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In this study, from 235 vaginal swabs, 45 GBS isolates were identified and screened; 12 representative isolates (strong biofilm producers with the complete virulence gene profile) were selected for downstream analyses. Molecular analysis showed high prevalence of virulence genes (fbsA 95.5%, lmb 91.1%, pavA 88.8%, fbsB 86.6%) and biofilm-related genes (pil-1 88.8%, pil-2a 91.1%, pil-2b 84.4%). Subsequently, a Bifidobacterium bifidum (designated B. bifidum BB-6; GenBank accession number PX474696) isolated from human breast milk was used to prepare a cell-free supernatant (CFS). Gas chromatography\u2013mass spectrometry analysis (GC\u2013MC) of the CFS of B. bifidum identified several bioactive compounds, including acetic acid, propionic acid, and lactic acid. Checkerboard assays indicated synergism between CFS and penicillin\/vancomycin, with a fractional inhibitory concentration index (FICI) of\u00a0\u2264\u00a00.5 in most cases. Sub-minimum inhibitory concentration (sub-MIC) of CFS significantly inhibited biofilm formation (P\u00a0&amp;lt;\u00a00.01), and quantitative real-time PCR (qRT-PCR) revealed downregulation of virulence (fbsB down \u22124.40-fold) and biofilm genes (pil-2b down \u22125.49-fold). These results highlight the therapeutic potential of B. bifidum CFS against GBS, warranting further studies to isolate active compounds and evaluate safety and efficacy in vivo.<\/jats:p>","DOI":"10.1093\/lambio\/ovaf133","type":"journal-article","created":{"date-parts":[[2025,11,29]],"date-time":"2025-11-29T12:28:33Z","timestamp":1764419313000},"source":"Crossref","is-referenced-by-count":0,"title":["Inhibitory effects of cell-free supernatant of\n                    <i>Bifidobacterium bifidum<\/i>\n                    on biofilm formation and virulence gene expression in Group B\n                    <i>Streptococcus<\/i>\n                    clinical isolates"],"prefix":"10.1093","volume":"78","author":[{"given":"Uosef","family":"Ramezani","sequence":"first","affiliation":[{"name":"Ar.C., Islamic Azad University Department of Microbiology, , Arak 3836119131 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