Antimicrobial and Anti-inflammatory Activities of Bacteriocin-like Inhibitory Substance-producing Lactiplantibacillus plantarum against Vaginitis-associated Pathogens

Abstract Vaginal dysbiosis is characterized by disruption of the vaginal microbial community, pathogen overgrowth, and mucosal inflammation. Here, we integrated in vitro screening, whole-genome sequencing, LC-MS/MS profiling, machine learning, and a murine bacterial vaginosis (BV) model to characterize potentially beneficial Lactiplantibacillus plantarum strains. Among 88 Lactobacillaceae isolates, 53 exhibited bacteriocin-like inhibitory substance activity, and 7 demonstrated broad antimicrobial effects against Gardnerella vaginalis , Neisseria gonorrhoeae , Streptococcus agalactiae , and Candida spp. Two L. plantarum strains, CYLB2146 and CYLB2189, were selected based on their antimicrobial activity, adhesion capacity (15.0% and 13.5%, respectively), and significant anti-inflammatory effects, including reductions in IL-6, IL-8, and TNF-α levels ( p < 0.05). Genomic analysis revealed plantaricin biosynthetic gene clusters and secondary metabolite pathways, while LC-MS/MS identified predominantly low-molecular-weight and cyclic peptides. Machine-learning models achieved ROC-AUC > 0.92 and prioritized candidate peptides. Cyclo(Tyr-Pro) inhibited multiple pathogens and reduced biofilm formation at sub-inhibitory concentrations without membrane disruption, supporting a potential anti-virulence role. In a murine BV model, probiotic treatment markedly reduced G. vaginalis compared with PBS and MRS controls (51.2 and 41.6-fold). After 5 days, levels decreased to 3.2 and 11.1-fold in live CYLB2146 and CYLB2189 groups and to 3.2 and 0.7-fold in CFS groups, with further reductions by day 10, while controls remained high (34.0 and 50.6-fold). Probiotic treatment also attenuated infection-associated inflammation, particularly TNF-α, and improved vaginal tissue morphology and histological scores. Collectively, CYLB2146 and CYLB2189 exhibited complementary antimicrobial, antibiofilm, and anti-inflammatory activities, supporting their potential as probiotic candidates for BV management.