ultra-deep sequencing, pyrosequencing, genome assembly, graphical models, Bayesian statistics, haplotype inference, HIV-1, drug resistance, virus evolution, antiretroviral therapy
Zagordi O, Geyrhofer L, Roth V, Beerenwinkel N (2012), Deep Sequencing of a Genetically Heterogeneous Sample: Local Haplotype Reconstruction and Read Error Correction, in RECOMB 2012
, Springer, New York.
Zagordi Osvaldo, Geyrhofer Lukas, Roth Volker, Beerenwinkel Niko (2010), Deep sequencing of a genetically heterogeneous sample: local haplotype reconstruction and read error correction., in Journal of computational biology : a journal of computational molecular cell biology
, 17(3), 417-28.
Zagordi O, Klein R, Daumer M, Beerenwinkel N (2010), Error correction of next-generation sequencing data and reliable estimation of HIV quasispecies, in NUCLEIC ACIDS RESEARCH
, 38(21), 7400-7409.
Saeed F, Khokhar A, Zagordi O, Beerenwinkel N (2010), Multiple Sequence Alignment System for Pyrosequencing Reads, in BICoB 2009 -- Bioinformatics and Computational Biology
, Springer, New York.
Zagordi O, Bhattacharya A, Eriksson N, Beerenwinkel N (2011), ShoRAH: estimating the genetic diversity of a mixed sample from next-generation sequencing data, in BMC BIOINFORMATICS
, 12, 119-119.
Niko Beerenwinkel and Osvaldo Zagordi (2011), Ultra-deep sequencing for the analysis of viral populations, in Current Opinion in Virology
, 1, 413-418.
Knoepfel SA, Di Giallonardo F, Daumerd M, Thielen A, Metzner KJ (2011), In-depth analysis of G-to-A hypermutation rate in HIV-1 env DNA induced by endogenous APOBEC3 proteins using massively parallel sequencing, in JOURNAL OF VIROLOGICAL METHODS
, 171(2), 329-338.
Töpfer A., Zagordi O., Prabhakaran S., Roth V., Halperin E. & Beerenwinkel N. (2013), Probabilistic inference of viral quasispecies subject to recombination, in J Comput Biol
, 20, 113-123.
Osvaldo Zagordi and Martin Däumer and Christian Beisel and Niko Beerenwinkel (2012), Read length versus depth of coverage for viral quasispecies reconstruction, in PLOS ONE
, 7, e47046..
Niko Beerenwinkel and Huldrych F. Günthard and Volker Roth and Karin J. Metzner (2012), Challenges and opportunities in estimating viral genetic diversity from next-generation sequencing data., in Front Microbio
, 3, 239.
Armin Töpfer and Dirk Höper and Sandra Blome and Martin Beer and Niko Beerenwinkel and Nicolas Ruggl (2013), Quasispecies analysis of classical swine fever virus by next-generation sequencing, in Virology
, 438(1), 14-19.
Li J. Z., Paredes R., Ribaudo H. J., Kozal M. J., Svarovskaia E. S., Johnson J. A., Geretti A. M., Metzner K. J., Jakobsen M. R., Hullsiek K. H., Ostergaard L., Miller M. D., Kuritzkes D. R. (2013), Impact of Minority Nonnucleoside Reverse Transcriptase Inhibitor Resistance Mutations on Resistance Genotype After Virologic Failure, in Journal of Infectious Diseases
, 207(6), 893-897.
Töpfer Armin, Höper Dirk, Blome Sandra, Beer Martin, Beerenwinkel Niko, Ruggli Nicolas, Leifer Immanuel (2013), Sequencing approach to analyze the role of quasispecies for classical swine fever, in Virology
, 438(1), 14-19.
Giallonardo Francesca Di, Töpfer Armin, Rey Melanie, Prabhakaran Sandhya, Duport Yannick, Leemann Christine, Schmutz Stefan, Campbell Nottania K., Joos Beda, Lecca Maria Rita, Patrignani Andrea, Däumer Martin, Beisel Christian, Rusert Peter, Trkola Alexandra, Günthard Huldrych F., Roth Volker, Beerenwinkel Niko, Metzner Karin J. (2014), Full-length haplotype reconstruction to infer the structure of heterogeneous virus populations, in Nucleic Acids Research
, 42(14), e115-e115.
McElroy Kerensa, Zagordi Osvaldo, Bull Rowena, Luciani Fabio, Beerenwinkel Niko (2013), Accurate single nucleotide variant detection in viral populations by combining probabilistic clustering with a statistical test of strand bias, in BMC Genomics
, 14(1), 501-501.
Di Giallonardo Francesca, Zagordi Osvaldo, Duport Yannick, Leemann Christine, Joos Beda, Künzli-Gontarczyk Marzanna, Bruggmann Rémy, Beerenwinkel Niko, Günthard Huldrych F., Metzner Karin J. (2013), Next-Generation Sequencing of HIV-1 RNA Genomes: Determination of Error Rates and Minimizing Artificial Recombination, in PLoS ONE
, 8(9), e74249-e74249.
Li Jonathan Z, Paredes Roger, Ribaudo Heather J, Svarovskaia Evguenia S, Metzner Karin J, Kozal Michael J, Hullsiek Kathy Huppler, Balduin Melanie, Jakobsen Martin R, Geretti Anna Maria, Thiebaut Rodolphe, Ostergaard Lars, Masquelier Bernard, Johnson Jeffrey A, Miller Michael D, Kuritzkes Daniel R (2011), Low-frequency HIV-1 drug resistance mutations and risk of NNRTI-based antiretroviral treatment failure: a systematic review and pooled analysis., in JAMA
, 305(13), 1327-35.
For many biological species, the traditional view of "one organism, one genome" is insufficient to explain their behavior. The genetic diversity within these species plays an important role for their survival in a given environment. In HIV-1 infection, a diverse population of viruses is maintained in each individual host, facilitating the evolutionary escape of HIV-1 from the host's immune response. The high genetic heterogeneity of HIV-1 quasispecies constitutes a major obstacle in the development of an effective vaccine and it limits therapeutic options due to drug resistant mutants.
In recent years, the next generation of high-throughput DNA sequencing technologies has been introduced. Because of their high coverage based on sequencing many short reads in parallel, this approach is referred to as ultra-deep sequencing. The new sequencing platforms have the potential to resolve the genetic variation in a sample at unprecedented detail by direct sequencing of the mixture of clones. While traditional Sanger sequencing can only infer a consensus sequence of a sample, ultra-deep sequencing generates a very large number of individual reads from the population of interest.
Beyond the standard usage of deep sequencing, we propose a combined approach of ultra-deep sequencing and computational modeling to infer the genetic diversity of intra-host HIV-1 populations. In this joint effort between physician scientists, molecular biologists, computational biologists, and computer scientists, our main goal is to reconstruct the individual full-length haplotypes of HIV-1 populations derived from infected patients and to characterize these quasispecies with respect to interactions among mutations and among individual variants in the context of drug resistance development.
To achieve this goal, we will (1) develop and optimize an experimental methodology for ultra-deep sequencing of full-length HIV-1 virus populations, (2) devise computational and statistical methods for haplotype reconstruction from a set of short, error-prone, observed sequence reads, and (3) analyze 100 pre- and post-treatment patient samples with this approach in order to determine the mechanisms driving the evolution of drug resistance. The comprehensiveness of this study due to the whole-genome and population-wide approach is a unique feature that is possible only with this interdisciplinary approach.
The proposed investigations will increase our understanding of the complex mechanisms of viral escape from the pressure of antiretroviral drugs. The establishment of the new experimental and computational methodologies will be widely applicable to HIV-1 infected patients in Switzerland and beyond. They will further increase the scientific value of the Swiss HIV Cohort Study, a large long-term Swiss collaboration, and they may considerably increase quality of patient care in the future.