Rapid Identification Of A Point Mutation Of The Mycobacterium tuberculosis Catalase-Peroxidase (katG) Gene Associated With Isoniazid Resistance

Cockerill, F. R.; Uhl, James R.; Temesgen, Zelalem; Zhang, Ying; Stockman, Leslie; Roberts, Glenn D.; Williams, Diana L.; Kline, Bruce C.

doi:10.1093/infdis/171.1.240

Cited by 106 publications

(72 citation statements)

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“…However, in previous studies, the mutation at codon 463 was found in 3 to 61% of INH-susceptible M. tuberculosis isolates (5,6,13,17,18,23). Also, it was found that the activity of catalase, a katG-encoded enzyme, did not differ among isolates having either Arg or Leu at codon 463 (21,22).…”

Section: Tuberculosis Isolates and Assessment Of Inh Resistancementioning

confidence: 87%

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The Susceptibility of Mycobacterium tuberculosis to Isoniazid and the Arg→Leu Mutation at Codon 463 of katG Are Not Associated

et al. 2001

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Tuberculosis is the leading cause of death due to infectious diseases worldwide (4), although various drugs against Mycobacterium tuberculosis are available. One of the mainstay drugs for the treatment of tuberculosis is isoniazid (INH). Its effectiveness against M. tuberculosis was initially reported in 1952 (3, 15). Today, INH-resistant M. tuberculosis organisms are not rare anymore; prevalence was reported to be 7% in a recent study performed in The Netherlands (25). The emergence of multidrug-resistant strains (resistant to at least INH and rifampin) (7,11,12,20) has further complicated the treatment of tuberculosis. Therefore, and because of the organism's slow growth rate, rapid methods for detecting drug resistance in clinical isolates of M. tuberculosis are required. The primary mechanism of resistance in M. tuberculosis is the accumulation of mutations in genes coding for drug targets or drug-converting enzymes (16).In the last decade, mutations in katG (24, 26) and inhA (2) have been found to account for 60 to 70% and 10 to 15% of INH resistance cases among M. tuberculosis isolates, respectively (11). The two predominant mutations of katG, and those most referred to, are found within codons 315 and 463 (17).The mutation at codon 315 has been found to be an important indicator for INH resistance as well as for multidrug resistance among isolates of M. tuberculosis organisms recovered from patients in The Netherlands (25).The aims of this study were to assess whether the Arg463Leu mutation is also predictive of INH resistance and, if so, to develop a diagnostic PCR-based screening method for this type of INH resistance.(This work was presented in part at the 40th Interscience Conference on Antimicrobial Agents and Chemotherapy, Toronto, Canada, 17 to 20 September 2000.) M. tuberculosis isolates and assessment of INH resistance.M. tuberculosis isolates from 395 patients who were diagnosed with tuberculosis in The Netherlands in the period of 1993 to 1997 were used in this study. The isolates were sent by medical microbiology laboratories in The Netherlands to the National Institute of Public Health and the Environment (RIVM, Bilthoven, The Netherlands) for routine typing and susceptibility tests. Susceptibility to INH was measured with the MIC method using 0.1, 0.2, 0.5, 1, 2, 5, 10, 20, and 50 g of INH/ml in Middlebrook 7H10 medium (8). Isolates were considered resistant if more than 1% of the bacteria in the inoculum grew in the presence of INH concentrations of Ն1 g/ml. If growth of more than 1% of the inoculum in the presence of 0.5 g of INH/ml occurred, then the isolates were classified as having intermediate susceptibility.DNA isolation. M. tuberculosis isolates were grown on Lö-wenstein-Jensen solid medium or Middlebrook 7H9 liquid medium for 7 days to an optical density corresponding to 10 8 bacteria/ml and were harvested by centrifugation (4,500 ϫ g for 15 min). Chromosomal DNA was isolated as described by Ausubel et al. (1). Briefly, the bacteria were killed by heating at 80°C for 20 min and the...

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Section: Tuberculosis Isolates and Assessment Of Inh Resistancementioning

confidence: 87%

“…The genetic mechanism of INH resistance remains unknown for 10 to 15% of the INH-resistant isolates. The mutations in katG occur in 50 to 60% of the INH-resistant isolates at codon 315 and in 25 to 45% of these isolates at codon 463 (5,16,17).…”

Section: Tuberculosis Isolates and Assessment Of Inh Resistancementioning

confidence: 99%

The Susceptibility of Mycobacterium tuberculosis to Isoniazid and the Arg→Leu Mutation at Codon 463 of katG Are Not Associated

et al. 2001

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“…By contrast, INHresistant strains exhibit mutations in several genes, such as katG, inhA, oxyR, and ahpC (22,27,30). The mutation of codon 315 (Ser) in the catalase-peroxidase (katG) gene is the most frequent site (30 to 65% of resistant strains) (4,11). Furthermore, EMB-resistant strains have a point mutation at codon 306 (Met) in embB (3,28), and the frequency is about 70% (3).…”

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Dual-Probe Assay for Rapid Detection of Drug-Resistant Mycobacterium tuberculosis by Real-Time PCR

et al. 2004

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Mutations in particular nucleotides of genes coding for drug targets or drug-converting enzymes lead to drug resistance in Mycobacterium tuberculosis. For rapid detection of drug-resistant M. tuberculosis in clinical specimens, a simple and applicable method is needed. Eight TaqMan minor groove binder (MGB) probes, which discriminate one-base mismatches, were designed (dual-probe assay with four reaction tubes). The target of six MGB probes was the rpoB gene, which is involved in rifampin resistance; five probes were designed to detect for mutation sites within an 81-bp hot spot of the rpoB gene, and one probe was designed as a tuberculosis (TB) control outside the rpoB gene hot-spot. We also designed probes to examine codon 315 of katG and codon 306 of embB for mutations associated with resistance to isoniazid and ethambutol, respectively. Our system was M. tuberculosis complex specific, because neither nontuberculous mycobacteria nor bacteria other than mycobacteria reacted with the system. Detection limits in direct and preamplified analyses were 250 and 10 fg of genomic DNA, respectively. The system could detect mutations of the rpoB, katG, and embB genes in DNAs extracted from 45 laboratory strains and from sputum samples of 27 patients with pulmonary TB. This system was much faster (3 h from DNA preparation) than conventional drug susceptibility testing (3 weeks). Results from the dual-MGB-probe assay were consistent with DNA sequencing. Because the dual-probe assay system is simple, rapid, and accurate, it can be applied to detect drug-resistant M. tuberculosis in clinical laboratories.

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“…The presence of R463 or L463 at codon 463 and the presence of S315T substitution in the katG gene was determined by amplification of the corresponding katG gene DNA region by PCR followed by restriction endonuclease digestion of the PCR amplified fragments to generate restriction fragment length polymorphism as described previously (1,5).…”

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confidence: 99%

Molecular Fingerprinting of Isoniazid‐Resistant Mycobacterium tuberculosis Isolates from Chest Diseases Hospital in Kuwait

Mokaddas

Ahmad

Abal

2002

Microbiology and Immunology

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Rapid Identification Of A Point Mutation Of The Mycobacterium tuberculosis Catalase-Peroxidase (katG) Gene Associated With Isoniazid Resistance

Cited by 106 publications

References 10 publications

The Susceptibility of Mycobacterium tuberculosis to Isoniazid and the Arg→Leu Mutation at Codon 463 of katG Are Not Associated

The Susceptibility of Mycobacterium tuberculosis to Isoniazid and the Arg→Leu Mutation at Codon 463 of katG Are Not Associated

Dual-Probe Assay for Rapid Detection of Drug-Resistant Mycobacterium tuberculosis by Real-Time PCR

Molecular Fingerprinting of Isoniazid‐Resistant Mycobacterium tuberculosis Isolates from Chest Diseases Hospital in Kuwait

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