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目的探讨临床患者痰标本中分离的结核分枝杆菌rpoB基因突变与利福平耐药之间的关系。方法对115株结核分枝杆菌应用L-J药敏培养法,检测对利福平、异烟肼、乙胺丁醇、链霉素、氧氟沙星的耐药情况;根据rpoB基因序列设计包含利福平耐药决定区(RRDR)的扩增引物,PCR法进行扩增,运用生物信息学方法对耐药决定区扩增产物序列进行比对分析,对耐药株与非耐药株基因突变率进行统计学分析。结果 115株结核分枝杆菌经比例法药敏试验,RIF耐药株53株,占所有菌株的46.09%;53株RIF耐药株中39株rpoB基因存在不同位点的变异,突变率为73.58%,突变位点包括531、526、522、516、513、511,最主要的突变位点集中在531位,引起氨基酸呈Ser531Gln、Ser531Leu、Ser531Trp 3种形式改变;RIF敏感株62株,其中有2株检测到rpoB基因的突变,敏感株与耐药株突变率经McNemar统计检验,差异有统计学意义(P<0.05)。结论岳阳地区rpoB基因变异与利福平耐药呈高度相关,rpoB基因最主要的突变位点位于531位和526位。
Objective To investigate the relationship between rpoB gene mutation and rifampicin resistance in sputum from clinical patients. Methods 115 strains of Mycobacterium tuberculosis were detected by LJ antimicrobial susceptibility culture method. The resistance to rifampicin, isoniazid, ethambutol, streptomycin and ofloxacin was detected. According to the sequence of rpoB gene, FDR-PCR primers were amplified by PCR. The bioinformatics methods were used to analyze the amplification product sequences in the drug-resistance-determining region. Mutations in the resistant and non-resistant strains Rate for statistical analysis. Results 115 mycobacterium tuberculosis strains were screened by proportional method. There were 53 strains of RIF-resistant strains, accounting for 46.09% of all strains. The 39 rpoB genes of 53 RIF-resistant strains showed different site mutation with a mutation rate of 73.58 %, And the mutation sites included 531,526,522,516,513,511. The major mutation sites were concentrated at 531 positions, causing the amino acids to be Ser531Gln, Ser531Leu and Ser531Trp. There were 62 RIF sensitive strains including The mutation of rpoB gene was detected in 2 strains. The mutation rate of sensitive strains and resistant strains was tested by McNemar statistical difference (P <0.05). Conclusion The mutation of rpoB gene in Yueyang region is highly correlated with the rifampicin resistance. The most frequent mutation sites of rpoB gene are located at 531 and 526.