This study was carried out to investigate effects of subsoiling on the diversity and composition of the bacterial community in a wheat–maize rotation field in the Guanzhong area of Shaanxi Province, China. After the wheat harvest, surface soil samples were collected under two tillage methods (single rotary tillage (RT) and subsoiling + rotary tillage (ST)) to perform high-throughput sequencing and bioinformatics analysis. Soil properties and root length density (RLD) of winter wheat at booting and flowering stages were also studied. Results showed that ST treatment significantly raised the water storage, organic carbon and total nitrogen contents of deep soil (>40 cm), and notably increased the total soil pH, ammonium nitrogen content and RLD in the tillage layer from 0–70 cm at booting stage and 0~100 cm at flowering stage, but the residual nitrate nitrogen significantly decreased by 17.74%. Compared with RT, soil bacterial richness and diversity in the 10~20 cm layer of ST treatment showed a significantly decreased trend. The relative abundances of GAL15, Actinobacteria, Nitrospirae, Rhizobiales, Burkholderiales, Pseudomonas and Serratia in the 10–20 cm layer were remarkably increased in ST. Principal Component Analysis (PCA) and Redundancy Analysis (RDA) results showed that surface soil pH, ammonium nitrogen and nitrate nitrogen contents have the strongest effect on the bacterial structure. In addition, there were positive correlations between the RLD and the relative abundances of Rhizobiales, Burkholderiales, Pseudomonas and the ammonium nitrogen content. In conclusion, although subsoiling was not conducive to improving soil bacterial community richness and diversity, it significantly increased soil beneficial bacteria (biological nitrogen-fixing bacteria, ammonifying bacteria, nitrobacteria) abundances, reduced the nitrogen loss caused by denitrifying bacteria, promoted earlier root development and improved the plant utilization ratio of soil nutrients.

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