Comparison and evaluation of three different molecular methods for detection of human Betapapillomaviruses in skin biopsies from patients with nonmelanoma skin cancer and precancerous lesions

Abstract

Betapapillomaviruses have been linked to the development of nonmelanoma skin cancers. A great diversity of these viruses in skin specimens requires the use of sensitive and reliable detection methods. There are currently no standardized assays for diagnostic purposes. A combination of several molecular methods has great practical significance and gives the opportunity to broaden the spectrum of detected Beta-HPV types. In the present study, different molecular methods for Beta-HPVs detection and genotyping were used: PCRs with different sets of primers, PCR followed by reverse hybridization and direct sequencing of PCR amplimers; all performed in skin biopsies from lesions and perilesional healthy area of 118 patients with NMSC or precancerous lesions. Beta-HPVs were detected in 41% of 261 biopsies examined. The RHA for 25 types of Beta-HPVs showed a significantly higher sensitivity than PCR-based methods and allowed to detect 172 genotypes in 86 samples, including 39 with multiple infections. The most frequently identified types were HPV23, HPV24 and HPV93. HPV5 and HPV8, considered high-risk carcinogen types, were detected only in a small percentage of samples. Direct sequencing confirmed the presence of Beta-HPV genotypes from outside of RHA panel in the analysed biopsies. This allowed detecting thirty-two additional genotypes in 5 samples, that were positive only in RHA with the universal probe, which failed to identify the virus genotypes. Our findings confirmed the need to apply different methods to detect Beta-HPV infections.