Progress 01/28/02 to 08/26/04
Outputs
1. What major problem or issue is being resolved and how are you resolving it (summarize project aims and objectives)? How serious is the problem? What does it matter? This project deals with Transmissible Spongiform Encephalopathies (TSE), which are fatal degenerative diseases of the central nervous system that can affect several animal species, including humans. The causal agent is believed to be a natural tissue protein, the prion protein, that has assumed an unnatural form. Because the altered protein is resistant to enzyme degradation, it accumulates in nervous tissue and the resulting dysfunction ultimately leads to death. The specific TSEs being investigated in this project are scrapie in sheep, transmissible mink encephalopathy (TME), bovine spongiform encephalopathy (BSE) in cattle, and chronic wasting disease (CWD) in deer and elk. The major concern about these diseases is that BSE has been shown to cross the species barrier to cause a unique TSE in human
beings. Although there has not been a similar demonstration that scrapie, TMS, or CWD could present any risk to human health, the BSE experience has raised many questions about the potential hazard these TSEs present for transmission to other animal species, especially domesticated livestock and wildlife. The current research is focused on direct experimental challenge of the species barrier effect by animal inoculation. These results are then compared to results obtained with a variety of laboratory procedures to determine if they can be used as predictive models for future risk assessments. These studies also provide information about the clinical and pathological disease characteristics that can be expected if a TSE crosses the species barrier, thus enabling animal health specialists to recognize such situations should they occur. Additional transmission studies in the natural host will focus on determining the modes of transmission and disease development in scrapie and CWD so
that appropriate intervention strategies can be devised that will control the spread of these diseases. Because of the risk for BSE transmission to human beings, the presence of scrapie, TME, CWD, and possibly BSE in the United States presents a potential liability to the U.S. livestock and hunting industries, because the safety of animals and animal products intended for domestic consumption and international trade may be questioned. Efforts are being made by both federal and state regulatory agencies to eradicate scrapie and CWD and to determine the prevalence, if any, of BSE. The effectiveness of these programs will depend heavily on having accurate information about the nature of these diseases, not only in the original hosts, but also in other species that may be affected. 2. List the milestones (indicators of progress) from your Project Plan. Animal experiments: Because of the long-term nature of most TSE experiments in animals, especially those involving cross-species
transmission, most of the studies outlined in this project will not be completed within 30 months. Therefore, the following time line primarily presents expectations for when experiments will be initiated. 2003 Cattle inoculated with white-tailed deer CWD Fallow and white-tailed deer inoculated with CWD Cattle inoculated with TME Sheep inoculated with AV136QR171 and Idaho scrapie isolates 2004 Cattle inoculated with elk CWD Raccoons inoculated with TME, scrapie, and CWD isolates Mice inoculated for strain typing of 10 TSE isolates Swine inoculated with scrapie and CWD Reindeer inoculated with CWD White-tailed deer inoculated with scrapie 2005 Study to assess scrapie and CWD amplification in market age swine completed Mice inoculated for strain typing of 15 TSE isolates Laboratory studies: 2003 Validation of method for genotyping from paraffin sections Methods developed for biochemical strain typing studies 2004 Genotyping of archived scrapie tissues from the National Veterinary
Services Laboratories (NVSL)/APHIS/VS/USDA Evaluation of biochemical methods for strain typing of scrapie isolates Development of mass spectrometry methods for characterization of protein expression in normal sheep brain 2005 Evaluation of biochemical methods for differentiation of TSE agents after cross-species transmission Comparison of protein expression (mass spectrometry) in scrapie brain to that in normal sheep brain 3. Milestones: A. List the milestones that were scheduled to be addressed in FY 2004. How many milestones did you fully or substantially meet in FY 2004 and indicate which ones were not fully or substantially met, briefly explain why not, and your plans to do so. Animal experiments: Studies are initiated as soon as appropriate animal space is available. Cattle inoculated with elk CWD - Started 18Aug04, projected study completion 18Aug10. Raccoons inoculated with TME, scrapie, and CWD isolates - Started early on 18Dec02, projected study completion 18Dec05. Mice
inoculated for strain typing of 10 TSE isolates: one of the re- derived inbred mouse strains, the VM strain, refused to breed; new embryos had to be obtained from the U.K. and are being re-derived. Several CWD isolates were inoculated into mice in Edinburgh, U.K., under a specific cooperative agreement. Swine inoculated with CWD - Will start 01Nov04 pending animal space availability. White-tailed deer inoculated with scrapie - Will start in autumn of 2004 pending animal space availability. Reindeer inoculated with CWD - On hold pending animal space availability over next 2 yrs. Laboratory studies: Evaluation of biochemical methods for strain typing of scrapie isolates: completed. Molecular mass determination and glycoform profile analysis methods were established as tools to differentiate TSE isolates. Preliminary results on naturally-occurring scrapie in 50 goats and 20 sheep indicate at least 2 PrPres molecular mass patterns within each species; PrPres glycoform analyses have
been uninformative in terms of identifying scrapie strains as each clinical case appears unique. Among 40 naturally occurring cases of CWD in elk and deer, preliminary findings indicate there is a molecular mass difference between elk and deer but no other differences detected thus far. Glycoform analyses among CWD isolates have been uninformative thus far. Development of mass spectrometry methods for characterization of protein expression in normal and diseased sheep brain: Not completed, however, this work has now been initiated, after a postdoctoral associate was hired to undertake these studies in FY 2004; additional equipment was obtained in FY2004 and methodologies are being developed. Initial experimentation indicate that detection of PrP in enriched samples is possible. This methodology will continue to be developed and should enable us to complete the milestone for next year. The majority of archived scrapie tissues provided by the NVSL/APHIS/VS/USDA have been genotyped
for their PrP alleles. All remaining samples (n=16) have been successfully amplified and have been submitted for sequencing. If some sequences are unreadable, then DNA will be re-extracted from fresh tissue sections from the same block(s). Anticipated completion by 01Oct04. B. List the milestones that you expect to address over the next 3 years (FY 2005, 2006, & 2007). What do you expect to accomplish, year by year, over the next 3 years under each milestone? FY-2005 Special lines of inbred mice, which have been used in Europe to differentiate scrapie strains in sheep, will be inoculated with several scrapie isolates and with additional CWD sources to determine if this strain typing procedure is useful for identification of unique TSE strains in the U.S. Cattle will be inoculated with the U.S. BSE isolate in order to amplify the BSE material for subsequent pathogenesis studies. In addition to continual monitoring of animals involved in transmission and strain typing experiments,
laboratory methods for TSE strain differentiation will be developed and evaluated. These studies will involve not only the examination of tissues from natural host species, but also the subsequent comparison of results from tissues of secondary hosts in cross-species transmission experiments. Inoculation of swine with CWD agent. FY-2006 and 2007 Because the incubation period for TSE transmission is typically 2 years or longer, it is anticipated that the major activity during this year will involve termination of many animal inoculation experiments that began in FY03. The work will include necropsies and tissue analysis to confirm TSE transmissions, followed by evaluations with the laboratory strain characterization methods developed and evaluated in FY05. 4. What were the most significant accomplishments this past year? A. Single Most Significant Accomplishment during FY 2004. CWD transmission to cattle: Findings of this study showed that although PrPres amplification occurred
following direct inoculation into the brain of CWD from mule deer, none of the affected animals had classic histopathologic lesions of spongiform encephalopathy. Furthermore, only 38% of the inoculated cattle demonstrated amplification of PrPres. Although intracerebral inoculation is an unnatural route of exposure, and is the most severe challenge possible, this experiment shows that CWD transmission in cattle could have long incubation periods (up to 5 years). Other CWDs (white-tailed and elk) may show different clinicopathological findings. B. Other Significant Accomplishments(s). Laboratory verification of the U.S. BSE case. Studies were conducted which confirmed the initial suspect BSE diagnosis. The PrPres profile from the first BSE case diagnosed in the United States showed similar molecular properties to the typical PrPres pattern described for the earlier Canadian and European BSE isolates, and a germline mutation in the bovine PrP gene was not evident. Prion protein
(PrPres) in tongue and muscles: In this study, samples of striated muscle tissues (tongue, heart, diaphragm and masseter muscle) from 20 animals (cattle, sheep, elk, and raccoons) were examined for PrPres by immunohistochemistry (IHC). All of the animals had developed a TSE after experimental inoculation. PrPres was found by IHC in the brains, but not in muscle tissues of all examined animals. In a separate study, tongues examined by a more sensitive technique (Western blot after PrP) enrichment did show presence of PrPres in some of the samples. Further testing of muscle tissues is needed to confirm these findings, as well to determine if PrP presence equals infectivitity. Transmission of CWD to cervids: Under free-ranging conditions, CWD has been observed in elk, mule deer and white-tailed deer. Since there is no available information to indicate that CWD of these 3 cervid species is the same (or if these are 3 different strains of cervid CWD), we inoculated CWD from these 3
sources to 3 groups of white-tailed deer fawns. At this time the experiment is still in-progress. However, preliminary data indicates that clinicopathologically, all 3 CWD sources appear to show more similarities than differences. Final results of the study should be available within the next 6 to 12 months. Developed a method to extract DNA from formalin-fixed paraffin-embedded brainstem tissue to determine prion gene polymorphisms in scrapie- affected sheep. Preliminary results indicate the method can deliver valid sequence data from approximately 90% of the samples tested thus far. Demonstrated the utility of raccoons as an animal model to distinguish scrapie, CWD and TME based on attack rate, IHC patterns, and time to clinical disease. C. Significant accomplishments/activities that support special target populations. Laboratory Verification of the U.S. BSE case. Assistance to the Animal and Plant Health Inspection Service (APHIS): Assignment to the Japan-U.S. BSE Working
Group. 5. Describe the major accomplishments over the life of the project, including their predicted or actual impact. Describe the major accomplishments over the life of the project, including their predicted or actual impact. This project was initiated January 28, 2002, as a result of the FY 2002 Appropriations Bill passed by Congress and signed by the President for research on Emerging & Exotic Diseases of Pests (BSE). The major objectives are to assess transmissibility of the TSEs that affect livestock and wildlife species, to develop methods for differentiation of TSE strains, and to determine the pathobiology of these diseases in the natural host and after cross-species transmission. Experiments are in progress to determine the transmissibility of CWD into cattle and sheep. Based on results of our studies, it may be concluded that under natural conditions cattle exposed to CWD would require a large dose of inoculum, and an extremely long incubation time to develop a
TSE-associated disease. Completed work has determined that scrapie can be transmitted to elk and that the resulting disease is indistinguishable from CWD in that species. White-tailed deer have been inoculated with CWD from elk, mule deer and white-tailed deer to determine if there are strain differences in the agent that depend on the host of origin. These same agents will be used in experiments to further assess the risk that CWD might pose to cattle, sheep, and swine. The experiments will also provide information and tissues that can be used to evaluate the effectiveness of current diagnostic protocols for the TSEs. An important contribution of this project has been the demonstration of the utility of raccoons as an animal model to distinguish scrapie, CWD and TME based on attack rate and time to clinical disease. Raccoons inoculated with TME develop disease within 6 months time and as such represent the fastest available, non- transgenic model of TSE disease. Through
collaborators we were able to compare the U.S. BSE isolate with other typical BSE isolates (Canadian, European) and found them to be indistinguishable. 6. What science and/or technologies have been transferred and to whom? When is the science and/or technology likely to become available to the end- user (industry, farmer, other scientists)? What are the constraints, if known, to the adoption and durability of the technology products? Presented to Agriculture Secretary, Ann M. Veneman, results of ARS research conducted in support of the first case of BSE reported in the United States. A project scientist was detailed on special assignment at the request of APHIS and the Agricultural Marketing Service (AMS) to serve in a scientific advisory capacity on the U.S. - Japan BSE Technical Working Group. This group met three times with a Japanese delegation to establish criteria to allow a return to normal U.S. beef export trade relations with Japan. As part of this special assignment, this
scientist authored a critical chapter entitled "Definition of BSE and the method of Testing", for the U.S. BSE Reference Book generated by this BSE Technical Working Group. This document is now guiding U.S. trade negotiations by the AMS and the International Trade Policy group of the Foreign Agricultural Service. At the request of NVSL/VS/APHIS, a project scientist developed a procedure for performing PrP gene sequencing from paraffin-embedded, formalin-fixed sheep brainstem samples. The methodology for extracting, amplifying, and sequencing PrP gene DNA from paraffin-embedded sheep brain was developed over the past year in our laboratory; collaboration with APHIS has confirmed their interest in this methodology and it is anticipated that transfer of this technology will be completed by the end of the year. This technology is important because it could eliminate the need for frozen tissue collection for gene sequencing from samples collected as part of the National scrapie
surveillance program. List your most important publications in the popular press and presentations to organizations and articles written about your work. 7. List your most important publications in the popular press and presentations to organizations and articles written about your work. A presentation on ARS research conducted in support of the first reported case of BSE identified in the United States was made at the National Cattlemen's Beef Association Production Research Committee Meeting in Phoenix, Arizona. Presented talk on animal TSE issues at the Annual Food Safety Symposium, Oklahoma Food and Agricultural Products Research and Technology Center, Stillwater, Oklahoma, April 19, 2004. Presented a seminar on CWD at the Annual Conference of the North American Deer and Elk Farming Association, Oconomowoc, Wisconsin, February 26, 2004. Presented a seminar on CWD at the Reindeer Owners and Breeders Association Annual Meeting, Ames, Iowa.
Impacts
(N/A)
Publications
- Hamir, A.N., Cutlip, R.C., Miller, J.M., Richt, J.A., Kunkle, R.A., Williams, E., Stack, M., Miller, M.W., O'Rourke, K.I., Chaplin, M.J. 2003. Experimental cross-species transmission of chronic wasting disease to domestic livestock at the National Animal Disease Center: An update [Abstract]. International Conference on Emerging Zoonoses. p. 91.
- Hamir, A.N., Cutlip, R.C., Miller, J.M., Richt, J.A., Kunkle, R.A., Williams, E., Stack, M., Chaplin, M., Miller, M., O'Rourke, K.I. 2003. Experimental cross-species transmission of chronic wasting disease (CWD- mule deer) to domestic livestock at the National Animal Disease Center: An update [Abstract]. 52nd Annual Wildlife Disease Association Conference. p. 134.
- Hamir, A.N., Miller, J.M., Cutlip, R.C., Stack, M., Chaplin, M., Bartz, J., Jenny, A., Williams, E. 2003. Experimental inoculation of TME, scrapie, and CWD to raccoons (Procyon lotor) and the utilization of raccoons for strain typing of unknown TSEs in the United States [Abstract]. 4th International Conference on Emerging Zoonoses. p. 93.
- Hamir, A.N., Miller, J.M., Cutlip, R.C., Stack, M., Chaplin, M., Bartz, J., Jenny, A., Williams, E. 2003. Experimental inoculation of TME, scrapie, and CWD to raccoons: An update [Abstract]. 52nd Annual Wildlife Disease Association Annual Meeting. p. 125.
- Hamir, A.N., Miller, J.M., Cutlip, R.C., Richt, J.A., Kunkle, R.A., O'Rourke, K.I., Jenny, A.L., Stack, M.J., Chaplin, M.J. 2003. Transmission of sheep scrapie to elk (Cervus elaphus nelsoni) by intracerebral inoculation [Abstract]. 52nd Annual Wildlife Disease Association Annual Meeting. p. 135.
- Hamir, A.N., Miller, J.M., Cutlip, R.C., Richt, J.A., Kunkle, R.A., O'Rourke, K.I., Jenny, A.L., Stack, M.J., Chaplin, M.J. 2003. Transmission of sheep scrapie to elk (Cervus elaphus nelsoni) by intracerebral inoculation. 46th Annual Meeting of the American Association of Veterinary Laboratory Diagnosticians. p. 210.
- Hamir, A.N., Cutlip, R.C., Miller, J.M., Richt, J.A., Kunkle, R.A., Williams, E., Stack, M.J., Chaplin, M.J., Miller, M., O'Rourke, K.I. 2003. Experimental cross-species transmission of chronic wasting disease (CWD- mule deer) to domestic livestock at the National Animal Disease Center: An update [Abstract]. 46th Annual Meeting of the American Association of Veterinary Laboratory Diagnosticians. p. 209.
- Richt, J.A., Hamir, A.N., Hoover, E.A., Bartz, J.C., Bessen, R.A. 2003. Molecular analysis of scrapie and chronic wasting disease isolates from the U.S. [Abstract]. 46th Annual Meeting of the American Association of Veterinary Laboratory Diagnosticians. p. 61.
- Young, A.J., Hamir, A.N., Richt, J.A. 2003. A potential role for B cell subsets in the pathogenesis of scrapie [Abstract]. Conference of Research Workers in Animal Diseases. p. 107.
- Richt, J.A., Kluge, J.P., Alt, D.P., Kunkle, R.A., Hamir, A.N., Czub, S., Davis, A.J., Hall, S.M. 2004. Identification and characterization of the bovine spongiform encephalopathy case diagnosed in the United States [Abstract]. XXth International Winter Meeting of the Swiss Society of Neuropathology. Paper No. 13, p. 29.
- Hamir, A.N., Miller, J.M., Cutlip, R.C., Stack, M.J., Chaplin, M.J., Jenny, A.L., Williams, E.S. 2003. Experimental inoculation of sheep scrapie and chronic wasting disease agents to raccoons (Procyon lotor). Veterinary Record. 153:121-123.
- Hamir, A.N., Miller, J.M., O'Rourke, K.I., Bartz, J.C., Stack, M.J., Chaplin, M.J. 2004. Transmission of transmissible mink encephalopathy (TME) to raccoons (Procyon lotor) by intracerebral inoculation. Journal of Veterinary Diagnostic Investigation. 16(1):57-63.
- Hamir, A.N., Stasko, J.A., Rupprecht, C.E. 2004. Observation of Helicobacter-like organisms in gastric mucosa of grey foxes (Urocyon cinereoargenteus) and bobcats (Lynx rufus). Canadian Journal of Veterinary Research. 68(2):154-156.
- Hamir, A.N., Miller, J.M., Cutlip, R.C. 2004. Failure to detect prion protein (PrPres) by immunohistochemistry in striated muscle tissues of animals experimentally inoculated with agents of transmissible spongiform encephalopathy. Veterinary Pathology. 41(1):78-81.
- Hamir, A.N., Miller, J.M., Cutlip, R.C., Richt, J.A., Kunkle, R.A., O'Rourke, K.I., Jenny, A.L., Stack, M.J., Chaplin, M.J. 2003. Transmission of sheep scrapie to elk (Cervus elaphus nelsoni) by intracerebral inoculation [Abstract]. International Conference on Emerging Zoonoses. p. 92.
- Hamir, A.N., Miller, J.M., Cutlip, R.C., Kunkle, R.A., Jenny, A.L., Stack, M.J., Chaplin, M.J., Richt, J.A. 2004. Transmission of sheep scrapie to elk (Cervus elaphus nelsoni) by intracerebral inoculation: Final outcome of the experiment. Journal of Veterinary Diagnostic Investigation. 16(4) :316-321.
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Progress 10/01/02 to 09/30/03
Outputs
1. What major problem or issue is being resolved and how are you resolving it? This project deals with Transmissible Spongiform Encephalopathies (TSE), which are fatal degenerative diseases of the central nervous system that can affect several animal species, including humans. The causal agent is believed to be a natural tissue protein, the prion protein, that has assumed an unnatural form. Because the altered protein is resistant to enzyme degradation, it accumulates in nervous tissue and the resulting dysfunction ultimately leads to death. The specific TSEs being investigated in this project are scrapie in sheep and chronic wasting disease (CWD) in deer and elk. The major concern about these diseases is that another TSE, bovine spongiform encephalopathy (BSE), has been shown to cross the species barrier to cause a unique TSE in human beings. Although there has not been a similar demonstration that scrapie or CWD could present any risk to human health, the BSE
experience has raised many questions about the potential hazard these TSEs present for transmission to other animal species, especially domesticated livestock and wildlife. The current research is focused on direct experimental challenge of the species barrier effect by animal inoculation. These results are then compared to results obtained with a variety of laboratory procedures to determine if they can be used as predictive models for future risk assessments. These studies also provide information about the clinical and pathological disease characteristics that can be expected if a TSE crosses the species barrier, thus enabling animal health specialists to recognize such situations should they occur. Additional transmission studies in the natural host will focus on determining the modes of transmission and disease development in scrapie and CWD so that appropriate intervention strategies can be devised that will control the spread of these diseases. 2. How serious is the problem?
Why does it matter? Because of the risk for BSE transmission to human beings, the presence of scrapie and CWD in the U.S. presents a potential liability to the U.S. livestock and hunting industries because these diseases also may be viewed by consumers as a potential public health risk. Furthermore, the safety of animals and animal products intended for international trade may be questioned. Efforts are being made by both federal and state regulatory agencies to eradicate scrapie and CWD, but the effectiveness of these programs will depend heavily on having accurate information about the nature of these diseases, not only in the original hosts, but also in other species that may be affected. 3. How does it relate to the National Program(s) and National Program Component(s) to which it has been assigned? National Program 103, Animal Health (100%) The work of this project supports National Program 103, Animal Health, and more specifically those components that deal with pathogen
detection and diagnosis, mechanisms of disease, and strategies to control disease. 4. What were the most significant accomplishments this past year? A. Single Most Significant Accomplishment during FY 2003: Three forms of TSE have been reported in the U.S. They are scrapie in sheep, transmissible mink encephalopathy (TME) in mink, and chronic wasting disease (CWD) in deer and elk. Because raccoons will consume tissues from dead animals, there was a concern about the possibility that they could act as reservoir hosts for transmission of TSE agents. To examine their susceptibility, groups of raccoons were inoculated with scrapie, TME, or CWD tissues. To date, after 3 years of observation, there is no evidence of CWD transmission, but all of the animals inoculated with TME and scrapie developed TSE. The incubation period for transmission of these 2 agents differed, however, being 2 years for scrapie but only 6 months for TME. This study shows that raccoons are susceptible to TME
and scrapie but probably not to CWD (longer observation is required to assure nonsusceptibility). The dramatic differences in response to the 3 agents also suggests that raccoons may provide an animal model for differentiation of TSE agents, as in the case of a suspected cross-species transmission event. B. Other Significant Accomplishments(s), if any: none. C. Significant Activities that Support Special Target Populations: none. 5. Describe the major accomplishments over the life of the project, including their predicted or actual impact. This project was initiated in January, 2002. The major objectives are to assess transmissibility of the TSEs that affect livestock and wildlife species, to develop methods for differentiation of TSE strains, and to determine the pathobiology of these diseases in the natural host and after cross-species transmission. Experiments are in progress to determine the transmissibility of CWD from mule deer into cattle and sheep. Completed work has
determined that scrapie can be transmitted to elk and that the resulting disease is indistinguishable from CWD in that species. White-tailed deer have been inoculated with CWD from elk, mule deer and white-tailed deer to determine if there are strain differences in the agent that depend on the host of origin. These same agents will be used in experiments to further assess the risk that CWD might pose to cattle, sheep, and swine. The experiments will also provide information and tissues that can be used to evaluate the effectiveness of current diagnostic protocols for the TSEs. 6. What do you expect to accomplish, year by year, over the next 3 years? FY04. Cattle will be inoculated with CWD from elk. Swine will be inoculated with CWD and scrapie. Animals inoculated in FY03 will be monitored for signs of TSE. These experiments include cattle inoculated with CWD from white-tailed deer and fallow deer inoculated with the elk and white-tailed deer CWD agents. Raccoons will be
inoculated with additional isolates of scrapie, CWD and TME to confirm and extend findings reported in the past year. Special lines of inbred mice, which have been used in Europe to differentiate scrapie strains in sheep, will be inoculated with several scrapie isolates and with each of the 3 CWD sources to determine if this strain typing procedure is useful for identification of unique TSE strains in the U.S. FY05. In addition to continual monitoring of animals involved in transmission and strain typing experiments, laboratory methods for TSE strain differentiation will be developed and evaluated. These studies will involve not only the examination of tissues from natural host species, but also the subsequent comparison of results from tissues of secondary hosts in cross-species transmission experiments. FY06. Because the incubation period for TSE transmission is typically 2 years or longer, it is anticipated that the major activity during this year will involve termination of
many animal inoculation experiments that began in FY03. The work will include necropsies and tissue analysis to confirm TSE transmissions, followed by evaluations with the laboratory strain characterization methods developed and evaluated in FY05.
Impacts
(N/A)
Publications
- Hamir, A.N., Miller, J.M., Stack, M.J., Chaplin, M.J. Failure to detect abnormal prion protein and scrapie-associated fibrils 6 wk after intracerebral inoculation of genetically susceptible sheep with scrapie agent. Canadian Journal of Veterinary Research. 2002. v. 66. p. 289-294.
- Hamir, A.N., Smith, B.B. Severe biliary hyperplasia associated with liver fluke infection in an adult alpaca. Veterinary Pathology. 2002. v. 39. p. 592-594.
- Hamir, A.N., Timm, K.I. Nodular hyperplasia and cysts in thyroid glands of llamas (Lama glama) from north-west USA. Veterinary Record. 2003. v. 152. p. 507-508.
- Hamir, A.N., Clark, W.W., Sutton, D.L., Miller, J.M., Stack, M.J., Chaplin M.J., Jenny, A.L. Resistance of domestic cats to a US sheep scrapie agent by intracerebral route. Journal of Veterinary Diagnostic Investigation. 2002. v. 14. p. 444-445.
- Hamir, A.N., Miller, J.M., Cutlip, R.C., Stack, M.J., Chaplin M.J., Jenny, A.L. Preliminary observations on the experimental transmission of scrapie to elk (Cervus elaphus nelsoni) by intracerebral inoculation. Veterinary Pathology. 2003. v. 40. p. 81-85.
- Hamir, A.N., Miller, J.M., Sonn, R.J. Meningoencephalitis and pneumonia associated with Cryptococcus neoformans infection in a free-ranging elk in the USA. Veterinary Record. 2002. v. 151. p. 332-333.
- Hamir, A.N., Miller, J.M., Cutlip, R.C., Stack, M.J., Chaplin, M.J., Bartz, J., Jenny, A.L., Williams, E.S. Experimental inoculation of TME, scrapie, and CWD to raccoons (Procyon lotor) and the utilization of raccoons for strain-typing of unknown TSEs in the United States. American Association of Veterinary Laboratory Diagnosticians. 2002. p. 117.
- Hamir, A., Miller, J., Cutlip, R., Stack, M., Chaplin, M., Bartz, J., Jenny, A., Williams, E. Experimental inoculation of TME, scrapie, and CWD to raccoons: An update. Conference: Diseases at the Interface Between Domestic Livestock and Wildlife Species. 2003. Abstract No. 5.
- Hamir, A.N., Miller, J.M., Cutlip, R.C., Richt, J.A., Kunkle, R.A., O'Rourke, K.I., Jenny, A.L., Stack, M.J., Chaplin, M.J. Transmission of sheep scrapie to elk (Cervus elaphus nelsoni) by intracerebral inoculation. Diseases at the Interface Between Domestic Livestock and Wildlife Species. 2003. Abstract No. 6.
- Hamir, A.N., Sonn, R.J., Franklin, S., Wesley, I. Small Intestinal Intussusception Associated with Campylobacter jejuni Infection in a Raccoon (Procyon lotor). Diseases at the Interface Between Domestic Livestock and Wildlife Species. 2003. Abstract No. 7.
- Hamir, A., Cutlip, R., Miller, J., Richt, J., Kunkle, R., Williams, E., Stack, M., Chaplin, M., Miller, M., O'Rourke, K. Experimental cross- species transmission of chronic wasting disease (CWD mule deer) to domestic livestock at the National Animal Disease Center: An update. Diseases at the Interface Between Domestic Livestock and Wildlife Species. 2003. Abstract No. 8.
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Progress 10/01/01 to 09/30/02
Outputs
1. What major problem or issue is being resolved and how are you resolving it? This project deals with Transmissible Spongiform Encephalopathies (TSE), which are fatal degenerative diseases of the central nervous system that can affect several animal species, including humans. The causal agent is believed to be a natural tissue protein, the prion protein, that has assumed an unnatural form. Because the altered protein is resistant to enzyme degradation, it accumulates in nervous tissue and the resulting dysfunction ultimately leads to death. The specific TSEs being investigated in this project are scrapie in sheep and chronic wasting disease (CWD) in deer and elk. The major concern about these diseases is that another TSE, bovine spongiform encephalopathy (BSE), has been shown to cross the species barrier to cause a unique TSE in human beings. Although there has not been a similar demonstration that scrapie or CWD could present any risk to human health, the BSE
experience has raised many questions about the potential hazard these TSEs present for transmission to other animal species, especially domesticated livestock and wildlife. The current research is focused on direct experimental challenge of the species barrier effect by animal inoculation. These results are then compared to results obtained with a variety of laboratory procedures to determine if they can be used as predictive models for future risk assessments. These studies also provide information about the clinical and pathological disease characteristics that can be expected if a TSE crosses the species barrier, thus enabling animal health specialists to recognize such situations should they occur. Additional transmission studies in the natural host will focus on determining the modes of transmission and disease development in scrapie and CWD so that appropriate intervention strategies can be devised that will control the spread of these diseases. 2. How serious is the problem?
Why does it matter? Because of the risk for BSE transmission to human beings, the presence of scrapie and CWD in the U.S. presents a potential liability to the U.S. livestock and hunting industries because these diseases also may be viewed by consumers as a potential public health risk. Furthermore, the safety of animals and animal products intended for international trade may be questioned. Efforts are being made by both federal and state regulatory agencies to eradicate scrapie and CWD, but the effectiveness of these programs will depend heavily on having accurate information about the nature of these diseases, not only in the original hosts, but also in other species that may be affected. 3. How does it relate to the national Program(s) and National Program Component(s) to which it has been assigned? The work of this project supports National Program 103, Animal Health, and more specifically those components that deal with pathogen detection and diagnosis, mechanisms of disease,
and strategies to control disease. 4. What was your most significant accomplishment this past year? A. Single Most Significant Accomplishment during FY 2002: A major concern with respect to the transmissible spongiform encephalopathies (TSE) of animals is to determine their capacity for interspecies transmissions. At the National Animal Disease Center, sheep were inoculated with the agent of chronic wasting disease (CWD), which is a TSE of deer and elk. During FY02 a sheep developed the clinical signs and microscopic lesions indicative of a TSE and laboratoy tests confirmed the presence of a TSE marker protein (prion) in the tissues. Results of this experiment show that CWD can be transmitted to sheep and that the resulting disease is indistinguishable from scrapie, a TSE that occurs naturally in U.S. sheep and for which a national eradication plan has been initiated. B. Other Significant Accomplishments(s), if any: none. C. Significant Accomplishments/Activities that Support
Special Target Populations: none. 5. Describe your major accomplishments over the life of the project, including their predicted or actual impact? Project was initiated January 28, 2002, as a result of the FY 2002 Appropriations Bill passed by Congress and signed by the President for research on Bovine Spongiform Encelphalopathies and Other Transmissible Spongiform Encelphalopathies. We will continue research on direct assessment of cross-species transmissibility among the different TSE agents. However, this activity will be considerably expanded in scope, with special emphasis on the transmissibility of CWD between different cervid species and to traditional livestock species. 6. What do you expect to accomplish, year by year, over the next 3 years? FY03. The CWD agent from white-tailed deer, mule deer, and elk will be inoculated into white-tails to determine if these 3 agents are equally pathogenic in that species. Species susceptibility experiments will be initiated to
determine the transmissibility of mule deer CWD to fallow deer, white-tailed deer CWD to cattle, and scrapie to swine. Special strains of inbred mice, which have been used to differentiate scrapie strains in sheep, will be inoculated with each of the 3 CWD sources to determine if this strain typing procedure is useful for identification of unique CWD strains. FY04. In addition to continual monitoring of the experiments from FY03, cattle will be inoculated with elk CWD as an additional transmissibility evaluation. Laboratory methods that have been proposed as methods for differentiation of scrapie and CWD will be conducted so that the results can be compared to results of direct animal inoculations. FY05. Because the incubation period for TSE transmissions are typically 2 years or longer, it is anticipated that the major activity during this year will involve termination of many animal inoculation experiments that began in FY03. The work will include necropsies and tissue analysis
to confirm TSE transmission, followed by comparisons with the laboratory strain characterization methods developed and evaluated in FY04.
Impacts
(N/A)
Publications
- Dubey, J.P., Saville, W.J., Stanek, J.F., Lindsay, D.S., Rosenthal, B.M., Oglesbee, M.J., Rosypal, A.C., Njoku, C.J., Stich, R.W., Kwok, O.C., Shen, S.K., Hamir, A.N., Reed, S.M. Sarcocystis neurona infections in raccoons (Procyon lotor): evidence for natural infection with sarcocysts, transmission of infection to opossums (Didelphis virginiana), and experimental induction of neurologic disease in raccoons. Veterinary Parasitology. 2001. v. 100. p. 117-129.
- Tornquist, S.J., Boeder, L.J., Mattson, D.E., Cebra, C.K., Bildfell, R.J., Hamir, A.N. Lymphocyte responses and immunophenotypes in horses with Sarcocystis neurona infection. Equine Veterinary Journal. 2001. v. 33. p. 726-729.
- Dubey, J.P., Hamir, A.N. Experimental toxoplasmosis in budgerigars (Melopsittacus undulatus). Journal of Parasitology. 2002. v. 88. p. 514- 519.
- Dubey, J.P., Hamir, A.N., Topper, M.J. Sarcocystis mephitisi n. sp. (Protozoa: Sarcocystidae), Sarcocystis neurona-like and Toxoplasma-like infections in striped skunks (Mephitis mephitis). Journal of Parasitology. 2002. v. 88. p. 113-117.
- Heidari, M., Hamir, A.N., Cutlip, R.C., Brogden, K.A. Antimicrobial anionic peptide binds in vivo to Mannheimia (Pasteurella) haemolytica attached to ovine alveolar epithelium. International Journal of Antimicrobial Agents. 2002. v. 20. p. 69-72.
- Wang, S., Cockett, N.E., Miller, J.M., Shay, T.L., Maciulis, A., Sutton, D. L., Foote, W.C., Holyoak, G.R., Evans, R.C., Bunc, T.D., Beever, J.E., Call, J.W., Taylor, W.D., Marshall, M.R. Polymorphic distribution of the ovine prion protein (PrP) gene in scrapie-infected sheep flocks in which embryo transfer was used to circumvent the transmissions of scrapie. Theriogenology. 2002. v. 57. p. 1865-75.
- Hamir, A.N., Miller, J.M. Experimental cross-species transmission of CWD at NADC, Ames, Iowa. Chronic Wasting Disease Symposium. 2002. p. 14.
- Hamir, A.N., Hanlon, C.A., Niezgoda, M., Rupprecht, C.E. The prevalence of interstitial nephritis and leptospirosis in 283 raccoons (Procyon lotor) from 5 different sites in the United States. Canadian Veterinary Journal. 2001. v. 42. p. 869-871.
- Hamir, A.N., Miller, J.M., Stack, M.J., Chaplin, M.J., Cutlip, R.C. Neuroaxonal dystrophy in raccoons (Procyon lotor) from Iowa. Journal of Veterinay Diagnostic Investigation. 2002. v. 14. p. 175-178.
- Hamir, A.N., Olsen, S., Rupprecht, C.E. Granulomatous orchitis associated with Histoplasma-like organisms in porcupines (Erethizon dorsatum). Veterinary Record. 2002. v. 150. p. 251-252.
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