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Seventy-five samples were collected from 15 beehives in the central highlands of Ecuador (Tungurahua-Chimborazo) to assess Africanization in managed bee populations using wing geometric morphometric and mitochondrial DNA analyses. The results indicated that when grouping the apiaries based on altitudinal floors into 2600-2800, 2801-3000, and 3001-3274 m above sea level, differences ( p < 0.001) were observed. The morphotypes were similar in the first two floors, but the third indicated that altitude plays a crucial role in the differentiation of populations. When comparing with the pure subspecies, we found differences ( p < 0.001); the nearest Mahalanobis distance was for Apis mellifera scutellata (D2 = 3.51), with 95.8% Africanization via father in the area. The maternal origin of all patterns belonged to lineage A ( A. m. scutellata ), with seven haplotypes. The most frequent haplotypes were A26 and A1; however, the A1q haplotype was not detected at the national level or in nearby countries. The identified haplotypes do not coincide with A4, which is predominant in South Africa and Brazil. The results indicate a double origin due to their presence in North Africa and the Iberian Peninsula. The formation of specific morphological groups within ecoregions is suggested.