title: Comparison of Two Widely Used Human Papillomavirus Detection and Genotyping Methods, GP5+/6+-Based PCR Followed by Reverse Line Blot Hybridization and Multiplex Type-Specific E7-Based PCR.
abstract: GP5+/6+-based PCR followed by reverse line blot hybridization (GP5+/6+RLB) and multiplex type-specific PCR (E7-MPG) are two human papillomavirus (HPV) genotyping methodologies widely applied in epidemiological research. We investigated their relative analytical performance in 4,662 samples derived from five studies in Bhutan, Rwanda, and Mongolia coordinated by the International Agency for Research on Cancer (IARC). A total of 630 samples were positive by E7-MPG only (13.5%), 24 were positive by GP5+/6+RLB only (0.5%), and 1,014 were positive (21.8%) by both methods. Ratios of HPV type-specific positivity of the two tests (E7-MPG:GP5+/6+RLB ratio) were calculated among 1,668 samples that were HPV positive by one or both tests. E7-MPG:GP5+/6+RLB ratios were >1 for all types and highly reproducible across populations and sample types. E7-MPG:GP5+/6+RLB ratios were highest for HPV53 (7.5) and HPV68 (7.1). HPV16 (1.6) and HPV18 (1.7) had lower than average E7-MPG:GP5+/6+RLB ratios. Among E7-MPG positive infections, median mean fluorescence intensity (MFI; a semiquantitative measure of viral load) tended to be higher among samples positive for the same virus type by GP5+/6+RLB than for those negative for the same type by GP5+/6+RLB. Exceptions, however, included HPV53, -59, and -82, for which the chances of being undetected by GP5+/6+RLB appeared to be MFI independent. Furthermore, the probability of detecting an additional type by E7-MPG was higher when another type was already detected by GP5+/6+RLB, suggesting the existence of masking effects due to competition for GP5+/6+ PCR primers. In conclusion, this analysis is not an evaluation of clinical performance but may inform choices for HPV genotyping methods in epidemiological studies, when the relative merits and dangers of sensitivity versus specificity for individual types should be considered, as well as the potential to unmask nonvaccine types following HPV vaccination.
authors.affiliation: International Agency for Research on Cancer, Lyon, France clifford@iarc.fr.
authors.forename: Gary M
authors.initials: GM
authors.lastname: Clifford
authors.affiliation: International Agency for Research on Cancer, Lyon, France.
authors.forename: Salvatore
authors.initials: S
authors.lastname: Vaccarella
authors.affiliation: International Agency for Research on Cancer, Lyon, France.
authors.forename: Silvia
authors.initials: S
authors.lastname: Franceschi
authors.affiliation: International Agency for Research on Cancer, Lyon, France.
authors.forename: Vanessa
authors.initials: V
authors.lastname: Tenet
authors.affiliation: Ministry of Health of Rwanda, Kigali, Rwanda.
authors.forename: M Chantal
authors.initials: MC
authors.lastname: Umulisa
authors.affiliation: Department of Obstetrics and Gynaecology, Jigme Dorji Wangchuck National Referral Hospital, Thimphu, Bhutan.
authors.forename: Ugyen
authors.initials: U
authors.lastname: Tshomo
authors.affiliation: Infection and Cancer Program, German Cancer Research Center, Heidelberg, Germany.
authors.forename: Bolormaa
authors.initials: B
authors.lastname: Dondog
authors.affiliation: Centre for the Evaluation of Vaccination, Vaccine and Infectious Disease Institute, University of Antwerp, Antwerp, Belgium.
authors.forename: Alex
authors.initials: A
authors.lastname: Vorsters
authors.affiliation: International Agency for Research on Cancer, Lyon, France.
authors.forename: Massimo
authors.initials: M
authors.lastname: Tommasino
authors.affiliation: Department of Pathology, VU University Medical Center, Amsterdam, the Netherlands.
authors.forename: Daniëlle A M
authors.initials: DA
authors.lastname: Heideman
authors.affiliation: Department of Pathology, VU University Medical Center, Amsterdam, the Netherlands.
authors.forename: Peter J F
authors.initials: PJ
authors.lastname: Snijders
authors.affiliation: International Agency for Research on Cancer, Lyon, France.
authors.forename: Tarik
authors.initials: T
authors.lastname: Gheit
mesh_terms: D000293:Adolescent; D000328:Adult; D000368:Aged; D001631:Bhutan; D005260:Female; D005838:Genotype; D060005:Genotyping Techniques / Q000379:methods*; D006801:Humans; D008875:Middle Aged; D025202:Molecular Diagnostic Techniques / Q000379:methods*; D008986:Mongolia; D009693:Nucleic Acid Hybridization / Q000379:methods; D027383:Papillomaviridae / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification*; D030361:Papillomavirus Infections / Q000175:diagnosis* / Q000821:virology*; D016133:Polymerase Chain Reaction / Q000379:methods; D012432:Rwanda; D055815:Young Adult