Biological soil crusts and slope gradient synergistically regulate the community structure and functional differentiation of nitrogen-cycling microorganisms in alpine meadow soils

<title>Abstract</title> <p> Background and aims Biological soil crusts (BSCs) and slope gradient regulate soil nitrogen cycling in alpine meadows, but their combined effects on nitrogen-cycling microbial communities remain unclear. This study investigated how BSCs and slope jointly influence soil nitrogen transformations and the community structure of nitrogen-cycling microorganisms. Methods Soils with and without BSC cover were collected from flat, gentle, and steep slopes in an alpine meadow on the Qinghai-Tibetan Plateau. Soil nitrogen nutrients were measured, and high-throughput sequencing was used to characterize the relative abundances, diversity, and co-occurrence networks of five functional genes (archaeal <italic>amoA</italic> , bacterial <italic>amoA</italic> , <italic>nirK</italic> , <italic>nirS</italic> , and <italic>nosZ</italic> ). Results BSCs significantly promoted nitrification, increasing nitrate nitrogen by 9.1%-56.8% and decreasing ammonium nitrogen by 71.0%-87.5%, with nitrate concentrations further increasing along slope gradient. Microbial groups responded differently to BSCs and slope. BSCs increased archaeal <italic>amoA</italic> and <italic>nirK</italic> abundances but reduced α diversity of AOB, <italic>nirK</italic> , and <italic>nosZ</italic> communities. Slope gradient primarily regulated AOA, <italic>nirK</italic> , and <italic>nosZ</italic> diversity, while their interaction reshaped community structure. Co-occurrence network analysis revealed that AOA communities maintained stable structures through core Thaumarchaeota, whereas AOB and denitrifier communities adapted through network modular differentiation: gentle slopes harbored high-abundance functional modules, while steep slopes enriched stress-tolerant rare modules. Structural equation modeling confirmed that BSCs and slope indirectly regulated microbial diversity by altering soil organic matter and ammonium nitrogen. Conclusions This study elucidates the mechanisms by which BSCs and slope gradient jointly regulate nitrogen-cycling microorganisms in alpine meadow soils, providing evidence for nutrient cycling maintenance and ecological conservation. </p>