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Abstract

Methyl substrates are important compounds for methanogenesis in marine
sediments but diversity and carbon utilization by methylotrophic
methanogenic archaea have not been clarified. Here, we demonstrate that
RNA-stable isotope probing (SIP) requires C-13-labeled bicarbonate as
co-substrate for identification of methylotrophic methanogens in
sediment samples of the Helgoland mud area, North Sea. Using lipid-SIP,
we found that methylotrophic methanogens incorporate 60-86% of dissolved
inorganic carbon (DIC) into lipids, and thus considerably more than what
can be predicted from known metabolic pathways (similar to 40%
contribution). In slurry experiments amended with the marine
methylotroph Methanococcoides methylutens, up to 12% of methane was
produced from CO2, indicating that CO2-dependent methanogenesis is an
alternative methanogenic pathway and suggesting that obligate
methylotrophic methanogens grow in fact mixotrophically on methyl
compounds and DIC. Although methane formation from methanol is the
primary pathway of methanogenesis, the observed high DIC incorporation
into lipids is likely linked to CO2-dependent methanogenesis, which was
triggered when methane production rates were low. Since methylotrophic
methanogenesis rates are much lower in marine sediments than under
optimal conditions in pure culture, CO2 conversion to methane is an
important but previously overlooked methanogenic process in sediments
for methylotrophic methanogens.