Molecular diagnosis of extensively drug-resistant tuberculosis by reverse blot hybridization assay

Dept. of Biomedical Laboratory Science/박사[한글] [영문]Tuberculosis (TB) remains one of the leading causes of death worldwide, and emergence of drug-resistant TB has become a major public health problem. Thus, rapid and accurate identification of drug-resistant TB is critical to lead appropriate therapy and prevent the spread of drug-resistant TB. However, conventional drug susceptibility test (DST) takes several weeks to get the final determination of drug-resistance. For rapid detection of drug resistant Mycobacterium tuberculosis (MTB), reverse blot hybridization assay (REBA) was designed to determine resistance to anti-TB drugs such as rifampin (RIF), isoniazid (INH), oflaxacin (OFX), kanamycin (KM), and streptomycin (SM) by detecting mutations of rpoB for RIF, katG, inhA promoter region, ahpC promoter region for INH, gyrA, gyrB for OFX, rrs, and rpsL for KM and SM. In the clinical evaluation using DNA extracted from cultured specimens, the sensitivities and specificities of the REBA to detect any sequence alterations in targeted gene were almost 100% when the results were compared with sequence analysis. However, the sensitivities for detection of resistance were 95.9% for RIF, 74.3% for INH, 41% for OFX, 62.2% for KM, and 90.3% for SM when compared with those of drug resistance by conventional DST. On the other hand, the specificities of the assay to detect drug resistance to the drugs were ranged from 91.9% to 100%. In the clinical evaluation using DNA extracted from sputum specimens, the sensitivities for detection of RIF-, INH-, OFX-, KM-, and SM-resistance were 50%, 87.5%, 80%, 33.3%, and 81.8%, respectively. The specificities were ranged from 95.5% to 100%. Despite the low sensitivity of the REBA when applied to sputum specimens, the REBA seems to be very attractive, since it is rapid compared to the fastest DST using liquid culture. Therefore, the REBA seems useful for rapid screening of drug-resistant TB for the effective management of TB patients and for the prevention of drug-resistant MTB transmission.restrictio