title: Full-length haplotype reconstruction to infer the structure of heterogeneous virus populations.
abstract: Next-generation sequencing (NGS) technologies enable new insights into the diversity of virus populations within their hosts. Diversity estimation is currently restricted to single-nucleotide variants or to local fragments of no more than a few hundred nucleotides defined by the length of sequence reads. To study complex heterogeneous virus populations comprehensively, novel methods are required that allow for complete reconstruction of the individual viral haplotypes. Here, we show that assembly of whole viral genomes of ∼8600 nucleotides length is feasible from mixtures of heterogeneous HIV-1 strains derived from defined combinations of cloned virus strains and from clinical samples of an HIV-1 superinfected individual. Haplotype reconstruction was achieved using optimized experimental protocols and computational methods for amplification, sequencing and assembly. We comparatively assessed the performance of the three NGS platforms 454 Life Sciences/Roche, Illumina and Pacific Biosciences for this task. Our results prove and delineate the feasibility of NGS-based full-length viral haplotype reconstruction and provide new tools for studying evolution and pathogenesis of viruses.
authors: Division of Infectious Diseases and Hospital Epidemiology, University Hospital Zurich, University of Zurich, 8091 Zurich, Switzerland Life Science Zurich Graduate School, University of Zurich, 8057 Zurich, Switzerland.
authors: Francesca Di
authors: FD
authors: Giallonardo
authors: Department of Biosystems Science and Engineering, ETH Zurich, 4058 Basel, Switzerland SIB Swiss Institute of Bioinformatics, 4058 Basel, Switzerland.
authors: Armin
authors: A
authors: Töpfer
authors: Department of Mathematics and Computer Science, University of Basel, 4056 Basel, Switzerland.
authors: Melanie
authors: M
authors: Rey
authors: Department of Mathematics and Computer Science, University of Basel, 4056 Basel, Switzerland.
authors: Sandhya
authors: S
authors: Prabhakaran
authors: Division of Infectious Diseases and Hospital Epidemiology, University Hospital Zurich, University of Zurich, 8091 Zurich, Switzerland.
authors: Yannick
authors: Y
authors: Duport
authors: Division of Infectious Diseases and Hospital Epidemiology, University Hospital Zurich, University of Zurich, 8091 Zurich, Switzerland.
authors: Christine
authors: C
authors: Leemann
authors: Division of Infectious Diseases and Hospital Epidemiology, University Hospital Zurich, University of Zurich, 8091 Zurich, Switzerland.
authors: Stefan
authors: S
authors: Schmutz
authors: Division of Infectious Diseases and Hospital Epidemiology, University Hospital Zurich, University of Zurich, 8091 Zurich, Switzerland Life Science Zurich Graduate School, University of Zurich, 8057 Zurich, Switzerland.
authors: Nottania K
authors: NK
authors: Campbell
authors: Division of Infectious Diseases and Hospital Epidemiology, University Hospital Zurich, University of Zurich, 8091 Zurich, Switzerland.
authors: Beda
authors: B
authors: Joos
authors: Functional Genomics Center Zurich, University of Zurich, ETH Zurich, 8057 Zurich, Switzerland.
authors: Maria Rita
authors: MR
authors: Lecca
authors: Functional Genomics Center Zurich, University of Zurich, ETH Zurich, 8057 Zurich, Switzerland.
authors: Andrea
authors: A
authors: Patrignani
authors: Institut für Immunologie und Genetik, 67655 Kaiserslautern, Germany.
authors: Martin
authors: M
authors: Däumer
authors: Department of Biosystems Science and Engineering, ETH Zurich, 4058 Basel, Switzerland.
authors: Christian
authors: C
authors: Beisel
authors: Institute of Medical Virology, University of Zurich, 8057 Zurich, Switzerland.
authors: Peter
authors: P
authors: Rusert
authors: Institute of Medical Virology, University of Zurich, 8057 Zurich, Switzerland.
authors: Alexandra
authors: A
authors: Trkola
authors: Division of Infectious Diseases and Hospital Epidemiology, University Hospital Zurich, University of Zurich, 8091 Zurich, Switzerland.
authors: Huldrych F
authors: HF
authors: Günthard
authors: Department of Mathematics and Computer Science, University of Basel, 4056 Basel, Switzerland Karin.Metzner@usz.ch.
authors: Volker
authors: V
authors: Roth
authors: Department of Biosystems Science and Engineering, ETH Zurich, 4058 Basel, Switzerland SIB Swiss Institute of Bioinformatics, 4058 Basel, Switzerland Karin.Metzner@usz.ch.
authors: Niko
authors: N
authors: Beerenwinkel
authors: Division of Infectious Diseases and Hospital Epidemiology, University Hospital Zurich, University of Zurich, 8091 Zurich, Switzerland Karin.Metzner@usz.ch.
authors: Karin J
authors: KJ
authors: Metzner
mesh_terms: D014644:Genetic Variation; D016679:Genome, Viral; D015658:HIV Infections / Q000821:virology; D015497:HIV-1 / Q000235:genetics*; D006239:Haplotypes; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D006801:Humans