Evaluation of Limosilactobacillus reuteri ATCC PTA 6127 Reveals Multilayered Protection Against Enteric Pathogens in a Canine Epithelial Model

Effective canine gastrointestinal health depends on suppression of enteric pathogens and maintenance of epithelial barrier integrity. Limosilactobacillus reuteri ATCC PTA 6127 (Lr6127) is a dog‑derived probiotic, but evidence supporting its protective capacity remains limited. Here, we evaluated the antimicrobial and epithelial‑supportive effects of Lr6127 using a canine epithelial cell model. Cell‑free supernatant (CFS) from Lr6127 significantly inhibited the growth of canine‑relevant pathogens, including Escherichia coli (52.0 ± 1.3%), Clostridium perfringens (54.0 ± 2.7%), and Salmonella typhimurium (48.6 ± 1.2%), compared with the medium control (P <  0.0001). Pathogen inhibition increased in a dose‑dependent manner with increasing CFS concentrations. Untargeted metabolomic analysis revealed enrichment of multiple antimicrobial‑associated metabolites, indicating multi‑component pathogen suppression, with genomic analysis supporting the aromatic amino acid–derived metabolite findings. In addition, viable Lr6127 significantly reduced epithelial adhesion of all tested pathogens (P <  0.01). Beyond direct antimicrobial effects, Lr6127 CFS promoted epithelial wound healing at later time points, accompanied by coordinated modulation of proteins associated with cytoskeletal remodeling and barrier repair. Collectively, these findings suggest that Lr6127 is associated with pathogen suppression, reduced adhesion, and epithelial wound repair mechanisms.