Barrier effect is a road‐related impact affecting several animal populations. It can be caused by behavioural responses towards roads (surface and/or gap avoidance), associated emissions (traffic‐emissions avoidance) and/or circulating vehicles (vehicle avoidance). Most studies so far have described road‐effect zones along major roads, without determining the actual factor inducing the behavioural response. The purpose of the present study was to assess the factors potentially causing road‐effect zones in a heterogeneous road network (with variations in road width, road surface and traffic volume) and eventually to estimate the reduction of habitat quality imposed by roads within a protected area (Doñana Biosphere Reserve, Spain). As model species, we used two ungulates, red deer Cervus elaphus and wild boar Sus scrofa. We surveyed the presence of both species along 200‐m transects. All transects … Palabras clave: Avoidance