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United States Department of Agriculture

Agricultural Research Service

Research Project: TRANSMISSION, DIFFERENTIATION, AND PATHOBIOLOGY OF TRANSMISSIBLE SPONGIFORM ENCEPHALOPATHIES

Location: Virus and Prion Research

Title: Use of bovine recombinant prion protein and real-time quaking-induced conversion to detect transmissible mink encephalopathy prions and discriminate classical and atypical L- and H-type bovine spongiform encephalopathy

Author
item Hwang, Soyoun
item Greenlee, Justin
item Nicholson, Eric

Submitted to: PLoS One
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 2/3/2017
Publication Date: 2/22/2017
Citation: Hwang, S., Greenlee, J.J., Nicholson, E.M. 2017. Use of bovine recombinant prion protein and real-time quaking-induced conversion to detect cattle transmissible mink encephalopathy prions and discriminate classical and atypical L- and H-type bovine spongiform encephalopathy. PLoS One. 12(2):e0172391.

Interpretive Summary: The prion protein (PrP) is normal protein that can change its shape into an abnormal protein that is the causative agent of transmissible spongiform encephalopathies, a disease that affects the central nervous system. The disease process involves conversion of the normal cellular PrP to a pathogenic misfolded conformation. This conversion process can be conducted in the lab using a misfolding amplification process known as real-time quaking induced conversion (RT-QuIC). RT-QuIC uses recombinant prion protein to detect minute amounts of the abnormal infectious form of the prion protein. Although RT-QuIC has been successfully used to detect pathogenic PrP with various substrates including hamster, human, sheep, bank vole, and chimeric form of prion protein, bovine prion protein has not been proved for its practical uses for RT-QuIC. We evaluated whether prions from transmissible mink encephalopathy (TME) and bovine spongiform encephalopathy (BSE) infected cattle can be detected with recombinant bovine prion proteins using RT-QuIC, and optimized the reaction conditions to specifically detect infectious TME over negative control and to discriminate different forms of BSE. We also found that BSE associated E211K mutant protein can be an effective substrate for the detection of TME in cattle and that wild type prion protein appears most practical to discriminate the different types of BSEs.

Technical Abstract: Prions are amyloid-forming proteins that cause transmissible spongiform encephalopathies through a process involving conversion from normal cellular prion protein to pathogenic misfolded conformation. This conversion has been used for in vitro assays including serial protein misfolding amplification and real-time quaking induced conversion (RT-QuIC). RT-QuIC can be used for the detection of prions in a variety of biological tissues from humans and animals. Extensive work has been done to demonstrate that RT-QuIC is a rapid, specific, and highly sensitive prion detection assay. RT-QuIC uses recombinant prion protein to detect minute amount of the abnormal infectious form of the prion protein. Although RT-QuIC has been successfully used to detect abnormal prion protein (PrPSc) with various substrates including hamster, human, sheep, bank vole and chimeric form of prion protein, bovine prion protein has not been proved for its practical uses for RT-QuIC. Furthermore, recombinant bovine prion protein has not been used to detect transmissible mink encephalopathy (TME) or differentiate types of bovine spongiform encephalopathy (BSE) in samples from cattle. We evaluated whether prions from TME and BSE infected cattle can be detected with recombinant bovine prion proteins using RT-QuIC, and optimized the reaction conditions to specifically detect infectious TME over negative control and to discriminate classical and atypical BSE. We also found that disease associated E211K mutant protein can be an effective substrate for the detection of TME in cattle and that wild type prion protein appears most practical to discriminate the different types of BSEs.

Last Modified: 08/18/2017
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