Endpoint Metagenomic Evidence for Shifts in Bulk Soil Microbial Communities Under Long-Term Nitrogen Addition in a Cold-Temperate Coniferous Forest

Atmospheric nitrogen (N) deposition is an important global change driver in forest ecosystems, yet its long-term effects on belowground microbial communities in cold-temperate coniferous forests remain insufficiently understood. In this study, endpoint shotgun metagenomic sequencing was used to evaluate bulk soil microbial communities after 12 years of experimental N addition in a Larix gmelinii-dominated forest in the Greater Khingan Mountains of northeastern China. Four treatments were included: control (0 kg N ha−1 yr−1), low N (25 kg N ha−1 yr−1), medium N (50 kg N ha−1 yr−1), and high N (75 kg N ha−1 yr−1). Microbial alpha diversity did not differ significantly among treatments, although moderate N addition showed a tendency to maintain relatively higher richness and diversity. In contrast, beta-diversity analysis indicated clear shifts in community composition along the N addition gradient. Pseudomonadota, Acidobacteriota, and Actinomycetota dominated the microbial communities, with Pseudomonadota tending to increase under N enrichment, whereas some oligotrophic groups showed reduced relative abundance. Functional annotation showed that metabolism-related genes remained dominant across treatments, and carbohydrate-active enzyme profiles suggested altered microbial potential for complex carbon decomposition under long-term N input. Nitrogen addition also modified the abundance patterns of some antibiotic resistance genes and mobile genetic elements, although overall resistome differentiation among treatments remained limited. These results provide endpoint metagenomic evidence that long-term N addition can reshape bulk soil microbial community composition and selected functional potentials in cold-temperate coniferous forest soils, even when overall alpha diversity remains relatively stable.