In this study, the pure culture method combined with metagenomic analysis was used to investigate the evolutionary relationship of 66 strains of Bacillus isolated from Pixian broad bean paste. The enzyme activities and antibiotic resistance of the selected representative strains were further analyzed. Totally 24 Bacillus strains were identified by metagenomics and were classified into 13 species according to full-length 16S rDNA gene sequences. As the salt content rose, the protease-producing and amylase-producing abilities of the 29 representative strains decreased. All the strains were susceptible to neomycin, but their sensitivity to 11 other antibiotics tested was largely different. Among the 12 antibiotics, the incidence of ampicillin resistance was the highest (66%). Our findings motivate the future utilization of the Bacillus strains from Pixian broad bean paste.