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2016-01-12Zeitschriftenartikel DOI: 10.1371/journal.pone.0146687
A Follow-Up of the Multicenter Collaborative Study on HIV-1 Drug Resistance and Tropism Testing Using 454 Ultra Deep Pyrosequencing
dc.contributor.authorJohn, Elizabeth P. St.
dc.contributor.authorSimen, Birgitte B.
dc.contributor.authorTurenchalk, Gregory S.
dc.contributor.authorBraverman, Michael S.
dc.contributor.authorAbbate, Isabella
dc.contributor.authorAerssens, Jeroen
dc.contributor.authorBouchez, Olivier
dc.contributor.authorGabriel, Christian
dc.contributor.authorIzopet, Jacques
dc.contributor.authorMeixenberger, Karolin
dc.contributor.authorGiallonardo, Francesca Di
dc.contributor.authorSchlapbach, Ralph
dc.contributor.authorParedes, Roger
dc.contributor.authorSakwa, James
dc.contributor.authorSchmitz-Agheguian, Gudrun G.
dc.contributor.authorThielen, Alexander
dc.contributor.authorVictor, Martin
dc.contributor.authorMetzner, Karin J.
dc.contributor.authorDäumer, Martin P.
dc.date.accessioned2018-05-07T18:49:26Z
dc.date.available2018-05-07T18:49:26Z
dc.date.created2016-02-01
dc.date.issued2016-01-12none
dc.identifier.otherhttp://edoc.rki.de/oa/articles/reJ7Kc8EIRzb6/PDF/24HJgCUuQWizQ.pdf
dc.identifier.urihttp://edoc.rki.de/176904/2249
dc.description.abstractBackground: Ultra deep sequencing is of increasing use not only in research but also in diagnostics. For implementation of ultra deep sequencing assays in clinical laboratories for routine diagnostics, intra- and inter-laboratory testing are of the utmost importance. Methods: A multicenter study was conducted to validate an updated assay design for 454 Life Sciences’ GS FLX Titanium system targeting protease/reverse transcriptase (RTP) and env (V3) regions to identify HIV-1 drug-resistance mutations and determine co-receptor use with high sensitivity. The study included 30 HIV-1 subtype B and 6 subtype non-B samples with viral titers (VT) of 3,940–447,400 copies/mL, two dilution series (52,129–1,340 and 25,130–734 copies/mL), and triplicate samples. Amplicons spanning PR codons 10–99, RT codons 1–251 and the entire V3 region were generated using barcoded primers. Analysis was performed using the GS Amplicon Variant Analyzer and geno2pheno for tropism. For comparison, population sequencing was performed using the ViroSeq HIV-1 genotyping system. Results: The median sequencing depth across the 11 sites was 1,829 reads per position for RTP (IQR 592–3,488) and 2,410 for V3 (IQR 786–3,695). 10 preselected drug resistant variants were measured across sites and showed high inter-laboratory correlation across all sites with data (P20% were missed, variants 2–10% were detected at most sites (even at low VT), and variants 1–2% were detected by some sites. All mutations detected by population sequencing were also detected by UDS. Conclusions: This assay design results in an accurate and reproducible approach to analyze HIV-1 mutant spectra, even at variant frequencies well below those routinely detectable by population sequencing.eng
dc.language.isoeng
dc.publisherRobert Koch-Institut, Infektionskrankheiten / Erreger
dc.subjectHumanseng
dc.subjectReproducibility of Resultseng
dc.subjectHIV-1/geneticseng
dc.subjectCooperative Behavioreng
dc.subjectFollow-Up Studieseng
dc.subjectDrug Resistance Viral/geneticseng
dc.subjectPlasmids/geneticseng
dc.subjectHigh-Throughput Nucleotide Sequencing/methodseng
dc.subjectAmino Acids/geneticseng
dc.subjectMutation/geneticseng
dc.subjectTropism/geneticseng
dc.subject.ddc610 Medizin
dc.titleA Follow-Up of the Multicenter Collaborative Study on HIV-1 Drug Resistance and Tropism Testing Using 454 Ultra Deep Pyrosequencing
dc.typeperiodicalPart
dc.identifier.urnurn:nbn:de:0257-10042878
dc.identifier.doi10.1371/journal.pone.0146687
dc.identifier.doihttp://dx.doi.org/10.25646/2174
local.edoc.container-titlePLoS ONE
local.edoc.container-textSt. John EP, Simen BB, Turenchalk GS, Braverman MS, Abbate I, Aerssens J, et al. (2016) A Follow-Up of the Multicenter Collaborative Study on HIV-1 Drug Resistance and Tropism Testing Using 454 Ultra Deep Pyrosequencing. PLoS ONE 11(1): e0146687.
local.edoc.fp-subtypeArtikel
local.edoc.type-nameZeitschriftenartikel
local.edoc.container-typeperiodical
local.edoc.container-type-nameZeitschrift
local.edoc.container-urlhttp://journals.plos.org/plosone/article?id=10.1371/journal.pone.0146687
local.edoc.container-publisher-namePublic Library of Science
local.edoc.container-volume11
local.edoc.container-issue1
local.edoc.container-year2016

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