Surfactin from Bacillus subtilis 1–4 disrupts the cell membrane of zoonotic staphylococcus intermedius in vitro and maintains gut microbiota community in urban pigeons

Urban pigeons frequently carry zoonotic, antibiotic-resistant Staphylococcus spp. and are increasingly recognized as reservoirs of multidrug-resistant (MDR) gut pathogens with One Health implications. Probiotic Bacillus strains and their lipopeptide products, such as surfactin, are promising antibiotic-sparing alternatives, but their in vivo effects on host gut microbiota and pathogen carriage in apparently healthy birds remain poorly characterized. To explore alternative antimicrobial strategies, Bacillus subtilis 1–4 was isolated from soil and shown to produce surfactin, a potent antimicrobial peptide. In vitro, surfactin exhibited bactericidal activity against S. aureus and S. intermedius by compromising membrane integrity. To evaluate its in vivo effects, pigeons (Columba livia) were treated with surfactin for seven days, and changes in their gut microbiota were characterized using full-length 16 S rRNA gene sequencing. While microbial α- and β-diversity remained largely unchanged, the surfactin-treated microbiota displayed enhanced compositional stability relative to controls. These findings support surfactin’s potential as a targeted antimicrobial agent with minimal disruption to the host gut microbiota.