Wing morphology has been used in taxonomic and systematic studies of insects, often enabling the identification of groups based on the variation. In this study, wing Geometric Morphometrics was used to verify if eastern and western populations of Partamona rustica, separated by the São Francisco River, are discriminated, thus confirming previous molecular data. The two groups of P. rustica exhibited significant differences in wing size and shape. Better differentiation of populations and groups was achieved with the centroid size. We generated dendrograms using Mahalanobis and Procrustes distances, which discriminated the eastern and western populations. Isolation by the distance between morphometric and geographic distances was found. The confirmation of the two population groups points out the need for further studies investigating the occurrence of barriers to gene flow and colonization history in the semiarid region by this stingless bee.