Acquisition of Anal Human Papillomavirus (HPV) Infection in Women: the Hawaii HPV Cohort Study

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1 MAJOR ARTICLE Acquisition of Anal Human Papillomavirus (HPV) Infection in : the Hawaii HPV Cohort Study Marc T. Goodman, 1 Yurii B. Shvetsov, 1 Katharine McDuffie, 1 Lynne R. Wilkens, 1 Xuemei Zhu, 1 Lily Ning, 2 Jeffrey Killeen, 4 Lori Kamemoto, 3 and Brenda Y. Hernandez 1 1 Cancer Research Center of Hawaii, 2 University Health Services, and 3 John A. Burns School of Medicine, University of Hawaii, and 4 Kapiolani Medical Center for and Children, Honolulu, Hawaii Background. The majority of anal cancer is associated with human papillomavirus (HPV) infection, yet little is known about women s risk of acquisition of anal HPV infection. Methods. Risk factors for the acquisition of anal HPV infection were examined in a longitudinal cohort study of 431 women, via repeated measurement of HPV DNA. Results. Seventy percent of women were positive for anal HPV infection at one or more clinic visits from baseline through a follow-up period that averaged 1.3 years. The incidence of a high-risk (HR) infection was 19.5 (95% confidence interval [CI], ) per 1000 woman-months. The most common incident HR HPV types were HPV-53, -52 and -16. The presence of an HR anal HPV infection at baseline increased the risk of an incident anal infection by 65%. Baseline HR cervical HPV infection also predicted the acquisition of an HR anal HPV infection (odds ratio, 1.81 [95% CI, ]). Nonviral risk factors for acquisition of HR HPV infection included younger age, lower socioeconomic status, greater lifetime number of sexual partners, past use of hormones, and condom use. Conclusions. The results of this study suggest that women s risk of anal HPV infection is as common as their risk of cervical HPV infection. Received 14 September 2007; accepted 2 November 2007; electronically published 4 March Potential conflicts of interest: none reported. Financial support: National Cancer Institute (grant CA to M.T.G.); Research Centers in Minority Institutions (award P20 RR11091 from the National Center for Research Resources, National Institutes of Health, to R. Yanagihara). Reprints or correspondence: Dr. Marc T. Goodman, Etiology Program, Cancer Research Center of Hawaii, University of Hawaii, 1236 Lauhala St., Honolulu, HI (marc@crch.hawaii.edu). The Journal of Infectious Diseases 2008; 197: by the Infectious Diseases Society of America. All rights reserved /2008/ $15.00 DOI: / Although anal cancer is an uncommon malignancy, occurring at a rate of 1/100,000, its incidence in the United States and other countries has increased during the past several decades [1 4]. Anal cancer is one of the few malignancies without a known hormonal etiology that is more frequent among women than among men [2]. During the past decade, epidemiological studies using state-of-the-art detection techniques have shown that the majority of anal cancers in both sexes are associated with human papillomavirus (HPV) infection [5]. In a Danish study, Frisch et al. [6] reported the presence of HPV in 93% of anal tumors among women, and 77% of these anal tumors were HPV-16, the most common high-risk (HR) HPV type associated with cervical cancer. In a US study, similar results were found by Daling et al. [7], detected HPV in 92% of anal-canal tumor specimens from women, with 74% of these specimens being positive for HPV-16. Not surprisingly, the risk of anal cancer is elevated in women with cervical and vulvar cancers, presumably because of a shared exposure to HR HPV infections [8, 9]. The anal canal shares with the cervix histological characteristics that may be due to a common etiology. Similar to what has been observed for cervical cancer, the majority of anal cancers develop in the transition zone from the glandular mucosa to the squamous mucosa of the rectum [10]. Furthermore, the natural history of anal malignancy has close parallels to that of cervical malignancy, in the initial development of a precursor dysplastic condition known as anal intraepithelial neoplasia [11]. Most investigations of the natural history of anal HPV disease have been limited to men, whereas investigations of women have largely studied HIV-infected cases. Of the few investigations of HPV disease in women, that by Moscicki et al. [12] found that 67% of study participants with abnormal anal cytology were positive for HPV, but it did not report the prevalence of HPV in cytologically normal women. Palefsky et al. [11] found that 42% of Acquisition of Anal HPV JID 2008:197 (1 April) 957

2 high-risk HIV-negative women were positive for anal HPV infection, higher than the 27% prevalence found in our earlier, baseline assessment of healthy women in Hawaii [13]. The objective of the present analysis was to examine the natural history of anal HPV infection in a longitudinal study of women in Hawaii. SUBJECTS AND METHODS Recruitment of subjects. Between 1998 and 2003, we conducted a longitudinal study of cervical and anal HPV infection [13, 14] in women attending 5 clinics on Oahu, Hawaii, were able to read, understand, and sign an informed-consent form and a medical-release form approved by the University of Hawaii Institutional Review Board. Potentially eligible patients include those with appointments either for new or annual gynecologic examinations and cervical cytological (Pap) smears or for family-planning services. At the first and subsequent visits, an exfoliated cervical cell specimen was obtained to test for HPV DNA. The collection of an anal specimen after gynecologic examination was optional and was obtained by use of a Dacron swab moistened with sterile water among willing women. The swab was inserted cm into the anus and was rotated 360 degrees both clockwise (5 times) and counterclockwise (5 times). The swab was placed in 1.0 ml of buffered medium (Digene). After completion of the examination, the study coordinator administered a study questionnaire to each participant. The baseline interview was concerned with demographic data and with a detailed history of tobacco and alcohol use. Selection and follow-up of cohort. The results of the baseline cervical smear and HPV-DNA testing were necessary to establish the women s final eligibility for participation in the study. se specimens were inadequate (i.e., were negative for the human -globin gene) at baseline were excluded; women were enrolled in the study were asked to return to the clinic every 4 months, for examination and testing. were treated for a cervical abnormality were excluded from the study. A more detailed interview was conducted during the second visit. This second questionnaire was concerned with gynecological, menstrual, reproductive and sexual history; hormone use; medical history; history of sexually transmitted infections; and income. Information collected during the first, baseline questionnaire (e.g., data regarding sexual activity and tobacco and alcohol use) also was updated. The questionnaire used at subsequent interviews was modified slightly for use during the follow-up period; questions focused on changes in sexual and reproductive information during the intervening period between clinic visits and checks on the reliability of certain data items. All women were treated for a squamous intraepithelial lesion or a more severe lesion (e.g., via conization) were removed from the study when they were treated and were not included in subsequent follow-up. Laboratory analyses. HPV DNA was extracted from exfoliated cervical cell specimens by use of commercial reagents (Qiagen). Specimens were analyzed for the presence or absence of HPV DNA by PCR using a modified version of the PGMY09/ PGMY11 primer system [15]. HPV DNA-positive specimens were genotyped, by use of a reverse-line-blot detection method [16], for 36 different HPV types, including HR HPV types -16, -18, -31, -33, -35, -39, -45, -51, -52, -53, -56, -58, -59, -66, -68, -70, -73, and -82; low-risk (LR) HPV types -6, -11, -42, -54, -61, -72, -84, and CP6108; and undetermined-risk HPV types -55, -62, -67, -71, -83, and -84 [17, 18] (Roche Molecular Systems). We defined the risk (oncogenic potential) associated with the various HPV types according to the International Agency for Research on Cancer definition. HPV-positive specimens that were subsequently found to be negative in the genotyping assay were considered to be unclassified HPV-positive specimens. All specimens were also for the human -globin gene, as an internal control for sample sufficiency. Specimens that negative for -globin were considered to be insufficient and were excluded from analyses. Statistical analysis. All analyses were limited to the 431 women completed both the detailed questionnaire during visit 2 and at least 2 clinic visits. The Kaplan-Meier method was used to construct cumulative incidence curves grouped by oncogenic risk and phylogenetic species, where the time to event was defined as the time from entry into the study until acquisition of the first incident anal HPV infection from a specified group. HPV type-specific incidence rates per 1000 womanmonths were calculated for all detected HPV genotypes, as well as grouped by the number of other genotypes present at the time of acquisition of the index infection. Woman-months of follow-up were calculated by adding all time intervals during which a woman was negative for a specific HPV type or group. For example, if a woman had acquired a new HPV-16 infection by visit 3, the time from entry into the study until visit 3 would be added to the follow-up time; if a woman was infected with HPV-16 at entry into the study and acquired HPV-18 by visit 2, this would not be counted as a new infection when the incidence rate of HR HPV was computed. Poisson exact confidence intervals (CIs) were constructed for all incidence rates [19]. The association between incident HPV infection and factors of interest was modeled by unconditional logistic regression, in which women with no incident HPV infection were considered to be nonevents. Odds ratios and 95% CIs were calculated. Incident HPV infections were classified according to oncogenic risk, as follows: all infections, HR, LR, and LR or unknown genotypes. Baseline HPV status was classified as either HR only, LR only, or both HR and LR (including and excluding the unknown types) and separately according to HPV genotype. All models were adjusted for age of the participants at entry into the study. Other adjustment factors were considered, but their inclusion in the models did not result in a significantly better fit as determined by 958 JID 2008:197 (1 April) Goodman et al.

3 Table 1. Prevalence at enrollment, incidence rates, and cumulative positivity of human papillomavirus (HPV) infection of the anus, stratified by genotype and number of coinfections, in 431 women. HPV category Prevalence at entry into study, no. (%) of women Follow-up, womanmonths a Cumulative incidence, no (%) of women Prevalence during follow-up, no. (%) of women b No. All incident infections No coinfections c coinfections c One or more Incidence, rate/1000 woman-months (95% CI) No. Incidence, rate/1000 woman-months (95% CI) No. Incidence, rate/1000 woman-months (95% CI) Genotype Any 183 (42.5) (49.9) 303 (70.3) ( ) ( ) ( ) High risk Any 96 (22.3) (25.5) 171 (39.7) ( ) ( ) ( ) (4.4) (3.7) 33 (7.7) ( ) ( ) ( ) 18 9 (2.1) (2.8) 21 (4.9) ( ) ( ) ( ) 31 5 (1.2) (1.9) 12 (2.8) ( ) ( ) ( ) 33 0 (0.0) (0.2) 1 (0.2) ( ) ( ) ( ) 35 3 (0.7) (0.5) 4 (0.9) ( ) ( ) ( ) (2.6) (1.4) 17 (3.9) ( ) ( ) ( ) 45 2 (0.5) (0.9) 6 (1.4) ( ) ( ) ( ) (3.7) (3.2) 24 (5.6) ( ) ( ) ( ) (3.0) (3.7) 29 (6.7) ( ) ( ) ( ) (3.5) (5.3) 36 (8.4) ( ) ( ) ( ) 56 6 (1.4) (3.0) 18 (4.2) ( ) ( ) ( ) 58 4 (0.9) (1.9) 11 (2.6) ( ) ( ) ( ) 59 5 (1.2) (1.6) 11 (2.6) ( ) ( ) ( ) 66 9 (2.1) (1.4) 14 (3.2) ( ) ( ) ( ) 68 3 (0.7) (1.6) 9 (2.1) ( ) ( ) ( ) 70 9 (2.1) (2.1) 17 (3.9) ( ) ( ) ( ) 73 2 (0.5) (2.1) 10 (2.3) ( ) ( ) ( ) 82 3 (0.7) (0.5) 5 (1.2) ( ) ( ) ( ) Low risk d Any 53 (12.3) (39.0) 90 (20.9) ( ) ( ) ( ) 6 8 (1.9) (1.6) 15 (3.5) ( ) ( ) ( ) 11 2 (0.5) (0.2) 3 (0.7) ( ) ( ) ( ) 42 5 (1.2) (1.6) 12 (2.8) ( ) ( ) ( ) (2.6) (2.1) 19 (4.4) ( ) ( ) ( ) (2.3) (3.2) 21 (4.9) ( ) ( ) ( ) 72 2 (0.5) (0.0) 2 (0.5) ( ) ( ) ( ) 81 6 (1.4) (2.1) 14 (3.2) ( ) ( ) ( ) CP (4.9) (1.9) 28 (6.5) ( ) ( ) ( ) Undetermined risk 55 7 (1.6) (1.2) 12 (2.8) ( ) ( ) ( ) (4.2) (2.8) 30 (7.0) ( ) ( ) ( ) 67 2 (0.5) (0.5) 4 (0.9) ( ) ( ) ( ) 71 4 (0.9) (1.2) 7 (1.6) ( ) ( ) ( ) (2.3) (2.3) 18 (4.2) ( ) ( ) ( ) (3.2) (4.4) 33 (7.7) ( ) ( ) ( ) Species e 1 25 (5.8) (3.2) 38 (8.8) ( ) ( ) ( ) 3 53 (12.3) (11.6) 95 (22.0) ( ) ( ) ( ) 5 18 (4.2) (3.5) 27 (6.3) ( ) ( ) ( ) 6 29 (6.7) (8.8) 63 (14.6) ( ) ( ) ( ) 7 37 (8.6) (7.9) 68 (15.8) ( ) ( ) ( ) 9 40 (9.3) (9.3) 74 (17.2) ( ) ( ) ( ) (3.9) (2.6) 28 (6.5) ( ) ( ) ( ) NOTE. CI, confidence interval. a For women at risk of acquisition of an infection of the specified type. b Includes prevalence at baseline. c Other HPV genotypes detected at time of acquisition of the index infection. d Includes undetermined-risk HPV types. e -Papillomavirus species; species 1 comprises types 42 and CP6108; species 3 comprises types 61, 62, 72, 81, 83, and 84; species 5 comprises types 26, 51, and 82; species 6 comprises types 53, 56, and 66; species 7 comprises types 18, 39, 45, 59, 68, and 70; species 9 comprises types 16, 31, 33, 35, 52, 58, and 67; and species 10 comprises types 6, 11, and 55.

4 the likelihood-ratio test. Baseline risk factors for acquisition of an anal HPV infection included a variety of sociodemographic, life-style, and reproductive variables. P values for trend were calculated from the Wald 2 statistic for all interval variables; for the categorical levels, the median value was used. All P values were 2-sided; P.05 was considered to be significant. RESULTS Characteristics of the cohort. A total of 650 women were recruited, for anal HPV DNA by PCR, and completed at least 1 follow-up visit. Of these 650 women, 431 (66%) contributed at least 2 -globin positive anal specimens. Of the remaining 431 women, 215 (50%) experienced at least 1 incident anal HPV infection, defined as an HPV genotype not identified on the previous visit, and 177 (41%) were HPV negative at all visits. Baseline and follow-up analyses included 1508 visits in which anal specimens were collected (median, 3.5 visits/woman). The cumulative follow-up experience for this cohort was 7004 woman-months (mean, 487 days). The cohort composition was multiethnic, including women of white (42%), Hawaiian (14%), Japanese (12%), Filipino (6%), and mixed (26%) ethnicity. The median age of the cohort was 40 years (mean, 39.4 years). Only 14% of the women were current tobacco smokers, and 23% were current alcohol drinkers at baseline. Cumulative probability of incident HPV infection. The prevalence of an anal HPV infection at enrollment was 42% (183 HPV-positive cases), with more than half of these being HR types (table 1). A total of 303 women were positive for anal HPV at least once, yielding a crude cumulative rate of 70%, including baseline infections. The incidence of HR HPV types (19.5/1000 woman-months) was more than twice that of LR HPV types (8.2/1000 woman-months). HR HPV types HPV-16, -51, -52, -53 were the most common high-risk types at entry into the study. The cumulative incidence of HR HPV infections rose significantly (P.001) more rapidly than the incidence of LR HPV infections (figure 1). Acquisition of anal HPV infection was most rapid for -papillomavirus species 3, followed by species 9, species 6, and species 7. Acquisition of an incident anal HPV infection: viral factors. The risk of acquisition of any anal HPV infection was 46.9 (95% CI, ) per 1000 woman-months (table 1). The most commonly acquired viral type in the anal canal was HPV-53 (3.7/1000 woman-months), followed by HPV-84 (2.8/1000 woman-months). The prevalence of multiple anal HPV-type infections was 15.8% at baseline and averaged 11.4% at subsequent visits. Overall, the presence of 1 HPV type did not increase the risk of acquisition of another HPV type, but some differences were found Figure 1. Cumulative incidence of anal human papillomavirus (HPV) infections, stratified by (A) oncogenic risk and (B) 4 -papillomavirus species: species 3 comprises types 61, 62, 72, 81, 83, and 84; species 6 comprises types 53, 56, and 66; species 7 comprises types 18, 39, 45, 59, 68, and 70; and species 9 comprises types 16, 31, 33, 35, 52, 58, and 67. the log-rank test result for low-risk HPV vs. high-risk HPV was P.001, and the log-rank test results for species 6, 7, and 9 vs. species 3 were P.0201,.0120, and.0811, respectively. in the type-specific analyses (table 1). The risk of acquisition of any anal HPV infection was 37.3/1000 woman-months (95% CI, ) in women with no coinfection and 9.6/1000 woman-months (95% CI, ) in women with 1 or more coinfections. The risk of acquisition of a new HPV-16 infection was 2-fold greater in women with 1 or more coinfections than in women with a single infection, although the CIs for the 2 groups overlapped. The only incident HPV-31, -33, -35, and -11 infections occurred in women with a coinfection. The risk of acquisition of a species 9 HPV infection, including HPV-16, -31, -33 and -35, was also significantly greater in women with multiple HPV types than in women with no coinfection. 960 JID 2008:197 (1 April) Goodman et al.

5 Table 2. Baseline human papillomavirus (HPV) status and acquisition of anal HPV infection. Variable, no. no. b High-risk HPV Low-risk HPV a Any HPV Odds ratio (95% CI) c no. b Odds ratio (95% CI) c no. b Odds ratio (95% CI) c Baseline anal-hpv status Any HPV (n 431) No (reference) (reference) (reference) Yes ( ) ( ) ( ) High-risk HPV (n 344) No (reference) (reference) (reference) Yes ( ) ( ) ( ) Low-risk HPV (n 334) No (reference) (reference) (reference) Yes ( ) ( ) ( ) High-risk low-risk HPV (n 295) No (reference) (reference) (reference) Yes ( ) ( ) ( ) Baseline cervical-hpv status Any HPV (n 431) No (reference) (reference) (reference) Yes ( ) ( ) ( ) High-risk HPV (n 409) No (reference) (reference) (reference) Yes ( ) ( ) ( ) Low-risk HPV (n 342) No (reference) (reference) (reference) Yes ( ) ( ) ( ) High-risk low-risk HPV (n 320) No (reference) (reference) (reference) Yes ( ) ( ) ( ) NOTE. CI, confidence interval. a Includes undetermined-risk HPV types. b experienced an incident anal HPV infection. c Adjusted for age of participants at entry into the study. The presence of an HR anal HPV infection at baseline increased the risk of acquisition of any HPV type by 65% (95% CI, 1% 170%), and the presence of an LR anal HPV infection increased the risk of acquisition of another HPV type by 80% (95% CI, 8% 200%) (table 2). The risk of acquisition of an HR anal HPV infection was also increased significantly in women with a cervical HPV infection at baseline: women positive for any cervical HPV at baseline had a 91% (95% CI, 18% 209%) greater risk of an incident HR anal HPV infection than did women were negative for any cervical HPV. Of new anal HPV infections, 46% occurred in women with no baseline or concurrent cervical infection (data not shown). Acquisition of an incident anal HPV infection: nonviral factors. A significant inverse relation was found for the acquisition of a new HR anal HPV infection and age: women were 45 years of age at baseline had a 57% (95% CI, 19% 77%) lower risk of acquisition of an HR anal HPV infection than did women were 25 years at baseline (table 3). White women represented the largest ethnic subgroup (42%), and they also were at greatest risk of acquisition of an anal HPV infection; Japanese women had the lowest risk of acquisition of an anal HPV infection. Indicators of low socioeconomic status, such as homelessness, limited education, and low income, tended to be associated with an increased risk of acquisition of anal HPV, especially HR types. Reproductive characteristics were not associated with the risk of an incident HR anal HPV infection. Acquisition of Anal HPV JID 2008:197 (1 April) 961

6 Table 3. Baseline risk factors for acquisition of anal human papillomavirus (HPV) infection. High-risk HPV (n 110) Low-risk HPV a (n 87) Any HPV (n 215) Factor Total b (n 431) Age 25 years (reference) (reference) (reference) years ( ) ( ) ( ) years ( ) ( ) ( ) 45 years ( ) ( ) ( ) 0.43 Ethnicity Japanese (reference) (reference) (reference) White ( ) ( ) ( ) Hawaiian ( ) ( ) ( ) Filipino ( ) ( ) ( ) Other ( ) ( ) ( ) Education High school or less (reference) (reference) (reference) Some college ( ) ( ) ( ) College graduate ( ) ( ) ( ) Graduate degree ( ) ( ) ( ) 0.34 Place of birth United States (reference) (reference) (reference) Other ( ) ( ) ( ) Homeless Never (reference) (reference) (reference) Ever ( ) ( ) ( ) Income $ (reference) (reference) (reference) $ , ( ) ( ) ( ) $20,000 49, ( ) ( ) ( ) $50, ( ) ( ) ( ) 0.46 Pregnancies (reference) (reference) (reference) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) 0.27 Age at first sexual intercourse 16 years (reference) (reference) (reference) years ( ) ( ) ( ) years ( ) ( ) ( ) 20 years ( ) ( ) ( ) 0.37 Lifetime sex no. of partners (reference) (reference) (reference) ( ) ( ) ( ) ( ) ( ) ( ) Oral contraceptive use at baseline Never (reference) (reference) (reference) Ever ( ) ( ) ( ) Past user ( ) ( ) ( ) Current user ( ) ( ) ( ) (continued)

7 Table 3. (Continued) High-risk HPV (n 110) Low-risk HPV a (n 87) Any HPV (n 215) Factor Total b (n 431) Oral-contraceptive pill use Never (reference) (reference) (reference) 2 years ( ) ( ) ( ) 2 4 years ( ) ( ) ( ) 5 9 years ( ) ( ) ( ) 10 years ( ) ( ) ( ) Noncontraceptivehormone use Never (reference) (reference) (reference) Ever ( ) ( ) ( ) Past user ( ) ( ) ( ) Current user ( ) ( ) ( ) History of tobacco smoking Never (reference) (reference) (reference) Ever ( ) ( ) ( ) Past user ( ) ( ) ( ) Current user ( ) ( ) ( ) Pack-years of tobacco use e (reference) (reference) (reference) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) 0.98 History of alcohol use Never (reference) (reference) (reference) Ever ( ) ( ) ( ) Past user ( ) ( ) ( ) Current user ( ) ( ) ( ) Lifetime ethanol intake f None (reference) (reference) (reference) 250 drinks ( ) ( ) ( ) drinks ( ) ( ) ( ) 1100 drinks ( ) ( ) ( ) 0.41 Condom use at baseline No (reference) (reference) (reference) Yes ( ) ( ) ( ) History of anal sex Never (reference) (reference) (reference) Ever ( ) ( ) ( ) Past ( ) ( ) ( ) Current ( ) ( ) ( ) a b c d e f Includes undetermined-risk HPV types. completed questionnaire and at least 2 clinical visits. acquired an incident anal HPV infection during the study period. Adjusted for age of participants at entry into the study. Defined as the product of years of smoking and average number of cigarette packs smoked daily. Assessed by the summation of individual drinks of each alcohol type, in which each drink was considered to contain 12 ml of ethanol. Of the sexual variables considered, only lifetime number of sexual partners was predictive of incident HR anal HPV infection. No association between oral contraceptive pill use at baseline and the risk of acquisition of an HR anal HPV infection was found. However, long-term use of oral contraceptives Acquisition of Anal HPV JID 2008:197 (1 April) 963

8 was positively associated with the risk of an incident anal HPV infection, although the trend in risk was not monotonic and was not significant for HR HPV types. A significantly increased risk of incident HR HPV infection was also found for past users, but not present users, of estrogens. Little association between tobacco smoking or alcohol drinking and the risk of acquisition of an HR HPV infection was found. Condom use during sexual intercourse increased the risk of incident HR HPV infection but not the risk of acquisition of an LR HPV infection. A history of anal sex increased the risk of acquisition of any anal HPV type, especially in women had recently engaged in this sexual practice; however, no significant association was found between anal sex and the risk of acquisition of an HR HPV infection. DISCUSSION In the present study of the natural history of anal HPV infection in adult women, 70% of the participants had at least 1 anal HPV infection during the average 1.3-year follow-up period. We found that the acquisition of a new infection was a relatively common event: the incidence of a new infection was almost 50/ 1000 woman-months, compared with the incidence rate of 30/ 1000 woman-months that Trottier and Franco [20] estimated for cervical HPV infection in their summary of cohort studies that used polymerase chain reaction for detection of HPV DNA. Although these estimates are dependent on a number of factors, such as age, sexual activity, and geographic area, the data reported in the present study suggest that women have a high cumulative risk of acquisition of anal HPV infection. The risk of an incident HR anal HPV infection was 19.5/1000 woman-months, compared with a risk of 8.2/1000 womanmonths for an incident LR anal HPV infection. The greater incidence of HR HPV infection is consistent with the findings of natural history studies of cervical HPV infection [20]. In a cohort study of Canadian female university students, Richardson et al. [21] reported that the incidence of HR cervical HPV infection was 14.0/1000 woman-months for HR HPV and that of LR cervical HPV infection was 12.4/1000 woman-months. In a study of another young cohort, comprising year-old American women, Giuliano et al. [22] reported that the 1-year cumulative risk of HR cervical HPV infection was 32%, compared with 18% for LR cervical HPV infection. Although these studies are not directly comparable, because of differences in their participants risk behaviors, their results suggest that the rates of acquisition of anal and cervical HPV infections are similar. In the present study, HPV-16 was the most common oncogenic type, affecting 4.4% of all cohort members at enrollment and having a period prevalence of 7.7% during the study. This relative frequency of HPV-16 is similar to that reported by most studies of HPV infection in women with normal cervical cytology, worldwide [23]. HPV-51, -53, and -52 were the next most common types infecting the anal canal. These results are in accord with those of prevalence studies of cervical infection in cytologically normal women in North America, in m HPV-53 and -51 are the second and fourth most common infections, and in cytologically normal women in Japan and Taiwan, in m HPV-52 and -51 are the first and third most frequent infections [24]. Anal coinfection with multiple HPV types was relatively common in the cohort in the present study. The risk of acquisition of a new HPV type was increased significantly by the presence of anal HPV infection at baseline, although not necessarily by a concurrent new infection. Specific viral types, such as HPV-16 and other types within species 9 of the -papillomavirus genus, were positively associated with the risk of a subsequent HPV infection. The relative-risk estimates reported in the present study could have been biased by the overlapping of exposure and outcome categories for example, a woman with a baseline HPV-16 anal infection was not at risk of acquisition of HPV-16 infection until the first infection had cleared. The present study s findings could have been confounded by the correlation between preexisting HPV infection and higher-risk sexual behavior. When we adjusted for other variables associated with acquisition of HPV infection, such as number of sexual partners, the risk estimates did not change. However, residual confounding by sexually related factors might still exist. The results of the present study do not support the notion of competition between viral types. The present study s findings are in agreement with those reported for the Ludwig-McGill cohort of cervical HPV, in showing that the acquisition of an HPV infection is increased in women in m any type of HPV infection is detected at baseline [24]. In this and other studies of the natural history of HPV infection of the cervix, the risk of acquisition of a new HPV infection was found to be increased in women previously had harbored or concurrently were harboring an HPV-16 or -18 infection [24 26]. Nonviral risk factors for the acquisition of an oncogenic anal HPV infection included young age, ethnicity, low socioeconomic status, greater lifetime number of sexual partners, past estrogen use, and condom use. The data of the present study suggest that age has a strong influence on the acquisition of anal HPV infection. This finding is concordant with studies of cervical HPV infection, which show a continuous decline in prevalence with increasing age [20, 23], perhaps reflecting reduced exposure to new partners. The positive association between lifetime number of sexual partners and the incidence of HR anal HPV infection supports the notion that sexual intercourse is the primary route of anal infection. Indeed, Frisch et al. [6] have reported that anal cancer is elevated in women had 10 or more lifetime sexual partners and in women had a history of receptive anal intercourse at 30 years of age. The positive association found by the 964 JID 2008:197 (1 April) Goodman et al.

9 present study is consistent with observations that having multiple sexual partners is the strongest determinant of the incidence of cervical HPV [27]. However, as was found in other studies [11, 28], a history of anal intercourse was not a significant risk factor for incident HR anal HPV infection. This, combined with the positive association between cervical and anal HPV infections, suggests that other sexual and nonsexual routes of transmission are possible. Nonpenetrative sex, including the use of fingers and sex toys, may increase the risk of anal transmission and may account for HPV transmission in lesbian couples [29]. Piketty et al. [30] found that, in heterosexual HIV-positive men, anal HPV infection and anal dysplasia may be acquired in the absence of anal intercourse. Moscicki et al. [12] speculated that HPV shed in cervical or vaginal discharge may be a risk factor for transmission of the virus to the anus. It is also possible that responses to our questions regarding anal sex were less than candid, which would contribute to a false-negative association. Although condom use may afford some protection against cervical neoplasia and cancer, it is unclear whether condoms reduce the risk of HPV transmission to the cervix [31]. The association between condom use and the risk of acquisition of anal HPV might be explained by use of the same condom during both vaginal intercourse and postcoital anal sex. This possibility is supported by the significant association between baseline cervical HPV infection and the risk of incident anal HPV. Unfortunately, we did not ask our subjects whether they used condoms during anal sex, so this conclusion is speculative. Several case-control studies [7, 32, 33] have reported that a history of tobacco smoking increases the risk of anal cancer, but we did not find an association between tobacco smoking and the risk of acquisition of an anal HPV infection. As with cervical cancer, it is likely that risk factors for acquisition of HPV in the anus may be distinct from those for anal cancer. The use of oral contraceptives has been found to increase the risk of cervical cancer and its precursors [18, 34, 35], although the results of a pooled analysis were unsupportive of a positive association [36]. We found little evidence for an association between oral-contraceptive use and the risk of acquisition of anal HPV, although past users of noncontraceptive hormones were at elevated risk. The sample population in the present study included women from both a university setting and a health maintenance organization in Honolulu, so the results may not be generalizable to the overall population. Furthermore, because collection of anal specimens was optional, only 66% of women agreed to participate in collection of both cervical and anal specimens [13]. A shortcoming that the present study shares with other investigations is the assumption that HPVs of the same genotype that are present at consecutive visits are part of the same infection. In studies of the association between coinfection and risk, such as ours, this assumption is conservative, because coinfections with the same HPV type are not considered. Analysis of sequence variants would permit distinction between the same or a different infection with the same genotype and may be included in future investigations. Exfoliation of cells from the perianal region may have contaminated our anal specimens and may partially explain the absence of an association between anal HPV infection and anal sex, although we had no means of examining this possibility. Swabs from participants backs, as well as from the examination table, were routinely taken as clinical and environmental controls to monitor possible HPV contamination during the collection process; HPV DNA was not detected in any of the clinical or environmental control specimens. Although, compared with the incidence of cervical cancer, the incidence of anal cancer in women is rare, the results of the present study suggest that HPV infection of the anus is relatively common and that several risk behaviors might be used as a basis for clinical screening. The observation that HPV-16 and -18 infections in the anal canal are highly prevalent implies that prophylactic vaccines targeting these oncogenic types may reduce the incidence of anal cancer. 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