DNA序列分析与PCR-SSCP分析结核分支杆菌利福平敏感性的比较

中国防痨杂志 ›› 2003, Vol. 25 ›› Issue (4): 260-263.

DNA序列分析与PCR-SSCP分析结核分支杆菌利福平敏感性的比较

黄海荣¹；何秀云²；张宗德¹；张小刚²；庄玉辉²；马玙¹；

1.北京市结核病胸部肿瘤研究所北京 101149；2.解放军第309医院北京 100091；

出版日期:2003-04-10 发布日期:2003-11-03
基金资助:
国家重大基础研究发展规划 (973 )“严重传染病防治基础研究”课题资助项目 (G1999054104)

Comparison of DNA sequencing and single-strand conformation polymorphism methods for identification of rpoB mutations in rifampin-resistant isolates of Mycobacterium tuberculosis

HUANG Hai-rong,HE Xiu-yun,ZHANG Zong-de,et al.

Beijing Tuberculosis & Thoracic Tumor Research Institute,Beijing 101149

Online:2003-04-10 Published:2003-11-03

摘要/Abstract

摘要： 目的　了解DNA序列分析与PCR-SSCP法分析利福平敏感性的价值。方法应用DNA序列分析与PCR-SSCP法检测利福平敏感的结核分支杆菌临床分离株 25株、耐药株 41株的利福平耐药相关基因rpoB基因核心区域的突变情况。结果 25株利福平敏感株DNA序列分析未检测到rpoB基因突变，41株耐利福平株中 38株发生突变，突变率为 92.7% (38/41)；25株利福平敏感的结核分支杆菌菌株PCR-SSCP带型与对照H₃₇R_v株相同，41株耐利福平结核分支杆菌菌株中38株SSCP带型不同于对照株，提示有突变存在。与DNA序列分析相比，PCR-SSCP检测准确率为 93.9% (62/66)，敏感度为 92.1% (35/38)；特异度是 96.4% (27/28)。结论 DNA序列分析对判断结核分支杆菌耐利福平非常有价值；PCR-SSCP可用于利福平耐药结核分支杆菌的初步筛选。

关键词: DNA序列分析, 聚合酶链反应-单链构象多态性, 药物耐受性, 分支杆菌,结核

Abstract: Objective To evaluate the significance of DNA sequencing and PCR-SSCP in susceptibility analysis of M.tuberculosis to rifampin.Methods Analyzed the susceptibility to refampin of 25 rifampin-sensitive and 41 rifampin-resistant clinical isolates of M.tuberculosis by DNA sequencing and PCR-SSCP.Results No mutation was found in 25 rifampin-sensitive strains by DNA sequencing.while 38 of 41 rifampin-resistant strains had mutations in rifampin resistant determination region (RRDR) of rpoB gene,the mutation rate was 92.7%(38/41);25 rifampin-sensitive strains had same PCR-SSCP gel profile with M.tuberculosis H₃₇R_v,while 38 of 41 rifampin-resistant strains showed different SSCP profile that could differentiate its from control,indicating the possible existence of sequence alterations in the region of rpoB gene.The accordance rate of DNA sequencing and PCR-SSCP was 93.9%(62/66),compared with DNA sequencing,the sensitivity of PCR-SSCP was 92.1%(35/38),the specialty of PCR-SSCP was 96.4%(27/28).Conclusion DNA sequencing is valuable in rifampin susceptibility test;PCR-SSCP is useful in rifampin resistance strains screening.

Key words: DNA sequencing, PCR-SSCP, Drug resistance, Mycobacterium tuberculosis

黄海荣,何秀云,张宗德,张小刚,庄玉辉,马玙,. DNA序列分析与PCR-SSCP分析结核分支杆菌利福平敏感性的比较[J]. 中国防痨杂志, 2003, 25(4): 260-263.

HUANG Hai-rong,HE Xiu-yun,ZHANG Zong-de,et al.. Comparison of DNA sequencing and single-strand conformation polymorphism methods for identification of rpoB mutations in rifampin-resistant isolates of Mycobacterium tuberculosis[J]. Chinese Journal of Antituberculosis, 2003, 25(4): 260-263.

参考文献

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