Single Nucleotide Polymorphisms in Genes Associated with Isoniazid Resistance in Mycobacterium tuberculosis

Abstract

Isoniazid (INH) is one of the primary chemotherapeutic and prophylactic drugs used against Mycobacterium tuberculosis, the causative agent of tuberculosis, which remains the leading single cause of death due to an infectious agent throughout the world. Recent studies indicate that the median rate of primary resistance to INH is 7.3% (range, 1.5 to 32%) and that the rates of acquired resistance range from 5.3 to 70% globally (9, 45). The overall rate of resistance to INH is 8.4% in the United States and has remained relatively stable in the last decade (23). Global reports of clusters of tuberculosis cases caused by drug-resistant strains together with the emergence and dissemination of multidrug-resistant tuberculosis have underscored the need for research into the mechanisms of drug resistance and the design of more effective antituberculous agents. Despite the use of INH for several decades, the molecular basis for its bactericidal action and the mechanisms by which INH resistance evolves in M. tuberculosis are only beginning to be understood.INH has a simple chemical structure consisting of a hydrazide group attached to a pyridine ring, but its mode of action is very complex (8). It is proposed that INH enters M. tuberculosis as a prodrug by passive diffusion and is activated by catalase-peroxidase, encoded by katG, to generate free radicals, which then attack multiple targets in the cells (6). Recent studies have shown that an NADH-dependent enoyl acyl carrier protein (ACP) reductase, encoded by inhA, and a ␤-ketoacyl ACP synthase, encoded by kasA, are two potential intracellular enzymatic targets for activated INH; and both of these enzymes are involved in the biosynthesis of mycolic acids (4,19,20). Resistance-associated amino acid substitutions have been identified in the katG, inhA, and kasA genes of INHresistant clinical isolates of M. tuberculosis (7,20,24,26,29). In addition, mutations in the oxyR-ahpC intergenic region have been identified in INH-resistant isolates (36). Additional genetic and biochemical studies have shown that certain promoter mutations of alkylhydroperoxide reductase, encoded by ahpC, in INH-resistant isolates result in overexpression of ahpC as a compensatory mechanism for the loss of catalase activity due to katG mutations (15, 32). Recently, missense mutations were identified in ndh, a gene encoding NADH dehydrogenase, which is an essential respiratory chain enzyme that regulates the NADH/NAD ϩ ratio in cells (18,22). The molecular mechanism by which mutations in ndh confer INH resistance in M. tuberculosis is poorly understood. In addition, low-level INH resistance in mycobacteria has been shown to be

Detection of Multidrug Resistance in Mycobacterium tuberculosis

“…INH resistance is related to mutation(s) in katG, inhA, and/or the promoter region of mabA-inhA (17,22,38,39). In the present study, we found 11 novel mutations and a CTA insertion at nt position 1170 in katG.…”

“…Isolates carrying the T324P, L48Q, or M257T mutation and Ϫ15C3T upstream of mabA were resistant to INH. However, it is unclear whether T324P, L48Q, or M257T is related to INH resistance because the Ϫ15C3T upstream of mabA is known to confer INH resistance (10,22,39).…”

“…The L48Q-R463L mutant also carried the Ϫ15C3T transition upstream of mabA (Table 4). Therefore, the INH resistance of this mutant is likely due to the Ϫ15C3T mutation, which is known to be related to INH resistance (10,22,39).…”

Primer design for multiplex PCR using a genetic algorithm

“…Previous studies show that a variety of single nucleotide polymorphisms in multiple genes are found exclusively in INHresistant clinical isolates. These genes are either involved in mycolic acid biosynthesis or are overexpressed as a response to the buildup or cellular toxicity of INH [5]. Up to the present, twenty-four polymorphisms have been published.…”

The use of biodiversity as source of new chemical entities against defined molecular targets for treatment of malaria, tuberculosis, and T-cell mediated diseases: a review

“…Deletions of, or missense mutations in, the katG gene have been associated with decreased susceptibility to INH in approximately 50% of clinical isolates of M. tuberculosis (Heym et al 1995, Ramaswamy et al 2003. It has recently been shown that the S315T KatG mutant, which is one of the most commonly encountered substitutions in clinical INH-resistant strains, has reduced affinity for INH, which would result in decreased drug activation and ensuing INH resistance (Yu et al 2003).…”