An epidemiological study on gynaecological morbidities among married women in rural Nepal with special emphasis on cervical cancer and related risk factors.

The study is a collaboration project between NTNU and Dhulikhel Hospital (DH) / Kathmandu University Hospital (KUH) in kavre region of Nepal. The current study aims primarily to investigate the prevalence of gynaecological morbidities among married women in rural areas in the Kavre district of Nepal, with special emphasis on cervical cancer and related risk factors, and secondarily to plan strategies for expanding awareness and screening programmes for women in other rural areas of Nepal.

Materials and Methods The study is a collaboration project between the Norwegian University of Science and Technology (NTNU) and Dhulikhel Hospital (DH) / Kathmandu University Hospital (KUH) in Kavre region of Nepal. Dhulikhel Hospital Kathmandu University Hospital (DH) is a non-governmental, not-for-profit, independent, tertiary level health facility with a catchment population of almost 1.9 million people in Nepal. The institution has been working in about twenty places throughout the country by developing novel approaches to a community based health and development programmes. The current study will be conducted in Bolde VDC (village development committee) where one of the outreach centres of Dhulikhel Hospital is located, and in neighbouring VDS’s. All married women between 15 years and above are eligible for inclusion in the study. Unmarried and pregnant women will be excluded. We aim to include about 1000 women. The information regarding the population in the area will be provided from District Health Office. An experienced and trained assistant nurse and a gynaecologist (and/or a medical officer who is trained in gynaecology and obstetrics department at least for six months after the completion of his/her medical studies), in coordination with the staff (who are also local inhabitants of Bolde) at Bolde Health Centre will identify the households. The women of the identified households will be approached to discuss the research project in detail. The women will also be informed about the benefits of the study (free clinical as well as diagnostic services in relation to the gynaecological problems as mentioned in the appendix), as well as the possible risks/difficulties. After thorough information of the women, an informed consent will be sought. No undue pressure will be exerted to include women in the study. After getting the informed consent, a date will be fixed to administer a questionnaire (ref. appendix, translated in Nepali) to gather demographic information, past and present medical history concerning gynaecological morbidities and risk factors related to cervical cancer. Thereafter the women will undergo a comprehensive general as well as gynaecological examination. The prevalence of precancerous lesions will be determined by cytological examination of cervical smear (Pap smear). The gynaecological examination and collection of cervical smear will be conducted by a gynaecologist and/or an experienced and trained nurse, using a spatula on ectocervix and a brush in endocervix in order to remove cells from the mucosa. In cases of clinically suspicious cervical cancer, biopsy will be taken for further histological examination. The prevalence of HPV infection will be determined by examination of cervical/vaginal smear. Vaginal smear will be collected for culture to investigate chlamydia and gonorrhoea infection. Blood samples will be collected in order to examine for HIV and hepatitis B infection. Data regarding other risk factors (age of marriage, age at first child birth, parity, number of sexual partners (of both wife and husband), occupation, symptoms of sexually transmitted infections, use of contraception, smoking habits) will be registered on a case-report form. All the data will be confidential and will not be disclosed to anybody without first consulting with the patient. All demographic date, survey answers and findings will be systematically registered and analysed in a database (SPSS).

Cytology: Cervical smears will be taken using cytobrush and fixed in alcohol immediately after sampling at the health centre. The smears will be stained with Papanicolaou’s staining method, examined and diagnosed and classified according to internationally accepted systems by pathologists at the Department of Pathology, Dhulikhel Hospital. An algorithm for follow-up of unsatisfactory or suspicious smears will be devised in cooperation with the Departments of Pathology and Gynaecology at Dhulikhel Hospital. HPV: cervical specimens are collected by flocked swab and transported to the molecular research laboratory, Dhulikhel Hospital as dry swab. DNA is then extracted using QIAamp DNA Mini Kit, before PCR amplification using two HPV specific primer sets (29) including all oncogenic HPV types. Further typing of specific oncogenic HPV types is done, after extracted DNA is transported to Norway, by additional PCR amplification and DNA sequencing at the Department of medical microbiology, St Olavs Hospital. In addition, a cervical specimen will be tested for HPV by use of the careHPV test (Qiagen), if this test is made available for the study. Chlamydia trachomatis: will be analysed by conventional PCR-amplification using published primers (30), of the same extracted DNA from cervical specimen as described for the HPV-test. HIV-, hepatitis B virus and syphilis infection: will be analysed in serum samples from all study participants by recognized immunological methods already available at Dhulikhel hospital (HIV antibodies, hepatitis B surface antigen (HBsAg) and RPR/TPPA). For confirmation of HIV-positivity, positive samples will be retested by an alternative method. Gonorrhea: cervical swab specimens are transported to the laboratory and inoculated on selective culture medium within the same day for incubation at 37 ºC 24-48 hrs before further typing. Confirmation of suspect colonies as Neisseria gonorrheae is done by microscopy, oxidase reaction, biochemical profile and, if available, by phadebact reaction.