| Objective:To analyze the dynamic change trends of drug-resistant tuberculosis in Zhejiang province. To investigate the characteristics of rpoB,katG and inhA conferring rifampicin or isonazid resistance. To explore the genotypic characteristics of M. tuberculosis in Zhejiang province, and analyze the related risk factors.Methods:Three tuberculosis drug resistance surveillances were performed from1998to2008in Zhejiang province. Gene sequencing was used to detect the drug-resistance gene mutations of rpoB,katG and inhA. Spoligotyping and MIRU-VNTR were performed among Mycobacterium tuberculosis strains collected from of the surveillance of drug-resistance in2008in Zhejiang province. The genotyping data was analyzed by Bionumerics5.0, and chi-square test and logistic regression were used to analyze the relationship among genotypes, drug susceptibility profiles and demographic characteristics.Results:1. From1998-2008, the any resistance rate of Mycobacterium tuberculosis from the smear-positive pulmonary tuberculosis patients increased from21.5%to28.9%. The resistant rate of new cases raised from14.8%to26.5%, and that of retreated cases declined from59.0%to50.0%. The rate of multi-drug resitance (MDR) among all cases, new cases or retreated cases declined from9.1%to5.9%,4.4%to3.3%and34.7%to28.1%, respectively.2. In all RIF resistant strains, the percentage of MDR was76.1%, and the rate in new cases patients and retreatment pulmonary tuberculosis was68.9%and82.7%, respectively.3. A total of55MDR-TB strains were isolated during the third drug resistance surveillance. The rate of rpoB gene mutation was94.6%(52/55) by DNA sequencing,92.7%(51/55) of the strains was detected with mutations located in rifampin resistance determining region (RRDR). Mutations at531codon of rpoB gene were the most frequent (51.9%,27/52), including S53IF and S531L change. Mutation at526codon had a frequency of21.2%(11/52), including amino acid changes H526D and H526Y. The rate of katG mutation was65.5%(36/55), and the most common mutation locus was315(AGC-ACC), accounting for77.8%of all mutations. Mutations were detected in six strains with5of C15T mutations and1of G17T mutation in inhA promoter region.4. Spoligotyping showed that the Beijing family and T family were the dorminate genotypes, and the prevalence was71.6%and16.6%, respectively. Beijing family had a heterogeneous distribution in Zhejiang province, and the prevalence was higher than that of the whole China (62.2%) and the southern China (53.2%),but lower than that of the northern China (76.5%). There is no correlation between the Beijing genotype and demographic characteristics of the patients, including gender, age and occupation. The strains with a special MIRU-VNTR genotype (42436) distinguished by ETR serial loci were found has a higher percentage among MDR-TB strains than that among non-MDR group, the difference of which was statistically significant.5. The total distinguishbility (H) value of Spoligotyping for Mycobacterim tuberculosis in Zhejiang province was0.53, while the H value became to0.93for non-Beijing family genotypes, which showed a relatively high resolution. When the21MIRU-VNTR loci were used to genotype the strains, the h values of21loci varied from0.1-0.88. The h value and clustering rate of5loci were0.9554and78%,0.9995and8%for10loci,0.9998and3%for12loci, respectively. The18loci and21loci combinations showed the same distinguishbility and clustering rate, which was0.99998and0.6%, almost every strain was unique.Conclusion:1. Only76.1%of rifampicin-resistant strains were identified as multidrug-resistant strains in Zhejiang Province, suggesting that RIF resistance may be not suitable to use as an indicator to screen multidrug-resistant tuberculosis in Zhejiang.2. There was no significant difference among the distribution of the MDR-TB gene mutations between Zhejiang Province and other regions, and several rare mutations were also observed.3. Beijing family genotype is one of the most dominate gentoype in Zhejiang province, with a variation in regional distribution, but no correlation with drug resistance and epidemiological characteristics of TB patients was found.4. Clustering rate of Mycobacterium tuberculosis is low, indicating a high rate of divergence. One genotype is42436which cluster by5ETR serial loci could be used to monitor the MDR-TB.In conclusion, our findings lay the foundation for the further molecular epidemiological studies of Mycobacterium tuberculosis in Zhejiang Province. In addition, our data will also provide scientific insights for generating effective TB control strategy for Zhejiang province. |
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