The giant anteater Myrmecophaga tridactyla is categorized as Vulnerable on the IUCN Red List, and is extinct in several locations as a result of environmental pressures. We present the first records of the giant anteater in the largest continuous expanse of the Atlantic Forest biome, in southern Brazil, highlighting its occurrence in the highlands of the Serra do Mar mountain range. During a camera-trapping survey of medium and large mammals we obtained two records of the species, one in 2013 and the other in 2014. These records from dense rainforest highlands indicate the importance of this environment for the giant anteater and highlight these areas as possible foci for future studies of the species. In addition, we present a review of the species’ occurrence in protected areas in Brazil, to provide a resource for the conservation of this species and for future re-evaluations.