Microbial degradation and assimilation of veratric acid in oxic and anoxic groundwaters.

Microbial communities are key players in groundwater ecosystems. In this dark environment, heterotrophic microbes rely on biomass produced by the activity of lithoautotrophs or on the degradation of organic matter seeping from the surface. Most studies on bacterial diversity in groundwater habitats are based on 16S gene sequencing and full genome reconstructions showing potential metabolic pathways used in these habitats. However, molecular-based studies do not allow for the assessment of population dynamics over time or the assimilation of specific compounds and their biochemical transformation by microbial communities. Therefore, in this study, we combined DNA-, phospholipid fatty acid-, and metabolomic-stable isotope probing to target and identify heterotrophic bacteria in the groundwater setting of the Hainich Critical Zone Exploratory (CZE), focusing on 2 aquifers with different physico-chemical conditions (oxic and anoxic). We incubated groundwater from 4 different wells using either 13C-labeled veratric acid (a lignin-derived compound) (single labeling) or a combination of 13CO2 and D-labeled veratric acid (dual labeling). Our results show that heterotrophic activities dominate all groundwater sites. We identified bacteria with the potential to break down veratric acid (Sphingobium or Microbacterium). We observed differences in heterotrophic activities between the oxic and anoxic aquifers, indicating local adaptations of bacterial populations. The dual labeling experiments suggested that the serine pathway is an important carbon assimilation pathway and that organic matter was an important source of hydrogen in the newly produced lipids. These experiments also yielded different labeled taxa compared to the single labeling experiments, showing that there exists a complex interaction network in the groundwater habitats.

Deep Isolated Aquifer Brines Harbor Atypical Halophilic Microbial Communities in Quebec, Canada.

The deep terrestrial subsurface, hundreds of meters to kilometers below the surface, is characterized by oligotrophic conditions, dark and often anoxic settings, with fluctuating pH, salinity, and water availability. Despite this, microbial populations are detected and active, contributing to biogeochemical cycles over geological time. Because it is extremely difficult to access the deep biosphere, little is known about the identity and metabolisms of these communities, although they likely possess unknown pathways and might interfere with deep waste deposits. Therefore, we analyzed rock and groundwater microbial communities from deep, isolated brine aquifers in two regions dating back to the Ordovician and Devonian, using amplicon and whole genome sequencing. We observed significant differences in diversity and community structure between both regions, suggesting an impact of site age and composition. The deep hypersaline groundwater did not contain typical halophilic bacteria, and genomes suggested pathways involved in protein and hydrocarbon degradation, and carbon fixation. We identified mainly one strategy to cope with osmotic stress: compatible solute uptake and biosynthesis. Finally, we detected many bacteriophage families, potentially indicating that bacteria are infected. However, we also found auxiliary metabolic genes in the viral genomes, probably conferring an advantage to the infected hosts.

Linking Groundwater to Surface Discharge Ecosystems: Archaeal, Bacterial, and Eukaryotic Community Diversity and Structure in Quebec (Canada).

Aquifer systems are composed of water flowing from surface recharge areas, to the subsurface and back to the surface in discharge regions. Groundwater habitats harbor a large microbial biomass and diversity, potentially contributing to surface aquatic ecosystems. Although this contribution has been widely studied in marine environments, very little is known about the connection between underground and surface microbial communities in freshwater settings. Therefore, in this study, we used amplicon sequencing to analyze the archaeal, bacterial, and eukaryotic community diversity and structure in groundwater and surface water samples, spanning the vast regions of the Laurentides and Lanaudières in the Quebec province (Canada). Our results show significant differences between subsurface and surface taxa; with more fungi, Amoebozoa, and chemolithoautotrophic prokaryotes involved in nitrogen-, sulfur-, and iron-cycling dominating the underground samples; while algae, ciliates, methanogens, and Actinobacteria dominate the surface discharge waters. Microbial source tracking suggested that only a small portion of the microbial communities in the groundwater contributed to the surface discharge communities. However, many taxa were shared between both habitats, with a large range of functional diversity, likely explaining their survival in both subsurface and surface water ecosystems.

Effect of Snowmelt on Groundwater Bacterial Community Composition and Potential Role of Surface Environments as Microbial Seed Bank in Two Distinct Aquifers from the Region of Quebec, Canada.

Events of groundwater recharge are associated with changes in the composition of aquifer microbial communities but also abiotic conditions. Modification in the structure of the community can be the result of different environmental condition favoring or hindering certain taxa, or due to the introduction of surface-derived taxa. Yet, in both cases, the local hydrogeochemical settings of the aquifer is likely to affect the amount of variation observed. Therefore, in our study, we used 16S rRNA gene sequencing to assess how microbial communities change in response to snowmelt and the potential connectivity between subsurface and surface microbiomes in two distinct aquifers located in the region of Vaudreuil-Soulanges (Québec, Canada). At both sites, we observed an increase in groundwater level and decrease in temperature following the onset of snow melt in March 2019. Bacterial community composition of each aquifer was significantly different (p < 0.05) between samples collected prior and after groundwater recharge. Furthermore, microbial source tracking results suggested a low contribution of surface environments to the groundwater microbiome except for in the months associated with recharge (March 2019 and April 2019). Overall, despite differences in soil permeability between both sites, the period of snow melt was followed by important changes in the composition of microbial communities from aquifers.

Ecological processes differ in community assembly of Archaea, Bacteria and Eukaryotes in a biogeographical survey of groundwater habitats in the Quebec region (Canada).

Aquifers are inhabited by microorganisms from the three major domains of life: Archaea, Eukaryotes and Bacteria. Although interest in the processes that govern the assembly of these microbial communities is growing, their study is almost systematically limited to one of the three domains of life. Archaea, Bacteria and Eukaryotes are however interconnected and essential to understand the functioning of their living ecosystems. We, therefore, conducted a spatial study of the distribution of microorganisms by sampling 35 wells spread over an area of 10,000 km2 in the Quebec region (Canada). The obtained data allowed us to define the impact of geographic distance and geochemical water composition on the microbial communities. A null model approach was used to infer the relative influence of stochastic and determinist ecological processes on the assembly of the microbial community from all three domains. We found that the organisms from these three groups are mainly governed by stochastic mechanisms. However, this apparent similarity does not reflect the differences in the processes that govern the phyla assembly. The results obtained highlight the importance of considering all the microorganisms without neglecting their individual specificities.

Laboratory-Controlled Experiments Reveal Microbial Community Shifts during Sediment Resuspension Events.

In freshwater ecosystems, dynamic hydraulic events (floods or dam maintenance) lead to sediment resuspension and mixing with waters of different composition. Microbial communities living in the sediments play a major role in these leaching events, contributing to organic matter degradation and the release of trace elements. However, the dynamics of community diversity are seldom studied in the context of ecological studies. Therefore, we carried out laboratory-induced leaching experiments, using sediments from the Villerest dam reservoir (Villerest, France). To assess whole microbial community diversity, we sequenced the archaeal and bacterial 16S rRNA genes using Illumina MiSeq. Our results suggest that the degree of dissolved oxygen found in the water during these resuspension episodes influenced community dynamics, with anoxic waters leading to drastic shifts in sedimentary communities compared to oxic waters. Furthermore, the release of microbial cells from sediments to the water column were more favorable to water colonization when events were caused by oxic waters. Most of the bacteria found in the sediments were chemoorganotrophs and most of the archaea were methanogens. Methylotrophic, as well as archaeal, and bacterial chemoorganotrophs were detected in the leachate samples. These results also show that organic matter degradation occurred, likely participating in carbonate dissolution and the release of trace elements during freshwater resuspension events.

From Recharge, to Groundwater, to Discharge Areas in Aquifer Systems in Quebec (Canada): Shaping of Microbial Diversity and Community Structure by Environmental Factors.

Groundwater recharge and discharge rates and zones are important hydrogeological characteristics of aquifer systems, yet their impact on the formation of both subterranean and surface microbiomes remains largely unknown. In this study, we used 16S rRNA gene sequencing to characterize and compare the microbial community of seven different aquifers, including the recharge and discharge areas of each system. The connectivity between subsurface and surface microbiomes was evaluated at each site, and the temporal succession of groundwater microbial communities was further assessed at one of the sites. Bacterial and archaeal community composition varied between the different sites, reflecting different geological characteristics, with communities from unconsolidated aquifers being distinct from those of consolidated aquifers. Our results also revealed very little to no contribution of surface recharge microbial communities to groundwater communities as well as little to no contribution of groundwater microbial communities to surface discharge communities. Temporal succession suggests seasonal shifts in composition for both bacterial and archaeal communities. This study demonstrates the highly diverse communities of prokaryotes living in aquifer systems, including zones of groundwater recharge and discharge, and highlights the need for further temporal studies with higher resolution to better understand the connectivity between surface and subsurface microbiomes.

Microbial diversity gradients in the geothermal mud volcano underlying the hypersaline Urania Basin.

Mud volcanoes transport deep fluidized sediment and their microbial communities and thus provide a window into the deep biosphere. However, mud volcanoes are commonly sampled at the surface and not probed at greater depths, with the consequence that their internal geochemistry and microbiology remain hidden from view. Urania Basin, a hypersaline seafloor basin in the Mediterranean, harbors a mud volcano that erupts fluidized mud into the brine. The vertical mud pipe was amenable to shipboard Niskin bottle and multicorer sampling and provided an opportunity to investigate the downward sequence of bacterial and archaeal communities of the Urania Basin brine, fluid mud layers and consolidated subsurface sediments using 16S rRNA gene sequencing. These microbial communities show characteristic, habitat-related trends as they change throughout the sample series, from extremely halophilic bacteria (KB1) and archaea (Halodesulfoarchaeum spp.) in the brine, toward moderately halophilic and thermophilic endospore-forming bacteria and uncultured archaeal lineages in the mud fluid, and finally ending in aromatics-oxidizing bacteria, uncultured spore formers, and heterotrophic subsurface archaea (Thermoplasmatales, Bathyarchaeota, and Lokiarcheota) in the deep subsurface sediment at the bottom of the mud volcano. Since these bacterial and archaeal lineages are mostly anaerobic heterotrophic fermenters, the microbial ecosystem in the brine and fluidized mud functions as a layered fermenter for the degradation of sedimentary biomass and hydrocarbons. By spreading spore-forming, thermophilic Firmicutes during eruptions, the Urania Basin mud volcano likely functions as a source of endospores that occur widely in cold seafloor sediments.

The endolithic bacterial diversity of shallow bedrock ecosystems.

Terrestrial subsurface microbial communities are not restricted to the fluid-filled void system commonly targeted during groundwater sampling but are able to inhabit and dwell in rocks. However, compared to the exploration of the deep biosphere, endolithic niches in shallow sedimentary bedrock have received little interest so far. Despite the potential contribution of rock matrix dwellers to matter cycling and groundwater resource quality, their identity and diversity patterns are largely unknown. Here, we investigated the bacterial diversity in twenty-two rock cores in common limestone-mudstone alternations that differed in rock permeabilities and other geostructural and petrological factors. 16S rRNA gene analysis showed the existence of a unique rock matrix microbiome compared to surrounding groundwater. Typically, shallow weathered limestones contained bacterial groups most likely originating from soil habitats. In low-permeable mudstones, we found similar communities of oligotrophic heterotrophs, and thiosulfate-oxidizing autotrophs, without relation to depth, rock type and bulk rock permeability. In fractured limestone, the bacterial communities of fracture surfaces were distinct from their matrix counterparts and ranged from organic matter decomposers in outcrop areas to autotrophs in downdip positions that receive limited surface input. Contrastingly, rock matrices from lithologically corresponding, but highly isolated environments, were dominated by spore-forming bacteria, oligotrophic heterotrophs and hydrogen-oxidizing autotrophs. Neither depth, matrix permeability nor major mineralogy dominantly controlled the endolithic bacterial diversity. Instead, a combination of subsurface factors drives the supply of niches by fluids, matter and energy as well as the (re)dispersal conditions that likely shape bacterial diversity.

Archaeal Diversity and CO2 Fixers in Carbonate-/Siliciclastic-Rock Groundwater Ecosystems.

Groundwater environments provide habitats for diverse microbial communities, and although Archaea usually represent a minor fraction of communities, they are involved in key biogeochemical cycles. We analysed the archaeal diversity within a mixed carbonate-rock/siliciclastic-rock aquifer system, vertically from surface soils to subsurface groundwater including aquifer and aquitard rocks. Archaeal diversity was also characterized along a monitoring well transect that spanned surface land uses from forest/woodland to grassland and cropland. Sequencing of 16S rRNA genes showed that only a few surface soil-inhabiting Archaea were present in the groundwater suggesting a restricted input from the surface. Dominant groups in the groundwater belonged to the marine group I (MG-I) Thaumarchaeota and the Woesearchaeota. Most of the groups detected in the aquitard and aquifer rock samples belonged to either cultured or predicted lithoautotrophs (e.g., Thaumarchaeota or Hadesarchaea). Furthermore, to target autotrophs, a series of 13CO2 stable isotope-probing experiments were conducted using filter pieces obtained after filtration of 10,000 L of groundwater to concentrate cells. These incubations identified the SAGMCG Thaumarchaeota and Bathyarchaeota as groundwater autotrophs. Overall, the results suggest that the majority of Archaea on rocks are fixing CO2, while archaeal autotrophy seems to be limited in the groundwater.

Genomic inference of the metabolism of cosmopolitan subsurface Archaea, Hadesarchaea.

The subsurface biosphere is largely unexplored and contains a broad diversity of uncultured microbes(1). Despite being one of the few prokaryotic lineages that is cosmopolitan in both the terrestrial and marine subsurface(2-4), the physiological and ecological roles of SAGMEG (South-African Gold Mine Miscellaneous Euryarchaeal Group) Archaea are unknown. Here, we report the metabolic capabilities of this enigmatic group as inferred from genomic reconstructions. Four high-quality (63-90% complete) genomes were obtained from White Oak River estuary and Yellowstone National Park hot spring sediment metagenomes. Phylogenomic analyses place SAGMEG Archaea as a deeply rooting sister clade of the Thermococci, leading us to propose the name Hadesarchaea for this new Archaeal class. With an estimated genome size of around 1.5 Mbp, the genomes of Hadesarchaea are distinctly streamlined, yet metabolically versatile. They share several physiological mechanisms with strict anaerobic Euryarchaeota. Several metabolic characteristics make them successful in the subsurface, including genes involved in CO and H2 oxidation (or H2 production), with potential coupling to nitrite reduction to ammonia (DNRA). This first glimpse into the metabolic capabilities of these cosmopolitan Archaea suggests they are mediating key geochemical processes and are specialized for survival in the subsurface biosphere.

Extremophile microbiomes in acidic and hypersaline river sediments of Western Australia.

We investigated the microbial community compositions in two sediment samples from the acidic (pH ∼3) and hypersaline (>4.5% NaCl) surface waters, which are widespread in Western Australia. In West Dalyup River, large amounts of NaCl, Fe(II) and sulfate are brought by the groundwater into the surface run-off. The presence of K-jarosite and schwertmannite minerals in the river sediments suggested the occurrence of microbial Fe(II) oxidation because chemical oxidation is greatly reduced at low pH. 16S rRNA gene diversity analyses revealed that sequences affiliated with an uncultured archaeal lineage named Aplasma, which has the genomic potential for Fe(II) oxidation, were dominant in both sediment samples. The acidophilic heterotrophs Acidiphilium and Acidocella were identified as the dominant bacterial groups. Acidiphilium strain AusYE3-1 obtained from the river sediment tolerated up to 6% NaCl at pH 3 under oxic conditions and cells of strain AusYE3-1 reduced the effects of high salt content by forming filamentous structure clumping as aggregates. Neither growth nor Fe(III) reduction by strain AusYE3-1 was observed in anoxic salt-containing medium. The detection of Aplasma group as potential Fe(II) oxidizers and the inhibited Fe(III)-reducing capacity of Acidiphilium contributes to our understanding of the microbial ecology of acidic hypersaline environments.

Genomic evidence for distinct carbon substrate preferences and ecological niches of Bathyarchaeota in estuarine sediments.

Investigations of the biogeochemical roles of benthic Archaea in marine sediments are hampered by the scarcity of cultured representatives. In order to determine their metabolic capacity, we reconstructed the genomic content of four widespread uncultured benthic Archaea recovered from estuary sediments at 48% to 95% completeness. Four genomic bins were found to belong to different subgroups of the former Miscellaneous Crenarcheota Group (MCG) now called Bathyarchaeota: MCG-6, MCG-1, MCG-7/17 and MCG-15. Metabolic predictions based on gene content of the different genome bins indicate that subgroup 6 has the ability to hydrolyse extracellular plant-derived carbohydrates, and that all four subgroups can degrade detrital proteins. Genes encoding enzymes involved in acetate production as well as in the reductive acetyl-CoA pathway were detected in all four genomes inferring that these Archaea are organo-heterotrophic and autotrophic acetogens. Genes involved in nitrite reduction were detected in all Bathyarchaeota subgroups and indicate a potential for dissimilatory nitrite reduction to ammonium. Comparing the genome content of the different Bathyarchaeota subgroups indicated preferences for distinct types of carbohydrate substrates and implicitly, for different niches within the sedimentary environment.

Genomic resolution of linkages in carbon, nitrogen, and sulfur cycling among widespread estuary sediment bacteria.

BACKGROUND: Estuaries are among the most productive habitats on the planet. Bacteria in estuary sediments control the turnover of organic carbon and the cycling of nitrogen and sulfur. These communities are complex and primarily made up of uncultured lineages, thus little is known about how ecological and metabolic processes are partitioned in sediments. RESULTS: De novo assembly and binning resulted in the reconstruction of 82 bacterial genomes from different redox regimes of estuary sediments. These genomes belong to 23 bacterial groups, including uncultured candidate phyla (for example, KSB1, TA06, and KD3-62) and three newly described phyla (White Oak River (WOR)-1, WOR-2, and WOR-3). The uncultured phyla are generally most abundant in the sulfate-methane transition (SMTZ) and methane-rich zones, and genomic data predict that they mediate essential biogeochemical processes of the estuarine environment, including organic carbon degradation and fermentation. Among the most abundant organisms in the sulfate-rich layer are novel Gammaproteobacteria that have genes for the oxidation of sulfur and the reduction of nitrate and nitrite. Interestingly, the terminal steps of denitrification (NO3 to N2O and then N2O to N2) are present in distinct bacterial populations. CONCLUSIONS: This dataset extends our knowledge of the metabolic potential of several uncultured phyla. Within the sediments, there is redundancy in the genomic potential in different lineages, often distinct phyla, for essential biogeochemical processes. We were able to chart the flow of carbon and nutrients through the multiple geochemical layers of bacterial processing and reveal potential ecological interactions within the communities.

Environmental controls on intragroup diversity of the uncultured benthic archaea of the miscellaneous Crenarchaeotal group lineage naturally enriched in anoxic sediments of the White Oak River estuary (North Carolina, USA).

Sediments of the White Oak River (WOR) estuary are situated on the coast of North Carolina harbour, one of the most diverse known populations of uncultured Archaea, specifically the miscellaneous Crenarchaeotal group (MCG). In order to constrain the environmental factors influencing the uncultured archaeal groups in the WOR estuary, biogeochemical profiles as well as archaeal 16S rRNA genes from sediment pushcores were analysed. The relative fraction of MCG Archaea in clone libraries decreased at shallow sediment depths (27% of the total MCG). A LINKTREE analysis of the MCG intragroup diversity reinforced the observation that the MCG subgroup 6 was found predominantly within sulfide-depleted shallow sediment layers; other subgroups (especially MCG-1 and MCG-5/8) occurred preferentially in deeper, more strongly reducing sediment layers. The available evidence from this study and published MCG distribution patterns indicates that the MCG-6 subgroup is a specialized MCG lineage that, in contrast to other MCG subgroups, prefers suboxic sediment horizons with minimal or no free sulfide. Collectively, our results reveal the habitat preferences of different MCG subgroups in the WOR sediments and suggest that physiological adaptations to distinct sedimentary geochemical niches evolved in different MCG subgroups.

Methanogenic activity and diversity in the centre of the Amsterdam Mud Volcano, Eastern Mediterranean Sea.

Marine mud volcanoes are geological structures emitting large amounts of methane from their active centres. The Amsterdam mud volcano (AMV), located in the Anaximander Mountains south of Turkey, is characterized by intense active methane seepage produced in part by methanogens. To date, information about the diversity or the metabolic pathways used by the methanogens in active centres of marine mud volcanoes is limited. (14)C-radiotracer measurements showed that methylamines/methanol, H(2)/CO(2) and acetate were used for methanogenesis in the AMV. Methylotrophic methanogenesis was measured all along the sediment core, Methanosarcinales affiliated sequences were detected using archaeal 16S PCR-DGGE and mcrA gene libraries, and enrichments of methanogens showed the presence of Methanococcoides in the shallow sediment layers. Overall acetoclastic methanogenesis was higher than hydrogenotrophic methanogenesis, which is unusual for cold seep sediments. Interestingly, acetate porewater concentrations were extremely high in the AMV sediments. This might be the result of organic matter cracking in deeper hotter sediment layers. Methane was also produced from hexadecanes. For the most part, the methanogenic community diversity was in accordance with the depth distribution of the H(2)/CO(2) and acetate methanogenesis. These results demonstrate the importance of methanogenic communities in the centres of marine mud volcanoes.

Distribution of anaerobic methane-oxidizing and sulfate-reducing communities in the G11 Nyegga pockmark, Norwegian Sea.

Pockmarks are seabed geological structures sustaining methane seepage in cold seeps. Based on RNA-derived sequences the active fraction of the archaeal community was analysed in sediments associated with the G11 pockmark, in the Nyegga region of the Norwegian Sea. The anaerobic methanotrophic Archaea (ANME) and sulfate-reducing bacteria (SRB) communities were studied as well. The vertical distribution of the archaeal community assessed by PCR-DGGE highlighted the presence of ANME-2 in surface sediments, and ANME-1 in deeper sediments. Enrichments of methanogens showed the presence of hydrogenotrophic methanogens of the Methanogenium genus in surface sediment layers as well. The active fraction of the archaeal community was uniquely composed of ANME-2 in the shallow sulfate-rich sediments. Functional methyl coenzyme M reductase gene libraries showed that sequences affiliated with the ANME-1 and ANME-3 groups appeared in the deeper sediments but ANME-2 dominated both surface and deeper layers. Finally, dissimilatory sulfite reductase gene libraries revealed a high SRB diversity (i.e. Desulfobacteraceae, Desulfobulbaceae, Syntrophobacteraceae and Firmicutes) in the shallow sulfate-rich sediments. The SRB diversity was much lower in the deeper section. Overall, these results show that the microbial community in sediments associated with a pockmark harbour classical cold seep ANME and SRB communities.

Methanogenic diversity and activity in hypersaline sediments of the centre of the Napoli mud volcano, Eastern Mediterranean Sea.

Submarine mud volcanoes are a significant source of methane to the atmosphere. The Napoli mud volcano, situated in the brine-impacted Olimpi Area of the Eastern Mediterranean Sea, emits mainly biogenic methane particularly at the centre of the mud volcano. Temperature gradients support the suggestion that Napoli is a cold mud volcano with moderate fluid flow rates. Biogeochemical and molecular genetic analyses were carried out to assess the methanogenic activity rates, pathways and diversity in the hypersaline sediments of the centre of the Napoli mud volcano. Methylotrophic methanogenesis was the only significant methanogenic pathway in the shallow sediments (0-40 cm) but was also measured throughout the sediment core, confirming that methylotrophic methanogens could be well adapted to hypersaline environments. Hydrogenotrophic methanogenesis was the dominant pathway below 50 cm; however, low rates of acetoclastic methanogenesis were also present, even in sediment layers with the highest salinity, showing that these methanogens can thrive in this extreme environment. PCR-DGGE and methyl coenzyme M reductase gene libraries detected sequences affiliated with anaerobic methanotrophs (mainly ANME-1) as well as Methanococcoides methanogens. Results show that the hypersaline conditions in the centre of the Napoli mud volcano influence active biogenic methane fluxes and methanogenic/methylotrophic diversity.

Archaeal populations in hypersaline sediments underlying orange microbial mats in the Napoli mud volcano.

Microbial mats in marine cold seeps are known to be associated with ascending sulfide- and methane-rich fluids. Hence, they could be visible indicators of anaerobic oxidation of methane (AOM) and methane cycling processes in underlying sediments. The Napoli mud volcano is situated in the Olimpi Area that lies on saline deposits; from there, brine fluids migrate upward to the seafloor. Sediments associated with a brine pool and microbial orange mats of the Napoli mud volcano were recovered during the Medeco cruise. Based on analysis of RNA-derived sequences, the "active" archaeal community was composed of many uncultured lineages, such as rice cluster V or marine benthic group D. Function methyl coenzyme M reductase (mcrA) genes were affiliated with the anaerobic methanotrophic Archaea (ANME) of the ANME-1, ANME-2a, and ANME-2c groups, suggesting that AOM occurred in these sediment layers. Enrichment cultures showed the presence of viable marine methylotrophic Methanococcoides in shallow sediment layers. Thus, the archaeal community diversity seems to show that active methane cycling took place in the hypersaline microbial mat-associated sediments of the Napoli mud volcano.

Active archaeal communities at cold seep sediments populated by Siboglinidae tubeworms from the Storegga Slide.

Siboglinid tubeworms in cold seep sediments can locally modify the geochemical gradients of electron acceptors and donors, hence creating potential microhabitats for prokaryotic populations. The archaeal communities associated with sediments populated by Oligobrachia haakonmosbiensis and Sclerolinum contortum Siboglinid tubeworms in the Storegga Slide were examined in this study. Vertical distribution of archaeal communities was investigated using denaturing gradient gel electrophoresis based on 16S rRNA genes. The active fraction of the archaeal community was assessed by using reverse-transcribed rRNA. Archaeal communities associated with sediments colonized by tubeworms were affiliated with uncultivated archaeal lineages of the Crenarchaeota and Euryarchaeota. The composition of the active archaeal populations changed with depth indicating a reorganization of microbial communities. 16S rRNA gene libraries were dominated by sequences affiliated to the Rice Cluster V which are unusual in marine sediment samples. Moreover, this study provides the first evidence of living Crenarchaeota of the Rice Cluster V in cold seep sediments. Furthermore, the Storegga Slide sediments harbored a high diversity of other minor groups of uncultivated lineages including Terrestrial Miscellaneous Euryarchaeotal Group, Marine Benthic Group (MBG)-D, MBG-E, Deep-Sea Hydrothermal Vent Euryarchaeotal Group, Lake Dagow Sediment, Val Kotinen Lake clade III, and Sippenauer Moor 1. Thus, we hypothesize that the vertical geochemical imprint created by the tubeworms could support broad active archaeal populations in the Siboglinidae-populated Storegga Slide sediments.