Prion Seeding Activity in DNA Extractions: Implications for Laboratory Biosafety

Gresch, S.; Morrill, T.; Ellis-Cramer, M.; Arifin, M.; Frank, L. E.; Bartz, J. C.; Schwabenlander, M.; Wolf, T. M.; Mitchell, G. B.; Guan, J.; Larsen, P. A.

doi:10.1101/2025.09.15.676309

bioRxiv · 10.1101/2025.09.15.676309

Prion Seeding Activity in DNA Extractions: Implications for Laboratory Biosafety

Gresch, S.·Morrill, T.·Ellis-Cramer, M.·Arifin, M.·Frank, L. E.·Bartz, J. C.·Schwabenlander, M.·Wolf, T. M.·Mitchell, G. B.·Guan, J.·Larsen, P. A.

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Abstract

Infectious prions (PrPSc) 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 PrPSc, 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). PrPSc 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 ({kappa}) = 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 PrPSc, enabling its carry-over into DNA eluates. Until infectivity is resolved by animal bioassay, DNA from PrPSc-positive tissues should be managed under biosafety protocols appropriate for the originating prion disease, with appropriate decontamination and containment procedures.

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Gresch, S., Morrill, T., Ellis-Cramer, M., Arifin, M., Frank, L. E., Bartz, J. C., Schwabenlander, M., Wolf, T. M., Mitchell, G. B., Guan, J., Larsen, P. A.. 2025-09-17. Prion Seeding Activity in DNA Extractions: Implications for Laboratory Biosafety. https://doi.org/10.1101/2025.09.15.676309

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