Leuconostoc mesenteroides TH-ST373 Cell-Free Supernatant Against Multidrug-Resistant Escherichia coli

Multidrug resistance (MDR) poses a critical global health threat, rendering conventional antibiotics increasingly ineffective and driving an urgent need for innovative alternatives. This study unveils the potent antibacterial and biofilm-inhibitory activity of cell-free supernatant (CFS) derived from Leuconostoc mesenteroides subsp. jonggajibkimchii TH-ST373 against MDR Escherichia coli, including clinically relevant strains resistant to ampicillin, norfloxacin, cephalothin, and gentamicin. TH-ST373 CFS consistently outperformed the reference strain, achieving dose-dependent growth inhibition with minimum inhibitory concentrations (MICs) of 0.63-1.25 mg/mL and minimum bactericidal concentrations (MBCs) ≥ 2.5 mg/mL against all tested MDR isolates. Remarkably, biofilm formation was suppressed even at sub-MIC levels. Preliminary mechanistic insights, based on pH neutralization, Proteinase K, and catalase treatments, suggest that organic acids are the primary contributors to the observed antibacterial activity. Beyond direct antimicrobial effects, TH-ST373 CFS significantly reduced E. coli adhesion to HaCaT keratinocytes and downregulated TLR4 expression elicited by bacterial stimulation. Furthermore, it reduced the mRNA expression of pro-inflammatory cytokines TNF-α and IL-6. Collectively, these findings suggest that TH-ST373 CFS has multi-faceted activity against MDR E. coli, combining direct antibacterial effects with biofilm inhibition and modulation of host inflammatory responses, and may represent a promising antibiotic-sparing adjunct strategy.