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ARS Home » Midwest Area » Wooster, Ohio » Application Technology Research » Research » Publications at this Location » Publication #410300
Research Project: Sustainable Production and Pest Management Practices for Nursery, Greenhouse, and Protected Culture Crops

Location: Application Technology Research

Title: A quantitative real-time PCR method to detect the quinoa downy mildew pathogen, Peronospora variabilis

Author
item Testen, Anna
item PURI, PURNIMA - Washington State University
item Shaw, Robert - Scott
item DOMSIC, EVAN - Washington State University
item GRIFFIN-LAHUE, DEIDRE - Washington State University
item MURPHY, KEVIN - Washington State University
item MATTUPALLI, CHAKRADHAR - Washington State University

Submitted to: Plant Disease
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 5/16/2024
Publication Date: 9/3/2024
Citation: Testen, A.L., Puri, P., Shaw, R.S., Domsic, E.C., Griffin-Lahue, D., Murphy, K.M., Mattupalli, C. 2024. A quantitative real-time PCR method to detect the quinoa downy mildew pathogen, Peronospora variabilis. Plant Disease. 108(9):2887-2893. https://doi.org/10.1094/PDIS-11-23-2308-RE.
DOI: https://doi.org/10.1094/PDIS-11-23-2308-RE

Interpretive Summary: Quinoa is a highly nutritious crop exported from the Andean Region of South America and is an emerging crop in the United States. Quinoa production is harmed by quinoa downy mildew, caused by Peronospora variabilis. This pathogen spreads by seed and cannot be grown in a lab, which makes its detection and quantification difficult. We developed a quantitative real-time PCR (qPCR) assay to detect P. variabilis in quinoa seeds and leaves. This qPCR assay will speed diagnosis of quinoa downy mildew, allow for detection and quantification of the pathogen in seeds, and provide us with a better understanding of how this pathogen infects quinoa to improve disease management.

Technical Abstract: Quinoa downy mildew, caused by Peronospora variabilis, is the most devastating disease of quinoa globally. Rapid and sensitive diagnostic methods are needed to detect and quantify this obligate seedborne pathogen in seeds and plant tissue. A hydrolysis probe-based quantitative real-time PCR (qPCR) assay including a competitive internal control was developed for P. variabilis detection. This assay could detect as low as 20 ag of DNA or approximately 25 ITS copies per reaction with efficiencies ranging from 93.9-98.2%. No non-target amplification was observed when the assay was tested against DNA from other downy mildew pathogens and related oomycetes. Peronospora variabilis strains from multiple countries were detected using this assay. The assay was successfully applied to evaluate pathogen load in quinoa seeds from a field trial conducted in 2022 in Washington state. Downy mildew disease was recorded on all 14 genotypes with the genotypes 106.49 and 104.88 recording the highest area under the disease progress values (285 ± 20 and 324 ± 30, respectively) while J6 and Dutchess recorded the lowest (44 ± 11 and 41± 13, respectively). Seed washes obtained from the field samples were subjected to the qPCR assay, and the pathogen was detected in all the samples tested. Highest pathogen ITS copy number was recorded with 106.49 (194,934 ± 38,171), while the lowest was observed with Pasto (5,971 ± 1,435) and Riobamba (9,954 ± 4,243). This qPCR assay could lead to improved detection and quantification of P. variabilis and increase understanding of quinoa–P. variabilis interactions and epidemiology of this pathosystem.