Scalable long-read nanopore HPV16 amplicon-based whole-genome sequencing
Human papillomavirus 16 (HPV16) drives precursor cervical lesions that often progress to cervical cancer (CC). Variation within the HPV16 genome has been associated with CC risk. Here, we developed an affordable and portable amplicon-based long-read whole genome sequencing (WGS) approach using Oxford Nanopore Technologies to investigate HPV16 genetic diversity among women in sub-Saharan African countries. Applied to a control CaSki cell line and clinical samples (n = 12), our method generated complete HPV16 genomes at high coverage (median read coverage: 5,899-15,279 ×). Benchmarking our HPV16 controls showed high accuracy for two variant calling pipelines (Clair3 and PEPPER-Margin DeepVariant). Phylogenetic analysis identified all four previously defined HPV16 lineages (A-D) and their high-risk sublineages. All lineages exhibited strong concordance across de novo assembly, reference-based phylogenetics, and unsupervised clustering. Our pipeline effectively captured the full extent of genomic variation, including putative lineage-informative SNPs. This method offers a robust amplicon-based WGS and analysis pipeline for HPV16, making it well-suited for integration into surveillance, diagnostics, and epidemiological efforts in low-resource areas.
Journal
Scientific ReportsAuthors
Funding
Providence/Boston Center for AIDS Research (CFAR)The East Africa Consortium for HPV and Cervical Cancer in Women living with HIV/AIDSBrown/Moi Training Program for the Prevention of HIV related Cervical CancerAMPATH-Oncology Institute: HPV and Cervical Cancer in Kenyan Women with HIV/AIDSNational Institutes of Health (NIH), National Cancer Institute (NCI), United States of America Grant 5U54CA190151-02 and 5U54CA254518-03Providence/Boston Center for AIDS Research (CFAR)Brown/Moi Training Program for the Prevention of HIV related Cervical Cancer