Location: Emerging Pests and Pathogens Research
Project Number: 8062-21000-042-006-S
Project Type: Non-Assistance Cooperative Agreement
Start Date: May 1, 2020
End Date: Apr 30, 2022
The overall objective of this project is to provide seed certification with protocols for determining disease incidence estimates using dormant tuber testing and to help with the implementation of the new procedures. Specific objectives are: 1. Optimize PCR-based diagnostics for important viruses and bacteria using the DNA/RNA extracted from the Flinders Technology Associates (FTA) cards. 2. Compare the results obtained from the new assay with current protocols used by seed certification.
Experimental Approach: Objective 1: The focus will be on further validating testing protocols by expanding PCR-based testing to include other viruses, bacteria, and pathogens and optimizing PCR-based detection assay using real-time quantitative PCR (qPCR) or high throughput sequencing (HTS). Based on the grower survey, other pathogens, including bacteria, were reported as a concern for growers. Objective 2: Our objective is to compare direct tuber testing with visual/ELISA protocols now in use. This comparison is limited to PVY since the current methods do not allow for accurate determination of levels of other pathogens. We will select two seed lots from a potato seed certification program (Colorado or Idaho), with an elevated PVY infection level (based on summer inspections) and subject the 400-tuber sample to a direct tuber testing using the developed FTA-card protocol, with a subsequent winter grow-out test of the same 400-tuber set in Hawaii. Leaves of the emerging plants will be collected and subjected to a conventional enzyme-linked immunosorbent assay (ELISA) testing, which is currently a “gold standard” of the potato certification. The data will be compared side-by-side for further evaluation and presentation to the industry panel. Testing of FTA cards will be conducted by various PCR-based methods, such as conventional RT-PCR, RT-qPCR, and HTS.