Development of Nanobodies against Mal de Río Cuarto virus major viroplasm protein P9-1 for diagnostic sandwich ELISA and immunodetection

Abstract Mal de Río Cuarto virus (MRCV) is a member of the genus Fijivirus of the family Reoviridae that causes a devastating disease in maize and is persistently and propagatively transmitted by planthopper vectors. Virus replication and assembly occur within viroplasms formed by viral and host pro...

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Autores principales:	Gabriela Llauger, Demián Monti, Matías Adúriz, Ema Romão, Analía Delina Dumón, María Fernanda Mattio, Andrés Wigdorovitz, Serge Muyldermans, Cécile Vincke, Viviana Parreño, Mariana del Vas
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2021
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/c0685c6872914794bdfc53f26ee2fc9c
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spelling	oai:doaj.org-article:c0685c6872914794bdfc53f26ee2fc9c2021-12-02T19:16:11ZDevelopment of Nanobodies against Mal de Río Cuarto virus major viroplasm protein P9-1 for diagnostic sandwich ELISA and immunodetection10.1038/s41598-021-99275-z2045-2322https://doaj.org/article/c0685c6872914794bdfc53f26ee2fc9c2021-10-01T00:00:00Zhttps://doi.org/10.1038/s41598-021-99275-zhttps://doaj.org/toc/2045-2322Abstract Mal de Río Cuarto virus (MRCV) is a member of the genus Fijivirus of the family Reoviridae that causes a devastating disease in maize and is persistently and propagatively transmitted by planthopper vectors. Virus replication and assembly occur within viroplasms formed by viral and host proteins. This work describes the isolation and characterization of llama-derived Nanobodies (Nbs) recognizing the major viral viroplasm component, P9-1. Specific Nbs were selected against recombinant P9-1, with affinities in the nanomolar range as measured by surface plasmon resonance. Three selected Nbs were fused to alkaline phosphatase and eGFP to develop a sandwich ELISA test which showed a high diagnostic sensitivity (99.12%, 95% CI 95.21–99.98) and specificity (100%, 95% CI 96.31–100) and a detection limit of 0.236 ng/ml. Interestingly, these Nanobodies recognized different P9-1 conformations and were successfully employed to detect P9-1 in pull-down assays of infected maize extracts. Finally, we demonstrated that fusions of the Nbs to eGFP and RFP allowed the immunodetection of virus present in phloem cells of leaf thin sections. The Nbs developed in this work will aid the study of MRCV epidemiology, assist maize breeding programs, and be valuable tools to boost fundamental research on viroplasm structure and maturation.Gabriela LlaugerDemián MontiMatías AdúrizEma RomãoAnalía Delina DumónMaría Fernanda MattioAndrés WigdorovitzSerge MuyldermansCécile VinckeViviana ParreñoMariana del VasNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 11, Iss 1, Pp 1-13 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Gabriela Llauger Demián Monti Matías Adúriz Ema Romão Analía Delina Dumón María Fernanda Mattio Andrés Wigdorovitz Serge Muyldermans Cécile Vincke Viviana Parreño Mariana del Vas Development of Nanobodies against Mal de Río Cuarto virus major viroplasm protein P9-1 for diagnostic sandwich ELISA and immunodetection
description	Abstract Mal de Río Cuarto virus (MRCV) is a member of the genus Fijivirus of the family Reoviridae that causes a devastating disease in maize and is persistently and propagatively transmitted by planthopper vectors. Virus replication and assembly occur within viroplasms formed by viral and host proteins. This work describes the isolation and characterization of llama-derived Nanobodies (Nbs) recognizing the major viral viroplasm component, P9-1. Specific Nbs were selected against recombinant P9-1, with affinities in the nanomolar range as measured by surface plasmon resonance. Three selected Nbs were fused to alkaline phosphatase and eGFP to develop a sandwich ELISA test which showed a high diagnostic sensitivity (99.12%, 95% CI 95.21–99.98) and specificity (100%, 95% CI 96.31–100) and a detection limit of 0.236 ng/ml. Interestingly, these Nanobodies recognized different P9-1 conformations and were successfully employed to detect P9-1 in pull-down assays of infected maize extracts. Finally, we demonstrated that fusions of the Nbs to eGFP and RFP allowed the immunodetection of virus present in phloem cells of leaf thin sections. The Nbs developed in this work will aid the study of MRCV epidemiology, assist maize breeding programs, and be valuable tools to boost fundamental research on viroplasm structure and maturation.
format	article
author	Gabriela Llauger Demián Monti Matías Adúriz Ema Romão Analía Delina Dumón María Fernanda Mattio Andrés Wigdorovitz Serge Muyldermans Cécile Vincke Viviana Parreño Mariana del Vas
author_facet	Gabriela Llauger Demián Monti Matías Adúriz Ema Romão Analía Delina Dumón María Fernanda Mattio Andrés Wigdorovitz Serge Muyldermans Cécile Vincke Viviana Parreño Mariana del Vas
author_sort	Gabriela Llauger
title	Development of Nanobodies against Mal de Río Cuarto virus major viroplasm protein P9-1 for diagnostic sandwich ELISA and immunodetection
title_short	Development of Nanobodies against Mal de Río Cuarto virus major viroplasm protein P9-1 for diagnostic sandwich ELISA and immunodetection
title_full	Development of Nanobodies against Mal de Río Cuarto virus major viroplasm protein P9-1 for diagnostic sandwich ELISA and immunodetection
title_fullStr	Development of Nanobodies against Mal de Río Cuarto virus major viroplasm protein P9-1 for diagnostic sandwich ELISA and immunodetection
title_full_unstemmed	Development of Nanobodies against Mal de Río Cuarto virus major viroplasm protein P9-1 for diagnostic sandwich ELISA and immunodetection
title_sort	development of nanobodies against mal de río cuarto virus major viroplasm protein p9-1 for diagnostic sandwich elisa and immunodetection
publisher	Nature Portfolio
publishDate	2021
url	https://doaj.org/article/c0685c6872914794bdfc53f26ee2fc9c
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Development of Nanobodies against Mal de Río Cuarto virus major viroplasm protein P9-1 for diagnostic sandwich ELISA and immunodetection

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