Inhibition of the respiration of multi-drug resistant clinical isolates of Mycobacterium tuberculosis by thioridazine: potential use for initial therapy of freshly diagnosed tuberculosis

ABSTRACT Effective tuberculosis treatment requires at least 6 months of combination therapy. Alterations in the physiological state of the bacterium during infection are thought to reduce drug efficacy and prolong the necessary treatment period, but the nature of these adaptations remain incompletely defined. To identify specific bacterial functions that limit drug effects during infection, we employed a comprehensive genetic screening approach to identify mutants with altered susceptibility to the first-line antibiotics in the mouse model. We identified many mutations that increase the rate of bacterial clearance, suggesting new strategies for accelerating therapy. In addition, the drug-specific effects of these mutations suggested that different antibiotics are limited by distinct factors. Rifampin efficacy is inferred to be limited by cellular permeability, whereas isoniazid is preferentially affected by replication rate. Many mutations that altered bacterial clearance in the mouse model did not have an obvious effect on drug susceptibility using in vitro assays, indicating that these chemical-genetic interactions tend to be specific to the in vivo environment. This observation suggested that a wide variety of natural genetic variants could influence drug efficacy in vivo without altering behavior in standard drug-susceptibility tests. Indeed, mutations in a number of the genes identified in our study are enriched in drug-resistant clinical isolates, identifying genetic variants that may influence treatment outcome. Together, these observations suggest new avenues for improving therapy, as well as the mechanisms of genetic adaptations that limit it. IMPORTANCE Understanding how Mycobacterium tuberculosis survives during antibiotic treatment is necessary to rationally devise more effective tuberculosis (TB) chemotherapy regimens. Using genome-wide mutant fitness profiling and the mouse model of TB, we identified genes that alter antibiotic efficacy specifically in the infection environment and associated several of these genes with natural genetic variants found in drug-resistant clinical isolates. These data suggest strategies for synergistic therapies that accelerate bacterial clearance, and they identify mechanisms of adaptation to drug exposure that could influence treatment outcome.

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Characterization of the Genetic Diversity of Extensively-Drug Resistant Mycobacterium tuberculosis Clinical Isolates from Pulmonary Tuberculosis Patients in Peru

PLoS ONE ◽

10.1371/journal.pone.0112789 ◽

2014 ◽

Vol 9 (12) ◽

pp. e112789 ◽

Cited By ~ 13

Author(s):

Omar Cáceres ◽

Nalin Rastogi ◽

Carlos Bartra ◽

David Couvin ◽

Marco Galarza ◽

...

Keyword(s):

Genetic Diversity ◽

Mycobacterium Tuberculosis ◽

Pulmonary Tuberculosis ◽

Clinical Isolates ◽

Drug Resistant ◽

Tuberculosis Patients ◽

Extensively Drug Resistant

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Evaluation of Pyrosequencing for Detecting Extensively Drug-Resistant Mycobacterium tuberculosis among Clinical Isolates from Four High-Burden Countries

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.03614-14 ◽

2014 ◽

Vol 59 (1) ◽

pp. 414-420 ◽

Cited By ~ 20

Author(s):

Kanchan Ajbani ◽

Shou-Yean Grace Lin ◽

Camilla Rodrigues ◽

Duylinh Nguyen ◽

Francine Arroyo ◽

...

Keyword(s):

Drug Resistance ◽

Mycobacterium Tuberculosis ◽

The Philippines ◽

Drug Susceptibility Testing ◽

Clinical Isolates ◽

Drug Resistant ◽

Second Line ◽

Additional Advantage ◽

Content Type ◽

Extensively Drug Resistant

ABSTRACTReliable molecular diagnostics, which detect specific mutations associated with drug resistance, are promising technologies for the rapid identification and monitoring of drug resistance inMycobacterium tuberculosisisolates. Pyrosequencing (PSQ) has the ability to detect mutations associated with first- and second-line anti-tuberculosis (TB) drugs, with the additional advantage of being rapidly adaptable for the identification of new mutations. The aim of this project was to evaluate the performance of PSQ in predicting phenotypic drug resistance in multidrug- and extensively drug-resistant tuberculosis (M/XDR-TB) clinical isolates from India, South Africa, Moldova, and the Philippines. A total of 187 archived isolates were run through a PSQ assay in order to identifyM. tuberculosis(via the IS6110marker), and to detect mutations associated with M/XDR-TB within small stretches of nucleotides in selected loci. The molecular targets includedkatG, theinhApromoter and theahpC-oxyRintergenic region for isoniazid (INH) resistance; therpoBcore region for rifampin (RIF) resistance;gyrAfor fluoroquinolone (FQ) resistance; andrrsfor amikacin (AMK), capreomycin (CAP), and kanamycin (KAN) resistance. PSQ data were compared to phenotypic mycobacterial growth indicator tube (MGIT) 960 drug susceptibility testing results for performance analysis. The PSQ assay illustrated good sensitivity for the detection of resistance to INH (94%), RIF (96%), FQ (93%), AMK (84%), CAP (88%), and KAN (68%). The specificities of the assay were 96% for INH, 100% for RIF, FQ, AMK, and KAN, and 97% for CAP. PSQ is a highly efficient diagnostic tool that reveals specific nucleotide changes associated with resistance to the first- and second-line anti-TB drug medications. This methodology has the potential to be linked to mutation-specific clinical interpretation algorithms for rapid treatment decisions.

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