Exercise stress is associated with an increased risk for upper respiratory tract infection (URTI). We have shown that consumption of the soluble oat fiber β-glucan (OβG) can offset the increased risk for infection and decreased macrophage antiviral resistance following stressful exercise; however, the direct role of macrophages is unknown. This study examined the effect of macrophage depletion on the benefits of orally administered OβG on susceptibility to infection (morbidity, symptom severity, and mortality) following exercise stress. CL2MDP (Ex- H2O-CL2MDP, Ex-OβG-CL2MDP, Con-H2O-CL2MDP, Con-OβG-CL2MDP)-encapsulated liposomes were administered intranasally to deplete macrophages, and PBS (Ex-H2O-PBS, Ex-OβG-PBS, Con-H2O-PBS, Con-OβG-PBS)-encapsulated liposomes were given to macrophage-intact groups. Ex mice ran to volitional fatigue on a treadmill for 3 consecutive days, and OβG mice were fed a solution of 50% OβG in their drinking water for 10 consecutive days before infection. Fifteen minutes following the final bout of Ex or rest, mice were intranasally inoculated with 50 μl of a standardized dose of herpes simplex virus-1. Ex increased morbidity ( P < 0.001) and symptom severity ( P < 0.05) but not mortality ( P = 0.09). The increase in morbidity and symptom severity was blocked by OβG consumption for 10 consecutive days before exercise and infection [morbidity ( P < 0.001) and symptom severity ( P < 0.05)]. Depletion of macrophages negated the beneficial effects of OβG on reducing susceptibility to infection following exercise stress, as evidenced by an increase in morbidity ( P < 0.01) and symptom severity ( P < 0.05). Results indicate that lung macrophages are at least partially responsible for mediating the beneficial effects of OβG on susceptibility to respiratory infection following exercise stress.