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Journal of Clinical Microbiology, October 2008, p. 3437-3445, Vol. 46, No. 10
0095-1137/08/$08.00+0     doi:10.1128/JCM.00620-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

Comparison of Two PCR-Based Human Papillomavirus Genotyping Methods ^,

Division of Cancer Epidemiology and Genetics, National Cancer Institute, National Institutes of Health, Bethesda, Maryland,¹ Proyecto Epidemiológico Guanacaste, Fundación INCIENSA, Costa Rica,² DDL Diagnostic Laboratory, Voorburg, The Netherlands,³ Departments of Epidemiology and Molecular Microbiology and Immunology, Johns Hopkins University, Baltimore, Maryland,⁴ Universidad de Costa Rica, Costa Rica⁵

Received 1 April 2008/ Returned for modification 3 August 2008/ Accepted 15 August 2008

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 SPF₁₀ 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 SPF₁₀ 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 SPF₁₀ method, cytology, and Hybrid Capture 2. LA results were extrapolated to the trial cohort (n = 5,659). The LA and SPF₁₀ 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 SPF₁₀ 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 SPF₁₀ method; inclusion of genotype-specific testing for HPV16 and HPV18 for those specimens testing positive for HPV by the SPF₁₀ method but for which no individual HPV genotype was detected abrogated any differences between the LA and SPF₁₀ methods. The LA method detected more carcinogenic-HPV-genotype infections per specimen than the SPF₁₀ method (P < 0.001). In conclusion, the LA method and the SPF₁₀ 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.

Published ahead of print on 20 August 2008.

Supplemental material for this article may be found at http://jcm.asm.org/.