Gui Jing, Wang Feng, Luo Dao-quan, Wang Hong-fu, Peng Yi, Zhang Li
Department of Pathogenic Laboratory, Shenzhen Center for Chronic Disease Control, Shenzhen 518020, China.
Zhonghua Jie He He Hu Xi Za Zhi. 2010 Apr;33(4):276-9.
To study the molecular characterization of drug-resistant and extensive-drug resistant isolates of Mycobacterium tuberculosis (MTB) in Shenzhen of China during 2007 - 2008.
According to the standards of WHO and 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, rrs((1)), gyrA/B, rrs((2))) in these MTB isolates were identified by PCR, followed by DNA sequencing of the purified PCR products.
A total of 123 isolates were collected. Seventy-three isolates were drug resistant, and 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 44/52, 44/47, 28/41, 11/11, and 11/18, respectively. For katG gene, the mutation detected was S315T or S315N. For rpoB, the most frequently found changes were S531L (30/44) and H526D (9/44) or H526R (1/44). For the reported mutations related with streptomycin-resistant strains, K43R and K88Q were found in the rpsL locus, and 491C-->T and 513A-->C were found in the rrs gene. For gyrA, all gyrA mutations were clustered in codons 90, 91, and 94 apart from the S95T that was natural polymorphism, accounting for 9/11 of the ofloxacin-resistant isolates, and condon 91 was the most frequently mutated. No mutations were found in gyrB. The most frequent substitutions were 1400A-->G (9/11) and 1483G-->T (2/11) in a specific region of the rrs gene related with kanamycin-resistant strains. No mutations except S95T of gyrA was detected in the drug- susceptible isolates.
The mutation characterization of drug-resistant and drug- susceptible isolates of MTB was shown to vary according to geographic regions, and these findings can help understanding the molecular characterization and establishing a novel method for the detection of drug-resistant MTB.
研究2007 - 2008年中国深圳结核分枝杆菌（MTB）耐药及广泛耐药菌株的分子特征。
按照世界卫生组织（WHO）和国际防痨和肺部疾病联盟（IUATLD）的标准，采用1%比例法在罗 - 琴培养基上对异烟肼、利福平、链霉素、氧氟沙星和卡那霉素进行药物敏感性试验（DST），收集136株MTB菌株。通过聚合酶链反应（PCR）鉴定这些MTB分离株相应耐药基因（rpoB、katG、rpsL、rrs((1))、gyrA/B、rrs((2))）中的基因突变，随后对纯化的PCR产物进行DNA测序。
共收集到123株分离株。73株耐药，50株敏感。在对异烟肼、利福平、链霉素、氧氟沙星和卡那霉素耐药的分离株中，各基因发生突变的分离株比例分别为44/52、44/47、28/41、11/11和11/18。对于katG基因，检测到的突变是S315T或S315N。对于rpoB基因，最常见的变化是S531L（30/44）和H526D（9/44）或H526R（1/44）。对于报道的与链霉素耐药菌株相关的突变，在rpsL位点发现K43R和K88Q，在rrs基因中发现491C→T和513A→C。对于gyrA基因，除了天然多态性的S95T外，所有gyrA突变都集中在密码子90、91和94，占氧氟沙星耐药分离株的9/11，密码子91是最常发生突变的。gyrB基因未发现突变。在与卡那霉素耐药菌株相关的rrs基因特定区域，最常见的替换是1400A→G（9/11）和1483G→T（2/11）。在药物敏感分离株中，除了gyrA的S95T外未检测到其他突变。
MTB耐药和敏感分离株的突变特征因地理区域而异，这些发现有助于了解分子特征并建立一种检测耐药MTB的新方法。
