Historical tillage promotes grass-legume mixtures establishment and accelerates soil microbial activity and organic carbon decomposition

Perennial grass-legume mixtures have been extensively used to restore degraded grasslands, increasing grassland productivity and forage quality. Tillage is crucial for seedbed preparation and sustainable weed management for the establishment of grass-legume mixtures. However, a common concern is that intensive tillage may alter soil characteristics, leading to losses in soil organic carbon (SOC). We investigated the plant community composition, SOC, soil microbial biomass carbon (MBC), soil enzyme activities, and soil properties in long-term perennial grass-legume mixtures under two different tillage intensities (once and twice) as well as in a fenced grassland (FG). The establishment of grass-legume mixtures increased plant species diversity and plant community coverage, compared with FG. Compared with once tilled grassland (OTG), twice tilled grassland (TTG) enhanced the coverage of high-quality leguminous forage species by 380.3%. Grass-legume mixtures with historical tillage decreased SOC and dissolved organic carbon (DOC) concentrations, whereas soil MBC concentrations in OTG and TTG increased by 16.0% and 16.4%, respectively, compared with FG. TTG significantly decreased the activity of N-acetyl-β-D-glucosaminidase (NAG) by 72.3%, whereas soil enzyme β-glucosidase (βG) in OTG and TTG increased by 55.9% and 27.3%, respectively, compared with FG. Correlation analysis indicated a close association of the increase in MBC and βG activities with the rapid decline in SOC. This result suggested that MBC was a key driving factor in soil carbon storage dynamics, potentially accelerating soil carbon cycling and facilitating biogeochemical cycling. The establishment of grass-legume mixtures effectively improves forage quality and boosts plant diversity, thereby facilitating the restoration of degraded grasslands. Although tillage assists in establishing legume-grass mixtures by controlling weeds, it accelerates microbial activity and organic carbon decomposition. Our findings provide a foundation for understanding the process and effectiveness of restoration management in degraded grasslands.