Results of the Use of Biochips to Determine the Drug Resistance of Mycobacterium tuberculosis in the Novosibirsk and Tomsk Regions

摘要

To evaluate the possibility of determining rifampicin resistance using a biochip developed at the Institute of Molecular Biology of the Russian Academy of Sciences (Moscow), 272 rifampicin-resistant isolates of M. tuberculosis, isolated from tuberculosis patients in the Novosibirsk and Tomsk regions in 2000-2005, were investigated. Rifampicin resistance was determined by the method of absolute concentrations on Lowen-stein-Jensen medium; mutations in the rpoB gene were determined with the biochip and simultaneously by the SSCP method. Moreover, the variable region of the rpoB gene was sequenced in 60 strains of a randomized sample. In 86% of the cases, the results of the determination of rifampicin resistance by the method of absolute concentrations and with the biochip were in full agreement. The results differed in cases when cultures isolated from a patient contained a mixture of sensitive and resistant strains. Four main codons in which mutations can cause resistance to rifampicin were detected: S531 (76.2%), H526 (7%), D516 (5.6%), and L511 (5.6%). Resistance to isoniazid was also established in the region among 94% of the rifampicin-resistant strains, i.e., they were resistant to a number of drugs. Consequently, the biochip for the determination of rifampicin resistance can also be used to determine multiple-drug-resistant (MDR) strains. The use of the biochip to diagnose rifampicin resistance is significantly simplified if the two steps of the polymerase chain reaction (PCR) are replaced by one-step variant, in which a single-stranded labeled PCR fragment is synthesized according to a "nested PCR" scheme in one test tube.