SUMMARYThe overall aim of this study was to (1) evaluate the adaptive value of mitochondrial DNA by comparing mitochondrial performance in populations possessing different haplotypes and distribution, and to (2) evaluate the sensitivity of different enzymes of the electron transport system (ETS) during temperature-induced changes. We measured the impact of temperature of mitochondrial respiration and several key enzymes of mitochondrial metabolism in two mitotypes (siII and siIII) of Drosophila simulans. The temperature dependencies of oxygen consumption for mitochondria isolated from flight muscle was assessed with complex I substrates (pyruvate + malate + proline) and with sn glycerol-3-phosphate (to reduce complex III via glycerophosphate dehydrogenase) in both coupled and uncoupled states. Activities of citrate synthase, cytochrome c oxidase (COX), catalase and aconitase, and the excess capacity of COX at high convergent pathway flux were also measured as a function of temperature. Overall, our results showed that functional differences between the two mitotypes are few. Results suggest that differences between the two mitotypes could hardly explain the temperature-specific differences measured in mitochondria performances. It suggests that some other factor(s) may be driving the maintenance of mitotypes. We also show that the different enzymes of the ETS have different thermal sensitivities. The catalytic capacities of these enzymes vary with temperature changes, and the corresponding involvement of the different steps on mitochondrial regulation probably varies with temperature. For example, the excess COX capacity is low, even non-existent, at high and intermediate temperatures (18°C, 24°C and 28°C) whereas it is quite high at a lower temperature (12°C), suggesting release of respiration control by COX at low temperature.