Following influenza virus infection, CD8 T cells encounter mature, Ag-bearing dendritic cells within the draining lymph nodes and undergo activation, programmed proliferation, and differentiation to effector cells before migrating to the lungs to mediate viral clearance. However, it remains unclear whether CD8 T cells continue their proliferation after arriving in the lungs. To address this question, we developed a novel, in vivo, dual-label system using intranasal CFSE and BrdU administration to identify virus-specific CD8 T cells that are actively undergoing cell division while in the lungs. With this technique we demonstrate that a high frequency of virus-specific CD8 T cells incorporate BrdU while in the lungs and that this lung-resident proliferation contributes significantly to the magnitude of the Ag-specific CD8 T cell response following influenza virus infection.

Mycobacterium bovis Bacillus Calmette-Guérin (BCG) is a live attenuated vaccine for use against tuberculosis (TB); however, it is known to reduce childhood mortality from infections other than TB. The unspecific protection induced by BCG vaccination has been associated with the induction of memory-like traits of the innate immune system identified as ‘trained’ immunity. In humans and mouse models, in vitro and in vivo BCG training leads to enhanced production of monocyte-derived proinflammatory cytokines in response to secondary unrelated bacterial and fungal pathogens. While BCG has been studied extensively for its ability to induce innate training in humans and mouse models, BCG’s nonspecific protective effects have not been defined in agricultural species. Here, we show that in vitro BCG training induces a functional change in bovine monocytes, characterized by increased transcription of proinflammatory cytokines upon restimulation with the toll-like receptor agonists. Importantly, in vivo, aerosol BCG vaccination in young calves also induced a ‘trained’ phenotype in circulating peripheral blood mononuclear cells (PBMCs), that lead to a significantly enhanced TLR-induced proinflammatory cytokine response and changes in cellular metabolism compared to PBMCs from unvaccinated control calves. Similar to the long-term training effects of BCG reported in humans, our results suggest that in young calves, the effects of BCG induced innate training can last for at least 3 months in circulating immune populations. Interestingly, however, aerosol BCG vaccination did not ‘train’ the innate immune response at the mucosal level, as alveolar macrophages from aerosol BCG vaccinated calves did not mount an enhanced inflammatory response to secondary stimulation, compared to cells isolated from control calves. Together, our results suggest that, like mice and humans, the innate immune system of calves can be ‘trained’; and that BCG vaccination could be used as an immunomodulatory strategy to reduce disease burden in juvenile food animals before the adaptive immune system has fully matured.

Antimicrobial resistance (AMR) is a significant problem in health care, animal health, and food safety. To limit AMR, there is a need for alternatives to antibiotics to enhance disease resistance and support judicious antibiotic usage in animals and humans. Immunomodulation is a promising strategy to enhance disease resistance without antibiotics in food animals. One rapidly evolving field of immunomodulation is innate memory in which innate immune cells undergo epigenetic changes of chromatin remodeling and metabolic reprogramming upon a priming event that results in either enhanced or suppressed responsiveness to secondary stimuli (training or tolerance, respectively). Exposure to live agents such as bacille Calmette-Guerin (BCG) or microbe-derived products such as LPS or yeast cell wall ß-glucans can reprogram or “train” the innate immune system. Over the last decade, significant advancements increased our understanding of innate training in humans and rodent models, and strategies are being developed to specifically target or regulate innate memory. In veterinary species, the concept of enhancing the innate immune system is not new; however, there are few available studies which have purposefully investigated innate training as it has been defined in human literature. The development of targeted approaches to engage innate training in food animals, with the practical goal of enhancing the capacity to limit disease without the use of antibiotics, is an area which deserves attention. In this review, we provide an overview of innate immunomodulation and memory, and the mechanisms which regulate this long-term functional reprogramming in other animals (e.g., humans, rodents). We focus on studies describing innate training, or similar phenomenon (often referred to as heterologous or non-specific protection), in cattle, sheep, goats, swine, poultry, and fish species; and discuss the potential benefits and shortcomings of engaging innate training for enhancing disease resistance.

Respiratory syncytial virus (RSV) infection is a leading cause of severe acute lower respiratory tract infection in infants and children worldwide. Vitamin A deficiency (VAD) is one of the most prevalent nutrition-related health problems in the world and is a significant risk factor in the development of severe respiratory infections in infants and young children. Bovine RSV (BRSV) is a primary cause of lower respiratory tract disease in young cattle. The calf model of BRSV infection is useful to understand the immune response to human RSV infection. We have previously developed an amphiphilic polyanhydride nanoparticle (NP)-based vaccine (i.e., nanovaccine) encapsulating the fusion and attachment proteins from BRSV (BRSV-NP). Calves receiving a single, intranasal dose of the BRSV-NP vaccine are partially protected from BRSV challenge. Here, we evaluated the impact of VAD on the immune response to the BRSV-NP vaccine and subsequent challenge with BRSV. Our results show that VAD calves are unable to respond to the mucosal BRSV-NP vaccine, are afforded no protection from BRSV challenge and have significant abnormalities in the inflammatory response in the infected lung. We further show that acute BRSV infection negatively impacts serum and liver retinol, rendering even well-nourished individuals susceptible to VAD. Our results support the use of the calf model for elucidating the impact of nutritional status on mucosal immunity and respiratory viral infection in infants and underline the importance of VA in regulating immunity in the respiratory mucosa.

The objectives of this study were to determine the effects of oral supplementation with Saccharomyces cerevisiae fermentation products (SCFP; SmartCare and NutriTek; Diamond V, Cedar Rapids, IA) on immune function and bovine respiratory syncytial virus (BRSV) infection in preweaned dairy calves. Twenty-four Holstein x Angus, 1-2 day old calves (38.46 ± 0.91 kg initial body weight) were assigned two treatment groups: control; or SCFP treated, milk replacer with 1 g/d SCFP (SmartCare) and calf starter top-dressed with 5 g/d SCFP (NutriTek). The study consisted of one 31 d period. On d 19-21 of the supplementation period, calves were challenged via aerosol inoculation with BRSV strain 375. Calves were monitored twice daily for clinical signs, including rectal temperature, cough, nasal and ocular discharge, respiration effort and lung auscultation. Calves were euthanized on d 10 post infection (d 29-31 of the supplementation period) to evaluate gross lung pathology and pathogen load. Supplementation with SCFP did not affect body weight (P = 0.762) or average daily gain (P = 0.750), percentages of circulating white blood cells (P < 0.05), phagocytic (P = 0.427 for neutrophils and P = 0.460 for monocytes) or respiratory burst (P = 0.119 for neutrophils and P = 0.414 for monocytes) activity by circulating leukocytes either before or following BRSV infection, or serum cortisol concentrations (P = 0.321) after BRSV infection. Calves receiving SCFP had reduced clinical disease scores compared to control calves (P = 0.030); reduced airway neutrophil recruitment (P < 0.002); reduced lung pathology (P = 0.031); and a reduced incidence of secondary bacterial infection. Calves receiving SCFP shed reduced virus compared to control calves (P = 0.049) and tended towards lower viral loads in the lungs (P = 0.051). Immune cells from the peripheral blood of SCFP treated calves produced increased (P < 0.05) quantities of IL-6 and TNFα in response to toll-like receptor stimulation; while cells from the bronchoalveolar lavage (BAL) of SCFP treated calves secreted less (P < 0.05) proinflammatory cytokines in response to the same stimuli. Treatment with SCFP had no effect on virus-specific T cell responses in the blood, but resulted in reduced (P = 0.045) virus-specific IL-17 secretion by T cells in the BAL. Supplementing with SCFP modulates both systemic and mucosal immune responses and may improve the outcome of an acute respiratory viral infection in preweaned dairy calves.