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Detection of drug resistance variants at minor frequencies can help to formulate successful antiretroviral therapy (ART) regimens for people living with HIV (PLWH) and reduce the emergence of drug resistance. Therefore, a pipeline which can accurately produce consensus nucleotide sequences and identify drug resistance mutations (DRMs) at defined frequency thresholds will be helpful in the treatment of PLWH, analysis of virus evolution, and the control of the pandemic.<\/jats:p>\n          <\/jats:sec>\n          <jats:sec>\n            <jats:title>Results<\/jats:title>\n            <jats:p>We have developed a pipeline, HIVGenoPipe, to determine HIV drug resistance variants within the <jats:italic>gag-pol<\/jats:italic> region above user-defined frequencies for HIV-1 samples sequenced using Illumina technology. The pipeline has been validated by comparing its results with the results generated by a widely used pipeline, HyDRA, which is limited to the <jats:italic>pol<\/jats:italic> region, and with the results generated by Sanger sequencing technology using the same set of 30 samples. The variant frequency used to generate ambiguous consensus sequences in HIVGenoPipe is more accurate than other pipelines because a sample-specific reference, which is generated in real-time with a novel hybrid strategy of de novo and reference-based assembly, is used for the frequency calculation, leading to more accurate drug resistance calls for use by clinicians. In addition, since Nextflow is used as the pipeline platform, HIVGenoPipe inherently has great portability, scalability and reproducibility; and the components can be updated or replaced independently if required.<\/jats:p>\n          <\/jats:sec>\n          <jats:sec>\n            <jats:title>Conclusions<\/jats:title>\n            <jats:p>We developed HIVGenoPipe for the detection of HIV-1 drug resistance. It constructs more accurate gag-pol consensus sequences, leading to improved detection of DRMs. HIVGenoPipe is open source and freely available under the MIT license at <jats:ext-link xmlns:xlink=\"http:\/\/www.w3.org\/1999\/xlink\" xlink:href=\"https:\/\/github.com\/LHRI-Bioinformatics\/HIVGenoPipe\" ext-link-type=\"uri\">https:\/\/github.com\/LHRI-Bioinformatics\/HIVGenoPipe<\/jats:ext-link>. The current release (v1.0.1) is archived and available at <jats:ext-link xmlns:xlink=\"http:\/\/www.w3.org\/1999\/xlink\" xlink:href=\"https:\/\/doi.org\/\" ext-link-type=\"uri\">https:\/\/doi.org\/<\/jats:ext-link>\n              <jats:ext-link xmlns:xlink=\"http:\/\/www.w3.org\/1999\/xlink\" xlink:href=\"10.5281\/zenodo.15528502\" ext-link-type=\"doi\">https:\/\/doi.org\/10.5281\/zenodo.15528502<\/jats:ext-link>.<\/jats:p>\n          <\/jats:sec>","DOI":"10.1186\/s12859-025-06201-5","type":"journal-article","created":{"date-parts":[[2025,7,7]],"date-time":"2025-07-07T12:50:49Z","timestamp":1751892649000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["HIVGenoPipe: a nextflow pipeline for the detection of HIV-1 drug resistance using a real-time sample-specific reference sequence"],"prefix":"10.1186","volume":"26","author":[{"given":"Thoai","family":"Dotrang","sequence":"first","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Brad T.","family":"Sherman","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Lisheng","family":"Dai","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Muhammad Ayub","family":"Khan","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Helene C.","family":"Highbarger","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Whitney","family":"Bruchey","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Sylvain","family":"Laverdure","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Michael W.","family":"Baseler","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Tomozumi","family":"Imamichi","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Robin L.","family":"Dewar","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Weizhong","family":"Chang","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"297","published-online":{"date-parts":[[2025,7,7]]},"reference":[{"issue":"4599","key":"6201_CR1","doi-asserted-by":"publisher","first-page":"868","DOI":"10.1126\/science.6189183","volume":"220","author":"F Barr\u00e9-Sinoussi","year":"1983","unstructured":"Barr\u00e9-Sinoussi F, Chermann JC, Rey F, Nugeyre MT, Chamaret S, Gruest J, Dauguet C, Axler-Blin C, V\u00e9zinet-Brun F, Rouzioux C, et al. 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Viruses. 2020;12(8):1045.","journal-title":"Viruses"}],"container-title":["BMC Bioinformatics"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/link.springer.com\/content\/pdf\/10.1186\/s12859-025-06201-5.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/link.springer.com\/article\/10.1186\/s12859-025-06201-5\/fulltext.html","content-type":"text\/html","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/link.springer.com\/content\/pdf\/10.1186\/s12859-025-06201-5.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,7,7]],"date-time":"2025-07-07T14:03:26Z","timestamp":1751897006000},"score":1,"resource":{"primary":{"URL":"https:\/\/bmcbioinformatics.biomedcentral.com\/articles\/10.1186\/s12859-025-06201-5"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,7,7]]},"references-count":46,"journal-issue":{"issue":"1","published-online":{"date-parts":[[2025,12]]}},"alternative-id":["6201"],"URL":"https:\/\/doi.org\/10.1186\/s12859-025-06201-5","relation":{},"ISSN":["1471-2105"],"issn-type":[{"type":"electronic","value":"1471-2105"}],"subject":[],"published":{"date-parts":[[2025,7,7]]},"assertion":[{"value":"18 February 2025","order":1,"name":"received","label":"Received","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"23 June 2025","order":2,"name":"accepted","label":"Accepted","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"7 July 2025","order":3,"name":"first_online","label":"First Online","group":{"name":"ArticleHistory","label":"Article History"}},{"order":1,"name":"Ethics","group":{"name":"EthicsHeading","label":"Declarations"}},{"value":"All experimental procedures in these studies were approved by the National Cancer Institute at Frederick and the National Institute of Allergy and Infectious Diseases (NIAID) Institutional Review Boards and performed in accordance with the relevant guidelines and regulations. Samples were obtained from 30 participants who were enrolled in following institutional review board-approved NIH studies: Clinical and Immunologic Monitoring of Patients With Known or Suspected HIV Infection (ClinicalTrials registration number NCT00789009), A Study to Evaluate the Safety and Effects of Repeated Doses of 3BNC117-LS and 10-1074-LS on Persistent Viral Reservoirs in People Living With HIV and on Suppressive Antiretroviral Therapy (ClinicalTrials registration number NCT05612178), A Trial of Anti-CD4 Antibody UB-421 in Combination With Optimized Background Antiretroviral Therapy in Patients With Multi-Drug Resistant HIV-1 Infection (ClinicalTrials registration number NCT05582694) and Tissue Biopsy and Imaging Studies in HIV-Infected Patients (ClinicalTrials registration number NCT00001471). The participants provided written informed consent.","order":2,"name":"Ethics","group":{"name":"EthicsHeading","label":"Ethics approval and consent to participate"}},{"value":"Not applicable.","order":3,"name":"Ethics","group":{"name":"EthicsHeading","label":"Consent for publication"}},{"value":"The authors declare no competing interests.","order":4,"name":"Ethics","group":{"name":"EthicsHeading","label":"Competing interests"}}],"article-number":"168"}}