Multiplex Solid-Phase Melt Curve Analysis for the Point-of-Care Detection of HIV-1 Drug Resistance.
Identifieur interne : 000554 ( PubMed/Corpus ); précédent : 000553; suivant : 000555Multiplex Solid-Phase Melt Curve Analysis for the Point-of-Care Detection of HIV-1 Drug Resistance.
Auteurs : Dana S. Clutter ; Gelareh Mazarei ; Ruma Sinha ; Justen Manasa ; Janin Nouhin ; Ellen Laprade ; Sara Bolouki ; Philip L. Tzou ; Jessica Hannita-Hui ; Malaya K. Sahoo ; Peter Kuimelis ; Robert G. Kuimelis ; Benjamin A. Pinsky ; Gary K. Schoolnik ; Arjang Hassibi ; Robert W. ShaferSource :
- The Journal of molecular diagnostics : JMD [ 1943-7811 ] ; 2019.
Abstract
A point-of-care HIV-1 genotypic resistance assay that could be performed during a clinic visit would enable care providers to make informed treatment decisions for patients starting therapy or experiencing virologic failure on therapy. The main challenge for such an assay is the genetic variability at and surrounding each drug-resistance mutation (DRM). We analyzed a database of diverse global HIV sequences and used thermodynamic simulations to design an array of surface-bound oligonucleotide probe sets with each set sharing distinct 5' and 3' flanking sequences but having different centrally located nucleotides complementary to six codons at HIV-1 DRM reverse transcriptase position 103: AAA, AAC, AAG, AAT, AGA, and AGC. We then performed in vitro experiments using 80-mer oligonucleotides and PCR-amplified DNA from clinical plasma HIV-1 samples and culture supernatants that contained subtype A, B, C, D, CRF01_AE, and CRF02_AG viruses. Multiplexed solid-phase melt curve analysis discriminated perfectly among each of the six reported reverse transcriptase position 103 codons in both 80-mers and clinical samples. The sensitivity and specificity for detecting targets that contained AAC mixed with targets that contained AAA were >98% when AAC was present at a proportion of ≥10%. Multiplexed solid-phase melt curve analysis is a promising approach for developing point-of-care assays to distinguish between different codons in genetically variable regions such as those surrounding HIV-1 DRMs.
DOI: 10.1016/j.jmoldx.2019.02.005
PubMed: 31026601
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Clutter</name><affiliation><nlm:affiliation>Division of Infectious Diseases and Geographic Medicine, Stanford University School of Medicine, Stanford, California.</nlm:affiliation></affiliation></author><author><name sortKey="Mazarei, Gelareh" sort="Mazarei, Gelareh" uniqKey="Mazarei G" first="Gelareh" last="Mazarei">Gelareh Mazarei</name><affiliation><nlm:affiliation>InSilixa Inc., Sunnyvale, California.</nlm:affiliation></affiliation></author><author><name sortKey="Sinha, Ruma" sort="Sinha, Ruma" uniqKey="Sinha R" first="Ruma" last="Sinha">Ruma Sinha</name><affiliation><nlm:affiliation>InSilixa Inc., Sunnyvale, California.</nlm:affiliation></affiliation></author><author><name sortKey="Manasa, Justen" sort="Manasa, Justen" uniqKey="Manasa J" first="Justen" last="Manasa">Justen Manasa</name><affiliation><nlm:affiliation>African Institute of Biomedical Science and Technology, Harare, Zimbabwe.</nlm:affiliation></affiliation></author><author><name sortKey="Nouhin, Janin" sort="Nouhin, Janin" uniqKey="Nouhin J" first="Janin" last="Nouhin">Janin Nouhin</name><affiliation><nlm:affiliation>Division of Infectious Diseases and Geographic Medicine, Stanford University School of Medicine, Stanford, California.</nlm:affiliation></affiliation></author><author><name sortKey="Laprade, Ellen" sort="Laprade, Ellen" uniqKey="Laprade E" first="Ellen" last="Laprade">Ellen Laprade</name><affiliation><nlm:affiliation>Division of Infectious Diseases and Geographic Medicine, Stanford University School of Medicine, Stanford, California.</nlm:affiliation></affiliation></author><author><name sortKey="Bolouki, Sara" sort="Bolouki, Sara" uniqKey="Bolouki S" first="Sara" last="Bolouki">Sara Bolouki</name><affiliation><nlm:affiliation>InSilixa Inc., Sunnyvale, California.</nlm:affiliation></affiliation></author><author><name sortKey="Tzou, Philip L" sort="Tzou, Philip L" uniqKey="Tzou P" first="Philip L" last="Tzou">Philip L. 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Clutter</name><affiliation><nlm:affiliation>Division of Infectious Diseases and Geographic Medicine, Stanford University School of Medicine, Stanford, California.</nlm:affiliation></affiliation></author><author><name sortKey="Mazarei, Gelareh" sort="Mazarei, Gelareh" uniqKey="Mazarei G" first="Gelareh" last="Mazarei">Gelareh Mazarei</name><affiliation><nlm:affiliation>InSilixa Inc., Sunnyvale, California.</nlm:affiliation></affiliation></author><author><name sortKey="Sinha, Ruma" sort="Sinha, Ruma" uniqKey="Sinha R" first="Ruma" last="Sinha">Ruma Sinha</name><affiliation><nlm:affiliation>InSilixa Inc., Sunnyvale, California.</nlm:affiliation></affiliation></author><author><name sortKey="Manasa, Justen" sort="Manasa, Justen" uniqKey="Manasa J" first="Justen" last="Manasa">Justen Manasa</name><affiliation><nlm:affiliation>African Institute of Biomedical Science and Technology, Harare, Zimbabwe.</nlm:affiliation></affiliation></author><author><name sortKey="Nouhin, Janin" sort="Nouhin, Janin" uniqKey="Nouhin J" first="Janin" last="Nouhin">Janin Nouhin</name><affiliation><nlm:affiliation>Division of Infectious Diseases and Geographic Medicine, Stanford University School of Medicine, Stanford, California.</nlm:affiliation></affiliation></author><author><name sortKey="Laprade, Ellen" sort="Laprade, Ellen" uniqKey="Laprade E" first="Ellen" last="Laprade">Ellen Laprade</name><affiliation><nlm:affiliation>Division of Infectious Diseases and Geographic Medicine, Stanford University School of Medicine, Stanford, California.</nlm:affiliation></affiliation></author><author><name sortKey="Bolouki, Sara" sort="Bolouki, Sara" uniqKey="Bolouki S" first="Sara" last="Bolouki">Sara Bolouki</name><affiliation><nlm:affiliation>InSilixa Inc., Sunnyvale, California.</nlm:affiliation></affiliation></author><author><name sortKey="Tzou, Philip L" sort="Tzou, Philip L" uniqKey="Tzou P" first="Philip L" last="Tzou">Philip L. Tzou</name><affiliation><nlm:affiliation>Division of Infectious Diseases and Geographic Medicine, Stanford University School of Medicine, Stanford, California.</nlm:affiliation></affiliation></author><author><name sortKey="Hannita Hui, Jessica" sort="Hannita Hui, Jessica" uniqKey="Hannita Hui J" first="Jessica" last="Hannita-Hui">Jessica Hannita-Hui</name><affiliation><nlm:affiliation>Division of Infectious Diseases and Geographic Medicine, Stanford University School of Medicine, Stanford, California.</nlm:affiliation></affiliation></author><author><name sortKey="Sahoo, Malaya K" sort="Sahoo, Malaya K" uniqKey="Sahoo M" first="Malaya K" last="Sahoo">Malaya K. Sahoo</name><affiliation><nlm:affiliation>Division of Infectious Diseases and Geographic Medicine, Stanford University School of Medicine, Stanford, California.</nlm:affiliation></affiliation></author><author><name sortKey="Kuimelis, Peter" sort="Kuimelis, Peter" uniqKey="Kuimelis P" first="Peter" last="Kuimelis">Peter Kuimelis</name><affiliation><nlm:affiliation>InSilixa Inc., Sunnyvale, California.</nlm:affiliation></affiliation></author><author><name sortKey="Kuimelis, Robert G" sort="Kuimelis, Robert G" uniqKey="Kuimelis R" first="Robert G" last="Kuimelis">Robert G. Kuimelis</name><affiliation><nlm:affiliation>InSilixa Inc., Sunnyvale, California.</nlm:affiliation></affiliation></author><author><name sortKey="Pinsky, Benjamin A" sort="Pinsky, Benjamin A" uniqKey="Pinsky B" first="Benjamin A" last="Pinsky">Benjamin A. Pinsky</name><affiliation><nlm:affiliation>Division of Infectious Diseases and Geographic Medicine, Stanford University School of Medicine, Stanford, California; Department of Pathology, Stanford University School of Medicine, Stanford, California.</nlm:affiliation></affiliation></author><author><name sortKey="Schoolnik, Gary K" sort="Schoolnik, Gary K" uniqKey="Schoolnik G" first="Gary K" last="Schoolnik">Gary K. Schoolnik</name><affiliation><nlm:affiliation>InSilixa Inc., Sunnyvale, California.</nlm:affiliation></affiliation></author><author><name sortKey="Hassibi, Arjang" sort="Hassibi, Arjang" uniqKey="Hassibi A" first="Arjang" last="Hassibi">Arjang Hassibi</name><affiliation><nlm:affiliation>InSilixa Inc., Sunnyvale, California.</nlm:affiliation></affiliation></author><author><name sortKey="Shafer, Robert W" sort="Shafer, Robert W" uniqKey="Shafer R" first="Robert W" last="Shafer">Robert W. Shafer</name><affiliation><nlm:affiliation>Division of Infectious Diseases and Geographic Medicine, Stanford University School of Medicine, Stanford, California. Electronic address: rshafer@stanford.edu.</nlm:affiliation></affiliation></author></analytic><series><title level="j">The Journal of molecular diagnostics : JMD</title><idno type="eISSN">1943-7811</idno><imprint><date when="2019" type="published">2019</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">A point-of-care HIV-1 genotypic resistance assay that could be performed during a clinic visit would enable care providers to make informed treatment decisions for patients starting therapy or experiencing virologic failure on therapy. The main challenge for such an assay is the genetic variability at and surrounding each drug-resistance mutation (DRM). We analyzed a database of diverse global HIV sequences and used thermodynamic simulations to design an array of surface-bound oligonucleotide probe sets with each set sharing distinct 5' and 3' flanking sequences but having different centrally located nucleotides complementary to six codons at HIV-1 DRM reverse transcriptase position 103: AAA, AAC, AAG, AAT, AGA, and AGC. We then performed in vitro experiments using 80-mer oligonucleotides and PCR-amplified DNA from clinical plasma HIV-1 samples and culture supernatants that contained subtype A, B, C, D, CRF01_AE, and CRF02_AG viruses. Multiplexed solid-phase melt curve analysis discriminated perfectly among each of the six reported reverse transcriptase position 103 codons in both 80-mers and clinical samples. The sensitivity and specificity for detecting targets that contained AAC mixed with targets that contained AAA were >98% when AAC was present at a proportion of ≥10%. Multiplexed solid-phase melt curve analysis is a promising approach for developing point-of-care assays to distinguish between different codons in genetically variable regions such as those surrounding HIV-1 DRMs.</div></front></TEI><pubmed><MedlineCitation Status="In-Process" Owner="NLM"><PMID Version="1">31026601</PMID><DateRevised><Year>2020</Year><Month>02</Month><Day>26</Day></DateRevised><Article PubModel="Print-Electronic"><Journal><ISSN IssnType="Electronic">1943-7811</ISSN><JournalIssue CitedMedium="Internet"><Volume>21</Volume><Issue>4</Issue><PubDate><Year>2019</Year><Month>07</Month></PubDate></JournalIssue><Title>The Journal of molecular diagnostics : JMD</Title><ISOAbbreviation>J Mol Diagn</ISOAbbreviation></Journal><ArticleTitle>Multiplex Solid-Phase Melt Curve Analysis for the Point-of-Care Detection of HIV-1 Drug Resistance.</ArticleTitle><Pagination><MedlinePgn>580-592</MedlinePgn></Pagination><ELocationID EIdType="pii" ValidYN="Y">S1525-1578(18)30274-5</ELocationID><ELocationID EIdType="doi" ValidYN="Y">10.1016/j.jmoldx.2019.02.005</ELocationID><Abstract><AbstractText>A point-of-care HIV-1 genotypic resistance assay that could be performed during a clinic visit would enable care providers to make informed treatment decisions for patients starting therapy or experiencing virologic failure on therapy. The main challenge for such an assay is the genetic variability at and surrounding each drug-resistance mutation (DRM). We analyzed a database of diverse global HIV sequences and used thermodynamic simulations to design an array of surface-bound oligonucleotide probe sets with each set sharing distinct 5' and 3' flanking sequences but having different centrally located nucleotides complementary to six codons at HIV-1 DRM reverse transcriptase position 103: AAA, AAC, AAG, AAT, AGA, and AGC. We then performed in vitro experiments using 80-mer oligonucleotides and PCR-amplified DNA from clinical plasma HIV-1 samples and culture supernatants that contained subtype A, B, C, D, CRF01_AE, and CRF02_AG viruses. Multiplexed solid-phase melt curve analysis discriminated perfectly among each of the six reported reverse transcriptase position 103 codons in both 80-mers and clinical samples. The sensitivity and specificity for detecting targets that contained AAC mixed with targets that contained AAA were >98% when AAC was present at a proportion of ≥10%. Multiplexed solid-phase melt curve analysis is a promising approach for developing point-of-care assays to distinguish between different codons in genetically variable regions such as those surrounding HIV-1 DRMs.</AbstractText><CopyrightInformation>Copyright © 2019 American Society for Investigative Pathology and the Association for Molecular Pathology. Published by Elsevier Inc. All rights reserved.</CopyrightInformation></Abstract><AuthorList CompleteYN="Y"><Author ValidYN="Y"><LastName>Clutter</LastName><ForeName>Dana S</ForeName><Initials>DS</Initials><AffiliationInfo><Affiliation>Division of Infectious Diseases and Geographic Medicine, Stanford University School of Medicine, Stanford, California.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Mazarei</LastName><ForeName>Gelareh</ForeName><Initials>G</Initials><AffiliationInfo><Affiliation>InSilixa Inc., Sunnyvale, California.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Sinha</LastName><ForeName>Ruma</ForeName><Initials>R</Initials><AffiliationInfo><Affiliation>InSilixa Inc., Sunnyvale, California.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Manasa</LastName><ForeName>Justen</ForeName><Initials>J</Initials><AffiliationInfo><Affiliation>African Institute of Biomedical Science and Technology, Harare, Zimbabwe.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Nouhin</LastName><ForeName>Janin</ForeName><Initials>J</Initials><AffiliationInfo><Affiliation>Division of Infectious Diseases and Geographic Medicine, Stanford University School of Medicine, Stanford, California.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>LaPrade</LastName><ForeName>Ellen</ForeName><Initials>E</Initials><AffiliationInfo><Affiliation>Division of Infectious Diseases and Geographic Medicine, Stanford University School of Medicine, Stanford, California.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Bolouki</LastName><ForeName>Sara</ForeName><Initials>S</Initials><AffiliationInfo><Affiliation>InSilixa Inc., Sunnyvale, California.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Tzou</LastName><ForeName>Philip L</ForeName><Initials>PL</Initials><AffiliationInfo><Affiliation>Division of Infectious Diseases and Geographic Medicine, Stanford University School of Medicine, Stanford, California.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Hannita-Hui</LastName><ForeName>Jessica</ForeName><Initials>J</Initials><AffiliationInfo><Affiliation>Division of Infectious Diseases and Geographic Medicine, Stanford University School of Medicine, Stanford, California.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Sahoo</LastName><ForeName>Malaya K</ForeName><Initials>MK</Initials><AffiliationInfo><Affiliation>Division of Infectious Diseases and Geographic Medicine, Stanford University School of Medicine, Stanford, California.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Kuimelis</LastName><ForeName>Peter</ForeName><Initials>P</Initials><AffiliationInfo><Affiliation>InSilixa Inc., Sunnyvale, California.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Kuimelis</LastName><ForeName>Robert G</ForeName><Initials>RG</Initials><AffiliationInfo><Affiliation>InSilixa Inc., Sunnyvale, California.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Pinsky</LastName><ForeName>Benjamin A</ForeName><Initials>BA</Initials><AffiliationInfo><Affiliation>Division of Infectious Diseases and Geographic Medicine, Stanford University School of Medicine, Stanford, California; Department of Pathology, Stanford University School of Medicine, Stanford, California.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Schoolnik</LastName><ForeName>Gary K</ForeName><Initials>GK</Initials><AffiliationInfo><Affiliation>InSilixa Inc., Sunnyvale, California.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Hassibi</LastName><ForeName>Arjang</ForeName><Initials>A</Initials><AffiliationInfo><Affiliation>InSilixa Inc., Sunnyvale, California.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Shafer</LastName><ForeName>Robert W</ForeName><Initials>RW</Initials><AffiliationInfo><Affiliation>Division of Infectious Diseases and Geographic Medicine, Stanford University School of Medicine, Stanford, California. Electronic address: rshafer@stanford.edu.</Affiliation></AffiliationInfo></Author></AuthorList><Language>eng</Language><GrantList CompleteYN="Y"><Grant><GrantID>R21 AI131918</GrantID><Acronym>AI</Acronym><Agency>NIAID NIH HHS</Agency><Country>United States</Country></Grant></GrantList><PublicationTypeList><PublicationType UI="D016428">Journal Article</PublicationType><PublicationType UI="D013485">Research Support, Non-U.S. Gov't</PublicationType><PublicationType UI="D052061">Research Support, N.I.H., Extramural</PublicationType></PublicationTypeList><ArticleDate DateType="Electronic"><Year>2019</Year><Month>04</Month><Day>23</Day></ArticleDate></Article><MedlineJournalInfo><Country>United States</Country><MedlineTA>J Mol Diagn</MedlineTA><NlmUniqueID>100893612</NlmUniqueID><ISSNLinking>1525-1578</ISSNLinking></MedlineJournalInfo><CitationSubset>IM</CitationSubset></MedlineCitation><PubmedData><History><PubMedPubDate PubStatus="received"><Year>2018</Year><Month>06</Month><Day>20</Day></PubMedPubDate><PubMedPubDate PubStatus="revised"><Year>2019</Year><Month>02</Month><Day>05</Day></PubMedPubDate><PubMedPubDate PubStatus="accepted"><Year>2019</Year><Month>02</Month><Day>19</Day></PubMedPubDate><PubMedPubDate PubStatus="pmc-release"><Year>2020</Year><Month>07</Month><Day>01</Day></PubMedPubDate><PubMedPubDate PubStatus="pubmed"><Year>2019</Year><Month>4</Month><Day>27</Day><Hour>6</Hour><Minute>0</Minute></PubMedPubDate><PubMedPubDate PubStatus="medline"><Year>2019</Year><Month>4</Month><Day>27</Day><Hour>6</Hour><Minute>0</Minute></PubMedPubDate><PubMedPubDate PubStatus="entrez"><Year>2019</Year><Month>4</Month><Day>27</Day><Hour>6</Hour><Minute>0</Minute></PubMedPubDate></History><PublicationStatus>ppublish</PublicationStatus><ArticleIdList><ArticleId IdType="pubmed">31026601</ArticleId><ArticleId IdType="pii">S1525-1578(18)30274-5</ArticleId><ArticleId IdType="doi">10.1016/j.jmoldx.2019.02.005</ArticleId><ArticleId IdType="pmc">PMC6643098</ArticleId></ArticleIdList><ReferenceList><Reference><Citation>Nat Methods. 2004 Nov;1(2):141-7</Citation><ArticleIdList><ArticleId IdType="pubmed">15782177</ArticleId></ArticleIdList></Reference><Reference><Citation>Bioinformatics. 2005 Oct 15;21(20):3940-1</Citation><ArticleIdList><ArticleId IdType="pubmed">16096348</ArticleId></ArticleIdList></Reference><Reference><Citation>J Mol Diagn. 2006 Sep;8(4):430-2; 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