Author:
Llauger Gabriela,Monti Demián,Adúriz Matías,Romão Ema,Dumón Analía Delina,Mattio María Fernanda,Wigdorovitz Andrés,Muyldermans Serge,Vincke Cécile,Parreño Viviana,del Vas Mariana
Abstract
AbstractMal 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.
Funder
Agencia Nacional de Promoción Científica y Tecnológica
Instituto Nacional de Tecnología Agropecuaria
Publisher
Springer Science and Business Media LLC
Reference81 articles.
1. Lenardón, S. L., March, G. J., Nome, S. F. & Ornaghi, J. A. Recent outbreak of “Mal de Río Cuarto” virus on corn in Argentina. Plant Dis. 82, 448–448 (1998).
2. Marino de Remes Lenicov, A. M., Tesón, A., Dagoberto, E. & Huguet, N. Hallazgo de uno de los vectores del Mal de Río Cuarto virus en maíz. Gac. Agron. 25, 251–258 (1985).
3. Truol, G. A. et al. Transmisión experimental del virus del Mal de Río Cuarto por Delphacodes kuscheli. Phytopatologia Bras. 26, 39–44 (2001).
4. Wang, H. D., Chen, J. P., Wang, A. G., Jiang, X. H. & Adams, M. J. Studies on the epidemiology and yield losses from rice black-streaked dwarf disease in a recent epidemic in Zhejiang province, China. Plant Pathol. https://doi.org/10.1111/j.1365-3059.2009.02091.x (2009).
5. Zhou, G., Xu, D., Xu, D. & Zhang, M. Southern rice black-streaked dwarf virus: A white-backed planthopper-transmitted fijivirus threatening rice production in Asia. Front. Microbiol. https://doi.org/10.3389/fmicb.2013.00270 (2013).
Cited by
6 articles.
订阅此论文施引文献
订阅此论文施引文献,注册后可以免费订阅5篇论文的施引文献,订阅后可以查看论文全部施引文献