Development of a novel three-bacteriophage cocktail for controlling Enterohemorrhagic Escherichia coli in iceberg lettuce and chicken-based sliced ham

Enterohemorrhagic Escherichia coli (EHEC) is a major foodborne pathogen responsible for severe intestinal diseases, including hemorrhagic colitis and hemolytic uremic syndrome. EHEC infections are primarily linked to the consumption of contaminated food, such as undercooked beef, raw vegetables, and unpasteurized dairy products, and even a low infectious dose can lead to illness. The increasing prevalence of antibiotic-resistant EHEC strains underscores the urgent need for alternative antimicrobial strategies. To address this challenge, three EHEC-infecting bacteriophages (phages), EH11, EP26, and EP27, were newly isolated from mixed environmental samples and utilized to construct a phage cocktail. Each phage exhibited a distinct host range, and their combination allowed targeting a broader range of host bacteria, including not only EHEC but also other pathogenic E. coli strains such as enteropathogenic and enterotoxigenic E. coli . The phage cocktail significantly delayed the emergence of bacterial resistance compared to single-phage treatments. The three phages exhibited high stability across a wide range of pH levels, temperatures, and NaCl concentrations, suggesting their suitability for application in diverse food matrices. The phage cocktail effectively disrupted biofilms and inhibited bacterial growth in food samples, including iceberg lettuce and chicken-based sliced ham, demonstrating its efficacy under practical conditions. These findings suggest that this phage cocktail has promising potential as an effective biocontrol agent for managing EHEC contamination in food systems and may serve as a viable alternative to conventional antibiotic treatments. • Phages EH11, EP26, and EP27 form a cocktail targeting EHEC. • Phage cocktail reduced EHEC biofilms by up to 91%. • Cocktail delayed bacterial resistance for 8 h, better than single phage treatment. • Phage cocktail reduced EHEC by ∼7 logs on iceberg lettuce and ∼1 log on sliced ham.