title: Prevalence of mutations at codon 463 of katG gene in MDR and XDR clinical isolates of Mycobacterium tuberculosis in Belarus and application of the method in rapid diagnosis.
abstract: Isoniazid (INH) is a central component of drug regimens used worldwide to treat tuberculosis. In respect to high GC content of Mycobacterium tuberculosis, nonsynonymous mutations are dominant in this group. In this study a collection of 145 M. tuberculosis isolates was used to evaluate the conferring mutations in nucleotide 1388 of katG gene (KatG463) in resistance to isoniazid. A PCR-RFLP method was applied in comparison with DNA sequencing and anti-mycobacterial susceptibility testing. From all studied patients, 98 (67.6%) were men, 47 (32.4%) were women, 3% were <15 and 9% were >65 years old; male to female ratio was 1:2.4. PCR result of katG for a 620-bp amplicon was successful for all purified M. tuberculosis isolates and there was no positive M. tuberculosis culture with PCR negative results (100% specificity). Subsequent PCR RFLP of the katG identified mutation at KatG463 in 33.3%, 57.8% and 59.2% of our clinically susceptible, multidrug resistant TB (MDR) and extensively drug resistant (XDR) isolates, respectively. Strains of H37Rv and Academic had no any mutations in this codon. M. bovis was used as a positive control for mutation in KatG463. Automated DNA sequencing of the katG amplicon from randomly selected INH-susceptible and resistant isolates verified 100% sequence accuracy of the point mutations detected by PCR-RFLP. We concluded that codon 463 was a polymorphic site that is associated to INH resistance (a missense or "quiet" mutation). RFLP results of katG amplicons were identical to those of sequence method. Our PCR-RFLP method has a potential application for rapid diagnosis of M. tuberculosis with a high specificity.
authors: Arak University of Medical Sciences, Tuberculosis and Pediatric Infectious Diseases Research Center, Arak Iran. mmatinam81@yahoo.com
authors: M
authors: M
authors: Arjomandzadegan
authors: P
authors: P
authors: Owlia
authors: R
authors: R
authors: Ranjbar
authors: A A
authors: AA
authors: Farazi
authors: Masume
authors: M
authors: Sofian
authors: Maryam
authors: M
authors: Sadrnia
authors: E
authors: E
authors: Ghaznavi-Rad
authors: Larisa K
authors: LK
authors: Surkova
authors: L P
authors: LP
authors: Titov
mesh_terms: D000293:Adolescent; D000328:Adult; D000368:Aged; D000995:Antitubercular Agents / Q000494:pharmacology; D001426:Bacterial Proteins / Q000235:genetics*; D002374:Catalase / Q000235:genetics*; D003062:Codon; D054908:Extensively Drug-Resistant Tuberculosis / Q000175:diagnosis / Q000382:microbiology*; D005260:Female; D006801:Humans; D007538:Isoniazid / Q000494:pharmacology; D008297:Male; D008826:Microbial Sensitivity Tests / Q000379:methods; D008875:Middle Aged; D020125:Mutation, Missense; D009169:Mycobacterium tuberculosis / Q000187:drug effects / Q000235:genetics* / Q000302:isolation & purification; D016133:Polymerase Chain Reaction / Q000379:methods; D012150:Polymorphism, Restriction Fragment Length; D015995:Prevalence; D002094:Republic of Belarus; D017422:Sequence Analysis, DNA; D018088:Tuberculosis, Multidrug-Resistant / Q000175:diagnosis / Q000382:microbiology*; D055815:Young Adult
chemical_list: D000995:Antitubercular Agents; D001426:Bacterial Proteins; D003062:Codon; D002374:Catalase; C480353:katG protein, Mycobacterium tuberculosis; D007538:Isoniazid