Guibinga G H, Coutlée F, Kessous A, Hankins C, Lapointe N, Richer G, Tousignant J
Département de Microbiologie-Immunologie, Université de Montréal, Québec, Canada.
J Clin Microbiol. 1996 Jul;34(7):1654-9. doi: 10.1128/JCM.34.7.1654-1659.1996.
A PCR assay for the sensitive detection of a transforming fragment of herpes simplex virus type 2 (HSV-2) was developed. Oligonucleotide primers were selected in Xho-2, a transforming region of the BglII N fragment of HSV-2. The assay reached a sensitivity endpoint of 10 copies of the Xho-2 subfragment and did not show cross-reactivity with other herpesviruses, including HSV-1. All 42 HSV-2 isolates scored positive in the assay. The Xho-2 PCR assay was evaluated with 216 clinical specimens and the results were compared with those of cell culture. The best protocol for processing specimens contained in viral transport medium included a centrifugation step followed by cell lysis. Of the 107 specimens positive for HSV-2 by culture, 105 were PCR positive (sensitivity, 98.1%). For one of the two falsely negative samples, beta-globin as well as sequences from the HSV-2 DNA polymerase gene could not be amplified. The other sample scored positive in both of these reactions but was indeterminate in duplicate tests by Xho-2 PCR. Two of 109 HSV-2 culture-negative specimens tested positive in the PCR assay (specificity, 98.2%). The latter two samples tested positive in a PCR test for the HSV-2 DNA polymerase gene. This novel tool was shown to be sensitive and specific for HSV-2 sequences and should allow for the investigation of the role of HSV-2 in genital cancers.
开发了一种用于灵敏检测2型单纯疱疹病毒(HSV - 2)转化片段的聚合酶链反应(PCR)检测方法。在HSV - 2的BglII N片段的转化区域Xho - 2中选择了寡核苷酸引物。该检测方法的灵敏度终点为Xho - 2亚片段的10个拷贝,并且与包括HSV - 1在内的其他疱疹病毒没有交叉反应。所有42株HSV - 2分离株在该检测中均呈阳性。用216份临床标本对Xho - 2 PCR检测方法进行了评估,并将结果与细胞培养结果进行了比较。处理病毒运输培养基中所含标本的最佳方案包括离心步骤,然后进行细胞裂解。在107份经培养检测为HSV - 2阳性的标本中,105份PCR检测呈阳性(灵敏度为98.1%)。对于两份假阴性样本中的一份,β - 珠蛋白以及HSV - 2 DNA聚合酶基因的序列均无法扩增。另一份样本在这两个反应中均呈阳性,但在Xho - 2 PCR的重复检测中结果不确定。109份HSV - 2培养阴性标本中有两份在PCR检测中呈阳性(特异性为98.2%)。后两份样本在HSV - 2 DNA聚合酶基因的PCR检测中呈阳性。这种新工具对HSV - 2序列显示出灵敏性和特异性,应该有助于研究HSV - 2在生殖器癌症中的作用。