Whole Genome Pyrosequencing of Rare Hepatitis C Virus Genotypes Enhances Subtype Classification and Identification of Naturally Occurring Drug Resistance Variants

Ruchi M Newman, Thomas Kuntzen, Brian Weiner, Andrew Berical, Patrick Charlebois, Carla Kuiken, Donald G Murphy, Peter Simmonds, Phil Bennett, Niall J Lennon, Bruce W Birren, Michael C Zody, Todd M Allen, Matthew R Henn

Research output: Contribution to journalArticlepeer-review

Abstract

Background. Infection with hepatitis C virus (HCV) is a burgeoning worldwide public health problem, with 170 million infected individuals and an estimated 20 million deaths in the coming decades. While six main genotypes generally distinguish the global geographic diversity of HCV, a multitude of closely related subtypes within these genotypes are poorly defined and may influence clinical outcome and treatment options. Unfortunately, the paucity of genetic data from many of these subtypes makes time-consuming primer walking the limiting step for sequencing understudied subtypes.Methods. Here we combined long-range PCR amplification with pyrosequencing for a rapid approach to generate the complete viral coding region of 31 samples representing poorly defined HCV subtypes.Results. Phylogenetic classification based on full genome sequences validated previously identified HCV subtypes, identified a recombinant sequence and a new distinct subtype of genotype 4. Unlike conventional sequencing methods, use of deep sequencing also facilitated characterization of minor drug resistance variants within these uncommon or, in some cases, previously uncharacterized HCV subtypes.Conclusions. These data aid in classification of uncommon HCV subtypes while also providing a high-resolution view of viral diversity within infected patients, which may be relevant to the development of therapeutic regimens to minimize drug resistance.
Original languageEnglish
Pages (from-to)17-31
Number of pages15
JournalThe Journal of Infectious Diseases
Volume208
Issue number1
Early online date6 Nov 2012
DOIs
Publication statusPublished - 1 Jul 2013

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