Delayed flooding after green manure incorporation decreases methane emissions and greenhouse gas intensity in rice paddy fields

Greenhouse gas (GHG) emissions from paddy fields are strongly influenced by practices such as green manure incorporation and water management. In this study, we examined the effects of incorporating Chinese milk vetch (Astragalus sinicus L.) and applying delayed flooding on methane (CH4) and nitrous oxide (N2O) emissions, global warming potential (GWP), greenhouse gas intensity (GHGI), soil and surface water properties, and methane-cycling microbial communities. Our approach combined a two-year field experiment in southern China with a complementary microcosm incubation. Green manure incorporation significantly increased CH4 emissions and boosted rice yield, whereas delayed flooding substantially reduced CH4 emissions without affecting N2O emissions. Among the tested treatments, withholding flooding for 10 days (10DbF) produced the lowest CH4 emissions and GWP, reducing GHGI by 51.6 % in 2022 and 36.1 % in 2023. CH4 emissions were positively correlated with soil total nitrogen (STN) and water-dissolved organic carbon (WDOC), indicating their key regulatory roles. Microcosm experiments further showed that 10DbF postponed the CH4 emission peak and reduced cumulative emissions by 405.1 % compared to immediate flooding (0DbF). Quantitative PCR analysis revealed a significant decline in mcrA gene abundance and in the mcrA/pmoA ratio under 10DbF, while pmoA gene abundance remained largely unaffected. Microbial community profiling indicated clear shifts in methanogenic and methanotrophic assemblages, with delayed flooding decreasing the relative abundances of amplicon sequence variants (ASVs) within key methanogenic taxa (Methanoregula and Methanobacterium) and enriching Methylomonas. These findings demonstrate that combining green manure incorporation with optimized water management can effectively mitigate CH4 emissions while sustaining rice productivity, offering a promising strategy for climate-resilient and sustainable rice farming.