ROS in wide temperature range

Temperature is a strong selection pressure, especially for ectothermic organisms where it affects everything from metabolic rate to behavior, development, reproduction, and survival. Temperature influences organismal fitness by affecting cellular processes and stress responses. Reactive oxygen species (ROS) are unavoidable by-products of cellular metabolism, but their production is markedly elevated under thermal stress, leading to oxidative damage and compromising cellular integrity. Previous studies have largely relied on indirect measurements or single-temperature point assessments of oxidative stress. This study provides the first systematic in vivo measurement of ROS production across a temperature gradient (4°C to 28°C), which is experienced by the insect Chironomus riparius in its natural environment. Using in vivo fluorescence measurements of various ROS, we revealed a U-shaped relationship between temperature and ROS production, with ROS levels increasing at extreme temperatures. At higher temperatures (24°C and 28°C), ROS composition shifted, with hydrogen peroxide, peroxynitrite anion, and nitric oxide more abundant while superoxide anion and hydroxyl radicals were dominating at 4°C. Transcriptome analysis revealed the organism's specific molecular response to oxidative stress, showing upregulation of specific antioxidant genes at higher temperatures, particularly those involved in hydrogen peroxide metabolism, and a more general response at lower temperatures. The correlation between the observed U-shaped ROS pattern and previously reported U-shaped mutation rates highlights the potential role of temperature-induced oxidative stress as an important driver of genomic variability in natural populations.