Application of protein misfolding cyclic amplification to detection of prions in anaerobic digestate.
The exceptional physio-chemical resistance of prions to established decontamination procedures poses a challenge to assessing the suitability of applied inactivation methods. Prion detection is limited by the sensitivity level of Western blotting or by the cost and time factors of bioassays. In addition, prion detection assays can be limited by either the unique or complex nature of matrices associated with environmental samples. To investigate anaerobic digestion (AD) as a practical and economical approach for potential conversion of specified risk materials (SRM) into value added products (i.e., renewable energy), challenges associated with detection of prions in a complex matrix need to be overcome to determine potential inactivation. Protein misfolding cyclic amplification (PMCA) assay, with subsequent Western blot visualization, was used to detect prions within the AD matrix. Anaerobic digestate initially inhibited the PMCA reaction and/or Western blot detection. However, at concentrations of ≤1% of anaerobic digestate, 263K scrapie prions could be amplified and semi-quantitatively detected. Infectious 263K prions were also proven to be bioavailable in the presence of high concentrations of digestate (10-90%). Development of the PMCA application to digestate provides extremely valuable insight into the potential degradation and/or fate of prions in complex biological matrices without requiring expensive and time-consuming bioassays.
School of Environmental Sciences, University of Guelph, Ridgetown N0P 2C0, Canada. Electronic address: bgilroye@uoguelph.ca.
Brandon H
BH
Gilroyed
School of Public Health, University of Alberta, Edmonton T6G 2T4, Canada.
Shannon L
SL
Braithwaite
School of Public Health, University of Alberta, Edmonton T6G 2T4, Canada.
Luke M
LM
Price
Alberta Agriculture and Rural Development, Lethbridge T1J 4V6, Canada.
Tim
T
Reuter
Canadian Food Inspection Agency, Lethbridge T1H 6P7, Canada.
Stefanie
S
Czub
Canadian Food Inspection Agency, Lethbridge T1H 6P7, Canada.
Catherine
C
Graham
Canadian Food Inspection Agency, Ottawa K2H 8P9, ON, Canada.
Arumuga
A
Balachandran
Agriculture and Agri-Food Canada, Lethbridge T1J 4B1, Canada.
Tim A
TA
McAllister
School of Public Health, University of Alberta, Edmonton T6G 2T4, Canada; Department of Biological Sciences, University of Alberta, Edmonton T6G 2E9, Canada.
Miodrag
M
Belosevic
School of Public Health, University of Alberta, Edmonton T6G 2T4, Canada; Alberta Provincial Laboratory for Public Health, Edmonton T6G 2J2, Canada.
Norman F
NF
Neumann
D000693:Anaerobiosis; D000818:Animals; D015153:Blotting, Western; D001923:Brain Chemistry; D002623:Chemistry Techniques, Analytical; D006224:Cricetinae; D011328:Prions / Q000032:analysis*; D017510:Protein Folding
D011328:Prions
Anaerobic digestion; Hamster scrapie; PMCA; Specified risk materials; Transmissible spongiform encephalopathies