Deep Placement of Nitrogen Fertilizer Mitigates Methane Emissions from Rice Paddies by Modulating Methanogenic and Methanotrophic Communities in a Rice–Wheat Rotation System

Deep placement of nitrogen fertilizer (DPN) is an effective fertilization strategy for improving nitrogen use efficiency in rice systems, but its effects on methane (CH4) emissions and the associated microbial mechanisms remain insufficiently understood. This study aimed to determine whether DPN mitigates CH4 emissions in a rice–wheat rotation system and to clarify how it regulates methanogenic and methanotrophic communities. A two-year field experiment was conducted in East China with two nitrogen management practices, i.e., conventional surface application and DPN. Compared with surface application, DPN significantly reduced cumulative CH4 emissions by 22.9% in 2023 and 17.0% in 2024, while tending to increase rice grain yield. During the tillering stage, DPN decreased soil dissolved organic carbon by 19.0% and increased NH4+-N and NO3−-N concentrations by 35.8% and 44.1%, respectively. These changes were accompanied by a 24.7% reduction in methanogen abundance and a 26.9% decrease in methanogenic activity. Although total methanotroph abundance was not significantly affected, DPN increased methanotrophic activity by 24.6%. Amplicon sequencing further showed that DPN shifted the methanogenic community from acetoclastic taxa toward hydrogenotrophic taxa, as indicated by the decline in Methanosarcina and Methanothrix and the enrichment of Methanobacterium and Methanoregula. In parallel, DPN promoted Type I methanotrophs, especially Methylomonas, while suppressing the Type II methanotroph Methylocystis. These results demonstrate that DPN mitigates CH4 emissions by reducing labile carbon availability, suppressing methanogenic abundance and activity, and enhancing the functional potential of methane oxidation through methanotrophic community restructuring. Overall, this study indicates that DPN mitigates CH4 emissions through coordinated regulation of carbon substrate availability and functional microbial community restructuring, suggesting that DPN is a promising strategy for sustainable rice production and greenhouse gas mitigation.