ABSTRACT

The importance of anaerobic oxidation of methane (AOM) as methane sink in freshwater systems is largely unexplored, particularly in peat ecosystems. Nitrite-dependent anaerobic methane oxidation (n-damo) was recently discovered and reported to be catalyzed by the bacterium ‘Candidatus Methylomirabilis oxyfera’ that is affiliated with the NC10 phylum. So far, several M. oxyfera enrichment cultures have been obtained using a limited number of fresh water sediments or wastewater treatment sludge as inoculum. In this study, using stable isotope measurements and pore water profiles, we investigated the potential of n-damo in a minerotrophic peat land in the south of the Netherlands that is infiltrated by nitrate-rich ground water. Methane and nitrate profiles suggested that all methane produced was oxidized before reaching the oxic layer, and NC10 bacteria could be active in the transition zone where counter-gradients of methane and nitrate occur. Quantitative PCR showed high NC10 bacterial cell numbers at this methane-nitrate transition zone. This soil section was used to enrich the prevalent NC10 bacteria in a continuous culture supplied with methane and nitrite at an in situ pH of 6.2. An enrichment of nitrite-reducing methanotrophic NC10 bacteria was successfully obtained. Phylogenetic analysis of retrieved 16S rRNA and pmoA genes showed that the enriched bacteria were very similar to the ones found in situ, and constituted a new branch of NC10 bacteria, with an identity percentage of less than 96% and 88% to the 16S rRNA and pmoA genes of M. oxyfera, respectively. The results of this study expand our knowledge of the diversity and distribution of NC10 bacteria in the environment, and highlight their potential contribution to nitrogen and methane cycles.