During FY2013: 1) We adapted RT-QuIC and eQuIC to various murine prions and evaluated how seeding activity depends on glycophosphatidylinositol (GPI) anchoring and the abundance of amyloid plaques and protease-resistant PrPSc (PrPRes). Scrapie brain dilutions up to 10-8 and 10-13 were detected by RT-QuIC and eQuIC, respectively. Comparisons of scrapie-affected wild-type mice and transgenic mice expressing GPI anchorless PrP showed that, although similar concentrations of seeding activity accumulated in brain, the heavily amyloid-laden anchorless mouse tissue seeded more rapid reactions. Next we compared seeding activities in the brains of mice with similar infectivity titers, but widely divergent PrPRes levels, and found that the former were similar. Thus, in this comparison, RT-QuIC seeding activity correlated more closely with infectivity than with PrPRes levels. We also found that eQuIC, which incorporates a PrPSc immunoprecipitation step, detected seeding activity in plasma from wild-type and anchorless PrP transgenic mice inoculated with 22L, 79A and/or RML scrapie strains. Overall, we conclude that these new mouse-adapted prion seeding assays detect diverse types of PrPSc. 2) We adapted the RT-QuIC assay to the detection of prion seeding activity in the saliva of deer infected with chronic wasting disease. 3) We have also collaborated with neurologists to begin evaluating the utility of our previously developed RT-QuIC assay for the detection and diagnosis of human sporadic Creutzfeldt-Jakob disease using specimens derived from blood or olfactory mucosa brushings. The data so far have shown that RT-QuIC sensitively detects prion seeding activity in nasal brushings from a wide variety of human CJD cases. Results from the blood testing are pending. We are continuing to test additional samples that are being collected by our collaborators. Conclusions as to the diagnostic utility of these tests must await further testing.

Project Start
Project End
Budget Start
Budget End
Support Year
5
Fiscal Year
2013
Total Cost
$656,042
Indirect Cost
City
State
Country
Zip Code
Orrú, Christina D; Hughson, Andrew G; Groveman, Bradley R et al. (2016) Factors That Improve RT-QuIC Detection of Prion Seeding Activity. Viruses 8:
Zanusso, Gianluigi; Monaco, Salvatore; Pocchiari, Maurizio et al. (2016) Advanced tests for early and accurate diagnosis of Creutzfeldt-Jakob disease. Nat Rev Neurol 12:325-33
Haley, Nicholas J; Siepker, Chris; Hoon-Hanks, Laura L et al. (2016) Seeded Amplification of Chronic Wasting Disease Prions in Nasal Brushings and Recto-anal Mucosa-Associated Lymphoid Tissues from Elk by Real-Time Quaking-Induced Conversion. J Clin Microbiol 54:1117-26
Masujin, Kentaro; Orrú, Christina D; Miyazawa, Kohtaro et al. (2016) Detection of Atypical H-Type Bovine Spongiform Encephalopathy and Discrimination of Bovine Prion Strains by Real-Time Quaking-Induced Conversion. J Clin Microbiol 54:676-86
Dassanayake, Rohana P; Orrú, Christina D; Hughson, Andrew G et al. (2016) Sensitive and specific detection of classical scrapie prions in the brains of goats by real-time quaking-induced conversion. J Gen Virol 97:803-12
Chesebro, Bruce; Striebel, James; Rangel, Alejandra et al. (2015) Early Generation of New PrPSc on Blood Vessels after Brain Microinjection of Scrapie in Mice. MBio 6:e01419-15
Orrú, Christina D; Favole, Alessandra; Corona, Cristiano et al. (2015) Detection and discrimination of classical and atypical L-type bovine spongiform encephalopathy by real-time quaking-induced conversion. J Clin Microbiol 53:1115-20
Orrú, Christina D; Groveman, Bradley R; Raymond, Lynne D et al. (2015) Bank Vole Prion Protein As an Apparently Universal Substrate for RT-QuIC-Based Detection and Discrimination of Prion Strains. PLoS Pathog 11:e1004983
Orrú, Christina D; Groveman, Bradley R; Hughson, Andrew G et al. (2015) Rapid and sensitive RT-QuIC detection of human Creutzfeldt-Jakob disease using cerebrospinal fluid. MBio 6:
Zanusso, Gianluigi; Bongianni, Matilde; Caughey, Byron (2014) A test for Creutzfeldt-Jakob disease using nasal brushings. N Engl J Med 371:1842-3

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