Impacts of sequential salinity and heat stress are recovery time-specific in freshwater crustacean, Daphnia pulicaria.

Stressors can interact to affect animal fitness, but we have limited knowledge about how temporal variation in stressors may impact their combined effect. This limits our ability to predict the outcomes of pollutants and future dynamic environmental changes. Elevated salinity in freshwater ecosystems has been observed worldwide. Meanwhile, heatwaves have become more frequent and intensified as an outcome of climate change. These two stressors can jointly affect organisms; however, their interaction has rarely been explored in the context of freshwater ecosystems. We conducted lab experiments using Daphnia pulicaria, a key species in lakes, to investigate how elevated salinity and heatwave conditions collectively affect freshwater organisms. We also monitored the impacts of various recovery times between the two stressors. Daphnia physiological conditions (metabolic rate, Na-K-ATPase (NKA) activity, and lipid peroxidation level) and life history traits (survival, fecundity, and growth) in response to salt stress as well as mortality in heat treatment were examined. We found that Daphnia responded to elevated salinity by upregulating NKA activity and increasing metabolic rate, causing a high lipid peroxidation level. Survival, fecundity, and growth were all negatively affected by this stressor. These impacts on physiological conditions and life history traits persisted for a few days after the end of the exposure. Heat treatments caused mortality in Daphnia, which increased with rising temperature. Results also showed that individuals that experienced salt exposure were more susceptible to subsequent heat stress, but this effect decreased with increasing recovery time between stressors. Findings from this work suggest that the legacy effects from a previous stressor can reduce individual resistance to a subsequent stressor, adding great difficulties to the prediction of outcomes of multiple stressors. Our work also demonstrates that cross-tolerance/susceptibility and the associated mechanisms remain unclear, necessitating further investigation.