Microbial methane turnover at mud volcanoes of the Gulf of Cadiz

Geomicrobiology Journal, 2008

Cadiz mud volcanoes (MV) is a mixture of deep sourced thermogenic CH 4 and shallow biogenic CH 4 . We directly investigated this possibility by comparing porewater CH 4 concentrations and their δ 13 C values with the potential for Archaeal methanogenesis in Gulf of Cadiz mud volcano (MV) sediments (Captain Arutyunov, Bonjardim, Ginsburg and Porto) using 14 C-rate measurements. The CH 4 has a deep sourced thermogenic origin (δ 13 C ∼ −49‰) but becomes 13 C-depleted in and beneath the zone of anaerobic oxidation of methane (AOM) where the rates of hydrogenotrophic methanogenesis increase. Thus we infer that a portion of AOMproduced CO 2 is being recycled to CH 4 by methanogens yielding further 13 C-depleted CH 4 , which might be misinterpreted as indicative of a fully shallow biogenic origin for this gas. Production of H 2 is related to compositional changes in sedimentary organic matter, or to upward flux of substrate-enriched fluids. In contrast to other MVs in the Gulf of Cadiz, Ginsburg MV fluids are enriched in SO 2− 4 and contain very high concentrations of acetate (2478 µM below 150 cmbsf); however, the high levels of acetate did not stimulate methanogenesis but instead were oxidized to CO 2 coupled to sulphate reduction. Both anaerobic oxidation of thermogenic CH 4 linked to shallow methanogenesis and fluid geochemistry control the recycling of deep-sourced carbon at Gulf of Cadiz MVs, impacting near-surface δ 13 C-CH 4 values.

Frontiers in Microbiology, 2023

The Gulf of Cádiz is a tectonically active continental margin with over sixty mud volcanoes (MV) documented, some associated with active methane (CH 4) seepage. However, the role of prokaryotes in influencing this CH 4 release is largely unknown. In two expeditions (MSM1-3 and JC10) seven Gulf of Cádiz MVs (Porto, Bonjardim, Carlos Ribeiro, Captain Arutyunov, Darwin, Meknes, and Mercator) were analyzed for microbial diversity, geochemistry, and methanogenic activity, plus substrate amended slurries also measured potential methanogenesis and anaerobic oxidation of methane (AOM). Prokaryotic populations and activities were variable in these MV sediments reflecting the geochemical heterogeneity within and between them. There were also marked differences between many MV and their reference sites. Overall direct cell numbers below the SMTZ (0.2-0.5 mbsf) were much lower than the general global depth distribution and equivalent to cell numbers from below 100 mbsf. Methanogenesis from methyl compounds, especially methylamine, were much higher than the usually dominant substrates H 2 /CO 2 or acetate. Also, CH 4 production occurred in 50% of methylated substrate slurries and only methylotrophic CH 4 production occurred at all seven MV sites. These slurries were dominated by Methanococcoides methanogens (resulting in pure cultures), and prokaryotes found in other MV sediments. AOM occurred in some slurries, particularly, those from Captain Arutyunov, Mercator and Carlos Ribeiro MVs. Archaeal diversity at MV sites showed the presence of both methanogens and ANME (Methanosarcinales, Methanococcoides, and ANME-1) related sequences, and bacterial diversity was higher than archaeal diversity, dominated by members of the Atribacterota, Chloroflexota, Pseudomonadota, Planctomycetota, Bacillota, and Ca. "Aminicenantes." Further work is essential to determine the full contribution of Gulf of Cádiz mud volcanoes to the global methane and carbon cycles.

The present study assesses the diversity and composition of sediment bacterial and microeukaryotic communities from deep-sea mud volcanoes (MVs) associated with strike-slip faults in the SouthWest Iberian Margin (SWIM). We used a 16S/18S rRNA gene based pyrosequencing approach to characterize and correlate the sediment bacterial and microeukaryotic communities from MVs with differing gas seep regimes and from an additional site with no apparent seeping activity. In general, our results showed significant compositional changes of bacterial and microeukaryotic communities in sampling sites with different seepage regimes. Sediment bacterial communities were enriched with Methylococcales (putative methanotrophs) but had lower abundances of Rhodospirillales, Nitrospirales and SAR202 in the more active MVs. Within microeukaryotic communities, members of the Lobosa (lobose amoebae) were enriched in more active MVs. We also showed a strong correlation between Methylococcales populations and lobose amoeba in active MVs. This study provides baseline information on the diversity and composition of bacterial and microeukaryotic communities in deep-sea MVs associated with strike-slip faults. Interactions between the Eurasian and African plates at the SWIM have resulted in a complex geological setting characterized by the extensive occurrence of mud volcanoes (MVs), at depths between 200 m and 5000 m 1–3. MVs are piercement structures, normally related to tectonic activity or petroleum reservoirs and are usually created by compression forces that promote the intrusion and seepage of pressurized fluids composed of gas, water, liquid hydrocarbons and solid particles 4. Methane is the most predominant gas expelled at MVs, which together with methane released from other types of seeps, increases atmospheric carbon content by 0.01–0.05 Gt yr −1 , accounting for 1 to 5% of global methane emissions to the atmosphere 5. Methane-oxidizing microorganisms mediate the methane flux in these ecosystems; providing energy sources to other microorganisms through predation and symbiosis, thus generating a hotspot of biomass in the deep-sea 6. Together, micro-and macroorganisms at methane seeps consume c. 75% of the methane that reaches the seafloor; thus serving as a biological filter that controls the emission of this greenhouse gas from the ocean to the atmosphere 5,7. In surface sediments of MV seeps, methane oxidation is usually performed by aerobic methanotrophic bacteria 8. In the subsurface deeper sediments it is mainly accomplished by anaerobic methanotrophic archaea in association with sulfate reducing bacteria, adding another reduced compound (hydrogen sulfide) to the seeping fluids 9,10. Some methanotrophic archaea and thiotrophic bacteria form symbiotic relationships with macroeukar-yotes, such as tubeworms and clams 9,11. There is evidence that the methane released by several MVs piercing a thick accretionary wedge in the Gulf of Cadiz (AWGC, SWIM) is almost totally consumed at the seabed, and therefore does not constitute a relevant source of methane to the hydrosphere and subsequently to the atmosphere 10. This is a strong indication of the presence of developed benthic communities involved in the uptake of methane. Although several studies have surveyed the microbial communities within MVs 10,12,13 , there is still considerable uncertainty regarding their

Sedimentary Geology, 2012

Methane oxidizing prokaryotes are ubiquitous in oxic and anoxic habitats wherever C 1 -compounds are present. Thus, methane saturated mud volcano fluids should be a preferred habitat of methane consuming prokaryotes, using the readily available electron donors. In order to understand the relevance of methane as a carbon and energy source in mud volcano communities, we investigate the diversity of prokaryotic organisms involved in oxidation of methane in fluid samples from the Salse di Nirano mud volcano field situated in the Northern Apennines. Cell counts were at approximately 0.7 × 10 6 microbial cells/ml. A fraction of the microbial biomass was identified as ANME (anaerobic methanotroph) archaea by fluorescence in situ hybridization (FISH) analysis. They are associated in densely colonized flakes, of some tens of μm in diameter, embedded in a hyaline matrix. Diversity analysis based on the 16S rDNA genes, retrieved from amplified and cloned environmental DNA, revealed a high proportion of archaea, involved in anaerobic oxidation of methane (AOM). Aerobic methane-oxidizing proteobacteria could be highly enriched from mud volcano fluids, indicating the presence of aerobic methanotrophic bacteria, which may contribute to methane oxidation, whenever oxygen is readily available. The results imply that biofilms, dominated by ANME archaea, colonize parts of the mud volcano venting system.

Many mud volcanoes were recently discovered in the euxinic bottom waters of the Sorokin Trough (NE Black Sea). Three of them, i.e., NIOZ, Odessa, and Kazakov, were selected for a detailed biogeochemical investigation. Four methane-related carbonate crusts covered with microbial mats, and sediments (dmud brecciaT) from these mud volcanoes were collected during the 11th Training-Through-Research cruise (TTR-11) in 2001, the first finding of methanotrophic microbial mats associated with authigenic carbonates in the deep Black Sea. We measured the concentrations and d 13 C values of methane and specific archaeal and bacterial lipids, and determined archaeal and bacterial 16S rRNA gene sequences. The d 13 C of the microbial lipids reflected the carbon isotopic values of the methane, indicating that methane was the main carbon source for microorganisms inducing carbonate formation. Anaerobic oxidation of methane (AOM) in these settings was performed by archaea affiliated with the so-called ANME-1 group. None of the identified archaeal sequences were closely related to known methanogens. The combined 16S rRNA gene sequence and biomarker data revealed a distinct difference in archaeal assemblage between the carbonate crusts and mud breccias. Besides gene sequences of sulfate-reducing bacteria, DNA analysis of bacterial communities revealed a diversity of bacteria with apparent contrasting metabolic properties. The methane utilization via AOM processes was detected in the uppermost sediments where it subsequently induces authigenic carbonate precipitation most probably below seafloor. The results of integrated biomarker and 16S rRNA gene study reveal a crucial role of AOM processes in formation of authigenic carbonates in methane seep environments. D

Applied and Environmental Microbiology, 2000

PLoS ONE, 2014

Numerous studies on marine prokaryotic communities have postulated that a process of anaerobic oxidation of methane (AOM) coupled with sulfate reduction (SR) is the main methane sink in the world's oceans. AOM has also been reported in the deep biosphere. But the responses of the primary microbial players in eliciting changes in geochemical environments, specifically in methane and sulfate supplies, have yet to be fully elucidated. Marine mud volcanoes (MVs) expel a complex fluid mixture of which methane is the primary component, forming an environment in which AOM is a common phenomenon. In this context, we attempted to identify how the prokaryotic community would respond to changes in methane and sulfate intensities, which often occur in MV environments in the form of eruptions, diffusions or seepage. We applied an integrated approach, including (i) biochemical surveys of pore water originated from MV, (ii) in vitro incubation of mud breccia, and (iii) prokaryotic community structure analysis. Two distinct AOM regions were clearly detected. One is related to the sulfate methane transition zone (SMTZ) at depth of 30-55 cm below the sea floor (bsf); the second is at 165-205 cm bsf with ten times higher rates of AOM and SR. This finding contrasts with the sulfide concentrations in pore waters and supports the suggestion that potential AOM activity below the SMTZ might be an important methane sink that is largely ignored or underestimated in oceanic methane budget calculations. Moreover, the incubation conditions below the SMTZ favor the growth of methanotrophic archaeal group ANME-2 compared to ANME-1, and promote the rapid growth and high diversity of bacterial communities. These incubation conditions also promote the increase of richness in bacterial communities. Our results provide direct evidence of the mechanisms by which deep AOM processes can affect carbon cycling in the deep biosphere and global methane biochemistry.

Biogeosciences

In this study, we report lipid biomarker patterns and phylogenetic identities of key microbial communities mediating anaerobic oxidation of methane (AOM) in active mud volcanoes (MVs) on the continental slope of the Canadian Beaufort Sea. The carbon isotopic compositions (δ 13 C) of sn-2and sn-3-hydroxyarchaeol showed the highly 13 Cdepleted values (−114 ‰ to −82 ‰) associated with a steep depletion in sulfate concentrations within 0.7 m of sediment depths. This suggested the presence of methanotrophic archaea involved in sulfate-dependent AOM, albeit in a small amount. The ratio of sn-2-hydroxyarchaeol to archaeol (> 1) and operational taxonomic units (OTUs) indicated that the anaerobic methanotrophic archaea (ANME) clades ANME-2c and ANME-3 were involved in AOM. Higher δ 13 C values of archaeol and biphytanes (BPs; −55.2 ± 10.0 ‰ and −39.3 ± 13.0 ‰, respectively) suggested that archaeal communities were also assimilating AOM-derived inorganic carbon. Furthermore, the distinct distribution patterns of methanotrophs in the three MVs appears to be associated with varying intensities of ascending gas fluids. Consequently, our results suggest that the niche diversification of active mud volcanoes has shaped distinct archaeal communities that play important roles in AOM in the Beaufort Sea.