The only method available to measure prion infectivity is by bioassay in an appropriate experimental animal, usually mice. We have developed an alternative prion bioassay whereby mice are replaced by the less sentient host Drosophila. To do so, we have generated PrP transgenic Drosophila and have shown that these flies are susceptible to mammalian prion infection, evidenced by a transmissible neurotoxic phenotype associated with accumulation of PK-resistant PrPSc. In this Skills and Knowledge Transfer to the APHA, we will validate the sensitivity of bovine PrP transgenic Drosophila for their bioassay for classical and atypical BSE prion infectivity. Prion inocula will consist of brain material from terminal clinical cases of cattle infected with classical BSE, or H-type or L-type atypical BSE. Prion-free bovine brain tissue will be used as control material. Prion inoculation of Drosophila will be at the larval stage while assessment of their phenotypic response by locomotor assay, detection of PrPSc by in vitro PMCA and IHC analysis of fly brains will be performed on adult flies. Specifically, we will: (a) Inoculate bovine PrP transgenic Drosophila with dilutions of either classical BSE- or atypical BSE-infected bovine brain homogenate to determine the sensitivity of these flies to bovine prions. The sensitivity of our Drosophila bioassay will be compared with existing data for these inocula obtained from titration in mice; (b) Detect PrPSc in BSE-exposed PrP transgenic Drosophila by in vitro PMCA & IHC. This will confirm the specificity of our novel fly-based prion bioassay; (c) Instruct the APHA on how to maintain and prion infect PrP transgenic Drosophila in an appropriate fly laboratory.
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