Detection of human papillomaviruses in paired healthy skin and actinic keratosis by next generation sequencing

Actinic keratosis (AK) arises on photo-damaged skin and is considered to be the precursor lesion of cutaneous squamous cell carcinoma (cSCC). Many findings support the involvement of ß human papillomaviruses (HPVs) in cSCC, while very little is known on ? HPV types. The objective of this study was to characterize the spectrum of PV types in healthy skin (HS) and AK samples of the same immunocompetent individuals using next-generation sequencing (NGS). Viral DNA of 244 AK and 242 HS specimens were amplified by PCR using two different sets of primers (FAP59/64 and FAPM1). Purified amplicons were pooled and sequenced using NGS. The study resulted in the identification of a large number of known ß and ? PV types. In addition, 27 putative novel ß and 16 ? and 4 unclassified PVs were isolated. Only HPV types of species ?-1 (e.g. HPV4) appeared to be strongly enriched in AK versus HS. The NGS analysis revealed that a large spectrum of known and novel PVs is present in HS and AK. The evidence that species ?-1 HPV types appear to be enriched in AK in comparison to HS warrants further biological and epidemiological studies to evaluate their role in the development of skin (pre)cancerous lesions.

光化性角化病（Actinic keratosis, AK）好发于光损伤皮肤，被认为是皮肤鳞状细胞癌（cutaneous squamous cell carcinoma, cSCC）的前驱病变。诸多研究证实β型人乳头瘤病毒（human papillomaviruses, HPVs）与皮肤鳞状细胞癌的发生相关，但目前对？型人乳头瘤病毒的了解仍十分有限。本研究旨在利用下一代测序（next-generation sequencing, NGS）技术，对同一免疫功能正常个体的健康皮肤（healthy skin, HS）及光化性角化病样本中的乳头瘤病毒（PV）型别谱进行表征。研究人员采用两套不同引物（FAP59/64与FAPM1）通过聚合酶链式反应（PCR）扩增了244份AK样本与242份HS样本的病毒DNA。将纯化后的扩增产物混合后，通过下一代测序进行测序。本研究共鉴定出大量已知的β型与？型乳头瘤病毒型别。此外，还分离得到27种潜在新型β型乳头瘤病毒、16种？型乳头瘤病毒以及4种未分类乳头瘤病毒。仅？-1种属的人乳头瘤病毒（如HPV4）在AK样本中的丰度显著高于HS样本。下一代测序分析结果显示，健康皮肤与光化性角化病样本中均存在广泛的已知及新型乳头瘤病毒。相较于健康皮肤，？-1种属人乳头瘤病毒在光化性角化病样本中富集这一发现，为进一步开展生物学与流行病学研究以评估其在皮肤（前驱）癌性病变发生中的作用提供了依据。