Long-term cultivation of grass–legume mixtures changed the assembly process of the microbial community and increased microbial community stability

Grass-legume mixtures are a common cultivation system on the Qinghai-Tibet Plateau, where the interactions between rhizosphere microorganisms and crops under long-term cultivation are complex and dynamic. Investigating the dynamic changes in microbial community structure and ecological functions is essential. This study investigated the dynamic interactions of rhizosphere microbial communities of <i>Elymus nutans</i> Griseb. cv. Aba and <i>Medicago sativa</i> L. cv. Beilin in a grass-legume mixture at a 1:1 ratio >4 years on the Qinghai-Tibet Plateau. The research focused on their long-term effects on plant productivity, soil health, and microbial functions. The results revealed a decline in grass yield and soil properties in the fourth year (<i>P</i> < .05) and significant year-to-year differences in bacterial α-diversity (<i>P</i> < .05). Molecular ecological network analysis showed greater stability in the bacterial network of legumes in the first year, with reduced robustness by the fourth year. Additionally, the average niche widths of bacterial and fungal communities were narrower in the first year than in the fourth, indicating microbial adaptation to the evolving environmental conditions within the mixture system. The transition of bacterial community assembly processes from stochastic to deterministic suggests a shift toward more structured and predictable microbial interactions over time. In conclusion, the results highlight the intricate interplay between plant productivity, soil health, microbial community dynamics, and ecosystem stability under long-term planting of grass-legume mixtures. Our results provide new insights into biomass changes and microbial dynamics in this planting system.