Diversified crop rotations exert a profound influence on the nosZI denitrifier community

Agricultural soils are a significant contributor to global nitrous oxide (N₂O) emissions, which is primarily driven by microbial nitrification and denitrification processes. Diversifying crop rotations can enhance soil nitrogen (N) utilization and influence N-cycling microbes, particularly the denitrifiers. Here, we evaluated the abundance, diversity, and community structure of soil denitrifiers by analyzing the denitrification genes (nirS, nirK, and nosZI) with a 14-year experiment of continuous and rotated crop systems. Compared to continuous cotton (C), crop rotations altered abundance of the nirS, nirK, and nosZI genes, but the responses varied among different rotation patterns (p < 0.05). Diversified crop rotations decreased Shannon index of the nirS and nirK gene communities, while increased that of the nosZI gene communities. Diversified crop rotations greatly changed community structure of the nirS, nirK and nosZI gene communities (p < 0.05). Among the responsive operational taxonomic units (OTUs), 2.62 %, 2.52 %, and 4.31 % of the nirS, nirK, and nosZI genes were involved in the responses, respectively. Notably, a significant increase in OTUs belonged to N-fixing groups (i.e., Herbaspirillum and Rhodanobacter), while a decrease in OTUs affiliated to Achromobacter and Bosea was observed in diversified crop rotations. Furthermore, soil pH and C/N ratio correlated significantly with the nirS gene community structure (p < 0.05), SOC and C/N ratio correlated with the nirK gene community (p < 0.05), and soil pH, TN, NO₃^--N, and C/N ratio correlated with the nosZI gene community (p < 0.05), respectively. In conclusion, our study demonstrates that long-term diversified crop rotations significantly altered the microbial communities related to denitrification, especially the nosZI gene community. These findings highlight the importance of diversified crop rotations in fostering soil denitrifiers communities that contribute to N₂O reduction, potentially supporting sustainable agricultural practices by reducing greenhouse gas emissions. However, future studies should be focus on both the nosZI and nosZII clades communities to offer a more complete picture of microbial contributions to soil N-cycling and N₂O emission reduction.