The Long-eared Owl (Asio otus) is a widely distributed predator with plastic foraging habits. Its diet has been studied across most of Europe and North America, but little information is available on intra-seasonal diet variation. We studied the winter diet of Long-eared Owls in a Mediterranean fragmented farmland. Diet was dominated by small mammals (mostly Apodemus spp.), with birds the second most abundant group. In our landscape, the amount of bird prey items in the diet was higher compared to values reported from other areas of Italy. There was relatively large variation in the diet among the three months of study. Our study confirms the pattern of higher Long-eared Owl diet diversity in a Mediterranean landscape compared to that reported for other regions of Europe where the diversity of prey species may be lower. The findings suggest that extensively-managed Mediterranean rural landscapes can provide habitat suitable for the Long-eared Owl, as they offer a wide diversity of food sources that may be exploited by this nocturnal predator.