Sublethal benzalkonium chloride exposure induced direct protection and cross-protection to freezing stress in Salmonella Typhimurium

Abstract

Benzalkonium chloride (BAC) is a widely used disinfectant, but its improper application can induce Salmonella Typhimurium (S. Typhimurium) to develop tolerance to subsequent stressors. In this study, we demonstrated that adaptation of S. Typhimurium to BAC conferred both direct protection and cross-protection against cold stress, resulting in significantly enhanced survival at -20°C compared with non-adapted cells. BAC-adapted S. Typhimurium also exhibited a survival advantage on chicken meat during frozen storage at -20 °C. Membrane fatty acid analysis indicated that adaptation of S. Typhimurium to BAC led to a 2.2-fold increase in the levels of cyclopropane fatty acids. Moreover, BAC-adapted S. Typhimurium showed a significant 8.0% increase in cell surface hydrophobicity. RT-qPCR analysis demonstrated upregulation of efflux pump genes tolC, acrB, and mdfA (3.6-, 3.4-, and 3.0-fold, respectively) and cold shock-related genes cspA, cspE, and rpoS (2.4-, 4.0-, and 2.6-fold, respectively). These findings suggest that upregulation of cold shock genes in BAC-adapted cells was associated with the observed cross-protection. Taken together, adaptation of S. Typhimurium to BAC promoted the development of direct and cross-protective phenotypes, which may increase the risk of cross-contamination and salmonellosis in the food industry, posing a serious threat to food safety and public health.