Stratified Effects of Tillage and Crop Rotations on Soil Microbes in Carbon and Nitrogen Cycles at Different Soil Depths in Long-Term Corn, Soybean, and Wheat Cultivation

Understanding the soil bacterial communities involved in carbon (C) and nitrogen (N) cycling can inform beneficial tillage and crop rotation practices for sustainability and crop production. This study evaluated soil bacterial diversity, compositional structure, and functions associated with C-N cycling at two soil depths (0–15 cm and 15–30 cm) under long-term tillage (conventional tillage [CT] and no-till [NT]) and crop rotation (monocultures of corn, soybean, and wheat and corn–soybean–wheat rotation) systems. The soil microbial communities were characterized by metabarcoding the 16S rRNA gene V4–V5 regions using Illumina MiSeq. The results showed that long-term NT reduced the soil bacterial diversity at 15–30 cm compared to CT, while no significant differences were found at 0–15 cm. The bacterial communities differed significantly at the two soil depths under NT but not under CT. Notably, over 70% of the tillage-responding KEGG orthologs (KOs) associated with C fixation (primarily in the reductive citric acid cycle) were more abundant under NT than under CT at both depths. The tillage practices significantly affected bacteria involved in biological nitrogen (N₂) fixation at the 0–15 cm soil depth, as well as bacteria involved in denitrification at both soil depths. The crop type and rotation regimes had limited effects on bacterial diversity and structure but significantly affected specific C-N-cycling genes. For instance, three KOs associated with the Calvin–Benson cycle for C fixation and four KOs related to various N-cycling processes were more abundant in the soil of wheat than in that of corn or soybean. These findings indicate that the long-term tillage practices had a greater influence than crop rotation on the soil bacterial communities, particularly in the C- and N-cycling processes. Integrated management practices that consider the combined effects of tillage, crop rotation, and crop types on soil bacterial functional groups are essential for sustainable agriculture.