Abstract:[Background] Previous transcriptomics studies suggested that the RNA polymerase sigma factor (sigW) and the anti-sigma factor W (rsiW) might be involved in the manganese stress response of Bacillus safensis. [Objective] To investigate the effects of sigW and rsiW deletion on the motility, biofilm formation, and manganese-oxidizing ability of B.safensis ST7. [Methods] We employed homologous recombination to construct the sigW- and rsiW-deleted mutant strains of B.safensis ST7. Furthermore, we compared the motility, biofilm formation, manganese oxidation, and expression of flagellum formation-related genes between the wild type and the mutant strains. [Results] The deletion of sigW and rsiW had no significant effect on the growth of ST7 cultured without manganese stress, whereas it retarded the growth of ST7 under manganese stress. Compared with the wild type, ∆sigW showed reduced motility and manganese-oxidizing ability and enhanced biofilm formation, while ∆rsiW showed reduced motility and enhanced biofilm formation and manganese-oxidizing ability. In addition, the expression levels of genes (flim, flgd and flii) related to flagellum formation were downregulated in ∆sigW and ∆rsiW. [Conclusion] sigW and rsiW can promote the flagellum formation, motility, and growth and co-regulate the manganese-oxidizing ability of B.safensis.