White-tailed deer are susceptible to the agent of sheep scrapie by intracerebral inoculation

Abstract

Interspecies transmission studies afford the opportunity to better understand the potential host range and origins of prion diseases. The purpose of this experiment was to determine susceptibility of white-tailed deer to the agent of scrapie after intracerebral inoculation and to compare clinical signs and lesions to those reported for chronic wasting disease (CWD). Deer (n = 5) were inoculated with 1 mL of a 10% (wt/vol) brain homogenate derived from a sheep clinically affected with scrapie. A non-inoculated deer was maintained as a negative control. Deer were observed daily for clinical signs of disease and euthanized and necropsied when unequivocal signs of scrapie were noted. One animal died 7 months post inoculation (pi) due to intercurrent disease. Examinations of brain tissue for the presence of the disease-associated abnormal prion protein (PrP(Sc)) by western blot (WB) and immunohistochemistry (IHC) were negative whereas IHC of lymphoid tissues was positive. Deer necropsied at 15-22 months pi were positive for scrapie by IHC and WB. Deer necropsied after 20 months pi had clinical signs of depression and progressive weight loss. Tissues with PrP(Sc) immunoreactivity included brain (at levels of cerebrum, hippocampus, colliculus, cerebellum, and brainstem), trigeminal ganglion, neurohypophysis, retina, spinal cord, and various lymphoid tissues including tonsil, retropharyngeal and mesenteric lymph nodes, Peyer's patches, and spleen. This work demonstrates for the first time that white-tailed deer are susceptible to sheep scrapie by intracerebral inoculation. To further test the susceptibility of white-tailed deer to scrapie these experiments will be repeated with a more natural route of inoculation.

Figure 1

Immunoreactivity for PrP^Sc occurs in lymphoid tissue prior to detection in brain. Retropharyngeal lymph node from deer 1 was immunoreactive for PrP^Sc as early as 6.8 months post-inoculation (pi). Immunoreactivity (red) is intense in the germinal centers of lymphoid follicles. PrP^Sc immunohistochemistry with monoclonal antibodies F99/97.6.1 and hematoxylin counterstain. Bar = 100 μm.

Figure 2

Brain is immunoreactive for PrP^Sc at 15.4 months pi. PrP^Sc immunoreactivity is present in brainstem of deer 2 just caudal to the obex. IR is most intense adjacent to the central canal rather than in the brainstem nuclei. PrP^Sc immunohistochemistry with monoclonal antibodies F99/97.6.1 and hematoxylin counterstain. Bar = 10 μm.

Figure 3

PrP^Sc immunoreactivity is widespread throughout the cerebral cortex in deer with clinical signs of scrapie. Within the cortex, course granular staining is abundant throughout the neuropil and frequently coalesces in a glial pattern (deer 3). PrP^Sc immunohistochemistry with monoclonal antibodies F99/97.6.1 and hematoxylin counterstain. Bar = 10 μm.

Figure 4

Representative western blot of deer with scrapie. This blot compares brain homogenates from the sheep scrapie inoculum (lane 2), a white-tailed deer with CWD [16] (lane 3), and white-tailed deer with scrapie (deer 5, lane 4-5). All brain homogenates are positive using monoclonal antibodies P4 and 6H4 and tissues with a characteristic 3-band pattern representing diglycosylated, monoglycosylated, and unglycosylated forms of PrP^Sc. Homogenates derived from cerebrum (lane 5) have a lower migration of the unglycosylated band than those derived from brainstem at the level of obex (lanes 4). Lanes were loaded as follows: sheep scrapie inoculums = 0.1 mg, white-tailed deer CWD = 0.175 mg; deer 5 obex = 1 mg, deer 5 cerebrum = 1 mg, Marker = molecular weight marker.

Figure 5

Deer with scrapie exhibit spongiform change (SE). SE was severe in the obex of deer clinically-affected with scrapie (deer 3). Parasympathetic nucleus of the vagus. Hematoxylin and eosin stain. Bar = 100 μm.

Figure 6

Gray matter vacuolation profiles for white-tailed deer inoculated intracerebrally with scrapie or CWD. Vacuolation scores were higher in the cerebral cortex and colliculus and lower in the hippocampus in CWD-affected deer. The converse was present in scrapie-affected deer. Abbreviations: IC, intracerebral; CWD, chronic wasting disease; DMNV, dorsal motor nucleus of the vagus nerve.