Abundance of HPV L1 Intra-Genotype Variants With Capsid Epitopic Modifications Found Within Low- and High-Grade Pap Smears With Potential Implications for Vaccinology.

Jane Shen-Gunther,Hong Cai,Hao Zhang,Yufeng Wang

doi:10.3389/fgene.2019.00489

Abstract

Background: The aim of this study was to explore the Human Papillomavirus (HPV) genotype composition and intra-genotype variants within individual samples of low- and high-grade cervical cytology by deep sequencing. Clinical, cytological, sequencing, and functional/structural data were forged into an integrated variant profiling pipeline for the detection of potentially vaccine-resistant genotypes or variants.Methods: Low- and high-grade intraepithelial lesion (LSIL and HSIL) cytology samples with +HPV were subjected to amplicon (L1 gene fragment) sequencing by dideoxy (Sanger) and deep methods. Taxonomic, abundance, diversity, and phylogenetic analyses were conducted to determine HPV genotypes/sub-lineages, relative abundance, species diversity and phylogenetic distances within and between samples. Variant detection and functional analysis of translated L1 amino acid sequences determined structural variations of interest.Results: Pure and mixed HPV infections were common among LSIL (n = 6) and HSIL (n = 6) samples. Taxonomic profiling revealed loss of species richness and gain of dominance by carcinogenic genotypes in HSIL samples. Phylogenetic analysis showed excellent correlation between HPV-type specific genetic distances and carcinogenic potential. For combined LSIL/HSIL samples (n = 12), 11 HPV genotypes and 417 mutations were detected: 375 single-nucleotide variants (SNV), 29 insertion/deletion (indel), 12 multi-nucleotide variants (MNV), and 1 replacement variant. The proportion of nonsynonymous mutations was lower for HSIL (0.38) than for LSIL samples (0.51) (p < 0.05). HPV variant analysis pinpointed nucleotide-level mutations and amino acid-level structural modifications.Conclusion: HPV L1 intra-host and intra-genotype variants are abundant in LSIL and HSIL samples with potential functional/structural consequences. An integrated multi-omics approach to variant analysis may provide a sensitive and practical means of detecting changes in HPV evolution and dynamics within individuals or populations.

Highlights

In 1932, Richard Shope isolated the first papillomavirus (PV) from crude extracts of “warty” tumors found on the skin of a wild cottontail rabbit (Shope, 1932)
We focused on the genetic and translated amino acid sequence variations of L1 informed by next-generation sequencing (NGS) for mapping onto the structure of Human Papillomavirus (HPV) antigenic loops as a means of variant profiling and visualization
The proportion of nonsynonymous mutations was lower for high-grade squamous intraepithelial lesion (HSIL) (0.38) than for low-grade squamous intraepithelial lesion (LSIL) samples (0.51) (p = 0.017, Fisher’s exact test) (Figure 7)

Summary

Introduction

In 1932, Richard Shope isolated the first papillomavirus (PV) from crude extracts of “warty” tumors found on the skin of a wild cottontail rabbit (Shope, 1932). Through phylogenetic analysis, Chen et al (2018a) demonstrated that viral nicheadaptation to host ecosystems (tissue tropism) anteceded viralhost codivergence. The PV-host tissue tropism apparently played a vital role in shaping the molecular evolution of oncogenic HPV from archaic hominins to modern humans. HPV-16, an extraordinary result of evolutionary processes over the last 40 million years (Chen et al, 2018a) has emerged as a highly potent carcinogen with a predilection for human mucosa. The aim of this study was to explore the Human Papillomavirus (HPV) genotype composition and intra-genotype variants within individual samples of low- and high-grade cervical cytology by deep sequencing. Cytological, sequencing, and functional/structural data were forged into an integrated variant profiling pipeline for the detection of potentially vaccine-resistant genotypes or variants

Objectives

Methods

Results

Discussion

Conclusion