To identify some of the mechanisms involved in the high resistance to Cd(2+) in the protist Euglena gracilis, we studied the effect of Cd(2+) exposure on its energy and oxidative stress metabolism as well as on essential heavy metals homeostasis. In E. gracilis heterotrophic cells, as in other organisms, CdCl(2) (50 microM) induced diminution in cell growth, severe oxidative stress accompanied by increased antioxidant enzyme activity and strong perturbation of the heavy metal homeostasis. However, Cd(2+) exposure did not substantially modify the cellular respiratory rate or ATP intracellular level, although the activities of respiratory complexes III and IV were strongly decreased. In contrast, an enhanced capacity of the alternative oxidase (AOX) in both intact cells and isolated mitochondria was determined under Cd(2+) stress; in fact, AOX activity accounted for 69-91% of total respiration. Western blotting also revealed an increased AOX content in mitochondria from Cd(2+)-exposed cells. Moreover, AOX was more resistant to Cd(2+) inhibition than cytochrome c oxidase in mitochondria from control and Cd(2+)-exposed cells. Therefore, an enhanced AOX seems to be a relevant component of the resistance mechanism developed by E. gracilis against Cd(2+)-stress, in addition to the usual increased antioxidant enzyme activity, that enabled cells to maintain a relatively unaltered the energy status.