Summary1. Natural populations are exposed to multiple stressors, including both anthropogenic challenges such as xenobiotics and natural stressors associated with exposure to parasites and predators. While there is increasing concern and interest in the combined impact of current exposure to multiple stressors, little attention has been given to how past exposure to a stressor and its evolutionary response shapes the effects of current stressors. 2. Here, we performed a life-table experiment using the water flea Daphnia magna to study combined effects of current exposure to the pesticide carbaryl, parasite spores and fish predation risk and how these effects depend upon past exposure to carbaryl using clones obtained from a previous carbaryl selection experiment. 3. The current exposure to all three treatments affected life-history traits. Exposure to fish kairomones increased intrinsic population growth rate, while carbaryl and parasite exposure decreased this fitness measure. The three treatments interacted only in a few cases: carbaryl and fish kairomone exposure interacted in shaping intrinsic population growth rate and its component individual reproductive performance, yet the latter only in the animals not exposed to carbaryl stress in the past. 4. Our data revealed not only adaptive evolution of carbaryl resistance but also associated evolutionary costs in terms of reduced resistance to parasites, corroborating results of an earlier study. Importantly, both the evolutionary benefits and costs of past exposure to carbaryl stress were conditional on current environmental conditions, exposure to predation risk and parasites, respectively. 5. The emerging pattern showed that past stress interacted with current stress in shaping life history. Such evolution-driven carry-over effects across generations have been often ignored and may complicate the prediction of effects of current exposure to single and combined stressors even long after the past stress has disappeared.

Microarrays are an ideal tool to screen for differences in gene expression of thousands of genes simultaneously. However, often commercial arrays are not available. In this study, we performed microarray analyses to evaluate patterns of gene transcription following exposure to two natural and one anthropogenic stressor. cDNA microarrays compiled of three life stage specific and three stressor-specific EST libraries, yielding 1734 different EST sequences, were used. We exposed juveniles of the water flea Daphnia magna for 48, 96 and 144 h to three stressors known to exert strong selection in natural populations of this species i.e. a sublethal concentration of the pesticide carbaryl, infective spores of the endoparasite Pasteuria ramosa, and fish predation risk mimicked by exposure to fish kairomones. A total of 148 gene fragments were differentially expressed compared to the control. Based on a PCA, the exposure treatments were separated into two main groups based on the extent of the transcriptional response: a low and a high (144 h of fish or carbaryl exposure and 96 h of parasite exposure) stress group. Firstly, we observed a general stress-related transcriptional expression profile independent of the treatment characterized by repression of transcripts involved in transcription, translation, signal transduction and energy metabolism. Secondly, we observed treatment-specific responses including signs of migration to deeper water layers in response to fish predation, structural challenge of the cuticle in response to carbaryl exposure, and disturbance of the ATP production in parasite exposure. A third important conclusion is that transcription expression patterns exhibit stress-specific changes over time. Parasite exposure shows the most differentially expressed gene fragments after 96 h. The peak of differentially expressed transcripts came only after 144 h of fish exposure, while carbaryl exposure induced a more stable number of differently expressed gene fragments over time.