniques were used for detection and mapping of gas jet areas in the coastal regions of the Crimean peninsula. Gas seep areas in the bays Laspi, Khersones, and Kazach'ya were chosen for detailed microbiological investigation. The first type of gas jets, observed in the Laspi Bay, was probably associated with discarge of deep thermogenic methane along the faults. Methane isotopic composition was char- acterized by Δ13C of -35.3 degrees. While elevated rates of aerobic methane oxidation were revealed in the sandy sediments adjacent to the methane release site, no evidence of bacterial mats was found. The second type of gas emission, observed in the Khersones Bay, was accompanied by formation of bacterial biofilms of the "Thiodendron" microbial community type, predominated by filamentous, spirochete-like organisms, in the areas of gas seepage. The isotopic composition of methane was there considerably lower (-60.4 degrees), indicating a considerable contribution of modern microbial methane to the gas bubbles discharged in this bay. Activity of the third type of gas emission, the seeps of the Kazach'ya Bay, probably depended directly on modern microbial processes of organic matter degradation in the upper sediment layers. The rates of sulfate reduction and methanogenesis were 260 and 34 μmol dm(-3) day(-1), respectively. Our results indicate different mechanisms responsible for formation of methane jets in the Laspi Bay and in the coastal areas of the Heracles Peninsula, where the bays Kazach'ya and Khersones are located.

Dissimilatory sulfate reduction plays an important role in removal of dissolved metals from acidic mine waters. Although this process was convincingly shown to occur in acidic waste of metal recovery, few isolates of acid-tolerant sulfate rducers are known. We isolated a new acidophilic sulfidogen, strain BG, from the oxidized acidic waste of the Bom-Gorkhon tungsten deposit, Transbaikalia, Russia. Phylogenetic analysis of its 16S rRNA gene sequence made it possible to identify it as a member of the genus Desulfosporosinus. Unlike other known acidophilic sulfate reducers of this genus, strain BG was tolerant to high copper concentrations (up to 5 g/L), could grow on organic acids at low ambient pH, and formed crystalline copper sulfides (covellite and chalcopyrite). Molecular analysis of the phenotypes predominating in oxidized waste and in enrichment cultures confirmed the presence of various Desulfosporosinus strains.

Biodiversity of sulfate-reducing bacterial communities in the water column of the Gdansk Deep, Baltic Sea, where H2S had been detected in near-bottom layers, was analyzed by PCR with primers for the 16S rRNA genes of six major phylogenetic subgroups of sulfate-reducing bacteria (SRB). Using denaturing gradient gel electrophoresis followed by sequencing, the nucleotide sequences of reamplified dsrB gene fragments from investigated water samples were determined. For the first time the presence of nucleotide sequences of the dsrB gene was detected by PCR in the water samples from all hydrochemical layers, including subsurface oxic waters. The presence of the 16S rRNA genes of representatives of Desulfotomaculum, Desulfococcus-Desulfonema-Desulfosarcina, and Desulfovibrio-Desulfomicrobium SRB subgroups was also revealed throughout the water column of the Gdansk Deep. Analysis of translated amino acid sequences encoded by the dsrB gene demonstrated the highest homology with the relevant sequences of uncultured SRB from various marine habitats.

A new species of bacteria oxidizing ammonium with nitrite under anoxic conditions was isolated from the activated sludge of a semi-industrial bioreactor treating digested sludge of the Kuryanovo wastewater treatment plant (Moscow, Russia). Physiological, morphological, and molecular genetic characterization of the isolate was carried out. The cells were ovoid (-0.5 x 0.8 μm), with the intracellular membrane structures characteristic of anammox bacteria (anammoxosome and paryphoplasm); unlike other anammox bacteria, it possessed extensive intracellular membrane structures located in layers parallel to the cytoplasmic membrane, but never close to the anammoxosome. The cells formed aggregates 5-28 μm in diameter and readily attached to solid surfaces. The cells were morphologically labile, easily plasmolyzed, and lost their content. Doubling time was 28 days, μ(max) = 0.025 day(-1); optimal temperature and pH for growth were 20-45 degrees C and 8.0, respectively. Phylogenetic analysis of the 16S rRNA gene sequences suggested its classification as a new species of the candidate genus Jettenia (order Planctomycetales). The name Candidatus "Jettenia moscovienalis" sp. nov. was proposed for the new bacterium.

Microbiological and biogeochemical investigation of the water column of oligotrophic Lake Baikal at the sites of the K2 and Bolshoy mud volcanoes and the Gorevoy Utes oil seep was carried out in July 2013. Total microbial numbers (TMN), cell numbers of type I and type II methanotrophs, and methane concentrations were measured; the rate of methane oxidation was determined. Methane concentrations in Lake Baikal water column varied from 0.09 to 1 μL/L, while methane oxidation rates varied from 0.007 to 0.9 nL/(L day). The highest rates of methane oxidation were revealed in the near-bottom water horizons at the sites of the Bolshoy mud volcano and the Gorevoy Utes oil seep. These were the sites where the most pronounced anomalies in methane concentration were also detected. TMN varied from 0.123 x 10(6) to 1.64 x 10(6) cells/mL. Methanotrophic bacteria were revealed in the water column at all sites, their abundance did not always correlate with methane concentrationsand the rates of methane oxidation. Methanotrophs constituted not more than 1.63% of the total microbial number, with their highest abundance in the upper 200 m of the water column.

Production of gaseous hydrocarbons by the microbial community of the Posolsky Bank methane seep bottom sediments (Southern Baikal) was studied at 4°C. Formation of both methane and a heavier gas- eous hydrocarbon, ethane, was detected in enrichment cultures. The highest methane concentrations (6.15 and 4.51 mmol L(-1)) were revealed in enrichments from the sediments from 55-cm depth incubated with-so- dium acetate and H2/CO2 gas mixture, respectively. A decrease in activity of aceticlastic methanogensand a decrease in methane concentration produced by hydrogenotrophic archaea occurred with depth. The highest concentration of ethane was revealed in enrichments from the microbial community of the layer close to gas hydrates (75 cm) incubated with CO2 as a substrate. According to analysis of the 16S rRNA gene fragments from the clone library, these enrichments were found to contain members of the phylum Crenarchaeota form- ing a separate cluster with members of the class Thermoprotei. The phylum Euryarchaeota was represented by nucleotide sequences of the organisms homologous to members of the orders Methanococcales, Methanosa- rcinales, and Thermoplasmatales.

The contribution of the major technologically important microbial groups (ammonium- and nitrite-oxidizing, phosphate-accumulating, foam-inducing, and anammox bacteria, as well as planctomycetes and methanogenic archaea) was characterized for the aeration tanks of the Moscow wastewater treatment facilities. FISH investigation revealed that aerobic sludges were eubacterial communities; the metabolically active archaea contributed insignificantly. Stage II nitrifying microorganisms and planctomycetes were significant constituents of the bacterial component of activated sludge, with Nitrobacter spp. being the dominant nitrifier. No metabolically active anammox bacteria were revealed in the sludge from aeration tanks. The sludge from the aeration tanks using different wastewater treatment technologies were found to differ in characteristics. Abundance of the nitrifying and phosphate-accumulating bacteria in the sludges generally correlated with microbial activity, in microcosms and with efficiency of nitrogen and phosphorus removal from wastewater. The highest microbial numbers and activity were found in the sludges of the tanks operating according to the technologies developed in the universities of Hanover and Cape Town. The activated sludge from the Novokur yanovo facilities, where abundant growth of filamentous bacteria resulted in foam formation, exhibited the lowest activity The group of foaming bacteria included Gordonia spp. and Acinetobacter spp., utilizing petroleum and motor oils, Sphaerotilus spp. utilizing unsaturated fatty acids, and Candidatus 'Microthrix parvicella'. Thus, the data on abundance and composition of metabolically active microorganisms obtained by FISH may be used for the technological control of wastewater treatment.

The rates of sulfate reduction (SR) and the diversity of sulfate-reducing bacteria (SRB) were studied in the sediments of the Posol'skaya banka elevation in the southern part of Lake Baikal. SR rates varied from 1.2 to 1641 nmol/(dm3 day), with high rates (> 600 nmol/(dm3 day)) observed at both deep-water stations and in subsurface silts. Integral SR rates calculated for the uppermost 50 cm of the sediments were higher for gas-saturated and gas hydrate-bearing sediments than in those with low methane content. Enrichment SRB cultures were obtained in Widdel medium for freshwater SRB. Analysis of the 16S rRNA gene fragments from clone libraries obtained from the enrichments revealed the presence of SRB belonged to Desulfosporosinus genus, with D. lacus as the most closely related member (capable of sulfate, sulfite, and thiosulfate reduction), as well as members of the order Clostridiales.

The community of anoxygenic phototrophic bacteria (APB) in the water column of the Kislo-Sladkoe stratified lake recently isolated from the sea (White Sea, Kandalaksha Bay) was investigated in September 2010. The water of the sulfide-rich zone was greenish-brown due to intense development of green sulfur bacteria (GSB). Nine APB strains were isolated from the water samples: three belonging to GSB, five, to purple sulfur bacteria (PSB), and one, to purple nonsulfur bacteria (PNB). GSB predominated in the phototrophic community of the chemocline. Unexpectedly, two morphologically different green-colored GSB strains were found to be phylogenetically identical and related to the brown-colored @Chlorobium phaeovibrioides (99% similarity according to the 16S rRNA gene sequencing). Homology to the closest green-colored species (Chlorobium luteolum) was 98%. Two morphologically and physiologically similar PSB strains (TcrPS10 and AmPS10) had rounded cells containing okenonokenonee and gas vesicles. According to the 16S rRNA gene sequencing, these strains were most closely related (99%) to two different Thiocapsa species: Tca. marina (containing okenonokenonee and no gas vesicles) and Tca. rosea (containing spirilloxanthin and gas vesicles). The remaining isolates of purple bacteria were similar to the already described APB species.

The rates of microbial processes of sulfate reduction and of the methane cycle were measured in the bottom sediments of the Sevastopol basin, where seeps of gaseous methane have been previously found. Typically for marine environments, sulfate reduction played the major role in the terminal phase of decomposition of organic matter (OM) in reduced sediments of this area. The rate of this process depended on the amount of available OM. The rate of methanogenesis in the sediments increased with depth, peaking in the subsurface horizons, where decreased sulfate concentration was detected in the pore water. The highest rates of sulfate-dependent anaerobic methane oxidation were found close to the methane-sulfate transition zone as is typical of most investigated marine sediments. The data on the carbon isotopic composition of gaseous methane from the seeps and dissolved CH4 from the bottom sediments, as well as on the rates of microbial methanogenesis and methane oxidation indicate that the activity of the methane seeps results from accumulation of biogenic methane in the cavities of the underlying geological structures with subsequent periodic release of methane bubbles into the water column.

Mass-spectrometric investigation of carbon isotope composition (delta13C) was carried out for suspended organic matter and dissolved mineral compounds for the water column of some meromictic water bodies differing in salinity and trophic state. As a rule, a more pronounced carbon isotope fractionation (resulting from the metabolism of phytoplankton and anoxygenic phototrophic bacteria) was revealed in the zones of enhanced oxygenic and anoxygenic photosynthesis. Carbon isotope fractionation at the border between oxidized and reduced waters depends both on the activity of microbial communities and on the dominant species of phototrophic microorganisms. Analysis of the distribution profiles of the isotopic composition of suspended organic matter and dissolved mineral carbon revealed active mineralization of the organic matter newly formed via anoxygenic photosynthesis in the monimolimnion by microbial communities, resulting in the release of isotopically light carbon dioxide. Mineral carbon in the anaerobic zones of highly productive meromictic water bodies is therefore enriched with the light 12C isotope.

Comprehensive microbiological and biogeochemical investigation of a pockmark within one of the sites of gas-saturated sediments in the Gdansk depression, Baltic Sea was carried out during the 87th voyage of the Professor Shtokman research vessel. Methane content in the near-bottom water and in the underlying sediments indicates stable methane flow from the sediment into the water. In the 10-m water layer above the pockmark, apart from methane anomalies, elevated numbers of microorganisms and enhanced rates of dark CO2 fixation (up to 1.15 micromol C/(1 day)) and methane oxidation (up to 2.14 nmol CH4/(1 day)) were revealed. Lightened isotopic composition of suspended organic matter also indicates high activity of the near-bottom microbial community. Compared to the background stations, methane content in pockmark sediments increased sharply from the surface to 40-60 ml/dm3 in the 20-30cm horizon. High rates of bacterial sulfate reduction (SR) were detected throughout the core (0-40 cm); the maximum of 74 micromol/(dm3 day) was located in subsurface horizons (15-20 cm). The highest rates of anaerobic methane oxidation (AMO), up to 80 micromol/(dm3 day), were detected in the same horizon. Good coincidence of the AMO and SR profiles with stoichiometry close to 1:1 is evidence in favor of a close relation between these processes performed by a consortium of methanotrophic archaea and sulfate-reducing bacteria. Methane isotopic composition in subsurface sediments of the pockmark (from -53.0 to -56.5% per hundred) does not rule out the presence of methane other than the biogenic methane from the deep horizons of the sedimentary cover.

The anoxygenic phototrophic bacterial community of the high-altitude meromictic Lake Gek-Gel (Azerbaijan) was investigated in September 2003. The highest concentration of bacteriochlorophyll e (48 microg/l) was detected at a depth of 30 m; the peak of bacteriochlorophyll a (4.5 microg/l) occurred at 29 m. Phylogenetic analysis revealed that brown-colored green sulfur bacteria Chlorobium phaeobacteroides predominated in the lake. Nonsulfur purple bacteria phylogenetically close to Blastochloris sulfoviridis were found in insignificant amounts; these organisms have not been previously reported in Lake Gek-Gel.

The possibility of measuring the rates of light and dark CO2 assimilation using 13C carbonate was demonstrated on Lake Kichier (Marii El). The application of methods utilizing the stable 13C and the radioactive 14C isotopes resulted in comparable values of the rates of light and dark CO2 fixation. Due to its absolute environmental safety, the method with 13C mineral carbon can be recommended as an alternative to radioisotope methods for qualitative measurements of CO2 fixation rates in aquatic ecosystems.

The research performed in August 2004 within the framework of the Russian-American Long-term Census of the Arctic (RUSALCA) resulted in the first data concerning the rates of the key microbial processes in the water column and bottom sediments of the Bering strait and the Chukchi Sea. The total bacterial counts in the water column varied from 30 x 10(3) cells ml(-1) in the northern and eastern parts to 245 x 10(3) cells ml(-1) in the southern part. The methane content in the water column of the Chukchi sea varied from 8 nmol CH4 l(-1) in the eastern part of the sea to 31 nmol CH4 l(-1) in the northern part of the Herald Canyon. Active microbial processes occurred in the upper 0-3 cm of the bottom sediments; the methane formation rate varied from 0.25 to 16 nmol CH4 dm(-3) day(-1). The rates of methane oxidation varied from 1.61 to 14.7 nmol CH4 dm(-3) day(-1). The rates of sulfate reduction varied from 1.35 to 16.2 micromol SO4(2-) dm(-3) day(-1). The rate of methane formation in the sediments increased with depth, while sulfate reduction rates decreased (less than 1 micromol SO4(2-) dm(-3) day(-1)). These high concentrations of biogenic elements and high rates of microbial processes in the upper sediment layers suggest a specific type of trophic chain in the Chukchi Sea. The approximate calculated balance of methane emission from the water column into the atmosphere is from 5.4 to 57.3 micromol CH4 m(-2) day(-1).

The rate of CO conversion by a pure culture of a thermophilic CO-oxidizing, H2-producing bacterium Carboxydocella sp. strain 1503 was determined by the radioisotopic method. The overall daily uptake of 14CO by the bacterium was estimated at 38-56 micromol CO per 1 ml of the culture. A radioisotopic method was developed to separate and quantitatively determine the products of anaerobic CO conversion by microbial communities in hot springs. The new method was first tested on the microbial community from a sample obtained from a hot spring in Kamchatka. The potential rate of CO conversion by the anaerobic microbial community was found to be 40.75 nmol CO/cm3 sediment per day. 85% of the utilized 14CO was oxidized to carbon dioxide; 14.5% was incorporated into dissolved organic matter, including 0.2% that went into volatile fatty acids; 0.5% was used for cell bio mass production; and only just over 0.001% was converted to methane.

The anoxygenic phototrophic bacterial community of the brackish meromictic Lake Shira (Khakassia) was investigated in August 2001, July 2002, and February-March 2003. In all the periods of investigation, the prevailing microorganisms were purple sulfur bacteria similar to Lamprocystis purpurea in morphology and pigment composition. Their highest number (3 x 10(5) cells/ml) was recorded in July 2002 at the depth of 15 m. According to 16S rRNA gene analysis, the strain of purple sulfur bacteria isolated in 2001 and designated ShAm01 exhibited 98.6% similarity to the type strain of Thiocapsa roseopersicina and 94.4-97.1% similarity to the type strains of Tca. pendens, Tca. litoralis, and Tea. rosea. The minor microorganisms of the anoxygenic phototrophic bacterial community within the period of investigation were nonsulfur purple bacteria phylogenetically close to Rhodovulum strictum (98.3% similarity, strain ShRb01), Ahrensia kielensis (of 93.9% similarity, strain ShRb02), Rhodomicrobium vannieli (of 99.7% similarity, strain ShRmc01), and green sulfur bacteria, phylogenetically close to Chlorobium limicola (of 98.7% similarity, strain ShCl03).

Seasonal studies of the anoxygenic phototrophic bacterial community of the water column of the saline eutrophic meromictic Lake Shunet (Khakassia) were performed in 2002 (June) and 2003 (February-March and August). From the redox zone down, the lake water was of dark green color. Green sulfur bacteria predominated in every season. The maximum number of green sulfur bacteria was 10(7) cells/ml in summer and 10(6) cells/ml in winter. A multi-syringe stratification sampler was applied for the study of the fine vertical distribution of phototrophs in August 2003; the sampling was performed every five centimeters. A five-centimeter-thick pink-colored water layer inhabited by purple sulfur bacteria was shown to be located above the layer of green bacteria. The species composition and ratio of purple bacterial species depended on the sampling depth and on the season. In summer, the number of purple sulfur bacteria in the layer of pink water was 1.6 x 10(8) cells/ml. Their number in winter was 3 x 10(5) cells/ml. In the upper oxygen-containing layer of the chemocline the cells of purple nonsulfur bacteria were detected in summer. The maximum number of nonsulfur purple bacteria, 5 x 10(2) cells/ml, was recorded in August 2003. According to the results of the phylogenetic analysis of pure cultures of the isolated phototrophic bacteria, which were based on 16S rDNA sequencing, green sulfur bacteria were close to Prosthecochloris vibrioformis, purple sulfur bacteria, to Thiocapsa and Halochromatium species, and purple nonsulfur bacteria, to Rhodovulum euryhalinum and Pinkicyclus mahoneyensis.

In order to assess the phylogenetic diversity of the endosymbiotic microbial community of the gills of marine shellfish Bathymodiolus azoricus, total DNA was extracted from the gills. The PCR fragments corresponding to the genes encoding 16S rRNA, ribulose-bisphosphate carboxylase (cbbL), and particulate methane monooxygenase (pmoA) were amplified, cloned, and sequenced. For the 16S rDNA genes, only one phylotype was revealed; it belonged to the cluster of Mytilidae thiotrophic symbionts within the Gammaproteobacteria. For the RuBisCO genes, two phylotypes were found, both belonging to Gammaproteobacteria. One of them was closely related to the previously known mytilid symbiont, the other, to a pogonophore symbiont, presumably a methanotrophic bacterium. One phylotype of particulate methane oxygenase genes was also revealed; this finding indicated the presence of a methanotrophic symbiont. Phylogenetic analysis of the pmoA placed this endosymbiont within the Gammaproteobacteria, in a cluster including the methanotrophic bacterial genus Methylobacter and other methanotrophic Bathymodiolus gill symbionts. These results provide evidence for the existence of two types of endosymbionts (thioautotrophic and methanotrophic) in the gills of B. azoricus and demonstrate that, apart from the phylogenetic analysis of 16S rRNA genes, parallel analysis of functional genes is essential.

The biogeochemical and molecular biological study of the chemocline and sediments of saline meromictic lakes Shira and Shunet (Khakass Republic, Russia) was performed. A marked increase in the rates of sulfate reduction and methanogenesis was revealed at the medium depths of the chemocline. The rates of these processes in the bottom sediments decreased with depth. The numbers of Bacteria, Archaea, and of sulfate-reducing bacteria (SRB) were determined by fluorescence in situ hybridization with rRNA specific oligonucleotide probes labeled with horseradish peroxidase and subsequent tyramide signal amplification. In the chemocline, both the total microbial numbers and those of Bacteria were shown to increase with depth. The archaea and SRB were present in almost equal numbers. In the lake sediments, a drastic decrease in microbial numbers with depth was revealed. SRB were found to prevail in the upper sediment layer and archaea in the lower one. This finding correlates with the measured rates of sulfate reduction and methanogenesis.

The distribution and activity of sulfate-reducing bacteria (SRB) in the water column of the alpine meromictic Gek-Gel lake were studied. Apart from traditional microbiological methods based on cultivation and on measuring the process rates with radioactive labels, in situ fluorescent hybridization (FISH) was used, which enables identification and quantification without cultivating organisms. The peak rate of sulfate reduction, 0.486 microg S/(l day), was found in the chemocline at 33 m. The peak SRB number of 2.5 x 106 cells/ml, as determined by the end-point dilutions method on selective media, was found at the same depth. The phylogenetic position of the SRB, as determined by FISH, revealed the predominance of the Desulfovibrio spp., Desulfobulbus spp., and Desulfoarculus spp./Desulfomonile spp. groups. The numbers of spore-forming Desulfotomaculum spp. increased with depth. The low measured rates of sulfate reduction accompanied with high SRB numbers and the predominance of the groups capable of reducing a wide range of substrates permit us to propose utilization of electron acceptors other than sulfate as the main activity of the SRB in the water column.

An anaerobic phototrophic bacterial community in Lake Mogilnoe, a relict lake on Kil'din Island in the Barents Sea, was studied in June 1999 and September 2001. Irrespective of the season, the upper layer of the anaerobic zone of this lake had a specific species composition of sulfur phototrophic bacteria, which were dominated by the brown-colored green sulfur bacterium Chlorobium phaeovibrioides. The maximum number of phototrophic sulfur bacteria was observed in June 1999 at a depth of 9 m, which corresponded to a concentration of bacteriochlorophyll (Bchl) e equal to 4.6 mg/l. In September 2001, the maximum concentration of this pigment (3.4 mg/l) was found at a depth of 10 m. In both seasons, the concentration of Bchl a did not exceed 3 microg/l. Purple sulfur bacteria were low in number, which can be explained by their poor adaptation to the hydrochemical and optical conditions of the Lake Mogilnoe water. In June 1999, the water contained a considerable number of Pelodictyon phaeum microcolonies and Prosthecochloris phaeoasteroides cell chains, which was not the case in September 2001. A 16S rDNA-based phylogenetic analysis of pure cultures of phototrophic bacteria isolated from the lake water confirmed that the bacterial community is dominated by Chl. phaeovibrioides and showed the presence of three minor species, Thiocvstis gelatinosa, Thiocapsa sp., and Thiorhodococcus sp., the last of which is specific to Lake Mogilnoe.

Microbiological and isotopic-geochemical investigations of the brackish meromictic lakes Shira and Shunet were performed in the steppe region of Khakasia in winter. Measurements made with a submersed sensor demonstrated that one-meter ice transmits light in a quantity sufficient for oxygenic and anoxygenic photosynthesis. As in the summer season, in the community of phototrophic bacteria found in Lake Shira, the purple sulfur bacteria Amoebobacter purpureus dominated, whereas, in Lake Shunet, the green sulfur bacteria Pelodictyon luteolum were predominant. Photosynthetic production, measured using the radioisotopic method, was several times lower than that in summer. The rates of sulfate reduction and production and oxidation of methane in the water column and bottom sediments were also lower than those recorded in summer. The process of anaerobic methane oxidation in the sediments was an exception, being more intense in winter than in summer. The data from radioisotopic measurements of the rates of microbial processes correlate well with the results of determination of the isotopic composition of organic and mineral carbon (delta13C) and hydrogen sulfide and sulfate (delta34S) and suggest considerable seasonal variations in the activity of the microbial community in the water bodies investigated.

A detailed study of the processes of anaerobic methane oxidation and sulfate reduction in the bacterial mats occurring on coral-like carbonate structures in the region of methane seeps in the Black Sea, as well as of the phenotypic diversity of sulfate-reducing bacteria developing in this zone, has been performed. The use of the radioisotopic method shows the microbial mat structure to be heterogeneous. The peak activity of the two processes was revealed when a mixture of the upper (dark) and underlying (intensely pink) layers was introduced into an incubation flask, which confirms the suggestion that methanotrophic archaea and sulfate-reducing bacteria closely interact in the process of anaerobic methane oxidation. Direct correlation between the rate of anaerobic methane oxidation and the methane and electron acceptor concentrations in the medium has been experimentally demonstrated. Several enrichment and two pure cultures of sulfate-reducing bacteria have been obtained from the near-bottom water and bacterial mats. Both strains were found to completely oxidize the substrates to CO2 and H2S. The bacteria grow at temperatures ranging from -1 to 18 (24) degrees C, with an optimum in the 10-18 degrees C range, and require the presence of 1.5-2.5% NaCl and 0.07-0.2% MgCl2 x 6H2O. Regarding the aggregate of their phenotypic characteristics (cell morphology, spectrum of growth substrates, the capacity for complete oxidation), the microorganisms isolated have no analogues among the psychrophilic sulfate-reducing bacteria already described. The results obtained demonstrate the wide distribution of psychrophilic sulfate-reducing bacteria in the near-bottom water and bacterial mats covering the coral-like carbonate structures occurring in the region of methane seeps in the Black Sea, as well as the considerable catabolic potential of this physiological group of psychrophilic anaerobes in deep-sea habitats.

In May 1998, during the fifty-first voyage on board the research vessel Professor Vodyanitskii, a comparative study was conducted of the species diversity of green and purple sulfur bacteria in the water column of the chemocline zone at deep-sea stations and on the bottom surface of the Black Sea shallow regions. At three deep-sea stations, the accumulation of photosynthesizing bacteria in the chemocline zone at a depth of 85-115 m was revealed on the basis of the distribution of potential values of carbon dioxide light fixation. The location of the site of potential carbon dioxide light fixation suggests that the photosynthesis may be determined by the activity of the brown Chlorobium sp., revealed earlier at these depths. Enrichment cultures of brown sulfur bacteria were obtained from samples taken at the deep-sea stations. By morphology, these bacteria, assigned to Chlorobium sp., appear as nonmotile straight or slightly curved rods 0.3-0.5 x 0.7-1.2 microm in size; sometimes, they form short chains. Ultrathin sections show photosynthesizing antenna-like structures, chlorosomes, typical of Chlorobiaceae. The cultures depended on the presence of NaCl (20 g/l) for growth, which corresponds to the mineralization of Black Sea water. The bacteria could grow photoautotrophically, utilizing sulfide, but the Black Sea strains grew much more slowly than the known species of brown sulfur bacteria isolated from saline or freshwater meromictic lakes. The best growth of the strains studied in this work occurred in media containing ethanol (0.5 g) or sodium acetate (1 g/l) and low amounts of sulfide (0.4 mM), which is consistent with the conditions of syntrophic growth with sulfidogens. The data obtained allow us to conclude that the cultures of brown sulfur bacteria are especially adapted to developing at large depths under conditions of electron donor deficiency owing to syntrophic development with sulfate reducers. The species composition of the photosynthetic bacteria developing in the bottom sediments of shallow stations differed substantially from that observed at deep-sea stations. Pure cultures of the green Chlorobium sp. BS 1C and BS 2C (chlorobactin as the carotenoid), purple sulfur bacteria Chromatium sp. BS 1Ch (containing spirilloxanthine series pigments), and Thiocapsa marina BS 2Tc (containing the carotenoid okenone) were obtained from samples of sediments at shallow-water stations. Brown sulfur bacteria were absent in the sediment samples obtained from the Black Sea shallow-water stations 1 and 2.

Methane emission from the following types of tundra soils was studied: coarse humic gleyey loamy cryo soil, peaty gley soil, and peaty gleyey midloamy cryo soil of the arctic tundra. All the soils studied were found to be potential sources of atmospheric methane. The highest values of methane emission were recorded in August at a soil temperature of 8-10 degrees C. Flooded parcels were the sources of atmospheric methane throughout the observation period. The rates of methane production and oxidation in tundra soils of various types at 5 and 15 degrees C were studied by the radioisotope method. Methane oxidation was found to occur in bog water, in the green part of peat moss, and in all the soil horizons studied. Methane formation was recorded in the horizons of peat, in clay with plant roots, and in peaty moss dust of the bogey parcels. At both temperatures, the methane oxidation rate exceeded the rate of methane formation in all the horizons of the mossy-lichen tundra and of the bumpy sinkhole complex. Methanogenesis prevailed only in a sedge-peat moss bog at 15 degrees C. Enrichment bacterial cultures oxidizing methane at 5 and 15 degrees C were obtained. Different types of methanotrophic bacteria were shown to be responsible for methane oxidation under these conditions. A representative of type I methylotrophs oxidized methane at 5 degrees C, and Methylocella tundrae, a psychroactive representative of an acidophilic methanotrophic genus Methylocella, at 15 degrees C.

Microbiological and biogeochemical measurements showed that the intensities of CO2 assimilation, methane oxidation, and sulfate reduction in the Lost City vent field (30 degrees N) reach 3.8 microg C/(1 day), 0.06 microg C/(1 day), and 117 microg S/(1 day), respectively. On the surface of the carbonate structures occurring in this field, two varieties of bacterial mats were found. The first variety, which is specific to the Lost City alkaline vent field, represents jelly bacterial mats dominated by slime-producing bacteria of several morphotypes. This mat variety also contains chemolithotrophic and heterotrophic microorganisms, either microaerobic or anaerobic. The intensities of CO2 assimilation, methane oxidation, and sulfate reduction in this variety reach 747 microg C/(dm3 day), 0.02 microg C/(dm3 day), and 28,000 microg S/(dm3 day), respectively. Bacterial mats of the second variety are formed by nonpigmented filamentous sulfur bacteria, which are close morphologically to Thiothrix. The intensities of CO2 assimilation, methane oxidation, and sulfate reduction in the second mat variety reach 8.2 microg C/(dm3 day), 5.8 microg C/(dm3 day), and 17,000 microg S/(dm3 day), respectively. These data suggest the existence of subsurface microflora in the Lost City vent field.

Microbiological and biogeochemical investigations of the processes of methane production (MP) and methane oxidation (MO) in the coastal waters and littoral of the Kandalaksha Bay of the White Sea were carried out. The studies were conducted in the coastal zones and in the water areas of the Kandalaksha Preserve, Moscow University White Sea Biological Station, and Zoological Institute (RAS) Biological Station in August, 1999, 2000, and 2001 and in March, 2001. The rate of CO2 assimilation in the shallow and littoral sediments was 35-27800 microg C/(dm3 day) in summer and 32.8-88.9 microg C/(dm3 day) in winter. The maximal rates of MP were observed in the littoral sediments in the zone of macrophyte decomposition, in local depressions, and in the estuary of a freshwater creak (up to 113 microl/(dm3 day)). The maximal level of MO was observed in the shallow estuarine sediments (up to 2450 microl/(dm3 day)). During the winter season, at the temperature of -0.5 to 0.5 degrees C, the MP rate in the littoral sediments was 0.02-0.3 microl/(dm3 day), while MO rate was 0.06-0.7 microl/(dm3 day). The isotopic data obtained indicate that the C(org) of the mats and of the upper sediment layers is enriched with the heavy 13C isotope by 1-4 per thousand as compared to the C(org) of the suspension, comprised on 33.5-34.3% of phytoplankton. A striking difference was found between the levels of methane emission by the typical littoral microlanscapes. In fine sediments, the average emission was 675 microl CH4/(m2 day), in the stormy discharge stretch sediments it was 1670 microl CH4/(m2 day), and under the stones and in silted pits, 1370 microl CH4/(m2 day). The calculation performed with consideration of the microlandscape areas with a high production allowed the CH4 production of 1 km2 of the littoral to be estimated as 192-300 1 CH4/(km2 day).

Microbiological and biogeochemical investigations of the coastal zone and the littoral of the Kandalaksha Bay of the White Sea were carried out. The material for investigations was obtained in the series of expeditions of the Institute of Microbiology, Russian Academy of Sciences, in August 1999, 2000, 2001, and in March 2003. The studies were conducted on the littoral and in the water area of the Kandalaksha Preserve, the Moscow University Belomorsk Biological Station, and the Zoological Institute Biological Station, Russian Academy of Sciences, Sediment sampling on the littoral was carried out in the typical microlandscapes differing in the sediment properties and macrobenthos distribution. The maximal sulfate reduction rate (SRR) was shown for the shallow part of the Chemorechenskaya Bay (up to 2550 micrograms S/(dm3 day)) and in the Bab'ye More Bay (up to 3191 micrograms S/(dm3 day)). During the winter season, at a temperature of -0.5-0.5 degrees C, the SRR in the sediments of the Kartesh Bay was 7.9-13 micrograms S/(dm3 day). In the widest limits, the SRR values varied in the sediment cores sampled on the littoral. The minimal values (11 mu]g S/(dm3 day)) were obtained in the core samples on the silt-sandy littoral. The littoral finely dispersed sediments rich in organic matter were characterized by high SRR values (524-1413 micrograms S/(dm3 day)). The maximal SRR values were shown for the sediments present within the stretch of decomposing macrophytes, in local pits at the lower littoral waterline, and in the mouth of a freshwater stream (51-159 mg S/(dm3 day)). A sharp difference in the level of H2S production in the type microlandscapes was shown. The average hydrogen sulfide production in finely dispersed sediments constituted 125 mg S/(m2 day); in stormy discharge deposits, 1950 mg S/(m2 day); in depressions under stones and in silted pits, 4300 mg S/(m2 day). A calculation made with regard to the area of microlandscapes with increased productivity shows that the daily H2S production per 1 km2 of the littoral (August) is 60.8 to 202 kg S/(km2 day), while the organic carbon consumption for sulfate reduction per 1 km2 of the littoral is 46 to 152 kg C(org)/(km2 day).

Microbiological and biogeochemical studies of the meromictic saline Lake Shira (Khakasia) were conducted. In the upper part of the hydrogen-sulfide zone, at a depth of 13.5-14 m, there was a pale pink layer of water due to the development of purple bacteria (6 x 10(5) cells/ml), which were assigned by their morphological and spectral characteristics to Lamprocystis purpureus (formerly Amoebobacter purpurea). In August, the production of organic matter (OM) in Lake Shira was estimated to be 943 mg C/(m2 day). The contribution of anoxygenic photosynthesis was insignificant (about 7% of the total OM production). The share of bacterial chemosynthesis was still less (no more than 2%). In the anaerobic zone, the community of sulfate-reducing bacteria played a decisive role in the terminal decomposition of OM. The maximal rates of sulfate reduction were observed in the near-bottom water (114 micrograms S/(1 day)) and in the surface layer of bottom sediments (901 micrograms S/(dm3 day)). The daily expenditure of Corg for sulfate reduction was 73% of Corg formed daily in the processes of oxygenic and anoxygenic photosynthesis and bacterial chemosynthesis. The profile of methane distribution in the water column and bottom sediments was typical of meromictic reservoirs. The methane content in the water column increased beginning with the thermocline (7-8 m), and reached maximum values in the near-bottom water (17 microliters/l). In bottom sediments, the greatest methane concentrations (57 microliters/l) were observed in the surface layer (0-3 cm). The integral rate of methane formation in the water column and bottom sediments was almost an order of magnitude higher than the rate of its oxidation by aerobic and anaerobic methanotrophic microorganisms.

Bivalve mollusks Bathymodiolus asoricus and Bathymodiolus puteoserpentis collected from the Rainbow and Logachev hydrothermal fields during dives of Mir 1 and Mir 2 deep-sea manned submersibles were studied. Rates of methane oxidation and carbon dioxide assimilation in mussel gill tissue were determined by radiolabel analysis. During oxidation of 14Ch4, radiocarbon was detected in significant quantities not only in carbon dioxide but also in dissolved organic matter, most notably 14C-formate and 14C-acetate, occurring in a 2:1 ratio. Activities of hexulose-phosphate synthase, phosphoribulokinase, and ribulose 1,5-bisphosphate carboxylase were shown in the soluble fraction of gill tissue cells. At the same time, no activity of hydroxypyruvate reductase--the key enzyme of the serine pathway of C1-assimilation--was detected. The results of PCR amplification using genetic probes for membrane-bound methane monooxygenase (pmoA) and methanol dehydrogenase (mxaF) attest to the presence of the genes of these enzymes in the total DNA extracted from gill samples. However, no appropriate PCR responses were obtained with the mmoX primer system, which is a marker for soluble methane monooxygenase. All samples studied showed amplification with primers for the genera Methylobacter and Methylosphaera. At the same time, no genes specific to the genera Methylomonas, Methylococcus, Methylomicrobium, or Methylosinus and Methylocystis were detected. Electron microscopic examinations revealed the presence of two groups of endosymbiotic bacteria in the mussel gill tissue. The first group was represented by large cells possessing a complex system of cytoplasmic membranes, typical of methanotrophs of morphotype I. The other type of endosymbionts, having much smaller cells and lacking intracellular membrane structures, is likely to be constituted by sulfur bacteria.

Seasonal investigations of methane distribution and rates of its oxidation and generation in the water column and sediments of the Black Sea northwestern shelf were carried out within the framework of the interdisciplinary projects "European River-Ocean Systems" (EROS-2000, EROS-21) and "Biogenic Gases Exchange in the Black Sea" (BigBlack) in August 1995, May 1997, and December 1999. Experiments that involved the addition of 14CH3COONa and 14CO2 to sediment samples showed the main part of methane to be formed from CO2. Maximum values of methane production (up to 559 mumol/(m2 day)) were found in coastal sediments in summer time. In winter and spring, methane production in the same sediments did not exceed 3.6-4.2 mumol/(m2 day). The delta 13C values of methane ranged from -70.7 to -81.8@1000, demonstrating its microbial origin and contradicting the concept of the migration of methane from cold seeps or from the oil fields located at the Black Sea shelf. Experiments that involved the addition of 14CH4 to water and sediment samples showed that a considerable part of methane is oxidized in the upper horizons of bottom sediments and in the water column. Nevertheless, it was found that, in summer, part of methane (from 6.8 to 320 mumol/(m2 day)) arrives in the atmosphere.

With the use of molecular ecology methods, the archaea component of microbial mats on coral-like structures associated with methane seeps occurring at a depth of about 200 m in the Black Sea was investigated without the isolation of pure cultures. Using archaea-specific 16S rRNA-targeted oligonucleotide primes, long fragments of genes were amplified, cloned, and sequenced and their phylogenetic analysis was carried out. It was shown that archaea in microbial mats on coral-like structures are represented by two dominant phylotypes that belong to the kingdoms Crenarchaeota and Euryarchaeota and are not specifically related to any described archaea species. The possible role of the revealed archaea in the process of anaerobic methane oxidation is discussed.

The aragonite constructions of the Black Sea are formed in a stable anaerobic zone and are a perfect object to study the natural mechanism of anaerobic methane oxidation. The most probable pathway of methane oxidation is its methanogen-mediated reaction with bicarbonates, dissolved in seawater, with the formation of water and acetate, which is then consumed by other components of the anaerobic community. Comparison of the delta 13C values of carbonate minerals and organic matter once more demonstrated that the formation of the organic matter of biomass is accompanied by intense fractionation of carbon isotopes, as a result of which the total organic matter of biomass acquires an extremely light isotopic composition, characterized by delta 13C values as low as -83.8@1000.

In the beginning of summer 1999, complex microbiological and biogeochemical investigations of meromictic Lake Mogil'noe (Kil'din Island, Barents Sea) were carried out. The analysis of the results shows clearly pronounced vertical zonality of the microbial processes occurring in the water column of the lake. To a depth of 8 m, the total number and activity of microorganisms was limited by the relatively low content of organic matter (OM). In the upper part of the hydrogen-sulfide zone of the lake (beginning at a depth of 8.25 m), the content of particulate OM and the microbial number sharply increased. In this zone, the daily production of OM during anaerobic photosynthesis at the expense of massive development of colored sulfur bacteria reached 620 mg C/m2, which was twofold greater than the daily production of phytoplankton photosynthesis and led to a considerable change in the isotopic composition (delta 13 C) of the particulate OM. In the same intermediate layer, the highest rates of sulfate reduction were recorded, and fractionation of stable sulfur isotopes occurred. Below 10 m was the third hydrochemical zone, characterized by maximum concentrations of H2S and CH4 and by a relatively high rate of autotrophic methanogenesis. The comparison of the results obtained with the results of investigations of previous years, performed in the end of summer, shows a decrease in the intensity of all microbial processes inspected. An exception was anoxygenic photosynthesis, which can utilize not only the de novo formed H2S but also the H2S accumulated in the lake during the winter period.