Responses of Soil Carbon and Nitrogen Cycling Gene Abundances to Winter Green Manure Incorporation in a Ratoon Rice System

Winter green manure incorporation is widely practised in southern China, yet its effect on soil carbon and nitrogen cycling gene abundances across a full ratoon rice cycle remains unclear. A field experiment combined three cropping systems differing in winter management (winter fallow–ratoon rice, FA; rapeseed–ratoon rice, RA; and Chinese milk vetch–ratoon rice, MV) with two ratoon rice varieties (Yliangyou 911 and Liangyou 6326). Soil properties and functional gene abundances were measured at three soil depths and three sampling stages, and theoretical grain yield was determined in the main and ratoon seasons. Green manure incorporation did not change the bulk physicochemical properties of the soil as a general effect but altered its labile carbon and nitrogen pools. Dissolved organic carbon under RA and MV was, respectively, 24.28% and 16.15% higher than under FA, and ammonium nitrogen under MV in the ratoon season was up to 193.04% higher than under FA, exceeding RA throughout the profile. The functional genes responded in contrasting directions. pmoA, mcrA, nirK and nosZ were lower under green manure (pmoA 20.00% and 14.53% lower under RA and MV), whereas AOA amoA, cnorB and nirS were higher (AOA amoA 0.42 and 0.55 log units higher), with the clearest shifts occurring in the ratoon season and the surface soil. Correlation, redundancy and path analyses indicated that the gene–environment associations were linked mainly to ammonium nitrogen, dissolved organic carbon and nitrate nitrogen, and that winter treatment was associated with yield indirectly, most closely through the soil carbon pool (standardized total effect 0.34–0.57), rather than acting on yield directly.