Single Nucleotide Polymorphisms in Genes Associated with Isoniazid Resistance in Mycobacterium tuberculosis (original) (raw)
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The evolution of resistant TB strain against isoniazid (INH) drug is alarming in the global population. The molecular mechanism of resistance for this compound is found to be associated with mutations in katG and inhA genes. However, types and frequency of mutations vary with geographical region. Therefore, this study aimed to explore new targets of katG and inhA gene mutations and their correlation with isoniazid (INH) minimal inhibitory concentration (MIC) in clinical isolates of Mycobacterium tuberculosis from Uttar Pradesh, India. Materials and methods: The level of INH resistance was measured by absolute concentration method. DNA sequencing was performed to determine mutations in INH-resistant and -sensitive isolates. Results: Totally, 44/50 INH-resistant isolates had the following mutations in katG gene at codon 315: Ser-Thr (41/44), Ser-Ile (1/44) and Ser-Asn (1/44), with INH MIC ranges 2 to >16 mg/ml. Only one isolate showed the following mutation at codon 299: Gly-Ser with MIC of 0.2 mg/ml. Two isolates showed the following novel mutation in inhA gene at codon 17:
PHENOTYPIC AND GENOTYPIC CHARACTERISTICS OF ISONIAZID RESISTANCE IN MYCOBACTERIUM TUBERCULOSIS
TJPRC, 2013
Tuberculosis (TB) is a single infectious disease, caused by Mycobacterium tuberculosis and represent high mortality of people in many countries. It is estimated that over three million people die a year of tuberculosis. Isoniazid (isonicotinic acid hydrazide; INH) is one of the structurally simplest primary chemotherapeutic and prophylactic drugs used to treat M. tuberculosis since 1952. INH is the most widely administered to tuberculosis patients than any other drug, and among the antituberculosis drugs, it is against INH emergence of resistance have been frequently reported. INHresistance is apparently controlled by a complex genetic system that involves several genes, namely, katG, inhA, oxyRahpC. The two predominant mutations of katG, and those most referred to, are found within codons 315 and 463. The primary target of activated INH is an NADH-dependent enoyl-acyl carrier protein reductase, designated InhA. The aim of this study is to confirm M. tuberculosis to PCR targeting MPB64 gene and IS6110 and to determine isoniazid resistance by in vitro phenotypic resistant to the drug by Broth Microdilution method and resistance associated mutation by DNA isolation, PCR amplification, DNA sequencing analysis to detect mutations within katG gene and inhA gene of the isolates to search further why the isolate become isoniazid-resistant. The study revealed INH-resistant isolates for which the INH MICs were 0.19 to 3.125 mg/ml had polymorphism in codon 463 (R463L), and insertions at 323, 719 and 720 with these mutations conferring low level INH resistance.
American Journal of Microbiological Research, 2014
The KatG gene of Mycobacterium Tuberculosis has been associated with isoniazid (INH) drug resistance. While isoniazid (INH) considered as corner stone and main chemotherapy used throughout the world to manage tuberculosis, thus the Progress in apprehension of principle concepts associated with resistance to isoniazid (INH) has allowed molecular tests in addition to bioinformatics tool for the detection of drug-resistant tuberculosis to be developed. In Consecutive isolates (n = 20) of multidrug-resistant Mycobacterium tuberculosis, part of the katG was sequenced for INH resistance analysis. BLAST analysis of all sequences revealed 100% identity with the available strain "EGY-K361" Mycobacterium tuberculosis with Accession No: KC49137.1 except 6 isolates :isolate1, 2, 4, 11, 15, and isolate 20 revealed 99% identity. Thosesix isolates (30%) have detected mutation in Catalase-peroxidase enzyme S315T; three isolate from six 3/6 (50%) of mutant isolates have SNP AGC>ACC substitution while others 3/6 have substitution C>G in position 1280 which may contributed in altering gene expression. The secondary structure of wild and mutant proteins had been done using phyre2 software while the three dimensional structures of them had been done by Chimera software. Stability of mutant protein was increased which detected by i-mutant. Phylogenetic tree of the sequences revealed two distinct phylogroups: mutant isolates and wild isolates phylogroups with controls from different countries retrieved from Gene bank. Serine at position 315 is one of potential drug active sites that proved via SiteEngine soft ware, therefore any substitution will change efficiency of INH.
The KatG gene of Mycobacterium Tuberculosis has been associated with isoniazid (INH) drug resistance. While isoniazid (INH) considered as corner stone and main chemotherapy used throughout the world to manage tuberculosis, thus the Progress in apprehension of principle concepts associated with resistance to isoniazid (INH) has allowed molecular tests in addition to bioinformatics tool for the detection of drug-resistant tuberculosis to be developed. In Consecutive isolates (n = 20) of multidrug-resistant Mycobacterium tuberculosis, part of the katG was sequenced for INH resistance analysis. BLAST analysis of all sequences revealed 100% identity with the available strain "EGY-K361" Mycobacterium tuberculosis with Accession No: KC49137.1 except 6 isolates :isolate1, 2, 4, 11, 15, and isolate 20 revealed 99% identity. Thosesix isolates (30%) have detected mutation in Catalase-peroxidase enzyme S315T; three isolate from six 3/6 (50%) of mutant isolates have SNP AGC>ACC substitution while others 3/6 have substitution C>G in position 1280 which may contributed in altering gene expression. The secondary structure of wild and mutant proteins had been done using phyre2 software while the three dimensional structures of them had been done by Chimera software. Stability of mutant protein was increased which detected by i-mutant. Phylogenetic tree of the sequences revealed two distinct phylogroups: mutant isolates and wild isolates phylogroups with controls from different countries retrieved from Gene bank. Serine at position 315 is one of potential drug active sites that proved via SiteEngine soft ware, therefore any substitution will change efficiency of INH.
Diagnostic Microbiology and Infectious Disease, 2003
The katG, inhA and ahpC genes, in 71 isoniazid (INH)-resistant and 26 INH-susceptible Mycobacterium tuberculosis isolates, from South Korea were examined by sequencing and MspI restriction enzyme analysis. Mutations in the katG 315 alone, katG 315 plus inhA, katG 315 plus ahpC, katG 309 alone, katG 309 plus inhA, inhA alone, and ahpC alone, were detected in 54.9, 2.8, 1.4, 1.4, 1.4, 19.7, and 5.6% of the 71 INH-resistant isolates, respectively. There was no statistically significant difference (p > 0.05) in the frequencies of these mutations for the INH-monoresistant compared with the multidrug-resistant isolates. Mutations in the katG codon 315 were associated with the high-level of INH resistance (MIC, >1 microg/ml), whereas the mutation in the inhA promoter region was associated with the low-level of INH resistance (MIC, >0.2 to 1 microg/ml). The previously undescribed GGT-->GAT (Gly-->Asp) mutation in the katG codon 309 was found in two rifampin, including-multidrug-resistant isolates, but we cannot assess if this is predictive of INH resistance. The sensitivity and specificity of molecular analysis of the katG codon 315 and/or the inhA promoter region were 80.3 and 100%, respectively. Therefore, mutations in these regions are highly predictive of INH resistance in South Korea.
Pathogens and Global Health, 2021
Tuberculosis (TB) is a global threat due to the emergence and spread of drug-resistant Mycobacterium tuberculosis (MTB). Isoniazid (INH) is the main antibiotic used for prevention and treatment of TB. Evidence shows that accumulated mutations can produce INH resistant (INHR) strains, resulting in the progression of multidrug-resistant (MDR) TB. Since point mutations in katG gene, inhA gene, and oxyR-ahpC region correlated with the INH resistance, in this study, we aimed to identify mutations in these three genes in INHR and MDR clinical isolates of MTB by Sanger DNA sequencing analysis. Thirty-three out of 438 isolates were resistant, including 66.7% INHR and 30.3% MDR isolates. In the katG gene, 68.2% INHR isolates had nonsynonymous point mutations, mainly R463L (63.6%), and non-synonymous point mutation KatG L587P was seen in one of the MDR isolate. A novel silent substitution L649L was identified in the inhA gene of the MDR isolates. The oxyR-ahpC intergenic region g-88a common mutations (63.6%) in INHR and two distinct novel mutations were found at positions −76 and −77 of the oxyR-ahpC intergenic region. The coexistence of katG non-codon 315 with oxyR-ahpC intergenic region mutations was highly frequent in INHR 59.1% and MDR isolates 70%. Since mutations of all three genes 95.5% lead to the detection of INHR, they might be useful for molecular detection. Our results indicated the continuous evolution and region-specific prevalence of INH resistance. Overall, identification of new mutations in INH resistance can improve the available strategies for diagnosis and control of TB.
Antimicrobial Agents and Chemotherapy, 2015
Ethionamide (ETH) is an antibiotic used for the treatment of multidrug-resistant (MDR) tuberculosis (TB) (MDR-TB), and its use may be limited with the emergence of resistance in the Mycobacterium tuberculosis population. ETH resistance in M. tuberculosis is phenomenon independent or cross related when accompanied with isoniazid (INH) resistance. In most cases, resistance to INH and ETH is explained by mutations in the inhA promoter and in the following genes: katG , ethA , ethR , mshA , ndh , and inhA . We sequenced the above genes in 64 M. tuberculosis isolates ( n = 57 ETH-resistant MDR-TB isolates; n = 3 ETH-susceptible MDR-TB isolates; and n = 4 fully susceptible isolates). Each isolate was tested for susceptibility to first- and second-line drugs using the agar proportion method. Mutations were observed in ETH-resistant MDR-TB isolates at the following rates: 100% in katG , 72% in ethA , 45.6% in mshA , 8.7% in ndh , and 33.3% in inhA or its promoter. Of the three ETH-susceptib...