{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,11,9]],"date-time":"2025-11-09T07:51:02Z","timestamp":1762674662181,"version":"build-2065373602"},"reference-count":32,"publisher":"MDPI AG","issue":"9","license":[{"start":{"date-parts":[[2023,8,25]],"date-time":"2023-08-25T00:00:00Z","timestamp":1692921600000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"D3\/Penta 21 trial","award":["NCT04337450","207742","MC_UU_00004\/03"],"award-info":[{"award-number":["NCT04337450","207742","MC_UU_00004\/03"]}]},{"name":"ViiV Healthcare","award":["NCT04337450","207742","MC_UU_00004\/03"],"award-info":[{"award-number":["NCT04337450","207742","MC_UU_00004\/03"]}]},{"name":"UCLH APDU","award":["NCT04337450","207742","MC_UU_00004\/03"],"award-info":[{"award-number":["NCT04337450","207742","MC_UU_00004\/03"]}]},{"DOI":"10.13039\/501100000265","name":"UK Medical Research Council core funding","doi-asserted-by":"publisher","award":["NCT04337450","207742","MC_UU_00004\/03"],"award-info":[{"award-number":["NCT04337450","207742","MC_UU_00004\/03"]}],"id":[{"id":"10.13039\/501100000265","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Viruses"],"abstract":"<jats:p>Modern HIV-1 treatment effectively suppresses viral amplification in people living with HIV. However, the persistence of HIV-1 DNA as proviruses integrated into the human genome remains the main barrier to achieving a cure. Next-generation sequencing (NGS) offers increased sensitivity for characterising archived drug resistance mutations (DRMs) in HIV-1 DNA for improved treatment options. In this study, we present an ultra-sensitive targeted PCR assay coupled with NGS and a robust pipeline to characterise HIV-1 DNA DRMs from buffy coat samples. Our evaluation supports the use of this assay for Pan-HIV-1 analyses with reliable detection of DRMs across the HIV-1 Pol region. We propose this assay as a new valuable tool for monitoring archived HIV-1 drug resistance in virologically suppressed individuals, especially in clinical trials investigating novel therapeutic approaches.<\/jats:p>","DOI":"10.3390\/v15091811","type":"journal-article","created":{"date-parts":[[2023,8,25]],"date-time":"2023-08-25T08:33:09Z","timestamp":1692952389000},"page":"1811","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":2,"title":["Sensitive HIV-1 DNA Pol Next-Generation Sequencing for the Characterisation of Archived Antiretroviral Drug Resistance"],"prefix":"10.3390","volume":"15","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-7635-7810","authenticated-orcid":false,"given":"Johannes C.","family":"Botha","sequence":"first","affiliation":[{"name":"Great Ormond Street Institute of Child Health, University College London, London WC1N 1EH, UK"},{"name":"Advanced Pathogen Diagnostics Unit, University College London Hospitals NHS Trust, London NW1 2PG, UK"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Matthew","family":"Byott","sequence":"additional","affiliation":[{"name":"Great Ormond Street Institute of Child Health, University College London, London WC1N 1EH, UK"},{"name":"Advanced Pathogen Diagnostics Unit, University College London Hospitals NHS Trust, London NW1 2PG, UK"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Moira J.","family":"Spyer","sequence":"additional","affiliation":[{"name":"Great Ormond Street Institute of Child Health, University College London, London WC1N 1EH, UK"},{"name":"Advanced Pathogen Diagnostics Unit, University College London Hospitals NHS Trust, London NW1 2PG, UK"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Paul R.","family":"Grant","sequence":"additional","affiliation":[{"name":"Health Services Laboratories, London WC1H 9AX, UK"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Kathleen","family":"G\u00e4rtner","sequence":"additional","affiliation":[{"name":"Great Ormond Street Institute of Child Health, University College London, London WC1N 1EH, UK"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Wilson X.","family":"Chen","sequence":"additional","affiliation":[{"name":"ViiV Healthcare, Durham, NC 27709, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"James","family":"Burton","sequence":"additional","affiliation":[{"name":"Great Ormond Street Hospital for Children NHS Foundation Trust, London WC1N 3JH, UK"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Alasdair","family":"Bamford","sequence":"additional","affiliation":[{"name":"Great Ormond Street Institute of Child Health, University College London, London WC1N 1EH, UK"},{"name":"Great Ormond Street Hospital for Children NHS Foundation Trust, London WC1N 3JH, UK"},{"name":"Medical Research Council Clinical Trials Unit, University College London, London WC1E 6BT, UK"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Laura J.","family":"Waters","sequence":"additional","affiliation":[{"name":"Central and North West London NHS Foundation Trust, Mortimer Market, London WC1E 6JB, UK"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Carlo","family":"Giaquinto","sequence":"additional","affiliation":[{"name":"Department of Women and Child Health, University of Padova, 35122 Padova, Italy"},{"name":"Fondazione Penta ETS, 35127 Padova, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-7008-4145","authenticated-orcid":false,"given":"Anna","family":"Turkova","sequence":"additional","affiliation":[{"name":"Great Ormond Street Hospital for Children NHS Foundation Trust, London WC1N 3JH, UK"},{"name":"Medical Research Council Clinical Trials Unit, University College London, London WC1E 6BT, UK"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Cindy L.","family":"Vavro","sequence":"additional","affiliation":[{"name":"ViiV Healthcare, Durham, NC 27709, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Eleni","family":"Nastouli","sequence":"additional","affiliation":[{"name":"Great Ormond Street Institute of Child Health, University College London, London WC1N 1EH, UK"},{"name":"Advanced Pathogen Diagnostics Unit, University College London Hospitals NHS Trust, London NW1 2PG, UK"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2023,8,25]]},"reference":[{"key":"ref_1","unstructured":"Department of Health and Human Services Panel on Antiretroviral Guidelines for Adults and Adolescents (2023, May 19). Guidelines for the Use of Antiretroviral Agents in Adults and Adolescents with HIV, Available online: https:\/\/clinicalinfo.hiv.gov\/sites\/default\/files\/guidelines\/documents\/AdultandAdolescentGL.pdf."},{"key":"ref_2","unstructured":"European AIDS Clinical Society (2023, May 19). Guidelines Version 11.0 October 2021. Available online: https:\/\/www.eacsociety.org\/media\/final2021eacsguidelinesv11.0_oct2021.pdf."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"13193","DOI":"10.1073\/pnas.94.24.13193","article-title":"Presence of an Inducible HIV-1 Latent Reservoir during Highly Active Antiretroviral\u2009Therapy","volume":"94","author":"Chun","year":"1997","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"1295","DOI":"10.1126\/science.278.5341.1295","article-title":"Identification of a Reservoir for HIV-1 in Patients on Highly Active Antiretroviral Therapy","volume":"278","author":"Finzi","year":"1997","journal-title":"Science"},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"190","DOI":"10.1016\/j.virol.2010.01.033","article-title":"Genetic Characterization of HIV-1 from Semen and Blood from Clade C-Infected Subjects from India and Effect of Therapy in These Body Compartments","volume":"401","author":"Shen","year":"2010","journal-title":"Virology"},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"6024","DOI":"10.1128\/JVI.00131-11","article-title":"Intercompartmental Recombination of HIV-1 Contributes to Env Intrahost Diversity and Modulates Viral Tropism and Sensitivity to Entry Inhibitors","volume":"85","author":"Brown","year":"2011","journal-title":"J. Virol."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"383","DOI":"10.1097\/COH.0000000000000281","article-title":"Diversity of HIV-1 Reservoirs in CD4+ T-Cell Subpopulations","volume":"11","author":"Lee","year":"2016","journal-title":"Curr. Opin. HIV AIDS"},{"key":"ref_8","doi-asserted-by":"crossref","unstructured":"Paquet, A.C., Baxter, J., Weidler, J., Lie, Y., Lawrence, J., Kim, R., Bates, M., Coakley, E., and Chappey, C. (2011). Differences in Reversion of Resistance Mutations to Wild-Type under Structured Treatment Interruption and Related Increase in Replication Capacity. PLoS ONE, 6.","DOI":"10.1371\/journal.pone.0014638"},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"1608","DOI":"10.1086\/503914","article-title":"Web Resources for HIV Type 1 Genotypic-Resistance Test Interpretation","volume":"42","author":"Liu","year":"2006","journal-title":"Clin. Infect. Dis."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"182","DOI":"10.1097\/COH.0000000000000344","article-title":"Future Technologies for Monitoring HIV Drug Resistance and Cure","volume":"12","author":"Parikh","year":"2017","journal-title":"Curr. Opin. HIV AIDS"},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"69","DOI":"10.1016\/j.virusres.2016.10.019","article-title":"Deep Sequencing for HIV-1 Clinical Management","volume":"239","author":"Paredes","year":"2017","journal-title":"Virus Res."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"1327","DOI":"10.1001\/jama.2011.375","article-title":"Low-Frequency HIV-1 Drug Resistance Mutations and Risk of NNRTI-Based Antiretroviral Treatment Failure: A Systematic Review and Pooled Analysis","volume":"305","author":"Li","year":"2011","journal-title":"JAMA"},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"104229","DOI":"10.1016\/j.jcv.2019.104229","article-title":"Performance Evaluation of the Vela Dx Sentosa Next-Generation Sequencing System for HIV-1 DNA Genotypic Resistance","volume":"122","author":"Raymond","year":"2020","journal-title":"J. Clin. Virol."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"104207","DOI":"10.1016\/j.jcv.2019.104207","article-title":"Routine Drug Resistance Testing in HIV-1 Proviral DNA, Using an Automated next- Generation Sequencing Assay","volume":"121","author":"Alidjinou","year":"2019","journal-title":"J. Clin. Virol."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"29","DOI":"10.1128\/JCM.39.1.29-33.2001","article-title":"Simple, Sensitive, and Specific Detection of Human Immunodeficiency Virus Type 1 Subtype B DNA in Dried Blood Samples for Diagnosis in Infants in the Field","volume":"39","author":"Beck","year":"2001","journal-title":"J. Clin. Microbiol."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"1917","DOI":"10.1093\/jac\/dky103","article-title":"Low Prevalence of Transmitted HIV-1 Drug Resistance Detected by a Dried Blood Spot (DBS)-Based next-Generation Sequencing (NGS) Method in Newly Diagnosed Individuals in Cameroon in the Years 2015\u201316","volume":"73","author":"Mbunkah","year":"2018","journal-title":"J. Antimicrob. Chemother."},{"key":"ref_17","doi-asserted-by":"crossref","unstructured":"Parkin, N.T., Avila-Rios, S., Bibby, D.F., Brumme, C.J., Eshleman, S.H., Harrigan, P.R., Howison, M., Hunt, G., Ji, H., and Kantor, R. (2020). Multi-Laboratory Comparison of Next-Generation to Sanger-Based Sequencing for HIV-1 Drug Resistance Genotyping. Viruses, 12.","DOI":"10.3390\/v12070694"},{"key":"ref_18","doi-asserted-by":"crossref","unstructured":"Arias, A., L\u00f3pez, P., S\u00e1nchez, R., Yamamura, Y., and Rivera-Amill, V. (2018). Sanger and Next Generation Sequencing Approaches to Evaluate HIV-1 Virus in Blood Compartments. Int. J. Environ. Res. Public. Health, 15.","DOI":"10.3390\/ijerph15081697"},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"1793","DOI":"10.1007\/s40121-022-00676-y","article-title":"HIV DNA Sequencing to Detect Archived Antiretroviral Drug Resistance","volume":"11","author":"Geretti","year":"2022","journal-title":"Infect. Dis. Ther."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"3555","DOI":"10.1093\/jac\/dkz347","article-title":"Switching to Bictegravir\/Emtricitabine\/Tenofovir Alafenamide Maintained HIV-1 RNA Suppression in Participants with Archived Antiretroviral Resistance Including M184V\/I","volume":"74","author":"Andreatta","year":"2019","journal-title":"J. Antimicrob. Chemother."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"5847","DOI":"10.1172\/JCI138099","article-title":"HIV-1 Viremia Not Suppressible by Antiretroviral Therapy Can Originate from Large T Cell Clones Producing Infectious Virus","volume":"130","author":"Halvas","year":"2020","journal-title":"J. Clin. Investig."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"58","DOI":"10.1016\/j.jcv.2018.07.008","article-title":"Quantification of Total HIV-1 DNA in Buffy Coat Cells, Feasibility and Potential Added Value for Clinical Follow-up of HIV-1 Infected Patients on ART","volume":"106","author":"Mortier","year":"2018","journal-title":"J. Clin. Virol."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"1209","DOI":"10.1038\/s41598-017-01221-5","article-title":"Instability of 8E5 Calibration Standard Revealed by Digital PCR Risks Inaccurate Quantification of HIV DNA in Clinical Samples by QPCR","volume":"7","author":"Busby","year":"2017","journal-title":"Sci. Rep."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"2114","DOI":"10.1093\/bioinformatics\/btu170","article-title":"Trimmomatic: A Flexible Trimmer for Illumina Sequence Data","volume":"30","author":"Bolger","year":"2014","journal-title":"Bioinformatics"},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"R25","DOI":"10.1186\/gb-2009-10-3-r25","article-title":"Ultrafast and Memory-Efficient Alignment of Short DNA Sequences to the Human Genome","volume":"10","author":"Langmead","year":"2009","journal-title":"Genome Biol."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"8","DOI":"10.1186\/s13059-018-1618-7","article-title":"An Amplicon-Based Sequencing Framework for Accurately Measuring Intrahost Virus Diversity Using PrimalSeq and IVar","volume":"20","author":"Grubaugh","year":"2019","journal-title":"Genome Biol."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"400","DOI":"10.1093\/bioinformatics\/16.4.400","article-title":"Detecting Hypermutations in Viral Sequences with an Emphasis on G \u2192 A Hypermutation","volume":"16","author":"Rose","year":"2000","journal-title":"Bioinformatics"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"8970","DOI":"10.1038\/s41598-019-45328-3","article-title":"A MiSeq-HyDRA Platform for Enhanced HIV Drug Resistance Genotyping and Surveillance","volume":"9","author":"Taylor","year":"2019","journal-title":"Sci. Rep."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"3291","DOI":"10.1038\/s41598-022-07196-2","article-title":"External Quality Assessment of HIV-1 DNA Quantification Assays Used in the Clinical Setting in Italy","volume":"12","author":"Vicenti","year":"2022","journal-title":"Sci. Rep."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"630","DOI":"10.1128\/JCM.38.2.630-634.2000","article-title":"PCR-Based Assay To Quantify Human Immunodeficiency Virus Type 1 DNA in Peripheral Blood Mononuclear Cells","volume":"38","author":"Christopherson","year":"2000","journal-title":"J. Clin. Microbiol."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"2167","DOI":"10.1128\/AAC.02710-13","article-title":"Sensitive Deep-Sequencing-Based HIV-1 Genotyping Assay To Simultaneously Determine Susceptibility to Protease, Reverse Transcriptase, Integrase, and Maturation Inhibitors, as Well as HIV-1 Coreceptor Tropism","volume":"58","author":"Gibson","year":"2014","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_32","unstructured":"U.S. Food and Drug Administration (2023, May 19). Submitting Next Generation Sequencing Data to the Division of Antiviral Products Guidance for Industry Technical Specifications Document, Available online: https:\/\/www.fda.gov\/regulatory-information\/search-fda-guidance-documents\/submitting-next-generation-sequencing-data-division-antiviral-products-guidance-industry-technical."}],"container-title":["Viruses"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1999-4915\/15\/9\/1811\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,10]],"date-time":"2025-10-10T20:39:06Z","timestamp":1760128746000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1999-4915\/15\/9\/1811"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2023,8,25]]},"references-count":32,"journal-issue":{"issue":"9","published-online":{"date-parts":[[2023,9]]}},"alternative-id":["v15091811"],"URL":"https:\/\/doi.org\/10.3390\/v15091811","relation":{},"ISSN":["1999-4915"],"issn-type":[{"type":"electronic","value":"1999-4915"}],"subject":[],"published":{"date-parts":[[2023,8,25]]}}}