Detection of Mutations Associated with Resistance to Second-line Drugs in Isolates of Mycobacterium tuberculosis

Abstract

Background: Mycobacterium tuberculosis (M. tuberculosis) develops resistance toward second-line anti-tuberculous drugs, mostly through mutations of the chromosome. The mechanism of resistance is complicated and accompanies several genes as rrs and eis promoter mutations. Objective: To assess the validity of the Real-time PCR (Rt-PCR) in recognizing the mutations of the resistance to second-line injectable agents (capreomycin, amikacin and kanamycin) in multidrug resistant tuberculosis (MDR-TB) clinical isolates in comparison with the gold standard proportion method (PM) using Lowenstein-Jensen (LJ) media. Methodology: This study was conducted on 48 MDR-TB clinical isolates (from sputum and bronchioalveolar lavage samples) obtained from the Egyptian National Central Laboratory of the Ministry of Health, Egypt. The isolates obtained were resistant to a minimum one of the second-line anti-tuberculous drugs (ofloxacin, capreomycin, amikacin, and kanamycin) identified by PM using LJ media. Isolates were tested by Rt-PCR to track mutations in rrs and the eis promoter. Results: isolates were positive for rrs, and 32 isolates for the promoter of eis using Rt-PCR. Comparing the results to the gold standard PM, an agreement of 100%, and 69% were found to rrs, and the eis promoter, respectively. Conclusion: Using the Real-time PCR for recognizing mutations related to second-line anti-tuberculous drugs highly agrees with the PM. This could help in MDR-TB early detection and screening for extensively drug-resistant TB (XDR-TB) strains.