Dehydration of prions on environmentally relevant surfaces protects them from inactivation by freezing and thawing

Chronic wasting disease (CWD) is an emerging prion disease in North America. Recent identification of CWD in wild cervids from Norway raises the concern of the spread of CWD in Europe. CWD infectivity can enter the environment through live animal excreta and carcasses where it can bind to soil. Well-characterized hamster prion strains and CWD field isolates in unadsorbed or soil-adsorbed forms that were either hydrated or dehydrated were subjected to repeated rounds of freezing and thawing. We found that 500 cycles of repeated freezing and thawing of hydrated samples significantly decreased the abundance of PrP^Sc and reduced protein misfolding cyclic amplification (PMCA) seeding activity that could be rescued by binding to soil. Importantly, dehydration prior to freezing and thawing treatment largely protected PrP^Sc from degradation, and the samples maintained PMCA seeding activity. We hypothesize that redistribution of water molecules during the freezing and thawing process alters the stability of PrP^Sc aggregates. Overall, these results have significant implications for the assessment of prion persistence in the environment.