Genetic Stability of Mycobacterium smegmatis under the Stress of First-Line Antitubercular Agents: Assessing Mutagenic Potential

The sustained success of Mycobacterium tuberculosis as a pathogen arises from its ability to persist within macrophages for extended periods and its limited responsiveness to antibiotics. Furthermore, the high incidence of resistance to the few available antituberculosis drugs is a significant concern, especially since the driving forces of the emergence of drug resistance are not clear. Drug-resistant strains of Mycobacterium tuberculosis can emerge through de novo mutations, however, mycobacterial mutation rates are low. To unravel the molecular mechanisms of antibiotic pressure on genetic variability and their impact on mycobacterial survival, we systematically explored the effects of current antibiotics on genome stability, DNA repair system activation, and the dNTP pool using Mycobacterium smegmatis . Whole-genome sequencing revealed no significant increase in mutation rates after prolonged exposure to first-line antibiotics. However, the phenotypic fluctuation assay indicated rapid adaptation to antibiotics, likely mediated by non-genetic factors. The upregulation of DNA repair genes measured using qPCR suggests that genomic integrity is maintained through the activation of specific DNA repair pathways. Our results, indicating that antibiotic exposure does not result in de novo adaptive mutagenesis under laboratory conditions, do not lend support to the model suggesting antibiotic resistance development through drug pressure-induced microevolution.