Azolla  returning application Drives the Reshaping of Nitrogen-Fixing Microbial Communities in Paddy Soils to Enhance Fertilization and Reduce Emissions

Aims Azolla , a robust nitrogen (N)-fixing plant, can be efficiently and ecologically cultivated when incorporated into rice fields, while the mechanisms remain inadequately understood. This study systematically explored these mechanisms by assessing the effects of Azolla returning on soil nitrogen forms, ammonia volatilization, and microbial community responses across key rice growth stages. Methods The effects of Azolla incorporation were compared against rice monoculture and urea application at various stages of rice cultivation. The study monitored changes in soil nitrogen forms and ammonia volatilization. Furthermore, the response of the soil microbial community was examined, with a detailed analysis conducted during the grain-filling stage. Results Azolla returning significantly increased the content of total soil N, organic matter, available potassium and available phosphorus, while also elevating the soil pH. It substantially elevated ammonium nitrogen and microbial biomass nitrogen (peaking at grain-filling) compared to monoculture or urea, while significantly reducing nitrous oxide emissions, ammonia volatilization and maintaining nitrogen fertilizer content over time. Soil microbial community during the grain-filling stage indicated that urea fertilizer favored the enrichment of nitrifying bacteria ( Nitrospira and Nitrosomonas ) but also increased the abundance of pathogen Pseudomonas syringae . Azolla returning potentially reduced the abundance of pathogens while significantly promoting that of beneficial bacteria involved in N fixation and denitrification, including Azospirillum and Bacillus bataviensis . These enhanced soil nitrogen fixation and metabolic capacity. Conclusions This study is the first to track the dynamic characteristics and potential mechanisms behind the changes in nitrogen source during Azolla incorporation, a process that enhances soil fertility and reduces environmental emissions. It provides a theoretical foundation for the adoption of this sustainable practice.