Location: Animal Parasitic Diseases Laboratory
2024 Annual Report
Objectives
Objective 1: Develop immunologic tools to evaluate swine immunity.
Sub-objective 1.A: Develop immunological assays and validate their use for assessing host immune responses to infectious diseases and vaccination.
Sub-objective 1.B: Use immunological tools to inform the design of novel innate immune intervention strategies to treat respiratory diseases of swine.
Objective 2: Elucidate immune and genomic determinants of host responses associated with porcine reproductive and respiratory syndrome virus infection.
Sub-objective 2.A: Discover genetic and biological determinants associated with fetal susceptibility, resilience, or resistance to porcine reproductive and respiratory syndrome.
Sub-objective 2.B: Discover genetic and biological determinants associated with good responders to porcine reproductive and respiratory syndrome vaccines.
Approach
This project will produce new reagents for analyses of swine immunity. This will include characterization of monoclonal antibodies (mAbs) reactive with swine cytokines and chemokines and development of assays that quantify their reactivity. These efforts will be coupled with NIFA grant supported Swine Toolkit efforts. Once characterized commercial access to these mAbs will be coordinated through ARS Technology Transfer Office. Basic studies will be expanded by establishing molecular tools to evaluate gene expression using NanoString Technologies, with codesets developed to explore relative importance of specific tissues and unique cell subsets in disease responses and pathogen resistance. These tools will help to identify alternate biomarkers for risk analyses to assess the impact of respiratory diseases and vaccine efficacy.
This project will apply the use of these innovative, next-generation, genomic, transcriptomic, and immunologic techniques to address Porcine Reproductive and Respiratory Syndrome (PRRS), focusing on our unique sample resources collected via the PRRS Host Genetics Consortium and the Canadian Pregnant Gilt Model projects. We will evaluate the influence of host genetics on antiviral response pathways and will aim to identify critical biomarkers of protective vaccine and infection responses. We will use RNAseq data analyses to provide a more complete picture and reveal details of regulatory mechanisms impacting pig responses to vaccination, viral infection, and differential growth effects. Advanced bioinformatics will be combined with new molecular studies, as well as previously collected genotypic and phenotypic data, to identify pathways and biomarkers associated with protective immune responses to respiratory and reproductive diseases that cause significant losses to the pork industry.
This project will provide alternate resources for improved respiratory disease control and elimination strategies. Overall, our project will inform vaccine evaluation and virus control strategies, identify disease-resistant pigs, and stimulate advances that may be of broad economic importance to pig breeders and the animal health industry.
Progress Report
Under Objective 1, the focus is on the development of new reagents for swine immunity; which are important for scientists to document the effectiveness of vaccine and disease responses and for basic porcine biomedical research efforts worldwide. Researchers at Beltsville, Maryland, characterized panels of monoclonal antibodies (mAbs) reactive with swine immune proteins (cytokines, chemokines) provided by a commercial partner. In collaboration with university partners, we tested tissues and stimulated cells for mAb staining and developed assays for quantifying native proteins using pairs of mAbs. The best mAbs were commercialized and shared with colleagues internationally.
Under Objective 2, the focus is on the development of focuses on porcine reproductive and respiratory syndrome (PRRS) which causes an estimated $600 million in annual losses to the U.S. pig industry. Scientists in Beltsville, Maryland, worked with university researchers to probe responses to PRRS virus infection transmitted from the sow to her fetuses late in gestation. Fetal anti-viral responses vary greatly within the litter; litters can include uninfected fetuses, those harboring high viral levels, and neighboring dead fetuses. We concentrated on fetal factors that predict disease severity and death or, instead, fetal resistance to infection. Using tissues (kidney, heart and liver) collected from fetuses of late term PRRS virus infected sows, we used focused molecular tests (NanoString codesets) to verify RNA gene expression. We identified specific genes expressed by resilient fetuses; these provide us with biomarkers that may help select for PRRS-resistant breeding stock.
Accomplishments
1. Discovery of genes that protect fetal pigs from viruses. Porcine reproductive and respiratory syndrome virus (PRRSV) infection harms many late term swine fetuses, but some fetuses of infected mothers fare much better than others. Researchers in Beltsville, Maryland, worked with university researcher to identify genes characteristic of those faring better despite their exposure to the virus. Increase expression of the gene CCL5 in fetal heart and kidney suggest improved immune function, and decreased gene expression of ACE2 in kidney may indicate improved blood pressure regulation. Fetal liver underwent the broadest changes in gene expression, including changes in angiotensin II receptor type 1 (AGTR1) which suggest effects on blood pressure regulation, in several transporter genes altering energy conversion, in transforming growth factor beta 1 (TGFB) affecting growth modulation, and in interleukin-10 (IL10) and C-X-C motif chemokine ligand 10 (CXCL10) suggesting effects on liver immune response. These results shed light on mechanisms of fetal protection against maternal PRRSV infection, the costliest disease for pork producers, and provide targets for breeding healthier piglets.
2. Better reagents to identify the swine version of an important component of immune signaling. The veterinary community requires commercial reagents and standards for studying swine immunity. Reagents for the human version of an important immune signaling chemokine (the C-X-C motif chemokine ligand 10, or CXCL10) don’t work well to identify the version in pigs. Scientists in Beltsville, Maryland, worked with commercial suppliers to express the recombinant porcine protein and generate monoclonal antibodies that bind well to this molecule. One proved especially effective for intracellular staining, whereas another proved especially effective when staining pig lymph nodes and spleen. By making these reagents generally available, clinicians and researchers can now better understand swine responses to infectious diseases and vaccination.
3. A repository for livestock immunology reagents. Decades of effort by researchers in Beltsville, Maryland, and their collaborators discovered monoclonal antibodies reactive against many immune proteins (cytokines and chemokines) from livestock. Unfortunately, their limited commercial market threatens their long-term availability to clinicians, researchers, and pharmaceutical companies who might benefit from them. Therefore, ARS scientists in Beltsville, Maryland, and university colleagues surveyed their commercial availability and devised a plain for long-term maintenance and storage of hybridomas and monoclonal antibodies derived from cattle, sheep, swine, chicken immune proteins are needed. Once implemented, this plan will ensure future benefits to animal health, welfare, and public health.
4. A transformative agricultural research framework. Various government agencies and philanthropies seek to transform agriculture and food systems in ways that demand researchers to overcome traditional research “silos” through deep and sustained systemic change. Therefore, ARS scientists at Beltsville, Maryland, identified intersections in federal and global initiatives encompassing climate adaptation and mitigation; human health and nutrition; animal health and welfare; food safety and security; and equity and inclusion. They then devised a research framework to catalyze agricultural transformation, proposing that transformation in agricultural research should incorporate (1) innovation, (2) integration, (3) implementation, and (4) evaluation. This framework seeks to facilitate sustainable, flexible, and coordinated transformation in the agricultural sector.
Review Publications
Hailstock, T., Dai, C., Aquino, J.F., Walker, K.E., Chick, S.T., Manirarora, J.N., Suresh, R., Patil, V., Renukaradhya, G.J., Sullivan, Y., Labresh, J., Lunney, J.K. 2023. Production and characterization of anti-porcine CXCL10 monoclonal antibodies. Cytokine. 174: Article e156449. https://doi.org/10.1016/j.cyto.2023.156449.
Boyd, A., Luo, Y., Lunney, J.K., Kustas, B., Fukagawa, N.K., Mattoo, A.K., Crow, W.T., Pachepsky, Y.A., Kim, M.S., Lillehoj, H.S., Van Tassell, C.P., Zhang, H.Q., Blomberg, L., Dubey, J.P. 2023. Cross-cutting concepts to transform agricultural research. Frontiers in Sustainable Food Systems. 7. Article e1242665. https://doi.org/10.3389/fsufs.2023.1242665.
Walker, K.E., Pasternak, J., Jones, A., Mulligan, M., Van Goor, A.G., Harding, J., Lunney, J.K. 2024. Gene expression in heart, kidney, and liver identifies mechanisms involved in fetal resilience to congenital PRRSV infection. Veterinary Microbiology. 295(1): Article e110154. https://doi.org/10.1016/j.vetmic.2024.110154.