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Background: Results from the Costa Rica Vaccine Trial (CVT) demonstrated partial cross-protection by the bivalent human papillomavirus (HPV) vaccine, which targets HPV-16 and HPV-18, against HPV-31, -33, and -45 infection and an increased incidence of HPV-51 infection., Methods: A study nested within the CVT intention-to-treat cohort was designed to assess high-risk HPV variant lineage-specific vaccine efficacy (VE). The 2 main end points were (1) long-term incident infections persisting for ≥2 years and/or progression to high-grade squamous intraepithelial lesions (ie, cervical intraepithelial neoplasia grade 2/3 [CIN 2/3]) and (2) incident transient infections lasting for <2 years. For efficiency, incident infections due to HPV-16, -18, -31, -33, -35, -45, and -51 resulting in persistent infection and/or CIN 2/3 were matched (ratio, 1:2) to the more-frequent transient viral infections, by HPV type. Variant lineages were determined by sequencing the upstream regulatory region and/or E6 region., Results: VEs against persistent or transient infections with HPV-16, -18, -33, -35, -45, and -51 did not differ significantly by variant lineage. As the possible exception, VEs against persistent infection and/or CIN 2/3 due to HPV-31 A/B and HPV-31C variants were -7.1% (95% confidence interval [CI], -33.9% to 0%) and 86.4% (95% CI, 65.1%-97.1%), respectively (P = .02 for test of equal VE). No difference in VE was observed by variant among transient HPV-31 infections (P = .68)., Conclusions: Overall, sequence variation at the variant level does not appear to explain partial cross-protection by the bivalent HPV vaccine., (© The Author 2015. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail journals.permissions@oup.com.)

Two trials of clinically approved human papillomavirus (HPV) vaccines, Females United to Unilaterally Reduce Endo/Ectocervical Disease (FUTURE I/II) and the Papilloma Trial Against Cancer in Young Adults (PATRICIA), reported a 22% difference in vaccine efficacy (VE) against cervical intraepithelial neoplasia grade 2 or worse in HPV-naïve subcohorts; however, serological testing methods and the HPV DNA criteria used to define HPV-unexposed women differed between the studies. We applied previously described methods to simulate these HPV-naïve subcohorts within the Costa Rica HPV16/18 Vaccine Trial and assessed how these criteria affect the estimation of VE. We applied 2 enzyme-linked immunosorbent assay (ELISA) thresholds for HPV16 and HPV18 seropositivity (8 and 7 ELISA units/mL, respectively, for PATRICIA; 54 and 65 ELISA units/mL, respectively, for FUTURE I/II (to approximate the competitive Luminex immunoassay)) and 2 criteria for HPV DNA positivity (12 oncogenic HPV types, plus HPV66 and 68/73 for PATRICIA; or plus HPV6 and 11 for FUTURE I/II). VE was computed in the 2 naïve subcohorts. Using the FUTURE I/II and PATRICIA criteria, VE estimates against cervical intraepithelial neoplasia grade 2 or worse, regardless of HPV type, were 69.0% (95% confidence interval: 40.3%, 84.9%) and 80.8% (95% confidence interval: 52.6%, 93.5%), respectively (P = 0.1). Although the application of FUTURE I/II criteria to our cohort resulted in the inclusion of more sexually experienced women, methodological differences did not fully explain the VE differences., (Published by Oxford University Press on behalf of the Johns Hopkins Bloomberg School of Public Health 2014. This work is written by (a) US Government employee(s) and is in the public domain in the US.)

Background: Little is known about the epidemiology of oral human papillomavirus (HPV) in Latin America., Methods: Women (N = 5838) aged 22-29 in the control and vaccine arms of an HPV-16/18 vaccine trial in Costa Rica had oral, cervical, and anal specimens collected. Samples were tested for alpha mucosal HPV types (SPF10/LiPA25 version 1); a subset of oral samples (n = 500) was tested for cutaneous HPV types in the genera alpha, beta, gamma, mu, and nu., Results: In the control arm (n = 2926), 1.9% of women had an oral alpha mucosal HPV detected, 1.3% had carcinogenic HPV, and 0.4% had HPV-16; similar patterns for non-16/18 HPV types were observed in the vaccine arm. Independent risk factors for any oral alpha mucosal HPV among women in the control arm included marital status (adjusted odds ratio [AOR], 3.2; 95% confidence interval [CI], 1.8-5.7 for single compared to married/living as married), number of sexual partners (AOR, 2.4; 95% CI, 1.0-6.1 for ≥4 partners compared to 0-1 partners), chronic sinusitis (AOR, 3.1; 95% CI, 1.5-6.7), and cervical HPV infection (AOR, 2.6; 95% CI, 1.4-4.6). Detection of beta HPV was common (18.6%) and not associated with sexual activity., Conclusions: Unlike cutaneous HPV types, alpha mucosal HPV types were uncommon in the oral region and were predominately associated with sexual behavior. Clinical Trials Registration. NCT00128661.

The Costa Rica HPV16/18 Vaccine Trial (CVT) showed that four-year vaccine efficacy against 12-month HPV16/18 persistent infection was similarly high among women who received one, two, or the recommended three doses of the bivalent HPV16/18 L1 virus-like particle (VLP) vaccine. Live-attenuated viral vaccines, but not simple-subunit vaccines, usually induce durable lifelong antibody responses after a single dose. It is unclear whether noninfectious VLP vaccines behave more like live-virus or simple-subunit vaccines in this regard. To explore the likelihood that efficacy will persist longer term, we investigated the magnitude and durability of antibodies to this vaccine by measuring HPV16- and HPV18-specific antibodies by VLP-ELISA using serum from enrollment, vaccination, and annual visits through four years in four vaccinated groups; one-dose (n = 78), two-doses separated by one month (n = 140), two doses separated by six months (n = 52), and three scheduled doses (n = 120, randomly selected). We also tested enrollment sera from n = 113 HPV16- or HPV18 L1-seropositive women prevaccination, presumably from natural infection. At four years, 100% of women in all groups remained HPV16/18 seropositive; both HPV16/18 geometric mean titers (GMT) among the extended two-dose group were non-inferior to the three-dose group, and ELISA titers were highly correlated with neutralization titers in all groups. Compared with the natural infection group, HPV16/18 GMTs were, respectively, at least 24 and 14 times higher among the two-dose and 9 and 5 times higher among one-dose vaccinees. Antibody levels following one-dose remained stable from month 6 through month 48. Results raise the possibility that even a single dose of HPV VLPs will induce long-term protection., (©2013 AACR.)

Sexually transmitted carcinogenic human papillomavirus (HPV) infections are extraordinarily prevalent worldwide. However, most incident HPV infections clear within a few years, whereas a small minority persists to invasive cancer. Recent studies indicate that detection of methylated viral DNA may distinguish women with cervical intraepithelial neoplasia grade 2+ (CIN2+) from those with a carcinogenic HPV-type infection that shows no evidence of CIN2+. Several studies have reported a positive association between methylation of CpG sites in the L1 gene and CIN2+, although there are inconclusive results about methylation of CpG sites in the upstream regulatory region (URR). In this review, we summarize the current state of knowledge on HPV DNA methylation in cervical carcinogenesis, and discuss the merits of different methods used to measure HPV DNA methylation. To follow the promising leads, we suggest future studies to validate the use of methylated carcinogenic HPV DNA as a predictive and/or diagnostic biomarker for risk of cervical cancer among HPV-positive women.

Carcinogenic human papillomavirus (HPV) infections are very common after sexual debut and nearly all become undetectable ("clear") within a few years. Following clearance, the long-term risks of type-specific HPV re-appearance and subsequent risk of cervical intraepithelial neoplasia grade 2 or worse (CIN2+) are not well defined. In the 7-year, population-based cohort study in Guanacaste, Costa Rica, we studied how often type-specific carcinogenic HPV infections re-appeared after clearance and how often re-appearance led to CIN2+. We considered 1,740 carcinogenic HPV infections detected by MY09/11 PCR among 2,805 women (18-91 years old, median 34) who were actively followed at 6- or 12-month intervals. We identified women with one or more type-specific HPV infections that cleared and re-appeared and further defined a subgroup of "definite clearance and re-appearance" (≥2 intervening negative results over a period of ≥1 year). We determined the absolute risk of CIN2+ among the different groups. p values are two-sided. Only 7.7% (81/1,052) of HPV-infected women had intervening negative results. Very few (3.7%, 39/1,052) had "definite clearance and re-appearance", of which 5.1% (2/39) subsequently persisted to a diagnosis of CIN2. There were zero CIN3+ lesions. Extremely few women (2/2,805 of women in our cohort) had a type-specific carcinogenic HPV infection clear, re-appear and lead to CIN2+. If confirmed, this argues against vaccination to avoid re-appearance that leads to precursor lesions and against the need of frequent HPV screening after initial negative results., (Copyright © 2011 UICC.)

Background: Few studies have addressed the timing of cervical cytologic abnormalities and human papillomavirus (HPV) positivity during the course of an infection. It remains largely unknown how infections detected by HPV and cytology wax and wane relative to each other. The aim of this analysis was to assess the longitudinal relationship of abnormal cytology and HPV positivity in a 7-year prospective study of 2500 women in Guanacaste, Costa Rica., Methods: At each semiannual or annual visit, cervical specimens were screened using liquid-based cytology and tested for >40 HPV types with use of MY09/MY11 L1 degenerate primer polymerase chain reaction-based methods. On the basis of previous work, we separated prevalent and newly detected infections in younger and older women., Results: Among newly detected HPV- and/or cytology-positive events, HPV and cytology appeared together ∼60% of the time; when discordant, HPV tended to appear before cytology in younger and older women. Combining newly and prevalently detected events, HPV and cytology disappeared at the same time >70% of the time. When discordant, HPV tended to disappear after cytology in younger and older women., Conclusions: Detection of HPV DNA and associated cytological abnormalities tend to come and leave together; however, when discordant, detection of HPV DNA tends to precede and/or last longer than associated cytologic abnormalities.

To explore alternative cervical cancer screening approaches in an underserved population, we compared the performance of human papillomavirus (HPV) DNA assays in combination with different sample collection methods for primary cervical screening in the Mississippi Delta region. Three specimens were collected from women aged 26 to 65 years who were either routinely undergoing screening (n = 252) or not (n = 191): clinician-collected cervical specimens, clinician-collected cervicovaginal specimens, and self-collected cervicovaginal specimens taken at home. A novel collection device and medium were used for cervicovaginal sampling. Specimens were tested by three HPV DNA assays: hybrid capture 2 (HC2; Qiagen Corp., Gaithersburg, MD), Linear Array (LA; Roche Molecular Systems, Pleasanton, CA), and Amplicor (Roche Molecular Systems, Pleasanton, CA). Liquid-based cytology was performed on cervical specimens. We compared the overall positivity (a proxy for clinical specificity) for any carcinogenic HPV genotype and calculated the agreement across assay and specimen type using McNemar's test for differences in test positivity. Across all three assays there were no significant differences between clinician-collected and self-collected cervicovaginal specimens (P > 0.01 for all comparisons). For both cervicovaginal specimens (clinician collected and self-collected), fewer women tested positive by HC2 than by LA or Amplicor (P < 0.01 for all comparisons). HC2 had the best agreement between specimens for all assays. HC2 is likely more clinically specific, although possibly less sensitive, than either PCR test. Thus, use of HC2 on cervicovaginal specimens for screening could result in fewer referrals compared to LA and Amplicor.

Background: Seropositivity to human papillomavirus (HPV)16 and 18 antibodies is used as a measure of cumulative HPV exposure and as a stratifier of HPV exposure for vaccine efficacy analyses. Overall performance of these assays, as a measure of HPV exposure, has not been evaluated., Methods: Using data from the enrollment phase of the HPV16/18 vaccine trial in Costa Rica, we evaluated the performance of the polyclonal enzyme-linked immunosorbent assay (ELISA) HPV16 and 18 serological assays as a measure of HPV exposure. Biologic (e.g., HPV infection at the cervix) and behavioral characteristics (e.g., lifetime number of sexual partners) with known associations with current and past HPV infection were used to define cases and controls (HPV exposed vs. not exposed). Prevaccination serum was measured for antibodies against HPV16 and 18 by ELISA; cervical samples were tested for HPV DNA using PCR SPF10/LiPA25. ELISA results were analyzed using receiver-operator characteristic curves; performance was evaluated at the manufacturer set cut point (HPV16 = 8, HPV18 = 7) and at cut points chosen to optimize sensitivity and specificity (HPV16 = 34, HPV18 = 60)., Results: Defining cases as type-specific HPV DNA positive with high-grade abnormal cytology (i.e., combined molecular and microscopic markers of infection), HPV16-ELISA gave sensitivity that was lower at the optimal cut point than the manufacturer cut point (62.2 compared with 75.7, respectively; P = 0.44). However, specificity was higher (85.3 compared with 70.4, respectively; P < 0.0001). Similarly, HPV18-ELISA gave sensitivity that was lower at the optimal cut point than the manufacturer cut point (34.5 compared with 51.7, respectively; P = 0.40), with higher specificities (94.9 compared with 72.6, respectively; P < 0.0001)., Conclusions: Modifying cut points did not improve the low sensitivity. The low sensitivity of this assay does not support its use for risk stratification or clinical settings.

Background: A competitive Luminex Immunoassay (cLIA) has been developed to measure neutralizing antibodies against human papillomavirus (HPV) types 6, 11, 16 and 18., Methods: In a cohort of 974 women from the Guanacaste Natural History Study, we studied the relationship of baseline cLIA and virus-like particle (VLP) enzyme-linked immunosorbent assay (ELISA) (HPV16 and HPV18 only) seropositivity to measures of HPV exposure, HPV DNA positivity, number of sexual partners, cytology findings, and age. We then studied immunity against subsequent infection with HPV6, 11, 16, 18 and related types over a 7-year period., Results: cLIA seroprevalence varied with previous exposure; the prevalence of cLIA results positive for HPV16 and HPV18 was lower than the prevalence of positive VLP ELISA responses. cLIA and VLP ELISA positivity predicted protection from subsequent infections with concordant types. The combined odds ratio for HPV16 and HPV18 cLIA positivity was 0.41 (95% confidence interval [CI], 0.21-0.80), and the combined odds ratio for the HPV16 and HPV18 VLP ELISA positivity was 0.65 (95% CI, 0.46-0.93). Of individual types, statistical significance was only reached for HPV16 cLIA positivity (odds ratio, 0.44; 95% CI, 0.15-0.94)., Conclusions: Both assays showed an association between positive results and significant protection from subsequent infections for HPV16 and HPV18 combined. cLIA seroprevalence was lower than VLP ELISA, suggesting that the assay detects a subset of antibodies following natural infection that are specifically linked to immunity against subsequent HPV infection.

Objective: We investigated coinfection patterns for 25 human papillomavirus (HPV) types and assessed the risk conferred by multiple HPV types toward cervical disease., Methods: Sexually active women (n=5,871) in the NCI-sponsored Costa Rica HPV Vaccine Trial's prevaccination enrollment visit were analyzed. Genotyping for 25 HPVs was performed using SPF(10)/LiPA(25). We calculated odds ratios (ORs) to assess coinfection patterns for each genotype with 24 other genotypes. These ORs were pooled and compared with pair-specific ORs to identify genotype combinations that deviated from the pooled OR. We compared risk of CIN2+/HSIL+between multiple and single infections and assessed additive statistical interactions., Results: Of the 2478 HPV-positive women, 1070 (43.2%) were infected with multiple types. Multiple infections occurred significantly more frequently than predicted by chance. However, this affinity to be involved in a coinfection (pooled OR for 300 type-type combinations=2.2; 95% confidence interval [CI]=2.1-2.4) was not different across HPV type-type combinations. Compared with single infections, coinfection with multiple α9 species was associated with significantly increased risk of CIN2+(OR=2.2; 95% CI=1.1-4.6) and HSIL+(OR=1.6; 95% CI=1.1-2.4). However, disease risk was similar to the sum of estimated risk from individual types, with little evidence for synergistic interactions., Conclusions: Coinfecting HPV genotypes occur at random and lead to cervical disease independently.

The presence of more than one human papillomavirus (HPV) genotype may influence the duration of prevalently detected infections. This analysis included 1,646 infections detected at enrollment in 980 women from the Guanacaste, Costa Rica, cohort who were actively followed up every 6-12 months for up to 8 years. We categorized HPV infections as single or multiple types. Persistence of infections was estimated using discrete-time survival analysis. The difference between the duration of single and that of concurrent multiple type-specific prevalent HPV infections was not significant (P = .9; log-rank test). Concurrent, prevalent detection of additional HPV types did not change the likelihood of viral persistence.

Human papillomavirus (HPV) types differ profoundly in cervical carcinogenicity. For the most carcinogenic type HPV16, variant lineages representing further evolutionary divergence also differ in cancer risk. Variants of the remaining 10 to 15 carcinogenic HPV types have not been well studied. In the first prospective, population-based study of HPV variants, we explored whether, on average, the oldest evolutionary branches within each carcinogenic type predicted different risks of >2-year viral persistence and/or precancer and cancer [cervical intraepithelial neoplasia grade 3+ (CIN3+)]. We examined the natural history of HPV variants in the 7-year, 10,049-woman Guanacaste Cohort Study, using a nested case-control design. Infections were assigned to a variant lineage determined by phylogenetic parsimony methods based on URR/E6 sequences. We used the Fisher's combination test to evaluate significance of the risk associations, cumulating evidence across types. Globally, for HPV types including HPV16, the P value was 0.01 for persistence and 0.07 for CIN3+. Excluding HPV16, the P values were 0.04 and 0.37, respectively. For HPV16, non-European viral variants were significantly more likely than European variants to cause persistence [odds ratio (OR), 2.6; P = 0.01] and CIN3+ (OR, 2.4; P = 0.004). HPV35 and HPV51 variant lineages also predicted CIN3+. HPV variants generally differ in risk of persistence. For some HPV types, especially HPV16, variant lineages differ in risk of CIN3+. The findings indicate that continued evolution of HPV types has led to even finer genetic discrimination linked to HPV natural history and cervical cancer risk. Larger viral genomic studies are warranted, especially to identify the genetic basis for HPV16's unique carcinogenicity., ((c)2010 AACR.)

Determining the causal attribution of human papillomavirus (HPV) genotypes to cervical disease is important to estimate the effect of HPV vaccination and to establish a type spectrum for HPV-based screening. We analyzed the prevalence of HPV infections and their attribution to cervical disease in a population of 1,670 women referred to colposcopy for abnormal cytology at the University of Oklahoma. HPV genotyping was performed from cytology specimens using the Linear Array assay that detects 37 HPV genotypes. We used different methods of type attribution to revised cervical disease categories. We found very high prevalence of multiple HPV infections with up to 14 genotypes detected in single specimens. In all disease categories except for cancers, there was a significant trend of having more infections at a younger age. We did not see type interactions in multiple genotype infections. HPV16 was the most frequent genotype at all disease categories. Based on different attribution strategies, the attribution of vaccine genotypes (6, 11, 16, 18) ranged from 50.5 to 67.3% in cancers (n = 107), from 25.6 to 74.8% in CIN3 (n = 305), from 15.2 to 52.2% in CIN2 (n = 427), and from 6.6 to 26.0% in

Background: The impact of loop electrosurgical excision procedure (LEEP) treatment for cervical precancerous lesions on subsequent acquisition of new human papillomavirus (HPV) infections is not well described., Methods: The acquisition of new HPV infections was compared in HPV-positive women who underwent colposcopy and were treated by LEEP (n = 195) and those who were untreated (n = 1625) at entry into a 2-year study. Cumulative incidence rate ratios (IRRs) for treated versus untreated women at 6- and 24-months of follow-up, with 95% confidence intervals (CIs), were calculated for infection by individual HPV genotypes, any HPV genotypes, any carcinogenic HPV genotypes, any noncarcinogenic HPV genotypes, and phylogenetic groups of HPV genotypes., Results: Treated women were 29% less likely than untreated women to have carcinogenic HPV genotypes detected at 6-month follow-up visits (IRR, 0.71; 95% CI, 0.50-1.00) and were 18% less likely to have these genotypes detected at 24-month follow-up visits (IRR, 0.82; 95% CI, 0.68-1.01). Treated women were 56% less likely to have HPV genotypes of the alpha9 phylogenetic species (which includes HPV-16) detected at 6-month follow-up visits (IRR, 0.44; 95% CI, 0.23-0.85) and were 40% less likely to have these genotype detected at 24-month follow-up visits (IRR, 0.60; 95% CI, 0.42-0.85)., Conclusion: LEEP may reduce the acquisition of certain carcinogenic HPV genotypes related to HPV-16.

Driven by findings that human papillomavirus (HPV)-induced degradation of p53 differs by a TP53 polymorphism at codon 72 (Pro72Arg), past studies of TP53 genetic variants and cervical cancer have focused on this nonsynonymous polymorphism, with mixed results. We analyzed common single nucleotide polymorphisms (SNP) across the TP53 locus in a population-based nested case-control study in Guanacaste, Costa Rica. We evaluated 11 SNPs, including Pro72Arg (rs1042522), among 1,281 women: 465 with cervical intraepithelial neoplasia grade 3/cancer (CIN3+), 380 with HPV persistence (median, 25 months), and 436 random population controls. We combined HPV persistence and CIN3+ into one case group because they did not differ in TP53 genotypic frequencies and calculated odds ratios and 95% confidence intervals (CI) for individual SNPs and inferred haplotypes. We observed that proline at codon 72 was associated with increased risk of CIN3+/persistence compared with population controls. Relative to GG (Arg), the CG (Pro/Arg) and CC (Pro) genotypes had a 1.3-fold (95% CI, 0.99-1.6) and 1.8-fold (95% CI, 1.2-2.7) increased risk, respectively (P(trend) < 0.01). rs12951053 and rs1642785 were also associated with CIN3+/persistence (P (trend), 0.05 and 0.04, respectively), as was a haplotype containing the codon 72 variant (rs1042522), rs12951053, rs1642785, and rs12947788 (odds ratio, 1.6; 95% CI, 1.1-2.3 versus the most common haplotype, which comprised the major alleles for all 11 SNPs). Although genetic variation in TP53 might affect the natural history of HPV and cervical cancer, further work is needed to elucidate the possible mechanism.

Diagnosis and treatment of cervical cancer precursors rely on colposcopic biopsy, which is sometimes hampered by incorrect biopsy placement and the unclear prognostic value of poorly reproducible diagnoses such as cervical intraepithelial neoplasia (CIN) Grade 1 and 2. Searching for discrete disease categories that incorporate the value of cytology and that reflect the causal role of particular HPV types, we analyzed histology, cytology and HPV genotype distributions in the Study to Understand Cervical Cancer Endpoints and Early Determinants (SUCCEED). This cross-sectional study comprises approximately 1,700 women referred to colposcopy or treatment for the spectrum of cervical disease, including 439 women with

Human papillomavirus (HPV) cofactors for cervical cancer include smoking, multiparity, and oral contraceptive use, but their mechanisms of action are not fully understood. It is also unknown whether cofactors vary by HPV genotypes. The Study to Understand Cervical Cancer Early Endpoints and Determinants (SUCCEED) is a cross-sectional study comprising women referred to the University of Oklahoma from November 2003 to September 2007 for abnormal cervical screening results. Detailed questionnaire data and liquid cytology specimens were collected and the latter was genotyped for HPV using the LINEAR ARRAY HPV Genotyping Test. The present analysis includes women with both questionnaire and HPV data and diagnosed with

Variations in biological behavior suggest that each carcinogenic human papillomavirus (HPV) type should be considered individually in etiologic studies. HPV genotyping assays might have clinical applications if they are approved for use by the FDA. A widely used genotyping assay is the Roche Linear Array HPV genotyping test (LA). We used LA to genotype the HPV isolates from cervical specimens from women with the full spectrum of cervical disease: cervical cancer, cervical intraepithelial neoplasia (CIN), and HPV infections. To explore the feasibility and value of the automated reading of the LA results, we custom-designed novel, optical imaging software that provides optical density measurements of LA bands. We compared unmagnified visual examination with the automated measurements. The two measurements were highly associated. By either method, the threshold between a negative and a positive result was fairly sharp, with a clear bimodal distribution. Visually, most positive results were judged to be strong or medium, with fewer equivocal results categorized as weak (9.5% of positive samples), very weak (6.5% of positive samples), or extremely weak (7.7% of positive samples). The automated measurements of the intensities were significantly associated with the strength of the visual categories (P < 0.001). At the extremes of the automated signal intensities (< or = 20 units or > or = 120 units), the bands were almost always categorized visually as negative and positive, respectively. In the equivocal zone (20 to 119 units), specimens were more increasingly likely to be judged to be visually positive as the number of other, definite infections on the same strip increased (P for trend < 0.001). Multiple, concurrent infections comprise > or = 25% of HPV infections; thus, any systematic visual tendency that influences their evaluation when the result is equivocal should be minimized. Therefore, automated reading is probably worth development if easy-to-calibrate hardware and software can be optimized.

Hybrid Capture 2 (hc2), a clinical test for carcinogenic human papillomavirus (HPV) DNA, has proven to be a sensitive but only modestly specific predictor of cervical precancer and cancer risk. Some of its nonspecificity for clinical end points can be ascribed to cross-reactivity with noncarcinogenic HPV genotypes. However, the reference genotyping tests that have been used for these comparisons are also imperfect. We therefore sought to describe further the HPV genotype specificity of hc2 by comparing the hc2 results to paired results from two related PGMY09/11 L1 primer-based HPV genotyping assays: Linear Array (LA) and its prototype predecessor, the line blot assay (LBA). LA and LBA results were considered separately and combined (detection by either assay or both assays) for 37 individual HPV genotypes and HPV risk group categories (carcinogenic HPV > noncarcinogenic HPV > negative). Baseline specimens from 3,179 of 3,488 (91.5%) women referred to ALTS (a clinical trial to evaluate the management strategies for women with atypical squamous cells of undetermined significance [ASCUS] or low-grade squamous intraepithelial lesions) because of an ASCUS Papanicolaou smear were tested by all three assays. Among single-genotype infections with genotypes targeted by hc2 as detected by either PCR assay, HPV genotype 35 (HPV35) (86.4%), HPV56 (84.2%), and HPV58 (76.9%) were the most likely to test positive by hc2. Among single-genotype infections with genotypes not targeted by hc2 as detected by either assay, HPV82 (80.0%), HPV66 (60.0%) (recently classified as carcinogenic), HPV70 (59.1%), and HPV67 (56.3%) were the most likely to test positive by hc2. Among women who tested negative for carcinogenic HPV by both PCR tests and were positive for noncarcinogenic HPV by either test, 28% of women were hc2 positive. Conversely, 7.8% of all hc2-positive results in this population were due to cross-reactivity of hc2 with untargeted, noncarcinogenic HPV genotypes. In conclusion, hc2 cross-reacts with certain untargeted, noncarcinogenic HPV genotypes that are phylogenetically related to the targeted genotypes, but the degree of cross-reactivity may be less than previously reported.

Objective: Cross-sectional analyses of our 10,000-woman, population-based Guanacaste cohort suggest a lag of > or =10 years between the peak of human papillomavirus (HPV) infection and the later peak of cervical intraepithelial neoplasia grade 3 (CIN 3). We wanted to explore early HPV natural history and CIN 3 prospectively., Study Design: As part of the Guanacaste cohort, we followed 206 initially virginal women aged 18 to 26 semiannually for a median of 3.6 years after initiation of sexual life., Results: A total of 53.4% of women tested positive during the study for > or =1 HPV type. Very few infections persisted for >1 to 2 years. Three women had histologically confirmed CIN 3, of which 2 showed persistent HPV 16. The other had serologic evidence of HPV 31., Conclusions: HPV infection occurs frequently and clears rapidly in most young women initiating sexual intercourse. Persistent HPV 16 can cause early CIN 3. The peak age for CIN 3 will decline with the increased screening intensity and sensitivity typical of longitudinal studies.

We compared human papillomavirus (HPV) prevalence in an age-stratified random sample of women who have undergone a hysterectomy (WH) (n=573) with the HPV prevalence in age-matched women with intact cervices (women who have not undergone a hysterectomy [WNH]) (n=581) participating in a study at Kaiser Permanente in Portland, Oregon. Testing cervicovaginal lavage fluids for >40 HPV genotypes using an MY09/11 L1 consensus primer polymerase chain reaction method, we found no statistical differences in the prevalence of HPV (16% for WNH vs. 13.9% for WH) or carcinogenic HPV (6.5% for WNH vs. 4.5% for WH) between the 2 groups of women. Although WH have a similar prevalence of carcinogenic HPV infection, compared with WNH without a cervix, they have minimal risk of HPV-induced cancer and are unlikely to benefit from HPV testing.

Persistent cervical infections with carcinogenic human papillomaviruses (HPV) cause virtually all cervical cancer. Cytologic abnormalities are the manifestations of HPV infections used to identify women at risk. To compare the potential of the full range of anogenital HPV genotypes to induce cytopathic effects, we examined the influences of HPV type, viral load, and age on cytopathology among 1,222 women having a single HPV type at enrollment into a 10,000-woman population-based study in Costa Rica. Cervical specimens were tested for approximately 40 HPV types by MY09/MY11 L1 primer PCR and type-specific dot blot hybridization. Types were organized by phylogenetic species and cancer risk. PCR signal strength served as a qualitative surrogate for viral load. Overall, 24.8% [95% confidence interval (95% CI), 22.4-27.3] of single prevalent HPV infections had concurrent abnormalities (atypical squamous cells or worse) ranging from 0.0% to 80.0% based on HPV type. Noncarcinogenic alpha3/alpha15 types, although highly prevalent, uncommonly caused cytologic abnormalities (13.1%; 95% CI, 9.8-17.0). In contrast, one quarter to nearly one half of infections with a single major carcinogenic species type (alpha9/alpha11/alpha7/alpha5/alpha6) produced abnormalities. Greater abnormalities were observed with increasing qualitative viral load of carcinogenic types; fewer abnormalities were observed among older women (>54 years). A high percentage (46.2%) of detected abnormalities in women infected with HPV16 or related alpha9 types were high grade or worse, consistent with strong carcinogenicity, compared with 10.7% in women infected with alpha7 types, including HPV18, a major cause of adenocarcinoma. The lack of evident severe abnormalities associated with HPV18 and related HPV types might have implications for screening for poorly detected glandular and alpha7-related lesions.

Background: Loop electrosurgical excision procedure (LEEP) is the predominant treatment for cervical intraepithelial neoplasia grade 2 or 3 (CIN2+) in the United States, yet following treatment approximately 10% of women are diagnosed again with CIN2+, necessitating close follow-up of such patients., Methods: Surveillance strategies using cytology and/or human papillomavirus (HPV) testing were compared among women who underwent LEEP (n = 610) in the Atypical Squamous Cells of Undetermined Significance (ASCUS) Low-Grade Squamous Intraepithelial Lesion (LSIL) Triage Study. Cervical specimens, collected at 6-month visits for 2 years, were used for cytology, Hybrid Capture 2 (HC2) detection of carcinogenic HPVs, and PCR for genotyping of carcinogenic and noncarcinogenic HPV types. At exit, women had colposcopy for safety and disease ascertainment., Results: At the visit post-LEEP (median time: 4.5 months after LEEP), 36.9% [95% confidence interval (95% CI), 32.7-41.1%] of women were positive for carcinogenic HPV by PCR and 33.7% (95% CI, 29.7-37.9) had ASCUS or more severe (ASCUS+) cytology. The overall 2-year cumulative incidence of histologically confirmed posttreatment CIN2+ was 7.0%; this could be further stratified by the HPV risk category detected at the 6-month visit after LEEP. The 2-year risk associated with HPV16 positivity was 37.0%, significantly higher than for other carcinogenic HPV types (10.8%, P < 0.001), noncarcinogenic types (1.5%, P < 0.001), or testing HPV negative (0%). Post-LEEP cytology (using a positive threshold of ASCUS+) was 78.1% (95% CI, 60.0-90.7%) sensitive for detection of posttreatment CIN2+. By comparison, PCR for carcinogenic HPV and combination testing (using a positive result from carcinogenic HPV testing or cytology as the test threshold with HPV-negative ASCUS not referred) were significantly more sensitive (96.9% for each, P = 0.03); HC2 alone was nonsignificantly more sensitive (90.6%, P = 0.3). Specificity was similar for ASCUS+ cytology (69.1%, 95% CI, 64.6-73.3%) and PCR for carcinogenic HPV (67.1%, P = 0.5), yet was lower for HC2 (63.8%, P = 0.048) and combination testing (62.9%, P = 0.02)., Conclusion: Women who tested positive after LEEP for carcinogenic HPV types, especially HPV16, had high risk of subsequent CIN2+. HPV-based detection methods, alone or in combination with cytology, may be useful to incorporate in post-LEEP management strategies.

Approximately 15 human papillomavirus (HPV) types cause virtually all cervical cancer whereas other HPV types are unrelated to cancer. We were interested in whether some noncarcinogenic types differ from carcinogenic in their affinity for the cervical transformation zone, where nearly all HPV-induced cancers occur. To examine this possibility, we tested cervical specimens from 8,374 women without cervical precancer and cancer participating in a population-based study in Guanacaste for >40 HPV types using PCR. We compared age-group specific prevalences of HPV types of the alpha9 species, which are mainly carcinogenic and include HPV16, to the genetically distinct types of the alpha3/alpha15 species (e.g., HPV71), which are noncarcinogenic and common in vaginal specimens from hysterectomized women. We related HPV detection of each group to the location of the junction between the squamous epithelium of the ectocervix and vagina and the columnar epithelium of the endocervical canal. Models evaluated the independent effects of amount of exposed columnar epithelium (ectopy) and age on the presence of alpha9 or alpha3/alpha15 types. Prevalence of alpha9 types (7.6%) peaked in the youngest women, declined in middle-aged women, and then increased slightly in older women. By contrast, prevalence of alpha3/alpha15 types (7.6%) tended to remain invariant or to increase with increasing age. Detection of alpha9 infections increased (P(trend) < 0.0005) but alpha3/alpha15 infections decreased (P(trend) < 0.0005) with increasing exposure of the columnar epithelia. Older age and decreasing cervical ectopy were independently positively associated with having an alpha3/alpha15 infection compared with having an alpha9 infection. These patterns need to be confirmed in other studies and populations. We suggest that these genetically distinct groups of HPV types may differ in tissue preferences, which may contribute to their differences in carcinogenic potential.

Background: Human papillomavirus (HPV) types 16 and 18 cause 60%-70% of cervical cancer worldwide, and other HPV types cause virtually all remaining cases. Pooled HPV testing for 13 oncogenic types, including HPV16 and 18, is currently used in clinical practice for triage of equivocal cytology and, in conjunction with Pap tests, is an option for general screening among women 30 years of age and older. It is not clear to what extent individual identification of HPV16 or HPV18 as an adjunct to pooled oncogenic HPV testing might effectively identify women at particularly high risk of cervical cancer or its immediate precursor, cervical intraepithelial neoplasia 3 (CIN3)., Methods: From April 1, 1989, to November 2, 1990, a total of 20 810 women in the Kaiser Permanente health plan in Portland, OR, enrolled in a cohort study of HPV and cervical neoplasia. Women were tested for 13 oncogenic HPV types by Hybrid Capture 2 (HC2), and those women with a positive HC2 test were tested for HPV16 and 18. Enrollment Pap smear interpretation and HPV test results were linked to histologically confirmed CIN3 and cervical cancer (> or = CIN3) occurring during 10 years of cytologic follow-up. We calculated cumulative incidence rates with 95% confidence intervals for each interval up to 122 months using Kaplan-Meier methods., Results: The 10-year cumulative incidence rates of > or = CIN3 were 17.2% (95% confidence interval [CI] = 11.5% to 22.9%) among HPV16+ women and 13.6% (95% CI = 3.6% to 23.7%) among HPV18+ (HPV16-) women, but only 3.0% (95% CI = 1.9% to 4.2%) among HC2+ women negative for HPV16 or HPV18. The 10-year cumulative incidence among HC2- women was 0.8% (95% CI = 0.6% to 1.1%). A subanalysis among women 30 years of age and older with normal cytology at enrollment strengthened the observed risk differences., Conclusions: HPV screening that distinguishes HPV16 and HPV18 from other oncogenic HPV types may identify women at the greatest risk of > or = CIN3 and may permit less aggressive management of other women with oncogenic HPV infections.

Persistent infections with carcinogenic human papillomaviruses (HPV) cause virtually all cervical cancers. Cervical HPV types (n > 40) also represent the most common sexually transmitted agents, and most infections clear in 1-2 years. The risks of persistence and neoplastic progression to cancer and its histologic precursor, cervical intraepithelial neoplasia grade 3 (CIN3), differ markedly by HPV type. To study type-specific HPV natural history, we conducted a 10,000-woman, population-based prospective study of HPV infections and CIN3/cancer in Guanacaste, Costa Rica. By studying large numbers of women, we wished to separate viral persistence from neoplastic progression. We observed a strong concordance of newly-revised HPV evolutionary groupings with the separate risks of persistence and progression to CIN3/cancer. HPV16 was uniquely likely both to persist and to cause neoplastic progression when it persisted, making it a remarkably powerful human carcinogen that merits separate clinical consideration. Specifically, 19.9% of HPV16-infected women were diagnosed with CIN3/cancer at enrollment or during the five-year follow-up. Other carcinogenic types, many related to HPV16, were not particularly persistent but could cause neoplastic progression, at lower rates than HPV16, if they did persist. Some low-risk types were persistent but, nevertheless, virtually never caused CIN3. Therefore, carcinogenicity is not strictly a function of persistence. Separately, we noted that the carcinogenic HPV types code for an E5 protein, whereas most low-risk types either lack a definable homologous E5 ORF and/or a translation start codon for E5. These results present several clear clues and research directions in our ongoing efforts to understand HPV carcinogenesis.

Background: Cross-sectional human papillomavirus (HPV) DNA prevalence peaks at young ages, reflecting sexual acquisition and typically rapid clearance. In some populations, HPV prevalence demonstrates a second peak in older women. Longitudinal data may help to explain this second peak., Methods: We followed a population-based cohort of 7237 women in Guanacaste, Costa Rica, in which we had previously observed a second peak in the baseline HPV prevalence in older women. We tested for >40 HPV types by polymerase chain reaction. We analyzed age-specific patterns of acquisition and persistence 5-7 years after enrollment for individual HPV types., Results: At enrollment and follow-up, cross-sectional data revealed U-shaped age-specific HPV prevalence curves for virtually every type, with higher prevalences in the younger and older women than in the middle-aged women. Prospectively, acquisition of types decreased significantly as women aged (PTrend<.05, for both), with the highest peak in young women and a secondary minor peak in older women. Type-specific persistence of HPV increased with age (PTrend<.0001). Overall, HPV acquisition predominated at younger ages, whereas persistent infections gradually became more prominent with age (PTrend<.0001)., Conclusions: Newly apparent infections decreased, whereas persistence increased, with age; this latter tendency supports the utility of HPV screening in older women.

Background: Detailed epidemiologic studies of cervical type-specific human papillomavirus (HPV) infection in large populations are scarce., Methods: We recruited a population-based cohort in Guanacaste, Costa Rica. Participants were interviewed, screened for cervical neoplasia, and tested for >40 HPV types by use of MY09/11 L1 consensus primer polymerase chain reaction. We estimated the risk factors for infection and the associations between type-specific HPV infections and cervical intraepithelial neoplasia (CIN) and cancer in 8514 sexually active women who had not undergone a hysterectomy., Results: The overall HPV prevalence was 26.5%. The most common type was HPV-16 (3.6% of the population). HPV prevalence showed a U-shaped age-specific curve. Sexual behaviors were the main determinants of oncogenic and nononcogenic infections; age at first sexual intercourse was not independently associated with infection. Barrier contraceptive use was somewhat protective against infection. Oncogenic infections were strongly associated with risk of all grades of CIN and of cancer. Types 16, 18, and 58 were the most common in women diagnosed with CIN3 and cancer. Except for those that included HPV-16, multiple-type infections were associated with an increased risk (compared with that for single-type infections) of all grades of CIN and of cancer., Conclusions: We confirmed the bimodal age pattern of HPV infection in Guanacaste and the sexually transmitted nature of both oncogenic and nononcogenic HPV types.

Use of human papillomavirus (HPV) testing in cervical cancer prevention is increasing rapidly. A DNA test for 13 HPV types that can cause cervical cancer is approved in the United States for co-screening with cytology of women >or=30 years old and for triage of women of all ages with equivocal cytology. However, most infections with HPV are benign. We evaluated trade-offs between specificity and sensitivity for approximately 40 HPV types in predicting cervical intraepithelial neoplasia 3 and cancer in two prospective studies: a population-based screening study that followed 6196 women aged 30-94 years from Costa Rica for 7 years and a triage study that followed 3363 women aged 18-90 years with equivocal cytology in four U.S. centers for 2 years. For both screening and triage, testing for more than about 10 HPV types decreased specificity more than it increased sensitivity. The minimal increases in sensitivity and in negative predictive value achieved by adding HPV types to DNA tests must be weighed against the projected burden to thousands of women falsely labeled as being at high risk of cervical cancer.

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.>

We compared point prevalences and determinants of human papillomavirus (HPV) DNA detection by testing enrollment vaginal specimens from hysterectomized women (n=569) and enrollment cervical specimens from nonhysterectomized women (n=6098) >or=30 years old, using MY09/MY11 L1 consensus-primer polymerase chain reaction. The subjects were participating in a population-based cohort study (n=10,049) in Guanacaste, Costa Rica, that was initiated in 1993. Non-cancer-associated HPV types, especially types 61, 71, and 72, were detected more frequently in the vaginal specimens from hysterectomized women (23.7% [95% confidence interval [CI], 20.3%-27.4%]) than in the cervical specimens from nonhysterectomized women (16.7% [95% CI, 15.7%-17.6%]) (P=.0001). There was no difference between the prevalences of cancer-associated HPV types in hysterectomized women and those in nonhysterectomized women; in both groups, the prevalence of HPV DNA was greater in women with multiple lifetime sex partners. We infer from our data that the cervical transformation zone may not be needed for cancer-associated HPV infection but may be uniquely susceptible to HPV-induced carcinogenesis; we also infer that specific phylogenetic groups of HPV (i.e., A3/A4/A15) may have a predilection for vaginal epithelium.

Increased understanding of human papillomavirus (HPV) infection as the central cause of cervical cancer has permitted the development of improved screening techniques. To evaluate their usefulness, we evaluated the performance of multiple screening methods concurrently in a large population-based cohort of >8500 nonvirginal women without hysterectomies, whom we followed prospectively in a high-risk region of Latin America. Using Youden's index as a measure of the trade-off between sensitivity and specificity, we estimated the performances of a visual screening method (cervicography), conventional cytology, liquid-based cytology (ThinPrep), and DNA testing for 13 oncogenic HPV types. The reference standard of disease was neoplasia > or = cervical intraepithelial neoplasia grade 3 (CIN 3), defined as histologically confirmed CIN 3 detected within 2 years of enrollment (n=90) or invasive cancer detected within 7 years (n=20). We analyzed each technique alone and in paired combinations (n=112 possible strategies), and evaluated the significance of differences between strategies using a paired Z test that equally weighted sensitivity and specificity. As a single test, either liquid-based cytology or HPV DNA testing was significantly more accurate than conventional cytology or cervicography. Paired tests incorporating either liquid-based cytology or HPV DNA testing were not substantially more accurate than either of those two test strategies alone. However, a possibly useful synergy was observed between the conventional smear and cervicography. Consideration of age or behavioral risk profiles did not alter any of these conclusions. Overall, we conclude that highly accurate screening for cervical cancer and CIN 3 is now technically feasible. The remaining vital issue is to extend improved cervical cancer prevention programs to resource-poor regions.

We compared the performance of a prototype version of the Hybrid Capture 3 (HC3) human papillomavirus (HPV) DNA assay to the current generation Hybrid Capture 2 (HC2) assay, both of which target 13 oncogenic HPV types, for the detection of cervical intraepithelial neoplasia grade 3 and cancer (CIN3+) with cervicovaginal lavage specimens collected at enrollment into a 10-year cohort study at Kaiser Permanente (Portland, Oreg.). HC3 results for a risk-stratified sample (n = 4,364) were compared to HC2 results for the entire cohort (n = 20,810) with receiver operating characteristics curves, and the optimal cut points for both tests (relative light units [RLU]/positive control [PC]) for the detection of CIN3+ were determined. Specimens were also tested for HPV16 and HPV18 with separate HC3 type-specific probes. The optimal cut point for detecting CIN3+ was 1.0 RLU/PC for HC2, as previously shown, and was 0.6 RLU/PC for HC3. At the optimal cut points, HC3 and HC2 had similar screening performance characteristics for CIN3+ diagnosed at the enrollment visit. In analyses that included cases CIN3+ at enrollment and those diagnosed during early follow-up, HC3 had nonsignificantly higher sensitivity and equal specificity for the detection of CIN3+ compared to HC2; this increase in sensitivity was primarily the result of increased detection of CIN3+ in women who were 30 years of age or older and were cytologically negative (P = 0.006). We also compared the performance of the hybrid capture tests to MY09/11 L1 consensus primer PCR results (n = 1,247). HC3 was less likely than HC2 to test positive for specimens that tested positive by PCR for any untargeted types (P < 0.001). HC3 was less likely than HC2 to test positive for untargeted PCR-detected single infections with HPV53 (P = 0.001) and HPV66 (P = 0.01). There was good agreement between test positivity by PCR and by single type-specific HC3 probes for HPV16 (kappa = 0.76; 95% confidence interval [CI] = 0.71 to 0.82) and for HPV18 (kappa = 0.73; 95% CI = 0.68 to 0.79). In conclusion, we suggest that HC3 (>/=0.6 RLU/PC) may be slightly more sensitive than and equally specific test as HC2 (>/=1.0 RLU/PC) for the detection of CIN3+ over the duration of typical screening intervals.

Appropriately sophisticated statistical approaches are crucial for addressing the increasingly complex set of critical questions that follow from the recognition that human papillomavirus (HPV) is a necessary causal factor for cervical cancer. Cervical cancer researchers have defined the major stages of cervical carcinogenesis, with HPV infection as the necessary cause. Focus of etiologic studies is shifting from establishing causality to determining risk factors for HPV persistence and neoplastic progression using serially collected biomarkers. Prevention-oriented epidemiology and trials of new screening strategies and vaccines will rely on surrogate endpoints because we cannot let women develop cancer when it can be prevented. Future epidemiologic and prevention studies of HPV infection and cervical carcinogenesis will exploit subtle pathologic distinctions and will employ improved measurements of complex molecular biologic phenomena. The anticipated statistical issues are highlighted in this discussion.

Background: Annual Pap smear screening has been favored over less frequent screening in the United States to minimize the risk of cervical cancer. We evaluated whether simultaneous screening with a Pap test and human papillomavirus (HPV) testing is useful for assessing the risk for cervical intraepithelial neoplasia (CIN) 3 or cervical cancer., Methods: We enrolled 23 702 subjects in a study of HPV infection at Kaiser Permanente, Northwest Division, Portland, OR. Data were analyzed for 20 810 volunteers who were at least 16 years old (mean = 35.9 years) with satisfactory baseline Pap tests and suitable samples for HPV testing. Women were followed for up to 122 months (from April 1, 1989, to June 30, 1999) to determine the risk for histopathologically confirmed CIN3 or cancer., Results: Among 171 women with CIN3 or cancer diagnosed over 122 months, 123 (71.9%, 95% confidence interval [CI] = 65.2% to 78.7%) had baseline Pap results of atypical squamous cells or worse and/or a positive HPV test, including 102 (86.4%, 95% CI = 80.3% to 92.6%) of the 118 cases diagnosed within the first 45 months of follow-up. During this 45-month period, the cumulative incidence of CIN3 or cancer was 4.54% (95% CI = 3.61% to 5.46%) among women with a Pap test result of atypical squamous cells or worse, positive HPV tests, or both compared with 0.16% (95% CI = 0.08% to 0.24%) among women with negative Pap and HPV tests. Age, screening behavior, a history of cervical cancer precursors, and a history of treatment for CIN minimally affected results., Conclusions: Negative baseline Pap and HPV tests were associated with a low risk for CIN3 or cancer in the subsequent 45 months, largely because a negative HPV test was associated with a decreased risk of cervical neoplasia. Negative combined test results should provide added reassurance for lengthening the screening interval among low-risk women, whereas positive results identify a relatively small subgroup that requires more frequent surveillance.

Background: The addition of human papillomavirus (HPV) DNA testing to cytologic screening for cervical carcinoma is now being considered. The majority of women in screening cohorts who test positive for oncogenic types of HPV DNA have concurrent negative Pap tests. The absolute risk of a subsequent abnormal Pap test for these women is uncertain. Therefore, the proper counseling and clinical management of these women is also uncertain., Methods: A subcohort of 2020 women with a negative Pap test who tested positive at enrollment for oncogenic HPV DNA types using the Hybrid Capture 2 Test were followed for 57 months at Kaiser Permanente (Portland, OR). Absolute risks of new abnormal cytologic interpretations were computed using Kaplan-Meier methods. Logistic regression models were used to evaluate determinants of a new abnormal Pap test., Results: The cumulative incidence for a Pap test interpreted as atypical squamous cells or more severe (>or= ASC) was 16.8% (95% confidence interval [CI] = 15.0-18.6%), 6.4% (95% CI = 5.2-7.6%) for low-grade squamous intraepithelial lesions or more severe, and 2.2% (95% CI = 1.5-2.9%) for high-grade squamous intraepithelial lesions or more severe. By comparison, the cumulative incidence of greater than or equal to ASC among HPV-negative women was 4.2% (95% CI = 3.9-4.6%). The highest viral load (100 relative light units per the positive control or greater) was associated with a greater risk of an abnormal Pap test (odds ratio= 2.7, 95% CI = 1.7-4.1) than lower viral loads., Conclusions: These results suggest that about 15% of women in annual screening programs who concurrently have a negative Pap test and a positive oncogenic HPV test will have a subsequent abnormal Pap test within 5 years. This risk estimate will be useful to the many clinicians and patients likely to be diagnosed with an HPV infection and negative cytology if HPV DNA is added to general screening., (Published 2002 by the American Cancer Society.)

Hybrid Capture 2 Test using probe B (HC2-B) is a clinical test for the detection of 13 human papillomavirus (HPV) types associated with cervical cancer (oncogenic types), but the potential clinical significance of HC2-B cross-reactivity with untargeted (nononcogenic) HPV types has not been fully evaluated. Thus, HC2-B test results on 954 clinical cervical specimens from a population-based natural history study of HPV in Costa Rica were compared with the data from testing of the same specimens twice by HPV type-specific MY09/MY11 L1 consensus primer PCR. Specimens positive by PCR for single HPV types not targeted by HC2-B were used for determining type-specific cross-reactivity. Effects of cross-reactivity on clinical performance were estimated by calculating sensitivity and specificity with and without cross-reactivity for the detection of high-grade cervical lesions. HC2-B tested positive for single infections by untargeted (cross-reactive) types 11, 53, 61, 66, 67, 70, 71, and 81. Cross-reactivity was strongly associated with PCR signal strength (P(Trend) = 0.0001) and cervical abnormalities (P = 0.0002, Pearson chi(2)). We estimated that HC2-B cross-reactivity resulted in minor changes in screening performance. Clinical sensitivity increased from 84.3% to 87.9%, clinical specificity decreased from 89.6% to 88.1%, and referral rates increased from 11.7% to 13.2% for detection of >or=cervical intraepithelial neoplasia grade 2. The clinical effect of cross-reactivity varied by cytologic interpretation. Among women with normal cytologic interpretations, cross-reactivity significantly improved the accuracy of identifying cytologically nonevident histology of >or=cervical intraepithelial neoplasia grade 2 because of increased sensitivity with maintained specificity. However, among women with equivocal or mildly abnormal cytologic interpretations, cross-reactivity decreased the accuracy of HPV testing because of substantial decreases in specificity. In summary, cross-reactivity with nononcogenic HPV types had little effect on the overall clinical performance of HC2-B as a general screening test, but reduction of cross-reactivity might improve the performance of HPV testing for triage of equivocal or mildly abnormal cytologic interpretations.

Background: In case-control studies, smoking, parity, and oral contraceptive use have been associated with an increased risk of cervical intraepithelial neoplasia grade 3 (CIN3) and cervical cancer among women who are infected with oncogenic human papillomavirus (HPV). However, these potential risk factors have not been adequately studied in prospective studies., Methods: We studied 1812 women who were enrolled in a 10-year prospective study of cervical neoplasia at Kaiser Permanente in Portland, Oregon, and who at enrollment had tested positive for oncogenic HPV DNA and had responded to a questionnaire that included questions on smoking, oral contraceptive use, and parity. Absolute risks and crude relative risks (RRs) with 95% confidence intervals (CIs) for CIN3 or cervical cancer were computed for three time intervals (0-8, 9-68, and 69-122 months after enrollment) using the Kaplan-Meier method. Conditional logistic regression models were used to control for factors that may have influenced our risk estimates, specifically the cytologic interpretation of baseline Pap smear, number of Pap smears during follow-up, age at enrollment, age at prediagnosis visit, and age at diagnosis. All statistical tests were two-sided., Results: Oral contraceptive use and parity were not associated with risk of CIN3 or cervical cancer. Former smokers, women who smoked less than one pack of cigarettes per day, and women who smoked one or more packs per day had crude RRs for CIN3 or cervical cancer for the entire follow-up period of 2.1 (95% CI = 1.1 to 3.9), 2.2 (95% CI = 1.2 to 4.2), and 2.9 (95% CI = 1.5 to 5.6), respectively, compared with never smokers. In the multivariable model, former smokers, women who smoked less than one pack/day, and women who smoked one or more packs/day had RRs of 3.3 (95% CI = 1.6 to 6.7), 2.9 (95% CI = 1.4 to 6.1), and 4.3 (95% CI = 2.0 to 9.3), respectively, for CIN3 or cervical cancer compared with never smokers., Conclusions: Smoking is associated with an increased risk of invasive cervical cancer in women who are infected with oncogenic HPV. Subsequent studies should examine the role of smoking in the multistage pathogenesis of cervical cancer.

Carcinogenic human papillomaviruses (HPV) are thought to be necessary for development of cervical cancer. We assessed whether higher viral loads of such viruses predicted future risk of CIN3 or cancer (CIN3+) in a cohort of 20810 women followed up for 10 years with cytological screening. We measured the viral load for 13 types of carcinogenic HPV (relative light units normalised to 1 pg/mL HPV 16 positive controls [RLU/PC]) using Hybrid Capture 2 testing of cervicovaginal lavages obtained at enrolment. Results were stratified into four groups (RLU/PC 1 to <10, 10 to <100, 100 to <1000, > or = 1000). Although presence of HPV strongly increased risk of CIN3+, high viral load did not further predict risk of CIN3+.

We examined factors associated with high-grade squamous intraepithelial lesions (HSIL) and cervical cancer among human papillomavirus (HPV)-infected women in a prevalent case-control study conducted within a population-based cohort of 10 077 women in Costa Rica. We compared 146 women with HPV-positive HSIL or cancer (HSIL/CA) against 843 HPV-positive women without evidence of HSIL/CA. Subjects completed a risk factor questionnaire. We evaluated the associations between exposures and HSIL/CA among women positive for any HPV and restricted to those positive for high-risk HPV types. Risk of HSIL/CA increased with increasing number of live births (P(trend)= 0.04). Women who smoked 6+ cigarettes/day had a RR for HSIL/CA of 2.7 (95% CI = 1.1-6.7) compared to non-smokers. Current use of barrier contraceptives was associated with a reduction in risk of HSIL/CA (RR = 0.39; 95% CI = 0.16-0.96). Sexual behaviour and a self-reported history of sexually transmitted diseases (STDs) other than HPV were not associated with HSIL/CA. Oral contraceptive use was associated with HSIL/CA among women with <3 pregnancies. Effects were similar in analysis restricted to women positive for high-risk HPV types. Among women positive for high-risk HPV types, 44% of HSIL/CA could be attributed to multiparity (>/=3 pregnancies) and/or smoking. Among HPV-positive women, multiparity and smoking are risk factors for HSIL/CA. Oral contraceptive use may be associated with HSIL/CA in subgroups of women., (Copyright 2001 Cancer Research Campaign http://www.bjcancer.com.)

Human papillomavirus (HPV)-16 causes about half the cases of cervical cancer worldwide and is the focus of HPV vaccine development efforts. Systematic data are lacking as to whether the prevention of HPV-16 could affect the equilibrium of infection with other HPV types and thus alter the predicted impact of vaccination on the occurrence of cervical neoplasia. Therefore, the associations of HPV-16 detection with subsequent acquisition of other HPV types and with the persistence of concomitantly detected HPV types were examined prospectively among 1124 initially cytologically normal women. Preexisting HPV-16 was generally associated with an increased risk for subsequent acquisition of other types. HPV-16 did not affect the persistence of concomitant infections, regardless of type. These findings suggest that the prevention or removal of HPV-16 is not likely to promote the risk of infection with other types, a theoretical concern with current vaccination efforts.

Background: Human papillomavirus (HPV) is the main cause of cervical neoplasia. Because few population-based studies have investigated the prevalence of type-specific infection in relation to cervical disease, we studied a high-risk population, estimating the prevalence of HPV infection and the risk associated with various HPV types., Methods: We screened 9175 women in Guanacaste, Costa Rica, to obtain a referent standard final diagnosis, and tested 3024 women for more than 40 types of HPV with a polymerase chain reaction-based system., Results: Among women with normal cytology, HPV infections peaked first in women younger than 25 years, and they peaked again at age 55 years or older with predominantly non-cancer-associated types of HPV and uncharacterized HPV types. Low-grade squamous intraepithelial lesions (LSILs) (n = 189) decreased consistently with age. The prevalence of high-grade squamous intraepithelial lesions (HSILs) (n = 128) peaked first around age 30 years and again at age 65 years or older. Seventy-three percent of LSILs were HPV positive, with HPV16 being the predominant type (16% of positive subjects). HPV was found in 89% of HSILs and 88% of cancers, with HPV16 being strongly predominant (51% and 53% of positive subjects). Virtually all HSILs and cancers had cancer-associated HPV types, with high odds ratios (ORs) and attributable fractions around 80%. Risk for HPV16 was particularly high (OR for HSILs = 320, 95% confidence interval [CI] = 97-1000; OR for cancer = 710, 95% CI = 110-4500)., Conclusions: We confirm the early decline of HPV infection with age but note increased prevalence after menopause, which could be related to a second peak of HSILs, an observation that warrants further investigation. At least 80% of HPVs involved in cervical carcinogenesis in this population have been characterized. Polyvalent vaccines including the main cancer-associated HPV types may be able to prevent most cases of cervical disease in this region.

Context: Human papillomaviruses (HPVs) are known to cause most cervical cancer worldwide, but the utility of HPV DNA testing in cervical cancer prevention has not been determined., Objective: To provide comprehensive data on the screening performance of HPV testing for the most common carcinogenic types, at different levels of analytic sensitivity., Design: Laboratory analysis conducted during 1993-1995, using 3 cytologic techniques and cervicography, followed by colposcopic examination of women with any abnormal cervical finding, to detect all high-grade intraepithelial lesions and cancer (reference standard of clinically significant disease). The HPV testing was performed subsequently with masking regarding clinical findings., Setting: Guanacaste Province, Costa Rica, a region with a high age-adjusted incidence of cervical cancer., Participants: Of 11742 randomly selected women, 8554 nonpregnant, sexually active women without hysterectomies underwent initial HPV DNA testing using the original Hybrid Capture Tube test; a stratified subsample of 1119 specimens was retested using the more analytically sensitive second generation assay, the Hybrid Capture II test., Main Outcome Measures: Receiver operating characteristic analysis of detection of cervical high-grade intraepithelial lesions and cancer by HPV DNA testing based on different cut points of positivity., Results: An analytic sensitivity of 1.0 pg/mL using the second generation assay would have permitted detection of 88.4% of 138 high-grade lesions and cancers (all 12 cancers were HPV-positive), with colposcopic referral of 12.3% of women. Papanicolaou testing using atypical squamous cells of undetermined significance as a cut point for referral resulted in 77.7% sensitivity and 94.2% specificity, with 6.9% referred. Specificity of the second generation assay for positivity for high-grade lesions and cancer was 89.0%, with 33.8% of remaining HPV DNA-positive subjects having low-grade or equivocal microscopically evident lesions. The higher detection threshold of 10 pg/mL used with the original assay had a sensitivity of 74.8% and a specificity of 93.4%. Lower levels of detection with the second generation assay (<1 pg/mL) proved clinically nonspecific without gains in diagnostic sensitivity., Conclusions: In this study population, a cut point of 1.0 pg/mL using the second generation assay permitted sensitive detection of cervical high-grade lesions and cancer, yielding an apparently optimal trade-off between high sensitivity and reasonable specificity for this test. The test will perform best in settings in which sensitive detection of high-grade lesions and cancer is paramount. Because HPV prevalence varies by population, HPV testing positive predictive value for detection of high-grade lesions and cancer will vary accordingly, with implications for utility relative to other cervical cancer screening methods.

Background: Human papillomavirus (HPV) infection has been strongly associated with cervical carcinoma and its cytologic precursors, squamous intraepithelial lesions (SIL). We investigated the risk of SIL prospectively following polymerase chain reaction (PCR)-based DNA testing for a wide range of genital HPV types in a cohort of initially cytologically normal women, to clarify the role of HPV in the etiology of SIL., Methods: Starting in April 1989, 17,654 women who were receiving routine cytologic screening at Kaiser Permanente (Portland, OR) were followed for the development of incident SIL. During follow-up, 380 incident case patients and 1037 matched control subjects were eligible for this nested case-control study. Cervical lavages collected at enrollment and, later, at the time of case diagnosis (or the corresponding time for selection of control subjects) were tested for HPV DNA using a PCR-based method. The data were analyzed as contingency tables with two-sided P values or, for multivariable analyses, using odds ratios (ORs) with 95% confidence intervals (CIs)., Results: In comparison with initially HPV-negative women, women who tested positive for HPV DNA at enrollment were 3.8 times (95% CI = 2.6-5.5) more likely to have low-grade SIL subsequently diagnosed for the first time during follow-up and 12.7 times more likely (95% CI = 6.2-25.9) to develop high-grade SIL. At the time of diagnosis, the cross-sectional association of HPV DNA and SIL was extremely strong (OR = 44.4 and 95% CI = 24.2-81.5 for low-grade SIL and OR = 67.1 and 95% CI = 19.3-233.7 for high-grade SIL). HPV16 was the virus type most predictive of SIL, even low-grade SIL., Conclusions: These findings are consistent with the hypothesis that HPV infection is the primary cause of cervical neoplasia. Furthermore, they support HPV vaccine research to prevent cervical cancer and efforts to develop HPV DNA diagnostic tests.

Human papillomavirus (HPV), particularly HPV 16, is linked to the development of cervical cancer. However, the role of HPV 16 in a number of extra-cervical epithelial tumours is controversial. To assess exposure to HPV 16 in patients with different cancers, we conducted a large serosurvey of 905 patients with 21 types of tumours and measured IgG to HPV 16 virus-like particles (VLPs) using a well characterized enzyme linked immunosorbent assay (ELISA). Patients with cervical cancer were considered 'positive controls', as about half were expected to be specifically HPV 16-related. A non-cancer study group consisting of 48 patients with endocrine disorders (eg diabetes) was also tested. HPV 16 antibody prevalence was highest in patients with cancers of the cervix (52% +/- 7%), vulva (27% +/- 9%), vagina (27% +/- 13%) and penis (63% +/- 16%). Seroprevalence was much lower in the non-cancer group (4% +/- 3%) and all other cancer patients: uterus (9% +/- 4%); ovary (4% +/- 3%); testis (5% +/- 4%); prostate (6% +/- 4%); squamous carcinoma (6% +/- 3%) and adenocarcinoma (4% +/- 3%) of the lung; rectum (2% +/- 2%); pancreas (8% +/- 4%); colon (2% +/- 2%); stomach (0%); oesophagus (8% +/- 4%); buccal cavity (12% +/- 5%); breast (10% +/- 4%); non-melanomatous (9% +/- 6%) and melanomatous (6% +/- 3%) skin tumours; bladder (8% +/- 4%); and kidney (2% +/- 2%). The results confirm the strong relation of HPV with several lower anogenital tract tumours, but, at least in this population, fail to identify additional epithelial tumours associated with high seroprevalence of HPV 16 VLP antibodies.

Several earlier case-control studies reported inverse associations of cervical squamous intraepithelial lesions (SIL) with high dietary or biomarker levels of carotenoids, folate, and vitamins C and E. However, most studies did not measure the primary causal factor, cancer-associated genital human papillomaviruses (HPV), now detected by sensitive viral DNA tests. This nested case-control study assessed whether high dietary intakes of these nutrients, plus zinc and vitamin A, reduced SIL risk in cancer-associated HPV DNA-positive women. Using a 60-item food-frequency questionnaire, nutrient estimates were obtained for 33 incident cases with high-grade lesions, 121 with low-grade lesions, 97 with equivocal SIL, and 806 cytologically normal controls sampled from a large prospective cohort study. Baseline cervicovaginal lavages were tested for HPV DNA by the polymerase chain reaction. Among DNA-positive cases (n = 68) and controls (n = 69), age-adjusted odds ratios (ORs) of SIL in the highest vs. the lowest nutrient quartiles were 1.4 [95% confidence interval (CI) = 0.5-4.2] for vitamin A, 0.6 (CI = 0.2-2.0) for beta-carotene, 1.3 (CI = 0.4-3.6) for vitamin C, 1.0 (CI = 0.4-3.6) for vitamin E, 0.7 (CI = 0.3-2.1) for folate, and 0.8 (CI = 0.3-2.2) for zinc. ORs in HPV DNA-negative women approximated 1.0, with the exception of vitamin E (OR = 0.5, CI = 0.3-0.9). These results do not support a protective role for the above nutrients against low-grade or equivocal SIL, which constituted the majority of diagnoses in this study.

Serological assays for measuring antibodies to human papillomavirus type 16 (HPV-16) virus-like particles (VLPs) have become important epidemiologic tools in recent years. However, the interlaboratory replicability of these assays has not been assessed. In this investigation, three laboratories tested a panel of specimens obtained from two different groups: 265 subjects in a vulvar cancer case-control study and 107 healthy volunteer blood donors. Each laboratory used an enzyme-linked immunosorbent assay (ELISA), but no attempt was made to standardize assay procedures among the three laboratories. The data showed good day-to-day intralaboratory replicability in laboratory 1 (correlation coefficient, > or = 0.88) and good intra-assay variability in laboratory 3 (correlation coefficient, > or = 0.93). Interlaboratory correlations, likewise, ranged between 0.61 and 0.80 in both case-control study subjects and healthy blood donors, indicating that ELISA optical density (OD) values between laboratories were linearly related regardless of the population. Kappa coefficients (kappa), based on each laboratory's categorical interpretation of its results (as positive or negative), showed good agreement (kappa, > 0.6) in case-control study subjects and moderate agreement (kappa, > or = 0.4) in blood donors, a population that had few strongly positive sera. When OD values near seropositive cutoffs were treated as indeterminates, there was little discordance between laboratories in either population. The data suggest that each laboratory measured the same humoral immune response and that their HPV-16 VLP ELISAs performed similarly (Pearson correlations). Interlaboratory differences, however, probably due to reagents and procedures, were considerably greater than intralaboratory day-to-day variability. Interlaboratory agreement in determining seropositivity (kappa) could be improved by sharing positive and negative serum controls and by treating marginal results as indeterminate. As part of continuing cooperation to improve interlaboratory agreement, we are preparing bulk serum control specimens to be shared and made available to interested researchers.

This paper reports on the enrollment phase of a population-based natural history study of cervical neoplasia in Guanacaste, a rural province of Costa Rica with consistently high rates of invasive cervical cancer. The main goals of the study are to investigate the role of human papillomavirus (HPV) infection and its co-factors in the etiology of high-grade cervical neoplasia, and to evaluate new cervical cancer screening technologies. To begin, a random sample of censal segments was selected and enumeration of all resident women 18 years of age and over was conducted with the aid of outreach workers of the Costa Rican Ministry of Health. Of the 10738 women who were eligible to participate, 10049 (93.6%) were interviewed after giving written informed consent. After the interview on cervical cancer risk factors was administered, a pelvic examination was performed on those women who reported previous sexual activity. The pelvic examination included a vaginal pH determination and collection of cervical cells for cytologic diagnosis using three different techniques. Additional cervical cells were collected for determination of the presence and amount of DNA from 16 different types of HPV, and two photographic images of the cervix were taken and interpreted offsite by an expert colposcopist. Finally, blood samples were collected for immunologic and micronutrient assays. Women with any abnormal cytologic diagnosis or a positive Cervigram, as well as a sample of the whole group, were referred for colposcopy, and biopsies were taken when lesions were observed. The enrollment screening will serve as the basis for a prevalent case-control study, and the members of the cohort free from serious disease will be followed actively, at intervals of no more than a year, to study the natural history of HPV infection and the origins of high-grade squamous intraepithelial lesions (HSIL). Details of the field operation are outlined, with particular reference to the realization of this kind of study in developing countries. Descriptive data on the prevalence of disease and exposure to various risk factors are also presented.