De Novo Emergence of Genetically Resistant Mutants of Mycobacterium tuberculosis from the Persistence Phase Cells Formed against Antituberculosis Drugs In Vitro

Bacterial persisters are a subpopulation of cells that can tolerate lethal concentrations of antibiotics. However, the possibility of the emergence of genetically resistant mutants from antibiotic persister cell populations, upon continued exposure to lethal concentrations of antibiotics, remained u...

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Bibliographic Details
Published in:Antimicrobial agents and chemotherapy 2017-02, Vol.61 (2)
Main Authors: Sebastian, Jees, Swaminath, Sharmada, Nair, Rashmi Ravindran, Jakkala, Kishor, Pradhan, Atul, Ajitkumar, Parthasarathi
Format: Article
Language:English
Subjects:
Antitubercular Agents
Antitubercular Agents - pharmacology
Bacterial Proteins - genetics
DNA Gyrase - genetics
DNA-Directed RNA Polymerases - genetics
Dose-Response Relationship, Drug
Drug Resistance, Bacterial
Drug Resistance, Bacterial - drug effects
Drug Resistance, Bacterial - genetics
Fluoroquinolones - pharmacology
Genome, Bacterial
Hydroxyl Radical - metabolism
Mechanisms of Resistance
Moxifloxacin
Mutation
Mycobacterium tuberculosis
Mycobacterium tuberculosis - drug effects
Mycobacterium tuberculosis - genetics
Mycobacterium tuberculosis - growth & development
Oxidative Stress - drug effects
Rifampin - administration & dosage
Rifampin - pharmacology
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Summary:Bacterial persisters are a subpopulation of cells that can tolerate lethal concentrations of antibiotics. However, the possibility of the emergence of genetically resistant mutants from antibiotic persister cell populations, upon continued exposure to lethal concentrations of antibiotics, remained unexplored. In the present study, we found that Mycobacterium tuberculosis cells exposed continuously to lethal concentrations of rifampin (RIF) or moxifloxacin (MXF) for prolonged durations showed killing, RIF/MXF persistence, and regrowth phases. RIF-resistant or MXF-resistant mutants carrying clinically relevant mutations in the rpoB or gyrA gene, respectively, were found to emerge at high frequency from the RIF persistence phase population. A Luria-Delbruck fluctuation experiment using RIF-exposed M. tuberculosis cells showed that the rpoB mutants were not preexistent in the population but were formed de novo from the RIF persistence phase population. The RIF persistence phase M. tuberculosis cells carried elevated levels of hydroxyl radical that inflicted extensive genome-wide mutations, generating RIF-resistant mutants. Consistent with the elevated levels of hydroxyl radical-mediated genome-wide random mutagenesis, MXF-resistant M. tuberculosis gyrA de novo mutants could be selected from the RIF persistence phase cells. Thus, unlike previous studies, which showed emergence of genetically resistant mutants upon exposure of bacteria for short durations to sublethal concentrations of antibiotics, our study demonstrates that continuous prolonged exposure of M. tuberculosis cells to lethal concentrations of an antibiotic generates antibiotic persistence phase cells that form a reservoir for the generation of genetically resistant mutants to the same antibiotic or another antibiotic. These findings may have clinical significance in the emergence of drug-resistant tubercle bacilli.
ISSN:0066-4804
1098-6596
DOI:10.1128/AAC.01343-16