Response of nitrite-dependent anaerobic methanotrophs to elevated atmospheric CO₂ concentration in paddy fields

Nitrite-dependent anaerobic methane oxidation (n-damo) catalyzed by Candidatus Methylomirabilis oxyfera (M. oxyfera)-like bacteria is a new pathway for the regulation of methane emissions from paddy fields. Elevated atmospheric CO₂ concentrations (e[CO₂]) can indirectly affect the structure and function of microbial communities. However, the response of M. oxyfera-like bacteria to e[CO₂] is currently unknown. Here, we investigated the effect of e[CO₂] (ambient CO₂ + 200 ppm) on community composition, abundance, and activity of M. oxyfera-like bacteria at different depths (0-5, 5-10, and 10-20 cm) in paddy fields across multiple rice growth stages (tillering, jointing, and flowering). High-throughput sequencing showed that e[CO₂] had no significant effect on the community composition of M. oxyfera-like bacteria. However, quantitative PCR suggested that the 16S rRNA gene abundance of M. oxyfera-like bacteria increased significantly in soil under e[CO₂], particularly at the tillering stage. Furthermore, ¹³CH₄ tracer experiments showed potential n-damo activity of 0.31-8.91 nmol CO₂ g^-1 (dry soil) d^-1. E[CO₂] significantly stimulated n-damo activity, especially at the jointing and flowering stages. The n-damo activity and abundance of M. oxyfera-like bacteria increased by an average of 90.9% and 50.0%, respectively, under e[CO₂]. Correlation analysis showed that the increase in soil dissolved organic carbon content caused by e[CO₂] had significant effects on the activity and abundance of M. oxyfera-like bacteria. Overall, this study provides the first evidence for a positive response of M. oxyfera-like bacteria to e[CO₂], which may help reduce methane emissions from paddy fields under future climate change conditions.