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Multi-Laboratory Comparison of Next-Generation to Sanger-Based Sequencing for HIV-1 Drug Resistance Genotyping

Next-generation sequencing (NGS) is increasingly used for HIV-1 drug resistance genotyping. NGS methods have the potential for a more sensitive detection of low-abundance variants (LAV) compared to standard Sanger sequencing (SS) methods. A standardized threshold for reporting LAV that generates dat...

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Autores principales: Parkin, Neil T., Avila-Rios, Santiago, Bibby, David F., Brumme, Chanson J., Eshleman, Susan H., Harrigan, P. Richard, Howison, Mark, Hunt, Gillian, Ji, Hezhao, Kantor, Rami, Ledwaba, Johanna, Lee, Emma R., Matías-Florentino, Margarita, Mbisa, Jean L., Noguera-Julian, Marc, Paredes, Roger, Rivera-Amill, Vanessa, Swanstrom, Ronald, Zaccaro, Daniel J., Zhang, Yinfeng, Zhou, Shuntai, Jennings, Cheryl
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2020
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7411816/
https://www.ncbi.nlm.nih.gov/pubmed/32605062
http://dx.doi.org/10.3390/v12070694
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author Parkin, Neil T.
Avila-Rios, Santiago
Bibby, David F.
Brumme, Chanson J.
Eshleman, Susan H.
Harrigan, P. Richard
Howison, Mark
Hunt, Gillian
Ji, Hezhao
Kantor, Rami
Ledwaba, Johanna
Lee, Emma R.
Matías-Florentino, Margarita
Mbisa, Jean L.
Noguera-Julian, Marc
Paredes, Roger
Rivera-Amill, Vanessa
Swanstrom, Ronald
Zaccaro, Daniel J.
Zhang, Yinfeng
Zhou, Shuntai
Jennings, Cheryl
author_facet Parkin, Neil T.
Avila-Rios, Santiago
Bibby, David F.
Brumme, Chanson J.
Eshleman, Susan H.
Harrigan, P. Richard
Howison, Mark
Hunt, Gillian
Ji, Hezhao
Kantor, Rami
Ledwaba, Johanna
Lee, Emma R.
Matías-Florentino, Margarita
Mbisa, Jean L.
Noguera-Julian, Marc
Paredes, Roger
Rivera-Amill, Vanessa
Swanstrom, Ronald
Zaccaro, Daniel J.
Zhang, Yinfeng
Zhou, Shuntai
Jennings, Cheryl
author_sort Parkin, Neil T.
collection PubMed
description Next-generation sequencing (NGS) is increasingly used for HIV-1 drug resistance genotyping. NGS methods have the potential for a more sensitive detection of low-abundance variants (LAV) compared to standard Sanger sequencing (SS) methods. A standardized threshold for reporting LAV that generates data comparable to those derived from SS is needed to allow for the comparability of data from laboratories using NGS and SS. Ten HIV-1 specimens were tested in ten laboratories using Illumina MiSeq-based methods. The consensus sequences for each specimen using LAV thresholds of 5%, 10%, 15%, and 20% were compared to each other and to the consensus of the SS sequences (protease 4–99; reverse transcriptase 38–247). The concordance among laboratories’ sequences at different thresholds was evaluated by pairwise sequence comparisons. NGS sequences generated using the 20% threshold were the most similar to the SS consensus (average 99.6% identity, range 96.1–100%), compared to 15% (99.4%, 88.5–100%), 10% (99.2%, 87.4–100%), or 5% (98.5%, 86.4–100%). The average sequence identity between laboratories using thresholds of 20%, 15%, 10%, and 5% was 99.1%, 98.7%, 98.3%, and 97.3%, respectively. Using the 20% threshold, we observed an excellent agreement between NGS and SS, but significant differences at lower thresholds. Understanding how variation in NGS methods influences sequence quality is essential for NGS-based HIV-1 drug resistance genotyping.
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spelling pubmed-74118162020-08-25 Multi-Laboratory Comparison of Next-Generation to Sanger-Based Sequencing for HIV-1 Drug Resistance Genotyping Parkin, Neil T. Avila-Rios, Santiago Bibby, David F. Brumme, Chanson J. Eshleman, Susan H. Harrigan, P. Richard Howison, Mark Hunt, Gillian Ji, Hezhao Kantor, Rami Ledwaba, Johanna Lee, Emma R. Matías-Florentino, Margarita Mbisa, Jean L. Noguera-Julian, Marc Paredes, Roger Rivera-Amill, Vanessa Swanstrom, Ronald Zaccaro, Daniel J. Zhang, Yinfeng Zhou, Shuntai Jennings, Cheryl Viruses Article Next-generation sequencing (NGS) is increasingly used for HIV-1 drug resistance genotyping. NGS methods have the potential for a more sensitive detection of low-abundance variants (LAV) compared to standard Sanger sequencing (SS) methods. A standardized threshold for reporting LAV that generates data comparable to those derived from SS is needed to allow for the comparability of data from laboratories using NGS and SS. Ten HIV-1 specimens were tested in ten laboratories using Illumina MiSeq-based methods. The consensus sequences for each specimen using LAV thresholds of 5%, 10%, 15%, and 20% were compared to each other and to the consensus of the SS sequences (protease 4–99; reverse transcriptase 38–247). The concordance among laboratories’ sequences at different thresholds was evaluated by pairwise sequence comparisons. NGS sequences generated using the 20% threshold were the most similar to the SS consensus (average 99.6% identity, range 96.1–100%), compared to 15% (99.4%, 88.5–100%), 10% (99.2%, 87.4–100%), or 5% (98.5%, 86.4–100%). The average sequence identity between laboratories using thresholds of 20%, 15%, 10%, and 5% was 99.1%, 98.7%, 98.3%, and 97.3%, respectively. Using the 20% threshold, we observed an excellent agreement between NGS and SS, but significant differences at lower thresholds. Understanding how variation in NGS methods influences sequence quality is essential for NGS-based HIV-1 drug resistance genotyping. MDPI 2020-06-27 /pmc/articles/PMC7411816/ /pubmed/32605062 http://dx.doi.org/10.3390/v12070694 Text en © 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Parkin, Neil T.
Avila-Rios, Santiago
Bibby, David F.
Brumme, Chanson J.
Eshleman, Susan H.
Harrigan, P. Richard
Howison, Mark
Hunt, Gillian
Ji, Hezhao
Kantor, Rami
Ledwaba, Johanna
Lee, Emma R.
Matías-Florentino, Margarita
Mbisa, Jean L.
Noguera-Julian, Marc
Paredes, Roger
Rivera-Amill, Vanessa
Swanstrom, Ronald
Zaccaro, Daniel J.
Zhang, Yinfeng
Zhou, Shuntai
Jennings, Cheryl
Multi-Laboratory Comparison of Next-Generation to Sanger-Based Sequencing for HIV-1 Drug Resistance Genotyping
title Multi-Laboratory Comparison of Next-Generation to Sanger-Based Sequencing for HIV-1 Drug Resistance Genotyping
title_full Multi-Laboratory Comparison of Next-Generation to Sanger-Based Sequencing for HIV-1 Drug Resistance Genotyping
title_fullStr Multi-Laboratory Comparison of Next-Generation to Sanger-Based Sequencing for HIV-1 Drug Resistance Genotyping
title_full_unstemmed Multi-Laboratory Comparison of Next-Generation to Sanger-Based Sequencing for HIV-1 Drug Resistance Genotyping
title_short Multi-Laboratory Comparison of Next-Generation to Sanger-Based Sequencing for HIV-1 Drug Resistance Genotyping
title_sort multi-laboratory comparison of next-generation to sanger-based sequencing for hiv-1 drug resistance genotyping
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7411816/
https://www.ncbi.nlm.nih.gov/pubmed/32605062
http://dx.doi.org/10.3390/v12070694
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