Insights into Cronobacter sakazakii adaptation to Benzalkonium chloride: virulence assessment and transcriptomics...

Cronobacter sakazakii is an opportunistic foodborne pathogen associated with significant health risks. Benzalkonium chloride (BC) is a widely used disinfectant in food processing and healthcare settings. Nonetheless, increasing evidence indicates that exposure to sublethal concentrations of disinfectants can alter virulence-associated traits in foodborne pathogens. Despite this, the fundamental mechanisms of bacterial adaptation and its influence on virulence in C. sakazakii remain incompletely understood, particularly from a global transcriptomic viewpoint. The impact of BC adaptation on virulence-associated phenotypes of C. sakazakii was evaluated using a combination of in vitro assays, including planktonic growth, biofilm formation, adhesion and invasion of Caco-2 cells, and endotoxin production, together with an in vivo Galleria mellonella infection model. BC adaptation resulted in significantly reduced planktonic growth, biofilm formation, and adhesion and invasion toward Caco-2 cells (P < 0.05). In contrast, BC-adapted strains exhibited increased endotoxin production and significantly enhanced lethality in G. mellonella compared with non-adapted cells. The molecular pathways of C. sakazakii adaptation to BC were examined in this study using high-throughput RNA sequencing. KEGG analysis revealed coordinated transcriptional changes associated with ABC transport systems, flagellar assembly, quorum sensing, amino acid metabolism, and fatty acid metabolism to adapt to external environmental changes. Overall, these findings indicate that BC adaptation leads to differential modulation of virulence-associated traits in C. sakazakii, indicating that sublethal disinfectant exposure may influence virulence-associated phenotypes relevant to food safety management.