Abstract
Infectious prions (PrP(Sc)) are largely resistant to proteolytic digestion, including proteinase K (PK) digestion. While nucleic acid extracts are generally considered non-infectious from a classical microbiology context (i.e. free of intact bacteria and viruses), we investigated whether standard DNA purification methods co-purify PrP(Sc), posing an unrecognized biosafety risk. Commercial DNA extraction kits can eliminate conventional pathogens but are likely ineffective against PrP(Sc) due to resistance to kit reagents and enzymatic degradation. 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, one manual), while MNPRO tested two manual kits (filter- and 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 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 carry-over into DNA eluates. Until infectivity is resolved by animal bioassay, DNA from PrP(Sc)-positive tissues should be handled under biosafety protocols appropriate for the originating prion disease, with decontamination and containment procedures.