Selective Enrichment of Methylococcaceae versus Methylocystaceae Methanotrophs via Control of Methane Feeding Schemes

Methanotrophs are crucial in keeping environmental CH₄ emissions in check. However, the contributions of different groups of methanotrophs at terrestrial CH₄-oxidation hotspots, such as the oxic-anoxic interface of rice paddies, have shown considerable inconsistency across observations. To address the knowledge gap regarding this inconsistency, methanotrophic microbiomes were enriched from paddy soils in well-mixed CH₄-fed batch reactors under six different incubation conditions, prepared as combinations of two CH₄ mixing ratios (0.5 and 10%) and three supplemented Cu²⁺ concentrations (0, 2, and 10 μM). Monitoring of temporal community shifts in these cultures revealed a dominance of Methylocystis spp. in all 0.5%-CH₄ cultures, while methanotrophs affiliated to Gammaproteobacteria dominated the 10%-CH₄ cultures that were less consistent both temporally and across conditions. The shotgun metagenome analyses of the 0.5%-CH₄ cultures corroborated the Methylocystis dominance and, interestingly, showed that copper deficiency did not select for mmoXYZ-possessing methanotrophs. Instead, a mbn cluster, accounting for approximately 5% of the Methylocystis population, was identified, suggesting the ecological significance of methanobactin in Cu-deficient methanotrophy. These findings underscore the important role of Methylocystis spp. in mitigating emissions from terrestrial CH₄ hotspots and suggest the feasibility of directed enrichment and/or isolation of Methylocystis spp. for utilization in, for example, methanobactin and polyhydroxybutyrate production.