Drainage culverts are known to be used by a diverse number of species. To date, most studies looking at culvert usage have been restricted to the dry season. This seasonal bias has limited our understanding of how different species respond to culverts and, consequently, our ability to find effective ways to promote the use of culverts as aids to species movement. The main goal of this study was to examine the role of highway culverts for mesocarnivores throughout the year. We addressed (1) the seasonality of culvert use, (2) the relative importance of culvert structure, highway features, and surrounding landscape on culvert use, (3) the influence of the water depth and cover on culvert use, and (4) the effect of culvert structure on individual behavior. Fifteen culverts were monitored along 2 highways in southern Portugal using video-surveillance cameras and marble dust for 10 consecutive days per season. We used generalized linear mixed models to determine which factors most affected the culvert use and behavior by mesocarnivores. Our results highlight the effect of seasonality and water on culvert use. Culvert use was positively related with species activity throughout the year. All species (except otters (Lutra lutra)) were less likely to use culverts that contained water more than 3 cm deep or covering more than 70 % of the culvert base. Based on our results, future surveys and culvert retrofit design should address (1) the importance of seasonality in the interpretation of results and (2) the complementarity of culvert-specific features (water, ledges, and naturalization).