Optimization and Clinical Validation of Colorimetric Reverse Transcription Loop-Mediated Isothermal Amplification, a Fast, Highly Sensitive and Specific COVID-19 Molecular Diagnostic Tool That Is Robust to Detect SARS-CoV-2 Variants of Concern

The coronavirus disease 2019 (COVID-19) pandemic unfolded due to the widespread severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) transmission reinforced the urgent need for affordable molecular diagnostic alternative methods for massive testing screening. We present the clinical validati...

Descripción completa

Guardado en:

Detalles Bibliográficos
Autores principales:	Pedro A. Alves, Ellen G. de Oliveira, Ana Paula M. Franco-Luiz, Letícia T. Almeida, Amanda B. Gonçalves, Iara A. Borges, Flávia de S. Rocha, Raissa P. Rocha, Matheus F. Bezerra, Pâmella Miranda, Flávio D. Capanema, Henrique R. Martins, Gerald Weber, Santuza M. R. Teixeira, Gabriel Luz Wallau, Rubens L. do Monte-Neto
Formato:	article
Lenguaje:	EN
Publicado:	Frontiers Media S.A. 2021
Materias:	COVID-19 RT-LAMP SARS-CoV-2 molecular test respiratory virus diagnostic test Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/c0a5c7b1038f408d910e62de6e30d448
Etiquetas:	Agregar Etiqueta Sin Etiquetas, Sea el primero en etiquetar este registro!

id	oai:doaj.org-article:c0a5c7b1038f408d910e62de6e30d448
record_format	dspace
spelling	oai:doaj.org-article:c0a5c7b1038f408d910e62de6e30d4482021-11-18T08:46:16ZOptimization and Clinical Validation of Colorimetric Reverse Transcription Loop-Mediated Isothermal Amplification, a Fast, Highly Sensitive and Specific COVID-19 Molecular Diagnostic Tool That Is Robust to Detect SARS-CoV-2 Variants of Concern1664-302X10.3389/fmicb.2021.713713https://doaj.org/article/c0a5c7b1038f408d910e62de6e30d4482021-11-01T00:00:00Zhttps://www.frontiersin.org/articles/10.3389/fmicb.2021.713713/fullhttps://doaj.org/toc/1664-302XThe coronavirus disease 2019 (COVID-19) pandemic unfolded due to the widespread severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) transmission reinforced the urgent need for affordable molecular diagnostic alternative methods for massive testing screening. We present the clinical validation of a pH-dependent colorimetric reverse transcription loop-mediated isothermal amplification (RT-LAMP) for SARS-CoV-2 detection. The method revealed a limit of detection of 19.3 ± 2.7 viral genomic copies/μL when using RNA extracted samples obtained from nasopharyngeal swabs collected in guanidine-containing viral transport medium. Typical RT-LAMP reactions were performed at 65°C for 30 min. When compared to reverse transcriptase–quantitative polymerase chain reaction (RT-qPCR), up to cycle-threshold (Ct) value 32, RT-LAMP presented 98% [95% confidence interval (CI) = 95.3–99.5%] sensitivity and 100% (95% CI = 94.5–100%) specificity for SARS-CoV-2 RNA detection targeting E and N genes. No cross-reactivity was detected when testing other non–SARS-CoV virus, confirming high specificity. The test is compatible with primary RNA extraction–free samples. We also demonstrated that colorimetric RT-LAMP can detect SARS-CoV-2 variants of concern and variants of interest, such as variants occurring in Brazil named gamma (P.1), zeta (P.2), delta (B.1.617.2), B.1.1.374, and B.1.1.371. The method meets point-of-care requirements and can be deployed in the field for high-throughput COVID-19 testing campaigns, especially in countries where COVID-19 testing efforts are far from ideal to tackle the pandemics. Although RT-qPCR is considered the gold standard for SARS-CoV-2 RNA detection, it requires expensive equipment, infrastructure, and highly trained personnel. In contrast, RT-LAMP emerges as an affordable, inexpensive, and simple alternative for SARS-CoV-2 molecular detection that can be applied to massive COVID-19 testing campaigns and save lives.Pedro A. AlvesPedro A. AlvesEllen G. de OliveiraAna Paula M. Franco-LuizLetícia T. AlmeidaAmanda B. GonçalvesIara A. BorgesFlávia de S. RochaRaissa P. RochaMatheus F. BezerraPâmella MirandaFlávio D. CapanemaHenrique R. MartinsHenrique R. MartinsGerald WeberSantuza M. R. TeixeiraGabriel Luz WallauRubens L. do Monte-NetoFrontiers Media S.A.articleCOVID-19RT-LAMPSARS-CoV-2molecular testrespiratory virusdiagnostic testMicrobiologyQR1-502ENFrontiers in Microbiology, Vol 12 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	COVID-19 RT-LAMP SARS-CoV-2 molecular test respiratory virus diagnostic test Microbiology QR1-502
spellingShingle	COVID-19 RT-LAMP SARS-CoV-2 molecular test respiratory virus diagnostic test Microbiology QR1-502 Pedro A. Alves Pedro A. Alves Ellen G. de Oliveira Ana Paula M. Franco-Luiz Letícia T. Almeida Amanda B. Gonçalves Iara A. Borges Flávia de S. Rocha Raissa P. Rocha Matheus F. Bezerra Pâmella Miranda Flávio D. Capanema Henrique R. Martins Henrique R. Martins Gerald Weber Santuza M. R. Teixeira Gabriel Luz Wallau Rubens L. do Monte-Neto Optimization and Clinical Validation of Colorimetric Reverse Transcription Loop-Mediated Isothermal Amplification, a Fast, Highly Sensitive and Specific COVID-19 Molecular Diagnostic Tool That Is Robust to Detect SARS-CoV-2 Variants of Concern
description	The coronavirus disease 2019 (COVID-19) pandemic unfolded due to the widespread severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) transmission reinforced the urgent need for affordable molecular diagnostic alternative methods for massive testing screening. We present the clinical validation of a pH-dependent colorimetric reverse transcription loop-mediated isothermal amplification (RT-LAMP) for SARS-CoV-2 detection. The method revealed a limit of detection of 19.3 ± 2.7 viral genomic copies/μL when using RNA extracted samples obtained from nasopharyngeal swabs collected in guanidine-containing viral transport medium. Typical RT-LAMP reactions were performed at 65°C for 30 min. When compared to reverse transcriptase–quantitative polymerase chain reaction (RT-qPCR), up to cycle-threshold (Ct) value 32, RT-LAMP presented 98% [95% confidence interval (CI) = 95.3–99.5%] sensitivity and 100% (95% CI = 94.5–100%) specificity for SARS-CoV-2 RNA detection targeting E and N genes. No cross-reactivity was detected when testing other non–SARS-CoV virus, confirming high specificity. The test is compatible with primary RNA extraction–free samples. We also demonstrated that colorimetric RT-LAMP can detect SARS-CoV-2 variants of concern and variants of interest, such as variants occurring in Brazil named gamma (P.1), zeta (P.2), delta (B.1.617.2), B.1.1.374, and B.1.1.371. The method meets point-of-care requirements and can be deployed in the field for high-throughput COVID-19 testing campaigns, especially in countries where COVID-19 testing efforts are far from ideal to tackle the pandemics. Although RT-qPCR is considered the gold standard for SARS-CoV-2 RNA detection, it requires expensive equipment, infrastructure, and highly trained personnel. In contrast, RT-LAMP emerges as an affordable, inexpensive, and simple alternative for SARS-CoV-2 molecular detection that can be applied to massive COVID-19 testing campaigns and save lives.
format	article
author	Pedro A. Alves Pedro A. Alves Ellen G. de Oliveira Ana Paula M. Franco-Luiz Letícia T. Almeida Amanda B. Gonçalves Iara A. Borges Flávia de S. Rocha Raissa P. Rocha Matheus F. Bezerra Pâmella Miranda Flávio D. Capanema Henrique R. Martins Henrique R. Martins Gerald Weber Santuza M. R. Teixeira Gabriel Luz Wallau Rubens L. do Monte-Neto
author_facet	Pedro A. Alves Pedro A. Alves Ellen G. de Oliveira Ana Paula M. Franco-Luiz Letícia T. Almeida Amanda B. Gonçalves Iara A. Borges Flávia de S. Rocha Raissa P. Rocha Matheus F. Bezerra Pâmella Miranda Flávio D. Capanema Henrique R. Martins Henrique R. Martins Gerald Weber Santuza M. R. Teixeira Gabriel Luz Wallau Rubens L. do Monte-Neto
author_sort	Pedro A. Alves
title	Optimization and Clinical Validation of Colorimetric Reverse Transcription Loop-Mediated Isothermal Amplification, a Fast, Highly Sensitive and Specific COVID-19 Molecular Diagnostic Tool That Is Robust to Detect SARS-CoV-2 Variants of Concern
title_short	Optimization and Clinical Validation of Colorimetric Reverse Transcription Loop-Mediated Isothermal Amplification, a Fast, Highly Sensitive and Specific COVID-19 Molecular Diagnostic Tool That Is Robust to Detect SARS-CoV-2 Variants of Concern
title_full	Optimization and Clinical Validation of Colorimetric Reverse Transcription Loop-Mediated Isothermal Amplification, a Fast, Highly Sensitive and Specific COVID-19 Molecular Diagnostic Tool That Is Robust to Detect SARS-CoV-2 Variants of Concern
title_fullStr	Optimization and Clinical Validation of Colorimetric Reverse Transcription Loop-Mediated Isothermal Amplification, a Fast, Highly Sensitive and Specific COVID-19 Molecular Diagnostic Tool That Is Robust to Detect SARS-CoV-2 Variants of Concern
title_full_unstemmed	Optimization and Clinical Validation of Colorimetric Reverse Transcription Loop-Mediated Isothermal Amplification, a Fast, Highly Sensitive and Specific COVID-19 Molecular Diagnostic Tool That Is Robust to Detect SARS-CoV-2 Variants of Concern
title_sort	optimization and clinical validation of colorimetric reverse transcription loop-mediated isothermal amplification, a fast, highly sensitive and specific covid-19 molecular diagnostic tool that is robust to detect sars-cov-2 variants of concern
publisher	Frontiers Media S.A.
publishDate	2021
url	https://doaj.org/article/c0a5c7b1038f408d910e62de6e30d448
work_keys_str_mv	AT pedroaalves optimizationandclinicalvalidationofcolorimetricreversetranscriptionloopmediatedisothermalamplificationafasthighlysensitiveandspecificcovid19moleculardiagnostictoolthatisrobusttodetectsarscov2variantsofconcern AT pedroaalves optimizationandclinicalvalidationofcolorimetricreversetranscriptionloopmediatedisothermalamplificationafasthighlysensitiveandspecificcovid19moleculardiagnostictoolthatisrobusttodetectsarscov2variantsofconcern AT ellengdeoliveira optimizationandclinicalvalidationofcolorimetricreversetranscriptionloopmediatedisothermalamplificationafasthighlysensitiveandspecificcovid19moleculardiagnostictoolthatisrobusttodetectsarscov2variantsofconcern AT anapaulamfrancoluiz optimizationandclinicalvalidationofcolorimetricreversetranscriptionloopmediatedisothermalamplificationafasthighlysensitiveandspecificcovid19moleculardiagnostictoolthatisrobusttodetectsarscov2variantsofconcern AT leticiatalmeida optimizationandclinicalvalidationofcolorimetricreversetranscriptionloopmediatedisothermalamplificationafasthighlysensitiveandspecificcovid19moleculardiagnostictoolthatisrobusttodetectsarscov2variantsofconcern AT amandabgoncalves optimizationandclinicalvalidationofcolorimetricreversetranscriptionloopmediatedisothermalamplificationafasthighlysensitiveandspecificcovid19moleculardiagnostictoolthatisrobusttodetectsarscov2variantsofconcern AT iaraaborges optimizationandclinicalvalidationofcolorimetricreversetranscriptionloopmediatedisothermalamplificationafasthighlysensitiveandspecificcovid19moleculardiagnostictoolthatisrobusttodetectsarscov2variantsofconcern AT flaviadesrocha optimizationandclinicalvalidationofcolorimetricreversetranscriptionloopmediatedisothermalamplificationafasthighlysensitiveandspecificcovid19moleculardiagnostictoolthatisrobusttodetectsarscov2variantsofconcern AT raissaprocha optimizationandclinicalvalidationofcolorimetricreversetranscriptionloopmediatedisothermalamplificationafasthighlysensitiveandspecificcovid19moleculardiagnostictoolthatisrobusttodetectsarscov2variantsofconcern AT matheusfbezerra optimizationandclinicalvalidationofcolorimetricreversetranscriptionloopmediatedisothermalamplificationafasthighlysensitiveandspecificcovid19moleculardiagnostictoolthatisrobusttodetectsarscov2variantsofconcern AT pamellamiranda optimizationandclinicalvalidationofcolorimetricreversetranscriptionloopmediatedisothermalamplificationafasthighlysensitiveandspecificcovid19moleculardiagnostictoolthatisrobusttodetectsarscov2variantsofconcern AT flaviodcapanema optimizationandclinicalvalidationofcolorimetricreversetranscriptionloopmediatedisothermalamplificationafasthighlysensitiveandspecificcovid19moleculardiagnostictoolthatisrobusttodetectsarscov2variantsofconcern AT henriquermartins optimizationandclinicalvalidationofcolorimetricreversetranscriptionloopmediatedisothermalamplificationafasthighlysensitiveandspecificcovid19moleculardiagnostictoolthatisrobusttodetectsarscov2variantsofconcern AT henriquermartins optimizationandclinicalvalidationofcolorimetricreversetranscriptionloopmediatedisothermalamplificationafasthighlysensitiveandspecificcovid19moleculardiagnostictoolthatisrobusttodetectsarscov2variantsofconcern AT geraldweber optimizationandclinicalvalidationofcolorimetricreversetranscriptionloopmediatedisothermalamplificationafasthighlysensitiveandspecificcovid19moleculardiagnostictoolthatisrobusttodetectsarscov2variantsofconcern AT santuzamrteixeira optimizationandclinicalvalidationofcolorimetricreversetranscriptionloopmediatedisothermalamplificationafasthighlysensitiveandspecificcovid19moleculardiagnostictoolthatisrobusttodetectsarscov2variantsofconcern AT gabrielluzwallau optimizationandclinicalvalidationofcolorimetricreversetranscriptionloopmediatedisothermalamplificationafasthighlysensitiveandspecificcovid19moleculardiagnostictoolthatisrobusttodetectsarscov2variantsofconcern AT rubensldomonteneto optimizationandclinicalvalidationofcolorimetricreversetranscriptionloopmediatedisothermalamplificationafasthighlysensitiveandspecificcovid19moleculardiagnostictoolthatisrobusttodetectsarscov2variantsofconcern
_version_	1718421346813214720

Optimization and Clinical Validation of Colorimetric Reverse Transcription Loop-Mediated Isothermal Amplification, a Fast, Highly Sensitive and Specific COVID-19 Molecular Diagnostic Tool That Is Robust to Detect SARS-CoV-2 Variants of Concern

Ejemplares similares