应用PCR-SSCP技术检测痰中结核分枝杆菌katG、rpoB、embB基因突变的研究

Detection of the mutations of katG,rpoB and embB Genes in Mycobacterium tuberculosis from sputum specimens with PCR-SSCP technique

目的应用PCR-SSCP技术检测痰样本中结核分枝杆菌katG、rpoB、embB基因突变,探索其临床应用价值。方法以结核分枝杆菌标准株H37RV为对照,应用套式PCR扩增目的基因,并使用SSCP技术直接检测100例耐药患者和10例敏感患者痰样本中结核分枝杆菌katG、rpoB、embB基因突变并进行基因测序,将SSCP结果及测序结果与细菌培养药敏结果进行比较分析。结果PCR-SSCP直接检测痰样本中的结核分枝杆菌katG基因突变的敏感性和特异性分别是55.9%和70.0%,rpoB为76.0%和90.0%,embB为46.4%和60.0%。结论PCR-SSCP操作快速、简单,敏感性和特异性较高,可用来直接检测临床痰样本中结核分枝杆菌katG、rpoB、embB基因突变。

Objective To detect the mutations of katG,rpoB and embB genes in Mycobacterium tuberculosis(MTB) from sputum specimens by PCR-SSCP method,and to evaluate the clinical value of this method.Methods 110 MTB isolates from sputum specimens were detected by PCR-SSCP and conventional drug susceptibility testing.The sequences of katG,rpoB and embB were also analyzed.Results The sensitivities of mutations of katG,rpoB and embB genes analyzed by PCR-SSCP were 55.9%,76.0% and 46.4%,respectively.Their specificities were 70.0%,90.0% and 60.0%,respectively.Conclusions PCR-SSCP technique is a rapid,simple,high sensitive and strongly specific method for detection of mutations of katG,rpoB and embB genes in MTB from sputum specimens.