Abstract
BackgroundHPV genomic variation may be involved in viral carcinogenesis.MethodsIn a national register-based nested case–control study, we retrieved archival smears from baseline cytologically normal women who later developed cancer in situ (CIS), squamous cervical cancer (SCC) or remained free of disease. These smears were previously HPV tested by PCR and HPV16 was the strongest risk factor. We now used the Illumina NextSeq platform to sequence HPV16 genomes in cervical smears from 242 women who later developed CIS/CIN3 (n = 134), SCC (n = 92) or remained healthy (n = 16).ResultsThe median sequence depth per sample was high (11,288×). For 218/242 samples (>90%), we covered ≥80% of the complete HPV16 genome with sequencing median depths of >200×. We identified a wide range of unique isolates and 147 novel SNPs across the 218 samples. Most women (97%) had HPV16 lineage A infection, with the sublineages being A1 (66.1%), A2 (28.9%) and A4 (1.8%), respectively. The least variable gene was the E7 (3.4% variability), where 170/204 case women (83%) displayed a fully conserved sequence. There were no obvious differences by disease outcome (CIS or SCC).ConclusionsWe found a high number of novel SNPs. The E7 gene was hypovariable both among women later developing CIN3/CIS, and SCC, respectively.
Highlights
human papillomavirus (HPV) genomic variation may be involved in viral carcinogenesis
Novel SNPs We identified up to 147 novel SNPs, including 26 SNPs located within non-coding regions, 26 silent substitutions, 86 non-synonymous nucleotide variations, 5 nonsense substitutions and 4 SNPs that were classified as both silent and non-synonymous, depending on the protein to translate (E4 vs E2 or L2 vs L1)
We identified mainly variant lineage A-positive women with a large amount of SNPs, 24.6% of which were classified as novel, and confirmed that the E7 was the HPV gene that exhibits the least variability across samples
Summary
METHODS: In a national register-based nested case–control study, we retrieved archival smears from baseline cytologically normal women who later developed cancer in situ (CIS), squamous cervical cancer (SCC) or remained free of disease. These smears were previously HPV tested by PCR and HPV16 was the strongest risk factor. We used the Illumina NextSeq platform to sequence HPV16 genomes in cervical smears from 242 women who later developed CIS/CIN3 (n = 134), SCC (n = 92) or remained healthy (n = 16). CONCLUSIONS: We found a high number of novel SNPs. The E7 gene was hypovariable both among women later developing CIN3/CIS, and SCC, respectively. The four identified variants of HPV16 are divided into sublineages A1–3 (formerly termed European), A4 (Asian); B (African-1), C (African-2) and D1–3
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