title: PCR testing of pooled longitudinally collected cervical specimens of women to increase the efficiency of studying human papillomavirus infection.
abstract: In large active cohort studies of women investigating human papillomavirus (HPV) and cervical neoplasia, many women will be HPV-negative at all time points and testing of all their cervical specimens is an inefficient use of laboratory resources. The aim of this pilot study was to evaluate whether pooling cervical specimens from the same woman might provide a useful pretest of specimens from women unlikely to have high-grade cervical neoplasia or significant HPV exposure. We selected women (n = 187) participating in the Guanacaste Project for whom we already had HPV testing data on all their specimens from multiple visits (median = 8 visits), who were HPV DNA-negative at enrollment and at their 5- to 7-year exit from the cohort, and had no evidence of high-grade cervical neoplasia. Equal aliquots of cervical specimens from these women were pooled to create a proportional pooled specimen. Aliquots of pooled specimens were tested in a masked fashion by MY09/11 L1 consensus primer PCR. Second aliquots of some pooled specimens (n = 83) were included to assess the reliability of pooled testing. Results were compared with the predicted (expected) results based on the obtained test results of the individual specimens collected at interim visits. There was good overall agreement between observed and expected HPV DNA positivity, with a kappa of 0.63 [95% confidence interval (95% CI), 0.51-0.75] and a percent agreement of 83.4% (95% CI, 77.3-88.5%) although the HPV DNA positivity in the pooled specimen was less than expected (P = 0.001). The agreement between observed and expected HPV DNA positivity was related to the number of aliquots pooled, suggesting that positivity was related to viral genome concentrations. The kappa and percent agreement for intra-batch reliability of testing pooled specimens were 0.68 (95% CI, 0.53-0.84) and 84.3% (95% CI, 74.7-91.4%), respectively. We conclude that pooling specimens and testing by PCR may be useful for discriminating HPV DNA-positive from completely negative specimen sets in women who are likely to have been HPV DNA-negative.>
authors.affiliation: Division of Cancer Epidemiology and Genetics, National Cancer Institute, Room 7074, 6120 Executive Boulevard, EPS MSC 7234, Bethesda, MD 20892-7234, USA. castlep@mail.nih.gov
authors.forename: Philip E
authors.initials: PE
authors.lastname: Castle
authors.forename: Mark
authors.initials: M
authors.lastname: Schiffman
authors.forename: Rolando
authors.initials: R
authors.lastname: Herrero
authors.forename: Allan
authors.initials: A
authors.lastname: Hildesheim
authors.forename: Ana-Cecilia
authors.initials: AC
authors.lastname: Rodriguez
authors.forename: M Concepcion
authors.initials: MC
authors.lastname: Bratti
authors.forename: Sholom
authors.initials: S
authors.lastname: Wacholder
authors.forename: Hortense
authors.initials: H
authors.lastname: Kendal
authors.forename: Anne M
authors.initials: AM
authors.lastname: Breheny
authors.forename: Andrew
authors.initials: A
authors.lastname: Prior
authors.forename: Ruth
authors.initials: R
authors.lastname: Pfeiffer
authors.forename: Robert D
authors.initials: RD
authors.lastname: Burk
mesh_terms: D005260:Female; D006801:Humans; D008137:Longitudinal Studies; D027383:Papillomaviridae / Q000302:isolation & purification*; D030361:Papillomavirus Infections / Q000150:complications* / Q000175:diagnosis*; D010865:Pilot Projects; D016133:Polymerase Chain Reaction / Q000379:methods*; D015203:Reproducibility of Results; D002583:Uterine Cervical Neoplasms / Q000821:virology*; D002578:Uterine Cervical Dysplasia / Q000821:virology*