The hepatitis E virus (HEV) is an emerging multi-host pathogen, with suids being the main reservoir. Humans are primarily infected through the consumption of contaminated water or food. In Argentina, HEV circulation has been confirmed in humans, domestic pigs, wild boar (Sus scrofa), and surface water. In El Palmar National Park, invasive wild boar and axis deer (Axis axis) are controlled, and their meat is released for public consumption, with trimmings and offal frequently fed to dogs. Between 2017 and 2019, we conducted a multi-species serological survey in this protected area to assess HEV exposure in invasive mammals and in dog and human consumers of game meat. We also evaluated associations between seropositivity and environmental variables, as well as behavioral risk factors among game-meat consumers. Total anti-HEV antibodies were detected in 29/75 (38.67%) wild boar, 1/134 (0.75%) deer, 1/18 (5.6%) dogs, and 6/59 (10.17%) humans. A spatial cluster of seropositive wild boar was identified in a low-lying, flood-prone area near the confluence of the El Palmar stream and the Uruguay river, suggesting increased risk of environmental transmission. This is the first report of HEV exposure in wild boar from this park and in axis deer and dogs in Argentina. Participation in culling and game meat handling and consumption may contribute to HEV exposure pathways among humans. These findings improve understanding of HEV epidemiology at the wildlife–domestic animal–human interface and highlight the influence of environmental factors and human behavior on zoonotic virus circulation.