Location: Animal Parasitic Diseases Laboratory
2019 Annual Report
Objectives
Highly infectious diseases of pigs present exceptional challenges to producers. Novel approaches are required to maintain animal health and welfare, particularly as scientists work to develop alternatives to the use of antibiotics for pathogen control. This project will explore immune and genomic-based approaches for understanding host-pathogen interactions. Probing the genetic variations associated with infection, immune evasion, innate and adaptive immune responses, and disease susceptibility and resistance will lead to improved animal health and alternatives for disease control and vaccine design. The goal of this research project is to develop effective countermeasures for preventing and controlling important respiratory diseases of pigs, such as Porcine Reproductive and Respiratory Syndrome (PRRS).
New genetic and immune markers will help producers and animal health professionals to prevent and control swine viral diseases; they will provide basic data to use for design of alternate control and vaccine strategies, thus decreasing production costs and improving trade potential. As a result of this work, animal health companies will have alternatives for discovering biotherapeutics and vaccines for swine respiratory diseases; pig breeding companies will have new tools to identify disease-resistant stock. Overall this project will stimulate advances in pig health that may be of broad economic importance.
Objective 1. Develop immunologic tools to evaluate swine immunity, including using immunological tools to enhance our understanding of swine immune system development [C4, PS4B], and using immunological tools to inform the design of novel innate immune intervention strategies to treat respiratory diseases of swine. [C2, PS2B]
Objective 2. Elucidate host response associated with swine respiratory disease and protective immunity, including discovering genetic and biological determinants associated with swine respiratory disease susceptibility, tolerance, or resistance, and discovering genetic and biologic determinants associated with good responders to swine respiratory disease vaccines. [C4, PS4B]
Approach
Characterize swine immune proteins (cytokines, chemokines) and monoclonal antibodies (mAbs) to these proteins and their receptors, and to antigens that define swine cell subsets and activation markers (CD antigens). To speed progress on reagent development, collaborate with commercial partners for protein expression and mAb production. Coordinate ARS efforts with NIFA supported US UK Swine Toolkit progress. Once panels of mAbs reactive with swine targets are available, test them for specificity and identify epitope reactivities to help develop sandwich ELISAs and Bead Based Multiplex Assays (BBMA). Work with USDA ARS and NIFA leadership to establish a veterinary immune reagent repository for relevant hybridomas and cell lines from various livestock species as well as to provide an updated website www.vetimm.org highlighting the availability of these reagents.
Emerging and re-emerging infectious diseases heighten the need to use the expanded swine toolkit to facilitate veterinary and biomedical research. Complex immune interactions determine the efficacy of a pig's response to infection, vaccination and therapeutics. New tools developed through this project, and the US UK Swine Toolkit grant, will expand options for probing mechanisms involved in disease and vaccine responses. Continue to assess samples archived through the PRRS Host Genetics Consortium (PHGC) for protein and metabalome alterations that may be predictive of PRRS viral levels or weight gain changes at different time-points post infection. Expand efforts to use pig as an important biomedical model including tuberculosis (TB) research. For TB test whether vaccination in neonatal minipigs leads to the development of immune responses similar to those described in human infants. Results from these trials will allow study of infant TB and TB vaccine efficacy. address biomedical
Following up on PHGC studies, as part of a USDA NIFA translational genomics grant, a more complex model, testing vaccination for PRRS followed by PRRSV and porcine circovirus (PCV) challenge [a PCV associated disease (PCVAD) model] was pursued. This approaches typical farm conditions and enables us to ask about the effectiveness of vaccination prior to PRRSV and PCV2 challenge. Additionally, data was collected on genetically defined pigs in true field trial conditions, providing data that is essential for transfer (and affirmation) of our disease genetic results to pig breeders. We expect that the combined models and genomic approaches will lead to identification of chromosomal regions, putative candidate genes and mechanisms involved in regulating pig responses to viral infections, vaccinations, and associated growth effects.For this ARS project we will evaluate the effect of anti-viral response pathways and biomarkers on vaccine and infection responses.
We will use RNAseq analyses to provide a more complete picture and reveal details of regulatory mechanisms impacting pig responses to vaccination, viral infection, and differential growth effects. Our proposed studies will expand analyses of samples collected on the grant funded 4 vaccination/PCVAD trials and 6 field trials (Appendix). As we identify
Progress Report
Progress was made on both project objectives and their subobjectives, all of which fall under National Program 103, Animal Health. They address NP103 Component 4: Respiratory Diseases, Problem Statement 4B: Porcine Respiratory Diseases and NP103 Component 2: Antimicrobial Resistance, Problem Statement 2B: Alternatives to Antibiotics.
All milestones were “Met” or “Substantially met” in FY19. Under Objective 1, swine immune proteins (cytokines, chemokines, transcription factors) were cloned and expressed by commercial partners; monoclonal antibodies (mAbs) to some of these proteins were developed by a second commercial partner and then characterized in collaboration with university partners. Once characterized these reagents are being transferred to commercial partners. These reagents are essential for advancing international pig health and vaccination research efforts, which require a broad range of immune reagents; unfortunately, immune reagents are not widely available for pigs. Refined molecular technologies, including NanoString arrays, Iso-seq, and RNAseq, and advanced biostatistical analyses were used to more effectively assess expression of important genes controlling immune responses to porcine reproductive and respiratory syndrome virus (PRRSV) infection and vaccination.
Under Objective 2, genetic and biological determinants associated with porcine reproductive and respiratory syndrome virus (PRRSV) infection were explored, expanding on the previously discovered swine chromosome 4 (SSC4) genetic viral resistance allele. Genetic and immune factors controlling PRRSV persistence in tonsil were analyzed. An expanded coinfection [PRRSV and porcine circovirus (PCV2)] model of swine respiratory disease was used to assess susceptibility and resistance as well as identify determinants associated with good vaccine responses. A model of reproductive PRRS has been evaluated to determine how fetuses resist congenital infection.
Accomplishments
1. A new Blueprint for Animal Genomics. Over the last decade, USDA funded scientists and their colleagues from across the globe used the 2008-2017 Genomic Blueprint to develop an animal genomics infrastructure that facilitated novel scientific discoveries, many of which have been implemented into commercial production and led to returns that far exceeded investment costs. The newly-published 2018-2027 Genomic Blueprint will continue this trajectory based on new technologies, new insights into animal biology, and new genome-enabled strategies that improve various aspects of animal production. Ultimately, animal genome technologies will become part of mainstream agricultural production strategies and used to improve animal health, well-being, production efficiency, and product quality in ways that meet the demands of growing global populations. Because the Nation values technology and innovation, the animal genomics community is well-positioned to address current and future agricultural challenges facing the planet.
2. Predicting PRRS viral persistence. Porcine Reproductive and Respiratory Syndrome (PRRS) is a globally important swine disease from both an economic and animal welfare standpoint. It has been documented to persist in tonsils for as long as 150 days. ARS scientists in Beltsville, Maryland, partnered with Iowa State and Kansas State University researchers to identify genomic regions controlling tonsil virus levels. Pigs infected with 2 PRRSV isolates did not pinpoint genes, but innate immune pathways and differential cell recruitment were found to be critical factors affecting PRRSV persistence. Importantly, the level of tonsil virus was found to be associated with traits related to viral clearance, i.e., pigs with low tonsil virus levels had an earlier and faster rate of maximal serum viral clearance; they also exhibited a stronger tonsil inflammatory and cell-mediated immune response. Thus, alternate immune mechanisms may be critical for reducing PRRSV persistence in the tonsil. These data may assist pig breeders, veterinarians, and pork producers better understand and manage this widespread problem.
3. Serum antibodies contribute to PRRS control. Porcine reproductive and respiratory syndrome virus (PRRS) is the most economically important disease of pigs in the U.S. and abroad. ARS scientists in Beltsville, Maryland, partnered with Iowa State and Kansas State University researchers to seek ways of predicting which pigs fare best against this infection. They found that the level of serum anti-PRRS antibody response 42 days after being infected predicted decreased blood viral level and improved weight gain. A region in the swine major histocompatibility complex was associated with antibody response level. Even lacking that precise information, breeders are using this discovery to select pigs with increased viral infection resistance, providing another tool for improving swine health.
4. New tools for swine immunology. Sophisticated immune tools are needed to analyze disease and vaccine responses, yet those for pigs are limited. ARS scientists at Beltsville, Maryland, worked with a commercial partner to clone and express immune proteins using yeast. A second commercial partner developed monoclonal antibodies (mAbs) to these proteins which were then characterized in collaboration with university partners at Ohio State and Tennessee State Universities. Panels of mAbs reactive with 7 different swine immune proteins were characterized and transferred to commercial partners. These antibodies provide new assays to quantitate each protein in body fluids as well as to measure their intracellular expression. Tools and reagents generated by this project are now available and being used, the world-over, for studying swine immunity and disease.
Review Publications
Hess, A.S., Lunney, J.K., Abrams, S., Choi, I., Trible, B., Hess, M.K., Rowland, R., Plastow, G., Dekkers, J. 2018. Identification of factors associated with virus level in tonsils of pigs experimentally infected with Porcine Reproductive and Respiratory Syndrome virus. Journal of Animal Science. 97:536-547. https://doi.org/10.1093/jas/sky446.
Hess, A.S., Trible, B., Hess, M.K., Rowland, R., Lunney, J.K., Plastow, G., Dekkers, J. 2018. Genetic relationships of antibody response, viremia level and weight gain in pigs experimentally infected with porcine reproductive and respiratory syndrome virus. Journal of Animal Science. 96:3565-3581. https://doi.org/10.1093/jas/sky229.
Rexroad III, C.E., Vallet, J.L., Matukumalli, L.K., Ernst, C., Van Tassell, C.P., Cheng, H.H., Reecy, J., Fulton, J., Taylor, J., Lunney, J.K., Liu, J., Cockett, N., Smith, T.P., Van Eenennaam, A., Clutter, A., Telugu, B., Purcell, C., Bickhart, D.M., Blackburn, H.D., Neibergs, H., Wells, K., Boggess, M.V., Sonstegard, T. 2019. Genome to phenome: improving animal health, production, and well-being: a new USDA blueprint for animal genome research 2018–2027. Frontiers in Genetics. 10:327. https://doi.org/10.3389/fgene.2019.00327.
