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A new model for sensitive detection of zoonotic prions by PrP transgenic Drosophila.

ABSTRACT: Prions are transmissible protein pathogens most reliably detected by a bioassay in a suitable host, typically mice. However, the mouse bioassay is slow and cumbersome, and relatively insensitive to low titers of prion infectivity. Prions can be detected biochemically in vitro by the protein misfolding cyclic amplification (PMCA) technique, which amplifies disease-associated prion protein but does not detect bona fide prion infectivity. Here, we demonstrate that Drosophila transgenic for bovine prion protein (PrP) expression can serve as a model system for the detection of bovine prions significantly more efficiently than either the mouse prion bioassay or PMCA. Strikingly, bovine PrP transgenic Drosophila could detect bovine prion infectivity in the region of a 10^-12 dilution of classical bovine spongiform encephalopathy (BSE) inoculum, which is 10⁶-fold more sensitive than that achieved by the bovine PrP mouse bioassay. A similar level of sensitivity was observed in the detection of H-type and L-type atypical BSE and sheep-passaged BSE by bovine PrP transgenic Drosophila. Bioassays of bovine prions in Drosophila were performed within 7 weeks, whereas the mouse prion bioassay required at least a year to assess the same inoculum. In addition, bovine PrP transgenic Drosophila could detect classical BSE at a level 10⁵-fold lower than that achieved by PMCA. These data show that PrP transgenic Drosophila represent a new tractable prion bioassay for the efficient and sensitive detection of mammalian prions, including those of known zoonotic potential.

SUBMITTER: Thackray AM

PROVIDER: S-EPMC8350378 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:IntroductionMammalian prions are proteinaceous pathogens responsible for a broad range of fatal neurodegenerative diseases in humans and animals. These diseases can occur spontaneously, such as Creutzfeldt-Jakob disease (CJD) in humans, or be acquired or inherited. Prions are primarily formed of macromolecular assemblies of the disease-associated prion protein PrP(Sc), a misfolded isoform of the host-encoded prion protein PrP(C). Within defined host-species, prions can exist as conformational variants or strains. Based on both the M/V polymorphism at codon 129 of PrP and the electrophoretic signature of PrP(Sc) in the brain, sporadic CJD is classified in different subtypes, which may encode different strains. A transmission barrier, the mechanism of which remains unknown, limits prion cross-species propagation. To adapt to the new host, prions have the capacity to 'mutate' conformationally, leading to the emergence of a variant with new biological properties. Here, we transmitted experimentally one rare subtype of human CJD, designated cortical MM2 (129 MM with type 2 PrP(Sc)), to transgenic mice overexpressing either human or the VRQ allele of ovine PrP(C).ResultsIn marked contrast with the reported absence of transmission to knock-in mice expressing physiological levels of human PrP, this subtype transmitted faithfully to mice overexpressing human PrP, and exhibited unique strain features. Onto the ovine PrP sequence, the cortical MM2 subtype abruptly evolved on second passage, thereby allowing emergence of a pair of strain variants with distinct PrP(Sc) biochemical characteristics and differing tropism for the central and lymphoid tissues. These two strain components exhibited remarkably distinct replicative properties in cell-free amplification assay, allowing the 'physical' cloning of the minor, lymphotropic component, and subsequent isolation in ovine PrP mice and RK13 cells.ConclusionsHere, we provide in-depth assessment of the transmissibility and evolution of one rare subtype of sporadic CJD upon homologous and heterologous transmission. The notion that the environment or matrix where replication is occurring is key to the selection and preferential amplification of prion substrain components raises new questions on the determinants of prion replication within and between species. These data also further interrogate on the interplay between animal and human prions.

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A new model for sensitive detection of zoonotic prions by PrP transgenic Drosophila.

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