Journal
PLANT DISEASE
Volume 105, Issue 10, Pages 3154-3161Publisher
AMER PHYTOPATHOLOGICAL SOC
DOI: 10.1094/PDIS-11-20-2339-RE
Keywords
pathogen detection and quantification; downy mildew; cucurbits; hop
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Funding
- Agriculture and Food Research Initiative Food Security [2016-68004-24931]
- Pickle Packers International
- U.S. Department of Agriculture Animal and Plant Health Inspection Service Awards [13-8130-0254-CA, 13-8130-0274-CA]
- U.S. Department of Agriculture National Institute of Food and Agriculture [2016-68004-024931]
- U.S. Department of Agriculture North Carolina Department of Agriculture Specialty Crop Block [12-25-B-16-88, 15SCBGP0003]
- North Carolina State Hatch Project [NC02418, NC02628]
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The study developed a highly specific multiplex TaqMan PCR assay that can accurately detect and quantify three aerially dispersed plant pathogens in a single amplification, improving management options for controlling the diseases they cause.
The ability to detect and quantify aerially dispersed plant pathogens is essential for developing effective disease control measures and epidemiological models that optimize the timing for control. There is an acute need for managing the downy mildew pathogens infecting cucurbits and hop incited by members of the genus Pseudoperonospora (Pseudoperonospora cubensis clade 1 and 2 isolates and Pseudoperonospora humuli, respectively). A highly specific multiplex TaqMan quantitative polymerase chain reaction (PCR) assay targeting unique sequences in the pathogens' mitochondrial genomes was developed that enables detection of all three taxa in a single multiplexed amplification. An internal control included in the reaction evaluated whether results were influenced by PCR inhibitors that can make it through the DNA extraction process. Reliable quantification of inoculum as low as three sporangia in a sample was observed. The multiplexed assay was tested with DNA extracted from purified sporangia, infected plant tissue, and environmental samples collected on impaction spore traps samplers. The ability to accurately detect and simultaneously quantify all three pathogens in a single multiplexed amplification should improve management options for controlling the diseases they cause.
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