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Next-Generation Sequencing for HIV Drug Resistance Testing: Laboratory, Clinical, and Implementation Considerations
Article

Development and Application of Performance Assessment Criteria for Next-Generation Sequencing-Based HIV Drug Resistance Assays

1
National HIV and Retrovirology Laboratories, National Microbiology Laboratory at JC Wilt Infectious Diseases Research Center, Public Health Agency of Canada, Winnipeg, MB R3E 3R2, Canada
2
ViroDx Clinical Diagnostics Laboratory, St. Louis, MO 63017, USA
3
SutanStats, St. Louis, MO 63017, USA
4
Department of Medical Microbiology and Infectious Diseases, University of Manitoba, Winnipeg, MB R3E 0J9, Canada
*
Author to whom correspondence should be addressed.
These authors contributed equally to this work.
Received: 20 April 2020 / Revised: 29 May 2020 / Accepted: 7 June 2020 / Published: 10 June 2020
(This article belongs to the Special Issue Next Generation Sequencing for HIV Drug Resistance Testing)
Next-generation sequencing (NGS)-based HIV drug resistance (HIVDR) assays outperform conventional Sanger sequencing in scalability, sensitivity, and quantitative detection of minority resistance variants. Thus far, HIVDR assays have been applied primarily in research but rarely in clinical settings. One main obstacle is the lack of standardized validation and performance evaluation systems that allow regulatory agencies to benchmark and accredit new assays for clinical use. By revisiting the existing principles for molecular assay validation, here we propose a new validation and performance evaluation system that helps to both qualitatively and quantitatively assess the performance of an NGS-based HIVDR assay. To accomplish this, we constructed a 70-specimen proficiency test panel that includes plasmid mixtures at known ratios, viral RNA from infectious clones, and anonymized clinical specimens. We developed assessment criteria and benchmarks for NGS-based HIVDR assays and used these to assess data from five separate MiSeq runs performed in two experienced HIVDR laboratories. This proposed platform may help to pave the way for the standardization of NGS HIVDR assay validation and performance evaluation strategies for accreditation and quality assurance purposes in both research and clinical settings. View Full-Text
Keywords: HIV; drug resistance; test; assay; next-generation sequencing; validation; assessment; criteria; benchmarks; standardization HIV; drug resistance; test; assay; next-generation sequencing; validation; assessment; criteria; benchmarks; standardization
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MDPI and ACS Style

Becker, M.G.; Liang, D.; Cooper, B.; Le, Y.; Taylor, T.; Lee, E.R.; Wu, S.; Sandstrom, P.; Ji, H. Development and Application of Performance Assessment Criteria for Next-Generation Sequencing-Based HIV Drug Resistance Assays. Viruses 2020, 12, 627. https://0-doi-org.brum.beds.ac.uk/10.3390/v12060627

AMA Style

Becker MG, Liang D, Cooper B, Le Y, Taylor T, Lee ER, Wu S, Sandstrom P, Ji H. Development and Application of Performance Assessment Criteria for Next-Generation Sequencing-Based HIV Drug Resistance Assays. Viruses. 2020; 12(6):627. https://0-doi-org.brum.beds.ac.uk/10.3390/v12060627

Chicago/Turabian Style

Becker, Michael G., Dun Liang, Breanna Cooper, Yan Le, Tracy Taylor, Emma R. Lee, Sutan Wu, Paul Sandstrom, and Hezhao Ji. 2020. "Development and Application of Performance Assessment Criteria for Next-Generation Sequencing-Based HIV Drug Resistance Assays" Viruses 12, no. 6: 627. https://0-doi-org.brum.beds.ac.uk/10.3390/v12060627

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