Usefulness of resistant gene markers for predicting treatment outcome on second-line anti-tuberculosis drugs

Abstract

Aim: Mutations in rrs [nucleotide (nt) 1401], gyrA gene (codons 90, 91 or 94), tlyA, ethA and thyA genes of Mycobacterium tuberculosis (MTB) were evaluated for their usefulness in predicting treatment outcome of kanamycin (KM), capreomycin (CPM), ofloxacin (OFX), ethionamide (ETH) and para‐aminosalicylic acid (PAS). Methods and Results: DNA sequence analyses of these genes were performed against 188 MTB isolates obtained from patients put on second‐line anti‐TB drugs (SLDs) with well‐documented clinical history and treatment outcome. Mutations in rrs and gyrA have 100% positive predictive value (PPV) in predicting treatment failure for KM and OFX, while 88·9 and 80% were obtained, respectively, when tlyA and rrs mutations were considered in CPM. For ETH and PAS, the PPV of using ethA and thyA mutations to predict treatment failure was 82·5 and 89·3%, respectively. Conclusions: Our study demonstrated high specificities of gene mutations in predicting poor treatment outcome; however, further technical advancement is required to make the molecular detection of resistances to other SLDs feasible in clinical laboratories. Significance and Impact of the Study: This is the first study to correlate different polymorphisms of major SLD resistance gene markers with predicted treatment outcome, using an international set of well‐documented clinical MTB strains.

Correlations between Gene Resistant Markers and Second-Line Anti-TB Drug Resistance in Pre-XDR and XDR-TB Patients

“…One study in 2001 reported that streptomycin resistance was related to mutations in the 500 region of rrs [22]. Our study followed primer patterns from one related study in Hong Kong that could predict 96.8% specificity and 64.2% sensitivity for kanamycin from rrs on gene marker 1401 [11]. showed 78% of kanamycin resistance correlated to SNPs in rrs at nt1401 [28] and concluded that the commercial diagnostic tool determining the aminoglycoside drug resistance target at rrs nt 1401 could detect 78% of kanamycin resistance in XDR-TB isolates [28].…”

“…PCR product comprised 665 base pair fragments, according to the following steps: beginning denaturation at 95˚C for 2 min, followed by 35 cycles of 94˚C for 1 min, 60˚C for 1min, 68˚C for 2 min, followed by elongation at 68˚C for 10 min [11]. The amplified products were purified with a multiscreen filter plate (Millipore Corp., Bedford, MA, USA) and DNA sequencing data were produced by the ABI 3730xl DNA analyzer at Macrogen.…”

“…The following steps were performed: one denature cycle at 95˚C for 2 min, followed by 30 cycles of 1 min at 95˚C, 1 min at 65˚C and 1 min at 72˚C, followed by elongation at 72˚C for 10 min. PCR product comprised 320 base pairs in size (11). To detect kanamycin resistance, the primers were designed to identify mutations in codon 1401 of rrs gene: 16s-1F…”

“…Related studies have shown specificities of gene mutations in predicting kanamycin [9] [10] [11] and ofloxacin resistance [11] [12] [13] [14]. Many studies have been conducted on genotypic and phenotypic characteristics of drug resistant M. tuberculosis; however, this kind of study is rare in Thailand [15].…”

Old and New TB Drugs: Mechanisms of Action and Resistance

“…However, only slightly more than a third of the evaluated PAS-resistant strains had mutations in thyA, suggesting the existence of additional mechanisms of PAS resistance. Thr202Ala has been reported as the most common mutation associated with PAS resistance, although this mutation has also been identified in several PAS-susceptible isolates (Leung, Yip et al 2010).…”