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Article: Long-Read Sequencing with Hierarchical Clustering for Antiretroviral Resistance Profiling of Mixed Human Immunodeficiency Virus Quasispecies

TitleLong-Read Sequencing with Hierarchical Clustering for Antiretroviral Resistance Profiling of Mixed Human Immunodeficiency Virus Quasispecies
Authors
Issue Date4-Aug-2023
PublisherOxford University Press
Citation
Clinical Chemistry, 2023, v. 69, n. 10, p. 1174-1185 How to Cite?
AbstractBackground HIV infections often develop drug resistance mutations (DRMs), which can increase the risk of virological failure. However, it has been difficult to determine if minor mutations occur in the same genome or in different virions using Sanger sequencing and short-read sequencing methods. Oxford Nanopore Technologies (ONT) sequencing may improve antiretroviral resistance profiling by allowing for long-read clustering. Methods A new ONT sequencing-based method for profiling DRMs in HIV quasispecies was developed and validated. The method used hierarchical clustering of long amplicons that cover regions associated with different types of antiretroviral drugs. A gradient series of an HIV plasmid and 2 plasma samples was prepared to validate the clustering performance. The ONT results were compared to those obtained with Sanger sequencing and Illumina sequencing in 77 HIV-positive plasma samples to evaluate the diagnostic performance. Results In the validation study, the abundance of detected quasispecies was concordant with the predicted result with the R-2 of > 0.99. During the diagnostic evaluation, 59/77 samples were successfully sequenced for DRMs. Among 18 failed samples, 17 were below the limit of detection of 303.9 copies/& mu;L. Based on the receiver operating characteristic analysis, the ONT workflow achieved an F1 score of 0.96 with a cutoff of 0.4 variant allele frequency. Four cases were found to have quasispecies with DRMs, in which 2 harbored quasispecies with more than one class of DRMs. Treatment modifications were recommended for these cases. Conclusions Long-read sequencing coupled with hierarchical clustering could differentiate the quasispecies resistance profiles in HIV-infected samples, providing a clearer picture for medical care.
Persistent Identifierhttp://hdl.handle.net/10722/337832
ISSN
2023 Impact Factor: 7.1
2023 SCImago Journal Rankings: 1.460
ISI Accession Number ID

 

DC FieldValueLanguage
dc.contributor.authorNg, TTL-
dc.contributor.authorSu, JH-
dc.contributor.authorLao, HY-
dc.contributor.authorLui, WW-
dc.contributor.authorChan, CTM-
dc.contributor.authorLeung, AWS-
dc.contributor.authorJim, SHC-
dc.contributor.authorLee, LK-
dc.contributor.authorShehzad, S-
dc.contributor.authorTam, KKG-
dc.contributor.authorLeung, KSS-
dc.contributor.authorTang, FR-
dc.contributor.authorYam, WC-
dc.contributor.authorLuo, RB-
dc.contributor.authorSiu, GKH-
dc.date.accessioned2024-03-11T10:24:14Z-
dc.date.available2024-03-11T10:24:14Z-
dc.date.issued2023-08-04-
dc.identifier.citationClinical Chemistry, 2023, v. 69, n. 10, p. 1174-1185-
dc.identifier.issn0009-9147-
dc.identifier.urihttp://hdl.handle.net/10722/337832-
dc.description.abstractBackground HIV infections often develop drug resistance mutations (DRMs), which can increase the risk of virological failure. However, it has been difficult to determine if minor mutations occur in the same genome or in different virions using Sanger sequencing and short-read sequencing methods. Oxford Nanopore Technologies (ONT) sequencing may improve antiretroviral resistance profiling by allowing for long-read clustering. Methods A new ONT sequencing-based method for profiling DRMs in HIV quasispecies was developed and validated. The method used hierarchical clustering of long amplicons that cover regions associated with different types of antiretroviral drugs. A gradient series of an HIV plasmid and 2 plasma samples was prepared to validate the clustering performance. The ONT results were compared to those obtained with Sanger sequencing and Illumina sequencing in 77 HIV-positive plasma samples to evaluate the diagnostic performance. Results In the validation study, the abundance of detected quasispecies was concordant with the predicted result with the R-2 of > 0.99. During the diagnostic evaluation, 59/77 samples were successfully sequenced for DRMs. Among 18 failed samples, 17 were below the limit of detection of 303.9 copies/& mu;L. Based on the receiver operating characteristic analysis, the ONT workflow achieved an F1 score of 0.96 with a cutoff of 0.4 variant allele frequency. Four cases were found to have quasispecies with DRMs, in which 2 harbored quasispecies with more than one class of DRMs. Treatment modifications were recommended for these cases. Conclusions Long-read sequencing coupled with hierarchical clustering could differentiate the quasispecies resistance profiles in HIV-infected samples, providing a clearer picture for medical care.-
dc.languageeng-
dc.publisherOxford University Press-
dc.relation.ispartofClinical Chemistry-
dc.rightsThis work is licensed under a Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License.-
dc.titleLong-Read Sequencing with Hierarchical Clustering for Antiretroviral Resistance Profiling of Mixed Human Immunodeficiency Virus Quasispecies-
dc.typeArticle-
dc.identifier.doi10.1093/clinchem/hvad108-
dc.identifier.pmid37537871-
dc.identifier.scopuseid_2-s2.0-85174080653-
dc.identifier.volume69-
dc.identifier.issue10-
dc.identifier.spage1174-
dc.identifier.epage1185-
dc.identifier.eissn1530-8561-
dc.identifier.isiWOS:001041969600001-
dc.publisher.placeCARY-
dc.identifier.issnl0009-9147-

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