Kidd I M, Clark D A, Bremner J A, Pillay D, Griffiths P D, Emery V C
Department of Virology, Royal Free Hospital School of Medicine, London, UK.
J Virol Methods. 1998 Jan;70(1):29-36. doi: 10.1016/s0166-0934(97)00165-1.
A multiplex polymerase chain reaction (PCR) method was developed for the simultaneous detection of human herpesviruses 6 and 7 (HHV-6; HHV-7) in clinical samples, using primers which amplify a section of the HHV-6 U67 gene and the HHV-7 homologue of the HHV-6 U42 gene. Comparison of the multiplex assay with the respective single PCR assays, using cloned HHV-6 and HHV-7 sequences as targets for amplification, showed equivalent sensitivity and specificity for the assays. To demonstrate the use of multiplex PCR for the analysis of clinical samples, serum and saliva from infants were analysed using this technique. The results showed that a clear distinction can be made between the amplicons of HHV-6 and HHV-7, without loss of sensitivity or specificity. There was complete concordance between the respective single PCR assays, and the multiplex PCR. HHV-6 amplicons derived from the multiplex PCR analysis were typed by differential AvaII restriction endonuclease digestion, in which HHV-6 variant A amplicons are cleaved but those of variant B remain undigested. These results were compared to HHV-6 variant typing by an established method, the results of which showed complete concordance between assays. It is proposed that this multiplex assay, where HHV-6 positive samples may be typed directly from the reaction products, is an efficient and cost-effective approach to the analysis of large numbers of samples to determine the epidemiological importance of HHV-6 and HHV-7.
开发了一种多重聚合酶链反应（PCR）方法，用于同时检测临床样本中的人类疱疹病毒6型和7型（HHV - 6；HHV - 7），该方法使用的引物可扩增HHV - 6 U67基因的一段序列以及HHV - 6 U42基因的HHV - 7同源序列。将多重检测方法与各自的单重PCR检测方法进行比较，以克隆的HHV - 6和HHV - 7序列作为扩增靶标，结果显示两种检测方法具有相同的灵敏度和特异性。为了证明多重PCR在临床样本分析中的应用，使用该技术对婴儿的血清和唾液进行了分析。结果表明，可以清楚地区分HHV - 6和HHV - 7的扩增子，且不会损失灵敏度或特异性。各自的单重PCR检测方法与多重PCR检测结果完全一致。通过差异AvaII限制性内切酶消化对多重PCR分析得到的HHV - 6扩增子进行分型，其中HHV - 6 A亚型扩增子可被切割，而B亚型扩增子则保持未消化状态。将这些结果与通过既定方法进行的HHV - 6亚型分型结果进行比较，结果显示两种检测方法完全一致。有人提出，这种多重检测方法可直接从反应产物中对HHV - 6阳性样本进行分型，是一种分析大量样本以确定HHV - 6和HHV - 7流行病学重要性的高效且经济有效的方法。
