Myiases are parasitic infections caused by the larval stages of some fly species. In American nonhuman primates (NHP), three bot fly species causing cutaneous myiasis have been reported: Cuterebra baeri, Cochliomyia hominivorax, and Dermatobia hominis. Studies on myiasis in NHP are scarce and mainly based on morphologic identification of larvae, while molecular approaches have been barely used. This study aimed to molecularly identify bot flies parasitizing two NHP species living sympatrically in central Colombia. Bot fly larvae were collected from two grey-legged night monkeys (Aotus griseimembra) and from a howler monkey (Alouatta seniculus). Larvae were measured and photographed for morphologic evaluation; subsequent molecular characterizations of a partial region of the cytochrome c oxidase subunit 1 mitochondrial gene were performed. Sequence analysis allowed the identification of all specimens as Cuterebra baeri, enabling confirmation of this species parasitizing Alouatta seniculus and Aotus griseimembra in Colombia.