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 Table of Contents  
ORIGINAL ARTICLE
Year : 2021  |  Volume : 14  |  Issue : 3  |  Page : 322-327

Detection of human papillomavirus genotypes in human immunodeficiency virus-negative women in Belagavi, Karnataka


1 Department of Microbiology, KAHER, Belagavi, Karnataka, India
2 Department of Microbiology, USM-KLE International Medical Programme, KAHER, Belagavi, Karnataka, India
3 Department of Microbiology, JN Medical College, KAHER, Belagavi, Karnataka, India
4 Department of Obstetrics and Gynecology, JN Medical College, KAHER, Belagavi, Karnataka, India
5 Division of Microbiology, National AIDS Research Institute, Pune, Maharashtra, India

Date of Submission08-Feb-2021
Date of Acceptance08-Jul-2021
Date of Web Publication30-Sep-2021

Correspondence Address:
Dr. Vinay S Pala
Department of Microbiology, KAHER, Belagavi, Karnataka
India
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/kleuhsj.kleuhsj_21_21

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  Abstract 

BACKGROUND: Women are known to be at high risk of human papillomavirus (HPV) infection and its associated cervical pathology. However, limited data are available on circulating genotypes, especially among human immunodeficiency virus (HIV)-negative women. Therefore, the present study was designed to gauge the prevalence of HPV genotypes in HIV-negative women.
MATERIALS AND METHODS: This cross-sectional study was conducted on a total of 96 HIV-negative women in the age range of 18–45 years. Cervical samples were collected from the transformation zone of the cervix using the deoxyribonucleic acid collection device (cytobrush), which was rinsed in 20 ml of PreservCyt® vial (Hologic, Inc.). HPV genotyping was done with the linear array HPV genotyping test (Roche, Branchburg, NJ, USA) at National Aids Research Institute, Pune.
RESULTS: A significant linear increasing trend in proportion of carcinogenic and noncarcinogenic genotypes over grade was observed (P = 0.039 and P = 0.0024, respectively). HPV 59 was reported to be the most common genotype followed by 16, 53, 62, and 72 but without any statistical significance.
CONCLUSION: Screening strategies incorporating HPV genotyping and vaccination should be effective in preventing cervical cancer in HIV-negative women.

Keywords: Carcinogens, genotype, Papillomaviridae, prevalence, vaccination


How to cite this article:
Pala VS, Patil C, Nagmoti MB, Dalal AD, Mane A. Detection of human papillomavirus genotypes in human immunodeficiency virus-negative women in Belagavi, Karnataka. Indian J Health Sci Biomed Res 2021;14:322-7

How to cite this URL:
Pala VS, Patil C, Nagmoti MB, Dalal AD, Mane A. Detection of human papillomavirus genotypes in human immunodeficiency virus-negative women in Belagavi, Karnataka. Indian J Health Sci Biomed Res [serial online] 2021 [cited 2021 Nov 28];14:322-7. Available from: https://www.ijournalhs.org/text.asp?2021/14/3/322/327251


  Introduction Top


Human papillomavirus (HPV) is nonenveloped deoxyribonucleic acid (DNA) virus with 7900 bp that infects keratinocytes of the stratified cutaneous and mucosal epidermis. HPV is classified into more than 200 genotypes, out of which more than 50 genotypes are associated with anogenital infections, and these genotypes are further subdivided into two broad groups as high-risk (HR) and low-risk (LR) genotypes based on its oncogenic potential[1],[2]as well as cutaneous and mucocutaneous based on tissue tropism.[3]

HPV infection is identified as one of the most common sexually transmitted infections worldwide along with human immunodeficiency virus (HIV) and syphilis. Certain genotypes of HPV are also identified as the leading cause of cervical cancer, which is the second most common cancer in middle-aged women. HPV genotypes have been associated with the development of several types of squamous epithelial cell tumors such as cervical, vulvar, anal, penile, and oropharyngeal cancer representing 6% of cases worldwide.[4],[5] The most common HR-HPV is 16 and 18, and approximately 70% of cervical cancer cases are due to these genotypes. LR-HPV, especially 6 and 11, are predominantly involved in the development of genital warts.[6]

Cervical cancer prevention programs consist mainly of cervical cancer screening by Papanicolaou (Pap) smear for early detection of precancerous lesions.[7] Although screening programs are in place and effective, cervical cancer remains a burden for any nation due to a large proportion of women who remain unscreened or unaware, as well as false-negative Pap test results due to poor sensitivity of the test.[8],[9],[10]

With the recent advancement in HPV vaccine introduction, there is a need for knowing the genotypes prevalent in the community.[11] Since the sensitivity and specificity of  Pap smear More Details are poor, cervical cancer screening programs are switching from cytology-based screening to HR-HPV testing. Therefore, PCR testing is a sensitive and noninvasive method for determining the presence of a cervical infection since HPV genotyping is important to determine the presence of HR types. However, limited data are available on circulating genotypes, especially among HIV-negative women.[12]

In light of the above knowledge, the present study was designed to gauge the prevalence of HPV genotypes in HIV-negative women in Belagavi, Karnataka, and investigate the types of HPV most frequently found in these patients.


  Materials and Methods Top


Study design

This cross-sectional study was approved by the Human Research Ethics Committee of the KLE University, Belagavi, India, in June 2013. The study participants included all HIV-negative women aged from 18 to 45 years (reproductive age), attending department of obstetrics and gynecology. Women were excluded if they had never been pregnant, had a hysterectomy, were in active labor, had abnormal bleeding, had history of sexually transmitted disease, or were unwilling to participate in the study. The study was explained to patients in their local language or English. The informed consent was obtained from all the study participants after providing the study fact sheet. A total of 96 HIV-negative women of age ≥19 years were included in the study.


  Data and sample collection Top


Demographic data of all the participants were collected and recorded in a structured questionnaire, which was approved by the ethics committee through interview by a gynecologist. After collecting data and informed consent, participants were subjected to routine gynecological examination which included general, pelvic, and speculum examination and sample collection. Based on the data provided by the participants, they were counseled on risk of sexual behavior and provided with contraceptives if needed. Return appointments and referrals were made as required by the gynecologist.

Samples for Papanicolaou test

The smears were collected with the help of Ayer's spatula and cytobrush from the squamocolumnar junction by following the American Society of Cytopathology guidelines. The cellular material obtained was quickly smeared on a clean grease-free glass slide. Two smears were prepared for each case. The glass slides were then fixed by using 95% ethyl alcohol. The smears were stained with Pap stain. Slides were examined under light microscope and were reported by a pathologist according to the 2001 Bethesda System.[13],[14],[15]

Samples for human papillomavirus genotyping

Cervical samples were collected from the transformation zone of the cervix using the DNA collection device (cytobrush) which is rinsed in 20 ml of PreservCyt® vial (Hologic, Inc.). Samples were immediately stored at 4°C and transported to the laboratory at the department of microbiology where they were further stored at −20°C until DNA extraction.

Human papillomavirus genotyping

HPV genotyping was done with the linear array HPV genotyping test (Roche, Branchburg, NJ, USA) at National Aids Research Institute (NARI), Pune.

Total DNA was extracted from cervical samples collected in PreservCyt medium solution and purified using the AmpliLute Liquid Media Extraction Amplicor kit, following the manufacturer instructions (Roche Molecular Diagnostics, Branchburg, NJ, USA). All samples were genotyped by Linear Array HPV Genotyping Test as previously described.[15],[16]

Statistical analysis

Data analysis was done using R i386.3.5.1. R Foundation for Statistical Computing Vienna, Austria statistical software z. Continuous data were represented in the form of mean ± standard deviation, and the categorical variables were represented by the frequency table. Trend analysis was done using Cochran Armitage trend test. Crude odds ratio was used to study the factors influencing the HPV. A P ≤ 0.05 was considered statistically significant.

Ethical clearance

Institutional Ethical Committee with Ref no KLEU/Ethic/2012-13/D-4573 dated 18.03.2013.


  Results Top


A total of 96 participants were enrolled in the study of mean age 32.07 ± 6.79 years, consisting of 13 HPV-positive and 83 HPV-negative cases.

Almost 84.38% of women were married of which 14.8% had HPV. Among the participants 74.47% used the contraceptives. The current study reported that age was significantly associated with non HPV 16 genotypes which is contradictory to the study by Teixeira et al. that stated higher HR HPV prevalence in older age groups than in younger age groups (<26 years: 7.3%; 26–30 years: 13.2%; 31–35 years: 11.9%). Condom followed by Copper-T was reported as the common type of contraceptive used [Table 1].
Table 1: Baseline characteristics

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It has been observed that out of 13 HPV-positive cases, 7 had Grade 1+, 5 had Grade 2+, 9 had Grade 3+, and 2 had Grade 4+. A significant linear increasing trend in proportion of carcinogenic and noncarcinogenic genotypes over grade was observed (P = 0.039; P = 0.0024, respectively). HPV 59 was reported to be the most common genotype followed by 16, 53, 62, and 72 but without any statistical significance [Table 2].
Table 2: Prevalence of human papillomavirus genotypes, overall and stratified by grading among human papillomavirus

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Using univariate logistic regression, it was concluded that age was significantly associated with non-HPV 16 genotypes. Using odds ratio, it has been concluded that, for 1-year increase in age, odds of absence of non-HPV 16 genotype increased by a factor 1.14 [Table 3].
Table 3: Logistic regression analysis

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  Discussion Top


During the last few years, Pap smear is being replaced with HPV genotyping due to high sensitivity and specificity. Linear array genotyping and PCR testing with several primers were commonly used in many laboratories for HPV detection.[17],[18] Linear array test, launched in 2006, is a PCR-based strategy with pooled primer sets, coupled to hybridization to specific probes for 37 anogenital HPV genotypes immobilized on a nylon strip.[19],[20],[21],[22],[23],[24]

The prevalence of HPV DNA in the present study was observed to be 13/96, which was in slight contrast with the study conducted by Sun et al.[25] with HPV DNA being detected in 36% of negative women, and Minkoff et al. found a prevalence of 43%.[26] A study by Veldhuijzen et al. showed the prevalence of HR-HPV as 31.8% and LR-HPV as 32.4% in HIV-negative women.[27] A meta-analysis conducted by Women's Interagency HIV Study, 1999 showed prevalence of 28% in HIV-negative women, respectively. Brazilian studies conducted by Campos et al.[28] found HPV DNA prevalence in 23.7% of cases. Nevertheless, Levi et al.[29] showed 100% of HIV-negative women positive to HPV DNA. In this study, control group women were selected in a cervical pathology clinic, and a high positivity was expected to HPV DNA.

Among the patients in our study who were positive, HPV DNA type 59 and 16 were most common in 10 cases, whereas in a study conducted by Queiroz et al., the most common genotypes found were HPV 16 and HPV 52.[22] In our study, most of the women had infection with single genotype, and this was in concordance with several studies carried out among HIV-negative women. Findings of several studies resonated with the findings of the present study, thus concluding that HIV-negative women were less prone to getting HPV infection.[22],[23],[24]

A study by Clifford et al. and Sahasrabuddhe et al. found that the prevalence of HPV 16 was lower in the general population and that other high-risk types (e.g., HPV 18, 31, 33, 51, 52, and 58) were prevalent.[30],[31] Many others have also demonstrated that other HPV subtypes, except 16 and 18, are clearly playing a role in HPV prevalence.[32],[33],[34] In a study conducted by Rodolfo et al., the most common HPV genotype in HIV-negative women was identified as HPV 16.[35]

The current study reported that age was significantly associated with non-HPV 16 genotypes which was not in harmony with the findings of the study by Teixeira et al. that stated higher HR-HPV prevalence in older age groups than in younger age groups (<26 years: 7.3%; 26–30 years: 13.2%; 31–35 years: 11.9%) but no trend with age was found (P = 0.10). These contrasting results could be due to variation in the study design.[36]

To our knowledge, our study is the first of its kind in our geographical location to know the genotypes circulating in HIV-negative women as very few prospective studies on HPV epidemiology in HIV-negative women have been conducted.

The study, however, had few potential limitations as well. First, the FDA has approved vaccines, which were not effective against the most common genotypes found in our study. Second, further studies with larger population are required to identify the most common genotypes prevalent in our geographic location so as to use vaccine in prevention of HPV diseases.


  Conclusion Top


Newer vaccines such as the nonavalent HPV vaccine present the possibility of better coverage for women and will need to be evaluated. Strengthening preventive efforts is necessary to improve early detection through increasing accessibility to screening programs, adherence to follow-up among those with lesions, and intensifying health education for women living with HIV.

Acknowledgments

We would like to the author Mrs. Linata Patil, Technician, of the NARI, Pune, Maharashtra, for carrying out the HPV genotyping studies.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.

 
  References Top

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