CAJ | 학술논문

目的研究2007-2008年深圳地区MTB耐药和广泛耐药分离株的分子特征.方法参照世界卫生组织和同际防痨与肺部疾病联盟的标准,使用改良罗氏药敏培养基,用1%比例法药敏试验,筛选出针对异烟肼、利福平、链霉素、氧氟沙星和卡那霉素5种药物耐药或敏感的临床分离株,通过PCR扩增筛选菌株的rpoB、katG、rpsL、rrs~((1))、gyrA/B和rrs~((2))基因的相关序列,运用DNAStar和Blastn进行序列分析.结果筛选出实验菌株123株,其中耐药株73株,全敏感株50株.异烟肼耐药株katG基因突变率为44/52,突变位点全部为S315T或S315N.利福平耐药株rpoB基因突变率为44/47,突变位点主要集中在S531L(30/44)、H526D(9/44)和H526R(1/44).链霉素耐药株以rpsl基因突变为主,突变位点为K43R(19/28)和KS8Q(6/28);rrs(1)基因突变较少见,仪有491C→T(2/28)和513A→C(1/28),2个基因突变率合计为28/41.氧氟沙星耐药株突变率为11/11,以gyrA基因突变为主,突变位点包括D94A(2/11)、S91P(4/11)和A90V(3/11),3个突变位点总突变率为9/11;
목적 연구2007-2008년심수지구MTB내약화엄범내약분리주적분자특정.방법 삼조세계위생조직화동제방로여폐부질병련맹적표준,사용개량라씨약민배양기,용1%비례법약민시험,사선출침대이연정、리복평、련매소、양불사성화잡나매소5충약물내약혹민감적림상분리주,통과PCR확증사선균주적rpoB、katG、rpsL、rrs~((1))、gyrA/B화rrs~((2))기인적상관서렬,운용DNAStar화Blastn진행서렬분석.결과 사선출실험균주123주,기중내약주73주,전민감주50주.이연정내약주katG기인돌변솔위44/52,돌변위점전부위S315T혹S315N.리복평내약주rpoB기인돌변솔위44/47,돌변위점주요집중재S531L(30/44)、H526D(9/44)화H526R(1/44).련매소내약주이rpsl기인돌변위주,돌변위점위K43R(19/28)화KS8Q(6/28);rrs(1)기인돌변교소견,의유491C→T(2/28)화513A→C(1/28),2개기인돌변솔합계위28/41.양불사성내약주돌변솔위11/11,이gyrA기인돌변위주,돌변위점포괄D94A(2/11)、S91P(4/11)화A90V(3/11),3개돌변위점총돌변솔위9/11;
Objective To study the molecular characterization of drug-resistant and extensive-drug resistant isolates of Mycobacterium tuberculosis (MTB) in Shenzhen of China during 2007-2008.Methods 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.Results 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.Conclusions 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.