Rapid detection of drug resistance in mycobacterium tuberculosis using Tag Array chip

Abstract

Objective To evaluate the sensitivity and specificity of Tag Array in detecting the clinical isolates of bone and joint tuberculosis. Methods Twenty-four strains of Mycobacterium tuberculosis having sequence differences in extracted plasmids of mutant strains. The plasmid was diluted into different concentrations, and then multiplex PCR amplification was performed to analyze the sensitivity and specificity of the chip system. A total of 187 clinical isolates were collected from patients with bone and joint tuberculosis. Among them, there were 50 strains of sensitive bacteria, and 137 strains of drug-resistant. Designed, optimized and prepared chip inspection system, sequencing and phenotypic drug susceptibility results were analyzed to evaluate the sensitivity and specificity of the gene chip. Results The mutants in accordance with sequencing result in the sets with 1×103copy/μl of template concentration and above; while 29 sets showed false negative result in the sets with 1×102 copy/μl. Among the 126 rifampicin phenotype drug resistance strains, 119 strains appeared mutation in rpoB gene. In the rifampicin phenotype sensitivity strains, 9 strains of rpoB gene showed mutation, the sensitivity was 94.40%, and specificity was 86.76%. Among the 118 isoniazid phenotype drug resistance strains, 109 strains appeared mutation in relevant drug resistance gene locus, the sensitivity was 92.37%, and specificity was 81.16%. Among the 44 ethambutol phenotype drug resistance strains, 27 strains appeared mutation in locus embB306, in the ethambutol phenotype sensitivity strains, the mutation in locus embB306 was detected in 6 strains, the sensitivity was 61.36%, and specificity was 95.80%. Among the 102 levofloxacin phenotype drug resistance strains, 81 strains appeared mutation in gyrA gene, 4 of the sensitivity strains were detected mutation in gyrA gene, the sensitivity was 79.41%, and specificity was 95.29%. Among the 112 streptomycin phenotype drug resistance strains, the chip detected 101 strains appeared mutation, the sensitivity of chip detection was 90.17%, and specificity was 84.00%. Since amikacin, capreomycin and kanamycin are in cross drug resistance, among the 67 phenotype drug resistance strains, 52 strains appeared mutation in relevant drug resistance gene locus, its sensitivity was 79.10% and specificity was 90.83%. Conclusion Tag Array chip can achieve rapid, accurate and high-throughput detection of tuberculosis resistance, which has important clinical significance and feasibility. Key words: Extensively drug-resistant tuberculosis; Genetic testing; Sensitivity and specificity