Isolation and characterization of persisters of the pathogenic microorganism Staphylococcus aureus

Staphylococcus aureus is a notorious pathogen responsible for significant morbidity and mortality in both human society and animal husbandry. The presence of S. aureus persisters is also one of the leading causes of recurrent and chronic diseases. Persisters are a subset of growth-arrested bacteria within a susceptible bacterial population that are able to tolerate antibiotic treatment and resuscitate after stress removal. Consequently, investigating their formation and characteristics is of crucial importance to provide mechanism-based options for their eradication. However, one challenge in mechanistic research on persisters is the enrichment of pure persisters. In this work, we validated a proposed method to isolate persisters from vancomycin and enrofloxacin generated persistent populations. With this, we analyzed the proteome profile of pure persisters and revealed the distinct mechanisms associated with vancomycin and enrofloxacin induced persisters. Furthermore, morphological and metabolic characterizations were performed, indicating further differences between these two persister populations. Finally, we assessed the effect of ATP repression, protein synthesis inhibition and reactive oxygen species (ROS) level on persister formation. In conclusion, this work provides a comprehensive understanding of S. aureus vancomycin and enrofloxacin induced persisters at the molecular, single cell and population levels, facilitating a better understanding of persisters and the development of effective strategies to combat them.