In situ hybridization for the localization of two pepino mosaic virus isolates in mixed infections

In situ hybridization (ISH) is an informative and relatively accessible technique for the localization of viral genomes in plant tissue and cells. However, simultaneous visualization of related plant viruses in mixed infections may be limited by the nucleotide similarity in the genomes and the singl...

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Bibliographic Details
Published in:Journal of virological methods 2019-05, Vol.267, p.42-47
Main Authors: Gómez-Aix, C., Alcaide, C., Gómez, P., Aranda, M.A., Sánchez-Pina, M.A.
Format: Article
Language:English
Subjects:
Coinfection - virology
Genome, Viral
In Situ Hybridization
Mixed infections
Nicotiana - virology
Nicotiana benthamiana
PepMV
Plant Diseases - virology
Plant Leaves - virology
Potexvirus - isolation & purification
RNA localization
RNA Probes
Viral RNA
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Summary:In situ hybridization (ISH) is an informative and relatively accessible technique for the localization of viral genomes in plant tissue and cells. However, simultaneous visualization of related plant viruses in mixed infections may be limited by the nucleotide similarity in the genomes and the single chromogenic detection over the same sample preparation. To address this issue, we used two Pepino mosaic virus isolates and performed ISH over consecutive serial cross-sections of paraffin-embedded leaf samples of single and mixed infected Nicotiana benthamiana plants. Moreover, the probe design was optimized to reduce cross-hybridisation, and co-localization was based on the overlapping of consecutive cross-sections from mixed infected leaves; thus, our results showed that both Pepino mosaic virus isolates co-localized in the same leaf tissue. In turn, both isolates were localized in the cytoplasm of the same cells. These results provide valuable information for studying mixed infections in plants by using a simple ISH procedure that is accessible to any pathology laboratory.
ISSN:0166-0934
1879-0984
DOI:10.1016/j.jviromet.2019.02.003