The sigma factor 54(σ~(54)) controls the expression of many genes in response to nutritional and environmental conditions. There are two σ~(54) genes, rpo N1(XAC1969) and rpo N2(XAC2972), in Xanthomonas citri subsp. citri. To investigate their functions, the deletion mutants ΔrpoN1, ΔrpoN2 and ΔrpoN1N2 were constructed in this study. All the mutants delayed canker development in low concentration inoculation in citrus plants. The bacterial growth of mutants was retarded in the medium supplemented with nitrogen and carbon resources. Under either condition, the influence degree caused by deletion of rpoN 2 was larger than the deletion of rpoN 1. Remarkably, the mutant ΔrpoN 1 showed a reduction in cell motility, while the mutant Δrpo N2 increased cell motility. Our data suggested that the rpoN 1 and rpoN 2 play diverse roles in X. citri subsp. citri. There are two σ ~ (54) genes, rpo N1 (XAC1969) and rpo N2 (XAC2972), in Xanthomonas citri All the mutants delayed canker development in low concentration inoculation in citrus plants. The bacterial growth of mutants was retarded in the medium supplemented with nitrogen and the carbon resources. Under either condition, the influence degree caused by deletion of rpoN 2 was larger than the deletion of rpoN 1. Remarkably, the mutant ΔrpoN 1 showed a reduction in cell motility, while the mutant Δrpo N2 increased cell motility. Our data suggested that the rpoN 1 and rpoN 2 play diverse roles in X. citri subsp. citri.