Abstract
Sensitive and specific genotyping of human papillomaviruses (HPVs) is important for population-based surveillance of carcinogenic HPV types and for monitoring vaccine effectiveness. Here we compare HPV genotyping by Next Generation Sequencing (NGS) to an established DNA hybridization method. In DNA isolated from urine, the overall analytical sensitivity of NGS was found to be 22% higher than that of hybridization. NGS was also found to be the most specific method and expanded the detection repertoire beyond the 37 types of the DNA hybridization assay. Furthermore, NGS provided an increased resolution by identifying genetic variants of individual HPV types. The same Modified General Primers (MGP)-amplicon was used in both methods. The NGS method is described in detail to facilitate implementation in the clinical microbiology laboratory and includes suggestions for new standards for detection and calling of types and variants with improved resolution.
Highlights
Persistent infection with certain human papillomavirus (HPV) types is strongly associated with cervical cancer development [1]
Positive control samples consisting of HPV type reference plasmids (HPV16 and 18) and cell lines showed 100% agreement in regard to the detection limits for the two methods and with the 20 read depth Next Generation Sequencing (NGS) cut-off
The 11 unconfirmed calls made by hybridization were for HPV types 6, 33, 67, 68, 82, 90 and 91 (S3 Table)
Summary
Persistent infection with certain human papillomavirus (HPV) types is strongly associated with cervical cancer development [1]. HPV types associated with cancer development in humans all belong to the genus alpha-HPVs [2,3]. Based on the degree of association with malignancy, HPVs can be divided into high-risk (HR) and low-risk types. 14 HPV types are regarded HR types (HPV16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 66 and 68) and 12 types as low-risk types (HPV6, 11, 40, 42, 43, 44, 54, 61, 72, 81, and 89) [2,4]. Types 16 and 18 are the most prevalent HR types and account for 70% of HPV infections found in invasive cervical cancer [5]. With the exception of HPV51 (alpha-5), HPV56 and PLOS ONE | DOI:10.1371/journal.pone.0169074. With the exception of HPV51 (alpha-5), HPV56 and PLOS ONE | DOI:10.1371/journal.pone.0169074 January 3, 2017
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