The comb cell structure of Apis dorsata in different ecoregions is rarely reported. This study compared the differences in the comb cell structure of Apis dorsata from Menghai of Yunnan Province and Chengmai of Hainan Province, China. Accordingly, parameters such as brood comb thickness, the width of ten consecutive brood cells, cell diameter, depth, inclination angle, and the side length of the cell mouth were measured. The results showed that within the same area, the widths of ten consecutive cells and single-cell diameter in the 0° direction were significantly bigger than that in the 60° and 120° directions, however, there was no significant difference for the same between the 60° and 120° directions. The average width of ten consecutive cells and the single-cell diameter showed significant differences between the two areas. These were significantly bigger in Menghai than in Chengmai. The average thickness of the brood comb, cell depth, and side length of the cell mouth were also significantly higher in Menghai than in Chengmai, however, the cell inclination angle showed no regional difference. These differences in the comb cell structure of Apis dorsata may be due to the difference in the ecological environment of the two areas. This study revealed the structural characteristics of the comb cells of Apis dorsata in different areas of China, increasing our biological knowledge of this bee species.