BACKGROUND: Ochlerotatus scapularis is a potential vector of filarias and arboviruses in the Neotropics. This species was once typically associated with sylvatic environments; however, cases of synanthropy and urbanization of this species have been increasingly reported in southeast Brazil. Despite the medical relevance of Oc. scapularis, its populational variability is not yet known. To our knowledge, this is the first report describing the morphological and genetic variabilities of this species. METHODS: Population samples were characterized using the cytochrome oxidase subunit I (COI) mitochondrial gene and wing geometrics. Adult mosquitoes were collected from five sampling sites from remnants of the Atlantic forest embedded in the urban or rural areas of southeast Brazil. RESULTS: In the 130 individuals analyzed, 46 COI haplotypes were detected. Haplotype diversity was high and ranged from 0.66 to 0.97. Six haplotypes were present in 61% of the individuals, whereas the remaining haplotypes were less frequent (39%). Wing shape was also highly polymorphic. Differentiation of populations across sampling sites according to genetic distances (Fst = -0.009 to 0.060) and morphological distances (Qst = 0.47) indicated that populations were not identical. No correlations were noted for phenetic and genetic diversities (p = 0.19) or for genetic or phenetic distances with geographical distances (p = 0.2 and p = 0.18, respectively). CONCLUSIONS: Our study results suggest that Oc. scapularis has a rich genetic patrimony, even though its habitat is fragmented. Implications of such genetic richness with respect to vectorial competence, plasticity, and ability to exploit urbanized areas need to be further investigated.