Specific Real-Time PCR for the Detection and Absolute Quantitation of Grapevine Roditis Leaf Discoloration-Associated Virus, an EPPO Alert Pathogen

Grapevine Roditis leaf discoloration-associated virus (GRLDaV) is an emerging grapevine pathogen included in the European and Mediterranean Plant Protection Organization (EPPO) alert list due to its ability to damage grapevine crops and cause production losses. This work aimed to develop a specific...

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Bibliographic Details
Published in:Plants (Basel) 2020-09, Vol.9 (9), p.1151
Main Authors: Morán, Félix, Sassalou, Chrysoula-Lito, Canales, Celia, Maliogka, Varvara I, Olmos, Antonio, Ruiz-García, Ana Belén
Format: Article
Language:English
Subjects:
Crop damage
Crop production
Deoxyribonucleic acid
Diagnostic software
Diagnostic systems
Discoloration
DNA
Flowers & plants
Genetic diversity
Genomes
Geography
grapevine
GRLDaV
Leaves
Methods
Nucleic acids
Pathogens
Plant protection
plant virus diagnostics
Quantitation
Real time
real-time PCR
Reproducibility
Selectivity
Sensitivity analysis
Viruses
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Summary:Grapevine Roditis leaf discoloration-associated virus (GRLDaV) is an emerging grapevine pathogen included in the European and Mediterranean Plant Protection Organization (EPPO) alert list due to its ability to damage grapevine crops and cause production losses. This work aimed to develop a specific and reliable diagnostic tool that would contribute to preventing the spread of this pathogen. Therefore, a TaqMan real-time quantitative PCR was developed. The method was validated according to EPPO guidelines showing a high degree of analytical sensitivity, analytical specificity, selectivity, and repeatability and reproducibility. The sensitivity of this method is much higher than the sensitivity reached by previously reported methods even when tested in crude extracts, which could allow rapid testing by avoiding nucleic acid extraction steps. The method was also able to detect GRLDaV isolates from all the geographic origins reported so far, despite their high degree of genetic diversity. In addition, this new technique has been successfully applied for the quantitative detection of GRLDaV in plant material and two mealybug species, and . In conclusion, the methodology developed herein represents a significant contribution to the diagnosis and control of this emerging pathogen in grapevine.
ISSN:2223-7747
2223-7747
DOI:10.3390/plants9091151