Early Detection of Mycobacterium Tuberculosis and Determination Rifampin-Resistance by PCR-Reverse Hybridization (Line Probe Assay) v

Park, Min Jeong; Kim, Hyoun Tae; Lee, Kyu Man; Lee, Kye Sook

Abstract

BACKGROUND: Recent worldwide increase of tuberculosis has been mainly caused by opportunistic infections in the patients with AIDS. Emergence of multidrug- resistant Mycobacterium tuberculosis has been also posing serious problems on the treatment of tuberculosis. The purpose of this study was to evaluate the usefulness of the PCR-reverse hybridization method (line probe assay, INNO-LIPA; Innogenetics, Belgium) for the identification of M. tuberculosis and detection of rifampicin- resistant M. tuberculosis. METHOD: Identification of 37 isolates was performed by both conventional method and line probe assay. Detection of rifampin resistance by line probe assay was based on the reaction of amplified gene products of isolates with either wild type probes (S1, S2, S3, S4, and S5) specific for rpoB gene or mutant type probes (R2, R4a, R4b, and R5) specific for the mutation sequence in the rpoB region, and the results were compared with those by the absolute concentration method.
RESULTS
All of the 37 isolates were identified as M. tuberculosis by both line probe assay and conventional method. Eight isolates susceptible to rifampin by absolute concentration method showed no mutation by line probe assay. Twenty seven of 29 isolates resistant to rifampin by absolute concentration method showed the following mutations in rpoB gene: 11, S5-R5+(loss of S5 with R5); 7, S4-R4a; 1, S4-R4b+; 4, S2-R2+; 1, S1-S2+: 1; S2-S4-; 1, S4-; and 1, R5+. The sensitivity of rifampin resistance by line probe assay was 95%.
CONCLUSION
Line probe assay is a useful tool for the early identification of M. tuberculosis as well as determination of its rifampin resistance, especially in cases of emergent need for rapid treatment for tuberculosis before getting the final of conventional antituberculous drug susceptibility test.