Longitudinal detection of prion shedding in nasal secretions of CWD-infected white-tailed deer

Abstract

Chronic wasting disease (CWD) is an emergent prion disease spreading in cervid populations in North America, South Korea and Scandinavia. Rapid detection of CWD prions shed by live animals using minimally invasive methods remains an important need. Previous studies in deer, elk and hamsters have demonstrated prion replication in the nasal olfactory mucosa, yet the temporal profile of CWD prion shedding in nasal secretions has not been well characterized. Here we report nasal prion shedding in 18 deer orally exposed to low doses of CWD prions and monitored longitudinally by several parameters. Serially collected nasal swabs were assayed for CWD prion seeding activity using iron oxide magnetic extraction and real-time quaking-induced conversion (IOME RT-QuIC). These findings were correlated with the results from longitudinal tonsil biopsies, terminal tissues and PRNP genotype. We detected nasal prion shedding 3-16 months after the first positive tonsil biopsy in ten of the 18 deer; detectable shedding persisted thereafter in nine of the ten animals. Surprisingly, nasal swabs were negative in eight deer, even though all were CWD-infected as determined by tonsil biopsies and terminal tissue assays. Nasal shedding was detected more often in deer that were homozygous for glycine at codon 96, and those that were near or demonstrating symptoms of clinical disease shed earlier and more frequently, irrespective of prion exposure dose. The results of this study demonstrate nasal shedding of CWD prions that can be detected using minimally invasive nasal swab sampling and RT-QuIC analysis.

Keywords: CWD; IOME; RT-QuIC; longitudinal secretion; nasal swabs.

Fig. 1.

White-tail deer brain sectioning for post-mortem processing. The caudal aspect of the frontal lobe (section 1) and obex (section 8) were analysed by RT-QuIC. Prion positivity has been shown to progress from caudal to rostral across the brain. Progression of CWD was characterized from tonsil, to S1 brain section/olfactory bulb, and to nasal swab secretions collected by cytology brush.

Fig. 2.

(a) Longitudinal timelines of subsections of 10 deer inoculated with 10 mg, 1 mg or 300 ng brain or saliva material. Terminal and longitudinal nasal swab samples were analysed by RT-QuIC (positive nasal swab=red tick mark, negative nasal swab=yellow tick mark) and compared to the first IHC(+) tonsil biopsy (blue tick mark). (b) Positively inoculated deer (n=8) with IHC(+) tonsil biopsy (range: 12–24 MPI) and negative in all nasal swab samples (n=48). Negative controls (n=2) remained negative in all samples (n=10).