Utilizing PPE for the detection of environmental chronic wasting disease prions

Introduction/Objective: Chronic wasting disease (CWD) infectious prion proteins (PrP^Sc) circulate within environmental settings for years, posing significant challenges to the effective management of CWD in cervid populations. Despite these challenges, the environmental persistence of infectious CWD prions can be leveraged for new CWD surveillance and management strategies. Recent advancements in seeded amplification technologies (Real-Time Quaking Induced Conversion and Protein Misfolding Cyclic Amplification, PMCA) are facilitating opportunities for environmental-based detection of CWD prions. We hypothesized that personal protective equipment (PPE) exposed to CWD-positive environmental matrices could be leveraged as active PrP^Sc sentinels. Methods: We evaluate the effectiveness of swab-based PrP^Sc recovery from PPE (rubber boots) worn while hiking in CWD-positive and -negative geographic areas for active monitoring of environmental CWD prion contamination. To assess swab-based PrP^Sc recovery efficiency, boots were spiked in laboratory experiments with CWD-positive and CWD-negative controls, then swabbed and tested using RT-QuIC. Following confirmation of PrP^Sc recovery, we performed field-testing, consisting of hiking in CWD-positive and -negative sites, with boot swabbing at three time points: pre-hiking, post-hiking, and post-decontamination. RT-QuIC results were analyzed using ANOVA followed by pairwise comparison. Results: Max point ratio, a fluorescent measure, and max slope were used to compare sample seeding. There was a significant difference between pre- and post-sampling at the positive site and between the negative and positive sites post-sampling with p<0.001 and p<0.0001, respectively. Discussion/Conclusion: We posit that RT-QuIC testing of boot swabs is a promising and accessible tool for environmental CWD surveillance in natural habitats and within cervid farming operations.