Dynamics of Gut Bacteria Across Different Zooplankton Genera in the Baltic Sea.

In aquatic ecosystems, zooplankton-associated bacteria potentially have a great impact on the structure of ecosystems and trophic networks by providing various metabolic pathways and altering the ecological niche of host species. To understand the composition and drivers of zooplankton gut microbiota, we investigated the associated microbial communities of four zooplankton genera from different seasons in the Baltic Sea using the 16S rRNA gene. Among the 143 ASVs (amplified sequence variants) observed belonging to heterotrophic bacteria, 28 ASVs were shared across all zooplankton hosts over the season, and these shared core ASVs represented more than 25% and up to 60% of relative abundance in zooplankton hosts but were present at low relative abundance in the filtered water. Zooplankton host identity had stronger effects on bacterial composition than seasonal variation, with the composition of gut bacterial communities showing host-specific clustering patterns. Although bacterial compositions and dominating core bacteria were different between zooplankton hosts, higher gut bacteria diversity and more bacteria contributing to the temporal variation were found in Temora and Pseudocalanus, compared to Acartia and Synchaeta. Diet diatom and filamentous cyanobacteria negatively correlated with gut bacteria diversity, but the difference in diet composition did not explain the dissimilarity of gut bacteria composition, suggesting a general effect of diet on the inner conditions in the zooplankton gut. Synchaeta maintained high stability of gut bacterial communities with unexpectedly low bacteria-bacteria interactions as compared to the copepods, indicating host-specific regulation traits. Our results suggest that the patterns of gut bacteria dynamics are host-specific and the variability of gut bacteria is not only related to host taxonomy but also related to host behavior and life history traits.

Host Phylogeny Structures the Gut Bacterial Community Within Galerucella Leaf Beetles.

Gut microbes play important roles for their hosts. Previous studies suggest that host-microbial systems can form long-term associations over evolutionary time and the dynamic changes of the intestinal system may represent major driving forces and contribute to insect dietary diversification and speciation. Our study system includes a set of six closely related leaf beetle species (Galerucella spp.) and our study aims to separate the roles of host phylogeny and ecology in determining the gut microbial community and to identify eventual relationship between host insects and gut bacteria. We collected adult beetles from their respective host plants and quantified their microbial community using 16S rRNA sequencing. The results showed that the gut bacteria community composition was structured by host beetle phylogeny, where more or less host-specific gut bacteria interact with the different Galerucella species. For example, the endosymbiotic bacteria Wolbachia was found almost exclusively in G. nymphaea and G. sagittariae. Diversity indicators also suggested that α- and ß-diversities of gut bacteria communities varied among host beetle species. Overall, our results suggest a phylogenetically controlled co-occurrence pattern between the six closely related Galerucella beetles and their gut bacteria, indicating the potential of co-evolutionary processes occurring between hosts and their gut bacterial communities.

Opportunities to reduce pollination deficits and address production shortfalls in an important insect-pollinated crop.

Pollinators face multiple pressures and there is evidence of populations in decline. As demand for insect-pollinated crops increases, crop production is threatened by shortfalls in pollination services. Understanding the extent of current yield deficits due to pollination and identifying opportunities to protect or improve crop yield and quality through pollination management is therefore of international importance. To explore the extent of "pollination deficits," where maximum yield is not being achieved due to insufficient pollination, we used an extensive dataset on a globally important crop, apples. We quantified how these deficits vary between orchards and countries and we compared "pollinator dependence" across different apple varieties. We found evidence of pollination deficits and, in some cases, risks of overpollination were even apparent for which fruit quality could be reduced by too much pollination. In almost all regions studied we found some orchards performing significantly better than others in terms of avoiding a pollination deficit and crop yield shortfalls due to suboptimal pollination. This represents an opportunity to improve production through better pollinator and crop management. Our findings also demonstrated that pollinator dependence varies considerably between apple varieties in terms of fruit number and fruit quality. We propose that assessments of pollination service and deficits in crops can be used to quantify supply and demand for pollinators and help to target local management to address deficits although crop variety has a strong influence on the role of pollinators.

Seasonally varying marine influences on the coastal ecosystem detected through molecular gut analysis.

Terrestrial predators on marine shores benefit from the inflow of organisms and matter from the marine ecosystem, often causing very high predator densities and indirectly affecting the abundance of other prey species on shores. This indirect effect may be particularly strong if predators shift diets between seasons. We therefore quantified the seasonal variation in diet of two wolf spider species that dominate the shoreline predator community, using molecular gut content analyses with general primers to detect the full prey range. Across the season, spider diets changed, with predominantly terrestrial prey from May until July and predominantly marine prey (mainly chironomids) from August until October. This pattern coincided with a change in the spider age and size structure, and prey abundance data and resource selection analyses suggest that the higher consumption of chironomids during autumn is due to an ontogenetic diet shift rather than to variation in prey abundance. The analyses suggested that small dipterans with a weak flight capacity, such as Chironomidae, Sphaeroceridae, Scatopsidae and Ephydridae, were overrepresented in the gut of small juvenile spiders during autumn, whereas larger, more robust prey, such as Lepidoptera, Anthomyidae and Dolichopodidae, were overrepresented in the diet of adult spiders during spring. The effect of the inflow may be that the survival and growth of juvenile spiders is higher in areas with high chironomid abundances, leading to higher densities of adult spiders and higher predation rates on the terrestrial prey next spring.

A random survival forest illustrates the importance of natural enemies compared to host plant quality on leaf beetle survival rates.

BACKGROUND: Wetlands are habitats where variation in soil moisture content and associated environmental conditions can strongly affect the survival of herbivorous insects by changing host plant quality and natural enemy densities. In this study, we combined natural enemy exclusion experiments with random survival forest analyses to study the importance of local variation in host plant quality and predation by natural enemies on the egg and larval survival of the leaf beetle Galerucella sagittariae along a soil moisture gradient. RESULTS: Our results showed that the exclusion of natural enemies substantially increased the survival probability of G. sagittariae eggs and larvae. Interestingly, the egg survival probability decreased with soil moisture content, while the larval survival probability instead increased with soil moisture content. For both the egg and larval survival, we found that host plant height, the number of eggs or larvae, and vegetation height explained more of the variation than the soil moisture gradient by itself. Moreover, host plant quality related variables, such as leaf nitrogen, carbon and phosphorus content did not influence the survival of G. sagittariae eggs and larvae. CONCLUSION: Our results suggest that the soil moisture content is not an overarching factor that determines the interplay between factors related to host plant quality and factors relating to natural enemies on the survival of G. sagittariae in different microhabitats. Moreover, the natural enemy exclusion experiments and the random survival forest analysis suggest that natural enemies have a stronger indirect impact on the survival of G. sagittariae offspring than host plant quality.

A heterogeneous landscape does not guarantee high crop pollination.

The expansion of pollinator-dependent crops, especially in the developing world, together with reports of worldwide pollinator declines, raises concern of possible yield gaps. Farmers directly reliant on pollination services for food supply often live in regions where our knowledge of pollination services is poor. In a manipulative experiment replicated at 23 sites across an Ethiopian agricultural landscape, we found poor pollination services and severe pollen limitation in a common oil crop. With supplementary pollination, the yield increased on average by 91%. Despite the heterogeneous agricultural matrix, we found a low bee abundance, which may explain poor pollination services. The variation in pollen limitation was unrelated to surrounding forest cover, local bee richness and bee abundance. While practices that commonly increase pollinators (restricted pesticide use, flower strips) are an integral part of the landscape, these elements are apparently insufficient. Management to increase pollination services is therefore in need of urgent investigation.

Getting the smell of it--odour cues structure pollinator networks.

Floral visitors vary greatly among plant species and depend on the volatiles emitted by the flowers. Creeping thistle is normally visited by bees and bumblebees while common yarrow is rather visited by flies. Manipulating the flower volatiles caused pollinator communities to become more similar among the two plant species. Image credit: Robert Junker and Anna-Amelie Larue. In Focus: Larue, A.-A.C., Raguso, R.A. & Junker, R.R. (2015) Experimental manipulation of floral scent bouquets restructures flower-visitor interactions in the field. Journal of Animal Ecology, 85, 396-408. Pollinators use multiple cues to locate suitable flowers, and recent studies argue that flower volatiles are more important than previously believed. However, the role of volatiles is seldom separated from other cues. Larue, Raguso & Junker (2015) manipulated the volatile profile of two plants that are normally visited by different pollinators. Achillea millefolium is normally not visited by honeybees and bumblebees, but these pollinator groups did visit plants that were sprayed with volatiles from Cirsium arvense. Cirsium arvense, on the other hand, was less visited by honeybees and bumblebees when sprayed with volatiles from A. millefolium. These findings highlight the potential role of volatiles in structuring pollinator communities on plants.

Geographic variation and trade-offs in parasitoid virulence.

Host-parasitoid systems are characterized by a continuous development of new defence strategies in hosts and counter-defence mechanisms in parasitoids. This co-evolutionary arms race makes host-parasitoid systems excellent for understanding trade-offs in host use caused by evolutionary changes in host immune responses and parasitoid virulence. However, knowledge obtained from natural host-parasitoid systems on such trade-offs is still limited. In this study, the aim was to examine trade-offs in parasitoid virulence in Asecodes parviclava (Hymenoptera: Eulophidae) when attacking three closely related beetles: Galerucella pusilla, Galerucella calmariensis and Galerucella tenella (Coleoptera: Chrysomelidae). A second aim was to examine whether geographic variation in parasitoid infectivity or host immune response could explain differences in parasitism rate between northern and southern sites. More specifically, we wanted to examine whether the capacity to infect host larvae differed depending on the previous host species of the parasitoids and if such differences were connected to differences in the induction of host immune systems. This was achieved by combining controlled parasitism experiments with cytological studies of infected larvae. Our results reveal that parasitism success in A. parviclava differs both depending on previous and current host species, with a higher virulence when attacking larvae of the same species as the previous host. Virulence was in general high for parasitoids from G. pusilla and low for parasitoids from G. calmariensis. At the same time, G. pusilla larvae had the strongest immune response and G. calmariensis the weakest. These observations were linked to changes in the larval hemocyte composition, showing changes in cell types important for the encapsulation process in individuals infected by more or less virulent parasitoids. These findings suggest ongoing evolution in parasitoid virulence and host immune response, making the system a strong candidate for further studies on host race formation and speciation.

Herbivory strongly influences among-population variation in reproductive output of Lythrum salicaria in its native range.

Herbivory can negatively affect several components of plant reproduction. Yet, because of a lack of experimental studies involving multiple populations, the extent to which differences in herbivory contribute to among-population variation in plant reproductive success is poorly known. We experimentally determined the effects of insect herbivory on reproductive output in nine natural populations of the perennial herb Lythrum salicaria along a disturbance gradient in an archipelago in northern Sweden, and we quantified among-population differentiation in resistance to herbivory in a common-garden experiment in the same area. The intensity of leaf herbivory varied >500-fold and mean female reproductive success >400-fold among the study populations. The intensity of herbivory was lowest in populations subject to strong disturbance from ice and wave action. Experimental removal of insect herbivores showed that the effect of herbivory on female reproductive success was correlated with the intensity of herbivory and that differences in insect herbivory could explain much of the among-population variation in the proportion of plants flowering and seed production. Population differentiation in resistance to herbivory was limited. The results demonstrate that the intensity of herbivory is a major determinant of flowering and seed output in L. salicaria, but that differences in herbivory are not associated with differences in plant resistance at the spatial scale examined. They further suggest that the physical disturbance regime may strongly influence the performance and abundance of perennial herbs and patterns of selection not only because of its effect on interspecific competition, but also because of effects on interactions with specialized herbivores.

Plant patch structure influences plant fitness via antagonistic and mutualistic interactions but in different directions.

Plant patch structure and environmental context can influence the outcome of antagonistic and mutualistic plant-insect interactions, leading to spatially variable fitness effects for plants. We investigated the effects of herbivory and pollen limitation on plant reproductive performance in 28 patches of the self-compatible perennial herb Scrophularia nodosa and assessed how such effects varied with plant patch size, plant density and tree cover. Both antagonistic and mutualistic interactions had strong effects on plant reproductive performance. Leaf feeding from herbivores reduced both fruit production and seed germination, and leaf herbivory increased with plant patch size. Experimentally hand-pollinated flowers produced more seeds than open-pollinated flowers, and pollen limitation was more severe in patches with fewer plants. Our study on S. nodosa is one of few which documents that plant patch structure influences the outcome of both antagonistic and mutualistic plant-insect interactions. The results thus provide an example of how variation in plant patch structure and environmental factors can lead to spatially variable fitness effects from mutualistic and antagonistic interactions.

Effects of plant neighborhoods on plant-herbivore interactions: resource dilution and associational effects.

Effects of neighboring plants on herbivore damage to a focal plant (associational effects) have been documented in many systems and can lead to either increased or decreased herbivore attack. Mechanistic models that explain the observed variety of herbivore responses to local plant community composition have, however, been lacking. We present a model of herbivore responses to patches that consist of two plant types, where herbivore densities on a focal plant are determined by a combination of patch-finding, within-patch redistribution, and patch-leaving. Our analyses show that the effect of plant neighborhood on herbivores depends both on how plant and herbivore traits combine to affect herbivore movement and on how experimental designs reveal the effects of plant density and plant relative frequency. Associational susceptibility should be the dominant pattern when herbivores have biased landing rates within patches. Other behavioral decision rules lead to mixed responses, but a common pattern is that in mixed patches, one plant type experiences associational resistance while the other plant experiences associational susceptibility. In some cases, the associational effect may shift sign along a gradient of plant frequency, suggesting that future empirical studies should include more than two plant frequencies to detect nonlinearities. Finally, we find that associational susceptibility should be commonly observed in experiments using replacement designs, whereas associational resistance will be the dominant pattern when using additive designs. Consequently, outcomes from one experimental design cannot be directly compared to studies with other designs. Our model can also be translated to other systems with foragers searching for multiple resource types.

Bayesian species delimitation reveals generalist and specialist parasitic wasps on Galerucella beetles (Chrysomelidae): sorting by herbivore or plant host.

BACKGROUND: To understand the ecological and evolutionary consequences of species interactions in food webs necessitates that interactions are properly identified. Genetic analyses suggest that many supposedly generalist parasitoid species should rather be defined as multiple species with a more narrow diet, reducing the probability that such species may mediate indirect interactions such as apparent competition among hosts. Recent studies showed that the parasitoid Asecodes lucens mediate apparent competition between two hosts, Galerucella tenella and G. calmariensis, affecting both interaction strengths and evolutionary feedbacks. The same parasitoid was also recorded from other species in the genus Galerucella, suggesting that similar indirect effects may also occur for other species pairs. METHODS: To explore the possibility of such interactions, we sequenced mitochondrial and nuclear genetic markers to resolve the phylogeny of both host and parasitoid and to test the number of parasitoid species involved. We thus collected 139 Galerucella larvae from 8 host plant species and sequenced 31 adult beetle and 108 parasitoid individuals. RESULTS: The analysis of the Galerucella data, that also included sequences from previous studies, verified the five species previously documented as reciprocally monophyletic, but the Bayesian species delimitation for A. lucens suggested 3-4 cryptic taxa with a more specialised host use than previously suggested. The gene data analyzed under the multispecies coalescent model allowed us to reconstruct the species tree phylogeny for both host and parasitoid and we found a fully congruent coevolutionary pattern suggesting that parasitoid speciation followed upon host speciation. CONCLUSION: Using multilocus sequence data in a Bayesian species delimitation analysis we propose that hymenopteran parasitoids of the genus Asecodes that infest Galerucella larvae constitute at least three species with narrow diet breath. The evolution of parasitoid Asecodes and host Galerucella show a fully congruent coevolutionary pattern. This finding strengthens the hypothesis that the parasitoid in host search uses cues of the host rather than more general cues of both host and plant.

Insect density-plant density relationships: a modified view of insect responses to resource concentrations.

Habitat area is an important predictor of spatial variation in animal densities. However, the area often correlates with the quantity of resources within habitats, complicating our understanding of the factors shaping animal distributions. We addressed this problem by investigating densities of insect herbivores in habitat patches with a constant area but varying numbers of plants. Using a mathematical model, predictions of scale-dependent immigration and emigration rates for insects into patches with different densities of host plants were derived. Moreover, a field experiment was conducted where the scaling properties of odour-mediated attraction in relation to the number of odour sources were estimated, in order to derive a prediction of immigration rates of olfactory searchers. The theoretical model predicted that we should expect immigration rates of contact and visual searchers to be determined by patch area, with a steep scaling coefficient, µ = -1. The field experiment suggested that olfactory searchers should show a less steep scaling coefficient, with µ ≈ -0.5. A parameter estimation and analysis of published data revealed a correspondence between observations and predictions, and density-variation among groups could largely be explained by search behaviour. Aphids showed scaling coefficients corresponding to the prediction for contact/visual searchers, whereas moths, flies and beetles corresponded to the prediction for olfactory searchers. As density responses varied considerably among groups, and variation could be explained by a certain trait, we conclude that a general theory of insect responses to habitat heterogeneity should be based on shared traits, rather than a general prediction for all species.

Wetland productivity determines trade-off between biodiversity support and greenhouse gas production.

Establishing wetlands for nutrient capture and biodiversity support may introduce trade-offs between environmentally beneficial functions and detrimental greenhouse gas emissions. Investigating the interaction of nutrient capture, primary production, greenhouse gas production and biodiversity support is imperative to understanding the overall function of wetlands and determining possible beneficial synergistic effects and trade-offs. Here, we present temporally replicated data from 17 wetlands in hemi-boreal Sweden. We explored the relationship between nutrient load, primary producing algae, production of methane and nitrous oxide, and emergence rates of chironomids to determine what factors affected each and how they related to each other. Chironomid emergence rates correlated positively with methane production and negatively with nitrous oxide production, where water temperature was the main driving factor. Increasing nutrient loads reduced methanogenesis through elevated nitrogen concentrations, while simultaneously enhancing nitrous oxide production. Nutrient loads only indirectly increased chironomid emergence rates through increased chlorophyll-a concentration, via increased phosphorus concentrations, with certain taxa and food preference functional groups benefitting from increased chlorophyll-a concentrations. However, water temperature seemed to be the main driving factor for chironomid emergence rates, community composition and diversity, as well as for greenhouse gas production. These findings increase our understanding of the governing relationships between biodiversity support and greenhouse gas production, and should inform future management when constructing wetlands.

Density-Dependent Effects of Simultaneous Root and Floral Herbivory on Plant Fitness and Defense.

Plants are attacked by multiple herbivores, and depend on a precise regulation of responses to cope with a wide range of antagonists. Simultaneous herbivory can occur in different plant compartments, which may pose a serious threat to plant growth and reproduction. In particular, plants often face co-occurring root and floral herbivory, but few studies have focused on such interactions. Here, we investigated in the field the combined density-dependent effects of root-chewing cebrionid beetle larvae and flower-chewing pierid caterpillars on the fitness and defense of a semiarid Brassicaceae herb. We found that the fitness impact of both herbivore groups was independent and density-dependent. Increasing root herbivore density non-significantly reduced plant fitness, while the relationship between increasing floral herbivore density and the reduction they caused in both seed number and seedling emergence was non-linear. The plant defensive response was non-additive with regard to the different densities of root and floral herbivores; high floral herbivore density provoked compensatory investment in reproduction, and this tolerance response was combined with aboveground chemical defense induction when also root herbivore density was high. Plants may thus prioritize specific trait combinations in response to varying combined below- and aboveground herbivore densities to minimize negative impacts on fitness.

Species composition of shoreline wolf spider communities vary with salinity, but their diets vary with wrack inflow.

Wolf spiders are typically the most common group of arthropod predators on both lake and marine shorelines because of the high prey availability in these habitats. However, shores are also harsh environments due to flooding and, in proximity to marine waters, to toxic salinity levels. Here, we describe the spider community, prey availabilities, and spider diets between shoreline sites with different salinities, albeit with comparatively small differences (5‰ vs. 7‰). Despite the small environmental differences, spider communities between lower and higher saline sites showed an almost complete species turnover. At the same time, differences in prey availability or spider gut contents did not match changes in spider species composition but rather changed with habitat characteristics within a region, where spiders collected at sites with thick wrack beds had a different diet than sites with little wrack. These data suggest that shifts in spider communities are due to habitat characteristics other than prey availabilities, and the most likely candidate restricting species in high salinity would be saline sensitivity. At the same time, species absence from low-saline habitats remains unresolved.

The role of inputs of marine wrack and carrion in sandy-beach ecosystems: a global review.

Sandy beaches are iconic interfaces that functionally link the ocean with the land via the flow of organic matter from the sea. These cross-ecosystem fluxes often comprise uprooted seagrass and dislodged macroalgae that can form substantial accumulations of detritus, termed 'wrack', on sandy beaches. In addition, the tissue of the carcasses of marine animals that regularly wash up on beaches form a rich food source ('carrion') for a diversity of scavenging animals. Here, we provide a global review of how wrack and carrion provide spatial subsidies that shape the structure and functioning of sandy-beach ecosystems (sandy beaches and adjacent surf zones), which typically have little in situ primary production. We also examine the spatial scaling of the influence of these processes across the broader land- and seascape, and identify key gaps in our knowledge to guide future research directions and priorities. Large quantities of detrital kelp and seagrass can flow into sandy-beach ecosystems, where microbial decomposers and animals process it. The rates of wrack supply and its retention are influenced by the oceanographic processes that transport it, the geomorphology and landscape context of the recipient beaches, and the condition, life history and morphological characteristics of the macrophyte taxa that are the ultimate source of wrack. When retained in beach ecosystems, wrack often creates hotspots of microbial metabolism, secondary productivity, biodiversity, and nutrient remineralization. Nutrients are produced during wrack breakdown, and these can return to coastal waters in surface flows (swash) and aquifers discharging into the subtidal surf. Beach-cast kelp often plays a key trophic role, being an abundant and preferred food source for mobile, semi-aquatic invertebrates that channel imported algal matter to predatory invertebrates, fish, and birds. The role of beach-cast marine carrion is likely to be underestimated, as it can be consumed rapidly by highly mobile scavengers (e.g. foxes, coyotes, raptors, vultures). These consumers become important vectors in transferring marine productivity inland, thereby linking marine and terrestrial ecosystems. Whilst deposits of organic matter on sandy-beach ecosystems underpin a range of ecosystem functions and services, they can be at variance with aesthetic perceptions resulting in widespread activities, such as 'beach cleaning and grooming'. This practice diminishes the energetic base of food webs, intertidal fauna, and biodiversity. Global declines in seagrass beds and kelp forests (linked to global warming) are predicted to cause substantial reductions in the amounts of marine organic matter reaching many beach ecosystems, likely causing flow-on effects for food webs and biodiversity. Similarly, future sea-level rise and increased storm frequency are likely to alter profoundly the physical attributes of beaches, which in turn can change the rates at which beaches retain and process the influxes of wrack and animal carcasses. Conservation of the multi-faceted ecosystem services that sandy beaches provide will increasingly need to encompass a greater societal appreciation and the safeguarding of ecological functions reliant on beach-cast organic matter on innumerable ocean shores worldwide.

Genome assemblies of three closely related leaf beetle species (Galerucella spp.).

Galerucella (Coleoptera: Chrysomelidae) is a leaf beetle genus that has been extensively used for ecological and evolutionary studies. It has also been used as biological control agent against invading purple loosestrife in North America, with large effects on biodiversity. Here, we report genome assembly and annotation of three closely related Galerucella species: G. calmariensis, G. pusilla, and G. tenella. The three assemblies have a genome size ranging from 460 to 588 Mbp, with N50 from 31,588 to 79,674 kbp, containing 29,202 to 40,929 scaffolds. Using an ab initio evidence-driven approach, 30,302 to 33,794 protein-coding genes were identified and functionally annotated. These draft genomes will contribute to the understanding of host-parasitoid interactions, evolutionary comparisons of leaf beetle species and future population genomics studies.

Differential Expression of Immune Genes between Two Closely Related Beetle Species with Different Immunocompetence following Attack by Asecodes parviclava.

Endoparasitoid wasps are important natural enemies of many insect species and are major selective forces on the host immune system. Despite increased interest in insect antiparasitoid immunity, there is sparse information on the evolutionary dynamics of biological pathways and gene regulation involved in host immune defense outside Drosophila species. We de novo assembled transcriptomes from two beetle species and used time-course differential expression analysis to investigate gene expression differences in closely related species Galerucella pusilla and G. calmariensis that are, respectively, resistant and susceptible against parasitoid infection by Asecodes parviclava parasitoids. Approximately 271 million and 224 million paired-ended reads were assembled and filtered to form 52,563 and 59,781 transcripts for G. pusilla and G. calmariensis, respectively. In the whole-transcriptome level, an enrichment of functional categories related to energy production, biosynthetic process, and metabolic process was exhibited in both species. The main difference between species appears to be immune response and wound healing process mounted by G. pusilla larvae. Using reciprocal BLAST against the Drosophila melanogaster proteome, 120 and 121 immune-related genes were identified in G. pusilla and G. calmariensis, respectively. More immune genes were differentially expressed in G. pusilla than in G. calmariensis, in particular genes involved in signaling, hematopoiesis, and melanization. In contrast, only one gene was differentially expressed in G. calmariensis. Our study characterizes important genes and pathways involved in different immune functions after parasitoid infection and supports the role of signaling and hematopoiesis genes as key players in host immunity in Galerucella against parasitoid wasps.