Microbial functional gene assembly is associated with soil carbon and nitrogen dynamics during grassland degradation.

Introduction: Grassland degradation is often accompanied by changes in the structure and function of soil microbial communities. However, the mechanisms by which the assembly of microbial functional communities is associated with alterations in soil carbon and nitrogen pools remain unclear. Methods: This study was conducted along a degradation gradient in a typical steppe of Inner Mongolia. Metagenomics, community null models, and structural equation modeling were used to examine microbial functional gene assembly, carbon and nitrogen cycling genes, and their associations with soil carbon and nitrogen pools. Results: The assembly of microbial functions shifted from being predominantly influenced by stochastic processes to deterministic processes, with the strongest deterministic filtering observed during the moderate degradation stage. The abundance of the aerobic oxidation gene porA decreased with increasing degradation, whereas fermentation genes, including ldh and atoB, increased significantly during moderate degradation. Denitrification genes, including narG, nirK, norB, and nosZ, reached their highest abundance during the heavy degradation stage. Mineral-associated organic carbon exhibited a nonlinear pattern characterized by an initial increase followed by a decrease. Structural equation modeling revealed that microbial biomass carbon was the central variable linking microbial functional differentiation with changes in soil carbon and nitrogen pools. During the heavy degradation stage, soil ammonium nitrogen showed a numerical increase, suggesting that nitrogen released from mineral-associated organic carbon decomposition may be predominantly converted into inorganic forms. Discussion: These findings indicate that the threshold-like decline of microbial biomass carbon, rather than specific restructuring of functional gene profiles, was closely associated with the collapse of stable carbon-nitrogen pool stability during grassland degradation. Changes in functional genes may therefore represent responsive signals accompanying microbial biomass carbon attenuation. The continuous decrease in microbial biomass carbon and associated shifts in functional gene ratios may serve as potential indicators of declining carbon-nitrogen stability in grassland soils.