Rapid Detection of Rifampicin- and Isoniazid-Resistant Mycobacterium tuberculosis using TaqMan Allelic Discrimination

Abstract

ObjectivesMultidrug-resistant tuberculosis (MDR-TB) is a global problem that many countries are challenged with. Rapid and accurate detection of MDR-TB is critical for appropriate treatment and controlling of TB. The aim of the present study was to evaluate the TaqMan allelic discrimination without minor groove binder (MGB) as a rapid, efficient, and low-cost method for detection of drug resistant strains of Mycobacterium tuberculosis.MethodsA total of 112 M. tuberculosis isolates from cases with diagnosed TB were subjected to drug susceptibility testing (DST), using the proportion method. Resistant isolates were tested for characterization of mutations in the rpoB and KatG genes by TaqMan genotyping.ResultsOf 112 M. tuberculosis isolates for which DST was performed, three, one, and two isolates were MDR, rifampin (RIF) resistant, and isoniazid (INH) resistant, respectively. According to the threshold cycle (Ct) and curve pattern of mutants, TaqMan probes detect all of the mutations in the analyzed genes (katG 315, AGC→ACC, rpoB 531, TCG→TTG, and rpoB 531, TCG→TGG).ConclusionThe present study suggests that drug-resistant strains of M. tuberculosis can be detected by pattern’s curve or Ct with TaqMan probes without MGB in real-time polymerase chain reaction (PCR).

The Value of the inhA Mutation Detection in Predicting Ethionamide Resistance Using Melting Curve Technology

“…According to data from domestic and overseas research, the majority of ETH/PTH-resistant strains also show INH resistance. 10 , 14 , 15 In the present study, almost all ETH-resistant strains were also found to be resistant to both INH and RIF (96.4%), and the ETH-resistance rate in MDR-TB was 27.8%, which is consistent with our previous studies and similar data (20%–24.8%) from TB treatment institutions in China. 16–19…”

“…A recent study in South Korea found that only 23 (67%) of 34 PTH-resistant strains had an inhA mutation, while data from a study in Guangzhou, China indicates that, of 46 PTH-resistant strains, 43.2% had an inhA promoter (12 strains were c-15t and 4 strains t-8c), and 6.2% had a coding gene mutation (all were S94A). 10 , 21 The results of the present study show an inhA mutation rate in ETH-resistant strains (42.9%) similar to that in the study in China referenced above.…”

“…Of the 28 phenotypically ETH-resistant strains, inhA mutations accounted for 42.9%, which is consistent with previous reports. 10,20 However, in this study, only 35.3% of the 34 inhA mutant strains were resistant to ETH, and only 42.9% of ETH-resistant strains had inhA mutations. A recent study in South Korea found that only 23 (67%) of 34 PTH-resistant strains had an inhA mutation, while data from a study in Guangzhou, China indicates that, of 46 PTH-resistant strains, 43.2% had an inhA promoter (12 strains were c-15t and 4 strains t-8c), and 6.2% had a coding gene mutation (all were S94A).…”

“…This technique is widely used in clinical work. 9 , 10 In this study, we analyze the correlation between inhA mutation test results and phenotypic ETH susceptibility through MeltPro TB assay and evaluate whether the inhA test can be used to guide the clinical application of ETH where phenotypic DST results are unavailable.…”

Drug resistance and the genotypic characteristics of rpoB and katG in rifampicin- and/or isoniazid-resistant Mycobacterium tuberculosis isolates in central Vietnam

“…Four TaqMan probes, which could distinguish one-base mismatches, were used to detect genetic variation in the wild-type M. tuberculosis strain (H37Rv) in both rpoB and katG . All primers and probes used in this study were manufactured by Integrated DNA Technologies Inc. ( Table 1 ) [ 14 , 15 ].…”