Considering physiological and reproductive differences between European (Apis mellifera) and Africanized (Apis mellifera L.) honeybees, understanding sperm characteristics is essential for the application of assisted reproductive techniques. This work aimed to compare the morphological, morphometric and ultrastructural characteristics of spermatozoa from European and Africanized honeybee drones in the Caatinga biome. The experiment was conducted at the Laboratory of Animal Germplasm Conservation, Federal Rural University of the Semi-Arid Region (UFERSA), in Mossoró, RN, Brazil (5°03′37′′S;37°23′50′′W), in July-August/2024. Semen was collected from 9 European and 16 Africanized mature honeybee drones using the endophallus eversion technique. Samples were stained with rose Bengal solution to assess the percentage of normal spermatozoa under a light microscope and to measure the head, flagellum and total length using digital analysis software. Scanning electron microscopy was used to analyze sperm ultrastructure. Data were expressed as the mean ± standard error and compared by Student’s t-test for sperm morphology and the Mann–Whitney test for morphometric parameters (P<0,05). The percentages of normal spermatozoa in the Africanized and European drones were 12.06±1.01% and 10.89±1.66%, respectively, with no statistical difference; curled tail was the most prevalent defect observed. Morphometric measurements revealed significantly longer sperm heads in European drones (10.04±0.03µm) compared to Africanized drones (9.33±0.04µm; P < 0.05), although the flagellum and total length did not show significant differences, with a total length of approximately 260µm in both groups, aligning with the literature values. Ultrastructural evaluation allowed for a detailed identification of the conical acrosomal vesicle and the defined nucleus surface, which was not easily possible with conventional rose Bengal staining. In the head–flagellum coupling, the major mitochondrial derivative and the axoneme were visualized in parallel, but no differences were identified between groups. Although there is a morphometric difference in spermatozoa between European and Africanized drones, particularly in terms of head length, the overall morphological and ultrastructural characteristics are similar, suggesting a comparable basis for future applications in reproductive biotechnologies.