The present study aimed to characterize Enterococcus faecalis (n = -6) and Enterococcus faecium (n = 1) isolated from healthy chickens to find a novel perspective probiotic candidate that antagonize Clostridium botulinum types A, B, D, and E. The isolated enterococci were characterized based on phenotypic properties, PCR, and matrix-assisted laser desorption/ionization time of flight (MALDI-TOF). The virulence determinants including hemolytic activity on blood agar, gelatinase activity, sensitivity to vancomycin, and presence of gelatinase (gelE) and enterococcal surface protein (esp) virulence genes were investigated. Also, the presence of enterocin structural genes enterocin A, enterocin B, enterocin P, enterocin L50A/B, bacteriocin 31, enterocin AS48, enterocin 1071A/1071B, and enterocin 96 were assessed using PCR. Lastly, the antagonistic effect of the selected Enterococcus spp. on the growth of C. botulinum types A, B, D, and E was studied. The obtained results showed that four out of six E. faecalis and one E. faecium proved to be free from the tested virulence markers. All tested enterococci strains exhibited more than one of the tested enterocin. Interestingly, E. faecalis and E. faecium significantly restrained the growth of C. botulinum types A, B, D, and E. In conclusion, although, the data presented showed that bacteriocinogenic Enterococcus strains lacking of virulence determinants could be potentially used as a probiotic candidate against C. botulinum in vitro; however, further investigations are still urgently required to verify the beneficial effects of the tested Enterococcus spp. in vivo.
Keywords: Clostridium botulinum; Enterocin; Enterococcus; MALDI-TOF; PCR; Probiotics; Virulence markers.