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United States Department of Agriculture

Agricultural Research Service

Research Project: FOOT-AND-MOUTH DISEASE VIRUS (FMDV) HOST-PATHOGEN INTERACTIONS
2011 Annual Report


1a.Objectives (from AD-416)
1: Develop a defined model to identify early (pre-viremic) events in the pathogenesis of FMDV in its natural hosts. 2: Develop tools to evaluate host-pathogen interactions at the genomics level. 3: Identify critical FMDV virulence determinants associated with mechanisms of invasion, infection, replication, persistence and transmission. 4: Understand the mechanisms of FMDV persistence and the viral carrier state in animals. 5: Determine mechanisms of FMDV interference with host responses.


1b.Approach (from AD-416)
1: A defined model will be accomplished by developing and validating an inoculation model that mimics natural FMDV infections in target hosts.

2: Development of tools to evaluate the host-pathogen interaction will be accomplished through the design and development of a whole genome bovine microarray.

3: Identification of virulence determinants will be accomplished by: a.) Developing molecular tools to assess viral virulence factors, b.) Identify molecular events during FMDV-receptor interactions, c.) Identify components of the FMDV replication complex and the role of viral NSPs (non-structural proteins) in virus replication and, d.) Determine the function of the FMDV 5' and 3' untranslated regions.

4: An understanding of FMDV persistence in the carrier state will be accomplished by: a.) Establishment and characterization of persistently infected cells, b.) Characterization of virus released from persistently infected cells and, c.) Determine the role of autophagy in persistent infection.

5: Determination of FMDV interfernce with host responses will be completed by: a.) Examining the interference with the innate response in FMDV-infected cells and, b.) Examining the mechanisms of FMDV immune evasion.


3.Progress Report
In FY 2011, studies continued on understanding the interactions between Foot-and-Mouth Disease Virus (FMDV) and its hosts using classic pathology, computational biology, functional genomics and immunological approaches. Utilizing cDNA-derived attenuated viral strains we studied viral interaction with the target tissues and showed that low viral loads combined with differential patterns of cytokine expression consistently accompanied infection with the attenuated viruses and may hold the key to understanding the basis for virulence of FMDV. Some of these attenuated strains are being tested as vaccine candidates under RESEARCH PROJECT 053. Microarray technology was used to determine the basis for tissue tropism of FMDV and showed differential expression of genes involved in innate response and viral uptake in targeted tissues. Continuing earlier work with the leader protein, we created new mutants in a FHA domain of the leader protein and showed that unlike wild type FMDV, the FHA- mutant does not induce degradation of NF-kappaB, a critical step in FMDV pathogenesis and causes reduced severity of disease in swine. Efforts will continue to translate this research into more potent and effective vaccine candidates. We continued to characterize host proteins interacting with FMDV genome. We determined that RNA-binding protein Sam68 binds to 5’ untranslated region of FMDV and interacts with viral protein 3C. These interactions play an essential role in the replication of FMDV. We initiated preliminary studies using a yeast-two-hybrid technique to analyze the interaction of FMDV structural and non-structural proteins among themselves and with host proteins. Initial results identified a number of newly recognized protein interactions that play roles in FMDV replication. We analyzed cellular immune responses in swine following infection with FMDV and demonstrated induction of cytotoxic T lymphocytes (CTLs). Using a novel technology (MHC tetramers) we demonstrated that FMDV infected swine, develop a specific, major histocompatibility complex (MHC) restricted CTL response to FMDV in vivo. This is the first report to directly measure FMDV specific CTL activity and shows FMDV does not block the CTL response. Under an EPA supported agreement we developed methodology for testing the virucidal effect against FMDV, CSFV, and ASFV of various disinfectants on porpus and non-porous surfaces. In FY12 we will continue this work and will use the information in developing better FMD countermeasures. This research addresses project objectives:.
3)Identify critical FMDV virulence determinants associated with mechanisms of invasion, infection, replication, persistence and transmission of FMDV,.
4)Understand the mechanisms of FMDV persistence and the viral carrier state in animals and .
5)Determine mechanisms of FMDV interference with the host response and addresses the biodefense research of foreign animal diseases through (1) the development and/or improvement of surveillance tools, (2) vaccine development, (3) improved immunological resistance to infection.


4.Accomplishments
1. Attenuated novel strains of foot-and-mouth disease virus are being developed as safer and more effective vaccines. ARS researchers at PIADC, Greenport, NY developed foot-and-mouth disease (FMD) viruses with changes in specific genome regions that determine virulence. These strains, with rationally introduced attenuating mutations and specific antigenic markers, are harmless when tested in animals but have the capacity of inducing immunity. Furthermore, the addition of antigenic markers allows the differentiation of vaccinated from infected animals which is a critical advantage for field deployment. These attenuated strains are currently being tested as safer platforms for FMD vaccine production through a CRADA with an industry partner. Once adopted by industry this new technology will have a tremendous impact in reducing the cost and increasing the adequacy of commercial FMD vaccines.

2. Identified gamma delta T cells that provide early immunity to Foot-and-Mouth Disease Virus (FMDV) infection. FMDV infection in cattle is highly acute, with rapid onset, rapid clearance of virus from the blood and resolution of disease. The mechanisms mediating rapid clearance of the virus from blood are poorly understood. ARS researchers at PIADC, Greenport, NY, in collaboration with scientists from the Warsaw University of Life Sciences in Poland have now shown that during the 5 days following infection of cattle, a subset of lymphocytes, called gamma/delta T cells, are activated to kill FMDV infected cells and potentiate the immune response. These findings are being applied to the development of more effective FMD countermeasures.

3. Developed methodologies to test and identify disinfectants against foreign animal disease viruses. Disinfection is a critical step to limit the spread and accelerate recovery from foreign animal disease (FAD) outbreaks. Little is known about the efficacy of disinfectants to inactivate FAD viruses such as foot-and-mouth disease (FMDV), African swine fever (ASFV) and classical swine fever (CSFV), on agriculturally relevant surfaces. ARS researchers at PIADC, Greenport, NY, under an interagency agreement with the Environmental Protection Agency, developed standardized methodologies to test disinfectant efficacy on nonporous and porous surfaces that mimic those found at farms and facilities in the field. Using they identified bleach, and citric acid as good disinfectants on nonporous surfaces, but citric acid was more effective in porous surfaces. This information, provided to USDA-APHIS and EPA is currently used in the decision-making process on decontamination during emergency response to FAD virus outbreaks.


Review Publications
Maree, F.F., Blignaut, B., Aschenbrenner, L., Burrage, T., Rieder, A.E. 2010. Analysis of sat type foot-and-mouth disease virus capsid proteins: influence of receptor usage on the properties of virus particles. Virus Research. 155:462-472.

Arzt, J., Pacheco Tobin, J., Rodriguez, L.L. 2010. The early pathogenesis of foot-and-mouth disease in cattle after aerosol innoculation: identification of the nasopharynx as the primary site of infection. Veterinary Pathology. 47(6):1048-1063.

O'Donnell, V., Pacheco Tobin, J., Larocco, M.A., Burrage, T., Jackson, W., Rodriguez, L.L., Borca, M.V., Baxt, B. 2010. Foot-and-mouth disease virus utilizes an autophagic pathway during viral replication. Virology. 410:142-150.

Piccone, M.E., Diaz-San Segundo, F., Kramer, E., Rodriguez, L.L., De Los Santos, T.B. 2011. Introduction of tag epitopes in the inter-AUG region of foot and mouth disease virus: effect on the L protein. Virus Research. 155(1):91-97.

Rodriguez, L.L., Gay, C.G. 2011. Development of vaccines toward the global control and eradication of foot-and-mouth disease. Expert Review of Vaccines. 10(3):377-387.

Toka, F.N., Kenney, M.A., Golde, W.T. 2011. Rapid and transient activation of gamma/delta T cells to interferon gamma production, NK cell-like killing and antigen processing during acute virus infection. Journal of Immunology. 186(8):4853-4861.

Brito, B.P., Perez, A.M., Konig, G.A., Cosentino, B., Rodriguez, L.L. 2011. Factors associated with within-herd transmission of serotype A foot-and-mouth disease virus during the 2001 outbreak in Argentina: a protective effect of vaccination. Transboundary and Emerging Diseases. 58(5):387-393.

Pacheco Tobin, J., Piccone, M.E., Rieder, A.E., Pauszek, S.J., Borca, M.V., Rodriguez, L.L. 2010. Domain disruptions of individual 3B proteins for foot-and-mouth disease virus do not alter growth in cell culture nor virulence in cattle. Virology. DOI: 10.1016/j.virol.2010.05.036.

Maree, F.F., Blignaut, B., De Beer, T.A., Visser, N., Rieder, A.E. 2011. Mapping of amino acid residues responsible for adhesion of cell culture-adapted foot-and-mouth disease SAT type viruses. Virus Research. 153(1):82-91.

Arzt, J., Juleff, N., Zhang, Z., Rodriguez, L.L. 2011. The pathogenesis of foot-and-mouth disease I; viral pathways in cattle. Transboundary and Emerging Diseases. 58(4):291-304.

Krug, P.W., Lee, L.J., Eslami, A.C., Larson, C.R., Rodriguez, L.L. 2011. Chemical disinfection of high-consequence transboundary animal disease viruses on nonporous surfaces. Biologicals. 39(4):231-235.

Arzt, J., Baxt, B., Grubman, M.J., Jackson, T., Juleff, N., Rhyan, J., Rieder, A.E., Waters, R., Rodriguez, L.L. 2011. The pathogenesis of foot-and-mouth disease II; viral pathways in swine, small ruminants, and wildlife, myotropism, chronic syndromes, and molecular virus-host interactions. Transboundary and Emerging Diseases. 58(4):305-326.

Pedersen, L.E., Harndahl, M.N., Rasmussen, M., Lamberth, K., Golde, W.T., Lund, O., Nielsen, M., Buus, S. 2011. Porcine major histocompatibility complex (MHC) class I molecules and analysis of their peptide-binding specificities. Immunogenetics. 63(12):821-831.

Maree, F.F., Blignaut, B., Esterhuysen, J.J., De Beer, T., Theron, J., O'Neill, H.G., Rieder, A.E. 2011. Predicting antigenic sites on the foot-and-mouth disease virus capsid of the South African Territories (SAT) types using virus neutralization data. Journal of General Virology. 92(10):2297-2309.

Last Modified: 12/18/2014
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