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ARS Home » Pacific West Area » Corvallis, Oregon » Forage Seed and Cereal Research Unit » Research » Publications at this Location » Publication #411578
Research Project: Development of Superior Hops and Resilient Hop Production Systems

Location: Forage Seed and Cereal Research Unit

Title: Detection of Podosphaera macularis in air samples by quantitative PCR

Author
item Gent, David - Dave
item Adair, Nanci
item HATLEN, ROSS - Michigan State University
item MILES, TIMOTHY - Michigan State University
item RICHARDSON, BRIANA - Oregon State University
item Rivedal, Hannah
item ROSS, CAMERON - Oregon State University
item WISEMAN, MICHELE - Oregon State University

Submitted to: Plant Disease
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 4/29/2024
Publication Date: 5/7/2024
Citation: Gent, D.H., Adair, N.L., Hatlen, R., Miles, T.D., Richardson, B.J., Rivedal, H.M., Ross, C., Wiseman, M.S. 2024. Detection of Podosphaera macularis in air samples by quantitative PCR. Plant Disease. https://doi.org/10.1094/PDIS-04-24-0894-RE.
DOI: https://doi.org/10.1094/PDIS-04-24-0894-RE

Interpretive Summary: Disease organisms that spread in the air create difficult management problems for growers because there is uncertainty on when the organism is present until after disease symptoms develop. Methods to detect disease organisms in the air are useful for informing of potential disease risk and as research tools to understand patterns of dispersal. In this research, we developed a DNA-based diagnostic assay that is capable of detecting and quantifying the hop powdery mildew fungus. After rigorous testing to ensure the assay was robust and fit for the intended purpose, we coupled the assay to air sampling devices to quantify the patterns of when and where the fungus is present. We were able to detect the pathogen at all locations, including field sites multiple kilometers away from the nearest known hop yard. The assay should be a useful tool for research and diagnostic applications.

Technical Abstract: Detection and quantification of pathogen propagules in the air and environmental samples is facilitated by culture-independent assays. We developed a quantitative PCR assay for the hop powdery mildew fungus, Podosphaera macularis, that utilizes primers and a Taqman probe designed to target species-specific sequences in the 28S large submit (LSU) of the nuclear ribosomal rDNA. Assay performance was little affected by the presence of an exogenous internal control or potential PCR inhibitors associated with DNA extracted from soil. The assay had analytical sensitivity equivalent to 24 to 42 conidia when DNA was extracted from a fixed number of conidia, amplified all isolates of P. macularis tested, and had minimal cross-reactivity with closely related Podosphaera species when assayed with biologically relevant quantities of DNA. Standard curves generated independently in two other laboratories indicated that assay diagnostic sensitivity was qualitatively similar and reproducible. All laboratories successfully detected eight unknown isolates of P. macularis and correctly discriminated Pseudoperonospora humuli and a water control. Fitness for purpose for air sampling for late-season inoculum of P. macularis was demonstrated in field studies in 2019 and 2020. In both years, airborne populations of P. macularis were present consistently and increased in size during bloom and cone development. Given these characteristics, we expect the assay will be a useful tool for biosurvellience of P. macularis.