Identification of Gene Mutation Sites of Mycobacterium Tuberculosis and the Relationship Between Isoniazid and Rifampicin Resistance in Ningbo

OBJECTIVE To elucidate the mutation characteristics of rpoB, katG and inhA genes of Mycobacterium tuberculosis in newly treated pulmonary tuberculosis patients in Ningbo, and to analyze the relationship between gene mutation and drug resistance to isoniazid and rifampicin, so as to provide scientific basis for clinical anti-tuberculosis treatment. METHODS Drug susceptibility of 2 455 clinical isolates of Mycobacterium tuberculosis was detected by 1% ratio method. DNA was extracted from drug-resistant strains and amplified by PCR. The katG, inhA and rpoB genes of drug-resistant strains were sequenced. The three gene characteristics and the association of gene mutations with single drug resistance and multi-drug resistance were analyzed. RESULTS Among 2 455 clinical isolates of Mycobacterium tuberculosis, 108 cases were found to be resistant to Mycobacterium tuberculosis, including 75 cases of single drug resistance (inh 51 cases, rfp 24 cases), the single drug resistance rate was 3.05%; 33 cases of multi-drug resistance, the multi-drug resistance rate was 1.34%. Among the 33 multi-drug resistance strains, 5 of them had mutations of inhA gene, the mutation rate was 15.15%(5/33). The mutation rate of rpoB gene at 450 locus(ser450leu) was 51.52%(17/33), the mutation rate of katG gene 315 locus(ser315thr) was 87.88%(29/33), the mutation rate of 463 locus(arg463leu) was 78.79%(26/33). Among the 24 rifampicin resistant strains, 17 of them had mutations in rpoB gene, the mutation rate was 70.83%(17/24), mainly at the 446 locus(lys446lysarg), the mutation rate was 45.83%(11/24). Among 51 isoniazid-resistant strains, 45 cases had missense mutation of katG gene, the mutation rate was 88.23%(45/51), the mutation rate of 315 locus(ser315thr) was 72.55%(37/51), the mutation rate of 463 locus(arg463leu) was 64.71%(33/51), 22 cases had inhA gene missense mutation, the mutation rate was 43.14%(22/51), the mutation rate of 145 locus(val145valgly) was 27.45%(14/51), 5 cases had rpoB gene missense, the mutation rate was 9.80%(5/51). In addition, 11 of 108 drug-resistant strains had combined mutations of rpoB, katG and inhA genes, and missense mutations occurred at most sites, the mutation rate was 10.18%(11/108). CONCLUSION The mutations of rpoB, katG and inhA genes at 450 locus(ser450leu), 315 locus(ser315thr), 446 locus(lys446lysarg), 463 locus(arg463leu), 145 locus(val145valgly) are closely related to the resistance of Mycobacterium tuberculosis to isoniazid and rifampicin in Ningbo. The main mutation sites and mutation frequencies of rpoB and inhA genes are significantly different.