Specific metabolic pathways affect immune cell differentiation and function, and accordingly have an impact on the overall immune response during health and disease. The metabolic state of leukocytes may provide a signature associated with a particular condition or treatment, and identifying the metabolic demand of a response may lead to the development of interventions for immune modulation. Blood is relatively easy to collect from live animals, can be repeatedly sampled from the same animal, and is an obvious source of leukocytes. Using an Extracellular Flux Analyzer (Seahorse Biosciences) we interrogated glycolytic and mitochondrial function of porcine peripheral blood mononuclear cells (PBMC) in vitro following stimulation with mitogen and ex vivo following infection of pigs with porcine reproductive and respiratory syndrome virus (PRRSV). In addition, the metabolic baseline and stressed phenotype of different CD3+ T cells was determined. Stimulation of PBMC with pokeweed mitogen resulted in a significant change in the metabolic phenotype of the cells, with a shift primarily in mitochondrial respiration. Sorted CD3+ T cell populations (CD4+CD8α-, CD4-CD8α+, and CD4+8α+) had varied responses to metabolic stress, indicating different metabolic pathways may be associated with specific CD3+ T cell populations (CD4 versus CD8). Basal glycolysis in PBMC was not significantly impacted by PRRSV infection, but glycolytic capacity and reserve were significantly increased in PBMC from PRRSV-infected pigs (day 14 post-infection) over non-infected pigs. The effect of PRRSV infection on PBMC mitochondrial respiration was minimal, with a slight increase in spare respiratory capacity detected. Collectively, these data indicate that activation alters the bioenergetic profile of porcine PBMC, and infection can significantly impact the metabolic signature of circulating leukocytes. Understanding the relationship between immune cell phenotype and function as it relates to metabolism will aid in development of intervention strategies to meet the metabolic demand of pigs during health challenges.