Prion seeding activity in DNA extractions: implications for laboratory biosafety

ABSTRACT Infectious prions (PrP Sc ) are largely resistant to proteolytic digestion, including proteinase K digestion. While nucleic acid extracts are generally considered non-infectious, we investigated whether standard DNA purification methods can co-purify PrP Sc , posing an unrecognized biosafety risk. Two laboratories, the University of Minnesota Center for Prion Research and Outreach (MNPRO) and the Canadian Food Inspection Agency (CFIA), independently tested filter-based and magnetic bead-based DNA extraction kits using tissues from chronic wasting disease (CWD)-positive and -negative white-tailed deer (WTD; Odocoileus virginianus), as well as prion-infected and control Syrian hamster (Mesocricetus auratus) brains. CFIA used two filter-based kits (one automated and one manual), while MNPRO tested two manual kits (one filter-based and one magnetic bead-based). PrP Sc seeding activity was measured in extracted DNA and source tissues using real-time quaking-induced conversion (RT-QuIC). MNPRO found substantial to almost perfect agreement (kappa (κ) = 0.789 - 0.816) between RT-QuIC seeding activity of DNA eluates from both extraction methods and that of the source WTD tissue homogenate. CFIA optimized RT-QuIC to a 30-hour runtime, achieving 74% sensitivity and 94% specificity in 88 archived WTD DNA samples. Both laboratories concluded that commercial DNA extraction kits do not eliminate PrP Sc , enabling its carry-over into DNA eluates. Until infectivity is resolved by animal bioassay, DNA from PrP Sc -positive tissues should be managed under biosafety protocols appropriate for the originating prion disease, with appropriate decontamination and containment procedures.