Evaluation of Sensitivity of Molecular Methods for Detection of Rifampin-Resistant Strains Amongst Drug-resistant Mycobacterium tuberculosis Isolates

Resistance to rifampin in multidrug-resistant M. tuberculosis (MDR-TB) is one of the major threats to the global public health system.
The aim of the present study was to explore the molecular characterization of resistance against rifampin amongst multidrug-resistant (MDR) and extensively drug-resistant (XDR) isolates obtained from patients.
In this study, we used XDR (n = 6), MDR (n = 9) and rifampin sensitive (n = 39) isolates whose drug susceptibility patterns were identified by proportion method. A simple single-step multiplex-allele specific-polymerase chain reaction (MAS-PCR) assay was optimized to detect the three most frequent mutations in rifampin resistance-determining region (RRDR) fragment of rpoB gene. Additionally, single-strand conformational polymorphism (SSCP)-PCR and Sequencing were utilized.
Out of nine MDR isolates, five were detected by MAS-PCR as rifampin resistant (sensitivity; 55.5%). In comparison with the proportion method, the sensitivity of SSCP for XDR, MDR and rifampin sensitive isolates were 83.3%, 77.7% and 97.4%, respectively. The DNA sequencing revealed that three of the 6 XDR isolates had several silent mutations, one isolate had one silent mutation, one strain had no mutations and only one isolate had mutations at codon 531.
In sum, although the molecular methods are rapid, they are not able to identify resistance against rifampin efficiently.