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2018 Annual Report
Objectives
1) Development of intervention strategies to control and eradicate FMDV including vaccines and biotherapeutics/adjuvants that rapidly induce long lasting and cross-protective immunity against multiple FMDV subtypes, and are capable of preventing infection and controlling/abrogating persistent infections.
1.1) Development of novel FMD vaccine platforms.
1.1.1) Development of marker FMDV LL3B3D vaccines against relevant outbreak strains.
1.1.2) Development of improved second-generation Ad5-FMD vaccines.
1.1.3) Discovery of modified live attenuated FMDV vaccine candidates (MLAV).
1.1.4) Discovery of cross-protective vaccines against multiple FMDV subtypes.
1.2) Development of novel biotherapeutics to prevent or control FMD prior to vaccine-induced protection.
1.2.1) Discovery/development of novel biotherapeutics with increased potency and extended systemic half-life.
1.2.2) Evaluation of combined delivery of biotherapeutics and vaccine in swine and cattle.
1.3) Evaluation of vaccine-induced immunity and FMDV carrier state.
1.3.1) Characterization of host immunity associated with novel vaccines against FMDV.
1.3.2) Evaluation of novel vaccines for ability to prevent the FMD carrier state in cattle and assess the host response associated with the carrier divergence.
2) Elucidation of host-pathogen interactions of FMDV in acute and persistent infections. The information derived will be used to devise effective anti-viral intervention strategies.
2.1) Determine the molecular basis for FMDV-host interactions that impact virulence.
2.1.1) Examination of virus factors contributing to FMDV virulence.
2.1.2) Examination of host factors contributing to FMDV virulence.
2.2) Identification of molecular mechanisms associated with the establishment of FMDV persistence.
2.2.1) Determination of host and/or other non-FMDV factors causing or associated with clearance of FMDV from bovine nasopharyngeal tissue.
2.2.2) Investigation of within-host FMDV genomic evolution to characterize site-specific mutational pressure, genomic variation, and potential adaptation to the host.
2.3) Determination of the immune mechanisms affecting protective immunity against FMDV.
2.3.1) Analysis of CD4 helper T-cell response to FMDV vaccination.
2.3.2) Analysis of the CD8 cytotoxic T-cell response to FMDV vaccination.
2.3.3) Analysis of B-cell responses to FMDV in peripheral blood and lymphoid tissue.
3) Understanding the ecology of FMDV in endemic regions, determining drivers of transmission and maintenance in endemic settings, and characterizing risk factors driving FMDV emergence and spread.
3.1) Characterize the ecology of FMDV in endemic regions in Asia and Africa, including determining the factors driving viral transmission and maintenance.
3.2) Characterize factors driving FMDV emergence and spread of novel FMDV strains in endemic settings.
3.3) Role of Asian buffalo in maintenance and transmission of FMDV in endemic settings.
Approach
1. The development of intervention strategies to control and eradicate FMDV will be achieved through the development of novel FMD vaccine platforms: including of marker FMDV-LL3B3D vaccines against relevant outbreak strains, second–generation Ad5-FMD vaccines, the discovery of modified live-attenuated FMDV vaccine candidates, and the discovery of cross-protective vaccines against multiple subtypes. Novel biotherapeutics to prevent or control FMD prior to vaccine-induced protection will be based on the discovery/development of novel biotherapeutics with increased potency and extended systemic half-life. An evaluation of combined delivery of biotherapeutics and vaccine in swine and cattle will be conducted. To evaluate vaccine-induced immunity and FMDV carrier state the host immunity associated with novel vaccines against FMDV will be characterized. Novel vaccines will be evaluated for their ability to prevent the FMD carrier state in cattle and assess the host response associated with the carrier divergence.
2. The host-pathogen interactions of FMDV will be determined through: the identification of viral determinants of FMDV that control virulence in susceptible hosts, determining virus/host interactions associated with the FMDV life cycle, and determining the mechanisms of protective immunity to FMDV. The molecular basis for FMDV-host interactions that impact virulence and their specific contributions to virulence will be determined. The molecular mechanisms associated with the establishment of FMDV persistence will be identified through the determination of host and/or other non-FMDV factors causing or associated with clearance of FMDV from bovine nasopharyngeal tissue. The within-host FMDV genomic evolution will be characterized through an examination of site-specific mutational pressure, genomic variation and potential adaption to the host. The immune mechanisms affecting protective immunity against FMDV will be determined through the analysis of CD4 helper and CD8 cytotoxic T-cell responses to FMDV vaccination and B-cell responses to FMDV in peripheral blood and lymphoid tissue.
3. The characterization of the ecology of FMDV in endemic regions, including determining drivers of FMDV transmission and maintenance in endemic regions, characterizing factors driving FMDV emergence and spread, and the characterization of the role of the Asian buffalo in the transmission and maintenance of FMDV in the context of tolerance to infection will be analyzed. Efforts will focus on the characterization of the ecology of FMDV in endemic regions in Asia and Africa, including determining the factors driving viral transmission and maintenance. Factors driving FMDV emergence and spread of novel FMDV strains in endemic settings will be characterized. The role of Asian buffalo in maintenance and transmission of FMDV in endemic settings will be assessed.
Progress Report
This year was challenging for the Foreign Animal Disease Research Unit (FADRU) due to the lack of access to animal facilities and a significant shortage in personnel due to multiple personnel departures and hiring freeze. However, the remaining personnel stepped up to the multiple challenges and was able to achieve most of the milestones for the year either by establishing research collaborations overseas or with university partners. Under Objective 1 (countermeasures), significant progress was made in further development of the FMD3B3D platform with select agent exclusion and approval for transfer the vaccine to our commercial partner. Additional progress was made on a next generation Ad5 vector with more robust safety and increased antigen expression. However, these vectors are awaiting in-vivo testing. Likewise, new modified live vaccine candidates made using codon deoptimization await animal testing. A new DIVA ELISA test was also developed in partnership with APHIS (Animal Plant Health Inspection Service), DHS (Department of Homeland Security), Academia and commercial partners. Several Ad5-biotherapeutics vectors were made that are still awaiting in-vivo testing. Immune assessment of FMD vaccines and testing for prevention of persistent infection was not possible due to our immunologist vacancy and lack of animal facility. For Objective 2 (virus-host interactions) the exploration of MiRNA as a antiviral mechanism was greatly limited by the departure of an ARS scientist. However, significant progress was made in understanding the molecular mechanisms mediating viral persistence in cattle. Also, some progress was made on functional genomics especially understanding the role in viral virulence of features on the viral 5’ terminus (S-fragment). Important progress was made on Objective 3 (ecology) specially with collaborations overseas in Asia and Africa, where deeper understanding of the virus emergence and transmission was achieved. Overall ARS scientists made great progress in all three objectives overcoming significant limitations in personnel and animal facilities.
Accomplishments
1. Safe Foot-and-Mouth Disease vaccine. Currently, production of Foot and Mouth Disease (FMD) vaccines relies on the use of live virulent virus that can escape manufacturing facilities and cause outbreaks. Live FMD virus is not allowed on the U.S. mainland. Recently, ARS researchers in Orient Point, New York used reverse genetics to create an attenuated FMD virus that is innocuous to animals but can be safely used for vaccine manufacturing. Secretary of Agriculture has authorized the movement of this modified, non-virulent version of the FMD virus from the Plum Island Animal Disease Center to the U.S. mainland for the purposes of continued vaccine development and eventually manufacturing. Vaccine manufacturer Zoetis Inc. requested and has been granted a license for this vaccine technology. Identifying a vaccine that uses a modified virus will enable vaccine manufacturing in the USA and will allow the USDA-APHIS to more quickly source and acquire FMD vaccine for the Strategic Veterinary Stockpile and in the event of an outbreak of this devastating disease.
2. Understanding the mechanisms of the Foot-and-Mouth Disease virus (FMDV) carrier state in cattle. Control and eradication of foot-and-mouth disease (FMD) is impeded by the existence of a prolonged persistent phase of infection in ruminant species which is generally referred to as the FMDV carrier state. Researchers at ARS in Orient Point, New York have spent much of the last 10 years characterizing the mechanisms of persistence in the nasopharynx (throat) of cattle. Now we have taken that information and shown that clearance of FMDV infection was associated with activation of anti-viral T cell responses, whereas the antibody-mediate immune response was stronger in animals that maintained persistent infection. Additionally, mechanisms associated with programmed cell killing (apoptosis) were activated in animals that cleared infection whereas pathways associated with prolonged cell survival were upregulated in FMDV carriers. These findings are critically important for design of novel FMD vaccines that may prevent the occurrence of persistent FMDV infection. The study was published in Scientific Reports.
3. New rapid diagnostic Kit for Foot-and-Mouth Disease. Current foot-and-mouth disease virus (FMDV) vaccines are compatible with a strategy based on “differentiating infected from vaccinated animals” (DIVA). ARS researchers at Orient Point, New York as part of a large research consortium of federal agencies, academia and animal health industry, developed and licensed a rapid-response (three-hour) Foot-and-Mouth Disease (FMD) diagnostic kit. A manuscript describing the validation of this assay that led to the licensure of the FMD diagnostic kit was published in the Journal of Veterinary Diagnostic Investigation.
4. Increased the knowledge about Foot-and-Mouth Disease virus (FMDV) emergence and transmission. Although foot-and-mouth disease (FMD) is widely described as the most important disease affecting international trade in animal products, many aspects of the natural cycle of the causative virus remain unknown. ARS researchers in Orient Point, New York have been working for several years to develop research partnerships in endemic areas of Asia and Africa to describe the behavior of FMDV under natural conditions. Some of these studies have recently culminated with critical findings from work in India which have quantitated the speed of spread of FMDV through herds of cattle during outbreaks, and the duration that the virus persists in those herds. Similarly, ARS researchers performed a study in Cameroon which demonstrated that there are benefits to introducing vaccines to herds of cattle, even if the virus is already present. Additionally, genetic sequences were acquired from viruses discovered in Vietnam which helped to show how the virus evolves over time and space to continuously create new viruses. These findings are critically important to understand how FMDV spreads within the current disease range, and to prepare for the possibility of an outbreak in the USA.
5. Understanding the virus factors critical for Foot-and-Mouth Disease Virus (FMDV) replication. ARS researchers in Orient Point, New York have used novel approaches to derive attenuated FMDV strains using modifications at the 5’terminus of the viral genome. In proof of concepts studies identified a new role attributed to a small segment of the beginning of the FMDV genome that is responsible for virus replication and also modulating the innate immune response in animals to the virus. This knowledge of the virus-unique replication processes has potential for the development of disease control strategies such as better vaccines.
Review Publications
Stenfeldt, C., Eschbaumer, M., Smoliga, G.R., Rodriguez, L.L., Zhu, J.J., Arzt, J. 2017. Clearance of a persistent Picornavirus infection is associated with enhanced pro-apoptotic and cellular immune responses. Scientific Reports. 7:17800. https://doi.org/10.1038/s41598-017-18112-4.
Kloc, A., Rai, D.K., Rieder, A.E. 2018. The roles of picornavirus untranslated regions in infection and innate immunity. Frontiers in Microbiology. 20(9):485. https://doi.org/10.3389/fmicb.2018.00485.
Stenfeldt, C., Arzt, J., Pacheco, J., Gladue, D.P., Smoliga, G.R., Silva, E., Rodriguez, L.L., Borca, M.V. 2018. A partial deletion within foot-and-mouth disease virus non-structural protein 3A causes clinical attenuation in cattle but does not prevent subclinical infection. Virology. 516:115-126. https://doi.org/10.1016/j.virol.2018.01.008.
Kenney, M.A., Waters, R.A., Rieder, A.E., Pega, J., Perez-Filguera, M., Golde, W.T. 2017. Enhanced sensitivity in detection of antiviral antibody responses using biotinylation of foot-and-mouth disease virus (FMDV) capsids. Journal of Immunological Methods. 450:1-9. https://doi.org/10.1016/j.jim.2017.07.001.
Rekant, S., Lyons, M., Pacheco Tobin, J., Arzt, J., Rodriguez, L.L. 2016. Veterinary applications of infrared thermography. American Journal of Veterinary Research. 77:98-107. https://doi.org/10.2460/ajvr.77.1.98.
Ahmed, Z., Pauszek, S.J., Ludi, A., Larocco, M.A., Khan, E., Afzal, M., Arshed, M.J., Farooq, U., Arzt, J., Bertram, M., Brito, B., Naeem, K., Abubakar, M., Rodriguez, L.L. 2017. Genetic diversity and comparison of diagnostic tests for characterization of foot-and-mouth disease virus strains from Pakistan 2008-2012. Transboundary and Emerging Diseases. 65(2):534-546. https://doi.org/10.1111/tbed.12737.
Brito, B.P., Pauszek, S.J., Hartwig, E.J., Smoliga, G.R., Vu, L.T., Vu, P.P., Stenfeldt, C., Rodriguez, L.L., King, D.P., Knowles, N.J., Bachanek-Bankowska, K., Long, N.T., Dung, D.H., Arzt, J. 2018. A traditional evolutionary history of foot-and-mouth disease viruses in Southeast Asia challenged by analyses of non-structural protein coding sequences. Scientific Reports. 8:6472. https://doi.org/10.1038/s41598-018-24870-6.
Bertram, M.R., Delgado, A., Pauszek, S.J., Smoliga, G.R., Brito, B.P., Stenfeldt, C., Hartwig, E.J., Dickmu-Jumbo, S., Abdoulmoumini, M., Abona Olivia, A., Salhine, R., Rodriguez, L.L., Gerabed, R., Arzt, J. 2018. Effect of vaccination on cattle herds previously exposed to foot and mouth disease in Cameroon. Preventive Veterinary Medicine. 155:1-10. https://doi.org/10.1016/j.prevetmed.2018.04.003.
Eschbaumer, M., Stenfeldt, C., Pacheco Tobin, J., Rekant, S.I., Arzt, J. 2016. Effect of storage conditions on subpopulations of peripheral blood T lymphocytes from naive cattle and cattle infected with foot-and-mouth disease virus. Veterinary Clinical Pathology. 45:110-115. https://doi.org/10.1111/vcp.12327.
Hayder, S.S., VanderWaal, K., Ranjan, R., Biswal, J.K., Subramaniam, S., Mohapatra, J.K., Sharma, G.K., Rout, M., Dash, B., Das, B., Prusty, B., Sharma, A.K., Stenfeldt, C., Perez, A., Rodriguez, L.L., Arzt, J. 2017. Foot-and-mouth disease virus transmission dynamics and persistence in a herd of vaccinated dairy cattle in India. Transboundary and Emerging Diseases. 65(2):e404-e415. https://doi.org/10.1111/tbed.12774.
