Phenotypic and Genomic Analysis of Swine-Derived Bacillus subtilis 1BP1Co1 as a Functional Probiotic Candidate to Mitigate Escherichia coli-Associated Diarrhea

Post-weaning diarrhea caused by pathogenic Escherichia coli is a significant economic challenge in global swine production. Rising antimicrobial resistance linked to traditional antibiotics necessitates safe, sustainable alternatives. This study isolated and characterized host-associated Bacillus species from the gastrointestinal tracts of healthy pigs to evaluate their probiotic potential. Fifty-nine Bacillus isolates were screened for safety, gastrointestinal tract survivability, mucosal adhesion, and antimicrobial efficacy. Several isolates demonstrated robust tolerance to simulated gastrointestinal stress (acidic pH and bile salts), non-hemolytic profiles, and strong mucosal adhesion. Multiple isolates displayed inhibitory activity against pathogenic Escherichia coli in vitro. The most promising candidate, Bacillus subtilis 1BP1Co1, underwent Nanopore whole-genome sequencing. Genomic profiling confirmed its safety, revealing a complete absence of known virulence factors and high-risk antimicrobial resistance genes. Furthermore, genome annotation identified multiple biosynthetic gene clusters responsible for the production of antimicrobial secondary metabolites, including surfactin, fengycin, bacillaene, and subtilosin A, which mechanistically supports its pathogen-inhibitory properties. Collectively, these findings highlight indigenous Bacillus strains, particularly Bacillus subtilis 1BP1Co1, as promising probiotic candidates with the potential to be developed to mitigate Escherichia coli-associated enteric disorders in swine production systems.