Genomic and Phenotypic Characterization of Antibiotic-Resistant Probiotic Strains from Entromax^®

This study evaluated the probiotic properties of poly-antibiotic-resistant Bacillus clausii derived from Entromax®, which was subjected to comprehensive phenotypic, biochemical, and molecular characterization, including antibiotic susceptibility testing. Key probiotic attributes, such as tolerance to gastric juice, bile salts, and osmotic stresses, were assessed alongside enzymatic activities like casein and starch hydrolysis. The Total Viable Count (TVC) of the tested Entromax® Bacillus clausii batches was determined to be 0.42 ± 0.2 billion CFU/mL, corresponding to approximately 2 billion CFU per 5 mL dose. Antibiotic resistance profiles revealed that the bacterial culture isolated from Entromax® exhibited resistance to various classes of antibiotics like phenicols, fluoroquinolones, macrolides, aminoglycosides, beta-lactams, and tetracyclines. 16S rRNA sequencing indicated a close relationship with Shouchella clausii, formerly known as Bacillus clausii, while whole-genome BLAST analysis confirmed 100% similarity with Bacillus clausii ENTPro. The observed amylolytic and proteolytic activities, resistance to acidic conditions, bile salts, and gastric juice highlight critical probiotic characteristics, supporting the potential of these strains for probiotic applications.