Original Milou-project b2013028 Indirect interactions where one species mediate interactions between two other species are common in food webs. Not surprisingly, research on such indirect interactions are common, but few studies recognize that such interactions are often evolutionary unstable and that the occurrence of such interactions therefore may vary over time. In evolutionary time, speciation and diet changes in either predator or prey may cause indirect interactions to either appear or disappear. In ecological time, range shifts or species exclusions may have similar consequences. In order to understand the temporal variation in predator-mediated interactions, we need to understand the evolution or species traits connected to the interaction as a consequence of the interaction and how this in turn affects the topology of the food web. In host-parasitoid system, the key traits involved include traits connected to parasitoid host selection, host immune responses and parasitoid virulence. Consequently, in order to understand the underlying mechanisms, it is necessary to connect ecology, evolution and immunology in studies of the same system. In this project, we will use a model system that is very promising for these studies. First, we have previously shown that the parasitoid Asecodes parviclava may mediate indirect interactions between its host species, larvae of three beetle species (Galerucella calmariensis, G. pusilla and G. tenella). Second, we have also characterised the immune system of the larvae, particularly the encapsulation response to parasitoid eggs, and shown fairly large differences in immunocompetence between the host species. Third, we have identified geographic variation in parasitoid virulence and parasitoid host race formation suggesting on-going evolution of relevant traits. Finally, we discovered that female parasitoids have at least limited capacities to identify host larval odors. Within this project, we will first attempt to answer the question why we find much higher parasitism rates in northern (>70%) compared to southern (<10%) localities and test if this is connected to changes in the food web structure. The highly resistant species (G. pusilla) only occurs in the south. Second, we will then study the geographic mosaic of parasitoid virulence that we have previously observed in the southern area, and test if this variation is connected to the frequency of resistant versus susceptible host species. Our hypothesis is that high frequency of the resistant species would cause selection of higher virulence in the parasitoid. Across the same gradient of species frequencies, we will also study changes in the strength of indirect interactions. To connect these observation to the genome, we will perform RNAsequencing for transcriptome assembly in order to study gene expression between sites, and between control and infested individuals. Finally, we will perform RAD-sequencing in order to study the spatial structure of parasitoid populations and the phylogeography of parasitoid virulence traits.