The intraspecific variability of Triatoma dimidiata Latreille, a major vector of Chagas disease, was studied in four departments of Guatemala. Insects were collected from either domestic and sylvatic habitats, and their cuticular hydrocarbon pattern and head morphology were analyzed using ordination and classification techniques. A significant discrimination was obtained both with morphometric and hydrocarbon analyses. Insects from northern departments were easily differentiated from southern conspecifics. Distinctive hydrocarbon pattern and head shape were detected for insects collected from caves in the north central region of the country, posing concern about their taxonomic status.