Polyploidy plays a major role in citrus plant breeding to improve the adaptation of polyploid rootstocks as well as scions to adverse conditions and to enhance agronomic characteristics. In Citrus breeding programs, triploidy could be a useful tool to react to environmental issues and consumer demands because the produced fruits are seedless. In this study, we compared the physiological, biochemical, morphological, and ultrastructural responses to water deficit of triploid and diploid citrus varieties obtained from 'Fortune' mandarin and 'Ellendale' tangor hybridization. One diploid clementine tree was included and used as a reference. All studied scions were grafted on C-35 citrange rootstock. Triploidy decreased stomatal density and increased stomata size. The number of chloroplasts increased in 3x varieties. These cytological properties may explain the greater photosynthetic capacity (Pnet, gs, Fv/Fm) and enhanced water-holding capacity (RWC, proline). In addition, reduced degradation of ultrastructural organelles (chloroplasts and mitochondria) and thylakoids accompanied by less photosynthetic activity and low oxidative damages were found in 3x varieties. Triploid varieties, especially T40-3x, had a better ability to limit water loss and dissipate excess energy (NPQ) to protect photosystems. Higher starch reserves in 3x varieties suggest a better carbon and energy supply and increases in plastoglobuli size suggest less oxidative damage (H2O2, MDA), especially in T40-3x, and preservation of photosynthetic apparatus. Taken together, our results suggest that desirable cytological and ultrastructural traits induced by triploidy improve water stress response and could be a useful stress marker during environmental constraints.