Bacterial persistence to antibiotics activated by tRNA mutations

Abstract Objectives Bacterial persistence is a significant cause of the intractability of chronic and relapsing infections. Despite its importance, many of the underlying mechanisms are still not well understood. Methods Antibiotic-tolerant mutants of Burkholderia thailandensis were isolated through exposure to lethal doses of AMP or MEM, followed by whole-genome sequencing to identify mutations. Subsequently, these mutants underwent comprehensive characterization via killing curves, growth curves, and persistence-fraction plots. Northern blot analysis was employed to detect uncharged tRNA, while the generation of relA and spoT null mutations served to confirm the involvement of the stringent response in this persistence mechanism. Phenotypic reversion of the persistence mutation was demonstrated by incubating the mutants without antibiotics for 2 weeks. Results We have discovered a novel mechanism of persistence triggered by specific mutations at positions 32 or 38 within the anticodon loop of tRNAAsp. This leads to heightened persistence through a RelA-dependent stringent response. Notably, this persistence can be easily reverted to wild-type physiology by losing the mutant tRNA allele within the tRNA gene cluster when persistence is no longer essential for survival. Conclusions This distinct form of persistence underscores the novel function of tRNA mutations at positions 32 or 38 within the anticodon loop, as well as the significance of the tRNA gene cluster in conferring adaptability to regulate persistence for enhanced survival.