Propionate Converting Anaerobic Microbial Communities Enriched from Distinct Biogeochemical Zones of Aarhus Bay, Denmark under Sulfidogenic and Methanogenic Conditions.
ABSTRACT: The relationship between predominant physiological types of prokaryotes in marine sediments and propionate degradation through sulfate reduction, fermentation, and methanogenesis was studied in marine sediments. Propionate conversion was assessed in slurries containing sediment from three different biogeochemical zones of Aarhus Bay, Denmark. Sediment slurries were amended with 0, 3, or 20 mM sulfate and incubated at 25 °C and 10 °C for 514-571 days. Methanogenesis in the sulfate zone and sulfate reduction in the methane zone slurries was observed. Both processes occurred simultaneously in enrichments originating from samples along the whole sediment. Bacterial community analysis revealed the dominance of Desulfobacteraceae and Desulfobulbaceae members in sulfate-amended slurries incubated at 25°C and 10°C. Cryptanaerobacter belonging to the Peptococcaceae family dominated sulfate-free methanogenic slurries at 25°C, whereas bacteria related to Desulfobacteraceae were dominant at 10°C. Archaeal community analysis revealed the prevalence of different genera belonging to Methanomicrobiales in slurries incubated at different temperatures and amended with different sulfate concentrations. Methanosarcinaceae were only detected in the absence of sulfate. In summary, Aarhus Bay sediment zones contain sulfate reducers, syntrophs, and methanogens interacting with each other in the conversion of propionate. Our results indicate that in Aarhus Bay sediments, Cryptanaerobacter degraded propionate in syntrophic association with methanogens.
Project description:The conventional perception that the zone of sulfate reduction and methanogenesis are separated in high- and low-sulfate-containing marine sediments has recently been changed by studies demonstrating their co-occurrence in sediments. The presence of methanogens was linked to the presence of substrates that are not used by sulfate reducers. In the current study, we hypothesized that both groups can co-exist, consuming common substrates (H2 and/or acetate) in sediments. We enriched butyrate-degrading communities in sediment slurries originating from the sulfate, sulfate-methane transition, and methane zone of Aarhus Bay, Denmark. Sulfate was added at different concentrations (0, 3, 20 mM), and the slurries were incubated at 10 °C and 25 °C. During butyrate conversion, sulfate reduction and methanogenesis occurred simultaneously. The syntrophic butyrate degrader Syntrophomonas was enriched both in sulfate-amended and in sulfate-free slurries, indicating the occurrence of syntrophic conversions at both conditions. Archaeal community analysis revealed a dominance of Methanomicrobiaceae. The acetoclastic Methanosaetaceae reached high relative abundance in the absence of sulfate, while presence of acetoclastic Methanosarcinaceae was independent of the sulfate concentration, temperature, and the initial zone of the sediment. This study shows that there is no vertical separation of sulfate reducers, syntrophs, and methanogens in the sediment and that they all participate in the conversion of butyrate.
Project description:Patterns of microbial biogeography result from a combination of dispersal, speciation and extinction, yet individual contributions exerted by each of these mechanisms are difficult to isolate and distinguish. The influx of endospores of thermophilic microorganisms to cold marine sediments offers a natural model for investigating passive dispersal in the ocean. We investigated the activity, diversity and abundance of thermophilic endospore-forming sulfate-reducing bacteria (SRB) in Aarhus Bay by incubating pasteurized sediment between 28 and 85?°C, and by subsequent molecular diversity analyses of 16S rRNA and of the dissimilatory (bi)sulfite reductase (dsrAB) genes within the endospore-forming SRB genus Desulfotomaculum. The thermophilic Desulfotomaculum community in Aarhus Bay sediments consisted of at least 23 species-level 16S rRNA sequence phylotypes. In two cases, pairs of identical 16S rRNA and dsrAB sequences in Arctic surface sediment 3000?km away showed that the same phylotypes are present in both locations. Radiotracer-enhanced most probable number analysis revealed that the abundance of endospores of thermophilic SRB in Aarhus Bay sediment was ca. 10(4) per cm(3) at the surface and decreased exponentially to 10(0) per cm(3) at 6.5?m depth, corresponding to 4500 years of sediment age. Thus, a half-life of ca. 300 years was estimated for the thermophilic SRB endospores deposited in Aarhus Bay sediments. These endospores were similarly detected in the overlying water column, indicative of passive dispersal in water masses preceding sedimentation. The sources of these thermophiles remain enigmatic, but at least one source may be common to both Aarhus Bay and Arctic sediments.
Project description:Salinity effects on microbial community structure and on potential rates of arsenate reduction, arsenite oxidation, sulfate reduction, denitrification, and methanogenesis were examined in sediment slurries from two California soda lakes. We conducted experiments with Mono Lake and Searles Lake sediments over a wide range of salt concentrations (25 to 346 g liter(-1)). With the exception of sulfate reduction, rates of all processes demonstrated an inverse relationship to total salinity. However, each of these processes persisted at low but detectable rates at salt saturation. Denaturing gradient gel electrophoresis analysis of partial 16S rRNA genes amplified from As(V) reduction slurries revealed that distinct microbial populations grew at low (25 to 50 g liter(-1)), intermediate (100 to 200 g liter(-1)), and high (>300 g liter(-1)) salinity. At intermediate and high salinities, a close relative of a cultivated As-respiring halophile was present. These results suggest that organisms adapted to more dilute conditions can remain viable at high salinity and rapidly repopulate the lake during periods of rising lake level. In contrast to As reduction, sulfate reduction in Mono Lake slurries was undetectable at salt saturation. Furthermore, sulfate reduction was excluded from Searles Lake sediments at any salinity despite the presence of abundant sulfate. Sulfate reduction occurred in Searles Lake sediment slurries only following inoculation with Mono Lake sediment, indicating the absence of sulfate-reducing flora. Experiments with borate-amended Mono Lake slurries suggest that the notably high (0.46 molal) concentration of borate in the Searles Lake brine was responsible for the exclusion of sulfate reducers from that ecosystem.
Project description:Desulfatiglans-related organisms comprise one of the most abundant deltaproteobacterial lineages in marine sediments where they occur throughout the sediment column in a gradient of increasing sulfate and organic carbon limitation with depth. Characterized Desulfatiglans isolates are dissimilatory sulfate reducers able to grow by degrading aromatic hydrocarbons. The ecophysiology of environmental Desulfatiglans-populations is poorly understood, however, possibly utilization of aromatic compounds may explain their predominance in marine subsurface sediments. We sequenced and analyzed seven Desulfatiglans-related single-cell genomes (SAGs) from Aarhus Bay sediments to characterize their metabolic potential with regard to aromatic compound degradation and energy metabolism. The average genome assembly size was 1.3 Mbp and completeness estimates ranged between 20 and 50%. Five of the SAGs (group 1) originated from the sulfate-rich surface part of the sediment while two (group 2) originated from sulfate-depleted subsurface sediment. Based on 16S rRNA gene amplicon sequencing group 2 SAGs represent the more frequent types of Desulfatiglans-populations in Aarhus Bay sediments. Genes indicative of aromatic compound degradation could be identified in both groups, but the two groups were metabolically distinct with regard to energy conservation. Group 1 SAGs carry a full set of genes for dissimilatory sulfate reduction, whereas the group 2 SAGs lacked any genetic evidence for sulfate reduction. The latter may be due to incompleteness of the SAGs, but as alternative energy metabolisms group 2 SAGs carry the genetic potential for growth by acetogenesis and fermentation. Group 1 SAGs encoded reductive dehalogenase genes, allowing them to access organohalides and possibly conserve energy by their reduction. Both groups possess sulfatases unlike their cultured relatives allowing them to utilize sulfate esters as source of organic carbon and sulfate. In conclusion, the uncultivated marine Desulfatiglans populations are metabolically diverse, likely reflecting different strategies for coping with energy and sulfate limitation in the subsurface seabed.
Project description:Analyses of microbial diversity in marine sediments have identified a core set of taxa unique to the marine deep biosphere. Previous studies have suggested that these specialized communities are shaped by processes in the surface seabed, in particular that their assembly is associated with the transition from the bioturbated upper zone to the nonbioturbated zone below. To test this hypothesis, we performed a fine-scale analysis of the distribution and activity of microbial populations within the upper 50 cm of sediment from Aarhus Bay (Denmark). Sequencing and qPCR were combined to determine the depth distributions of bacterial and archaeal taxa (16S rRNA genes) and sulfate-reducing microorganisms (SRM) (dsrB gene). Mapping of radionuclides throughout the sediment revealed a region of intense bioturbation at 0-6 cm depth. The transition from bioturbated sediment to the subsurface below (7 cm depth) was marked by a shift from dominant surface populations to common deep biosphere taxa (e.g., Chloroflexi and Atribacteria). Changes in community composition occurred in parallel to drops in microbial activity and abundance caused by reduced energy availability below the mixed sediment surface. These results offer direct evidence for the hypothesis that deep subsurface microbial communities present in Aarhus Bay mainly assemble already centimeters below the sediment surface, below the bioturbation zone.
Project description:Most sulfate-reducing microorganisms (SRMs) present in subsurface marine sediments belong to uncultured groups only distantly related to known SRMs, and it remains unclear how changing geochemical zones and sediment depth influence their community structure. We mapped the community composition and abundance of SRMs by amplicon sequencing and quantifying the dsrB gene, which encodes dissimilatory sulfite reductase subunit beta, in sediment samples covering different vertical geochemical zones ranging from the surface sediment to the deep sulfate-depleted subsurface at four locations in Aarhus Bay, Denmark. SRMs were present in all geochemical zones, including sulfate-depleted methanogenic sediment. The biggest shift in SRM community composition and abundance occurred across the transition from bioturbated surface sediments to nonbioturbated sediments below, where redox fluctuations and the input of fresh organic matter due to macrofaunal activity are absent. SRM abundance correlated with sulfate reduction rates determined for the same sediments. Sulfate availability showed a weaker correlation with SRM abundances and no significant correlation with the composition of the SRM community. The overall SRM species diversity decreased with depth, yet we identified a subset of highly abundant community members that persists across all vertical geochemical zones of all stations. We conclude that subsurface SRM communities assemble by the persistence of members of the surface community and that the transition from the bioturbated surface sediment to the unmixed sediment below is a main site of assembly of the subsurface SRM community.IMPORTANCE Sulfate-reducing microorganisms (SRMs) are key players in the marine carbon and sulfur cycles, especially in coastal sediments, yet little is understood about the environmental factors controlling their depth distribution. Our results suggest that macrofaunal activity is a key driver of SRM abundance and community structure in marine sediments and that a small subset of SRM species of high relative abundance in the subsurface SRM community persists from the sulfate-rich surface sediment to sulfate-depleted methanogenic subsurface sediment. More generally, we conclude that SRM communities inhabiting the subsurface seabed assemble by the selective survival of members of the surface community.
Project description:The factors controlling the relative abundances of Archaea and Bacteria in marine sediments are poorly understood. We determined depth distributions of archaeal and bacterial 16S rRNA genes by quantitative PCR at eight stations in Aarhus Bay, Denmark. Bacterial outnumber archaeal genes 10-60-fold in uppermost sediments that are irrigated and mixed by macrofauna. This bioturbation is indicated by visual observations of sediment color and faunal tracks, by porewater profiles of dissolved inorganic carbon and sulfate, and by distributions of unsupported 210Pb and 137Cs. Below the depth of bioturbation, the relative abundances of archaeal genes increase, accounting for one third of 16S rRNA genes in the sulfate zone, and half of 16S rRNA genes in the sulfate-methane transition zone and methane zone. Phylogenetic analyses reveal a strong shift in bacterial and archaeal community structure from bioturbated sediments to underlying layers. Stable isotopic analyses on organic matter and porewater geochemical gradients suggest that macrofauna mediate bacterial dominance and affect microbial community structure in bioturbated sediment by introducing fresh organic matter and high-energy electron acceptors from overlying seawater. Below the zone of bioturbation, organic matter content and the presence of sulfate exert key influences on bacterial and archaeal abundances and overall microbial community structure.
Project description:We further developed the stable isotope probing, magnetic-bead capture method to make it applicable for linking microbial community function to phylogeny at the class and family levels. The main improvements were a substantial decrease in the protocol blank and an approximately 10-fold increase in the detection limit by using a micro-elemental analyzer coupled to isotope ratio mass spectrometry to determine (13)C labeling of isolated 16S rRNA. We demonstrated the method by studying substrate utilization by Desulfobacteraceae, a dominant group of complete oxidizing sulfate-reducing Deltaproteobacteria in marine sediments. Stable-isotope-labeled [(13)C]glucose, [(13)C]propionate, or [(13)C]acetate was fed into an anoxic intertidal sediment. We applied a nested set of three biotin-labeled oligonucleotide probes to capture Bacteria, Deltaproteobacteria, and finally Desulfobacteraceae rRNA by using hydrophobic streptavidin-coated paramagnetic beads. The target specificities of the probes were examined with pure cultures of target and nontarget species and by determining the phylogenetic composition of the captured sediment rRNA. The specificity of the final protocol was generally very good, as more than 90% of the captured 16S rRNA belonged to the target range of the probes. Our results indicated that Desulfobacteraceae were important consumers of propionate but not of glucose. However, the results for acetate utilization were less conclusive due to lower and more variable labeling levels in captured rRNA. The main advantage of the method in this study over other nucleic acid-based stable isotope probing methods is that (13)C labeling can be much lower, to the extent that delta(13)C ratios can be studied even at their natural abundances.
Project description:Cold marine sediments harbor endospores of fermentative and sulfate-reducing, thermophilic bacteria. These dormant populations of endospores are believed to accumulate in the seabed via passive dispersal by ocean currents followed by sedimentation from the water column. However, the magnitude of this process is poorly understood because the endospores present in seawater were so far not identified, and only the abundance of thermophilic sulfate-reducing endospores in the seabed has been quantified. We investigated the distribution of thermophilic fermentative endospores (TFEs) in water column and sediment of Aarhus Bay, Denmark, to test the role of suspended dispersal and determine the rate of endospore deposition and the endospore abundance in the sediment. We furthermore aimed to determine the time course of reactivation of the germinating TFEs. TFEs were induced to germinate and grow by incubating pasteurized sediment and water samples anaerobically at 50°C. We observed a sudden release of the endospore component dipicolinic acid immediately upon incubation suggesting fast endospore reactivation in response to heating. Volatile fatty acids (VFAs) and H2 began to accumulate exponentially after 3.5 h of incubation showing that reactivation was followed by a short phase of outgrowth before germinated cells began to divide. Thermophilic fermenters were mainly present in the sediment as endospores because the rate of VFA accumulation was identical in pasteurized and non-pasteurized samples. Germinating TFEs were identified taxonomically by reverse transcription, PCR amplification and sequencing of 16S rRNA. The water column and sediment shared the same phylotypes, thereby confirming the potential for seawater dispersal. The abundance of TFEs was estimated by most probable number enumeration, rates of VFA production, and released amounts of dipicolinic acid during germination. The surface sediment contained ?105-106 inducible TFEs cm-3. TFEs thus outnumber thermophilic sulfate-reducing endospores by an order of magnitude. The abundance of cultivable TFEs decreased exponentially with sediment depth with a half-life of 350 years. We estimate that 6 × 109 anaerobic thermophilic endospores are deposited on the seafloor per m2 per year in Aarhus Bay, and that these thermophiles represent >10% of the total endospore community in the surface sediment.
Project description:Marine surface sediments, which are replete with sulfate, are typically considered to be devoid of endogenous methanogenesis. Yet, methanogenic archaea are present in those sediments, suggesting a potential for methanogenesis. We used an isotope dilution method based on sediment bag incubation and spiking with 13C-CH4 to quantify CH4 turnover rates in sediment from Aarhus Bay, Denmark. In two independent experiments, highest CH4 production and oxidation rates (>200 pmol cm-3 d-1) were found in the top 0-2 cm, below which rates dropped below 100 pmol cm-3 d-1 in all other segments down to 16 cm. This drop in overall methane turnover with depth was accompanied by decreasing rates of organic matter mineralization with depth. Molecular analyses based on quantitative PCR and MiSeq sequencing of archaeal 16S rRNA genes showed that the abundance of methanogenic archaea also peaked in the top 0-2 cm segment. Based on the community profiling, hydrogenotrophic and methylotrophic methanogens dominated among the methanogenic archaea in general, suggesting that methanogenesis in surface sediment could be driven by both CO2 reduction and fermentation of methylated compounds. Our results show the existence of elevated methanogenic activity and a dynamic recycling of CH4 at low concentration in sulfate-rich marine surface sediment. Considering the common environmental conditions found in other coastal systems, we speculate that such a cryptic methane cycling can be ubiquitous.