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

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60bp；其敏感度为10拷贝／μ1。将FQPCR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：FQPCR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。

Experimental investigation and clinical application of fluorescent quantitative Polymerase chain reaction in detection of Haemophilus ducreyi

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. [