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Producers and consumers of methane in an underwater mud volcano

Down to several meters below the bottom of the sea, microorganisms produce methane; on the contrary, others, may be the same, consume it… Researchers recovered sediment cores from a mud volcano at a depth of 2000 m in the Mediterranean, to learn more about these microbial communities, their diversity and their functioning.

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On continental margins separating tectonic plates, mud volcanoes are formed on the sea bottom by the expulsion of fluids, gas and mud originating from deep subsurface reservoirs. Located south of Turkey, the Amsterdam mud volcano studied here is part of the Anaximander Mountains associated with a very active tectonic deformation. Large amounts of methane (CH4) seep out of it; this gas is mainly produced by microorganisms belonging to the Archaea and is the final output of the biochemical degradation of the organic matter contained in the underwater sediments.

These species are the only known living organisms able to produce methane. Little is known about their diversity and their distribution within sediment layers. Yet it is known he some Archaea use methane for their own metabolism, with or without oxygen (aerobic or anaerobic conditions). The producers and consumers of methane (so called methanogens and methanotrophs) are phylogenetically so close that they could possibly belong to the same populations which could be able to invert their metabolism.

One reason why the study of these mechanisms is important is that the submarine emissions of methane (in particular those associated with mud volcanoes) have an obvious influence on climate: the contribution (over 100 years and for an equal mass) of methane to greenhouse effect is about 25 times greater than that of carbon dioxide.
The analysis of sediment cores several meters long, collected at depths of about 2000 m near the active center of the Amsterdam mud volcano, helped to know more about these microbial communities, particularly on their functioning and their role in the environment. The core was cut into 50 cm-long sections where samples were collected to analyze the chemical composition of the porewater circulating between the particles, measure the rate of methane production and study the molecular diversity of microorganisms.

 

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Lowering the core tube

Techniques of molecular biology and laboratory cultures were applied. The total DNA contained in each sediment sample were extracted and purified, then a particular gene were isolated and identically replicated to be studied, using the "polymerase chain reaction" (PCR) technique. This gene has the same function in all Bacteria and Archaea but with regions highly variable from species to species, making it a very interesting specific marker to study microbial diversity. The different versions of the gene were separated by migration on a support containing a chemical gradient (electrophoresis).

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Result of the electrophoresis: the black bands appearing at various depths (0 to 6.50 m) correspond to different sequences of one of the analyzed genes. Their number is related to the richness and diversity of the microbial community..

Another gene, found only in anaerobic methanogen and methanotroph Archaea, was amplified by PCR to be sequenced. For each sample, it was therefore possible to determine the sequence of amino-acids of the corresponding protein coded by the specific gene. The differences between these sequences lead to a phylogenetic classification, i.e. a tree describing their greater or lesser genetic proximity.

 

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One part of the phylogenetic classification where samples collected on the Amsterdam mud volcano are compared with Archean sequences indexd in international data bases..

 

The methanogen species find the carbon they use in methylated compounds (one of which is methanol), in carbon dioxide or acetate. In order to better understand the respective importance of these metabolic paths and their distribution according to depth within the sediment, methane production was measured in samples cultivated in enrichment media supplemented with one or another of these three sources of carbon.

The community of Archaea living in the sediments is very rich and very different in abundance and diversity according to depth. Producers (methanogens) and consumers (methanotrophs) can coexist, but the chemical composition of interstitial water in the shallowest layers results from a net consumption of methane. The heterogeneity of water composition and species distribution shows that there are very different ecological niches within the sediment.

The depth distribution of microorganisms representative of the metabolic paths leading to methane production is consistent with that of the genetic affiliations of the sequences. The path based on methylated compounds is found at all depths with genetic sequences very similar to species recorded in international culture collections. The acetate path is globally more important than the carbon dioxide one, which is quite uncommon in cold seepage sediments. Acetate concentration was also very high in interstitial water. However the processes active in the sediment between methane and carbon sources are highly complex and involve interactions between methanogen and methanotroph Archaea and between Archaea et Bacteria.

These results emphasize the possible interaction between the deep underlying reservoir and the surface sediments of the center of Amsterdam mud volcano. The acetate present there in abundance could have been generated by high temperature cracking of organic matter in the deeper sediment layers. The fluid flux carrying it towards the sediment surface could also bring the microorganisms present in the deep subsurface biosphere.

 

The paper

C. Lazar, R. Parkes, B. Cragg, S. L’Haridon, L. Toffin, 2012. Methanogenic activity and diversity in the centre of the Amsterdam Mud Volcano, Eastern Mediterranean Sea. FEMS Microbiology Ecology 81 : 243–254.
See the first page

 

The authors

This work was conducted by scientists from IUEM and Ifremer (Laboratoire de microbiologie des environnements extrêmes, UMR6197 CNRS/Ifremer/UBO), University of North Carolina (USA) and Université of Cardiff (Great Britain).

 

The journal

FEMS Microbiology Ecology is published by Blackwell and is one of the five journals of the Federation of European Microbiological Soceities (FEMS). Its field covers microorganism (bacteria, archaea, filamentous fungi, yeasts, protozoa, cyanobacteria, algae or viruses) and their environments (natural soil, aquatic and atmospheric habitats, including extreme environments, artificial or managed environments). Papers deal with topics as diverse as ecology, evolutionary biology, phylogenetic diversity, interactions with the environment and other organisms, etc.

 

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