To the best of our knowledge, this study was the first to identify VMB biomarkers by performing a systematic exploration of the potential confounding variables of HPV infection . After obtaining metagenomics sequences and other factors through the eHealth platform, a correlation approach was utilized to explore the association between the candidate biomarkers and personal factors. We define stable microbiomes as biomarkers that are not influenced by the status of other crucial factors. The correlation p-value is utilized to select the stable biomarker panels as overlapping for each category.
Participants recruitment and sample collection
This research was approved by the ethics committee of the Institutional Review Board at Beijing Genomics Institution (BGI-IRB 21054). This research is recorded with , ChiCTR2100049221. The recruitment of participants for this study began on and was carried out in a community setting in Shenzhen, Mainland China. Permission gay hookup apps was given by the participants for the research team to use their eHealth data for both health record data including current and previous (within the last 12 months) HPV test results, as well as social personal factors. Based on the HPV test results, participants were grouped into three cohorts: HPV-negative (both samples were HPV negative), HPV-negative conversion (conversion from HPV positive to HPV negative), and HPV-positive subjects (both samples were HPV positive, suggesting persistent infection).
Once the subjects filled in their information in a registration form (S1 Text), they received a metagenomic self-sampling kit via mail, including clear instructions. Participants were requested to abstain from vaginal intercourse 24 hours before sampling, to wait for at least three days after menstrual blood was cleared and to avoid using vaginal douches and any vaginally administered medical treatments [13–16]. A sample of vaginal mucus was collected by inserting a swab into the vagina. The swab was then stirred/placed inside a special tube with a DNA preservative solution N-octylpyridinium bromide (NOPB) [17,18]. The participants were then instructed to close the tube and place it in a plastic bag until the pick-up was arranged, and the sample was collected at room temperature for up to at least 14 days. Both the tube and the bag had a barcode/QR code for identification.
Personal factor and eHealth platform
PROs (personal record outcomes) are defined as reports directly from the participants about the health condition status of the patient’s response without interpretation or amendment by doctor or anyone else. Participants in the study were required to upload their personal PROs, which were divided into five categories with 32 personal factors (S2 Text), on the eHealth platform (CanSeq). It covers several factors, including demographics, medical history, lifestyle (S3–S6 Text), sexual history and behavior and substance abuse factors. The eHealth platform enables noninteractive support for the participants for multiple purposes. First, video and written instructions were provided on the eHealth platform to guide the participants for sample collection. Second, HPV infection records since 2016 are recorded, as authorized by users. Third, the registration of the participant?s information, including evaluated eligibility for participating in the screening program and for the collection of PROs for analytical purposes.
After the metagenomic self-sampling kits were returned, they were sent to the China National GeneBank DataBase (CNGB). DNA was extracted for 38 samples as formerly mentioned [19–21]. Furthermore, DNA libraries were prepared as one paired-end (PE) with 350 bp insert size for individual sample . A length of each read is from 75bp at stage I to 90 bp at stage II. A shotgun of metagenomic was sequenced on BGISEQ-500 platform that is equivalent with other sequencing platforms [22–24]. Data (S7 Text) analysis was carried out using an onsite pipeline, and profiles were uploaded on the online cloud pipeline .