霍乱弧菌O139实时荧光PCR检测方法的建立及应用

目的：建立检测霍乱弧菌O139的实时荧光PCR检测方法，为霍乱弧菌0139的检测提供快速、敏感、特异的检测手段。方法：用复合探针法荧光PCR检测技术，检测0139特异基因——编码糖基转移酶的基因。结果：本方法特异性好，检测非0139肠道菌均无响应；灵敏度高，可检测低至10CFU的细菌；重复性较好，批间批内变异系数均小于3％。对采自浙江省0139霍乱弧菌感染患者的335份标本进行检测，共有133份为阳性，而采用常规细菌培养法，仅检测出90份阳性。结论：本方法的建立为霍乱的诊断提供了又一有效手段。

Development and application of real-time PCR for detection of Vibrio cholerae O139

Objective:To develop a method for rapid,sensitive and specific detection of Vibrio cholerae O139.(Method:) Using complex probe,a real-time PCR assay targeted at the sequence encoding glycosyltransferases was developed.(Results:)The assay showed good specificity,sensitivity and repeatability.All of the strains of V.cholerae O139 tested were positively detected and none of the other bacteria was detected by the assay.The sensitivity of the assay was about 10 CFU/ml in pure cultures.The coefficient of variation of intra-assay and inter-assay was less than 3%.The real-time PCR was used to screen 335 diarrheal stool specimens.All the 90 V.cholerae O139 culture-positive and 43 V.cholerae O139 culture-negative stool specimens were positive by real-time PCR,and the remaining specimens were all negative by PCR.Conclusion:The results indicated that the real-time PCR assay is a reproducible,specific and sensitive method for the detection of V.cholerae O139.