title: Comparison of two PCR-based human papillomavirus genotyping methods.
abstract: We compared two consensus primer PCR human papillomavirus (HPV) genotyping methods for the detection of individual HPV genotypes and carcinogenic HPV genotypes as a group, using a stratified sample of enrollment cervical specimens from sexually active women participating in the NCI/Costa Rica HPV16/18 Vaccine Efficacy Trial. For the SPF(10) method, DNA was extracted from 0.1% of the cervical specimen by using a MagNA Pure LC instrument, a 65-bp region of the HPV L1 gene was targeted for PCR amplification by using SPF(10) primers, and 25 genotypes were detected by reverse-line blot hybridization of the amplicons. For the Linear Array (LA) method, DNA was extracted from 0.5% of the cervical specimen by using an MDx robot, a 450-bp region of the HPV L1 gene was targeted for PCR amplification by using PGMY09/11 L1 primers, and 37 genotypes were detected by reverse-line blot hybridization of the amplicons. Specimens (n = 1,427) for testing by the LA method were randomly selected from strata defined on the basis of enrollment test results from the SPF(10) method, cytology, and Hybrid Capture 2. LA results were extrapolated to the trial cohort (n = 5,659). The LA and SPF(10) methods detected 21 genotypes in common; HPV16, -18, -31, -33, -35, -39, -45, -51, -52, -56, -58, -59, -66, -68, and -73 were considered the carcinogenic HPV genotypes. There was no difference in the overall results for grouped detection of carcinogenic HPV by the SPF(10) and LA methods (35.3% versus 35.9%, respectively; P = 0.5), with a 91.8% overall agreement and a kappa value of 0.82. In comparisons of individual HPV genotypes, the LA method detected significantly more HPV16, HPV18, HPV39, HPV58, HPV59, HPV66, and HPV68/73 and less HPV31 and HPV52 than the SPF(10) method; inclusion of genotype-specific testing for HPV16 and HPV18 for those specimens testing positive for HPV by the SPF(10) method but for which no individual HPV genotype was detected abrogated any differences between the LA and SPF(10) methods. The LA method detected more carcinogenic-HPV-genotype infections per specimen than the SPF(10) method (P < 0.001). In conclusion, the LA method and the SPF(10) method with HPV16 and HPV18 genotype-specific detection among ungenotyped HPV-positive specimens were comparable for detection of HPV16 and HPV18, the two HPV genotypes targeted by current prophylactic HPV vaccines. Both approaches are suitable for monitoring the impact of HPV16/18 vaccines in clinical trials.
authors: Division of Cancer Epidemiology and Genetics, National Cancer Institute, 6120 Executive Blvd., Room 5004, EPS MSC 7234, Bethesda, MD 20892-7234. castlep@mail.nih.gov.
authors: Philip E
authors: PE
authors: Castle
authors: Carolina
authors: C
authors: Porras
authors: Wim G
authors: WG
authors: Quint
authors: Ana Cecilia
authors: AC
authors: Rodriguez
authors: Mark
authors: M
authors: Schiffman
authors: Patti E
authors: PE
authors: Gravitt
authors: Paula
authors: P
authors: González
authors: Hormuzd A
authors: HA
authors: Katki
authors: Sandra
authors: S
authors: Silva
authors: Enrique
authors: E
authors: Freer
authors: Leen-Jan
authors: LJ
authors: Van Doorn
authors: Silvia
authors: S
authors: Jiménez
authors: Rolando
authors: R
authors: Herrero
authors: Allan
authors: A
authors: Hildesheim
mesh_terms: D000293:Adolescent; D000328:Adult; D002584:Cervix Uteri / Q000821:virology; D003364:Costa Rica; D004279:DNA, Viral / Q000235:genetics / Q000302:isolation & purification; D005260:Female; D006801:Humans; D027383:Papillomaviridae / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification*; D030361:Papillomavirus Infections / Q000175:diagnosis* / Q000821:virology*; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity
chemical_list: D004279:DNA, Viral