Assessment of Human Papillomavirus (HPV) type 16 and 18 status by nested multiplex PCR in cervical cancer patients and in healthy women from a tertiary care hospital

Abstract

Background: Cervical cancer is the most common malignancy in Indian women and approximately 130,000 women fall prey to cervical cancer every year. HPV DNA testing can be used as an adjunct to cytological screening of cervical samples of above 30 year old women and can be used as an optional test for follow-up in women who have slightly abnormal Pap test results. The aim of the study was to assess the HPV 16 and 18 status in cervical cancer patients and in healthy women visiting JIPMER, Pondicherry.Methods & Materials: This study was approved by JIPMER Institute Ethics Committee. 171 biopsies from cervical cancer patients and 152 cytobrush samples from healthy women were collected. DNA was extracted using QIAamp DNA mini kit. The quality and quantity of the extracted DNA was determined by a spectrophotometer. The integrity of genomic DNA was checked by amplifying a 248-bp product of the human beta-globin housekeeping gene. A nested multiplex PCR was standardized and carried out to detect 2 high risk types, HPV 16 and HPV 18, further representative positive PCR amplicons were sequenced for confirmation.Results: Out of 171 cervical cancer cases, 149 (87%) patient samples tested positive. Overall HPV-16 and18 occurrence was found to be 87% in which HPV-16 was 64%, HPV 18 was 3% and mixed infection was 23%. Out of 152 healthy women cases, 78 samples were positive. Overall HPV-16 and18 occurrence was found to be 51% in which HPV-16 was 22%, HPV 18 was 5% and mixed infection was 25%. Sequencing results were analyzed in NCBI BLAST to reconfirm the HPV genotype. Conclusion:In our study, HPV -16 found to be the most common genotype in our region. Our study suggests that the HPV 16 and 18 mixed infection could be due to the consequence of poor genital hygiene and unsafe sexual behavior. In healthy women cases, it may be too early to come to a definitive conclusion about the HPV positivity because women under 30 who are sexually active are likely to have an HPV infection that will disappear on its own in future. But those women may need regular follow up.