Tropical moist forests are complex and diverse ecosystems threatened by human-induced disturbance. Despite recent advances in the study of the impact of land-use change on mammal diversity, there is a lack of research about which factors affect mammal assemblages within fragmented landscapes in Colombia. Herein, we investigate how medium and large-bodied mammals in a fragmented landscape respond to landscape characteristics, identifying how they affect species richness and functional diversity. We sampled eight forested patches in the Middle Magdalena Valley in central Colombia and modeled mammal functional diversity (functional dispersion and evenness) and species richness as a function of landscape variables (i.e., patch size, patch shape, landscape connectivity, and forest fragmentation). Forest fragmentation was the most important variable explaining mammal species richness and functional diversity (measured as functional dispersion). Species richness, functional dispersion, and the number of functional groups were increased in less fragmented forest patches. Conversely, functional evenness exhibited the opposite trend. Taken together, these results show that mammal assemblages in forest patches within forest-dominated neighborhoods support both a higher diversity and a broader spectrum of ecological functions but are less resilient to environmental perturbations. We highlight the importance of forest cover for providing critical habitat for both generalized and specialized mammals in fragmented landscapes. Overall, our fndings reinforce the importance of defining conservation actions for minimizing the impact of human activities on the remaining native forests, as they are fundamental for maintaining wild mammals and the ecological processes in which they participate.