Antimicrobial resistance, virulence characteristics and genotypes of Bacillus spp. from probiotic products of diverse origins

Spore-forming probiotic Bacillus spp. have received extensively increasing scientific and commercial interest, but raised the concerns in the potential risks and pathogenesis. In this study, 50 commercial probiotic products were collected from all over the country and Bacillus spp. isolated from products were evaluated for the safety on the aspects of hemolytic activity, contamination profiles, toxin genes, cytotoxicity, antimicrobial resistance, and genotyping. 34 probiotic products (68%) exhibited hemolysis, including 19 human probiotics, 9 animal probiotics, and 6 plant probiotics. 28 products (56%) contained other bacteria not labeled in the ingredients. 48 strains in Bacillus spp. including 17 B. subtilis group isolates, 28 B. cereus, and 3 other Bacillus spp. were isolated from human, food animal, and plant probiotic products. Detection rates of enterotoxin genes, nheABC and hblCDA, and cytotoxin cytK2 in 48 Bacillus spp. isolates were 58%, 31%, and 46%, respectively. Also, one isolate B. cereus 34b from an animal probiotic product was positive for ces, encoding cereulide. 28 of 48 Bacillus spp. isolates were cytotoxic. 19 of 28 B. cereus isolates maintained to exhibit hemolysis after heat treatment. All 48 Bacillus spp. isolates exhibited resistance to lincomycin, and 5 were resistant to tetracycline. The genotyping of commercial probiotic Bacillus spp. reported in this study showed that ces existed in B. cereus 34b with the specific sequence type (ST1066). These findings support the hypothesis that probiotic products were frequently contaminated and that some commercial probiotics consisted of Bacillus spp. may possess toxicity and antimicrobial resistance genes. Thus, the further efforts are needed in regarding the surveillance of virulence factors, toxins, and antibiotic resistance determinants in probiotic Bacillus spp.