{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,8,29]],"date-time":"2025-08-29T09:43:40Z","timestamp":1756460620664,"version":"3.41.2"},"reference-count":18,"publisher":"Wiley","issue":"7","license":[{"start":{"date-parts":[[2021,7,8]],"date-time":"2021-07-08T00:00:00Z","timestamp":1625702400000},"content-version":"vor","delay-in-days":7,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/100000060","name":"National Institute of Allergy and Infectious Diseases","doi-asserted-by":"publisher","award":["R01AI108538"],"award-info":[{"award-number":["R01AI108538"]}],"id":[{"id":"10.13039\/100000060","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100000026","name":"National Institute on Drug Abuse","doi-asserted-by":"publisher","award":["R61DA047038"],"award-info":[{"award-number":["R61DA047038"]}],"id":[{"id":"10.13039\/100000026","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["onlinelibrary.wiley.com"],"crossmark-restriction":true},"short-container-title":["J Int AIDS Soc"],"published-print":{"date-parts":[[2021,7]]},"abstract":"<jats:title>Abstract<\/jats:title><jats:sec><jats:title>Introduction<\/jats:title><jats:p>HIV rebounds after cessation of antiretroviral therapy, representing a barrier to cure. To better understand the virus reservoir, analysis pipelines have been developed that categorize proviral sequences as intact or defective, and further determine the precise nature of the sequence defects that may be present. We investigated the effects that different analysis pipelines had on the characterization of HIV\u20101 proviral sequences.<\/jats:p><\/jats:sec><jats:sec><jats:title>Methods<\/jats:title><jats:p>We used single genome amplification to generate near full\u2010length (NFL) HIV\u20101 proviral DNA sequences, defined as amplicons greater than 8000 base pairs in length, isolated from peripheral blood mononuclear cells (PBMC) of treated suppressed participants with HIV\u20101. Amplicons underwent direct next\u2010generation single genome sequencing and were analysed using four HIV\u20101 proviral characterization pipelines. Sequences were characterized as intact or defective; defective sequences were assessed for the number and types of defects present. To confirm and extend our findings, 691 proviruses from the Proviral Sequence Database (PSD) were analysed and the ProSeq\u2010IT tool of the PSD was used to characterize both the participant and PSD proviruses.<\/jats:p><\/jats:sec><jats:sec><jats:title>Results and discussion<\/jats:title><jats:p>Virus sequences derived from thirteen ART\u2010treated virologically suppressed participants with HIV were studied. A total of 693 HIV\u20101 proviral sequences were generated, 282 of which were NFL. An average of 53 sequences per participant was analysed. We found that proviruses often harbour multiple sequence defect types (mean 2.7, 95% confidence interval [CI] 2.5, 3.0); the elimination order used by each pipeline affected the percentage of proviruses allotted into each defect category. These differences varied between participants, depending on the number of defect categories present in a given provirus sequence. Pipeline\u2010specific differences in characterizing the HIV\u20101 5\u2032 untranslated region (5\u2032 UTR) led to an overestimation of the number of intact NFL proviral sequences, a finding corroborated in the independent PSD analysis. A comparison of the four published pipelines to ProSeq\u2010IT found that ProSeq IT was more likely to characterize proviruses as intact.<\/jats:p><\/jats:sec><jats:sec><jats:title>Conclusions<\/jats:title><jats:p>The choice of pipeline used for HIV\u20101 provirus landscape analysis may bias the classification of defective sequences. To improve the comparison of provirus characterizations across research groups, the development of a consensus elimination pipeline should be prioritized.<\/jats:p><\/jats:sec>","DOI":"10.1002\/jia2.25725","type":"journal-article","created":{"date-parts":[[2021,7,8]],"date-time":"2021-07-08T07:23:34Z","timestamp":1625729014000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":3,"title":["HIV\u20101 proviral landscape characterization varies by pipeline analysis"],"prefix":"10.1002","volume":"24","author":[{"given":"Fernanda A","family":"Ferreira","sequence":"first","affiliation":[{"name":"Virology Program Graduate School of Arts and Sciences Harvard University Cambridge MA USA"},{"name":"Division of Infectious Diseases Brigham and Woman\u2019s Hospital Boston MA USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Qianjing","family":"He","sequence":"additional","affiliation":[{"name":"Division of Infectious Diseases Brigham and Woman\u2019s Hospital Boston MA USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Stephanie","family":"Banning","sequence":"additional","affiliation":[{"name":"Division of Infectious Diseases Brigham and Woman\u2019s Hospital Boston MA USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Olivia","family":"Roberts\u2010Sano","sequence":"additional","affiliation":[{"name":"Division of Infectious Diseases Brigham and Woman\u2019s Hospital Boston MA USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Olivia","family":"Wilkins","sequence":"additional","affiliation":[{"name":"Division of Infectious Diseases Brigham and Woman\u2019s Hospital Boston MA USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Daniel R.","family":"Kuritzkes","sequence":"additional","affiliation":[{"name":"Division of Infectious Diseases Brigham and Woman\u2019s Hospital Boston MA USA"},{"name":"Harvard Medical School Boston MA USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2681-3216","authenticated-orcid":false,"given":"Athe","family":"Tsibris","sequence":"additional","affiliation":[{"name":"Division of Infectious Diseases Brigham and Woman\u2019s Hospital Boston MA USA"},{"name":"Harvard Medical School Boston MA USA"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"311","published-online":{"date-parts":[[2021,7,8]]},"reference":[{"doi-asserted-by":"publisher","key":"e_1_2_9_2_1","DOI":"10.1038\/387183a0"},{"doi-asserted-by":"publisher","key":"e_1_2_9_3_1","DOI":"10.1126\/science.278.5341.1295"},{"doi-asserted-by":"publisher","key":"e_1_2_9_4_1","DOI":"10.1126\/science.278.5341.1291"},{"doi-asserted-by":"publisher","key":"e_1_2_9_5_1","DOI":"10.1016\/j.cell.2013.09.020"},{"key":"e_1_2_9_6_1","first-page":"1","article-title":"Amplification of near full\u2010length HIV\u20101 proviruses for next\u2010generation sequencing","volume":"140","author":"Hiener B","year":"2018","journal-title":"J Vis Exp"},{"doi-asserted-by":"publisher","key":"e_1_2_9_7_1","DOI":"10.1038\/nm.4156"},{"doi-asserted-by":"publisher","key":"e_1_2_9_8_1","DOI":"10.1172\/JCI93289"},{"doi-asserted-by":"publisher","key":"e_1_2_9_9_1","DOI":"10.1073\/pnas.1609057113"},{"doi-asserted-by":"publisher","key":"e_1_2_9_10_1","DOI":"10.1128\/JVI.00915-10"},{"doi-asserted-by":"publisher","key":"e_1_2_9_11_1","DOI":"10.1126\/science.1083338"},{"doi-asserted-by":"publisher","key":"e_1_2_9_12_1","DOI":"10.1016\/j.chom.2017.03.008"},{"doi-asserted-by":"publisher","key":"e_1_2_9_13_1","DOI":"10.1128\/JVI.01231-06"},{"unstructured":"Los Alamos National Laboratory.HIV Sequence Database \u2013 Hypermut tool[cited 2020 May 1]. 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