DOI: 10.57760/sciencedb.11565
title: A DATASET OF HPV GENOTYPES AMONG WOMEN ATTENDING REPRODUCTIVE HEALTH CLINICS IN LAKE VICTORIA BASIN KENYA IN 2023
title: A DATASET OF HPV GENOTYPES AMONG WOMEN ATTENDING REPRODUCTIVE HEALTH CLINICS IN LAKE VICTORIA BASIN KENYA IN 2023

description: Sample Collection/Storage After obtaining informed consent from the study participants, each participant was given standardized questionnaires in English or local languages (Luo) to fill and those who needed assistance were helped by the research assistants. The structured questionnaires focused on their socio-demographic characteristics, past reproductive health history, sexual behaviors, and other risk factors associated with potential HR/HR HPV (see supplementary information). The HIV status of each woman was confirmed from the hospital data registry where each woman receiving reproductive health services is encouraged to undergo HIV testing before getting routine services and their results are recorded in Ministry of Health registry for HIV Surveillance. Each woman consented to have this information obtained for the purpose of this study only. To maintain privacy, a registered Reproductive Health Clinic nurse conducted a physical examination and sample collection in a separate room.  A sterile vaginal speculum was inserted into the vagina to obtain a cervical swab for HPV testing. The specimens were obtained using the multi-collect specimen collection kit (Abbott). The collection swab was rotated twice in the ectocervical and endocervical region and then deposited into 1.2ml of transport buffer that contains guanidine thiocyanate (for DNA stabilization), according to manufacturer instructions. The specimens were then stored at -80℃ in the Western Kenya Cancer Care and Research Center laboratory for preservation till shipment to Antwerp, Belgium for molecular analysis.RIATOL HPV genotyping qPCR assayThe RIATOL qPCR HPV assay was used to extract and genotype HPV-DNA at AML, Sonic Healthcare Benelux (Antwerp, Belgium) under ISO15189 accreditation, as previously described (1). The choice to use RIATOL HPV genotyping qPCR assay was decided on because of its proven clinical accuracy, reproducibility and quantifiable results as it provides viral load information. Briefly, cervical samples were collected with the multi-Collect Specimen Collection Kit (Abbott) and 300 µl of each sample was transferred into the 96-deep well plate by using Janus G3 (Perkin Elmer) robot for batch processing. DNA was extracted in the automatic nucleic preparation in Chemargic360 (Perkin Elmer) using the viral DNA/RNA 360 H96 extraction kit. The DNA extract was then stored at 4℃ awaiting PCR. As an internal cellular control, beta-globin was employed to assess the validity of the PCR run. To achieve relative quantification of data and confirm the qPCR acceptable efficiency ranges of 90-110%, standard curves were generated by amplification of a series of synthetic DNA ten-fold dilutions. Subsequently, 384-PCR well plates were prepared by the Janus Liquid handler robot (Perkin Elmer) by mixing 2 µL of extracted DNA or specific control with 4 µL of the assigned master mix solutions, as described (1). In total, each sample was tested with 8 different master mixes, including primers and probes for detection of HPV 6, 11, 16, 18, 31, 33,35, 39, 45,51, 52, 53 56, 58, 59, 66, 67, 68 and beta-globin. RT-amplification was carried out on the Light Cycler 480 (Roche) instrument. The thermo-cycling profile consisted of 45 two-step cycles: 10s at 95℃ and 30s at 60℃. At the end of the amplification run, the RT-PCR run file automatically was transferred to the Fast-Finder online AI platform (Velsera) for analysis where clinical cut-offs were applied to determine positive and negative samples.The Genotypes have been entered into the Excel sheet.
description: Sample Collection/Storage After obtaining informed consent from the study participants, each participant was given standardized questionnaires in English or local languages (Luo) to fill and those who needed assistance were helped by the research assistants. The structured questionnaires focused on their socio-demographic characteristics, past reproductive health history, sexual behaviors, and other risk factors associated with potential HR/HR HPV (see supplementary information). The HIV status of each woman was confirmed from the hospital data registry where each woman receiving reproductive health services is encouraged to undergo HIV testing before getting routine services and their results are recorded in Ministry of Health registry for HIV Surveillance. Each woman consented to have this information obtained for the purpose of this study only. To maintain privacy, a registered Reproductive Health Clinic nurse conducted a physical examination and sample collection in a separate room.  A sterile vaginal speculum was inserted into the vagina to obtain a cervical swab for HPV testing. The specimens were obtained using the multi-collect specimen collection kit (Abbott). The collection swab was rotated twice in the ectocervical and endocervical region and then deposited into 1.2ml of transport buffer that contains guanidine thiocyanate (for DNA stabilization), according to manufacturer instructions. The specimens were then stored at -80℃ in the Western Kenya Cancer Care and Research Center laboratory for preservation till shipment to Antwerp, Belgium for molecular analysis.RIATOL HPV genotyping qPCR assayThe RIATOL qPCR HPV assay was used to extract and genotype HPV-DNA at AML, Sonic Healthcare Benelux (Antwerp, Belgium) under ISO15189 accreditation, as previously described (1). The choice to use RIATOL HPV genotyping qPCR assay was decided on because of its proven clinical accuracy, reproducibility and quantifiable results as it provides viral load information. Briefly, cervical samples were collected with the multi-Collect Specimen Collection Kit (Abbott) and 300 µl of each sample was transferred into the 96-deep well plate by using Janus G3 (Perkin Elmer) robot for batch processing. DNA was extracted in the automatic nucleic preparation in Chemargic360 (Perkin Elmer) using the viral DNA/RNA 360 H96 extraction kit. The DNA extract was then stored at 4℃ awaiting PCR. As an internal cellular control, beta-globin was employed to assess the validity of the PCR run. To achieve relative quantification of data and confirm the qPCR acceptable efficiency ranges of 90-110%, standard curves were generated by amplification of a series of synthetic DNA ten-fold dilutions. Subsequently, 384-PCR well plates were prepared by the Janus Liquid handler robot (Perkin Elmer) by mixing 2 µL of extracted DNA or specific control with 4 µL of the assigned master mix solutions, as described (1). In total, each sample was tested with 8 different master mixes, including primers and probes for detection of HPV 6, 11, 16, 18, 31, 33,35, 39, 45,51, 52, 53 56, 58, 59, 66, 67, 68 and beta-globin. RT-amplification was carried out on the Light Cycler 480 (Roche) instrument. The thermo-cycling profile consisted of 45 two-step cycles: 10s at 95℃ and 30s at 60℃. At the end of the amplification run, the RT-PCR run file automatically was transferred to the Fast-Finder online AI platform (Velsera) for analysis where clinical cut-offs were applied to determine positive and negative samples.The Genotypes have been entered into the Excel sheet.