荧光定量聚合酶链反应检测杜克雷嗜血杆菌的实验研究及临床应用  

Experimental investigation and clinical application of fluores cent quantitative poly-merase chain reaction in detection of Haemop hilus ducreyi

作　　者：朱慧兰[1] 胡斌[2] 赖维[3] 曹文苓[1] 武明昌[1] 颜景兰[1] 

机构地区：[1]广州市皮肤病防治所性病科,510095 [2]中山大学达安基因诊断中心 [3]中山大学附属三院皮肤科

出　　处：《中国抗感染化疗杂志》2004年第6期340-342,共3页Chinese Journal of Infection and Chemotherapy

基　　金：广东省科技厅重点攻关项目 (编号 :2 0 0 3C 3 42 0 3 )

摘　　要：目的 :建立荧光定量聚合酶链反应 (FQ PCR)检测杜克雷嗜血杆菌 (HD)的方法 ,并作临床应用评价。方法　根据HD的基因库序列设计引物对 2株参考株进行扩增 ,证实其特异性 ;再以 10倍稀释不同浓度模板扩增测定其敏感度 ;同时检测 76份生殖器溃疡处分泌物标本 ,每份标本同时作HD涂片和培养。结果 :2株不同来源的HD参考株均出现特异性扩增片段 ,扩增片段大小为 6 0bp ;其敏感度为 10拷贝 /μ1。将FQ PCR用于检测 76例HD培养阴性的生殖器溃疡分泌物标本 ,结果4例阳性。结论 :FQ PCR是一种高灵敏度、高特异性和高精确性的检测生殖器溃疡标本中HD的实验诊断方法 ,可用于临床诊断HD感染。Objective:To develop a method of fluorescent qu antitative polymerase chain reaction (FQ-PCR) for the detection of Haemophilu s ducreyi from genital ulcers among patients in STD clinics and evaluate the c linical use of FQ-PCR for the diagnosis of chancroid. Methods:Primers were designed with reference to the DNA sequence of H. ducreyi. T he specificity of the primers was confirmed in two reference strains of H. duc reyi. The template was diluted ten-fold serially to test the sensitivity of F Q-PCR. The developed method was used to test 76 samples from genital ulcers. Ea ch sample was examined by smear and culture simultaneously. Results: Specific 60 bp product was produced from the two reference strains of H. ducreyi by FQ-PCR. The sensitivity of FQ-PCR was 10 copies per microlit er. Four of the 76 clinical samples were positive for H. ducreyi by FQ-PCR. Conclusions: FQ-PCR is a highly sensitive, specific and acc u rate laboratory method to detect H. ducreyi in genital ulcers. It could be u seful for the diagnosis of chancroid. [

关 键 词：荧光定量聚合酶链反应 杜克雷嗜血杆菌 诊断 

分 类 号：R446.5[医药卫生—诊断学]