Genetic and phenotypic characterization of drug-resistant Mycobacterium tuberculosis isolates in Shenzhen of China
10.3760/cma.j.issn.0254-5101.2010.05.017
- VernacularTitle:深圳地区结核分枝杆菌耐药分离株分子特征与表型特征相关性研究
- Author:
Jing GUI
;
Feng WANG
;
Jinli LI
;
Daoquan LUO
;
Litai YAO
- Publication Type:Journal Article
- Keywords:
Mycobacterium tuberculosis;
Drug resistant;
PCR;
Gene;
Molecular characterization
- From:
Chinese Journal of Microbiology and Immunology
2010;30(5):466-471
- CountryChina
- Language:Chinese
-
Abstract:
Objective To characterize the relationship between such phenotypes and the patterns of genetic mutations in the corresponding resistance genes in drug-resistant Mycobacterium tuberculosis (MTB)isolates in Shenzhen of China during 2007-2008.Methods According to standard of WHO,International Union Against Tuberculosis and Lung Disease(IUATLD),136 strains of MTB were collected by performing drug sensitivity test(DST)to isoniazid,rifampicin,streptomycin,ofloxacin and kanamycin on Lowenstein-Jensen in 1%proportion method.Genetic mutations in the corresponding resistance genes (rpoB,katG,rpsL,rrs1,gyrAB,rrs2)in these MTB isolates were identified by PCR,followed by DNA sequencing of the purified PCR products.The minimal inhibitory concentration(MIC)values of the aforementioned anti-tuberculosis drugs were determined for these MTB clinical isolates by two-fold dilution method in vitro.Results A total of 123 isolates were collected,73 isolates were drug resistant.50 isolates were drug susceptible.Among the isolates that were resistant to isoniazid,rifampicin,streptomycin.ofloxacin and kanamycin,the proportion of isolates that harboured mutations in the respective genes was 84.6%,93.6%,65.9%,100%,61.1%.For katG gene,the mutation detected were S315T or S315N.For rpoB,the most frequently found changes were S531L(30/44,68.2%)and H526D(9/44,20.5%)or H526R(1/44,2.3%).The reported mutations that K43R and KS8Q were founded in the rpsL locus and 491C→T and 513A→C were founded in the rrs1 gene related with streptomycin-resistant strains.For gyrA,all gyrA mutations were clustered in codons 90,91,and 94 apart from the S95T that was natural polymorphism.accounted for 81.1% of the ofloxacin-resistant isolates,and condon 91 was the most frequently mutated.No mutation were found in gyrB.The most frequent substitution were 1400 A→G(9/11,81.8%)and 1483 G→T(2/11,18.2%)in a specific region of the rrs2 gene related with kanamycin-resistant strains.No mutations except S95T of gyrA detected in the drug-susceptible isolates.The MIC values of clinical drug-resistant strains that the same drug-resistant group contains a different resistance phenotype are basically the same with the relevant resistance genes in the same mutation.Associated resistance mutations in different sites varied significantly with their MIC values.Conclusion The mutation characterization of drug-resistant and drug-suscep-tible isolates of MTB have been shown to vary according to geographic region,phenotypic characteristics exist difference in resistance levels due to different muntants of drug-resistant gene.
