The C-Terminal Half of SARS-CoV-2 Nucleocapsid Protein, Industrially Produced in Plants, Is Valid as Antigen in COVID-19 Serological Tests

Background: The fight against the current coronavirus disease 2019 (COVID-19) pandemic has created a huge demand of biotechnological, pharmaceutical, research and sanitary materials at unprecedented scales. One of the most urgent demands affects the diagnostic tests. The growing need for rapid and a...

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Autores: Williams, Laura, Jurado, Silvia, Llorente, Francisco, Romualdo, Alberto, González, Sara, Saconne, Ana, Bronchalo, Isabel, Martínez-Cortés, Mercedes, Pérez-Gómez, Beatriz, Ponz Ascaso, Fernando, Jiménez-Clavero, Miguel Ángel, Lunello, Pablo
Formato: artículo
Estado:Versión publicada
Fecha de publicación:2021
País:España
Recursos:Consejo Superior de Investigaciones Científicas (CSIC)
Repositorio:DIGITAL.CSIC. Repositorio Institucional del CSIC
OAI Identifier:oai:digital.csic.es:10261/268912
Acesso em linha:http://hdl.handle.net/10261/268912
Access Level:acceso abierto
Palavra-chave:COVID-19
SARS CoV-2 nucleocapsid protein
Biofactories
Plant expression
Molecular farming
ELISA coronavirus
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spelling The C-Terminal Half of SARS-CoV-2 Nucleocapsid Protein, Industrially Produced in Plants, Is Valid as Antigen in COVID-19 Serological TestsWilliams, LauraJurado, SilviaLlorente, FranciscoRomualdo, AlbertoGonzález, SaraSaconne, AnaBronchalo, IsabelMartínez-Cortés, MercedesPérez-Gómez, BeatrizPonz Ascaso, FernandoJiménez-Clavero, Miguel ÁngelLunello, PabloCOVID-19SARS CoV-2 nucleocapsid proteinBiofactoriesPlant expressionMolecular farmingELISA coronavirusBackground: The fight against the current coronavirus disease 2019 (COVID-19) pandemic has created a huge demand of biotechnological, pharmaceutical, research and sanitary materials at unprecedented scales. One of the most urgent demands affects the diagnostic tests. The growing need for rapid and accurate laboratory diagnostic tests requires the development of biotechnological processes aimed at producing reagents able to cope with this demand in a scalable, cost-effective manner, with rapid turnaround times. This is particularly applicable to the antigens employed in serological tests. Recombinant protein expression using plants as biofactories is particularly suitable for mass production of protein antigens useful in serological diagnosis, with a neat advantage in economic terms. Methods: We expressed a large portion of the nucleoprotein (N) derived from SARS-CoV-2 in Nicotiana benthamiana plants. After purification, the recombinant N protein obtained was used to develop an indirect enzyme-linked immunosorbent assay (ELISA) for detection of antibodies to SARS-CoV-2 in human sera. To validate the ELISA, a panel of 416 sera from exposed personnel at essential services in Madrid City Council were tested, and the results compared to those obtained by another ELISA, already validated, used as reference. Furthermore, a subset of samples for which RT-PCR results were available were used to confirm sensitivity and specificity of the test. Results: The performance of the N protein expressed in plants as antigen in serologic test for SARS-CoV-2 antibody detection was shown to be highly satisfactory, with calculated diagnostic sensitivity of 96.41% (95% CI: 93.05–98.44) and diagnostic specificity of 96.37 (95% CI: 93.05–98.44) as compared to the reference ELISA, with a kappa (K) value of 0.928 (95% CI:0.892–0.964). Furthermore, the ELISA developed with plant-derived N antigen detected SARS-CoV-2 antibodies in 84 out of 93 sera from individuals showing RT-PCR positive results (86/93 for the reference ELISA). Conclusion: This study demonstrates that the N protein part derived from SARS-CoV-2 expressed in plants performs as a perfectly valid antigen for use in COVID-19 diagnosis. Furthermore, our results support the use of this plant platform for expression of recombinant proteins as reagents for COVID-19 diagnosis. This platform stands out as a convenient and advantageous production system, fit-for-purpose to cope with the current demand of this type of biologicals in a cost-effective manner, making diagnostic kits more affordable.Work at Agrenvec and at the CBGP was funded in part by grant COV20-00114 from the Autonomous Region of Madrid (CAM) to FP. The CBGP thanks the Spanish Ministry of Science for the Severo Ochoa Excellence Accreditation (SEV-2016-0672).Peer reviewedFrontiers MediaComunidad de MadridMinisterio de Economía y Competitividad (España)Consejo Superior de Investigaciones Científicas [https://ror.org/02gfc7t72]202220222021info:eu-repo/semantics/articlehttp://purl.org/coar/resource_type/c_6501Publisher's versioninfo:eu-repo/semantics/publishedVersionhttp://hdl.handle.net/10261/268912reponame:DIGITAL.CSIC. Repositorio Institucional del CSICinstname:Consejo Superior de Investigaciones Científicas (CSIC)Inglés#PLACEHOLDER_PARENT_METADATA_VALUE#info:eu-repo/grantAgreement/MINECO//SEV-2016-067https://doi.org/10.3389/fpls.2021.699665Síinfo:eu-repo/semantics/openAccessoai:digital.csic.es:10261/2689122026-05-22T06:33:51Z
dc.title.none.fl_str_mv The C-Terminal Half of SARS-CoV-2 Nucleocapsid Protein, Industrially Produced in Plants, Is Valid as Antigen in COVID-19 Serological Tests
title The C-Terminal Half of SARS-CoV-2 Nucleocapsid Protein, Industrially Produced in Plants, Is Valid as Antigen in COVID-19 Serological Tests
spellingShingle The C-Terminal Half of SARS-CoV-2 Nucleocapsid Protein, Industrially Produced in Plants, Is Valid as Antigen in COVID-19 Serological Tests
Williams, Laura
COVID-19
SARS CoV-2 nucleocapsid protein
Biofactories
Plant expression
Molecular farming
ELISA coronavirus
title_short The C-Terminal Half of SARS-CoV-2 Nucleocapsid Protein, Industrially Produced in Plants, Is Valid as Antigen in COVID-19 Serological Tests
title_full The C-Terminal Half of SARS-CoV-2 Nucleocapsid Protein, Industrially Produced in Plants, Is Valid as Antigen in COVID-19 Serological Tests
title_fullStr The C-Terminal Half of SARS-CoV-2 Nucleocapsid Protein, Industrially Produced in Plants, Is Valid as Antigen in COVID-19 Serological Tests
title_full_unstemmed The C-Terminal Half of SARS-CoV-2 Nucleocapsid Protein, Industrially Produced in Plants, Is Valid as Antigen in COVID-19 Serological Tests
title_sort The C-Terminal Half of SARS-CoV-2 Nucleocapsid Protein, Industrially Produced in Plants, Is Valid as Antigen in COVID-19 Serological Tests
dc.creator.none.fl_str_mv Williams, Laura
Jurado, Silvia
Llorente, Francisco
Romualdo, Alberto
González, Sara
Saconne, Ana
Bronchalo, Isabel
Martínez-Cortés, Mercedes
Pérez-Gómez, Beatriz
Ponz Ascaso, Fernando
Jiménez-Clavero, Miguel Ángel
Lunello, Pablo
author Williams, Laura
author_facet Williams, Laura
Jurado, Silvia
Llorente, Francisco
Romualdo, Alberto
González, Sara
Saconne, Ana
Bronchalo, Isabel
Martínez-Cortés, Mercedes
Pérez-Gómez, Beatriz
Ponz Ascaso, Fernando
Jiménez-Clavero, Miguel Ángel
Lunello, Pablo
author_role author
author2 Jurado, Silvia
Llorente, Francisco
Romualdo, Alberto
González, Sara
Saconne, Ana
Bronchalo, Isabel
Martínez-Cortés, Mercedes
Pérez-Gómez, Beatriz
Ponz Ascaso, Fernando
Jiménez-Clavero, Miguel Ángel
Lunello, Pablo
author2_role author
author
author
author
author
author
author
author
author
author
author
dc.contributor.none.fl_str_mv Comunidad de Madrid
Ministerio de Economía y Competitividad (España)
Consejo Superior de Investigaciones Científicas [https://ror.org/02gfc7t72]
dc.subject.none.fl_str_mv COVID-19
SARS CoV-2 nucleocapsid protein
Biofactories
Plant expression
Molecular farming
ELISA coronavirus
topic COVID-19
SARS CoV-2 nucleocapsid protein
Biofactories
Plant expression
Molecular farming
ELISA coronavirus
description Background: The fight against the current coronavirus disease 2019 (COVID-19) pandemic has created a huge demand of biotechnological, pharmaceutical, research and sanitary materials at unprecedented scales. One of the most urgent demands affects the diagnostic tests. The growing need for rapid and accurate laboratory diagnostic tests requires the development of biotechnological processes aimed at producing reagents able to cope with this demand in a scalable, cost-effective manner, with rapid turnaround times. This is particularly applicable to the antigens employed in serological tests. Recombinant protein expression using plants as biofactories is particularly suitable for mass production of protein antigens useful in serological diagnosis, with a neat advantage in economic terms. Methods: We expressed a large portion of the nucleoprotein (N) derived from SARS-CoV-2 in Nicotiana benthamiana plants. After purification, the recombinant N protein obtained was used to develop an indirect enzyme-linked immunosorbent assay (ELISA) for detection of antibodies to SARS-CoV-2 in human sera. To validate the ELISA, a panel of 416 sera from exposed personnel at essential services in Madrid City Council were tested, and the results compared to those obtained by another ELISA, already validated, used as reference. Furthermore, a subset of samples for which RT-PCR results were available were used to confirm sensitivity and specificity of the test. Results: The performance of the N protein expressed in plants as antigen in serologic test for SARS-CoV-2 antibody detection was shown to be highly satisfactory, with calculated diagnostic sensitivity of 96.41% (95% CI: 93.05–98.44) and diagnostic specificity of 96.37 (95% CI: 93.05–98.44) as compared to the reference ELISA, with a kappa (K) value of 0.928 (95% CI:0.892–0.964). Furthermore, the ELISA developed with plant-derived N antigen detected SARS-CoV-2 antibodies in 84 out of 93 sera from individuals showing RT-PCR positive results (86/93 for the reference ELISA). Conclusion: This study demonstrates that the N protein part derived from SARS-CoV-2 expressed in plants performs as a perfectly valid antigen for use in COVID-19 diagnosis. Furthermore, our results support the use of this plant platform for expression of recombinant proteins as reagents for COVID-19 diagnosis. This platform stands out as a convenient and advantageous production system, fit-for-purpose to cope with the current demand of this type of biologicals in a cost-effective manner, making diagnostic kits more affordable.
publishDate 2021
dc.date.none.fl_str_mv 2021
2022
2022
dc.type.none.fl_str_mv info:eu-repo/semantics/article
http://purl.org/coar/resource_type/c_6501
Publisher's version
info:eu-repo/semantics/publishedVersion
format article
status_str publishedVersion
dc.identifier.none.fl_str_mv http://hdl.handle.net/10261/268912
url http://hdl.handle.net/10261/268912
dc.language.none.fl_str_mv Inglés
language_invalid_str_mv Inglés
dc.relation.none.fl_str_mv #PLACEHOLDER_PARENT_METADATA_VALUE#
info:eu-repo/grantAgreement/MINECO//SEV-2016-067
https://doi.org/10.3389/fpls.2021.699665
Sí
dc.rights.none.fl_str_mv info:eu-repo/semantics/openAccess
eu_rights_str_mv openAccess
dc.publisher.none.fl_str_mv Frontiers Media
publisher.none.fl_str_mv Frontiers Media
dc.source.none.fl_str_mv reponame:DIGITAL.CSIC. Repositorio Institucional del CSIC
instname:Consejo Superior de Investigaciones Científicas (CSIC)
instname_str Consejo Superior de Investigaciones Científicas (CSIC)
reponame_str DIGITAL.CSIC. Repositorio Institucional del CSIC
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