{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,8,24]],"date-time":"2026-08-24T23:24:09Z","timestamp":1787613849773,"version":"build-2736575974"},"reference-count":26,"publisher":"Oxford University Press (OUP)","issue":"5","license":[{"start":{"date-parts":[[2025,12,16]],"date-time":"2025-12-16T00:00:00Z","timestamp":1765843200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/academic.oup.com\/pages\/standard-publication-reuse-rights"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"published-print":{"date-parts":[[2026,5,15]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                  <jats:sec>\n                    <jats:title>Background<\/jats:title>\n                    <jats:p>Hepatitis C virus (HCV) diagnosis usually requires detection of antibody followed by HCV RNA. The requirement for 2 tests leads to major drop-offs in the cascade of care. Existing near-care HCV RNA tests have slow turnaround time and are expensive with limited availability. We aim to develop a cost-effective, rapid, and sensitive test for detection of HCV RNA to enhance screening, particularly in marginalized and remote populations.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Methods<\/jats:title>\n                    <jats:p>After RNA extraction from plasma, HCV RNA is reverse-transcribed and amplified using loop-mediated isothermal amplification with HCV-specific primers. The amplified HCV DNA is then detected via CRISPR-Cas12b with a fluorescence readout.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Results<\/jats:title>\n                    <jats:p>HCV RNA from patient samples with genotypes 1a, 1b, 2, 3a, and 4 was detected with high sensitivity and specificity. The lower limit of detection (LLOD) with HCV JFH1 plasmid (genotype 2) is 250 plasmid copies\/mL (approximately 100\u2005IU\/mL). For clinical samples, we determined the LLOD for genotypes 1 and 3, the most common in North America. Using 500\u2005\u03bcL of plasma, genotype 1 RNA \u2265100\u2005IU\/mL was detected within 40\u201345\u2005minutes, while genotype 3 had an LLOD of 5000\u2005IU\/mL. The clinical sensitivity was 100% in 72 HCV patient samples, including acute HCV and HCV\/hepatitis B virus (HBV) coinfection. The specificity was 100%, with no false-positives in 33 HCV-negative samples, including those with HBV or human immunodeficiency virus\/HBV coinfection.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Conclusions<\/jats:title>\n                    <jats:p>Our assay shows high specificity and sensitivity to detect HCV RNA directly from plasma within 45\u2005minutes and hence could be used for efficient screening and diagnosis of HCV infection globally.<\/jats:p>\n                  <\/jats:sec>","DOI":"10.1093\/infdis\/jiaf609","type":"journal-article","created":{"date-parts":[[2025,12,3]],"date-time":"2025-12-03T13:09:00Z","timestamp":1764767340000},"page":"840-847","source":"Crossref","is-referenced-by-count":1,"title":["A Rapid Assay for Hepatitis C Virus RNA Detection Using Reverse-Transcription Loop-Mediated Isothermal Amplification\u2013Coupled CRISPR-Cas12b\u2013Based Strategy"],"prefix":"10.1093","volume":"233","author":[{"ORCID":"https:\/\/orcid.org\/0000-0001-5301-2185","authenticated-orcid":false,"given":"Avisha","family":"Chowdhury","sequence":"first","affiliation":[{"name":"Toronto Centre for Liver Disease, Toronto General Hospital, University Health Network, University of Toronto , Toronto, Ontario ,","place":["Canada"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-1081-5452","authenticated-orcid":false,"given":"Bianca Garcia","family":"Garcia","sequence":"additional","affiliation":[{"name":"Department of Molecular Genetics, University of Toronto , Toronto, Ontario ,","place":["Canada"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2302-3689","authenticated-orcid":false,"given":"Muhammad Atif","family":"Zahoor","sequence":"additional","affiliation":[{"name":"Toronto Centre for Liver Disease, Toronto General Hospital, University Health Network, University of Toronto , Toronto, Ontario ,","place":["Canada"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0000-6201-1512","authenticated-orcid":false,"given":"Nahla Fadl","family":"ElMawla","sequence":"additional","affiliation":[{"name":"Toronto Centre for Liver Disease, Toronto General Hospital, University Health Network, University of Toronto , Toronto, Ontario ,","place":["Canada"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0744-9199","authenticated-orcid":false,"given":"Alan R","family":"Davidson","sequence":"additional","affiliation":[{"name":"Department of Molecular Genetics, University of Toronto , Toronto, Ontario ,","place":["Canada"]},{"name":"Department of Biochemistry, University of Toronto , Toronto, Ontario ,","place":["Canada"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9995-1057","authenticated-orcid":false,"given":"Haley D M","family":"Wyatt","sequence":"additional","affiliation":[{"name":"Department of Biochemistry, University of Toronto , Toronto, Ontario ,","place":["Canada"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0211-5480","authenticated-orcid":false,"given":"Karen L","family":"Maxwell","sequence":"additional","affiliation":[{"name":"Department of Biochemistry, University of Toronto , Toronto, Ontario ,","place":["Canada"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0006-0345-5053","authenticated-orcid":false,"given":"Ayoub","family":"Mahassine","sequence":"additional","affiliation":[{"name":"Toronto Centre for Liver Disease, Toronto General Hospital, University Health Network, University of Toronto , Toronto, Ontario ,","place":["Canada"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1150-5840","authenticated-orcid":false,"given":"Adam","family":"Gehring","sequence":"additional","affiliation":[{"name":"Toronto Centre for Liver Disease, Toronto General Hospital, University Health Network, University of Toronto , Toronto, Ontario ,","place":["Canada"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2640-2211","authenticated-orcid":false,"given":"Jordan J","family":"Feld","sequence":"additional","affiliation":[{"name":"Toronto Centre for Liver Disease, Toronto General Hospital, University Health Network, University of Toronto , Toronto, Ontario ,","place":["Canada"]}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"286","published-online":{"date-parts":[[2025,12,16]]},"reference":[{"key":"2026051419575118100_jiaf609-B1","author":"World Health Organization"},{"key":"2026051419575118100_jiaf609-B2","doi-asserted-by":"crossref","first-page":"265","DOI":"10.1128\/JCM.02407-15","article-title":"Role of serologic and molecular diagnostic assays in identification and management of hepatitis C virus infection","volume":"54","author":"Cloherty","year":"2016","journal-title":"J Clin Microbiol"},{"key":"2026051419575118100_jiaf609-B3","doi-asserted-by":"crossref","first-page":"25","DOI":"10.5501\/wjv.v4.i1.25","article-title":"Current molecular methods for the detection of hepatitis C virus in high risk group population: a systematic review","volume":"4","author":"Firdaus","year":"2015","journal-title":"World J Virol"},{"key":"2026051419575118100_jiaf609-B4","doi-asserted-by":"crossref","first-page":"440","DOI":"10.1016\/j.ijid.2020.10.082","article-title":"Rapid visual detection of hepatitis C virus using a reverse transcription loop-mediated isothermal amplification assay","volume":"102","author":"Hongjaisee","year":"2021","journal-title":"Int J Infect Dis"},{"key":"2026051419575118100_jiaf609-B5","first-page":"100490","article-title":"Rapid hepatitis C virus point-of-care RNA testing and treatment at an integrated supervised consumption service in Toronto, Canada: a prospective, observational cohort study","volume":"22","author":"Lettner","year":"2023","journal-title":"Lancet Reg Health Am"},{"key":"2026051419575118100_jiaf609-B6","doi-asserted-by":"crossref","first-page":"171","DOI":"10.3138\/canlivj.2019-0007","article-title":"Hepatitis C virus (HCV) care in Canadian correctional facilities: where are we and where do we need to be?","volume":"2","author":"Kronfli","year":"2019","journal-title":"Can Liver J"},{"key":"2026051419575118100_jiaf609-B7","doi-asserted-by":"crossref","first-page":"2986","DOI":"10.1038\/s41596-019-0210-2","article-title":"SHERLOCK: nucleic acid detection with CRISPR nucleases","volume":"14","author":"Kellner","year":"2019","journal-title":"Nat Protoc"},{"key":"2026051419575118100_jiaf609-B8","doi-asserted-by":"crossref","first-page":"870","DOI":"10.1038\/s41587-020-0513-4","article-title":"CRISPR\u2013Cas12-based detection of SARS-CoV-2","volume":"38","author":"Broughton","year":"2020","journal-title":"Nat Biotechnol"},{"key":"2026051419575118100_jiaf609-B9","doi-asserted-by":"crossref","first-page":"2228","DOI":"10.1021\/acssynbio.9b00209","article-title":"HOLMESv2: a CRISPR-Cas12b-assisted platform for nucleic acid detection and DNA methylation quantitation","volume":"10","author":"Li","year":"2019","journal-title":"ACS Synth Biol"},{"key":"2026051419575118100_jiaf609-B10","doi-asserted-by":"crossref","first-page":"20","DOI":"10.1038\/s41421-018-0028-z","article-title":"CRISPR-Cas12a-assisted nucleic acid detection","volume":"4","author":"Li","year":"2018","journal-title":"Cell Discov"},{"key":"2026051419575118100_jiaf609-B11","doi-asserted-by":"crossref","first-page":"103926","DOI":"10.1016\/j.ebiom.2022.103926","article-title":"A thermostable Cas12b from Brevibacillus leverages one-pot discrimination of SARS-CoV-2 variants of concern","volume":"77","author":"Nguyen","year":"2022","journal-title":"EBioMedicine"},{"key":"2026051419575118100_jiaf609-B12","doi-asserted-by":"crossref","first-page":"63e","DOI":"10.1093\/nar\/28.12.e63","article-title":"Loop-mediated isothermal amplification of DNA","volume":"28","author":"Notomi","year":"2000","journal-title":"Nucleic Acids Res"},{"key":"2026051419575118100_jiaf609-B13","doi-asserted-by":"crossref","first-page":"e204","DOI":"10.1371\/journal.pbio.0040204","article-title":"DNA detection using recombination proteins","volume":"4","author":"Piepenburg","year":"2006","journal-title":"PLoS Biol"},{"key":"2026051419575118100_jiaf609-B14","doi-asserted-by":"crossref","first-page":"340938","DOI":"10.1016\/j.aca.2023.340938","article-title":"Fast and visual detection of nucleic acids using a one-step RPA-CRISPR detection (ORCD) system unrestricted by the PAM","volume":"1248","author":"Lin","year":"2023","journal-title":"Anal Chim Acta"},{"key":"2026051419575118100_jiaf609-B15","doi-asserted-by":"crossref","first-page":"62","DOI":"10.1016\/j.jhep.2019.02.011","article-title":"Deriving the optimal limit of detection for an HCV point-of-care test for viraemic infection: analysis of a global dataset","volume":"71","author":"Freiman","year":"2019","journal-title":"J Hepatol"},{"key":"2026051419575118100_jiaf609-B16","doi-asserted-by":"crossref","first-page":"376","DOI":"10.1016\/j.ijbiomac.2020.01.079","article-title":"A novel thermal Cas12b from a hot spring bacterium with high target mismatch tolerance and robust DNA cleavage efficiency","volume":"147","author":"Tian","year":"2020","journal-title":"Int J Biol Macromol"},{"key":"2026051419575118100_jiaf609-B17","doi-asserted-by":"crossref","first-page":"829","DOI":"10.1016\/j.jhep.2021.04.056","article-title":"Sofosbuvir\/velpatasvir for 12 vs. 6 weeks for the treatment of recently acquired hepatitis C infection","volume":"75","author":"Matthews","year":"2021","journal-title":"J Hepatol"},{"key":"2026051419575118100_jiaf609-B18","doi-asserted-by":"crossref","first-page":"18","DOI":"10.1007\/s13337-012-0067-2","article-title":"Loop-mediated isothermal amplification assay for rapid detection of hepatitis C virus","volume":"23","author":"Kargar","year":"2012","journal-title":"Indian J Virol Off Organ Indian Virol Soc"},{"key":"2026051419575118100_jiaf609-B19","doi-asserted-by":"crossref","first-page":"1524","DOI":"10.3390\/diagnostics12071524","article-title":"Highly specific and rapid detection of hepatitis C virus using RT-LAMP-coupled CRISPR\u2013Cas12 assay","volume":"12","author":"Kham-Kjing","year":"2022","journal-title":"Diagnostics"},{"key":"2026051419575118100_jiaf609-B20","doi-asserted-by":"crossref","first-page":"e0276582","DOI":"10.1371\/journal.pone.0276582","article-title":"Rapid detection of hepatitis C virus using recombinase polymerase amplification","volume":"17","author":"Chia","year":"2022","journal-title":"PLoS One"},{"key":"2026051419575118100_jiaf609-B21","doi-asserted-by":"crossref","first-page":"134278","DOI":"10.1016\/j.snb.2023.134278","article-title":"Rapid and sensitive LAMP\/CRISPR-powered diagnostics to detect different hepatitis C virus genotypes using an ITO-based EG-FET biosensing platform","volume":"394","author":"Ho","year":"2023","journal-title":"Sens Actuators B Chem"},{"key":"2026051419575118100_jiaf609-B22","doi-asserted-by":"crossref","first-page":"103926","DOI":"10.1016\/j.ebiom.2022.103926","article-title":"A thermostable Cas12b from Brevibacillus leverages one-pot discrimination of SARS-CoV-2 variants of concern","volume":"77","author":"Nguyen","year":"2022","journal-title":"eBioMedicine"},{"key":"2026051419575118100_jiaf609-B23","doi-asserted-by":"crossref","first-page":"113493","DOI":"10.1016\/j.bios.2021.113493","article-title":"A chemical-enhanced system for CRISPR-based nucleic acid detection","volume":"192","author":"Li","year":"2021","journal-title":"Biosens Bioelectron"},{"key":"2026051419575118100_jiaf609-B24","doi-asserted-by":"crossref","first-page":"198129","DOI":"10.1016\/j.virusres.2020.198129","article-title":"iSCAN: an RT-LAMP-coupled CRISPR-Cas12 module for rapid, sensitive detection of SARS-CoV-2","volume":"288","author":"Ali","year":"2020","journal-title":"Virus Res"},{"key":"2026051419575118100_jiaf609-B25","doi-asserted-by":"crossref","first-page":"994","DOI":"10.1111\/j.1537-2995.2005.04390.x","article-title":"Dynamics of viremia in early hepatitis C virus infection","volume":"45","author":"Glynn","year":"2005","journal-title":"Transfusion (Paris)"},{"key":"2026051419575118100_jiaf609-B26","doi-asserted-by":"crossref","first-page":"959","DOI":"10.1086\/428583","article-title":"Acute-phase hepatitis C virus infection: implications for research, diagnosis, and treatment","volume":"40","author":"Busch","year":"2005","journal-title":"Clin Infect Dis"}],"container-title":["The Journal of Infectious Diseases"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/academic.oup.com\/jid\/advance-article-pdf\/doi\/10.1093\/infdis\/jiaf609\/65900923\/jiaf609.pdf","content-type":"application\/pdf","content-version":"am","intended-application":"syndication"},{"URL":"https:\/\/academic.oup.com\/jid\/article-pdf\/233\/5\/840\/65900923\/jiaf609.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"syndication"},{"URL":"https:\/\/academic.oup.com\/jid\/article-pdf\/233\/5\/840\/65900923\/jiaf609.pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,5,14]],"date-time":"2026-05-14T23:58:00Z","timestamp":1778803080000},"score":1,"resource":{"primary":{"URL":"https:\/\/academic.oup.com\/jid\/article\/233\/5\/840\/8380346"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,12,16]]},"references-count":26,"journal-issue":{"issue":"5","published-online":{"date-parts":[[2025,12,16]]},"published-print":{"date-parts":[[2026,5,15]]}},"URL":"https:\/\/doi.org\/10.1093\/infdis\/jiaf609","relation":{},"ISSN":["0022-1899","1537-6613"],"issn-type":[{"value":"0022-1899","type":"print"},{"value":"1537-6613","type":"electronic"}],"subject":[],"published-other":{"date-parts":[[2026,5,15]]},"published":{"date-parts":[[2025,12,16]]}}}