Annual legume–grass overseeding and grassland management affect POC–MAOC partitioning in subtropical cultivated grasslands

Grassland carbon sequestration plays a critical role in regulating terrestrial carbon cycling and mitigating climate change. Annual forage overseeding is increasingly used to improve soil fertility and enhance soil organic carbon (SOC) storage in managed grasslands; however, how this management practice affects the formation and stabilization of particulate organic carbon (POC) and mineral-associated organic carbon (MAOC) under contrasting grassland management regimes remains poorly understood. We conducted a field experiment from 2023 to 2025 in a perennial cultivated grassland in southern China evaluate the effects of annual grass, annual legume, and legume–grass mixture overseeding under grazing and exclosure. We measured belowground biomass, root C and N pools, SOC, total nitrogen (TN), microbial biomass carbon and nitrogen (MBC and MBN), POC and MAOC. Our results showed that, compared with the control, SOC, MBC, and MBN increased significantly across all overseeding treatments, whereas TN increased significantly under annual legume and legume–grass mixture overseeding. Annual legume–grass mixture overseeding increased belowground biomass by approximately 15% compared with annual legume or annual grass overseeding alone. Grassland management altered the relative responses of SOC fractions to overseeding, with POC increasing by approximately 36% under grazing and MAOC by approximately 18% under exclosure. Redundancy analysis showed that root C and N pools, MBC, and TN were closely associated with variation in POC and MAOC. Random forest analysis further identified root C and N pools as important predictors of POC under both management regimes, whereas MAOC was more strongly associated with microbial biomass under exclosure and with root C pool and soil C: N stoichiometry under grazing. Overall, annual forage overseeding increased belowground C and N inputs, microbial biomass, and SOC content, with the legume–grass mixture showing the strongest overall response. The contrasting responses of POC and MAOC demonstrate that grassland management plays an important role in regulating overseeding-induced SOC accumulation and partitioning. Understanding how annual forage overseeding interacts with grassland management to regulate SOC accumulation and its partitioning between POC and MAOC provides new insights for developing grassland management strategies that enhance soil carbon storage while sustaining forage productivity.