Contributions of large and agricultural animal models to immunology

Abstract

While studies with laboratory rodent models have defined molecular and cellular components of the human immune system, experiments in these small mammals can’t capture all aspects of human immunity. This review focuses on immunologic research in large and agricultural species, highlighting how understanding immunity in a range of large animals can expand awareness of immunological processes critical for maintenance of human health. Large animal studies are reviewed and their unique contributions to knowledge of infectious disease spread, antibody production and vaccine development, organ transplantation, and immunotherapies are demonstrated. The review concludes with a brief discussion of limits of large models and their importance not only to the health of humans, but to agricultural animals themselves and human food systems.

Graphical Abstract

Introduction

Mouse models predominate the study of immunology and immunologic diseases for practical reasons. Maintaining mice is relatively low cost, mice have short life spans and reproduce rapidly, and genetic manipulation techniques are well established in laboratory strains. Additionally, many well-characterized and validated reagents are available for mouse studies, providing researchers with an extensive tool kit.

While it is less common to study immunology in large animals or agricultural species including pigs, sheep, goats, cows, horses, llamas, rabbits, and chickens than it is to use small rodents, characteristics of these animals can provide insights into the immunology of humans that cannot be captured using mouse models. Historically, this has been exemplified by Peter Medawar’s notion of acquired immunologic tolerance and description of the paradox of the fetal allograft from observations of dizygotic twins in cattle.^1,2 Although his original hypotheses have not stood the test of time, the work of many researchers testing them has led to much of what is known about maternal-fetal tolerance today. In recent years, details of maternal-fetal tolerance have been clarified using equine models, as horse embryos have unique features that make them amenable to pregnancy and transplant studies.^3,4 The chicken was instrumental in the identification of 2 classes of lymphocytes; thymic T-lymphocytes responsible for graft rejection and cell-mediated immunity, and antibody-producing B-lymphocytes derived from the bursa of Fabricius in birds and bone marrow in other vertebrates.⁵ Additionally, experiments in chickens and quail definitively established the hematopoietic stem cell origin of lymphocytes, an observation that was extended to all vertebrates, including humans.⁵

It has become recognized that agricultural species exhibit greater genetic diversity than most strains of experimental mice, thus more accurately represent the genetic diversity of humans.⁶ Farm animals, like cows, also share more homologous genes with humans than mice do.⁷ With the collective identification and quantification of large groups of biologic molecules, epigenomic, transcriptomic, proteomic, metabolomic, and other -omics data sets can be combined with classical genomic information to characterize specific immunologic pathways and processes. Multi-omics data from studies in model organisms has the potential to address clinical problems at a systems level that previously was not possible.⁸ Omics databases from large and agricultural animals are increasingly available and can be used to enhance experimental data generated in targeted studies designed to answer immunologic questions that are relevant to human health.^9–11

Additional benefits of agricultural animal models in immunology are that farm animals large, allowing for experimental designs in which “controls” and “treatments” can be tested in the same individual, and for collection of adequate numbers of cells and tissues for extensive downstream analysis. This feature has been capitalized on for experiments designed to test immunotherapies for skin wound healing and bacterial associated tissue damage.¹² The size and physiology of organs from agricultural animals is generally more representative of human counterparts than mouse organs, making them ideal models in which to study organ transplantation, and in some cases to serve as organ donors to humans.^13,14 Unique anatomic features of antibodies produced by certain agricultural animals can be leveraged to develop antibody therapies and vaccines for humans. Certain human diseases that cannot be reproduced in mouse models can be studied in large animals, as can the kinetics of a range of naturally occurring immunologic diseases and specific cancers.^13,15 Naturally occurring zoonotic diseases of agricultural animals are of particular importance to human health, as breakthrough cases can lead to human infection.

Finally, immunologic studies in large and agricultural animals benefit not only humans, but the health of animals and human food systems as well. This is in alignment with the global One Health approach, which postulates that general human public health, animal health, and environmental health are interconnected and recognizes that collaboration of experts across disciplines is critical to understanding the dynamic relationships that exist between humans, animals and the world we live in.¹⁶ Likewise, coordination of data collected by human and veterinary immunologists is needed to maximize human, animal, and food system health.

Agricultural animals contribute to knowledge of infectious disease

One way in which agricultural animals contribute to what is known about immunology in humans is by advancing our understanding of infectious diseases, especially those with a zoonotic character. Humans and agricultural animals are outbred populations with immune systems that have evolved in response to exposure to genetically related pathogens¹⁷ and multiple bacterial and viral diseases have a history of transfer between farm animals and humans.

Zoonotic infectious diseases

In 2021, experts in the fields of livestock and human infectious diseases gathered at an emerging animal infectious disease conference at Pennsylvania State University. Their primary objective was to outline a plan based on lessons learned from past experiences with high-consequence animal infectious diseases to inform preparations for future epizootics and panzootics.¹⁸ The overall conclusion was that the biosecurity of livestock operations is critical for understanding and minimizing the negative impacts of emerging animal infectious diseases. Recommendations were made to develop next-generation diagnostic tools to not only rapidly detect pathogens in clinical samples, but to detect variants of known pathogens and identify novel pathogens. The importance of a One Health approach for working with animal and human infectious diseases was stressed as well, recognizing the interconnections between people, animals, plants and our shared environment.¹⁸

Monitoring zoonotic infections in animals can shed light on infection patterns in humans and provide data to manage pathogens in the human population. The community-associated Clostridium difficile RT078 lineage has emerged as a human diarrheal pathogen characterized by highly transmissible spores and the acquisition of antimicrobial resistance genes (AMGs). It is isolated from both humans and farm animals, but transmission networks between species are not fully defined. Phylogenetic analysis of 248 C. difficile RT078 strains from 22 countries demonstrated extensive co-clustering of agricultural animal and human strains, suggesting a highly linked cross-species transmission network. Comparative whole-genome analyses revealed homologous accessory genomes in human and animal strains, including a variety of AMGs and the authors speculated that the bidirectional spread of C. difficile RT078 between agricultural animals and humans also provides a route for the circulation of AMGs through species and proposed a comprehensive approach to monitoring and managing C. difficile.¹⁹

Bacterial infections such as zoonotic tuberculosis, Leptospirosis, and Q fever can be transmitted from farm animals to humans through contact with sick animals, animal products or inhaled aerosols.^20–22 Genetic modification of livestock is being explored to reduce the prevalence of zoonotic disease in animals, with the goal of decreasing human infection. In 1 study, a homology-mediated end-joining (HMEJ)-based genome editing method was employed to integrate the natural resistance-associated macrophage protein-1 (NRAMP1) gene into bovine fetal fibroblasts. Calves produced by somatic cell nuclear transfer (SCNT) of these modified nuclei into enucleated eggs expressed NRAMP1, a metal ion transporter that increases resistance to zoonotic tuberculosis, verifying the HMEJ technique and exploring the possibility of genetic modification as a method of controlling zoonotic bacterial disease.²³

Disease monitoring and genetic modification can be used in tandem to minimize zoonotic outbreaks in humans. Only one transmissible spongiform encephalopathy (TSE) or prion disease affects humans, bovine spongiform encephalopathy (BSE), the cause of variant Creutzfeldt-Jakob disease. Other animal prion diseases such as scrapie, chronic wasting disease and atypical forms of BSE have not been documented in humans, but transmission experiments in nonhuman primates and mice suggest that they have zoonic potential.²⁴ Surveillance of TSE spread and infection kinetics in agricultural and wild animal species are critical to understand the links between animal and human prion disease.²⁵ In a proof-of-concept study, CRISPR-Cas genome editing was used to introduce novel prion protein (PRPN) gene variants, which provide protection against inherited prion diseases into cattle. The immediate aims of the study were to optimize the editing technique and generate viable calves with the desired genetic variation, with the long-term goal to reduce the likelihood of human prion pandemics.²⁶

Due to the remarkable ability of viruses to mutate and adapt to novel hosts, zoonotic viral diseases are particularly important to monitor. A review of zoonotic viral diseases highlights the prevalence of Rift Valley fever (RVF) and Middle East respiratory syndrome virus (MERSV) from livestock, Hendra virus (HeV), and equine encephalitis viruses (EEVs) from horses, Nipa viruses from pigs, rabies from most warm-blooded animals, and influenza viruses from domestic poultry, swine, horses and currently cattle.²⁷ A dramatic recent example of zoonotic viral disease spread in agricultural, wild, and companion animals is the highly pathogenic avian influenza viruses H5N1 clade 2.3.4.4b, which was identified in poultry and dairy facilities across the United States in 2022.^28–30 According to the Centers for Disease Control and Prevention, over 166 million poultry and 985 dairy herds have been affected. Seventy people have tested positive, and one person died, most of whom were exposed through work on farms or culling operations.^31–33 The first human infections were attributed to exposure to infected dairy cattle, with cattle serving as a mammalian link between birds and humans.³⁴ Recommendations to prevent human outbreaks include frequent metagenomic sequencing of cattle samples to understand viral genomic changes that allow for adaption to human hosts, risk of disease spread and disease severity.³⁵

Spontaneous infectious diseases

The One Health approach described in the introduction originates from the classic One Medicine concept, a central tenet of which is to promote health with parallel studies of naturally occurring spontaneous diseases in animals and humans.³⁶ The COVID-19 pandemic that began in 2019 emphasizes the importance of this work model. Even though livestock species such as pigs, cattle and poultry are mostly resistant to the causative agent of the pandemic, severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2), studies of naturally occurring coronavirus infections in these species already had created a base of knowledge about this virus subfamily that could inform the pandemic response. The first described coronavirus was isolated from chickens in 1931, and pigs are also susceptible to coronaviruses, including Middle Eastern Respiratory Syndrome coronavirus (MERS-CoV).³⁷ Bovine coronavirus (BCoV) belongs to the Betacoronavirus genus like SARS-CoV-2, and likewise, shares disease characteristics and tropism.³⁸ Following the One Medicine concept, these related spontaneous diseases in humans and livestock can be used to learn more about the mechanisms of Betacoronavirus infections. Medical researchers could generate research questions about a human pathogen, in this example SARS-Co-V-2, with veterinary scientists designing experiments using cattle naturally infected with BCoV to answer these research questions. Validation of the results could be translated back to human medicine.³⁸

Large animal models for antibody production and vaccine development

Anatomic features of antibodies produced by certain agricultural animal make them ideal for therapeutics, whereas physiological features of other agricultural animals render them relevant models in which to evaluate the efficacy of novel vaccines.

Llamas and other camelids produce functional heavy chain-only IgG antibodies (HCAbs), lacking the light chains typically associated with antigen recognition. Named nanobodies, these antibodies can reach a very high neutralization potency and due to their morphology, are less likely to obstruct access to hidden and essential epitopes on pathogenic agents like viruses, bacteria and parasites. The small size and relatively simple structure of nanobodies renders them stable, easy to produce and easily modifiable.³⁹ These features infer special therapeutic advantages of HCAbs as compared to conventional heterotetrametric antibodies.^40–43 Current clinical trials are investigating nanobodies as targeted therapies against programmed death-ligand 1 (PD-L1) and epidermal growth factor receptor (EGFR), surface proteins that are commonly overexpressed on cancer cells and correspond with tumor development and metastasis.^44,45 Nanobody therapies for infectious diseases include SARS-CoV-2, hepatitis B, human cytomegalovirus (HCMV), norovirus, and human immunodeficiency virus (HIV), which are being explored in human preclinical and clinical settings.^46–48

Rabbit antibodies are also intensively used for therapeutic purposes, as rabbits respond to infection or immunization by producing highly specific, high affinity antibodies through gene conversion and somatic hypermutation.^49,50 Rabbit Antithymocyte Globulin (Thymoglobulin^®) (rATG) is used in immunosuppressed kidney transplant patients to prevent and treat acute renal graft rejection. rATG further suppresses the immune system by depleting T-cells, modulating lymphocyte surface antigens, activating transcription factors and interfering with immune cell processes such as cytokine production, chemotaxis and endocytosis.⁵¹ Chimeric and humanized rabbit monoclonal antibodies have been also developed as potential specific immunotherapies for non-immunosuppressed patients. These include antibodies against CD40, hepatocyte growth factor (HGF) and vascular endothelial growth factor (VEGF).⁵⁰ The anti-VEGF antibody (brolucizumab) has been approved by the US Food and Drug Administration for use in patients with macular degeneration, and other humanized rabbit antibodies are being tested in clinical trials.^52,53

One of the most relevant and reliable animal models for human vaccine testing is the pig, which is similar to humans in physiology and immune system composition.^54–56 In a comparative analysis of various aspects of the immune system, the porcine immune system was found to resemble the human immune system for >80% of analyzed parameters, while mice resembled humans in <10% of analyzed parameters.⁵⁷ Pigs have been used to test Bordetella pertussis, Chlamydia trachomatis, Norovirus, and influenza virus vaccines.^58–61 Sheep are also a useful model for vaccine development, based on their relatively low maintenance cost, and they are extensively used for studies on mucosal immunization.⁶² An example is vaccine testing against the zoonotic pathogen Rift Valley fever virus (RVFV), where single vaccination induced sterile immunity in lambs and neutralizing antibodies were observed in vaccinated lambs without any anamnestic responses following RVFV challenge.⁶³

In some cases, multiple agricultural animals can serve as appropriate models for a single disease of humans. Human babies and young children are highly susceptible to respiratory syncytial virus (RSV) infection, which is the leading cause of infant hospitalization in the US. While mouse models are used to study RSV pathogenesis, they do not address the physiology and immunology that make infants especially prone to infection.⁶⁴ The epidemiological, pathological and clinical aspects of bovine respiratory syncytial virus (RSV) infection in calves is used as a comparative model for pediatric RSV.⁶⁵ Additionally, a perinatal lamb model of RSV infection parallels the disease in newborn infants, as both lung development and structure and pathology of RSV infection are similar in both species. Innate and adaptive immune responses of perinatal lambs in response to RSV infection resemble those of infants, further validating the model.⁶⁶ Consequently, both calf and lamb models are being used to test vaccines intended for infants.^65,66

Agricultural animal models for organ transplantation

Over 7 million women between the ages of 15 and 34 in the United States have lost their uteri to benign causes or obstetric complications, and uterine transplantation has been explored to compensate for these losses.⁶⁷ In the 1960s, several animal models including pigs and sheep were used to develop this technique. In 2000, a human-into-human uterine transplant was carried out, to test the transplantation protocols previously optimized in agricultural animal models. The technique was successfully performed, but the graft was removed after 99 d due to complications with the blood supply.⁶⁸ Since that initial attempt, researchers continue to employ pig, sheep, and goat models to understand aspects of mammalian immunology that determine the long term success or failure of uterine transplants.⁶⁷

Humans share more immune-system related genes and proteins with pigs than they do with other animals, including laboratory mice,⁶⁹ prompting researchers to create a pig model for the study of human lung transplantation. This pig model is particularly relevant for the study of primary graft dysfunction (PGD), a leading cause of morbidity and mortality soon after transplant.⁷⁰ It enables a clinical assessment of recovery from the surgery, including the extent of early inflammatory injury within the first 3 d post-transplant, and the effects of acquired immune responses from 3 to 7 d after surgery, making it an ideal model in which to test therapeutics prior to use in human clinical settings.⁷⁰

In addition to serving as a translational model for the study of organ transplantation, pig tissues soon may routinely be transplanted into human recipients. In the United States, over 100,000 patients are on waiting lists for organ transplantation and organ shortages continue to rise.⁷¹ Xenotransplantation from pigs to humans could help narrow the gap between available human organs and patients needing organs for survival. Although pigs and humans are physiologically and immunologically similar, genetic disparities between species need to be addressed for successful transplantation. Pigs are amenable to genomic engineering, which has allowed for the generation of transgenic lines with organs that are more compatible with the human immune system.⁷¹ To avoid hyperacute graft rejection in 1 set of pig-to-human kidney transplantation experiments, knockout pigs lacking the alpha-1,3-galactosyltransferase (GGTA1) gene were bred, and thymic autografts were inserted under the kidney capsule prior to transplantation. Kidneys were transplanted into 2 deceased human recipients whose circulatory and respiratory activity was maintained for the 54-h experiment. Transplanted kidneys made urine almost immediately after reperfusion and no signs of hyperacute or antibody-mediated rejection were detected during the study.⁷² These experiments are just a preliminary step toward permanently transplanting organs from pigs into humans, but the lack of hyperacute rejection suggests that the removal of GGTA1 plays a critical role in that process. The study duration was too short for the thymus transplant to exert the desired effect on the recipient T-cell repertoire, but preservation of thymic architecture and revascularization encouraged the authors that the use of the “thymokidney” may facilitate reduction in immunosuppression in future procedures.⁷²

Immunotherapies being explored in large animal models

Bacterial infection leads to tissue damage and impairs wound healing, both directly and because of inflammation initiated by host immune cell activity aimed at reducing bacterial load.⁷³ Agricultural animals are well-suited as models for antimicrobial and host-directed immunotherapies (HDTs), that target host immune cells with the goal of altering the immune response to reduce tissue damage and improve healing. HDTs tested in agricultural animal models include bacterial products, mammalian cell surface proteins, cytokines, stem cells and stem cell products, and engineered nanoparticles, all of which are relevant to human medicine.

Skin wound healing therapies

Horses and humans exhibit similarities in normal skin wound healing and both species naturally develop hard to heal chronic wounds, often because of secondary bacterial contamination. These features make the horse a physiologically relevant model to test novel treatments for human skin injury.⁷⁴ An established equine experimental excisional bacterial aggregate-infected skin wound model allows both normal and impaired wound healing to be studied over time, providing a unique situation in which to test antimicrobial compounds or HDTs.⁷⁵ A stem cell-based biologic alternative to traditional antibiotics with cross-species antimicrobial and HDT potential has been tested in a similar equine infected skin wound model.¹² A key advantage of both models is based on the body size of horses. For example, 20 surgically induced wounds were created at different sites on a single animal, resulting in wounds that heal at different rates, and allowing for treatments and controls to be tested in the same individual.⁷⁵

Cancer therapies

Certain types of cancer also present similarly in horses and humans. Ocular surface squamous neoplasia (OSSN) is the most common type of conjunctival cancer in both species^76,77 and many features of the disease, including ocular pathology, risk factors and lesion progression, are similar,^76,78 making horses with OSSN an appropriate large animal model in which to study immunotherapies to induce tumor regression in humans.⁷⁹ Liposomal TLR complex (LTC) immunotherapy has been shown to initiate an innate immune response that may inhibit OSSN progression.^80,81 A study designed to evaluate the effectiveness of LTC immunotherapy in an equine model of spontaneous OSSN model showed that treatment with LTC resulted in an overall tumor response rate of 67%, including regression of large OSSN tumors. Repeated treatment was well tolerated clinically, leading the authors to conclude that ocular immunotherapy with LTC warrants investigation as a novel approach to controlling OSSN in humans.⁷⁹ Bacterial-based intratumoral cancer immunotherapy, based on the rationale that a systemic response is initiated when the immune system recognizes conserved pathogen-associated molecular patterns (PAMPs), has been tested in another equine cancer model of melanoma. Injection of PAMPs directly into tumors was shown to have the potential to induce a host immune response against cancer cells.⁸² A slow release emulsion of killed mycobacteria with complete Freund’s adjuvant (CFA) was injected into naturally occurring equine melanomas that led to a 27% clinical response based on reduction in mass size.⁸³ These data, combined with the outcomes from similar experiments carried out in mice, dogs, and a preliminary human trial, demonstrates that intratumoral injection of CFA has antitumor effects in some treated animals and is safe for human use.⁸³

Melanoma is the deadliest human skin cancer with increasing incidence globally. Pigs, like humans, spontaneously develop melanomas, but unlike the human disease, porcine melanomas often regress, even after metastasis.⁸⁴ Three different pig models (MeLiM, Sinclair, and MMS-Troll) have been used to understand the role of the adaptive immune response in tumor regression.⁸⁵ A recent study of melanoma in MeLiM pigs found that CD4-CD8+ cytotoxic T cells and NK cells were the most abundant tumor infiltrating immune cells, leading the authors to speculate their role in tumor regression and propose leveraging them as therapies for human disease.⁸⁶

Therapies for bacterial-associated tissue damage

Bovine mastitis is another agricultural animal disease with features that render it a useful model in which to study HDTs for humans. The strength of this model is that preventative and curing anti-microbial therapies, as well as treatments to restore damaged tissue function, can be examined simultaneously. Effective treatments for bovine mastitis may be relevant to many diseases in which bacterial infection leads to tissue damage. Cytokine-based HDTs have been tested in bovine mastitis models using various protocols and pathogens. In a study designed to determine the utility of a prophylactic cytokine treatment, recombinant human granulocyte colony-stimulating factor (GCSF) was administered daily to lactating cows by subcutaneous injection for 15 d. Ten days after the first treatment, cows were challenged with Staphylococcus (S.) aureus via intracisternal inoculation. Compared to placebo-treated controls, infection was reduced by 47% in cows treated with GCSF, which the authors suggested could be related to the recruitment of neutrophils into the mammary tissue prior to the challenge.⁸⁷ Recombinant bovine interferon gamma (rbIFN-ɣ) has also been tested as a mastitis preventative. Cows given an intramammary dose of rbIFN-ɣ 24 h prior to an E. coli challenge had fewer infected quarters, less severe clinical signs of mastitis and shorter periods of infection as compared to placebo-treated controls. Additionally, all rbIFN-ɣ-treated cows survived the challenge, while 42% of placebo controls died within 3 d post challenge. The authors concluded that the infusion of rbIFN-ɣ prevented rapid E. coli growth in the mammary gland tissue and inhibited the development of a detrimental inflammatory response.⁸⁸ In an experimental S. aureus infection model, mastitic cows were treated with recombinant bovine interleukin 2 (IL-2) or interleukin 1β (IL-1β) after infection. The percentage of cured udder quarters given each cytokine treatment was not different than the percentage of cured udder quarters treated with antibiotics, suggesting the interleukin treatment has the potential to replace or complement antibiotics, and all treatment groups showed a significantly higher cure percentage than untreated controls.⁸⁹

Tissue damage from direct bacterial activity and host immune cell responses to pathogens is a long-term negative consequence of infectious disease in general, and mastitis in particular, as disrupted mammary epithelial cells typically never produce as much milk as they did pre-infection. Stem cell-based HDTs present the promise of treatments composed of molecules with direct antimicrobial and immunomodulatory activity, as well as factors that promote tissue regeneration. In one study, mesenchymal stromal cells (MSCs) were delivered intramammarily to healthy cows and cows that were previously inoculated with S. aureus. MSCs did not induce clinical or immunological responses in healthy cows and although MSC treatment in infected cows did not induce detectable changes in somatic cell counts in milk when compared to antibiotic- or vehicle control-treated cows, MSC-treated cows did have lower colony forming units of bacteria in milk compared to vehicle control-treated cows. Based on these results, the authors concluded that their work provided evidence that allogeneic MSC intramammary therapy is both safe and effective for the treatment of bovine mastitis.⁹⁰ A global hypothesis that can be generated from these studies is that similar immunotherapies can reduce bacterial load and infection-associated tissue damage in humans as well.

Limitations of immunologic studies in large and agricultural animals

Experiments in agricultural animals that advance the understanding of immunology in humans, and lead to the development of improved therapies, are not trivial to conduct. Due to the diversity in large animal physiology, particularly mechanisms of innate and adaptive immune responses, selection of relevant models must be carried out thoughtfully. What is known for one species cannot always be extrapolated to another species, and studies need to include appropriate controls and enough participants to provide adequate statistical power. In addition, ethical considerations associated with large animal experiments need to be addressed when designing studies and applying for funding. Agricultural species not only need to be well-cared for during studies, but often researchers are encouraged to adopt animals out to appropriate “homes” when experiments are finished.

The expenses associated with large animal care can be prohibitive and the availability of facilities to house farm animals appropriately is limited.⁹¹ Livestock species often need outdoor areas for freedom of movement and intraspecies interactions. Moving individual animals to contained areas for observation, sample collection and treatment can be time consuming and dangerous to those running the study. Because it can be physically risky to handle large animal species, researchers working directly with them may need specific training in both experimental methods and animal behavior. Removal of large animals from studies can be more difficult and costly than it is for common laboratory species and may involve specialized veterinary care, quarantine and/or shipping to alternate locations.

While beneficial in many cases, the large sizes, long gestation periods and extensive lifespans of agricultural animals can hinder timely acquisition of data. Finally, resources and reagents for these species have historically been limited as compared to what is available for humans and traditional laboratory animal models. However, as mentioned in the introduction, this is becoming less of an issue as -omics databases for many large and agricultural animals are increasingly curated and accessible. In parallel, the range of antibodies used to study immune cell populations in large and agricultural animals is growing but is not nearly as extensive as the inventory that is available for mice or humans.

Immunologic studies in agricultural animals benefit animals and food systems health

To emphasize the importance of farm animals in immunology research, this review describes contributions of select agricultural animal studies to inform our knowledge of the human immunologic diseases, and how farm animals aid in the development of human vaccines and therapies. It would be negligent, however, to not also discuss the importance of large animal models for their own health benefit and to human food systems health.

According to literature published by the USDA, there are millions of horses, cattle, sheep, goats, hogs and hens in the United States, for a total number of farm animals far exceeding the approximately 330 million people currently living in the country.⁹² It is generally accepted that humans have a moral obligation to consider and optimize the well-being of animals we use for food, fiber, work and sport. While the ethics of animal use are beyond the scope of this review, it can be argued that unhealthy animals negatively impact human health and the sustainability of human food supplies.

Agricultural animal immunomodulatory therapeutics are an area of active research, with several being approved and commercially available. Table 1 lists immunomodulators that are commercially available for use in agricultural animals and provides select examples of immunomodulators that are under investigation, with the long-term goal of administering them to agricultural species.^90,93–101 Some of these immunotherapies were discussed in detail earlier in this review, demonstrating their translational potential for minimizing the effects of bacterial associated tissue damage in humans. Here the focus is on animal well-being.

Table 1.

Select agricultural animal immunomodulators.

Commercially available immunomodulators
Product	Composition	Recipient species	Purpose	Mechanism	References
Amplimune®	Mycobacterium cell wall fraction	Calves	Reduction of mortality associated with scours caused by E. coli K99	Activates innate immune responses	93
Zylexis™	Inactivated parapoxvirus	Horses	Reduce effects of equine herpesviruses 1 and 4	Increases cytokines associated with Th1 cell mediated response	94
Imrestor™	Granulocyte colony stimulated factor	Cattle	Reduce clinical mastitis	Increases blood neutrophils, modifies neutrophil and monocyte phenotypes	95
ZelNate™	Liposome DNA complex	Cattle	Treatment of bovine respiratory disease	Activates stimulator of interferon gene pathway	96

Actively researched immunomodulators
Immunomodulator	Recipient species	Purpose	Mechanism	References
β-Glucans	Broiler chickens	Enhance immune status	Increases innate immunity	97
Pattern recognition receptor agonists	Chickens	Enhance vaccine response	Enhances antibody response, induces CD8+ T cell response	98
Probiotics	Piglets	Induce host defense peptide production	Recognized by TLR2, activate ERK1/2, JNK	99
Immunobiotics	Piglets	Modulate intestinal immunity	Reduces blood complement activity, C reactive protein concentrations	100
Fecal microbiota	Chicks	Reduce intestinal inflammation	Decreases Th17 cell associated transcription factors/cytokines, increases Treg transcription factors/cytokines	101
Stem cell products	Cattle	Reduce clinical mastitis	Reduces colony forming units of bacteria in milk	90

A prime example of basic immunological research uncovering mechanisms to understand and improve animal and food systems health can be found in the poultry industry, where producers must focus on disease control to be successful. The respiratory tract is a major point of entry for avian pathogens such as avian influenza virus, Newcastle Disease virus, infectious bronchitis virus and avian pathogenic Escherichia coli. Accordingly, many poultry vaccines are delivered as aerosols. The avian respiratory system differs anatomically from the mammalian respiratory system, and little is known about the ontogeny and function of the immune cells in that compartment.¹⁰² Research in transgenic chickens expressing a reporter gene allowing for visualization of cells derived of the mononuclear phagocyte (MNP) lineage has shed light on the development and distribution of germinal centers and MNPs in the bronchus-associated lymphoid tissue, parabronchi, trachea and air sacs.¹⁰² Additional experiments using the same model have provided previously unknown details about the development and function of chicken dendritic cells.¹⁰³ This work provides a base for additional functional characterization of avian MNP, macrophage and dendritic cell subpopulations, including antigen presentation to T and B cells. A better understanding of avian respiratory tract immunity will lead to a better understanding of respiratory disease pathogenesis and support the development of more effective vaccines. As discussed earlier, updates to avian influenza vaccines are imperative not only to prevent breakthrough infections in humans, but to maintain chicken health, and provide poultry and eggs as safe human food sources.

In the interest of producing a reliable source of safe, hypoallergenic food products, gene editing techniques have been applied to cattle, goats, chickens and pigs.¹⁰⁴ This primarily involves knocking out genes coding for proteins that illicit an allergic response themselves or lead to the production of immunogenic molecules. In the quest to produce milk with decreased allergenicity, the beta-lactoglobulin (BLG) gene has been edited in cows and goats.^105,106 Chickens with ovalbumin (OVA) and ovomucoid (OVM) knockouts have been generated, resulting in eggs that are nearly devoid of these antigenic proteins.¹⁰⁷ Pigs lacking the alpha-1,3-galactosyltransferase (GGTA1) gene, discussed earlier as organ donors, can be used to produce meat without alpha-1,3-galactose (α-gal). This carbohydrate is generally safe for human consumption, but some tick species transfer α-gal antigens into the human hosts on which they feed, triggering and IgE-mediated antibody reaction to meat. The US Food and Drug Administration has approved these “GalSafe” pigs for human consumption.¹⁰⁴ The company that produces these pigs does so on a relatively small scale, with people who are allergic to α-gal as their target market.

These are just a few select examples of how understanding the specific immunology of farm animals may have significant positive impacts on animal and food systems health.

Conclusions

Studies in large and agricultural animals advance our understanding of the human immune system and immunological diseases, filling in gaps that cannot be addressed using rodent models. Following a One Health approach, comparative immunologic studies across agricultural animals and humans will lead to improved human, animal, and food systems health.

Author contributions

R.M.H. conceptualization (lead), writing original draft, reviewing and editing (equal); G.R.V.d.W. conceptualization (support), reviewing and editing (equal).

Gerlinde R. Van de Walle (Conceptualization [Supporting], Writing—review & editing [Equal]) and Rebecca M. Harman (Conceptualization [Lead], Writing—original draft [Lead], Writing—review & editing [Equal]).

Funding

This study received funding from the Foundation for Food and Agriculture Research (grant number CA20-SS-0000000004), which includes matching funds from Elanco Animal Health and the NY Farm Viability Institute; the National Cancer Institute from the National Institutes of Health (grant number 5R21CA285521-02) and the US Department of Agriculture, National Institute of Food and Agriculture (grant number 2022-67015-36351) for support of immunology studies in large and agricultural animals.

Conflict of interest

The authors declare no conflicts of interest.