Feasibility of infectious prion digestion using mild conditions and commercial subtilisin

Authors

John L. Pilon, National Wildlife Research CenterFollow
Paul B. Nash, National Wildlife Research CenterFollow
Terry Arver, Prion Tech
Don Hoglund, Prion Tech
Kurt C. Vercauteren, USDA-APHIS-Wildlife ServicesFollow

Document Type

Article

Date of this Version

2009

Citation

Journal of Virological Methods 161 (2009) 168–172; doi:10.1016/j.jviromet.2009.04.040

Abstract

Two serine protease enzymes, subtilisin 309 and subtilisin 309-v,were used to digest brain homogenates containing high levels of prion infectivity using mildly alkaline conditions to investigate prion decontamination methods. To establish that PrP^res infectivity was eliminated, we utilized the Rocky Mountain Laboratory (RML) mouse-adapted scrapie model system for bioassay. Only one digestion condition (subtilisin 309 at 138mAU/ml, 55^◦C and 14 h digestion time pH 7.9) was considered to be highly relevant statistically (P < 0.001) compared to control, with 52% of challenged mice surviving until the end of the study period. In contrast, treatment of PrP^res by autoclaving at 134^◦C or treatment with hypochlorite at a concentration of 20,000ppm completely protected mice from prionosis. Further, in vitro assays suggest that potential proteolytic based PrP^res decontamination methods must use high enzyme concentration, pH values >9.0, and elevated temperatures to be a safely efficacious, thereby limiting applicability on delicate surgical instruments and use in the environment.