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Sexually Transmitted Diseases:
doi: 10.1097/OLQ.0b013e318158af65
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Detection of Human Papillomavirus From Self-Collected Vaginal Samples of Women in Chiang Mai, Thailand

Wongworapat, Kanlaya MSc*; Keawvichit, Rassamee BSc, MPA*; Sirirojn, Bangorn MSc*; Dokuta, Sirikwan BSc*; Ruangyuttikarn, Cholticha MSc*; Sriplienchan, Somchai MD†; Sontirat, Auchara MSc*; Kla, Kanitta Thai MSc*; Gravitt, Patti E. PhD†; Celentano, David D. ScD, MHS†

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From the *Research Institute for Health Sciences, Chiang Mai University, Chiang Mai, Thailand; and †Department of Epidemiology, Johns Hopkins Bloomberg School of Public Health, Baltimore, Maryland

The authors gratefully acknowledge the Roche Molecular System, USA, for the donation of reagents and supplies for HPV detection and HPV typing.

This work was supported, in part, by a fellowship/grant from the Fogarty International Center/USNIH, Johns Hopkins University, Grant 2 D 43 TW000010-18-AITRP.

Correspondence: Kanlaya Wongworapat, MSc, 110 Intavaroros Road, Tambol Sriphum, Chiang Mai 50202 Thailand. E-mail: kanlaya@chiangmai.ac.th.

Received for publication November 10, 2006, and accepted August 16, 2007.

IT HAS BEEN SHOWN THAT some types of human papillomavirus (HPV) are the major risk factor for high-grade squamous intraepithelial lesion and invasive cervical carcinoma.1,2 The detection of HPV has been used as an adjunct to cervical cytology to identify women who are at risk of developing cervical cancer.2 Currently, the most widely used method to detect HPV DNA is by obtaining a specimen from cervix. However, some studies have shown that HPV detection by self-collected vaginal swab is comparable to virus detection in swabs obtained directly from the cervix.3,4 The self-collected vaginal samples also offer advantages over endocervical swabs in term of acceptability to clients and in reducing the need for trained personnel and clinical materials required for a cervical sample collection.4,5 The self-collected sample for HPV detection seems to offer a potential opportunity for more widely accessible cancer prevention and for population-based molecular epidemiologic research.

We conducted the study of HPV infection among women from communities in Chiang Mai, Thailand, using self-collected vaginal samples. Five hundred thirty-one sexually active women aged 20 to 65 years participated in the study. They were selected by nonprobability sampling method from 8 communities located within 50 km of Chiang Mai city. Written informed consent was obtained from all women; the protocol was reviewed and approved by the relevant Institutional Review Boards. The women were asked to provide their vaginal sample for HPV testing. The women were given a sample collection kit (swab specimen collection kit, Digene Corp, Beltsville, MD) that was composed of a sterile Dacron swab in a wrapper and a plastic vial containing specimen transport media. Verbal and written instructions for cell collection were provided. In brief, women were asked to insert a collection swab gently into the vagina about halfway in or until meeting with resistance; then they were to rotate the swab 3 turns in the same direction, take the swab out, and put it in the transport vial. The samples were transported to the laboratory and tested for HPV DNA and HPV typing.

The DNA of vaginal samples was extracted and amplified using PGMY09-PGMY11 biotinylated L1 consensus primers, generating a 450-base pair amplicon.6 To determine specimen adequacy, the biotinylated β-GH20/β-PC04 human β-globin target was coamplified, generating a 250-base pair amplicon. The results of amplification were confirmed by agarose gel electrophoresis. The genotypes of HPV were determined by the reverse line blot hybridization method.7 The HPV genotyping strip detects 2 β-globin concentrations (high and low) and 38 HPV types: 22 putative high-risk types (16, 18, 26, 31, 33, 35, 39, 45, 51, 52, 53, 56,58, 59, 66, 67, 68, 69, 70, 73, 82, IS39) and 16 low-risk types (6, 11, 40, 42, 54, 55, 57, 61, 62, 64, 71, 72, 81, 83, 84, CP6108). The classification of high- and low-risk types was primarily based on International Agency for Research on Cancer (IARC)8 and also referred to the HPV reference guide provided by Roche.

All of the women we approached were interested in collecting their vaginal samples for HPV testing. About 9.4% (50 of 531) of the samples lacked β-globin control amplification. We therefore diluted the DNA template of these 50 samples with TE buffer (1:10) (TE = 20 mM Tris-HC1, 1 mM EDTA [pH 8.0]) and repeated the polymerase chain reaction (PCR), and subsequently every sample could be amplified and produced a β-globin PCR product. Of 531 women tested, 61 were positive for HPV DNA, and the overall HPV prevalence was 11.5% (Table 1). The prevalence of high- and low-risk types was 5.1% (27 of 531) and 7.5% (40 of 531), respectively. Twenty-two HPV types were found. Types 52, 16, and 68 were common among the group of high-risk types, and types 72, 71, and 62 were common among the group of low-risk types (Table 2). Multiple infections were found in 2.6% (14 of 531). The age-specific prevalence of HPV infection in participants was presented (Table 3). The prevalence of HPV infection was similar across all age groups. The women of age 40 to 49 had the highest prevalence of 13.8% (32 of 231), whereas the women of the age 30 to 39 had the highest prevalence of high-risk type HPV at 6.7% (8 of 119).

Table 1
Table 1
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Table 2
Table 2
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Table 3
Table 3
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To the best of our knowledge, this is the first study to investigate the prevalence of HPV using self-collected vaginal samples from the asymptomatic population of reproductive-age women in the northern part of Thailand. The prevalence of HPV infection in this study is approximately the same as that detected in exfoliated cervical cells collected from women in Lampang (a neighboring province), which was 9.1%.9 The most common types were 72, 52, 71, and 16, which were also similar to the types found in Lampang. It is notable that only 1 (HPV 16) of the 4 most commonly detected high-risk HPV types are currently considered for inclusion in the newly licensed HPV preventive vaccine cocktails (typically including HPV 6, 11, 16, and 18). It will be important to determine whether the other commonly detected types (HPV 52, 68, and 58) are contributing to the development of a significant number of cervical cancers in this population. The prevalence of HPV is shown to be higher in younger woman in other studies.10 However, we cannot clearly see the difference of the age-specific prevalence of HPV in our study; this may be due to the small number of participants enrolled in some age groups. Self-collection of vaginal samples is highly accepted by the women in the communities, with 100% of eligible women were willing to participate in our study. The self-collected sampling combined with PCR testing seems to be a rapid and simple method for HPV detection in the community-based epidemiologic studies.

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References

1. Chichareon S, Herrero R, Munoz N, et al. Risk factors for cervical cancer in thailand: A case-control study. J Natl Cancer Inst 1998; 90:50–57.

2. Nobbenhuis MA, Walboomers JM, Helmerhorst TJ, et al. Relation of human papillomavirus status to cervical lesions and consequences for cervical-cancer screening: a prospective study. Lancet 1999; 354:20–25.

3. Gravitt PE, Lacey JV, Brinton LA, et al. Evaluation of Self-collected cervicovaginal cell samples for human papillomavirus testing by polymerase chain reaction. Cancer Epidemiol Biomarkers Prev 2001; 10:95–100.

4. Sellors JW, Lorincz AT, Mahony JB, et al. Comparison of self-collected vaginal, vulvar and urine samples with physician-collected cervical samples for human papillomavirus testing to detect high-grade squamous intraepithelial lesions. CMAJ 2000; 163:513–518.

5. Dzuba IG, Díaz EY, Allen B, et al. The acceptability of self-collected samples for HPV testing vs. the pap test as alternatives in cervical cancer screening. J Womens Health Gend Based Med 2002; 11:265–275.

6. Gravitt PE, Peyton CL, Alessi TQ, et al. Improved amplification of genital human papillomaviruses. J Clin Microbiol 2000; 38:357–361.

7. Gravitt PE, Peyton CL, Apple RJ, et al. Genotyping of 27 humanpapillomavirus types by using L1 consensus PCR products by a single-hybridization, reverse line blot detection method. J Clin Microbiol 1998; 36:3020–3027.

8. Munoz N, Bosch FX, de Sanjose S, et al; International Agency for Research on Cancer Multicenter Cervical Cancer Study Group. Epidemiologic classification of human papillomavirus types associated with cervical cancer. N Engl J Med 2003; 348:518–527.

9. Sukvirach S, Smith JS, Tunsakul S, et al. Population-based human papillomavirus prevalence in Lampang and Songkla, Thailand. J Infect Dis 2003; 187:1246–1256.

10. Peyton CL, Gravitt PE, Hunt WC, et al. Determinants of genital human papillomavirus detection in a US population. J Infect Dis 2001; 183:1554–1564.

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