HPV Genotyping of Modified General Primer-Amplicons Is More Analytically Sensitive and Specific by Sequencing than by Hybridization

Roger Meisal; Trine Ballestad Rounge; Irene Kraus Christiansen; Alexander Kirkeby Eieland; Merete Molton Worren; Tor Faksvaag Molden; Øyvind Kommedal; Eivind Hovig; Truls Michael Leegaard; Ole Herman Ambur

doi:10.1371/journal.pone.0169074

HPV Genotyping of Modified General Primer-Amplicons Is More Analytically Sensitive and Specific by Sequencing than by Hybridization

PLoS One. 2017 Jan 3;12(1):e0169074. doi: 10.1371/journal.pone.0169074. eCollection 2017.

Authors

Affiliations

¹ Department of Microbiology and Infection Control, Akershus University Hospital, Lørenskog, Norway.
² Department of Research, Cancer Registry of Norway, Oslo, Norway.
³ Bioinformatics Core Facility, Institute for Cancer Genetics and Informatics, The Norwegian Radium Hospital, Oslo University Hospital, Oslo, Norway.
⁴ Department of Vaccines, Norwegian Institute of Public Health, Oslo, Norway.
⁵ Department of Microbiology, Haukeland University Hospital, Bergen, Norway.
⁶ Department of Informatics, University of Oslo, Oslo, Norway.
⁷ Department of Tumor Biology, Institute of Cancer Research and Institute for Cancer Genetics and Informatics, The Norwegian Radium Hospital, Oslo University Hospital, Oslo, Norway.
⁸ Division of Medicine and Laboratory Sciences, University of Oslo, Oslo Norway.
⁹ Department of Life Sciences and Health, Oslo and Akershus University College of Applied Sciences, Oslo, Norway.

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.

MeSH terms

Adolescent
Adult
Chlamydia Infections
DNA Primers / genetics*
DNA, Viral / genetics
Female
Genetic Variation
Genotyping Techniques / methods
High-Throughput Nucleotide Sequencing / methods*
Humans
Hybridization, Genetic
Nucleic Acid Hybridization*
Papillomaviridae / genetics*
Papillomavirus Infections / diagnosis
Papillomavirus Infections / virology*
Sensitivity and Specificity
Uterine Cervical Neoplasms / diagnosis
Uterine Cervical Neoplasms / virology*
Young Adult

Substances

DNA Primers
DNA, Viral

Grants and funding

Head of Dept. Merete Holth and the Dept. Microbiology and Infection Control for additional financial support. The majority of the work was funded by a Norwegian governmental grant to the National Institute of Public Health for the national surveillance of the HPV vaccination program.