Animal Health (NP 103) Annual Report for 2005

Introduction

The mission of the Animal Health Program is to conduct basic and applied research on selected diseases of economic importance to the United States livestock and poultry industries. The goals of the research mission are to produce knowledge and technology to reduce economic losses from infectious, genetic, and metabolic diseases of livestock and poultry. Dr. Cyril G. Gay, National Program Leader, Animal Health and Safety, and Dr. Robert Heckert, National Program Leader, Animal Health, continue to lead this ARS research program. Dr. Gay is lead on domestic disease issues and vaccine and drug discovery projects, and Dr. Heckert is lead on projects involving foreign and emerging diseases. 

The Animal Health National Program is coming to the end of its five-year program cycle.  The program assessment of past performance was completed in 2005 and a national Stakeholder’s workshop was held in Kansas City September 2005.  In 2005, scientists in the Animal Health program will begin writing their project plans that will set the research plans for the next five years up until 2012.

The Animal Health National Program currently includes 50 research projects supported by 106 scientists located at 11 research sites throughout the country.  The ARS research budget for the Animal Health Program Fiscal Year (FY) 2005 was $52.2 million (NTL).  Critical to the success of the research conducted at ARS is the ability to build and maintain the infrastructure of our laboratories to ensure the highest level of biosecurity and quality research.  Accordingly, we are continuing to develop plans to upgrade the animal research facilities at the National Animal Disease Center (NADC) in Ames Iowa. Construction of a high containment animal barn began in the fall of 2003 and is progressing.  Design of a consolidated laboratory that will be used by NADC and the Animal and Plant Health Inspection Service (APHIS) is progressing.

Several of the scientists in the Animal Health National Program again received accolades this past year.  Dr. W. Ray Waters, in the Bacterial Diseases of Livestock Research Unit at the NADC working on M. bovis in wildlife and cattle, received the Midwest Area Early Career Scientist Award. Dr. William (Bill) Huff with the Poultry Production and Product Safety Research Unit, in Fayetteville, AR was awarded the Broiler Research Award for research demonstrating that bacteriophage may provide an alternative to antibiotics at the annual meeting of the Poultry Science Association. Dr. Hyun Lillehoj, in the Animal Parasitic Disease Laboratory in Beltsville, received a $2 million, 5 year CSREES NRI grant as a co-PI. Dr. Barbara Drolet at the Arthropod borne Animal Disease Research Laboratory in Laramie, WY received a $33,500 grant from the Max Kade Foundation in New York. Dr. David Swayne at the Southeast Poultry Research Laboratory in Athens, GA received the Lamplighter Award from the U.S. Poultry and Egg Association for sustained exemplary service to the poultry industry.

Scientists within the National Animal Health Program were very active in their fields Fiscal Year (FY) 2005, with 186 articles published in peer-reviewed scientific journals.  Many of the discoveries and findings were published in the popular press to reach our customers and stakeholder, including 23 articles in trade journals and book chapters.  Technology transfer activities for the National Animal Health Program included 12 invention disclosures, 8 new Cooperative Research and Development Agreements (CRADA), 33 active Specific Cooperative Agreements (SCA), 99 Material Transfer Agreements (MTA), and 2 licenses from ARS inventions.

The following section of the report summarizes high impact research results addressing objectives in the current national program action plan.

Animal Health Research Highlights

Avian Coccidiosis and Determinants of Cross-Protective Immunity

Coccidiosis is a ubiquitous intestinal protozoan infection of poultry that seriously impairs the growth and feed utilization of infected birds.  This enteric disease is caused by several distinct apicomplexan species of obligate intracellular parasites of the genus Eimeria.  The use of anti-coccidial drugs is the primary control method but the worldwide emergence of drug-resistant coccidia strains are limiting the effectiveness of existing therapeutics.  This problem is compounded by our lack of understanding of the mechanism(s) that confer(s) drug resistance and the host-parasite and environmental factors that influence coccidiosis susceptibility.  Vaccines provide an important alternative to drug therapy but existing vaccines, which are comprised of one or more live coccidian species do not provide cross-protection against all seven species of Eimeria.  ARS scientists in Beltsville Maryland have discovered a conserved protein named SZ-1 that is highly conserved in E. tenella and two other important protozoan:  Toxoplasma gondii and Neospora caninum. The full length gene from T. gondii was characterized, expressed in a bacterial system, and the cloned protein was used to make antibodies to T. gondii  SZ1. These antibodies are providing important functional genomics research tools that can be used to determine the function of the SZ1 protein, and determine whether this protein might confer cross-protective immunity across Eimeria strains.  (National Program 103 and Performance Measure 3.2.1)

Scientific Publications:

Allen, P.C., Jenkins, M.C., Miska, K.B. 2005. Cross protection studies with eimeria maxima strains. Parasitology Research. Available: http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?CMD=Pager&DB=pubmed.

Fetterer, R.H., Miska, K.B., Jenkins, M.C., Barfield, R.C. 2004. A conserved 19 kda Eimeria tenella antigen is a profilin-like protein. Journal of Parasitology. 90:1321-1328.

Matsubayashi, M., Kimata, I., Iseki, M., Lillehoj, H.S., Matsuda, H., Nakanishi, T., Tani, H., Sasai, K., Baba, E. 2005. Cross-reactivities with cryptosporidium spp. by chicken monoclonal antibodies that recognize avian Eimeria spp. Veterinary Parasitology. 128:47-57.

Dallul, R.A., Lillehoj, H.S. 2005. Recent advances in immunomodulation and vaccination strategies against coccidiosis. Avian Diseases. 48:1-8.

Genetic Determinants of Chronic Wasting Disease (CWD) Susceptibility in Elk

Current control measures for transmissible spongiform encephalopathies in livestock depend on identification of the most appropriate tissue for diagnostic testing and identification of candidate resistant genotypes.  ARS scientists in Pullman Washington in collaboration with Colorado State University and the Canadian Food Inspection Agency have described the distribution of the CWD prion protein in elk and the genotypes in elk susceptible to disease.  The genetic analysis performed by ARS scientists provides the scientific basis for selecting the brain as the most reliable indicator of disease in elk, in contrast to the tests for deer, which rely on lymphoid tissue.  This work also demonstrated the first confirmed case of CWD in an elk of the relatively rare genotype 132LL, thereby ruling out this genotype as conferring resistance to disease under field conditions.  .  (National Program 103 and Performance Measure 3.2.2)

Scientific Publications:

Spraker, T.R., Balachandran, A., Zhuang, D., O'Rourke, K.I. 2004. Variable patterns of distribution of PrP(CWD) in the obex and cranial lymphoid tissues of Rocky Mountain elk (Cervus elaphus nelsoni) with subclinical chronic wasting disease. Veterinary Record. 155(10):295-302.

The Pathogenesis of Tibial Dyschondroplasia in Poultry

Intensive genetic selection for fast-growing high-yield birds has led to tremendous improvements in production efficiency.  However, selection pressure for rapid growth has negatively affected the normal development of the musculoskeletal system.  ARS scientists have been investigating the molecular mechanisms of tibial dyschondroplasia (TD), a common poultry skeletal problem, using a disease model that employs thiram (a fungicide) to disrupt chondrocyte growth and differentiation with the goal of finding whether this important skeletal problem can be prevented using nutritional means.  ARS scientists examined the changes in gene expression and the cellular and metabolic alterations in the growth plate during early periods of the onset of TD.  These studies revealed that TD was not induced by an aberrant pattern of gene expression in the growth plate, but was due to the death of endothelial cells that cause capillary vessel degeneration and leads to subsequent chondrocyte death.  These studies provide mechanistic insight into the pathogenesis of TD and will be useful to identify nutritional factors that may help prevent blood vessel death and the pathogenesis of TD.  (National Program 103 and Performance Measure 3.2.1)

Scientific Publications:

Rath, N.C., Richards, M.P., Huff, W.E., Huff, G.R., Balog, J.M. 2005. Changes in the tibial growth plates of chickens with thiram induced dyschondroplasia. Journal of Comparative Pathology. 133:41-52.

Optimizing the Delivery of Poultry Mycoplasma gallisepticum (MG) Vaccines.

Research into live Mycoplasma gallisepticum (MG) vaccine administration to layer chickens has shown that the pressure (psi) utilized for spray administration is extremely important in impacting subsequent seroconversion (positive blood tests).  A survey of the layer chicken industry showed that pressure settings used for the administration of live MG vaccine varied from 35-70 psi.  Research conducted by ARS scientists using the CPJ Vaccinator to determine the optimum pressure setting to dispense live MG vaccined demonstrated dramatic increases in MG colony counts resulting from using the lower (40 psi) setting as compared to the higher 60 psi setting.  This information is important as it explains one factor (pressure setting of the vaccinator) that can impact the administration of live MG vaccines that may, in turn, result in poor vaccination results.  As a result of poor vaccine test results, re-vaccination must take place, which entails costs of additional vaccine (approximately $1500/75,000 bird house) and labor.  (National Program 103 and Performance Measure 3.2.3)

Scientific Publications:

Branton, S.L., Lott, B.D., Evans, J.D., Collier, S.D., Roush, W.B., Bearson, S.M., Bearson, B.L., Pharr, G.T. 2005. A self-propelled, constant speed spray vaccinator for commercial layer chickens. Avian Diseases. 49:147-151.

The Diseases Agents Responsible for Poult enteritis mortality syndrome (PEMS)

Poult enteritis mortality syndrome (PEMS) is a highly infectious disease of young turkeys. PEMS was first reported in North Carolina in 1991. Since then, PEMS and similar disease conditions have been reported in most regions where turkeys are commercially produced and are costing the poultry industry millions of dollars annually.  The major impact of PEMS is due to mortality and decreased production as turkeys are stunted and grow poorly when affected by the disease.  Currently the agent or agents that cause PEMS are unknown.  ARS scientists have discovered an avian rotavirus in specimens from turkeys with poult enteritis and broiler chickens with runting-stunting syndrome.  Rotaviruses are well known enteric pathogens that have been minimally characterized in poultry.  Initial pathogenesis studies were performed with clinical specimens containing rotavirus.  Identification of the viral agents associated with poult enteritis and the new similar condition in chickens, broiler runting-stunting syndrome, is critical to controlling the disease.  (National Program 103 and Performance Measure 3.2.1)

Scientific Publications:

Spackman, E., Pantin Jackwood, M.J., Day, J.M., Sellers, H. 2005. The pathogenesis of turkey origin reoviruses in turkeys and chickens. Avian Pathology 34(4):291-296.

Spackman, E., Kapczynski, D.R., Sellers, H.,  2005. Multiplex real-time rt-pcr for the detection of three viruses associated with poult enteritis complex: turkey astrovirus, turkey coronavirus, and turkey reovirus. Avian Diseases. 49:86-91.

Innate Immune Response of Porcine Reproductive and Respiratory Syndrome (PRRS) Infections

ARS scientists have determined critical innate immune markers required for effective immune responses against porcine reproductive and respiratory syndrome (PRRS). Vaccine trials with the University Illinois-Urbana scientists showed that the use of interferon-alpha (IFNA) as a novel cytokine vaccine adjuvant did not improve protection against viral challenge. Ongoing genetic studies with University Nebraska-Lincoln scientists indicate that there may be correlations between levels of certain innate cytokines and resistance to PRRS associated pathologies.  ARS scientists have demonstrated that PRRS virus infection does not result in the induction of type I interferons in MARC 145 cells as would be expected with most RNA viruses.  These results are significant because both IFNA and IFNB (type I interferons) are members of the innate immune system, which is typically viewed as the first responder of the immune system.  Activation of this response signals other branches of the immune system to become activated and mount a protective immune response. The fact that PRRS virus is capable of suppressing the activation of this response may explain the general delayed immune response to PRRS virus infection.  Elucidation of the mechanism of PRRS virus suppression of the type I interferon response may provide targets for novel vaccination approaches to control this important disease.  (National Program 103 and Performance Measure 3.2.1)

Scientific Publications:

Miller, L.C., Fox, J.M. 2004. Apoptosis and porcine reproductive and respiratory syndrome virus. Veterinary Immunology and Immunopathology 102(3):131-142.

Miller, L.C., Laegreid, W.W., Bono, J.L., Chitko McKown, C.G., Fox, J.M. 2004. Interferon type I response in porcine reproductive and respiratory syndrome virus-infected MARC-145 cells. Archives of Virology. 149(12):2453-2463.

Royaee, A.R., Husmann, R.J., Dawson, H.D., Calzada-Nova, G., Schnitzlein, W.M., Zuckermann, F., Lunney, J.K. 2004. Deciphering the involvement of innate immune factors in the development of the host responses to Porcine Respiratory and Reproductive Syndrome Virus (PRRSV) vaccination. Veterinary Immunology and Immunopathology. 102(3):199-216.

New Identification Methods to Assess Parasitic Infections of Ruminants

New methods were developed and validated by ARS scientists for geographically extensive and site intensive survey and inventory of parasites in ungulate hosts based on application of molecular sequence data.  Protostrongyle nematodes include pathogenic parasites that reside in the pulmonary system, skeletal musculature, or the central nervous system of their ruminant hosts.  Identification based on either adults in tissue and tissue spaces, or larval parasites in feces has remained problematic, and has hampered a detailed understanding of host distribution and geographic range. Such information is critical in defining the potential for disease, and the degree to which parasites may be shared among a number of different ungulates.  A combination of comparative morphology and molecular analyses were applied to define the host and geographic range for Parelaphostrongylus odocoilei in North America.  Molecular identification of larvae indicates that the protostrongylid parasite occupies a broader geographic range in western North America than previously reported.  A total of 2,124 fecal samples from 29 locations from thinhorn sheep, bighorn sheep, mountain goats, woodland caribou, mule deer, and black-tailed deer were tested.  This study provided significant molecular epidemiological data and represents the first study to combine extensive fecal surveys, comparative morphology, and molecular diagnostic techniques to comprehensively describe the host associations and geographic distribution of a parasitic helminth.  The development of such “epidemiological probes” will have significant applications in veterinary and conservation medicine. (National Program 103 and Performance Measure 3.2.1)

Scientific Publications:

Hoberg, E.P., Lichtenfels, J.R., Gibbons, L. 2004. Phylogeny for species of the genus Haemonchus (Nematoda: Trichostrongyloidea): considerations of their evolutionary history and global biogeography among Camelidae and Pecora (Artiodactyla). Journal of Parasitology. 90:1085-1102.
 
Hoberg, E.P., Lichtenfels, J.R., Rickard, L.G. 2004. Phylogeny for genera of Nematodirinae (Nematoda: Tichostronoylina). Journal of Parasitology. 91:382-389.

Hoberg, E.P. 2004. Coevolution and biogeography among Nematodirinae (Nematoda: Trichostrongylina) Lagomorpha and Artiodactyla (Mammalia): exploring determinants of history and structure for the northern fauna across the Holarctic. Journal of Parasitology. 91:358-369.

Hoberg, E.P., Abrams, A. 2004. Pseudostertagia bullosa (Nematoda: Trichostrongyloidea) in artiodactyl hosts from North America: Redescription and comments on systematics. Journal of Parasitology. 91:370-381.

Jenkins, E.J., Appleyard, G.D., Hoberg, E.P., Rosenthal, B.M., Kutz, S.J., Veitch, A.M., Schwantje, H.M., Elkin, B.T., Polley, L. 2004. Geographic distribution of the muscle-dwelling nematode Parelaphostrongylus odocoilei in North America, using molecular identification of first-stage larvae. Journal of Parasitology. 91:574-584.

Genetically Engineered Vaccines to Control Swine Influenza Virus (SIV)

ARS scientists successfully used reverse genetics to develop a novel cross-protective vaccine for swine flu. Swine influenza is a re-emerging disease around the world as a result of several genetic changes in the viral populations being isolated from swine. This has resulted in a reduced efficacy of current commercially available vaccines.  An experimental modified live swine influenza virus (SIV) vaccine was developed as part of a collaborative project with Mount Sinai School of Medicine and St. Jude's Children's Hospital.  Preliminary studies indicate this vaccine may have a broader level of cross protection when compared to currently available SIV vaccines that are inactivated or killed.  These studies have also demonstrated a virulence mechanism and will impact the design of future commercially available SIV vaccines.  (National Program 103 and Performance Measure 3.2.1)

Scientific Publications:

Solorzano, A., Webby, R.J., Lager, K.M., Janke, B.H., Garcia-Sastre, A., Richt, J.A. 2005. Mutations in the NS1 protein of swine influenza virus impair anti-interferon activity and confer attenuation in pigs. Journal of Virology. 79(12):7535-7543.

Wesley, R.D., Lager, K.M., Tang, M. 2004. Protection of weaned pigs by vaccination with human adenovirus 5 recombinant viruses expressing the hemagglutinin and the nucleoprotein of H3N2 swine influenza virus. Vaccine. 22:3427-3434.

New vaccine to prevent mastitis in dairy cows

ARS scientists completed a large study evaluating the efficacy of a trivalent vaccine in preventing new intramammary infections in first calve heifers.  This study required the development of a new challenge model and involved a total of 72 animals over a one year period.  Compared to a sham immunization, the trivalent vaccine demonstrated efficacy in protecting against S. aureus intramammary infection using a teat-dip challenge model.  Since S. aureus is responsible for ~40% of all cases of mastitis and annual economic losses of ~$800 million, the development of an efficacious vaccine should substantially reduce these economic losses to producers. (National Program 103 and Performance Measure 3.2.3)

Scientific Publications:

Bannerman, D.D., Eiting, K.T., Winn, R.K., Harlan, J.M. 2004. Flice-like inhibitory protein (flip) protects against apoptosis and suppresses nf-kb activation induced by bacterial lipopolysaccharide. American Journal of Pathology. 165(4):1423-1431.
 
Wall, R.J., Powell, A.M., Paape, M.J., Kerr, D.E., Bannerman, D.D., Pursel, V.G., Wells, K.D., Talbot, N.C., Hawk, H.W. 2005. Genetically enhanced cows resist intramammary staphylococcus aureus infection. Nature Biotechnology. 23(4):445-451.

Lee, J., Obrien, C.N., Guidry, A.J., Paape, M.J., Schafer-Weaver, K.A., Zhao, X. 2005. The effect of a trivalent vaccine against staphylococcus aureus mastitis on lymphocyte subpopulations, antibody production and neutrophil phagocytosis. Canadian Journal of Veterinary Research. 69(1):11-18.

New diagnostic test for Johne’s disease

ARS scientists developed and optimized a new PCR test for the detection of M. paratuberculosis in fecal samples using a newly identified gene sequence.  The gene sequence is an insertion element that is present in 6 copies within the genome.  This element, ISMap02, was utilized in a nested PCR assay in conventional and real-time formats.  The sensitivity of this assay was demonstrated to be equivalent to culture and comparable to the widely used IS900 gene element for M. paratuberculosis, which is present in 17 copies within the genome.  There have been concerns about the specificity of the IS900 element in the past.  Therefore, use of the ISMap02, which is highly specific, will provide a sensitive alternative for the detection of M. paratuberculosis in fecal samples.  This assay has also demonstrated efficacy for the detection of M. paratuberculosis in milk and tissues of infected animals. (National Program 103 and Performance Measure 3.2.3)

Scientific Publications:

Huntley, J., Whitlock, R., Bannantine, J.P., Stabel, J.R. 2005. Comparison of diagnostic detection methods for mycobacterium avium subsp.  paratuberculosis from North American bison. Journal of Veterinary Diagnostic Investigation. 42(1):42-51.

Stabel, J.R., Goff, J.P. 2004. Efficacy of immunolgic assays for the detection of Johne's disease in dairy cows fed additional energy during the periparturient period. Journal of Veterinary Diagnostic Investigation. 16(5):412-20.

International work on controlling Avian Influenza

As part of an international research effort, ARS scientists characterized a highly pathogenic outbreak of H5N1 from poultry from South Korea for its relationship to other outbreaks in the region and its potential for crossing the species barrier.  The South Korean outbreak was the first reported outbreak of H5N1 from the large epidemic that swept Asia in 2003-05, and the timely dissemination of information from this outbreak was important for understanding the disease outbreak in the region.  In conjunction with the Centers for Disease Control in Atlanta and the Veterinary Research and Quarantine Service in South Korea, we coordinated and helped perform the experimental studies in several animal species and of the sequencing of the South Korean virus to characterize this strain of avian influenza.  The scientific information from these studies were used by federal and foreign governments to understand that the South Korean outbreak was similar to but distinctly different from outbreaks that were simultaneously occurring in other Asian countries.  This suggested a multiple point source of introduction which helped shape the regulatory response to the outbreaks. (National Program 103 and Performance Measure 3.2.2)

Scientific Publications:
Lee, C.W., Suarez, D.L., Tumpey, T.M., Sung, H., Kwon, Y., Lee, Y., Choi, J., Joh, S., Kim, M., Lee, E., Park, J., Lu, X., Katz, J.M., Spackman, E., Swayne, D.E., Kim, J. 2005. Characterization of highly pathogenic H5N1 avian influenza a viruses isolated from korean poultry. Journal of Virology. 79:3692-3702.

 Nguyen, D.C., Uyeki, T.M., Jadhao, S., Maines, T., Shaw, M., Matsuoka, Y., Rowe, T., Lu, X., Hall, H., Balish, A., Klimov, A., Tumpey, T., Swayne, D.E., Huynh, L.T., Nghiem, H.K., Hguyen, H.T., Hoang, L.T., Cox, N.J., Katz, J.M. 2005. Isolation and characterization of avian influenza viruses, including highly pathogenic H5N1 viruses, from poultry in live bird markets in hanoi, vietnam - 2001. Journal of Virology, p.4201-4212.

Avian Influenza antibody detection assay

ARS scientists developed an ELISA test to detect antibodies against the non-structural (NS) protein of influenza virus that can be used a DIVA (differentiate infected from vaccinated animals) test. A limitation on use of killed avian influenza vaccines is the need to identify infected poultry within a vaccinated population. This is necessary to assess the success of a vaccination program, and to continue trade in poultry and poultry products.  In chickens vaccinated with killed experimental and commercial H7N2 vaccines, vaccinated or non-vaccinated chickens infected with the live virus had anti-NS antibodies while chickens receiving only the vaccines lacked anti-NS antibodies. These results demonstrate the potential benefit of a simple, specific ELISA to easily identify infected poultry within vaccinated populations, which will provide assurance to trading partners of a safe product. (National Program 103 and Performance Measure 3.2.3)

Scientific Publications:
Lee, C.W., Suarez, D.L. 2005. Avian influenza virus: prospects for prevention and control by vaccination. Animal Health Research Reviews. 6(1):1-15.

Determining the transmission of Avian Pneumovirus

Vertical transmission of APV from turkey hen to egg was demonstrated for the first time.  The possibility of vertical transmission has been suspected but never proven experimentally. Turkey hens were inoculated with APV and eggs harvested for reisolation of virus. Infectious virus was isolated from the embryo and genomic RNA was detected on the egg shell for up to 1 week post inoculation. These results demonstrate a previously uncharacterized mode of transmission of APV and underscore the need further diagnostic and vaccine development for control of APV. (National Program 103 and Performance Measure 3.2.1)

Scientific Publications:
Alvarez, R., Jones, L.P., Seal, B.S., Kapczynski, D.R., Tripp, R.A. 2004. Serological cross-reactivity among members of the metapneumovirinae genus. Virus Research. 105:67-73.

Kapczynski, D.R. 2005. Experimental infection of SPF laying turkeys with avian pneumovirus subtype c: possible role of vertical transmission. In:Proceedings of the American Association of Avian Pathologists, July 16-20, 2005, Minneasplis, Minnesota. p.34.

Lwamba, H.M., Alvarez, R., Wise, M., Yu, Q., Halvorson, D., Njenga, M., Seal, B.S.2005. Comparison of the Full Length Genome Sequence of Avian Metapneumovirus Subtype C With Other Paramyxoviruses Including Highly Virulent Newcastle Disease Viruses. Virus Research. 107(1):83-92.