Layered nitrogen application drives changes in nitrogen nutrition status of wheat by affecting key soil microbial clusters

Abstract

Optimizing nitrogen (N) fertilizer management is essential for improving wheat yield and nitrogen use efficiency, yet the mechanisms by which layered N application regulates crop N nutrition through soil microbial processes across growth stages remain unclear. Here, we conducted a four-year field experiment to examine how layered N fertilization modulates soil microbial communities and nutrient dynamics to better match wheat N demand and enhance productivity. Treatments included conventional N application at 8 cm soil depth (N8), layered N application at 8, 16, and 24 cm in a 1:2:1 ratio (N1–2–1), and a no-N control (NCK). Layered N fertilization increased grain yield by 18.7 % and total N accumulation by 19.0 % compared with conventional N application, while maintaining an optimal nitrogen nutrition index throughout wheat development. These yield gains were associated with a marked increase in soil available phosphorus (AP), which enhanced bacterial diversity (Shannon index) and richness (Chao1 index). Co-occurrence network analysis identified two key microbial modules (Module 1 and Module 5) that strongly predicted wheat N accumulation and yield (p < 0.01). The functional roles of these modules shifted from saprotrophic and nitrifying processes at the regreening stage to aromatic compound degradation and root symbiosis at anthesis. Dominant taxa within these modules, particularly Stanjemonium and Coniochaeta, were the strongest contributors to wheat N concentration at the regreening and anthesis stages, respectively. Random forest analysis further indicated that AP exerted a direct regulatory effect on microbial module abundance, while soil N availability influenced wheat N nutrition indirectly through its interactions with AP and microbial network structure. Structural equation modeling confirmed that these pathways ultimately determined wheat nitrogen nutritional status and yield. Overall, layered N fertilization enhances wheat N uptake and productivity by reshaping soil microbial network organization through AP-mediated mechanisms, highlighting the importance of microbial ecological clusters in synchronizing crop N demand with nutrient supply across growth stages.