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Aerobic methanotrophs are crucial in ombrotrophic peatlands, driving the methane and nitrogen cycles. Peat mining adversely affects methanotrophs, but activity and community composition/abundances may recover after restoration. Considering that the methanotrophic activity and growth are significantly stimulated in the presence of other microorganisms, the methane-driven interaction network, which encompasses methanotrophs and nonmethanotrophs (i.e., the methanotrophic interactome), may also be relevant in conferring community resilience. Yet, little is known of the methanotrophic interactome's response to and recovery from disturbances. Here, we determined the recovery of the methanotrophic interactome as inferred by a co-occurrence network analysis comparing pristine and restored peatlands. We coupled a DNA-based stable isotope probing (SIP) approach using [ 13C]CH4 to a co-occurrence network analysis derived from the 13C-enriched 16S rRNA gene sequences to relate the response in methanotrophic activity to the structuring of the interaction network. Methanotrophic activity and abundances recovered after peat restoration since 2000. "Methylomonaceae" taxa were the predominantly active methanotrophs in both peatlands, but the peatlands differed in the relative abundances of Methylacidiphilaceae and Methylocystis. However, bacterial community compositions were distinct in both peatlands. Likewise, the methanotrophic interactome was profoundly altered in the restored peatland. Structuring of the interaction network after peat mining resulted in the loss of complexity and modularity, indicating a less connected and efficient network, which may have consequences in the event of recurring/future disturbances. Therefore, determining the response of the methane-driven interaction network, in addition to relating methanotrophic activity to community composition/abundances, provided a more comprehensive understanding of the resilience of the methanotrophs. [ABSTRACT FROM AUTHOR]

Microorganisms interact in complex communities, affecting microbially-mediated processes in the environment. Particularly, aerobic methanotrophs showed significantly stimulated growth and activity in the presence of accompanying microorganisms in an interaction network (interactome). Yet, little is known of how the interactome responds to disturbances, and how community functioning is affected by the disturbance-induced structuring of the interaction network. Here, we employed a time-series stable isotope probing (SIP) approach using 13C–CH 4 coupled to a co-occurrence network analysis after Illumina MiSeq sequencing of the 13C-enriched 16S rRNA gene to directly relate the response in methanotrophic activity to the network structure of the interactome after desiccation-rewetting of a paddy soil. Methane uptake rate decreased immediately (<5 days) after short-term desiccation-rewetting. Although the methanotroph subgroups differentially responded to desiccation-rewetting, the metabolically active bacterial community composition, including the methanotrophs, recovered after the disturbance. However, the interaction network was profoundly altered, becoming more complex but, less modular after desiccation-rewetting, despite the recovery in the methanotrophic activity and community composition/abundances. This suggests that the legacy of the disturbance persists in the interaction network. The change in the network structure may have consequences for community functioning with recurring desiccation-rewetting. • SIP-network analysis provided a direct link between interacting community members. • Methanotrophic activity and community were resilient to desiccation-rewetting. • Desiccation-rewetting structures the active interaction network. • Interaction network became more complex but, less modular after disturbance. • Legacy of disturbance persisted in the interaction network. [ABSTRACT FROM AUTHOR]