Analyses of spatial distributions of sulfate-reducing bacteria and their activity in aerobic wastewater biofilms.

The vertical distribution of sulfate-reducing bacteria (SRB) in aerobic wastewater biofilms grown on rotating disk reactors was investigated by fluorescent in situ hybridization (FISH) with 16S rRNA-targeted oligonucleotide probes. To correlate the vertical distribution of SRB populations with their activity, the microprofiles of O(2), H(2)S, NO(2)(-), NO(3)(-), NH(4)(+), and pH were measured with microelectrodes. In addition, a cross-evaluation of the FISH and microelectrode analyses was performed by comparing them with culture-based approaches and biogeochemical measurements. In situ hybridization revealed that a relatively high abundance of the probe SRB385-stained cells (approximately 10(9) to 10(10) cells per cm(3) of biofilm) were evenly distributed throughout the biofilm, even in the oxic surface. The probe SRB660-stained Desulfobulbus spp. were found to be numerically important members of SRB populations (approximately 10(8) to 10(9) cells per cm(3)). The result of microelectrode measurements showed that a high sulfate-reducing activity was found in a narrow anaerobic zone located about 150 to 300 microm below the biofilm surface and above which an intensive sulfide oxidation zone was found. The biogeochemical measurements showed that elemental sulfur (S(0)) was an important intermediate of the sulfide reoxidation in such thin wastewater biofilms (approximately 1,500 microm), which accounted for about 75% of the total S pool in the biofilm. The contribution of an internal Fe-sulfur cycle to the overall sulfur cycle in aerobic wastewater biofilms was insignificant (less than 1%) due to the relatively high sulfate reduction rate.     

Successional development of sulfate-reducing bacterial populations and their activities in an activated sludge immobilized agar gel film

A combination of fluorescence in situ hybridization (FISH), microprofiles, and denaturing gradient gel electrophoresis (DGGE) analysis of PCR-amplified 16S rDNA fragments followed by hybridization analysis with specific probes was applied to investigate successional development of sulfate-reducing bacteria (SRB) community structure and in situ sulfide production activity within an activated sludge immobilized agar gel film. In this model biofilm system, since biases arising from biofilm heterogeneity can be ignored, the population dynamics of SRB in the agar gel is directly related to physiological capability and in situ activity of SRB. Microelectrode measurements showed that an anoxic zone was already developed at the beginning (0 day), a first sulfide production of 0.054 mumol H2S m(-2) x s(-1) was detected during the first week, and the rate increased gradually to 0.221 mumol H2S m(-2) x s(-1) in the fifth week. The most active sulfide production zone moved upward to the chemocline and intensified with time to form a narrow zone with high volumetric sulfide production rates. This result coincided with the shift of the spatial distributions of SRB populations determined by FISH. In situ hybridization with probe SRB385 for mainly general SRB of the delta Proteobacteria plus some gram-positive bacteria and probe 660 for Desulfobulbus indicated that the most abundant populations of SRB were primarily restricted to near the oxic/anoxic interface (chemocline). A close observation of the development of the vertical distributions of SRB populations revealed that the cell numbers of Desulfobulbus tripled (from 0.5 x 10(8) to 1.5 x 10(8) cells cm(-3)) near the oxic/anoxic interface. Similar growth (from 1.0 x10(8) to 4.5 x 10(8) cells cm(-3)) of Desulfovibrio-like SRB that hybridized with probe SRB385 was observed. PCR-DGGE followed by hybridization analysis revealed that one Desulfobulbus strain was detected from the beginning, and another strain appeared after 1 week, coinciding with the first detected sulfide production. In addition, three strains hybridizing with probe 687 (possibly Desulfovibrio) were also dominant SRB in the agar gel.     

Successional development of sulfate-reducing bacterial populations and their activities in a wastewater biofilm growing under microaerophilic conditions

A combination of fluorescence in situ hybridization, microprofiles, denaturing gradient gel electrophoresis of PCR-amplified 16S ribosomal DNA fragments, and 16S rRNA gene cloning analysis was applied to investigate successional development of sulfate-reducing bacteria (SRB) community structure and in situ sulfide production activity within a biofilm growing under microaerophilic conditions (dissolved oxygen concentration in the bulk liquid was in the range of 0 to 100 microM) and in the presence of nitrate. Microelectrode measurements showed that oxygen penetrated 200 microm from the surface during all stages of biofilm development. The first sulfide production of 0.32 micromol of H(2)S m(-2) s(-1) was detected below ca. 500 microm in the 3rd week and then gradually increased to 0.70 micromol H(2)S m(-2) s(-1) in the 8th week. The most active sulfide production zone moved upward to the oxic-anoxic interface and intensified with time. This result coincided with an increase in SRB populations in the surface layer of the biofilm. The numbers of the probe SRB385- and 660-hybridized SRB populations significantly increased to 7.9 x 10(9) cells cm(-3) and 3.6 x 10(9) cells cm(-3), respectively, in the surface 400 microm during an 8-week cultivation, while those populations were relatively unchanged in the deeper part of the biofilm, probably due to substrate transport limitation. Based on 16S rRNA gene cloning analysis data, clone sequences that related to Desulfomicrobium hypogeium (99% sequence similarity) and Desulfobulbus elongatus (95% sequence similarity) were most frequently found. Different molecular analyses confirmed that Desulfobulbus, Desulfovibrio, and Desulfomicrobium were found to be the numerically important members of SRB in this wastewater biofilm.     

微生态制剂改善对虾养殖池塘底质的效果

研究了在117 d的养殖周期中微生态制剂对南美白对虾池塘底质的改良效果.结果表明,与对照组相比,施用微生态制剂可使底质中总氮、总磷和硫化物的含量显著下降;总菌数量无显著变化,而芽孢杆菌、氨化细菌以及硫氧化细菌、硫还原细菌、弧菌数量差异显著,其中弧菌数量在施用微生态制剂处理和对照条件下分别为3.65×10³ cfu·g^-1和1.16×10⁵ cfu·g^-1.表明施用微生态制剂可以减少氮、磷、硫等营养物质的积累,改善池塘底质的菌相,为南美白对虾的健康养殖提供良好的池塘底质环境.     

海岸盐沼湿地土壤硫循环中的微生物及其作用

硫及硫化合物的动态循环是海岸盐沼湿地的重要组成部分,硫酸盐还原菌(SRB)和硫氧化菌(SOB)是推动硫循环的重要微生物。硫酸盐还原菌把硫酸盐还原为硫化物,同时消耗土壤中的有机物质;硫氧化菌把还原性硫化合物氧化为硫酸盐,缓解土壤中硫化物的积累,它们共同维持硫循环的动态平衡。本文综述了海岸盐沼湿地土壤中硫的存在形式、硫的地球化学循环以及在硫循环过程中扮演重要角色的硫酸盐还原菌和硫氧化菌的生物多样性、活性测定方法及其生态学意义等的最新研究进展,并提出了存在的问题及研究展望。     

Vertical Migration in the Sediment-Dwelling Sulfur Bacteria Thioploca spp. in Overcoming Diffusion Limitations

In order to investigate the environmental requirements of the filamentous sulfur bacteria Thioploca spp., we tested the chemotactic responses of these sedimentary microorganisms to changes in oxygen, nitrate, and sulfide concentrations. A sediment core with a Thioploca mat, retrieved from the oxygen-minimum zone on the Chilean shelf, was incubated in a recirculating flume. The addition of 25 (mu)mol of nitrate per liter to the seawater flow induced the ascent of the Thioploca trichomes (length, up to 70 mm) in their mostly vertically oriented gelatinous sheaths. The upper ends of the filaments penetrated the sediment surface and protruded 1 to 3 mm into the flowing water before they bent downstream. By penetrating the diffusive boundary layer, Thioploca spp. facilitate efficient nitrate uptake in exposed trichome sections that are up to 30 mm long. The cumulative length of exposed filaments per square centimeter of sediment surface was up to 92 cm, with a total exposed trichome surface area of 1 cm(sup2). The positive reaction to nitrate overruled a negative response to oxygen, indicating that nitrate is the principal electron acceptor used by Thioploca spp. in the anoxic environment; 10-fold increases in nitrate fluxes after massive emergence of filaments strengthened this hypothesis. A positive chemotactic response to sulfide concentrations of less than 100 (mu)mol liter(sup-1) counteracted the attraction to nitrate and, along with phobic reactions to oxygen and higher sulfide concentrations, controlled the vertical movement of the trichomes. We suggest that the success of Thioploca spp. on the Chilean shelf is based on the ability of these organisms to shuttle between the nitrate-rich boundary layer and the sulfidic sediment strata.     

Methylated sulfur compounds in microbial mats: in situ concentrations and metabolism by a colorless sulfur bacterium

The concentrations of the volatile organic sulfur compounds methanethiol, dimethyl disulfide, and dimethyl sulfide (DMS) and the viable population capable of DMS utilization in laminated microbial ecosystems were evaluated. Significant levels of DMS and dimethyl disulfide (maximum concentrations of 220 and 24 nmol cm3 of sediment-1, respectively) could be detected only at the top 20 mm of the microbial mat, whereas methanethiol was found only at depth horizons from 20 to 50 mm (maximum concentration of 42 nmol cm3 of sediment-1). DMS concentrations in the surface layer doubled after cold hydrolysis of its precursor, dimethylsulfoniopropionate. Most-probable-number counts revealed 2.2 x 10(5) cells cm3 of sediment-1, in the 0- to 5-mm depth horizon, capable of growth on DMS as the sole source of energy. An obligately chemolithoautotrophic bacillus designated strain T5 was isolated from the top layer of the marine sediment. Continuous culture studies in which DMS was the growth-limiting substrate revealed a maximum specific growth rate of 0.10 h-1 and a saturation constant of 90 mumol liter-1 for aerobic growth on this substrate.     

Methylated sulfur compounds in microbial mats: in situ concentrations and metabolism by a colorless sulfur bacterium.

The concentrations of the volatile organic sulfur compounds methanethiol, dimethyl disulfide, and dimethyl sulfide (DMS) and the viable population capable of DMS utilization in laminated microbial ecosystems were evaluated. Significant levels of DMS and dimethyl disulfide (maximum concentrations of 220 and 24 nmol cm3 of sediment-1, respectively) could be detected only at the top 20 mm of the microbial mat, whereas methanethiol was found only at depth horizons from 20 to 50 mm (maximum concentration of 42 nmol cm3 of sediment-1). DMS concentrations in the surface layer doubled after cold hydrolysis of its precursor, dimethylsulfoniopropionate. Most-probable-number counts revealed 2.2 x 10(5) cells cm3 of sediment-1, in the 0- to 5-mm depth horizon, capable of growth on DMS as the sole source of energy. An obligately chemolithoautotrophic bacillus designated strain T5 was isolated from the top layer of the marine sediment. Continuous culture studies in which DMS was the growth-limiting substrate revealed a maximum specific growth rate of 0.10 h-1 and a saturation constant of 90 mumol liter-1 for aerobic growth on this substrate.     

Sulfide oxidation at halo-alkaline conditions in a fed-batch bioreactor

A biotechnological process is described to remove hydrogen sulfide (H(2)S) from high-pressure natural gas and sour gases produced in the petrochemical industry. The process operates at halo-alkaline conditions and combines an aerobic sulfide-oxidizing reactor with an anaerobic sulfate (SO(4) (2-)) and thiosulfate (S(2)O(3) (2-)) reducing reactor. The feasibility of biological H(2)S oxidation at pH around 10 and total sodium concentration of 2 mol L(-1) was studied in gas-lift bioreactors, using halo-alkaliphilic sulfur-oxidizing bacteria (HA-SOB). Reactor operation at different oxygen to sulfide (O(2):H(2)S) supply ratios resulted in a stable low redox potential that was directly related with the polysulfide (S(x) (2-)) and total sulfide concentration in the bioreactor. Selectivity for SO(4) (2-) formation decreased with increasing S(x) (2-) and total sulfide concentrations. At total sulfide concentrations above 0.25 mmol L(-1), selectivity for SO(4) (2-) formation approached zero and the end products of H(2)S oxidation were elemental sulfur (S(0)) and S(2)O(3) (2-). Maximum selectivity for S(0) formation (83.3+/-0.7%) during stable reactor operation was obtained at a molar O(2):H(2)S supply ratio of 0.65. Under these conditions, intermediary S(x) (2-) plays a major role in the process. Instead of dissolved sulfide (HS(-)), S(x) (2-) seemed to be the most important electron donor for HA-SOB under S(0) producing conditions. In addition, abiotic oxidation of S(x) (2-) was the main cause of undesirable formation of S(2)O(3) (2-). The observed biomass growth yield under SO(4) (2-) producing conditions was 0.86 g N mol(-1) H(2)S. When selectivity for SO(4) (2-) formation was below 5%, almost no biomass growth was observed.     

Colorless Sulfur Bacteria, Beggiatoa spp. and Thiovulum spp., in O(2) and H(2)S Microgradients

The interactions between colorless sulfur bacteria and the chemical microgradients at the oxygen-sulfide interface were studied in Beggiatoa mats from marine sediments and in Thiovulum veils developing above the sediments. The gradients of O(2), H(2)S, and pH were measured by microelectrodes at depth increments of 50 mum. An unstirred boundary layer in the water surrounding the mats and veils prevented microturbulent or convective mixing of O(2) and H(2)S. The two substrates reached the bacteria only by molecular diffusion through the boundary layer. The bacteria lived as microaerophiles or anaerobes even under stirred, oxic water. Oxygen and sulfide zones overlapped by 50 mum in the bacterial layers. Both compounds had concentrations in the range of 0 to 10 mumol liter and residence times of 0.1 to 0.6 s in the overlapping zone. The sulfide oxidation was purely biological. Diffusion calculations showed that formation of mats on solid substrates or of veils in the water represented optimal strategies for the bacteria to achieve a stable microenvironment, a high substrate supply, and an efficient competition with chemical sulfide oxidation. The continuous gliding movement of Beggiatoa cells in mats or the flickering motion of Thiovulum cells in veils were important for the availability of both O(2) and H(2)S for the individual bacteria.     

Diversity and vertical distribution of magnetotactic bacteria along chemical gradients in freshwater microcosms

The vertical distribution of magnetotactic bacteria along various physico-chemical gradients in freshwater microcosms was analyzed by a combined approach of viable cell counts, 16S rRNA gene analysis, microsensor profiling and biogeochemical methods. The occurrence of magnetotactic bacteria was restricted to a narrow sediment layer overlapping or closely below the maximum oxygen and nitrate penetration depth. Different species showed different preferences within vertical gradients, but the largest proportion (63-98%) of magnetotactic bacteria was detected within the suboxic zone. In one microcosm the community of magnetotactic bacteria was dominated by one species of a coccoid "Alphaproteobacterium", as detected by denaturing gradient gel electrophoresis in sediment horizons from 1 to 10 mm depth. Maximum numbers of magnetotactic bacteria were up to 1.5 x 10(7) cells/cm3, which corresponded to 1% of the total cell number in the upper sediment layer. The occurrence of magnetotactic bacteria coincided with the availability of significant amounts (6-60 microM) of soluble Fe(II), and in one sample with hydrogen sulfide (up to 40 microM). Although various trends were clearly observed, a strict correlation between the distribution of magnetotactic bacteria and individual geochemical parameters was absent. This is discussed in terms of metabolic adaptation of various strains of magnetotactic bacteria to stratified sediments and diversity of the magnetotactic bacterial communities.     

Linked redox precipitation of sulfur and selenium under anaerobic conditions by sulfate-reducing bacterial biofilms

A biofilm-forming strain of sulfate-reducing bacteria (SRB), isolated from a naturally occurring mixed biofilm and identified by 16S rDNA analysis as a strain of Desulfomicrobium norvegicum, rapidly removed 200 micro M selenite from solution during growth on lactate and sulfate. Elemental selenium and elemental sulfur were precipitated outside SRB cells. Precipitation occurred by an abiotic reaction with bacterially generated sulfide. This appears to be a generalized ability among SRB, arising from dissimilatory sulfide biogenesis, and can take place under low redox conditions and in the dark. The reaction represents a new means for the deposition of elemental sulfur by SRB under such conditions. A combination of transmission electron microscopy, environmental scanning electron microscopy, and cryostage field emission scanning electron microscopy were used to reveal the hydrated nature of SRB biofilms and to investigate the location of deposited sulfur-selenium in relation to biofilm elements. When pregrown SRB biofilms were exposed to a selenite-containing medium, nanometer-sized selenium-sulfur granules were precipitated within the biofilm matrix. Selenite was therefore shown to pass through the biofilm matrix before reacting with bacterially generated sulfide. This constitutes an efficient method for the removal of toxic concentrations of selenite from solution. Implications for environmental cycling and the fate of sulfur and selenium are discussed, and a general model for the potential action of SRB in selenium transformations is presented.     

Alkaliphilic sulfidogenesis on cellulose by combined cultures

Soda lakes are characterized by an intense sulfur cycle that begins with sulfidogenesis. Model laboratory experiments that involved combining of pure cultures showed that, during anaerobic decomposition of cellulose by Clostridium alkalicellulosi, the sulfate-reducing bacteria (SRB) of the species Desulfonatronovibrio hydrogenovorans, Desulfonatronum lacustre, and Desulfonatronum cooperativum, different in their nutritional requirements, may directly use the cellulose fermentation products for sulfidogenesis without mediatory microorgansims. In binary cocultures with SRB, the amount of the H2S formed constituted from one-third to two-thirds of the cellulose [H] equivalents; acetate was among the products formed. When the syntrophic Contubernalis alkalaceticum, capable of acetate oxidation, was incorporated into the trophic chain along with hydrogenotrophic SRB, the amount of the H2S formed exceeded by 33-42% the amount of the [H] equivalents in the utilized cellulose, water being the source of additional hydrogen. Thus, the trophic pathway from plant residues to sulfide, previously considered to be the longest in the alkaliphilic microbial community, may involve a minimal number of stages and do without intermediate participation of dissipotrophic fermenters.     

Succession of internal sulfur cycles and sulfur-oxidizing bacterial communities in microaerophilic wastewater biofilms

The succession of sulfur-oxidizing bacterial (SOB) community structure and the complex internal sulfur cycle occurring in wastewater biofilms growing under microaerophilic conditions was analyzed by using a polyphasic approach that employed 16S rRNA gene-cloning analysis combined with fluorescence in situ hybridization, microelectrode measurements, and standard batch and reactor experiments. A complete sulfur cycle was established via S(0) accumulation within 80 days in the biofilms in replicate. This development was generally split into two phases, (i) a sulfur-accumulating phase and (ii) a sulfate-producing phase. In the first phase (until about 40 days), since the sulfide production rate (sulfate-reducing activity) exceeded the maximum sulfide-oxidizing capacity of SOB in the biofilms, H(2)S was only partially oxidized to S(0) by mainly Thiomicrospira denitirificans with NO(3)(-) as an electron acceptor, leading to significant accumulation of S(0) in the biofilms. In the second phase, the SOB populations developed further and diversified with time. In particular, S(0) accumulation promoted the growth of a novel strain, strain SO07, which predominantly carried out the oxidation of S(0) to SO(4)(2-) under oxic conditions, and Thiothrix sp. strain CT3. In situ hybridization analysis revealed that the dense populations of Thiothrix (ca. 10(9) cells cm(-3)) and strain SO07 (ca. 10(8) cells cm(-3)) were found at the sulfur-rich surface (100 microm), while the population of Thiomicrospira denitirificans was distributed throughout the biofilms with a density of ca. 10(7) to 10(8) cells cm(-3). Microelectrode measurements revealed that active sulfide-oxidizing zones overlapped the spatial distributions of different phylogenetic SOB groups in the biofilms. As a consequence, the sulfide-oxidizing capacities of the biofilms became high enough to completely oxidize all H(2)S produced by SRB to SO(4)(2-) in the second phase, indicating establishment of the complete sulfur cycle in the biofilms.     

Inhibition of microbial H<Subscript>2</Subscript>S production in an oil reservoir model column by nitrate injection

The effect of nitrate addition on microbial H2S production in a seawater-flooded oil reservoir model column with crude oil as carbon and energy source was investigated. Injection of 0.5 mM nitrate for 2.5-3.5 months led to complete elimination of H2S (initially 0.45-0.67 mM). The major decline in H2S level coincided with the first complete nitrate consumption and production of nitrite. When nitrate was excluded, H2S production resumed after approximately 2.5 months and reached previous levels after approximately 5 months. Using a fluorescent antibody technique, three populations each of sulfate-reducing bacteria (SRB) and nitrate-reducing bacteria (NRB) were monitored. SRB dominated the anoxic zone prior to nitrate addition, comprising 64-93% of the total bacterial population. The monitored NRB constituted less than 6% and no increase was observed during nitrate addition (indicating that other, unidentified, NRB populations were present). After 1-3 months without significant H2S production (3.5-5.5 months with nitrate), the SRB population collapsed, the fraction being reduced to 9-25%. The dominant SRB strain in the column, which constituted on average 94% of the monitored SRB population, was partly/completely inhibited by 50/75 microM nitrite in batch culture tests. Similar nitrite concentrations (50-150 microM) were detected in the column when the H2S level declined, indicating that nitrite inhibition was the main cause of H2S elimination. The results from this study indicate that nitrate/nitrite can be used to prevent detrimental SRB activity in oil reservoirs.     

Community structure, cellular rRNA content, and activity of sulfate-reducing bacteria in marine arctic sediments

The community structure of sulfate-reducing bacteria (SRB) of a marine Arctic sediment (Smeerenburgfjorden, Svalbard) was characterized by both fluorescence in situ hybridization (FISH) and rRNA slot blot hybridization by using group- and genus-specific 16S rRNA-targeted oligonucleotide probes. The SRB community was dominated by members of the Desulfosarcina-Desulfococcus group. This group accounted for up to 73% of the SRB detected and up to 70% of the SRB rRNA detected. The predominance was shown to be a common feature for different stations along the coast of Svalbard. In a top-to-bottom approach we aimed to further resolve the composition of this large group of SRB by using probes for cultivated genera. While this approach failed, directed cloning of probe-targeted genes encoding 16S rRNA was successful and resulted in sequences which were all affiliated with the Desulfosarcina-Desulfococcus group. A group of clone sequences (group SVAL1) most closely related to Desulfosarcina variabilis (91.2% sequence similarity) was dominant and was shown to be most abundant in situ, accounting for up to 54. 8% of the total SRB detected. A comparison of the two methods used for quantification showed that FISH and rRNA slot blot hybridization gave comparable results. Furthermore, a combination of the two methods allowed us to calculate specific cellular rRNA contents with respect to localization in the sediment profile. The rRNA contents of Desulfosarcina-Desulfococcus cells were highest in the first 5 mm of the sediment (0.9 and 1.4 fg, respectively) and decreased steeply with depth, indicating that maximal metabolic activity occurred close to the surface. Based on SRB cell numbers, cellular sulfate reduction rates were calculated. The rates were highest in the surface layer (0.14 fmol cell(-1) day(-1)), decreased by a factor of 3 within the first 2 cm, and were relatively constant in deeper layers.     

Biogeochemistry and community composition of iron- and sulfur-precipitating microbial mats at the Chefren mud volcano (Nile Deep Sea Fan, Eastern Mediterranean)

In this study we determined the composition and biogeochemistry of novel, brightly colored, white and orange microbial mats at the surface of a brine seep at the outer rim of the Chefren mud volcano. These mats were interspersed with one another, but their underlying sediment biogeochemistries differed considerably. Microscopy revealed that the white mats were granules composed of elemental S filaments, similar to those produced by the sulfide-oxidizing epsilonproteobacterium "Candidatus Arcobacter sulfidicus." Fluorescence in situ hybridization indicated that microorganisms targeted by a "Ca. Arcobacter sulfidicus"-specific oligonucleotide probe constituted up to 24% of the total the cells within these mats. Several 16S rRNA gene sequences from organisms closely related to "Ca. Arcobacter sulfidicus" were identified. In contrast, the orange mat consisted mostly of bright orange flakes composed of empty Fe(III) (hydr)oxide-coated microbial sheaths, similar to those produced by the neutrophilic Fe(II)-oxidizing betaproteobacterium Leptothrix ochracea. None of the 16S rRNA gene sequences obtained from these samples were closely related to sequences of known neutrophilic aerobic Fe(II)-oxidizing bacteria. The sediments below both types of mats showed relatively high sulfate reduction rates (300 nmol x cm(-3) x day(-1)) partially fueled by the anaerobic oxidation of methane (10 to 20 nmol x cm(-3) x day(-1)). Free sulfide produced below the white mat was depleted by sulfide oxidation within the mat itself. Below the orange mat free Fe(II) reached the surface layer and was depleted in part by microbial Fe(II) oxidation. Both mats and the sediments underneath them hosted very diverse microbial communities and contained mineral precipitates, most likely due to differences in fluid flow patterns.     

Biogeochemistry and biodiversity of methane cycling in subsurface marine sediments (Skagerrak, Denmark)

This biogeochemical, molecular genetic and lipid biomarker study of sediments ( approximately 4 m cores) from the Skagerrak (Denmark) investigated methane cycling in a sediment with a clear sulfate-methane-transition zone (SMTZ) and where CH(4) supply was by diffusion, rather than by advection, as in more commonly studied seep sites. Sulfate reduction removed sulfate by 0.7 m and CH(4) accumulated below. (14)C-radiotracer measurements demonstrated active H(2)/CO(2) and acetate methanogenesis and anaerobic oxidation of CH(4) (AOM). Maximum AOM rates occurred near the SMTZ ( approximately 3 nmol cm(-3) day(-1) at 0.75 m) but also continued deeper, overall, at much lower rates. Maximum rates of H(2)/CO(2) and acetate methanogenesis occurred below the SMTZ but H(2)/CO(2) methanogenesis rates were x 10 those of acetate methanogenesis, and this was consistent with initial values of (13)C-depleted CH(4) (delta(13)C c.-80 per thousand). Areal AOM and methanogenic rates were similar ( approximately 1.7 mmol m(-2) day(-1)), hence, CH(4) flux is finely balanced. A 16S rRNA gene library from 1.39 m combined with methanogen (T-RFLP), bacterial (16S rRNA DGGE) and lipid biomarker depth profiles showed the presence of populations similar to some seep sites: ANME-2a (dominant), ANME-3, Methanomicrobiales, Methanosaeta Archaea, with abundance changes with depth corresponding to changes in activities and sulfate-reducing bacteria (SRB). Below the SMTZ to approximately 1.7 m CH(4) became progressively more (13)C depleted (delta(13)C -82 per thousand) indicating a zone of CH(4) recycling which was consistent with the presence of (13)C-depleted archaeol (delta(13)C -55 per thousand). Pore water acetate concentrations decreased in this zone (to approximately 5 microM), suggesting that H(2), not acetate, was an important CH(4) cycling intermediate. The potential biomarkers for AOM-associated SRB, non-isoprenoidal ether lipids, increased below the SMTZ but this distribution reflected 16S rRNA gene sequences for JS1 and OP8 bacteria rather than those of SRB. At this site peak rates of methane production and consumption are spatially separated and seem to be conducted by different archaeal groups. Also AOM is predominantly coupled to sulfate reduction, unlike recent reports from some seep and gassy sediment sites.     

Identity and abundance of active sulfate-reducing bacteria in deep tidal flat sediments determined by directed cultivation and CARD-FISH analysis

The identity and abundance of potentially active sulfate-reducing bacteria (SRB) in several metre deep sediments of a tidal sand flat in the German Wadden Sea were assessed by directed cultivation and cultivation-independent CARD-FISH analysis (catalysed reporter deposition fluorescence in situ hybridization). Presumably abundant SRB from different sediment layers between 0.5 and 4 m depth were selectively enriched in up to million-fold diluted cultures supplemented with lactate, acetate or hydrogen. Partial 16S rRNA gene sequences obtained from highest dilution steps showing sulfide formation indicated growth of deltaproteobacterial SRB belonging to the Desulfobulbaceae and the Desulfobacteraceae as well as of members of the Firmicutes. Subsequent isolation resulted in 10 novel phylotypes of both litho- and organotrophic sulfate-reducing Deltaproteobacteria. Molecular pre-screening identified six isolates as members of the Desulfobulbaceae, sharing highest identities with either candidatus 'Desulfobacterium corrodens' (95-97%) or Desulfobacterium catecholicum (98%), and four isolates as members of Desulfobacteraceae, being related to either Desulfobacter psychrotolerans (98%) or Desulfobacula phenolica (95-97%). Relatives of D. phenolica were exlusively isolated from 50 and 100 cm deep sediments with 10 and 2 mM of pore water sulfate respectively. In contrast, relatives of D. corrodens, D. psychrotolerans and D. catecholicum were also obtained from layers deeper than 100 cm and with less than 2 mM sulfate. The high in situ abundance of members of both families in sediment layers beneath 50 cm could be confirmed via CARD-FISH analysis performed with a set of six SRB-specific oligonucleotide probes. Moreover, SRB represented a numerically significant fraction of the microbial community throughout the sediment (up to 7%) and reached even higher cell numbers in deep, sulfate-poor layers than in the sulfate-rich surface sediment. This relatively large community size of potentially active SRB in deep sandy sediments might on the one hand be a result of their syntrophic association with other anaerobes. Our results furthermore support the hypothesis that enhanced advective pore water transport might supply nutrients to microbial communities in deep sandy sediments and point to their so far unrecognized contribution to biogeochemical processes in Wadden Sea sediments.     

Major contribution of sulfide-derived sulfur to the benthic food web in a large freshwater lake

In freshwater systems, contributions of chemosynthetic products by sulfur-oxidizing bacteria in sediments as nutritional resources in benthic food webs remain unclear, even though chemosynthetic products might be an important nutritional resource for benthic food webs in deep-sea hydrothermal vents and shallow marine systems. To study geochemical aspects of this trophic pathway, we sampled sediment cores and benthic animals at two sites (90 and 50 m water depths) in the largest freshwater (mesotrophic) lake in Japan: Lake Biwa. Stable carbon, nitrogen, and sulfur isotopes of the sediments and animals were measured to elucidate the sulfur nutritional resources for the benthic food web precisely by calculating the contributions of the incorporation of sulfide-derived sulfur to the biomass and of the biogeochemical sulfur cycle supporting the sulfur nutritional resource. The recovered sediment cores showed increases in ³⁴S-depleted sulfide at 5 cm sediment depth and showed low sulfide concentration with high δ³⁴S in deeper layers, suggesting an association of microbial activities with sulfate reduction and sulfide oxidation in the sediments. The sulfur-oxidizing bacteria may contribute to benthic animal biomass. Calculations based on the biomass, sulfur content, and contribution to sulfide-derived sulfur of each animal comprising the benthic food web revealed that 58%–67% of the total biomass sulfur in the benthic food web of Lake Biwa is occupied by sulfide-derived sulfur. Such a large contribution implies that the chemosynthetic products of sulfur-oxidizing bacteria are important nutritional resources supporting benthic food webs in the lake ecosystems, at least in terms of sulfur. The results present a new trophic pathway for sulfur that has been overlooked in lake ecosystems with low-sulfate concentrations.