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United States Department of Agriculture

Agricultural Research Service

Related Topics

Research Project: TRANSMISSION, DIFFERENTIATION, AND PATHOBIOLOGY OF TRANSMISSIBLE SPONGIFORM ENCEPHALOPATHIES

Location: Virus and Prion Research

2016 Annual Report


1a. Objectives (from AD-416):
1. Investigate the pathobiology of atypical transmissible spongiform encephalopathies (TSEs) in natural hosts. A. Investigate the pathobiology of atypical scrapie. B. Investigate the pathobiology of atypical bovine spongiform encephalopathy (BSE). 2. Investigate the horizontal transmission of TSEs. A. Assess the horizontal transmission of sheep scrapie in the absence of lambing. B. Determine routes of transmission in chronic wasting disease (CWD) infected premises. C. Assess oral transmission of CWD in reindeer. 3. Investigate determinants of CWD persistence. A. Determine CWD host range using natural routes of transmission. B. Investigate the pathobiology of CWD.


1b. Approach (from AD-416):
The studies will focus on three animal transmissible spongiform encephalopathy (TSE) agents found in the United States: bovine spongiform encephalopathy (BSE); scrapie of sheep and goats; and chronic wasting disease (CWD) of deer, elk, and moose. The research will address sites of accumulation, routes of infection, environmental persistence, and ante mortem diagnostics with an emphasis on controlled conditions and natural routes of infection. Techniques used will include clinical exams, histopathology, immunohistochemistry and biochemical analysis of proteins. The enhanced knowledge gained from this work will help mitigate the potential for unrecognized epidemic expansions of these diseases in populations of animals that could either directly or indirectly affect food animals.


3. Progress Report:
This is the final report for the project 5030-32000-103-00D that started 10/01/2011 and terminates 09/30/2016. Research efforts directed toward meeting objective 1 of our project plan include work in previous years starting with the inoculation of animals for studies designed to address the pathobiology of atypical scrapie, atypical bovine spongiform encephalopathy (BSE), and a genetic version of BSE. Postmortem examination of the sheep inoculated with atypical scrapie has been initiated and laboratory analysis of the tissues is ongoing. Cattle inoculated with atypical BSE developed disease and evaluation of samples is currently underway. Cattle inoculated with a genetic form of BSE developed disease, and a manuscript reporting these results was published (2012). Comparisons of the stability of the disease agent from various BSE isolates have been completed and a study published (2013). Comprehensive laboratory comparisons of genetic BSE to atypical and classical BSE are ongoing. In research pertaining to objective 2, "Investigate the horizontal transmission of TSEs", we initiated a study to determine the risk of horizontal transmission of scrapie to neonatal or adult sheep in the absence of lambing of scrapie-infected ewes. At this time, scrapie-free ewes have lambed in the presence of sheep preclinically affected with scrapie. Both the lambs and ewes remain cohoused with the scrapie-affected sheep and are regularly examined for the onset of clinical signs suggestive of scrapie. Additional research demonstrated for the first time that chronic wasting disease is horizontally transmissible amongst reindeer, and a manuscript was published (2016) showing differences in susceptibility of sheep to the scrapie disease agent based on the prion genotype of the host animal, an important factor in horizontal transmission. Additional published research (2016) reports that the high degree of sequence conservation in the prion protein is not based on linkage to another gene and is specific to the prion protein. The fact that the observed sequence conservation of the prion gene is not due to linkage to another gene has implications for our understanding the normal functions of the prion protein.


4. Accomplishments
1. Different evolutionary pressures in domestic cattle influence the prion protein (PrP) and its nearest neighbor, a homolog of PrP known as doppel encoded by the PRND gene. The mammalian prion gene (PRNP) expresses a highly conserved protein termed the prion protein (PrP), best known as the causative agent of prion disease. Studies of PRNP knockout animals (animals that have had the PRNP gene deleted from their genome), including both mice and cattle, have failed to result in an overt change in animal behavior, health, or development. Some have used this to suggest that PrP is dispensable and that the high degree of sequence conservation is due to linkage to a nearby essential gene rather than due to high pressure to maintain the PrP sequence. In this work, ARS researchers in Ames, Iowa demonstrated the degree of sequence conservation in the nearest neighbor of PRNP, a gene known as PRND. We determined that in cattle PRND is not under any selective pressure and is not influencing PRNP nor is it being influenced in a manner similar to PRNP. This indicates an essential role for the protein PrP suggesting that the loss of prion function is simply not manifesting in the controlled environmental conditions used to raise the knockout animals. This has implications for understanding the normal function of the prion protein and for the use of PRNP knockout cattle in the production of prion free bovine products.

2. Reindeer are susceptible to chronic wasting disease (CWD) from various cervid sources and infected reindeer can transmit the disease to healthy reindeer. Chronic wasting disease (CWD) is a naturally-occurring, fatal prion disease of cervids. Reindeer are susceptible to CWD following experimental oral challenge. This study investigated the susceptibility of reindeer to intracranial inoculation of CWD from white-tailed deer, mule deer, or elk, and assessed for transmission to non-inoculated reindeer placed in direct (same pen) or indirect contact (adjacent pen but without nose-to-nose contact). Intracranially inoculated reindeer developed clinical disease from 21 months post-inoculation, and a specific disease associated protein was detected in 5 out of 6 reindeer that were in either direct or indirect contact. ARS researchers in Ames, Iowa demonstrated that reindeer are susceptible to CWD from various cervid sources and can transmit CWD to naïve reindeer through direct contact or indirectly through the environment. This has important regulatory implications for the transport of this species, especially following the recent discovery of CWD in a free-ranging reindeer of Norway.

3. Sheep prion protein genetics influences disease progression in sheep orally inoculated with scrapie. Sheep scrapie is a transmissible spongiform encephalopathy that can be transmitted through contact with affected animals resulting in significant economic losses in affected flocks. The prion protein gene (PRNP) profoundly influences the susceptibility of sheep to the scrapie agent and the tissue levels and distribution of the disease that is associated with the form of the prion protein (PrPSc) in affected sheep. ARS researchers in Ames, Iowa demonstrated the failure to detect PrPSc in nervous or lymphoid tissues of neonatal sheep of a specific PRNP genotype (ARQ/ARR) after oral inoculation with a U.S. scrapie isolate. Sheep of the ARQ/ARQ genotype receiving the same inoculum developed scrapie within 24 months. Lambs of the ARQ/ARR genotype that received the same inoculum by intracranial inoculation developed scrapie with a prolonged incubation period and with abnormal prion present within the central nervous system, but not peripheral lymphoid tissues. Results suggest that ARQ/ARR sheep are resistant to oral infection with the scrapie isolate used even during the neonatal period. This enhances the understanding of natural transmission of scrapie to sheep of specific genotypes in a production relevant setting including neonatal lambs.


5. Significant Activities that Support Special Target Populations:
None.


Review Publications
Greenlee, J.J., Smith, J.D., Hamir, A.N. 2016. Oral inoculation of neonatal Suffolk sheep with the agent of classical scrapie results in PrPSc accumulation in sheep with the PRNP ARQ/ARQ but not the ARQ/ARR genotype. Research in Veterinary Science. 105:188–191.

Haley, N.J., Siepker, C., Walter, W.D., Thomsen, B.V., Greenlee, J.J., Lehmkuhl, A.D., Richt, J.A. 2016. Antemortem detection of chronic wasting disease prions in nasal brush collections and rectal biopsy specimens from white-tailed deer by real time quaking-induced conversion. Journal of Clinical Microbiology. 54(4):1108-1116.

Moore, S.J., Smith, J.D., West Greenlee, M.H., Nicholson, E.M., Richt, J.A., Greenlee, J.J. 2016. Comparison of two US sheep scrapie isolates supports identification as separate strains. Veterinary Pathology. 53(6):1187-1196. doi: 10.1177/0300985816629712.

Vermette, M.S., Schleining, J.A., Greenlee, J.J., Smith, J.D. 2016. Genetic variation of the prion protein gene (PRNP) in alpaca (Vicugna pacos). Gene Reports. 4:213–217.

Brunelle, B.W., Hannen, A.M., Nicholson, E.M., Seabury, C.M. 2016. Disparate modes of evolution shaped modern prion (PRNP) and prion-related doppel (PRND) variation in domestic cattle. PLoS One. 11(5):e0155924. doi: 10.1371/journal.pone.0155924.

Haley, N.J., Siepker, C., Greenlee, J.J., Richt, J.A. 2016. Limited amplification of chronic wasting disease prions in the peripheral tissues of intracerebrally inoculated cattle. Journal of General Virology. 97:1720-1724.

West Greenlee, M.H., Lind, M., Kokemuller, R., Mammadova, N., Kondru, N., Manne, S., Smith, J., Kanthasamy, A., Greenlee, J. 2016. Temporal resolution of misfolded prion protein transport, accumulation, glial activation, and neuronal death in the retinas of mice inoculated with scrapie. American Journal of Pathology. 186(9):2302–2309.

Last Modified: 08/18/2017
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