The role of divergent host use and geography in the evolution of habitat isolation and sexual isolation among sister species of Belonocnema gall wasps.

Ecology and geography can play important roles in the evolution of reproductive isolation across the speciation continuum, but few studies address both at the later stages of speciation. This notable gap in knowledge arises from the fact that traditional ecological speciation studies have predominantly focused on the role of ecology in initiating the speciation process, while many studies exploring the effect of geography (e.g., reinforcement) concentrate on species pairs that lack divergent ecological characteristics. We simultaneously examine the strength of habitat isolation and sexual isolation among three closely related species of Belonocnema gall-forming wasps on two species of live oaks, Quercus virginiana and Q. geminata, that experience divergent selection from their host plants and variable rates of migration due to their geographic context. We find that the strength of both habitat isolation and sexual isolation is lowest among allopatric species pairs with the same host plant association, followed by allopatric species with different host plant associations, and highest between sympatric species with different host-plant associations. This pattern suggests that divergent selection due to different host use interacts with geography in the evolution of habitat isolation and sexual isolation during the later stages of speciation of Belonocnema wasps.

A test of trade-offs in dispersal and reproduction within and between a sister species pair of specialist insect herbivores.

Understanding the drivers of trade-offs among traits is vital for comprehending the evolution and maintenance of trait variation. Theoretical frameworks propose that evolutionary mechanisms governing trade-offs frequently exhibit a scale-dependent nature. However, empirical tests of whether trade-offs exhibited across various biological scales (i.e. individuals, populations, species, genera, etc.) remains scarce. In this study, we explore trade-off between dispersal and reproductive effort among sympatric sister species of wasps in the genus Belonocnema (Hymenoptera: Cynipini: Cynipidae) that form galls on live oaks: B. fossoria, which specializes on Quercus geminata, and B. treatae, which specializes on Q. virginiana. Specifically, our results suggest that B. fossoria has evolved reduced flight capability and smaller wings, but a larger abdomen and greater total reproductive effort than B. treatae, which has larger wings and is a stronger flier, but has a smaller abdomen and reduced total reproductive effort. These traits and the relationships among them remain unchanged when B. fossoria and B. treatae are transplanted and reared onto the alternative host plant, suggesting that trait divergence is genetically based as opposed to being a plastic response to the different rearing environments. However, when looking within species, we found no evidence of intraspecific trade-offs between wing length and reproductive traits within either B. fossoria or B. treatae. Overall, our results indicate that observed trade-offs in life history traits between the two gall former species are likely a result of independent adaptations in response to different environments as opposed to the amplified expression of within species intrinsic tradeoffs.

Environment and shipping drive environmental DNA beta-diversity among commercial ports.

The spread of nonindigenous species by shipping is a large and growing global problem that harms coastal ecosystems and economies and may blur coastal biogeographical patterns. This study coupled eukaryotic environmental DNA (eDNA) metabarcoding with dissimilarity regression to test the hypothesis that ship-borne species spread homogenizes port communities. We first collected and metabarcoded water samples from ports in Europe, Asia, Australia and the Americas. We then calculated community dissimilarities between port pairs and tested for effects of environmental dissimilarity, biogeographical region and four alternative measures of ship-borne species transport risk. We predicted that higher shipping between ports would decrease community dissimilarity, that the effect of shipping would be small compared to that of environment dissimilarity and shared biogeography, and that more complex shipping risk metrics (which account for ballast water and stepping-stone spread) would perform better. Consistent with our hypotheses, community dissimilarities increased significantly with environmental dissimilarity and, to a lesser extent, decreased with ship-borne species transport risks, particularly if the ports had similar environments and stepping-stone risks were considered. Unexpectedly, we found no clear effect of shared biogeography, and that risk metrics incorporating estimates of ballast discharge did not offer more explanatory power than simpler traffic-based risks. Overall, we found that shipping homogenizes eukaryotic communities between ports in predictable ways, which could inform improvements in invasive species policy and management. We demonstrated the usefulness of eDNA metabarcoding and dissimilarity regression for disentangling the drivers of large-scale biodiversity patterns. We conclude by outlining logistical considerations and recommendations for future studies using this approach.

A genomic test of subspecies in the Eunota togata species group (Coleoptera: Cicindelidae): Morphology masks evolutionary relationships and taxonomy.

Most of the world's biodiversity is described primarily or exclusively using morphological traits that may not always reflect the true evolutionary units. Accurate taxonomy is critical for conservation efforts and re-evaluation of traditional taxonomy may often be warranted since species and subspecies are frequently the focus of conservation and faunistic studies. Here, we test comprehensive taxonomic hypotheses of morphologically defined subspecies in the tiger beetle, Eunota togata (LaFerté-Sénectère, 1841). The four recognized subspecies were delineated based mainly on the dorsal coloration and extent of white markings termed maculations. We combine inferences from mtDNA genealogies and genome-wide multilocus data to elucidate the evolutionary relationships within the group and assess the taxonomic implications. Three of the four subspecific taxa delineated by morphology were not supported by the genomic or mtDNA data. In fact, the species-level diversity in this group was underestimated, as E. togata was found to represent three well-supported distinct species in all genetic analyses. Emerging from these analyses, we also document an intriguing example of convergent evolution in lighter colored E. togata adapting to similar white saline backgrounds. Our collective work underscores the importance of using molecular methods to reevaluate morphological based taxonomy for species and subspecies delimitation and conservation.

Suspended Materials Affect Particle Size Distribution and Removal of Environmental DNA in Flowing Waters.

Environmental DNA (eDNA) in aquatic systems is a complex mixture that includes dissolved DNA, intracellular DNA, and particle-adsorbed DNA. Information about the various components of eDNA and their relative proportions could be used to discern target organism abundance and location. However, a limited knowledge of eDNA adsorption dynamics and interactions with other materials hinders these applications. To address this gap, we used recirculating stream mesocosms to investigate the impact of suspended materials (fine particulate organic matter, plankton, clay, and titanium dioxide) on the eDNA concentration and particle size distribution (PSD) from two fish species in flowing water. Our findings revealed that eDNA rapidly adsorbs to other materials in the water column, affecting its concentration and PSD. Nonetheless, only particulate organic matter affected eDNA removal rate after 30 h. Moreover, we observed that the removal of larger eDNA components (≥10 µm) was more strongly influenced by physical processes, whereas the removal of smaller eDNA components was driven by biological degradation. This disparity in removal mechanisms between larger and smaller eDNA components could explain changes in eDNA composition over time and space, which have implications for modeling the spatial distribution and abundance of target species and optimizing eDNA detection in high turbidity systems.

Novel Field-Based Protein Detection Method Using Light Transmission Spectroscopy and Antibody Functionalized Gold Nanoparticles.

Protein detection is a universal tool critical to many applications in medicine, agriculture, and biotechnology. We developed a novel protein detection method combining light transmission spectroscopy and particle-size analysis of gold nanospheres monovalently functionalized with polyclonal antibodies and applied it to an emerging challenge for such technologies─the monitoring of environmental proteins (eProteins) present in natural aquatic systems. These are an underreported source of pollution and include the pseudopersistent Cry toxins that enter aquatic ecosystems from surrounding genetically engineered crops. The assay is capable of detecting proteins in complex matrices, such as water samples collected in the field, making it a competitive assay for eProtein detection. It is sensitive, reaching 1.25 ng mL-1, and we demonstrate its application to the detection of Cry1Ab from subsurface tile-drain and streamwater samples from agricultural waterways. The assay can also be quickly adapted for other protein detection applications in the future.

Tissue-specific gene expression shows a cynipid wasp repurposes oak host gene networks to create a complex and novel parasite-specific organ.

Every organism on Earth depends on interactions with other organisms to survive. In each of these interactions, an organism must utilize the limited toolbox of genes and proteins it possesses to successfully manipulate or cooperate with another species, but it can also co-opt the genome machinery of its partner to expand its available tools. Insect-induced plant galls are an extreme example of this, wherein an insect hijacks the plant's genome to direct the initiation and development of galls consisting of plant tissue. However, previous transcriptomic studies have not evaluated individual tissues within a gall to determine the full extent to which a galling insect manipulates its host plant. Here we demonstrate that the cynipid wasp Dryocosmus quercuspalustris creates a complex parasite-specific organ from red oak tissue via massive changes in host gene expression. Our results show that the gall wasp is not merely modifying oak leaf tissue but creating extensive changes in gene expression between galled and ungalled tissue (differential expression in 28% of genes) and distinct gall tissue types (20% of genes). The outer gall tissue shows increases in various plant defence systems, which is consistent with its predicted functional role of protecting the wasp larva. The inner larval capsule shows suppression of large parts of the plant innate immune system and evidence for the wasp utilizing the plant's RNA interference mechanisms, which may be a potential mechanism for the wasp's control on gall growth.

Delimiting the cryptic diversity and host preferences of Sycophila parasitoid wasps associated with oak galls using phylogenomic data.

Cryptic species diversity is a major challenge regarding the species-rich community of parasitoids attacking oak gall wasps due to a high degree of sexual dimorphism, morphological plasticity, small size and poorly known biology. As such, we know very little about the number of species present, nor the evolutionary forces responsible for generating this diversity. One hypothesis is that trait diversity in the gall wasps, including the morphology of the galls they induce, has evolved in response to selection imposed by the parasitoid community, with reciprocal selection driving diversification of the parasitoids. Using a rare, continental-scale data set of Sycophila parasitoid wasps reared from 44 species of cynipid galls from 18 species of oak across the USA, we combined mitochondrial DNA barcodes, ultraconserved elements (UCEs), morphological and natural history data to delimit putative species. Using these results, we generate the first large-scale assessment of ecological specialization and host association in this species-rich group, with implications for evolutionary ecology and biocontrol. We find most Sycophila target specific subsets of available cynipid host galls with similar morphologies, and generally attack larger galls. Our results suggest that parasitoid wasps such as Sycophila have adaptations allowing them to exploit particular host trait combinations, while hosts with contrasting traits are resistant to attack. These findings support the tritrophic niche concept for the structuring of plant-herbivore-parasitoid communities.

Ultraconserved elements-based systematics reveals evolutionary patterns of host-plant family shifts and phytophagy within the predominantly parasitoid braconid wasp subfamily Doryctinae.

Phytophagy has promoted species diversification in many insect groups, including Hymenoptera, one of the most diverse animal orders on Earth. In the predominantly parasitoid family Braconidae, an association with insect-induced, plant galls in angiosperms have been reported in three subfamilies, but in particular in the Doryctinae, where it has been recorded to occur in species of ten genera. Allorhogas Gahan is the most species-rich of these genera, with its species having different phytophagous strategies. Here we conducted a comprehensive phylogenomic study for the doryctine gall-associated genera, with an emphasis on Allorhogas, using ultraconserved elements (UCEs). Based on this estimate of phylogeny we: (1) evaluated their taxonomic composition, (2) estimated the timing of origin of the gall-associated clade and divergence of its main subclades, and (3) performed ancestral state reconstruction analyses for life history traits related to their host-plant association. Our phylogenetic hypothesis confirmed Allorhogas as polyphyletic, with most of its members being nested in a main clade composed of various subclades, each comprising species with a particular host-plant family and herbivorous feeding habit. The origin of gall-association was estimated to have occurred during the late Oligocene to early Miocene, with a subsequent diversification of subclades during the middle to late Miocene and Pliocene. Overlap in divergence timing appears to occur between some taxa and their host-associated plant lineages. Evolution of the feeding strategies in the group shows "inquilinism-feeding" as the likely ancestral state, with gall-formation in different plant organs and seed predation having independently evolved on multiple occasions.

Genomically correlated trait combinations and antagonistic selection contributing to counterintuitive genetic patterns of adaptive diapause divergence in Rhagoletis flies.

Adaptation to novel environments can result in unanticipated genomic responses to selection. Here, we illustrate how multifarious, correlational selection helps explain a counterintuitive pattern of genetic divergence between the recently derived apple- and ancestral hawthorn-infesting host races of Rhagoletis pomonella (Diptera: Tephritidae). The apple host race terminates diapause and emerges as adults earlier in the season than the hawthorn host race, to coincide with the earlier fruiting phenology of their apple hosts. However, alleles at many loci associated with later emergence paradoxically occur at higher frequencies in sympatric populations of the apple compared to the hawthorn race. We present genomic evidence that historical selection over geographically varying environmental gradients across North America generated genetic correlations between two life history traits, diapause intensity and diapause termination, in the hawthorn host race. Moreover, the loci associated with these life history traits are concentrated in genomic regions in high linkage disequilibrium (LD). These genetic correlations are antagonistic to contemporary selection on local apple host race populations that favours increased initial diapause depth and earlier, not later, diapause termination. Thus, the paradox of apple flies appears due, in part, to pleiotropy or linkage of alleles associated with later adult emergence and increased initial diapause intensity, the latter trait strongly selected for by the earlier phenology of apples. Our results demonstrate how understanding of multivariate trait combinations and the correlative nature of selective forces acting on them can improve predictions concerning adaptive evolution and help explain seemingly counterintuitive patterns of genetic diversity in nature.

Context-Dependent Reproductive Isolation: Host Plant Variability Drives Fitness of Hybrid Herbivores.

AbstractThe role of divergent selection between alternative environments in promoting reproductive isolation (RI) between lineages is well recognized. However, most studies view each divergent environment as homogenous, thereby overlooking the potential role within-environment variation plays in RI between differentiating lineages. Here, we test the importance of microenvironmental variation in RI by using individual trees of two host plants, each harboring locally adapted populations of the cynipid wasp Belonocnema treatae. We compared the fitness surrogate (survival) of offspring from hybrid crosses with resident crosses across individual trees on each of two primary host plants, Quercus virginiana and Q. geminata. We found evidence of weak hybrid inviability between host-associated lineages of B. treatae despite strong genomic differentiation. However, averaging across environments masked great variation in hybrid fitness on individual trees, where hybrids performed worse than, equal to, or better than residents. Thus, considering the environmental context of hybridization is critical to improving the predictability of divergence under variable selection.

Fate of Environmental Proteins (eProteins) from Genetically Engineered Crops in Streams is Controlled by Water pH and Ecosystem Metabolism.

Environmental proteins (eProteins), such as Cry proteins associated with genetically engineered (GE) organisms, are present in ecosystems worldwide, but only rarely reach concentrations with detectable ecosystem-level impacts. Despite their ubiquity, the degradation and fate of Cry and other eProteins are mostly unknown. Here, we report the results of an experiment where we added Cry proteins leached from GE Bt maize to a suite of 19 recirculating experimental streams. We found that Cry exhibited a biphasic degradation with an initial phase of rapid and variable degradation within 1 h, followed by a slow and steady phase of degradation with traces of protein persisting after 48 h. The initial degradation was correlated with heterotrophic respiration and water column dissolved oxygen, confirming a previously documented association with stream metabolism. However, protein degradation persisted even with no biofilm and was faster at a more acidic pH, suggesting that water chemistry is also a critical factor in both degradation and subsequent detection. We suggest that Cry, as well as other eProteins, will have a rapid degradation caused by denaturation of proteins and pH changes, which confirms that the detection of Cry proteins in natural streams must be the result of steady and consistent leaching into the environment.

Host plant associations and geography interact to shape diversification in a specialist insect herbivore.

Disentangling the processes underlying geographic and environmental patterns of biodiversity challenges biologists as such patterns emerge from eco-evolutionary processes confounded by spatial autocorrelation among sample units. The herbivorous insect, Belonocnema treatae (Hymenoptera: Cynipidae), exhibits regional specialization on three plant species whose geographic distributions range from sympatry through allopatry across the southern United States. Using range-wide sampling spanning the geographic ranges of the three host plants and genotyping-by-sequencing of 1,217 individuals, we tested whether this insect herbivore exhibited host plant-associated genomic differentiation while controlling for spatial autocorrelation among the 58 sample sites. Population genomic structure based on 40,699 SNPs was evaluated using the hierarchical Bayesian model entropy to assign individuals to genetic clusters and estimate admixture proportions. To control for spatial autocorrelation, distance-based Moran's eigenvector mapping was used to construct regression variables summarizing spatial structure inherent among sample sites. Distance-based redundancy analysis (dbRDA) incorporating the spatial variables was then applied to partition host plant-associated differentiation (HAD) from spatial autocorrelation. By combining entropy and dbRDA to analyse SNP data, we unveiled a complex mosaic of highly structured differentiation within and among gall-former populations finding evidence that geography, HAD and spatial autocorrelation all play significant roles in explaining patterns of genomic differentiation in B. treatae. While dbRDA confirmed host association as a significant predictor of patterns of genomic variation, spatial autocorrelation among sites explained the largest proportion of variation. Our results demonstrate the value of combining dbRDA with hierarchical structural analyses to partition spatial/environmental patterns of genomic variation.

Temporal isolation between sympatric host plants cascades across multiple trophic levels of host-associated insects.

Phenological differences between host plants can promote temporal isolation among host-associated populations of insects with life cycles tightly coupled to plant phenology. Divergence in the timing of spring budbreak between two sympatric sister oak species has been shown to promote temporal isolation between host plants and their host-associated populations of a cynipid gall wasp. Here, we examined the generality of this mechanism by testing the hypothesis of cascading temporal isolation for five additional gall-formers and three natural enemy species associated with these same oak species. The timing of adult emergence from galls differed significantly between host-associated populations for all nine species and parallels the direction of the phenological differences between host plants. Differences in emergence timing can reduce gene flow between host-associated populations by diminishing mating opportunities and/or reducing the fitness of immigrants due to differences in the availability of ephemeral resources. Our study suggests that cascading temporal isolation could be a powerful 'biodiversity generator' across multiple trophic levels in tightly coupled plant-insect systems.

Human-mediated disturbance in multitrophic interactions results in outbreak levels of North America's most venomous caterpillar.

Anthropogenic environmental change is predicted to disrupt multitrophic interactions, which may have drastic consequences for population-level processes. Here, we investigate how a large-scale human-mediated disturbance affects the abundance of North America's most venomous caterpillar species, Megalopyge opercularis. Specifically, we used a natural experiment where netting was deployed to cover the entire canopies of a subset of mature southern live oak trees (Quercus virginiana) to exclude urban pest birds (grackles and pigeons), throughout an 8.1 km2 area encompassing a medical centre in Houston, Texas. We used this experimental exclusion to test the following hypothesis: release from avian predators increases caterpillar abundance to outbreak levels, which increases the risk to human health. Results from a multi-year survey show that caterpillar abundance increased, on average, more than 7300% on netted versus non-netted trees. Thus, increases in caterpillar abundance due to anthropogenic enemy release increase human exposure to this venomous pest, and should be considered a health threat in the area. This study emphasizes the unforeseen consequences of ecological disturbance for species interactions and highlights the importance of considering ecology in urban planning.

A keeper of many crypts: a behaviour-manipulating parasite attacks a taxonomically diverse array of oak gall wasp species.

Parasites of animals and plants can encounter trade-offs between their specificity to any single host and their fitness on alternative hosts. For parasites that manipulate their host's behaviour, the added complexity of that manipulation may further limit the parasite's host range. However, this is rarely tested. The recently described crypt-keeper wasp, Euderus set, changes the behaviour of the gall wasp Bassettia pallida such that B. pallida chews a significantly smaller exit hole in the side of its larval chamber and 'plugs' that hole with its head before dying. Euderus set benefits from this head plug, as it facilitates the escape of the parasitoid from the crypt after it completes development. Here, we find direct and indirect evidence that E. set attacks and manipulates the behaviour of at least six additional gall wasp species, and that these hosts are taxonomically diverse. Interestingly, each of E. set's hosts has converged upon similarities in their extended phenotypes: the galls they induce on oaks share characters that may make them vulnerable to attack by E. set. The specialization required to behaviourally manipulate hosts may be less important in determining the range of hosts in this parasitoid system than other dimensions of the host-parasitoid interaction, like the host's physical defences.

Rapid quantitative protein detection by light transmission spectroscopy.

Rapid, sensitive, and quantitative protein detection is critical for many applications in medicine, environmental monitoring, and the food industry. Advancements in detection of proteins include the use of antigen-antibody binding; however, many current methods are time-consuming and have limiting factors such as low sensitivity and the inability to provide absolute values. We present a new high-throughput method for protein detection using light transmission spectroscopy (LTS), which can quantify and size nanoparticles in fluid suspension. LTS can quantify proteins directly and target specific proteins through antigen-antibody binding. This work shows that LTS can distinguish between and quantify bovine serum albumin, its antibody, and the BSA + Ab complex and determine BSA protein concentrations down to 5 µg/mL. We use both Mie and discrete dipole approximation models to provide geometric insight into the binding process.

Diversity and distribution of Wolbachia in relation to geography, host plant affiliation and life cycle of a heterogonic gall wasp.

BACKGROUND: The maternally inherited endosymbiont Wolbachia is widespread in arthropods and nematodes and can play an important role in the ecology and evolution of its host through reproductive manipulation. Here, we survey Wolbachia in Belonocnema treatae, a widely distributed North American cynipid gall forming wasp that exhibits regional host specialization on three species of oaks and alternation of sexually and asexually reproducing generations. We investigated whether patterns of Wolbachia infection and diversity in B. treatae are associated with the insect's geographic distribution, host plant association, life cycle, and mitochondrial evolutionary history. RESULTS: Screening of 463 individuals from 23 populations including sexual and asexual generations from all three host plants across the southern U.S. showed an average infection rate of 56% with three common Wolbachia strains: wTre1-3 and an additional rare variant wTre4. Phylogenetic analysis based on wsp showed that these strains are unrelated and likely independently inherited. We found no difference in Wolbachia infection frequency among host plant associated populations or between the asexual and sexual generations, or between males and females of the sexual generation. Partially incomplete Wolbachia transmission rates might explain the occurrence of uninfected individuals. A parallel analysis of the mitochondrial cytochrome oxidase I gene in B. treatae showed high mtDNA haplotype diversity in both infected and uninfected populations suggesting an ancestral infection by Wolbachia as well as a clear split between eastern and western B. treatae mtDNA clades with a sequence divergence of > 6%. The strain wTre1 was present almost exclusively in the western clade while wTre2 and wTre3 occur almost exclusively in eastern populations. In contrast, the same strains co-occur as double-infections in Georgia and triple-infections in two populations in central Florida. CONCLUSIONS: The diversity of Wolbachia across geographically and genetically distinct populations of B. treatae and the co-occurrence of the same strains within three populations highlights the complex infection dynamics in this system. Moreover, the association of distinct Wolbachia strains with mitochondrial haplotypes of its host in populations infected by different Wolbachia strains suggests a potential role of the endosymbiont in reproductive isolation in B. treatae.

Water Flow and Biofilm Cover Influence Environmental DNA Detection in Recirculating Streams.

The increasing use of environmental DNA (eDNA) for determination of species presence in aquatic ecosystems is an invaluable technique for both ecology as a field and for the management of aquatic ecosystems. We examined the degradation dynamics of fish eDNA using an experimental array of recirculating streams, also using a "nested" primer assay to estimate degradation among eDNA fragment sizes. We introduced eDNA into streams with a range of water velocities (0.1-0.8 m s-1) and substrate biofilm coverage (0-100%) and monitored eDNA concentrations over time (∼10 d) to assess how biophysical conditions influence eDNA persistence. We found that the presence of biofilm significantly increased initial decay rates relative to previous studies conducted in nonflowing microcosms, suggesting important differences in detection and persistence in lentic vs lotic systems. Lastly, by using a nested primer assay that targeted different size eDNA fragments, we found that fragment size altered both the estimated rate constant coefficients, as well as eDNA detectability over time. Larger fragments (>600 bp) were quickly degraded, while shorter fragments (<100 bp) remained detectable for the entirety of the experiment. When using eDNA as a stream monitoring tool, understanding environmental factors controlling eDNA degradation will be critical for optimizing eDNA sampling strategies.

Sequential divergence and the multiplicative origin of community diversity.

Phenotypic and genetic variation in one species can influence the composition of interacting organisms within communities and across ecosystems. As a result, the divergence of one species may not be an isolated process, as the origin of one taxon could create new niche opportunities for other species to exploit, leading to the genesis of many new taxa in a process termed "sequential divergence." Here, we test for such a multiplicative effect of sequential divergence in a community of host-specific parasitoid wasps, Diachasma alloeum, Utetes canaliculatus, and Diachasmimorpha mellea (Hymenoptera: Braconidae), that attack Rhagoletis pomonella fruit flies (Diptera: Tephritidae). Flies in the R. pomonella species complex radiated by sympatrically shifting and ecologically adapting to new host plants, the most recent example being the apple-infesting host race of R. pomonella formed via a host plant shift from hawthorn-infesting flies within the last 160 y. Using population genetics, field-based behavioral observations, host fruit odor discrimination assays, and analyses of life history timing, we show that the same host-related ecological selection pressures that differentially adapt and reproductively isolate Rhagoletis to their respective host plants (host-associated differences in the timing of adult eclosion, host fruit odor preference and avoidance behaviors, and mating site fidelity) cascade through the ecosystem and induce host-associated genetic divergence for each of the three members of the parasitoid community. Thus, divergent selection at lower trophic levels can potentially multiplicatively and rapidly amplify biodiversity at higher levels on an ecological time scale, which may sequentially contribute to the rich diversity of life.