Journal of Womens Health, Issues and Care ISSN: 2325-9795

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Research Article, J Womens Health Issues Care Vol: 5 Issue: 5

Acetowhite Test for Detection of Papilloma Virus Infection in Men Sexual Partner of Women Diagnosed of High Grade Cervical Lesions

Elena López-Diez1*, Sonia Pérez2, Amparo Iñarrea3, Angel dela Orden3, Máximo Castro1, J Carlos Diz4, Sheila Almuster1, Moises Rodríguez1, Ruben Montero1, Miguel Perez Schoch1 and Antonio Ojea1
1Department of Urology, University Hospital of Vigo, Spain
2Department of Microbiology, University Hospital of Vigo, Spain
3Department of Obstetrics and Gynecology, University Hospital of Vigo, Spain
4Department of Anesthesia, University Hospital of Vigo, Spain
Corresponding author : Elena López-Diez
Department of Urology, University Hospital of Vigo, Alvaro Cunqueiro s/n 36200 EOXI de Vigo Apartado oficial, Spain
Tel: 0034986811111 Ext 514639
E-mail: [email protected]
Received: April 25, 2016 Accepted: June 15, 2016 Published: June 20, 2016
Citation: López-Diez E, Pérez S, Iñarrea A, Orden A, Castro M, et al. (2016) Acetowhite Test for Detection of Papilloma Virus Infection in Men Sexual Partner of Women Diagnosed of High Grade Cervical Lesions. J Womens Health, Issues Care 5:5. doi:10.4172/2325-9795.1000244

Abstract

Background: Traditionally, men asked urologist for treatment of genital warts caused by low risk Papilloma virus (LR-HPV) but, recently, asymptomatic men also want to know if they are infected by high risk HPV (HR-HPV) and the benefits of HPV vaccine. The aims of this cross-sectional study were: (1) to evaluate acetowhite test usefulness for detection of asymptomatic and subclinical HRHPV infection in a high risk population of men and (2) to assess risk factors of HR-HPV infection in this population.
Methods: Healthy men (n=137) sexual partners of women with preneoplastic cervical lesions were recruited. Acetowhite test was performed (5% acetic acid solution, colposcopic examination under 4-fold and 7-fold magnification). Genital samples were obtained for HR-HPV DNA detection from different penile areas. (Linear Array HPV Genotyping Test, Roche Diagnostics, Mannheim, Germany). SPSS 19.0 (IBM, Chicago, USA) was used for statistical analysis.
Results: HR-HPV prevalence was 68/137 (49.6%). AW procedure was positive in 36/137 (26.3%) patients. Genital warts were present in 18/137 (13.1%) patients. Acetowhite test showed 25% sensitivity (95% CI: 13.9-36), 72.4% specificity (95% CI: 61.2-83.7), 47.2% predictive positive value (95% CI: 29.5-64.9) and 49.5% negative predictive value (95% CI: 39.2- 59.7) for the identification of HR-HPV infection. Subclinical lesions were detected in 16/68 (23.5%) infected males. Genital warts (p=0.010) were associated with an increased risk for HRHPV infection in men.
Conclusion: In our experience, acetowhite procedure sensitivity was low for HR-HPV detection in asymptomatic/subclinical genital infection and it was not specific indicator of HR-HPV infection. Genital warts detection was associated with HR-HPV infection in men.

Keywords: Human papilloma virus; DNA test; Male; Prevention; Cervical intraepithelial neoplasia; Mass Screening; Genital warts; Diagnosis

Keywords

Human papilloma virus; DNA test; Male; Prevention; Cervical intraepithelial neoplasia; Mass Screening; Genital warts; Diagnosis

Background

Human papillomavirus (HPV) infection is the most common sexually transmitted infection in men and women worldwide [1]. HPVs are a very large family of double stranded DNA viruses (DsDNA), very resistant that can survive in the environment without a host and is able to infect humans. These viruses are not classified as serotypes, but as genotypes on the basis of DNA sequence. Currently, over 200 genotypes have been identified and about 40 genotypes (the alpha genus) can be transmitted through sexual contact and infect the anogenital region [2]. HPV genotypes have been classified into low-risk genotypes, associated with anogenital warts, low-grade cervical lesions and recurrent respiratory papillomatosis, and highrisk genotypes (HR-HPV) [3] (Table 1) which eventually can lead to malignant transformation. HR-HPV are strongly associated with cancer and high-grade neoplasia of the anogenital tract, including the anus (AIN), penis (PeIN), uterine cervix (CIN) and vulva (VIN) and also a proportion of oropharyngeal cancer [4]. In total 20 HPV genotypes are described causing cervical cancer [5]. It has been learned a lot about the epidemiology of HR-HPV infection in women, found in 99.7% of cervical cancer worldwide and in 96.8% of cervical preneoplastic and neoplastic lesions in our community [6]. Transmission of HR-HPV between sexual partners might maintain viral infection and consequently may influence the clinical course of CIN. HPV infection in sexual partners of women infected by HPV may be responsible for the woman reinfection [7] and plays an important role in anogenital cancers in men and women [8].
Table 1: Oncogenic HPV genotypes.
Relatively little is known about the natural history of HPV related anogenital infection in men. Anogenital HPV male infections are categorized as asymptomatic (latent), subclinical or clinical. Most infections are asymptomatic, detectable only with test for viral DNA and become undetectable over time. Although the prevalence of HPV DNA in the male genital tract appears to be similar to that observed in the cervix uterus of the same population and age group [9], the presence of HPV in the male genital tract produces generally latent or subclinical infections, more than 10 times common than clinical (apparent) infection [10]. Latent and subclinical infections can be detected only after the application of acetic acid solution, a procedure known as Acetowhite test (AW, peniscopy) [11]. Due to these diagnostic difficulties, male genital infections usually escape clinical observation and have limited pathological consequences for healthy men.
Genital warts represent a significant public health problem associated with clinical symptoms (burning, bleeding and pain) and psychosocial problems (embarrassment, anxiety and decreased selfesteem). Several studies have suggested that the occurrence of genital warts has been increasing over time [12], except in countries with a high HPV vaccination coverage [13-15]. The prevalence of genital warts in sexual partners of women having cervical intraepithelial neoplasia is 17% [12]. Among 1788 HPV-positive men in the HIM Study, 86 developed genital warts during follow-up. During the first 12 months of follow-up, 16% of men with a genital HPV 6 infection developed an HPV 6-positive condyloma, and 22% of genital HPV 11 infections progressed to an HPV 11-positive condiloma [16].The management of male sexual partners of women with preneoplastic lesions without the visible presence of penile, scrotal or endourethral lesions is still unclear [17]. In this context, focusing in asymptomatic men, studies are necessary to improve the understanding of viral transmission and prevention of HPV diseases in both men and women.
Since the American Society for Colposcopy and Cervical Pathology recommended cotesting (cytology+HPV DNA testing) in women aged 30-65 years, and several countries have started HPV DNA testing in primary screening, a considerable number of couples want to know the presence of HPV. A screening test for detection of asymptomatic and subclinical genital HPV infection in men at a reasonable price and causing minimal discomfort to the patient would be very valuable.
The aims of this cross-sectional study were: 1) to evaluate Acetowhite test usefulness in the detection of asymptomatic and subclinical HR-HPV infection in a high risk population of men, sexual partners of women infected by HPV and 2) to assess risk factors of HR-HPV infection in this population.

Materials and Methods

Population
A cross-sectional study was conducted by the Urology Department of the University Hospital of Vigo. We recruited 137 asymptomatic men more than 18 years old selected because their current couple had presented high grade squamous cervical lesions (CIN II, CIN III-Carcinoma in situ) in the previous six months. The institutional review board approved the study (cod 2013/470 from the ethics committee of clinical investigation of Galicia Santiago de Compostela, Spain). Information concerning the research project was provided to all participants. Written informed consent was obtained from all patients. Men were invited to fill in a questionnaire on life-style habits, including sexual behavior (SB). Because the significance of a positive HPV test in men is unknown, study personnel spent a considerable amount of time educating men about HPV. We explained that a positive test for the virus do not necessarily put them at risk for disease. Participants did not receive incentive for study involvement.
All patients were examined and after applying 5% acetic acid solution we enhanced visualization of the skin by a colposcope under 4-fold and 7-fold magnification. AW lesions were classified as typical (presence of well demarcated lesions with a slightly elevated border and the occurrence centrally of punctuated capillaries with or without an associated epithelial depression) and non-typical (presence of lesions exhibiting a ragged border and lacking punctuated capillaries).
Specimen collection for HPV DNA detection
Specimen for HPV DNA detection were taken with three cytobrushes from the preputial cavity, the glans and the sulcus coronarius, scrotum and urethral meatus and suspended together into one single vial containing TE buffer pH 8.0 Molecular Biology grade (AppliChem GmbH, Darmstadt, Germany). DNA was isolated using QIA amp MinElute Media Kit (Qiagen, Hilden, Germany). Amplification and detection were carried out using the Linear Array HPV Genotyping Test (Linear Array Roche Diagnostics, Mannheim, Germany) according to the manufacturer’s instructions. We described the distribution of 21 HR-HPV genotypes classified as HR (HR-HPV, IARC Group 1 carcinogens) or probable/possible HR (pHR-HPV, IARC Group 2A/B carcinogens) by the International Agency for Research on Cancer Monograph Working Group [3]. This test also detects human beta-globin in order to control the sample adequacy and quality as well as success in DNA extraction and PCR (internal control).
Statistical analysis
Sensitivity, specificity, positive and negative predictive values and their 95% confidence interval (CI) were calculated. Qualitative variables were compared using the chi-square test. Odds ratio (OR) and corresponding 95% CI were calculated. SPSS version 19.0 for Windows IBM, Chicago, USA was used for statistical analysis. A p value <0.05 were considered statistically significant. Multivariate logistic regression analysis was used to evaluate the factors independently associated with the detection of HR-HPV infection in male by use Step-Wise Logistic regression models. Only variables with p ≤ 0.1 in the bivariate analysis were included in the multivariate analysis.

Results

Average age of men was 38.0 ± 8.7 years. Human beta-globin was detected in all cases. Prevalence of HR-HPV infection in men was 49.6% (68/137). HR-HPV single infection was detected in 38.2% (26/68) and HR-Multiplex infection in 61.8% (42/68). HPV16 was the most frequent type, detected in 45.6% (31/68) of infected men.
AW procedure was positive in 36/137 (26.3%) patients. AW procedure showed 25% sensitivity (95% CI: 13.9-36), 72.4% specificity (95% CI: 61.2-83.7), 47.2% predictive positive value (95% CI: 29.5- 64.9) and 49.5% negative predictive value (95% CI: 39.2-59.7) for the identification of HR-HPV infection. Subclinical lesions were detected in 16/68 (23.5%) infected males (Table 2).
Table 2: Genital lesions viewed by peniscopy (AW) in asymptomatic sexual partners of women with high grade cervical lesions, according to the presence of HR-HPV.
Genital warts were present in 18/137 (13.1%) patients. Prevalence of HR-HPV infection was 14/18 (77.8%) in patients with genital warts vs. 54/119 (45.4%) in patients without them (OR 4.2, 95% CI 1.3-13.5, p=0.010) (Table 3). AW procedure showed 83.3% sensitivity (95% CI: 63.3-100), 82.3% specificity (95% CI: 75.7- 89.6), 41.6% predictive positive value (95% CI: 24.1-59.1) and 97% negative predictive value (95% CI: 93.2-100) for genital warts detection.
Table 3: Acetowhite lesions according to genital warts detection.

Discussion

In the present study sexually active healthy males, partners of women diagnosed of high grade cervical lesions were tested for HRHPV infection with HPV DNA and AW test. HR-HPV infection was diagnosed in half of this male subpopulation. True prevalence of HPV infection in asymptomatic men is difficult to estimate, mainly due to the silent behavior of this virus and to the fact that it depends on the nature of their sexual activities [18]. Our data falls within the range reported in other studies for sexual partners of women with CIN (30%-68%) [11,19-22].
The currently available methods for evaluating HPV infection in male are HPV DNA test and AW test [23]. This study found that 52% of positive AW results were not related with HR-HPV infection. Some authors have published similar results [24,25]. False positive results may be due to low risk HPV infection or inflammatory conditions such as folliculitis, lichen, psoriasis, contact dermatitis, candidiasis, genital herpes and microtraumatic lesions from recent intercourse [17,26]. Nevertheless the need for detecting subclinical genital HPV infection, associated with detectable AW lesions [27] have been emphasized. Our results, almost twenty four percent of the infected population by HR-HPV presented AW lesions at the same time, are consistent with previous studies [27]. Penile lesions, the majority of which are subclinical [19] may be present in 3.6% to 27.3% of the sexual partners of HPV positive women had presented high grade squamous cervical lesions [17,20]. HPV infection affects many sites in both males and females and shows a significant prevalence of oropharyngeal infection in couples performing oral sex. In coinfected couples, avoiding oral sex reduces the number of infected sites diminished faster during the follow-up and a higher number of couples had clearance [28]. In a Norwegian study consistent condom use increases the regression rate of cervical intraepithelial neoplasia 2-3 [29]. These acetowhite lesions on male genitalia are in fact squamous intraepithelial lesions and should not be ignored. There is a risk of further development of high-grade neoplasia and penis carcinoma [16,20]. We encourage the importance of the clinical follow-up of this men and if indicated, taking biopsy.
In the present study, the prevalence of genital warts (GW) was 13.1%. In a systematic review of the litterature, the overall prevalence of GWs based on retrospective administrative databases or medical chart reviews or prospectively collected physician reports ranged from 0.13% to 0.56%, whereas it ranged from 0.2% to 5.1% based on genital examinations [30]. Nevertheless, patients with GW had 4 folds more risk of HR-HPV infection. This is reasonable because low risk HPV genotypes causing GW share the transmission path with high risk HPV genotypes [31]. They are generally regarded as a benign condition not associated with mortality, but they can be difficult to treat and recurrence of these lesions is often seen. Although test for the detection of HPV are not recommended for diagnosis of GW [32], a Danish study of 50.000 people with GW found an elevated risk of HPV-associated cancers in people with GW compared with the general population [12]. The AW test was specific for genital warts detection, showing a high negative predictive value. This procedure could avoid missing small clinical lesions AW test can be helpful in the diagnosis of GW, in particular in suspicious lesions without classic features. Taking a biopsy might also be indicated if diagnosis is uncertain, the lesions do not respond to standard therapy or the disease worsens during therapy [32]. Visible warts represent only the tip of the iceberg and low and high-risk HPV infection contribute to the genital lesion burden in men [33].
A limitation of our study is the small sample size of studied men as well and the lack of data of the duration and clearance of the HPV infection in men. Because we studied a population at high risk of HPV infection our results might be not extrapolable to low risk men subpopulations. Moreover, penile lesions diagnosed by colposcopy could have been misclassified due to observational bias. Differences in collection, DNA extraction and HPV genotyping methods may also varies between countries influencing the results. In spite of there are no licensed test for HPV detection in men, polymerase chain reaction (PCR) has emerged as the most sensitive available method for the detection of latent HPV infection. The infectious diseases literature supports the lack of the US Food and Drug Administration (FDA) approval of HPV tests for HPV detection in men and the absence of adequate therapy for established HPV infection in this population.
Until recently, no highly effective primary prevention strategy to reduce the risk of HPV acquisition existed. However, research has demonstrated that nonavalent, quadrivalent and bivalent HPV vaccines are effective at preventing HPV-infections and precancer reducing the risk of HPV-related cancer in both males and females [29]. On October 16, 2009, the US Food and Drug Administration (FDA) approved the use of quadrivalent vaccine in males 9-26 years old for the prevention of genital warts. Most European countries offer HPV vaccination for girls. The HPV-vaccine is also recommended for boys, but only boys in Australia, Austria, parts of Canada and Italy are offered the vaccine in the vaccination program. Potential interventions such as a therapeutic HPV vaccine may avert new HPV infections. Moreover, vaccinating boys would reduce HPV related diseases in both sexes to a greater extent than herd immunity. Future trials of HPV vaccines in men should take into account not only the presence of penile HPV infection but also the presence of penile subclinical lesions as an outcome measure for the efficacy of a vaccine [34]. The HPV that we detected within the lesions could be prevented by a vaccine.
Although there are no recommended or approved screening programs for asymptomatic infections in men our finding suggests the need for greater attention to sexual partners of HPV infected individuals. Further prospective and controlled studies in different populations are needed to provide adequate counseling to men that demand to know if they are infected by HR-HPV. Although similar studies have been conducted in different countries, it is important to investigate the different aspects regarding HPV infection in men, attempting to reproduce and confirm results.
Follow-up for men with DNA HPV test and positive acetowhite represents the best route to the diagnosis of preneoplastic lesions in males. HPV vaccination may reduce the risk of recurrence of CIN2-3 after treament [35].

Conclusion

In our experience, acetowhite test sensitivity was low for HRHPV detection in asymptomatic infection of genital areas and it was not a specific indicator of HR-HPV infection. Nevertheless, acetowhite test allowed the diagnosis of subclinical lesions in quart of HPV infected males. Increased risk of HR-HPV infection was found in men presenting genital warts.

Authors`Contribution

ELD carried out the collection in male, design of study data and drafted the manuscript. SPC performed HPV detection, participated in the study design and drafted the manuscript. All authors read and approved the final manuscript.

Acknowledgment

We thank Vigo, Spain Research Unit for the statistical analysis and review of the manuscript. We also thank nursery team, especially Carmen Lago from the Urology Department of the University Hospital of Vigo, for their excellent help in this research, collecting patients. We thank M Consuelo Reboredo from the Gynecology Department and the laboratory technicians of the Microbiology Department of the University Hospital of Vigo, for their support of the study.

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