Tuesday, June 19, 2012

Experimental Oral Transmission of Chronic Wasting Disease to Reindeer (Rangifer tarandus tarandus)

Experimental Oral Transmission of Chronic Wasting Disease to Reindeer (Rangifer tarandus tarandus)

Gordon B. Mitchell1, Christina J. Sigurdson2,3, Katherine I. O’Rourke4, James Algire1, Noel P. Harrington1, Ines Walther1, Terry R. Spraker5, Aru Balachandran1*

1 National and OIE Reference Laboratory for Scrapie and CWD, Canadian Food Inspection Agency, Ottawa Laboratory – Fallowfield, Ottawa, Ontario, Canada,

2 Departments of Pathology and Medicine, University of California, San Diego, La Jolla, California, United States of America, 3 Department of Pathology, Microbiology and Immunology, University of California, Davis, California, United States of America, 4 Animal Disease Research Unit, Agricultural Research Service, United States Department of Agriculture, Pullman, Washington, United States of America, 5 Veterinary Diagnostic Laboratory, Colorado State University, Fort Collins, Colorado, United States of America


Chronic wasting disease (CWD), a transmissible spongiform encephalopathy of cervids, remains prevalent in North American elk, white-tailed deer and mule deer. A natural case of CWD in reindeer (Rangifer tarandus tarandus) has not been reported despite potential habitat overlap with CWD-infected deer or elk herds. This study investigates the experimental transmission of CWD from elk or white-tailed deer to reindeer by the oral route of inoculation. Ante-mortem testing of the three reindeer exposed to CWD from white-tailed deer identified the accumulation of pathological PrP (PrPCWD) in the recto-anal mucosa associated lymphoid tissue (RAMALT) of two reindeer at 13.4 months post-inoculation. Terminal CWD occurred in the two RAMALT-positive reindeer at 18.5 and 20 months post-inoculation while one other reindeer in the white-tailed deer CWD inoculum group and none of the 3 reindeer exposed to elk CWD developed disease. Tissue distribution analysis of PrPCWD in CWD-affected reindeer revealed widespread deposition in central and peripheral nervous systems, lymphoreticular tissues, the gastrointestinal tract, neuroendocrine tissues and cardiac muscle. Analysis of prion protein gene (PRNP) sequences in the 6 reindeer identified polymorphisms at residues 2 (V/M), 129 (G/S), 138 (S/N) and 169 (V/M). These findings demonstrate that (i) a sub-population of reindeer are susceptible to CWD by oral inoculation implicating the potential for transmission to other Rangifer species, and (ii) certain reindeer PRNP polymorphisms may be protective against CWD infection.


The importance of Rangifer species to the culture of aboriginal peoples cannot be underestimated with many components of hunted animals being consumed as food. Although relatively limited in comparison to elk and deer industries in North America, reindeer and caribou farming does occur, producing consumables such as meat, hides and antler velvet. The human health risks of consuming meat or other products derived from CWD-infected animals remain uncertain, although epidemiological evidence indicates transmission has not yet occurred [30,31,32] and transgenic mouse studies suggest the risk is remote in humans expressing common PRNP sequences [33,34,35]. The finding that CWD can be transmitted to squirrel monkeys by intracranial inoculation [36] raises concern for human transmissibility, however a study in macaques failed to demonstrate transmission after 70 months [37]. Since prion strains may undergo changes in transmission characteristics following passage through different species and strain selection pressures can be exerted by host genetic factors during passage within a species [24,38,39,40], caution is warranted when predicting the risks of CWD transmission from reindeer to other species.

This is the first evidence of CWD transmission to the sub-species Rangifer tarandus tarandus, implicating the potential for transmission to others in this genus. Current diagnostic tests, including antemortem RAMALT testing, appear capable of detecting CWD in Rangifer species and increased surveillance would be required to determine if natural transmission has indeed occurred. Additional studies are ongoing to chart the distribution of infectivity during the course of disease and determine the influence of PRNP polymorphisms in disease susceptibility.

Citation: Mitchell GB, Sigurdson CJ, O’Rourke KI, Algire J, Harrington NP, et al. (2012) Experimental Oral Transmission of Chronic Wasting Disease to Reindeer (Rangifer tarandus tarandus). PLoS ONE 7(6): e39055. doi:10.1371/journal.pone.0039055

Editor: Anthony E. Kincaid, Creighton University, United States of America

Received May 4, 2012; Accepted May 15, 2012; Published June 18, 2012

This is an open-access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 public domain dedication.

Funding: This work was supported by the Canadian Food Inspection Agency and Agriculture and Agri-Food Canada under C00233, the United States

Department of Agriculture Agricultural Research Service under CRIS 5348-32000-026-00-D and the United States National Institutes of Health under NS069566 and U54AI0359. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

Competing Interests: The authors have declared that no competing interests exist.

* E-mail: Aru.Balachandran@inspection.gc.ca


natural cases of CWD in eight Sika deer (Cervus nippon) and five Sika/red deer crossbreeds Korea and Experimental oral transmission to red deer (Cervus elaphus elaphus)


A Quarterly Newsletter from the Southeastern Cooperative Wildlife Disease Study College of Veterinary Medicine The University of Georgia Athens, Georgia 30602

Volume 27 January 2012 Number 4

Red deer susceptibility to CWD via oral inoculation was demonstrated in a study conducted by collaborators from the U.S. and Canada. Red deer developed clinical signs and had spongiform changes in the brain when euthanatized at 20 MPI. The CWD prion was detectable in neural and lymphoid tissues, endocrine organs, cardiac muscle, nasal mucosa, and other tissues. Although field cases of CWD in red deer have not been reported, results of this study indicate that it could occur, which is not surprising given that elk and red deer are subspecies of Cervus elaphus. The results of this study can be found in the Canadian Veterinary Journal 51: 169-178.

In addition, it was reported in May 2011 that natural cases of CWD were found in eight Sika deer (Cervus nippon) and five Sika/red deer crossbreeds during epidemiological investigations of CWD cases in captive elk in Korea.

May 2011 natural cases of CWD were found in eight Sika deer (Cervus nippon) and five Sika/red deer crossbreeds during epidemiological investigations of CWD cases in captive elk in Korea

Friday, May 13, 2011

Chronic Wasting Disease (CWD) outbreaks and surveillance program in the Republic of Korea Chronic Wasting Disease (CWD) outbreaks and surveillance program in the Republic of Korea

Hyun-Joo Sohn, Yoon-Hee Lee, Min-jeong Kim, Eun-Im Yun, Hyo-Jin Kim, Won-Yong Lee, Dong-Seob Tark, In- Soo Cho, Foreign Animal Disease Research Division, National Veterinary Research and Quarantine Service, Republic of Korea

Chronic wasting disease (CWD) has been recognized as an important prion disease in native North America deer and Rocky mountain elks. The disease is a unique member of the transmissible spongiform encephalopathies (TSEs), which naturally affects only a few species. CWD had been limited to USA and Canada until 2000. On 28 December 2000, information from the Canadian government showed that a total of 95 elk had been exported from farms with CWD to Korea. These consisted of 23 elk in 1994 originating from the so-called “source farm” in Canada, and 72 elk in 1997, which had been held in pre export quarantine at the “source farm”.Based on export information of CWD suspected elk from Canada to Korea, CWD surveillance program was initiated by the Ministry of Agriculture and Forestry (MAF) in 2001. All elks imported in 1997 were traced back, however elks imported in 1994 were impossible to identify. CWD control measures included stamping out of all animals in the affected farm, and thorough cleaning and disinfection of the premises. In addition, nationwide clinical surveillance of Korean native cervids, and improved measures to ensure reporting of CWD suspect cases were implemented. Total of 9 elks were found to be affected. CWD was designated as a notifiable disease under the Act for Prevention of Livestock Epidemics in 2002. Additional CWD cases - 12 elks and 2 elks - were diagnosed in 2004 and 2005. Since February of 2005, when slaughtered elks were found to be positive, all slaughtered cervid for human consumption at abattoirs were designated as target of the CWD surveillance program. Currently, CWD laboratory testing is only conducted by National Reference Laboratory on CWD, which is the Foreign Animal Disease Division (FADD) of National Veterinary Research and Quarantine Service (NVRQS). In July 2010, one out of 3 elks from Farm 1 which were slaughtered for the human consumption was confirmed as positive. Consequently, all cervid – 54 elks, 41 Sika deer and 5 Albino deer – were culled and one elk was found to be positive. Epidemiological investigations were conducted by Veterinary Epidemiology Division (VED) of NVRQS in collaboration with provincial veterinary services. Epidemiologically related farms were found as 3 farms and all cervid at these farms were culled and subjected to CWD diagnosis. Three elks and 5 crossbreeds (Red deer and Sika deer) were confirmed as positive at farm 2. All cervids at Farm 3 and Farm 4 – 15 elks and 47 elks – were culled and confirmed as negative. Further epidemiological investigations showed that these CWD outbreaks were linked to the importation of elks from Canada in 1994 based on circumstantial evidences. In December 2010, one elk was confirmed as positive at Farm 5. Consequently, all cervid – 3 elks, 11 Manchurian Sika deer and 20 Sika deer – were culled and one Manchurian Sika deer and seven Sika deer were found to be positive. This is the first report of CWD in these sub-species of deer. Epidemiological investigations found that the owner of the Farm 2 in CWD outbreaks in July 2010 had co-owned the Farm 5. In addition, it was newly revealed that one positive elk was introduced from Farm 6 of Jinju-si Gyeongsang Namdo. All cervid – 19 elks, 15 crossbreed (species unknown) and 64 Sika deer – of Farm 6 were culled, but all confirmed as negative. : Corresponding author: Dr. Hyun-Joo Sohn (+82-31-467-1867, E-mail: shonhj@korea.kr)

2011 Pre-congress Workshop: TSEs in animals and their environment 5

Friday, May 13, 2011

Chronic Wasting Disease (CWD) outbreaks and surveillance program in the Republic of Korea


Title: Experimental oral transmission of chronic wasting disease to red deer (Cervus elaphus elaphus): Early detection and late stage distribution of protease-resistant prion protein


Balachandran, A - CANADIAN FOOD INSPCTN AG Harrington, Noel - CANADIAN FOOD INSPCTN AG Algire, James - CANADIAN FOOD INSPCTN AG Souyrine, Andre - CANADIAN FOOD INSPCTN AG Spraker, Terry - COLORADO ST UNIV Jeffrey, Martin - Gonzalez, Lorenzo - Orourke, Katherine

Submitted to: Canadian Veterinary Journal Publication Type: Peer Reviewed Journal Publication Acceptance Date: December 1, 2008 Publication Date: March 11, 2010 Repository URL: http://ddr.nal.usda.gov/dspace/bitstream/10113/40677/1/IND44334511.pdf Citation: Balachandran, A., Harrington, N., Algire, J., Souyrine, A., Spraker, T., Jeffrey, M., Gonzalez, L., Orourke, K.I. 2010. Experimental oral transmission of chronic wasting disease to red deer (Cervus elaphus elaphus): Early detection and late stage distribution of protease-resistant prion protein. Canadian Veterinary Journal. Canadian Veterinary Journal. 51:169-178.

Interpretive Summary: Farmed cervids may be exposed to the prion disorder chronic wasting disease through contact with free ranging or farmed infected Rocky Mountain elk, white tailed deer, mule deer, or moose. This is the first report of experimental transmission of chronic wasting disease to red deer, an economically important agricultural commodity in parts of North America. Brain tissue from infected Rocky Mountain elk was administered by the oral route of red deer. Deer were examined at 18 months after infection for evidence of abnormal prion protein, the marker for the disease. The abnormal protein was found throughout the brain, spinal cord and lymphoid tissues, with variable distribution in other organ systems. This finding confirms the potential susceptibility of this species to disease under natural conditions and the reliability of the current testing format for identifying the abnormal protein in the tissues routinely collected in surveillance programs. The widespread distribution of the abnormal protein in red deer indicates the potential for shedding of the agent into the environment. Technical Abstract: Chronic wasting disease CWD is the transmissible spongiform encephalopathy or prion disease of wild and farmed cervid ruminants, including Rocky Mountain elk (Cervus elaphus nelsoni), white tailed deer (Odocoileus virginianus), mule deer (Odocoileus hemionus), or moose (Alces alces). Reliable data on the susceptibility of other farmed cervid species, the distribution of the abnormal prion protein marker in brain and lymphoid tissues collected in surveillance programs, and the role of prion genotype are necessary for design of control programs for CWD in farmed cervids. In this study, red deer (Cervus elaphus elaphus) were exposed to the prion agent by oral administration of brain homogenates from infected Rocky Mountain elk. Antemortem testing was performed at 7 months post infection and the deer were euthanized when clinical disease was observed at approximately 18 months after infection. The abnormal prion protein was assayed by immunohistochemistry, enzyme linked immunosorbent assay and western blot. Abnormal prion protein was found in the spinal cord, brainstem, cerebellum, midbrain, thalamus, and cerebrum in all 4 infected red deer. Most of the lymph nodes throughout the body were positive for abnormal prion proteins. Abnromal prion protein was observed in some additional peripheral tissues in some but not all of the deer. In particular, most areas of the gastrointestinal tract were positive for abnormal prions, although the salivary glands were rarely positive. This study demonstrates the potential for oral transmission of chronic wasting disease to red deer and confirms the usefulness of the current testing methods for post mortem diagnosis of the disease in this species.

Project Team Orourke, Katherine Knowles, Donald - Don White, Stephen Schneider, David

Publications Publications Related National Programs Animal Health (103)

Last Modified: 06/18/2012

Saturday, June 09, 2012

USDA Establishes a Herd Certification Program for Chronic Wasting Disease in the United States

Wednesday, June 13, 2012

TAHC Modifies Entry Requirements Effective Immediately for Cervids DUE TO CWD


Tuesday, June 05, 2012

Captive Deer Breeding Legislation Overwhelmingly Defeated During 2012 Legislative Session

Sunday, January 22, 2012

Chronic Wasting Disease CWD cervids interspecies transmission

Thursday, May 31, 2012

CHRONIC WASTING DISEASE CWD PRION2012 Aerosol, Inhalation transmission, Scrapie, cats, species barrier, burial, and more


2011 Annual Report

1a.Objectives (from AD-416) The project has 4 objectives: (1) Identify conformational and biological correlates of strain variation in the transmissible spongiform encephalopathies, (2) Identify genetic factors associated with horizontal transmission efficiency and susceptibility to the transmissible spongiform encephalopathies (3) Characterize the influence of genetics, strain, and multiple births on placental transmission of small ruminant TSEs; and (4) Devise a model system for assessing methods to reduce persistent environmental contaminations by prions.

1b.Approach (from AD-416) The current proposal addresses methods for characterizing and controlling classical and novel transmissible spongiform encephalopathies (TSEs)of domestic sheep and of farmed and free ranging deer and elk. The project includes discovery of unique identifiers for the North American TSE strain of small and wild ruminants and development of standardized methods suitable for use by the federal diagnostic reference laboratory and federally approved diagnostic laboratories. The genetic basis for relative transmission efficiency between and within the affected species, a critical element in design of control programs, will be is addressed through identification of haplotypes associated with naturally occurring disease. Allelic frequencies and disease associations are determined from tissue samples of naturally infected sheep, goats, deer, and elk. Genomic DNA is analyzed for the sequence of genomic regions including Prnp, Prnd, and Prnp' (when applicable). Samples of brain from infected animals are evaluated for relatively large changes in the apparent molecular weight of the proteinase K resistant core and for changes in the relative abundance of the variously glycosylated isoforms. The distribution and processing of disease associated PrP will be examined with a panel of monoclonal antibodies using single and double label immunohistochemistry assay. Samples with novel genotypes or prion protein isoforms will be evaluated in vivo when applicable. If novel strains are identified by these methods, standardized reagents and protocols for rapid strain typing of field samples will be developed and transferred to the national reference laboratory and the federally approved veterinary diagnostic laboratories. The role of the shed placenta and other environmental factors in TSE transmission and prion persistence will be examined.

3.Progress Report The project (1) addresses critical gaps in our understanding of the distribution of abnormal prion proteins in the tissues of sheep, goats, deer, elk, and small carnivores; (2) determines whether the current diagnostic methods are suitable for animals in each genotype; (3) identifies the limits of genetic resistance to the prion diseases; and (4) identifies environmental reservoirs of the infectious agent. This project provided reagents and methods for the first generation live animal test in sheep and participated in two international collaborations to extend those methods to diagnosis of bovine spongiform encephalopathy by assay of brain tissue. The methods were used to characterize atypical (Nor98) scrapie in Canada and the US. In association with Colorado State University, we described the distribution of the abnormal prion protein in the tissues of Rocky Mountain elk; this work contributed to our understanding of disease transmission and early diagnosis of prion disease in captive elk. In collaboration with the Canadian Food Inspection Agency, we reported the experimental transmission of prion disease from elk to red deer. The project provided information on the role of prion genotype in disease transmission in sheep and information on the effects of chimerism on genotype testing. In collaboration with the Canadian Food Inspection Agency, we described the prion genotypes of scrapie infected Canadian sheep and reported that the susceptibility patterns are similar to those observed in the US sheep. We provided baseline information on prion gene variation in US goats. In collaboration with the Veterinary Genetics Laboratory, we developed methods for determining relatedness in captive herds of white tailed deer with prion disease. In addition, we described the association between tissue mineral levels and prion disease in Rocky Mountain elk. Taken together, studies provided information being used by the regulatory agencies of the US and Canada in developing scrapie eradication programs. New project 5348-32000-030-00D to start 10-1-11.

4.Accomplishments 1. Association analysis of prion gene region sequences with chronic wasting disease in Rocky Mountain elk. Genetic resistance to ovine scrapie is a cornerstone of the current control program and a similar approach to chronic wasting disease (CWD) would be beneficial to the industry. In association with Colorado State University and USDA APHIS, ARS scientists in Pullman, WA, analyzed prion gene sequences in a sample of 559 captive and free ranging Rocky Mountain elk, of which 120 were considered positive for CWD. The previously reported mutation at codon 132 cut the odds of CWD by half but no association with CWD was found for any additional variants in the PRNP region (P > 0.05). This finding supports the current control programs, which consider that all elk exposed to CWD must be considered at risk of CWD.

2. Prion genotypes of scrapie-infected Canadian sheep. Selection for genetic resistance to scrapie is a critical element in the North American scrapie control programs although data supporting this approach are derived largely from European studies. The Canadian Food Inspection Agency and ARS collaborated on a retrospective study of the prion genotypes of a sample of 249 sheep with confirmed classical scrapie infection representing 98 flocks from 6 provinces and a further case-control analysis of 3 of these flocks comparing 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 a single haplotype (91.8%), findings similar to those reported by USDA APHIS for US scrapie-infected sheep. This finding supports the current control program that specifies removal or permanent quarantine of all scrapie-exposed sheep homozygous for that genotype.

3. The effects of autolysis on detection of abnormal prion proteins. Diagnosis and characterization of scrapie strains is made by a combination of biochemical tests for the abnormal prion protein PrP-Sc. Tissues submitted for analysis are often of poor quality, with varying levels of autolysis noted but the effect of autolysis on detection and characterization of the prion protein is not known. The Canadian Food Inspection Agency, in collaboration with ARS, completed a study of experimentally autolyzed tissues to address this knowledge gap. This study showed that the amount of PrP-Sc in lymphoid and central nervous system (CNS) tissues from elk and sheep decreased gradually as a result of autolysis, but PrP-Sc was still detectable after 5 and 15 d incubation at 37°C using either of two detection methods. The study demonstrated that the western blot technique can be used for diagnosis of scrapie using autolyzed samples.

4. Scrapie control program. The scrapie control program is based on the observations that ovine scrapie is transmitted effectively at the time of lambing and that transmission by unbred ewes or males rarely occurs. Similar data on transmission of scrapie in goats is lacking. In this study, ARS scientists in Pullman, WA examined the placentas of goats with naturally acquired scrapie and determined that the levels of abnormal prion protein are significantly lower than those observed in infected sheep, although the progeny of these goats were shown to develop the disease. This finding suggests that control programs specific for goats may be warranted, with attention to alternative modes of transmission.

5. Resistance of fallow deer to chronic wasting disease under field conditions. Chronic wasting disease in farmed cervids is managed on a species basis by a coordinated series of state and federal regulations. The regulations were written for the most commonly affected species (mule deer, white tailed deer and Rocky Mountain elk). The susceptibility of fallow deer, another commonly held species, to experimental infection by the intracerebral route has been established but the susceptibility of the species to natural challenge is not known. ARS contributed to a study conducted by USDA APHIS and the Colorado Division of Wildlife, in which fallow deer were held in pens with mule deer; all mule deer in the study died with evidence of CWD infection and none of the fallow deer showed either clinical disease or evidence of abnormal prions in any of the examined tissues. This finding suggests that the species is relatively resistant to natural transmission and may represent the only naturally resistant deer species reported to date.

Review Publications White, S.N., Spraker, T.R., Reynolds, J.O., Orourke, K.I. 2010. Association analysis of PRNP gene region with chronic wasting disease in Rocky Mountain elk. BMC Research Notes. 3:314.

Orourke, K.I., Zhuang, D., Truscott, T.C., Yan, H., Schneider, D.A. 2011. Scant PrP-Sc accumulation in the placentas of goats with naturally acquired scrapie. BioMed Central (BMC) Veterinary Research. 7(1):7.

Huang, H., Soutyrine, A., Rendulich, J., Orourke, K.I., Balachandran, A. 2011. Investigation of the effects of experimental autolysis on the detection of abnormal prion protein in lymphoid and central nervous system tissues from elk and sheep using the Western blotting method. Canadian Journal of Veterinary Research. 75(1)69-72.

Harrington, N., Orourke, K.I., Feng, Y., Rendulich, J., Difruscio, C., Balachandran, A. 2010. Prion genotypes of scrapie-infected Canadian sheep 1998-2008. Canadian Journal of Veterinary Research. 74(3):228-232.

Rhyan, J.C., Spraker, T.R., Mccollum, M., Nol, P., Wolfe, L., Miller, M.W., Davis, T., Creekmore, L., Orourke, K.I. 2009. Resistance of fallow deer (dama dama) to chronic wasting disease under natural exposure in a heavily contaminated environment. Journal of Wildlife Diseases. 47(3):739-744.

Project Team Orourke, Katherine Knowles, Donald - Don White, Stephen Schneider, David Project Annual Reports FY 2011 FY 2010 FY 2009 FY 2008 FY 2007

Publications Publications Related National Programs Animal Health (103)

Last Modified: 06/18/2012

Monday, June 18, 2012

natural cases of CWD in eight Sika deer (Cervus nippon) and five Sika/red deer crossbreeds captive Korea and Experimental oral transmission to red deer (Cervus elaphus elaphus)



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