Pleiotropic cellular responses underlying antibiotic tolerance in Campylobacter jejuni

Antibiotic tolerance enables antibiotic-susceptible bacteria to withstand prolonged exposure to high concentrations of antibiotics. Although antibiotic tolerance presents a major challenge for public health, its underlying molecular mechanisms remain to be elucidated. Previously, we have demonstrated that Campylobacter jejuni develops tolerance to clinically important antibiotics, including ciprofloxacin and tetracycline. To identify cellular responses associated with antibiotic tolerance, we conducted RNA-sequencing analysis on C. jejuni following the induction of antibiotic tolerance through exposure to elevated levels of ciprofloxacin or tetracycline. Additionally, we constructed knockout mutants for genes that showed significant changes in expression levels during antibiotic tolerance. We observed a significant upregulation of genes involved in protein chaperones, bacterial motility, DNA repair system, drug efflux pump, and iron homeostasis during antibiotic tolerance. Furthermore, the viability of these knockout mutants was significantly reduced compared to the wild-type strain, indicating the critical role of these cellular responses in sustaining antibiotic tolerance. Notably, we also found that the protein chaperone mutants Δ dnaK , Δ clpB , and Δ groESL exhibited increased protein aggregates under antibiotic treatment, suggesting protein chaperones play a critical role in managing protein aggregation and facilitating the survival of C. jejuni during antibiotic tolerance. In conclusion, our findings demonstrate that various cellular defense mechanisms collectively contribute to sustaining antibiotic tolerance in C. jejuni , providing novel insights into the molecular mechanisms of antibiotic tolerance that enable C. jejuni to withstand the lethal effects of antibiotics.