Defaunation is a key ecological issue that has only recently been given sufficient attention. As predicted, evidence so far indicates loss of larger species followed by medium-sized species, leading to cascading effects that propagate throughout entire communities and ecosystems. The Atlantic Forest is among the most important global biodiversity hotspots. These regions have historically been impacted by habitat loss and fragmentation, resulting in landscape changes and negative impacts upon animal communities. This study evaluates community characteristics of medium- and large-sized mammals in subtropical Atlantic Forest, southern Brazil. We gathered data on mammal occurrence using 108 cameras traps located across 8 protected areas. We then tested whether landscape differences impact mammal richness, composition, and community complexity. Specifically, we used a regression tree to evaluate compositional differences as a function of landscape configuration. We analyzed data for 26 species in total, with the number of species per area ranging from 9 to 17. Changes in mammal composition at the landscape scale were most strongly associated with human occupation. Areas with strong human occupation had low species richness, with a predominance of medium-sized omnivores and insectivores species; these conditions led to high defaunation indices. Community complexity was greater in areas with low human occupation, where carnivores (Felidae) were more abundant. Differences in species composition were also linked to altitudinal bands and the ratio of period of time with protected status versus history of land exploitation in a particular area. Analysis of functional groups indicated that intense human occupation had negative effects on larger species, a process that may have impending consequences. Despite defaunation being a serious ecological issue, we assert that taking prompt action may limit or potentially reverse effects of defaunation before the most dramatic changes take place.