Green manuring combined with nitrogen reduction sustained rice productivity and expanded soil organic nitrogen pool...

Green manuring can improve soil fertility and sustain rice productivity, yet its effects on rhizosphere metabolite-microbiome interactions regulating soil nitrogen pools remain unclear. Based on a six-year field experiment in the green manure-rice rotation system, we investigated how green manure combined with reduced nitrogen fertilization regulated soil nitrogen pool, enzyme activities, rhizosphere metabolites and microbial communities along the soil-root continuum. Green manuring combined with 20% reduction of nitrogen fertilizer (GM2) increased rice yield by approximately 8.7% and effectively expanded soil organic nitrogen pool. In contrast, green manuring without nitrogen reduction (GM3) enhanced inorganic nitrogen accumulation but accelerated organic nitrogen depletion. GM2 promoted the accumulation of soil amino sugar nitrogen, amino acid nitrogen and the activities of organic nitrogen-hydrolyzing enzymes (protease, leucine aminopeptidase and β-N-acetylglucosaminidase), indicating accelerated organic nitrogen depolymerization and turnover. Green manuring (GM) exerts an increasing influence on microbial community assembly along the soil-root continuum and promotes nitrogen-cycling functional potentials, most notably in the root endosphere of GM3. GM treatments enriched amino acids and signaling-related metabolites of rice rhizosphere soil, including L-isoleucine, sarcosine, 3-[(2-chloro-1,3-thiazol-5-yl) methyl]-5-methyl-1,3,5-oxadiazin-4-imine, and N-sulfo-D-glucosamine, all of which showed significant positive associations with nitrogen-cycling related bacterial taxa and saprotrophic fungi. Path modeling further demonstrated that metabolite-driven bacterial pathways dominated soil organic nitrogen accumulation. Overall, moderate nitrogen reduction under green manuring promoted coordinated rhizosphere metabolite-microbiome interactions that simultaneously sustained rice productivity and improved soil nitrogen stability.