This study analyzed sheep prion protein (PrP) genotypes of samples submitted from Ontario and other provinces of Canada to the Animal Health Laboratory at the University of Guelph, Guelph, Ontario, between 2005 and 2012. In Ontario, the proportion of scrapie-resistant sheep increased from 2005 to 2012 as evidenced by an increase in the ARR haplotype. When Canadian provinces (Alberta, Ontario, Quebec, and Nova Scotia) were compared from 2008 to 2012, a high proportion of scrapie-resistant sheep was found in all the provinces. The proportions of resistant sheep were lower in Alberta and Quebec than in Ontario and Nova Scotia. Alberta had higher proportions of susceptible sheep and a higher frequency of VRQ alleles, and Quebec had a higher frequency of the ARQ allele.

Final observations on experimental transmission of chronic wasting disease (CWD) from elk (Cervus elaphus nelsoni) and white-tailed deer (Odocoileus virginianus) to fallow deer (Dama dama) are reported herein. During the 5-year study, 13 fawns were inoculated intracerebrally with CWD-infected brain material from white-tailed deer (n = 7; Group A) or elk (n = 6; Group B), and 3 other fawns were kept as uninoculated controls (Group C). As described previously, 3 CWD-inoculated deer were euthanized at 7.6 mo post-inoculation (MPI). None revealed presence of abnormal prion protein (PrPd) in their tissues. At 24 (Group A) and 26 (Group B) MPI, 2 deer were necropsied. Both animals had a small focal accumulation of PrPd in their midbrains. Between 29 and 37 MPI, 3 other deer (all from Group A) were euthanized. The 5 remaining deer became sick and were euthanized between 51 and 60 MPI (1 from Group A and 4 from Group B). Microscopic lesions of spongiform encephalopathy (SE) were observed in only these 5 animals; however, PrPd was detected in tissues of the central nervous system by immunohistochemistry, Western blot, and by commercial rapid test in all animals that survived beyond 24 MPI. This study demonstrates that intracerebrally inoculated fallow deer not only amplify CWD prions, but also develop lesions of spongiform encephalopathy.

Chronic wasting disease (CWD) is a transmissible spongiform encephalopathy (TSE) of Rocky Mountain elk in North America. Recent studies suggest that tissue and blood mineral levels may be valuable in assessing TSE infection in sheep and cattle. The objectives of this study were to examine baseline levels of copper, manganese, magnesium, zinc, selenium, and molybdenum in the brains of Rocky Mountain elk with differing prion genotypes and to assess the association of mineral levels with CWD infection. Elk with leucine at prion position 132 had significantly lower magnesium levels than elk with 2 copies of methionine. Chronic wasting disease-positive elk had significantly lower magnesium than control elk. The incorporation of manganese levels in addition to magnesium significantly refined explanatory ability, even though manganese alone was not significantly associated with CWD. This study demonstrated that mineral analysis may provide an additional disease correlate for assessing CWD risk, particularly in conjunction with genotype.

This report describes the genetics of the prion protein gene (PRNP) at codons 136, 154, and 171 for sheep diagnosed with naturally acquired classical scrapie in Canada between 1998 and 2008. Genotyping analysis was performed on 249 sheep with confirmed classical scrapie infection representing 98 flocks from 6 provinces. A further case-control analysis of 3 of these flocks compared the genotypes between infected sheep (n = 72) and those of their healthy flockmates (n = 1990). The incidence of classical scrapie in the Canadian sheep population was highly associated with the ARQ haplotype (91.8%) and the ARQ/ARQ genotype (91.6%). In addition, the ARQ haplotype was found at significantly higher frequency in scrapie-infected sheep when compared with their healthy flockmates. Comparison with other published data suggests that the scrapie risk of PRNP genotypes differs between Canada and countries where the VRQ allele is associated with the highest susceptibility to infection.

Early epidemiological information indicated that bovine spongiform encephalopathy (BSE) originated from scrapie in sheep. The question arose if scrapie in North America would induce a BSE-like disease in cattle. Six years ago, we reported that brain tissue from sheep with scrapie caused a neurologic disease when injected directly into the brains of cattle, but the disease induced was different from BSE as it occurs in the United Kingdom and Europe. Here, we report that cattle fed raw brain or meat and bone meal and tallow prepared from sheep with scrapie remained normal for 8 years after exposure. This work indicates that cattle are highly resistant to North American scrapie by the oral route.

Reciprocal embryo transfers were made between scrapie-inoculated and scrapie-free sheep (Cheviot and Suffolk breeds) to measure scrapie transmission via the embryo (using offspring from embryos of scrapie-inoculated donors and scrapie-free recipients) and via the uterus (using offspring from embryos of scrapie-free donors and scrapie-inoculated recipients taken by cesarean section). Two control groups of offspring, 1 from scrapie-free parents (negative) and 1 from scrapie-inoculated parents (positive), also were included. All sheep were observed for clinical signs of scrapie until death or for a minimum of 60 months. Final diagnosis was made on the basis of histopathologic findings or results of mouse inoculation and/or proteinase-K-resistant protein analysis. Thirty to 61% of the scrapie-inoculated donor/recipient sheep within groups developed scrapie within 8 to 44 months after inoculation. None of the scrapie-free donor/recipients, including those gestating embryos from scrapie-inoculated donors, developed scrapie. Also, none of the offspring observed to larger than or equal to 24 months of age from reciprocal cross, via embryo (0/67), or via the uterus (0/25), or from the negative-control group (0/33) developed scrapie. Fifty-six of the offspring via embryo, 19 of these via the uterus, and 31 negative controls survived to larger than or equal to 60 months of age. Of the 21 sheep in the positive-control group, 2 (9.5%) developed scrapie, 1 at 31 months of age and 1 at 42 months of age. In the Cheviot offspring, the percentage of sheep carrying the short incubation allele ranged from 24 to 44% and the percentage in the Suffolk offspring ranged from 61 to 83%. These proportions indicate high degree of susceptibility to the disease. Results indicate that under the conditions of these experiments, scrapie was not transmitted to the offspring via the embryo or the uterus.

The duration of the incubation period for scrapie, a fatal transmissible neurodegenerative disorder of sheep and goats, is mainly determined by the Sip gene, which has 2 alleles (sA-susceptible and pA-resistant). A diagnostic test is not available to detect scrapie in live animals. We analyzed genomic DNA extracted from frozen sheep brains collected from Cheviot sheep of the United States that had been inoculated with the SSBP/1 scrapie inoculum. Digestion of the DNA with EcoRI or HindIII followed by the addition of a scrapie-associated fibril protein (PrP)-specific marker probe, yielded fragments of 6.8 (e1) and 4.0 (e3) kb, or 5.0 (h1) and 3.4 (h2) kb, respectively. Fragments e1 and h2 were associated with the histopathologic diagnosis of scrapie, and fragments e3 and h1 were associated with survival. A valine/alanine polymorphism within the PrP coding region that resulted in a BspHI site was further used to determine the genotype of these Cheviot sheep. Digestion of polymerase chain reaction fragments with BspHI resulted in an undigested fragment b- (0.840 kb), digested fragments b+ (0.460 and 0.380 kb), or both types of fragments. Survival time of b+/b+ homozygous sheep was significantly (P < 0.01) shorter (218 +/- 26.0 days) than survival time for b-/b- sheep (> 700 days after inoculation). Results indicated that b+ and b- are markers for the Sip sA and pA alleles, respectively. The intermediate duration of the incubation period for heterozygous sheep (b+/b-; 342.9 +/- 25.3 days) indicated that the Sip sA allele is expressed codominantly to the Sip pA allele.

Objective-To determine the usefulness of retina samples for detection of disease-associated prion protein by use of a commercially available enzyme immunoassay (EIA) intended for rapid identification of sheep and cattle with transmissible spongiform encephalopathies (TSEs). Samples-Retina, brainstem at the level of the obex, and retropharyngeal lymph node samples obtained from 15 TSE-inoculated sheep (scrapie [n = 13] or transmissible mink encephalopathy passaged through a bovid [2]); retina and brainstem samples obtained from 11 TSE-inoculated cattle (transmissible mink encephalopathy passaged through a bovid [7] or classical BSE [4]); and negative control tissue samples obtained from 2 sheep and 2 cattle that were not inoculated with TSEs. Procedures-Tissue samples were homogenized and analyzed for detection of abnormally folded disease-associated prion protein with a commercially available EIA and 2 confirmatory assays (western blot analysis or immunohistochemical analysis). Results-Retina sample EIA results were in agreement with results of brainstem sample EIA or confirmatory assay results for negative control animals and TSE-inoculated animals with clinical signs of disease. However, TSE-inoculated animals with positive confirmatory assay results that did not have clinical signs of disease had negative retina sample EIA results. Retina sample EIA results were in agreement with brainstem sample immunohistochemical results for 4 TSE-inoculated sheep with negative retropharyngeal lymph node EIA results. Conclusions and Clinical Relevance-Results of this study suggested that retina samples may be useful for rapid EIA screening of animals with neurologic signs to detect TSEs.

This report describes the genetics of the prion protein gene (PRNP) at codons 136, 154, and 171 for sheep diagnosed with naturally acquired classical scrapie in Canada between 1998 and 2008. Genotyping analysis was performed on 249 sheep with confirmed classical scrapie infection representing 98 flocks from 6 provinces. A further case-control analysis of 3 of these flocks compared the genotypes between infected sheep (n = 72) and those of their healthy flockmates (n = 1990). The incidence of classical scrapie in the Canadian sheep population was highly associated with the ARQ haplotype (91.8%) and the ARQ/ARQ genotype (91.6%). In addition, the ARQ haplotype was found at significantly higher frequency in scrapie-infected sheep when compared with their healthy flockmates. Comparison with other published data suggests that the scrapie risk of PRNP genotypes differs between Canada and countries where the VRQ allele is associated with the highest susceptibility to infection.

Susceptibility to chronic wasting disease (CWD) in cervid species has been associated with polymorphisms in the prion protein gene (PRNP). The single nucleotide polymorphisms (SNPs) were found in the PRNP of the Korean subspecies of Chinese water deer via analyses of the DNA sequences obtained from 34 individual deer. Two SNPs were detected at codons 77 and 100. One SNP at codon 77 encoding Glycine was determined to be a silent mutation but the other SNP detected at codon 100 induced an amino acid change, from Asparagine to Serine. The prion protein (PrP) amino acid sequence of the water deer showed 98.8-99.2% homology with those of American elk, white-tailed deer and mule deer. The PrP of the water deer contained amino acid residues closely related with CWD-susceptibility. This study is the first to describe genetic variations in the PRNP of the Korean subspecies of Chinese water deer.