Physiological, microscopic and proteomic performance of Escherichia coli O157:H7 biofilms challenged with antagonistic lactic acid bacteria as a bio-decontamination tool for the food industry

Enterohemorrhagic Escherichia coli O157:H7 (EHEC) biofilms are a critical concern in food industry due to their resilience, persistence, and ability to enhance pathogen survival on processing surfaces. Controlling these bio- films is paramount to mitigating the risks associated with EHEC. This work evaluates the potential of three lactic acid bacteria (LAB) strains—Lactiplantibacillus plantarum CRL1075, L. plantarum CRL1482, and Pediococcus pentosaceus CRL2145—as biocontrol agents to inhibit EHEC biofilms through exclusion, competition, and displacement strategies under conditions mimicking meat industry environment (12 ◦C, stainless steel, and meat- based culture medium). EHEC biofilms showed by scanning electron microscopy and confocal laser scanning microscopy high adherence and robustness. Introducing LAB strains led to substantial reductions in EHEC biofilm formation, with the competition strategy emerging most effective, reducing biofilms by up to 6.5 log units. Exclusion and displacement strategies also decreased pathogen viability, with P. pentosaceus CRL2145 showing consistent inhibitory effects across all conditions. EHEC-CRL2145 mixed biofilms exhibited reduced biomass and covered surface area, with LAB clustered around E. coli cells, limiting their growth and adhesion. Proteomic analysis revealed repression of 85 EHEC proteins associated with essential metabolic pathways, stress responses, and biofilm maintenance, which compromised the pathogen’s ability to sustain biofilm integrity and viability. Additionally, some EHEC proteins involved in adhesion and virulence were upregulated, possibly as stress response to LAB antagonism. Importantly, while the antimicrobial potential of LAB against foodborne pathogens was recently reported, this is the first study to describe the metabolic response of EHEC biofilms when challenged by LAB.