Preprint Article Version 1 Preserved in Portico This version is not peer-reviewed

Optimization of RT-QuIC Assay Duration for Screening Chronic Wasting Disease in White-Tailed Deer

Gokhan Yilmaz
ORCID logo ,
Tamara Morrill
,
Cian Ward
,
Gordon Mitchell
,
Andrei Soutyrine
,
Hanhong Dan
,
Min Lin
,
Jiewen Guan
*
Version 1 : Received: 6 September 2023 / Approved: 7 September 2023 / Online: 7 September 2023 (15:45:02 CEST)

A peer-reviewed article of this Preprint also exists.

Yilmaz, G.; Morrill, T.; Pilot, W.; Ward, C.; Mitchell, G.; Soutyrine, A.; Dan, H.; Lin, M.; Guan, J. Optimization of RT-QuIC Assay Duration for Screening Chronic Wasting Disease in White-Tailed Deer. Vet. Sci. 2024, 11, 60. Yilmaz, G.; Morrill, T.; Pilot, W.; Ward, C.; Mitchell, G.; Soutyrine, A.; Dan, H.; Lin, M.; Guan, J. Optimization of RT-QuIC Assay Duration for Screening Chronic Wasting Disease in White-Tailed Deer. Vet. Sci. 2024, 11, 60.

Abstract

Real-time quaking-induced conversion (RT-QuIC) assays have become common in the detection of chronic wasting disease (CWD) and are very sensitive provided the assay duration is sufficient. However, a prolonged assay duration may lead to non-specific signal amplification. The wide range of pre-defined assay durations in current RT-QuIC applications presents a need for optimization of the RT-QuIC assay duration. In this study, receiver operating characteristic (ROC) analysis was applied to optimize assay duration for detection of CWD in obex and retropharyngeal lymph node (RLN) tissue specimens. Two different fluorescence thresholds were used: a fixed threshold based on background fluorescence (Tstdev) and a max-point ratio (maximum/background fluorescence) threshold (TMPR) to determine CWD positivity. The optimal assay duration was 27 h for both obex and RLN based on Tstdev, and 27 and 28 h for obex and RLN, respectively, based on TMPR. The optimized assay durations were then evaluated for screening CWD in white-tailed deer from an affected farm. Results by RT-QuIC using optimized duration based on Tstdev and TMPR were in 100% and 92.3 % or higher agreement with those by the widely used screening assay, ELISA. In comparison, when using a 40 h assay duration, the agreement between RT-QuIC and ELISA reduced to 89.2% or higher, and the RT-QuIC results were significantly (p < 0.05) different from those using optimum durations. These findings demonstrated that the application of ROC analysis for the optimization of assay duration could improve the RT-QuIC assay for screening CWD in white-tailed deer.

Keywords

RT-QuIC; Chronic Wasting Disease; Diagnostics; Optimization

Subject

Medicine and Pharmacology, Veterinary Medicine

Copyright: This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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