Determination of Causative Human Papillomavirus Type in Tissue Specimens of Common Warts Based on Estimated Viral Loads.

Background: Assessment of human papillomavirus (HPV) type-specific viral load (VL) is a valid tool for determining the etiology of HPV-related skin tumors, especially when more than one HPV type is detected within one lesion. Methods: The causative HPV type was determined in 185 fresh-frozen tissue specimens of histologically confirmed common warts (CWs) collected from 121 immunocompetent patients. All tissues were tested using the type-specific quantitative real-time polymerase chain reactions (PCR) for the most common wart-associated Alpha-PV (HPV2/27/57) and Mu-PV types (HPV1/63/204). The presence of 23 additional low-risk HPVs was evaluated using a conventional wide-spectrum PCR. Results: HPV DNA was detected in 176/185 (95.1%) CWs and multiple HPV types in 71/185 (38.4%) lesions. Using the VL approach and a robust cutoff of one viral copy/cell established in this study, HPV2/27/57 were determined as causative agents in 41/53 (77.3%) and 53/71 (74.7%) CWs with single and multiple HPVs, respectively. Conclusions: CWs are mostly etiologically associated with HPV2/27/57 and only rarely with HPV1. In the majority of CWs containing multiple HPVs, a single HPV type was present in high concentration, indicating etiological association. No significant differences in VLs of lesion-causing HPV types in CWs containing single or multiple HPVs were found.

Molecular characterization, tissue tropism, and genetic variability of the novel Mupapillomavirus type HPV204 and phylogenetically related types HPV1 and HPV63.

HPV204 is the only newly identified Mupapillomavirus (Mu-PV) type in more than a decade. To comprehensively characterize HPV204, we performed a detailed molecular analysis of the viral genome and evaluated its clinical relevance in comparison to the other Mu-PVs, HPV1 and HPV63. The 7,227-bp long genome of HPV204 exhibits typical genomic organization of Mu-PVs with eight open reading frames (ORFs) (E6, E7, E1, E2, E8, E4, L2, and L1). We developed three type-specific quantitative real-time PCRs and used them to test a representative collection (n = 1,006) of various HPV-associated benign and malignant neoplasms, as well as samples of clinically normal cutaneous, mucosal, and mucocutaneous origins. HPV204, HPV1, and HPV63 were detected in 1.1%, 2.7%, and 1.9% of samples tested, respectively, and were present in skin and mucosa, suggesting dual tissue tropism of all Mu-PVs. To evaluate the etiological role of Mu-PVs in the development of HPV-associated neoplasms, Mu-PV viral loads per single cell were estimated. HPV1 and HPV63 were present in high viral copy numbers in 3/43 and 1/43 cutaneous warts, respectively, and were identified as the most likely causative agents of these warts. HPV204 viral load was extremely low in a single HPV204-positive cutaneous wart (7.4 × 10-7 viral copies/cell). Hence, etiological association between HPV204 and the development of cutaneous warts could not be established. To the best of our knowledge, this is the first study to evaluate the genetic variability of Mu-PVs by sequencing complete LCR genomic regions of HPV204, HPV1, and HPV63. We detected several nucleotide substitutions and deletions within the LCR genomic regions of Mu-PVs and identified two genetic variants of HPV204 and HPV63 and five genetic variants of HPV1.

Analysis of the genetic diversity and phylogenetic relationships of putative human papillomavirus types.

More than 170 human papillomavirus (HPV) types have been completely sequenced, curated and divided into five genera: Alphapapillomavirus, Betapapillomavirus, Gammapapillomavirus, Mupapillomavirus and Nupapillomavirus. With the application of PCR methods, hundreds of putative novel HPV types have been identified as PCR amplicons in mucosa and skin. However, at present there are no studies reporting a systematic search of the currently known L1 amplicons and their phylogenetic relationships. This survey revealed the existence of at least 202 different putative HPV types that are pending for full-genome characterization: five alphapapillomaviruses, 37 betapapillomaviruses, 159 gammapapillomaviruses and one mupapillomavirus. All potential viruses of the genera Alphapapillomavirus and Betapapillomavirus were grouped in the defined species, while 59 putative gammapapillomaviruses types were segregated in 21 unidentified putative species. These data highlight the need for progress in the identification of additional taxa of the family Papillomaviridae in order to elucidate the diversity, evolution and medical implications of these viruses.