Lai J-Y, Doyle R J, Bluhm J M, Johnson J C
Department of Dermatology, Mayo Clinic College of Medicine, Rochester, MN 55905, United States.
J Clin Virol. 2006 Apr;35(4):435-41. doi: 10.1016/j.jcv.2005.11.012. Epub 2006 Jan 18.
Plantaris verrucae are a common diagnosis in childhood, consume a significant amount of health-care resources, have many painful treatment options and many recurrences.
The objective of this study was to design and test a single site-anchored, multiplexed and expandable PCR assay for common types of cutaneous HPVs.
Common forward and unique reverse primers were selected from the E2 open reading frames of five cutaneous HPV genotypes. These were analyzed for sensitivity and selectivity using pHPV plasmids and several control DNAs in an optimized multiplexed assay. This standardized assay was used to analyze human verruca plantaris tissue for genome type and to evaluate the effect of a commonly used treatment protocol.
A sensitive, multiplexed PCR assay for human cutaneous HPV genotypes 1a, 2a and 4 was developed. Specific-unique primers and a consensus anchor primer were selected within the HPV E2 region to produce amplicons varying by greater than 100bp. In analytical sensitivity studies, fewer than 100 genome copies of HPV1a and 2a were detected, and fewer than 1000 copies of HPV4 were detected. The multiplexed assay did not amplify regions of human placenta, calf thymus, CaSki or SiHa DNA and E. coli, pBR322 or non-HPV virus DNAs. In combination with a forensic DNA extraction procedure, the multiplexed HPV assay detected and identified HPV types in 23 of 51 (45%) deep plantaris verrucae. Two patients were found with two different genotypes in single deep plantaris verruca. Detection of the HPV genome was followed as a function of tissue ablation and Mediplast treatment in one patient. In healing tissue, the genome content was reduced but had not totally disappeared.
The multiplexed HPV assay can be used to determine genotype prevalence that may correlate with treatment efficacy.
跖疣是儿童期的常见诊断，消耗大量医疗资源，有多种痛苦的治疗选择且复发率高。
本研究的目的是设计并测试一种针对常见皮肤型人乳头瘤病毒（HPV）的单位点锚定、多重且可扩展的聚合酶链反应（PCR）检测方法。
从五种皮肤HPV基因型的E2开放阅读框中选择通用正向引物和独特反向引物。在优化的多重检测中，使用人乳头瘤病毒（pHPV）质粒和几种对照DNA对其进行敏感性和选择性分析。该标准化检测用于分析人跖疣组织的基因组类型，并评估常用治疗方案的效果。
开发了一种针对人皮肤HPV基因型1a、2a和4的敏感多重PCR检测方法。在HPV E2区域内选择了特异性独特引物和共有锚定引物，以产生长度相差大于100bp的扩增子。在分析敏感性研究中，检测到的HPV1a和2a基因组拷贝数少于100个，HPV4的拷贝数少于1000个。多重检测未扩增人胎盘、小牛胸腺、CaSki或SiHa DNA以及大肠杆菌、pBR322或非HPV病毒DNA的区域。结合法医DNA提取程序，多重HPV检测在51例（45%）深部跖疣中的23例中检测并鉴定出HPV类型。在单个深部跖疣中发现两名患者感染了两种不同的基因型。在一名患者中，随着组织消融和Mediplast治疗跟踪检测HPV基因组。在愈合组织中，基因组含量减少但并未完全消失。
多重HPV检测可用于确定可能与治疗效果相关的基因型流行情况。
