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. 2024 May 26;13(6):452.
doi: 10.3390/pathogens13060452.

Prion Seeding Activity in Plant Tissues Detected by RT-QuIC

Kate Burgener  1   2 , Stuart Siegfried Lichtenberg  3   4 , Daniel P Walsh  5 , Heather N Inzalaco  6 , Aaron Lomax  7   8 , Joel A Pedersen  1   2   7
Affiliations

Prion Seeding Activity in Plant Tissues Detected by RT-QuIC

Kate Burgener et al. Pathogens. .

Abstract

Prion diseases such as scrapie, bovine spongiform encephalopathy (BSE), and chronic wasting disease (CWD) affect domesticated and wild herbivorous mammals. Animals afflicted with CWD, the transmissible spongiform encephalopathy of cervids (deer, elk, and moose), shed prions into the environment, where they may persist and remain infectious for years. These environmental prions may remain in soil, be transported in surface waters, or assimilated into plants. Environmental sampling is an emerging area of TSE research and can provide more information about prion fate and transport once shed by infected animals. In this study, we have developed the first published method for the extraction and detection of prions in plant tissue using the real-time quaking-induced conversion (RT-QuIC) assay. Incubation with a zwitterionic surfactant followed by precipitation with sodium phosphotungstate concentrates the prions within samples and allows for sensitive detection of prion seeding activity. Using this protocol, we demonstrate that prions can be detected within plant tissues and on plant surfaces using the RT-QuIC assay.

Keywords: chronic wasting disease; environmental transmission; plants; prions; real-time quaking induced conversion; seeded amplification.

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Conflict of interest statement

Author Aaron Lomax was employed by the company Varizymes. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Figures

Figure 1
Figure 1
Real-time quaking-induced conversion (RT-QuIC) analysis of the prion extraction method efficiency. Box plots indicate the median time-to-threshold with a dashed horizontal red line, the mean with a solid horizontal red line, the second and third quartiles with the box, and the first and fourth quartiles with the whiskers. The horizontal line at 48 h indicates the end time of the assay. Each condition was performed with 16 technical replicates. (A) Threshold results from brain homogenate (BH) run at dilutions of 10−3 to 10−6. (B) Time-to-threshold results for BH subjected to the prion extraction method at dilutions of 10−2 to 10−5. (C) Threshold measurements for BH-spiked plant tissue (Brachypodium distachyon) subjected to the prion extraction method at dilutions of 10−2 to 10−5.
Figure 2
Figure 2
Prion deposition and removal from Elymus repens leaves. The box plots indicate the median time-to-threshold with a horizontal red line, the mean with a solid horizontal red line, the second and third quartiles with the box, and the first and fourth quartiles with the whiskers. E. repens leaves were exposed to CWD-positive brain homogenate and either soaked or rinsed with water, and then analyzed with real-time quacking-induced conversion (RT-QuIC).
Figure 3
Figure 3
Confirmation of prion extraction method efficacy with plants that demonstrate prion seeding activity. Alfalfa and barley plants that were hydroponically grown in prion-spiked water were subjected to prion extraction and analyzed by real-time quaking-induced conversion (RT-QuIC). The box plots indicate the median time-to-threshold with a horizontal red line, the mean with a solid horizontal red line, the second and third quartiles with the box, and the first and fourth quartiles with the whiskers.
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