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BioMed Research International
Volume 2013 (2013), Article ID 626083, 10 pages
http://dx.doi.org/10.1155/2013/626083
Research Article

Ultradeep Pyrosequencing of Hepatitis C Virus Hypervariable Region 1 in Quasispecies Analysis

1Department of Immunopathology of Infectious and Parasitic Diseases, Medical University of Warsaw, 3c Pawińskiego Street, 02-106 Warsaw, Poland
2Postgraduate School of Molecular Medicine, Żwirki i Wigury 61 Street, 02-091 Warsaw, Poland
3Institute of Medical Virology, University of Zurich, Winterthurerstrasse, 190 8057 Zurich, Switzerland
4Department of Medical Genetics, Medical University of Warsaw, 3c Pawińskiego Street, 02-106 Warsaw, Poland
5Hospital for Infectious Diseases, 37 Wolska Street, 01-201 Warsaw, Poland

Received 22 October 2012; Accepted 12 February 2013

Academic Editor: Ozgur Cogulu

Copyright © 2013 Kamila Caraballo Cortés et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

Genetic variability of hepatitis C virus (HCV) determines pathogenesis of infection, including viral persistence and resistance to treatment. The aim of the present study was to characterize HCV genetic heterogeneity within a hypervariable region 1 (HVR1) of a chronically infected patient by ultradeep 454 sequencing strategy. Three independent sequencing error correction methods were applied. First correction method (Method I) implemented cut-off for genetic variants present in less than 1%. In the second method (Method II), a condition to call a variant was bidirectional coverage of sequencing reads. Third method (Method III) used Short Read Assembly into Haplotypes (ShoRAH) program. After the application of these three different algorithms, HVR1 population consisted of 8, 40, and 186 genetic haplotypes. The most sensitive method was ShoRAH, allowing to reconstruct haplotypes constituting as little as 0.013% of the population. The most abundant genetic variant constituted only 10.5%. Seventeen haplotypes were present in a frequency above 1%, and there was wide dispersion of the population into very sparse haplotypes. Our results indicate that HCV HVR1 heterogeneity and quasispecies population structure may be reconstructed by ultradeep sequencing. However, credible analysis requires proper reconstruction methods, which would distinguish sequencing error from real variability in vivo.