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Acetogens and Acetoclastic Methanosarcinales Govern Methane Formation in Abandoned Coal Mines

Beckmann, Sabrina, Lueders, Tillmann, Krüger, Martin, von Netzer, Frederick, Engelen, Bert, Cypionka, Heribert
Applied and environmental microbiology 2011 v.77 no.11 pp. 3749-3756
Clostridium, DNA probes, Methanosarcina barkeri, Pelobacter acetylenicus, acetates, bacteria, coal, denaturing gradient gel electrophoresis, hydrogen, methane, methane production, methanogens, microbial communities, mining, polymerase chain reaction
In abandoned coal mines, methanogenic archaea are responsible for the production of substantial amounts of methane. The present study aimed to directly unravel the active methanogens mediating methane release as well as active bacteria potentially involved in the trophic network. Therefore, the stable-isotope-labeled precursors of methane, [¹³C]acetate and H₂-¹³CO₂, were fed to liquid cultures from hard coal and mine timber from a coal mine in Germany. Guided by methane production rates, samples for DNA stable-isotope probing (SIP) with subsequent quantitative PCR and denaturing gradient gel electrophoretic (DGGE) analyses were taken over 6 months. Surprisingly, the formation of [¹³C]methane was linked to acetoclastic methanogenesis in both the [¹³C]acetate- and the H₂-¹³CO₂-amended cultures of coal and timber. H₂-¹³CO₂ was used mainly by acetogens related to Pelobacter acetylenicus and Clostridium species. Active methanogens, closely affiliated with Methanosarcina barkeri, utilized the readily available acetate rather than the thermodynamically more favorable hydrogen. Thus, the methanogenic microbial community appears to be highly adapted to the low-H₂ conditions found in coal mines.