title: Development of a general-primer-PCR-reverse-line-blotting system for detection of beta and gamma cutaneous human papillomaviruses.
abstract: The beta and gamma genera of papillomaviruses consist of epidermodysplasia verruciformis-related human papillomaviruses (HPVs) and phylogenetically related cutaneous HPVs. Here, we have developed a consensus primer PCR assay and reverse line blot typing system coupled thereto (referred to as beta and gamma cutaneous HPV PCR [BGC-PCR]) for detection and typing of 24 beta and gamma HPVs (HPV types 4, 5, 8, 9, 12, 14, 15, 17, 19, 20, 21, 22, 23, 24, 25, 36, 37, 38, 47, 48, 49, 50, 60, and 65). Because the HPV-specific PCR products are only 72 bp in size, the system is suitable for formalin-fixed, paraffin-embedded specimens and other samples in which the DNA is of suboptimal quality. This system was able to detect and type as little as 100 ag to 1 fg HPV DNA per reaction (depending on the HPV type) in a background of 100 ng human DNA without any cross-reactivity between the tested types. Beta and gamma HPVs were detected in DNA extracted from plucked eyebrow hairs of 31 of 34 renal transplant recipients. In addition, formalin-fixed, paraffin-embedded specimens from nonmelanoma skin tumors of renal transplant recipients (n = 25) and immunocompetent individuals (n = 15) scored BGC-PCR positive in 21 and 6 cases, respectively, with HPV type 5 (HPV5) and HPV8 being the predominant types. The data indicate that this method can be a valuable, user-friendly tool for the detection and typing of cutaneous HPV in clinical specimens and may have implications for future monitoring of vaccines or alternative treatment modalities for diseases caused by these cutaneous HPVs.
authors.affiliation: Department of Pathology, VU medical center, De Boelelaan 1117, 1081 HV Amsterdam, The Netherlands.
authors.forename: Antoinette A T P
authors.initials: AA
authors.lastname: Brink
authors.forename: Belen
authors.initials: B
authors.lastname: Lloveras
authors.forename: Ingo
authors.initials: I
authors.lastname: Nindl
authors.forename: Daniëlle A M
authors.initials: DA
authors.lastname: Heideman
authors.forename: Debbie
authors.initials: D
authors.lastname: Kramer
authors.forename: René
authors.initials: R
authors.lastname: Pol
authors.forename: M Jose
authors.initials: MJ
authors.lastname: Fuente
authors.forename: Chris J L M
authors.initials: CJ
authors.lastname: Meijer
authors.forename: Peter J F
authors.initials: PJ
authors.lastname: Snijders
mesh_terms: D001483:Base Sequence; D001706:Biopsy; D017931:DNA Primers / Q000235:genetics; D006197:Hair / Q000821:virology; D006801:Humans; D007121:Immunocompetence; D016030:Kidney Transplantation; D008969:Molecular Sequence Data; D027383:Papillomaviridae / Q000235:genetics / Q000302:isolation & purification*; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D016415:Sequence Alignment; D012867:Skin / Q000473:pathology / Q000821:virology; D014860:Warts / Q000175:diagnosis* / Q000209:etiology / Q000473:pathology / Q000821:virology
chemical_list: D017931:DNA Primers