Aphid populations are frequently infected with facultative endosymbionts.

The occurrence of facultative endosymbionts has been studied in many commercially important crop pest aphids, but their occurrence and effects in non-commercial aphid species in natural populations have received less attention. We screened 437 aphid samples belonging to 106 aphid species for the eight most common facultative aphid endosymbionts. We found one or more facultative endosymbionts in 53% (56 of 106) of the species investigated. This likely underestimates the situation in the field because facultative endosymbionts are often present in only some colonies of an aphid species. Oligophagous aphid species carried facultative endosymbionts significantly more often than monophagous species. We did not find a significant correlation between ant tending and facultative endosymbiont presence. In conclusion, we found that facultative endosymbionts are common among aphid populations. This study is, to our knowledge, the first of its kind in the Netherlands and provides a basis for future research in this field. For instance, it is still unknown in what way many of these endosymbionts affect their hosts, which is important for determining the importance of facultative endosymbionts to community dynamics.

Variation in sex determination mechanisms may constrain parthenogenesis-induction by endosymbionts in haplodiploid systems.

Endosymbionts are maternally transmitted, and therefore benefit from maximizing female offspring numbers. Parthenogenesis-induction (PI) is the most effective type of manipulation for transmission, but has solely been detected in haplodiploid species, whereas cytoplasmic incompatibility (CI) is detected frequently across the arthropod phylum, including haplodiploids. This puzzling observation led us to hypothesize that the molecular sex-determination mechanism of the haplodiploid host may be a constraining factor in the ability of endosymbionts to induce parthenogenesis. Recent insights indicate that PI-endosymbionts may be able to directly manipulate sex-determination genes to induce the necessary steps required for PI in haplodiploids. However, sex-determination cascades vary extensively, so PI-induction would require a specialized and host-dependent tool set. Contrastingly, CI-related genes target conserved cell-cycle mechanisms, are located on mobile elements, and spread easily. Finally, endosymbiont-manipulations may have a strong impact on the effectiveness of haplodiploid biocontrol agents, but can also be used to enhance their efficacy.

Do aphids in Dutch sweet pepper greenhouses carry heritable elements that protect them against biocontrol parasitoids?

Biological control (biocontrol) of crop pests is a sustainable alternative to the use of biodiversity and organismal health-harming chemical pesticides. Aphids can be biologically controlled with parasitoid wasps; however, variable results of parasitoid-based aphid biocontrol in greenhouses are reported. Aphids may display genetically encoded (endogenous) defences that increase aphid resistance against parasitoids as under high parasitoid pressure there will be selection for parasitoid-resistant aphids, potentially affecting the success of parasitoid-based aphid biocontrol in greenhouses. Additionally, aphids may carry secondary bacterial endosymbionts that protect them against parasitoids. We studied whether there is variation in either of these heritable elements in aphids in greenhouses of sweet pepper, an agro-economically important crop in the Netherlands that is prone to aphid pests and where pest management heavily relies on biocontrol. We sampled aphid populations in organic (biocontrol only) and conventional (biocontrol and pesticides) sweet pepper greenhouses in the Netherlands during the 2019 crop growth season. We assessed the aphid microbiome through both diagnostic PCR and 16S rRNA sequencing and did not detect any secondary endosymbionts in the two most encountered aphid species, Myzus persicae and Aulacorthum solani. We also compared multiple aphid lines collected from different greenhouses for variation in levels of endogenous-based resistance against the parasitoids commonly used as biocontrol agents. We found no differences in the levels of endogenous-based resistance between different aphid lines. This study does not support the hypothesis that protective endosymbionts or the presence of endogenous resistant aphid lines affects the success of parasitoid-based biocontrol of aphids in Dutch greenhouses. Future investigations will need to address what is causing the variable successes of aphid biocontrol and what (biological and management-related) lessons can be learned for aphid control in other crops, and biocontrol in general.

A single QTL with large effect is associated with female functional virginity in an asexual parasitoid wasp.

During the transition from sexual to asexual reproduction, a suite of reproduction-related sexual traits become superfluous, and may be selected against if costly. Female functional virginity refers to asexual females resisting to mate or not fertilizing eggs after mating. These traits appear to be among the first that evolve during transitions from sexual to asexual reproduction. The genetic basis of female functional virginity remains elusive. Previously, we reported that female functional virginity segregates as expected for a single recessive locus in the asexual parasitoid wasp Asobara japonica. Here, we investigate the genetic basis of this trait by quantitative trait loci (QTL) mapping and candidate gene analyses. Consistent with the segregation of phenotypes, we found a single QTL of large effect, spanning over 4.23 Mb and comprising at least 131 protein-coding genes, of which 15 featured sex-biased expression in the related sexual species Asobara tabida. Two of the 15 sex-biased genes were previously identified to differ between related sexual and asexual population/species: CD151 antigen and nuclear pore complex protein Nup50. A third gene, hormone receptor 4, is involved in steroid hormone mediated mating behaviour. Overall, our results are consistent with a single locus, or a cluster of closely linked loci, underlying rapid evolution of female functional virginity in the transition to asexuality. Once this variant, causing rejection to mate, has swept through a population, the flanking region does not get smaller owing to lack of recombination in asexuals.

Bracon brevicornis Genome Showcases the Potential of Linked-Read Sequencing in Identifying a Putative Complementary Sex Determiner Gene.

Bracon brevicornis is an ectoparasitoid of a wide range of larval-stage Lepidopterans, including several pests of important crops, such as the corn borer, Ostrinia nubilalis. It is also one of the earliest documented cases of complementary sex determination in Hymenoptera. Here, we present the linked-read-based genome of B. brevicornis, complete with an ab initio-derived annotation and protein comparisons with fellow braconids, Fopius arisanus and Diachasma alloeum. We demonstrate the potential of linked-read assemblies in exploring regions of heterozygosity and search for structural and homology-derived evidence of the complementary sex determiner gene (csd).

Seasonal morphotypes of Drosophila suzukii differ in key life-history traits during and after a prolonged period of cold exposure.

Seasonal polyphenism in Drosophila suzukii manifests itself in two discrete adult morphotypes, the "winter morph" (WM) and the "summer morph" (SM). These morphotypes are known to differ in thermal stress tolerance, and they co-occur during parts of the year. In this study, we aimed to estimate morph-specific survival and fecundity in laboratory settings simulating field conditions. We specifically analyzed how WM and SM D. suzukii differed in mortality and reproduction during and after a period of cold exposure resembling winter and spring conditions in temperate climates. The median lifespan of D. suzukii varied around 5 months for the WM flies and around 7 months for the SM flies. WM flies showed higher survival during the cold-exposure period compared with SM flies, and especially SM males suffered high mortality under these conditions. In contrast, SM flies had lower mortality rates than WM flies under spring-like conditions. Intriguingly, reproductive status (virgin or mated) did not impact the fly survival, either during the cold exposure or during spring-like conditions. Even though the reproductive potential of WM flies was greatly reduced compared with SM flies, both WM and SM females that had mated before the cold exposure were able to continuously produce viable offspring for 5 months under spring-like conditions. Finally, the fertility of the overwintered WM males was almost zero, while the surviving SM males did not suffer reduced fertility. Combined with other studies on D. suzukii monitoring and overwintering behavior, these results suggest that overwintered flies of both morphotypes could live long enough to infest the first commercial crops of the season. The high mortality of SM males and the low fertility of WM males after prolonged cold exposure also highlight the necessity for females to store sperm over winter to be able to start reproducing early in the following spring.

Hybrid Genome Assembly and Evidence-Based Annotation of the Egg Parasitoid and Biological Control Agent Trichogramma brassicae.

Trichogramma brassicae (Bezdenko) are egg parasitoids that are used throughout the world as biological control agents and in laboratories as model species. Despite this ubiquity, few genetic resources exist beyond COI, ITS2, and RAPD markers. Aided by a Wolbachia infection, a wild-caught strain from Germany was reared for low heterozygosity and sequenced in a hybrid de novo strategy, after which several assembling strategies were evaluated. The best assembly, derived from a DBG2OLC-based pipeline, yielded a genome of 235 Mbp made up of 1,572 contigs with an N50 of 556,663 bp. Following a rigorous ab initio-, homology-, and evidence-based annotation, 16,905 genes were annotated and functionally described. As an example of the utility of the genome, a simple ortholog cluster analysis was performed with sister species T. pretiosum, revealing over 6000 shared clusters and under 400 clusters unique to each species. The genome and transcriptome presented here provides an essential resource for comparative genomics of the commercially relevant genus Trichogramma, but also for research into molecular evolution, ecology, and breeding of T. brassicae.

Next-generation biological control: the need for integrating genetics and genomics.

Biological control is widely successful at controlling pests, but effective biocontrol agents are now more difficult to import from countries of origin due to more restrictive international trade laws (the Nagoya Protocol). Coupled with increasing demand, the efficacy of existing and new biocontrol agents needs to be improved with genetic and genomic approaches. Although they have been underutilised in the past, application of genetic and genomic techniques is becoming more feasible from both technological and economic perspectives. We review current methods and provide a framework for using them. First, it is necessary to identify which biocontrol trait to select and in what direction. Next, the genes or markers linked to these traits need be determined, including how to implement this information into a selective breeding program. Choosing a trait can be assisted by modelling to account for the proper agro-ecological context, and by knowing which traits have sufficiently high heritability values. We provide guidelines for designing genomic strategies in biocontrol programs, which depend on the organism, budget, and desired objective. Genomic approaches start with genome sequencing and assembly. We provide a guide for deciding the most successful sequencing strategy for biocontrol agents. Gene discovery involves quantitative trait loci analyses, transcriptomic and proteomic studies, and gene editing. Improving biocontrol practices includes marker-assisted selection, genomic selection and microbiome manipulation of biocontrol agents, and monitoring for genetic variation during rearing and post-release. We conclude by identifying the most promising applications of genetic and genomic methods to improve biological control efficacy.

Genetic Variation in the Feeding Behavior of Isofemale Lines of Nesidiocoris tenuis.

Zoophytophagous predators provide biocontrol services in various major crops of modern horticulture due to the combination of its predatory capacity and the induction of plant defenses derived from its phytophagy. However, under certain conditions of prey scarcity, these natural enemies can inflict plant damage. Exploitation of genetic variation and subsequent selective breeding on foraging traits is a potential alternative to overcome this inconvenience. In this study, we quantified the genetic variation of phytophagy and zoophagy of Nesidiocoristenuis (Reuter) (Hemiptera: Miridae), a zoophytophagous predator widely used in tomato crops to suppress key pests. We compared nine isofemale lines on their capacity to produce necrotic rings and wilting on tomato plants as a proxy for phytophagy, as well as their efficacy to prey on Ephestia kuehniella Zeller (Lepidoptera: Pyralidae) eggs, as a proxy for zoophagy. Differences between isofemale lines in phytophagy and zoophagy indicated a genetic basis. Variation found in the zoophagy levels was larger than that in phytophagy levels. Our results showed that there is a genetic basis for the variation observed in the feeding behavior of isofemale lines of N.tenuis, highlighting the potential importance of selective breeding for such traits of biocontrol interest.

Genomics of sex allocation in the parasitoid wasp Nasonia vitripennis.

BACKGROUND: Whilst adaptive facultative sex allocation has been widely studied at the phenotypic level across a broad range of organisms, we still know remarkably little about its genetic architecture. Here, we explore the genome-wide basis of sex ratio variation in the parasitoid wasp Nasonia vitripennis, perhaps the best studied organism in terms of sex allocation, and well known for its response to local mate competition. RESULTS: We performed a genome-wide association study (GWAS) for single foundress sex ratios using iso-female lines derived from the recently developed outbred N. vitripennis laboratory strain HVRx. The iso-female lines capture a sample of the genetic variation in HVRx and we present them as the first iteration of the Nasonia vitripennis Genome Reference Panel (NVGRP 1.0). This panel provides an assessment of the standing genetic variation for sex ratio in the study population. Using the NVGRP, we discovered a cluster of 18 linked SNPs, encompassing 9 annotated loci associated with sex ratio variation. Furthermore, we found evidence that sex ratio has a shared genetic basis with clutch size on three different chromosomes. CONCLUSIONS: Our approach provides a thorough description of the quantitative genetic basis of sex ratio variation in Nasonia at the genome level and reveals a number of inter-related candidate loci underlying sex allocation regulation.

Quantitative genetics of wing morphology in the parasitoid wasp Nasonia vitripennis: hosts increase sibling similarity.

The central aim of evolutionary biology is to understand patterns of genetic variation between species and within populations. To quantify the genetic variation underlying intraspecific differences, estimating quantitative genetic parameters of traits is essential. In Pterygota, wing morphology is an important trait affecting flight ability. Moreover, gregarious parasitoids such as Nasonia vitripennis oviposit multiple eggs in the same host, and siblings thus share a common environment during their development. Here we estimate the genetic parameters of wing morphology in the outbred HVRx population of N. vitripennis, using a sire-dam model adapted to haplodiploids and disentangled additive genetic and host effects. The results show that the wing-size traits have low heritability (h2 ~ 0.1), while most wing-shape traits have roughly twice the heritability compared with wing-size traits. However, the estimates increased to h2 ~ 0.6 for wing-size traits when omitting the host effect from the statistical model, while no meaningful increases were observed for wing-shape traits. Overall, host effects contributed to ~50% of the variation in wing-size traits. This indicates that hosts have a large effect on wing-size traits, about fivefold more than genetics. Moreover, bivariate analyses were conducted to derive the genetic relationships among traits. Overall, we demonstrate the evolutionary potential for morphological traits in the N. vitripennis HVRx-outbred population, and report the host effects on wing morphology. Our findings can contribute to a further dissection of the genetics underlying wing morphology in N. vitripennis, with relevance for gregarious parasitoids and possibly other insects as well.

Validating the Demethylating Effects of 5-aza-2'-deoxycytidine in Insects Requires a Whole-Genome Approach.

We previously demonstrated that treatment with the demethylating agent 5-aza-2'-deoxycytidine (5-aza-dC) alters the offspring sex ratios produced by females of the parasitoid wasp Nasonia vitripennis. Females allocate offspring sex ratio in line with local mate competition theory, producing more or less female-biased sex ratios as the number of other females laying eggs on a patch varies, thereby reducing competition among their sons for mates. Interestingly, treatment with 5-aza-dC did not ablate the facultative sex allocation response. Instead, sex ratios became less female biased, a shift in the direction of the optimum sex ratio for paternally inherited alleles according to genomic conflict theory. This was the first (albeit indirect) experimental evidence for genomic conflict over sex allocation. In their comment, Ellers and colleagues assayed the effects of 5-aza-dC on DNA methylation in 10 Nasonia genes, finding no evidence of demethylation in these 10 genes, from which they conclude that 5-aza-dC has no demethylating capability in N. vitripennis. Quantifying the efficacy of 5-aza-dC in terms of demethylation is indeed crucial to in-depth interpretation of studies using 5-aza-dC to link phenotypes to epigenetic regulation. Here we outline the mode of action of 5-aza-dC and demonstrate that determining the efficacy of 5-aza-dC in insect systems requires a whole-genome approach.

Overwintered Drosophila suzukii Are the Main Source for Infestations of the First Fruit Crops of the Season.

The mechanisms allowing the widespread invasive pest Drosophila suzukii to survive from early spring until the availability of the first fruit crops are still unclear. Seasonal biology and population dynamics of D. suzukii were investigated in order to better understand the contribution of the early spring hosts to the infestation of the first fruit crops of the season. We identified hosts available to D. suzukii in early spring and assessed their suitability for the pest oviposition and reproductive success under field and laboratory conditions. The natural infestation rate of one of these hosts, Aucuba japonica, was assessed over springtime and the morphology of the flies that emerged from infested A. japonica fruits was characterized under field conditions. Then, these findings were correlated with long-term monitoring data on seasonal reproductive biology and morphology of the pest, using a cumulative degree-days (DD) analysis. Field sampling revealed that overwintered D. suzukii females were physiologically able to lay eggs at 87 DD which coincided with the detection of the first infested early spring hosts. The latter were continuously and increasingly infested by D. suzukii eggs in nature from early spring until the end of May, in particular Aucuba japonica. Individuals emerged from most of these hosts were characterized by a poor fitness and a rather low success of emergence. In the field, only few summer morphs emerged from naturally infested A. japonica fruits around the end of May-beginning of June. However, field monitoring in orchards revealed that D. suzukii individuals consisted solely of winter morphs until mid-June. These observations indicate that overwintered D. suzukii females are the predominant source for the infestations in the first available fruit crops of the season. We discuss these findings in the context of possible pest control strategies.

Differential gene expression is not required for facultative sex allocation: a transcriptome analysis of brain tissue in the parasitoid wasp Nasonia vitripennis.

Whole-transcriptome technologies have been widely used in behavioural genetics to identify genes associated with the performance of a behaviour and provide clues to its mechanistic basis. Here, we consider the genetic basis of sex allocation behaviour in the parasitoid wasp Nasonia vitripennis. Female Nasonia facultatively vary their offspring sex ratio in line with Hamilton's theory of local mate competition (LMC). A single female or 'foundress' laying eggs on a patch will lay just enough sons to fertilize her daughters. As the number of 'foundresses' laying eggs on a patch increases (and LMC declines), females produce increasingly male-biased sex ratios. Phenotypic studies have revealed the cues females use to estimate the level of LMC their sons will experience, but our understanding of the genetics underlying sex allocation is limited. Here, we exposed females to three foundress number conditions, i.e. three LMC conditions, and allowed them to oviposit. mRNA was extracted from only the heads of these females to target the brain tissue. The subsequent RNA-seq experiment confirmed that differential gene expression is not associated with the response to sex allocation cues and that we must instead turn to the underlying neuroscience to reveal the underpinnings of this impressive behavioural plasticity.

Development of microsatellite markers and estimation of inbreeding frequency in the parasitoid wasp Melittobia.

The parasitoid wasp Melittobia is an important insect for basic and applied biology. Specifically, their extremely female-biased sex ratios, which contrast to the prediction of pre-existing theories, are needed to be explained from the aspect of evolutionary biology. In this study, using next-generation sequencing, 20 microsatellite loci were developed and characterized for M. australica. The developed loci were used to analyze two populations, one from a mainland Japan and one from the Okinawa island region. Both populations showed a smaller observed heterozygosity than expected, and a high inbreeding coefficient. Deviations from Hardy-Weinberg equilibrium were recorded for the majority of the 20 loci, suggesting that both the populations are subdivided due to inbreeding as is expected by the reproductive biology in Melittobia. The sib-mating frequency in the two populations was calculated as 0.873 and 0.996, which is higher than the values estimated by the number of females observed in a host cocoon or cell, implying that closely related females lay eggs on a host. The microsatellite loci developed in this study can be used for more comprehensive analyses to identify genetic structure in natural populations for understanding their sex allocation behavior and for more generally evolutionary and population genetic studies.

DNA Methylation and Sex Allocation in the Parasitoid Wasp Nasonia vitripennis.

The role of epigenetics in the control and evolution of behavior is being increasingly recognized. Here we test whether DNA methylation influences patterns of adaptive sex allocation in the parasitoid wasp Nasonia vitripennis. Female N. vitripennis allocate offspring sex broadly in line with local mate competition (LMC) theory. However, recent theory has highlighted how genomic conflict may influence sex allocation under LMC, conflict that requires parent-of-origin information to be retained by alleles through some form of epigenetic signal. We manipulated whole-genome DNA methylation in N. vitripennis females using the hypomethylating agent 5-aza-2'-deoxycytidine. Across two replicated experiments, we show that disruption of DNA methylation does not ablate the facultative sex allocation response of females, as sex ratios still vary with cofoundress number as in the classical theory. However, sex ratios are generally shifted upward when DNA methylation is disrupted. Our data are consistent with predictions from genomic conflict over sex allocation theory and suggest that sex ratios may be closer to the optimum for maternally inherited alleles.

Oviposition but Not Sex Allocation Is Associated with Transcriptomic Changes in Females of the Parasitoid Wasp Nasonia vitripennis.

Linking the evolution of the phenotype to the underlying genotype is a key aim of evolutionary genetics and is crucial to our understanding of how natural selection shapes a trait. Here, we consider the genetic basis of sex allocation behavior in the parasitoid wasp Nasonia vitripennis using a transcriptomics approach. Females allocate offspring sex in line with the local mate competition (LMC) theory. Female-biased sex ratios are produced when one or a few females lay eggs on a patch. As the number of females contributing offspring to a patch increases, less female-biased sex ratios are favored. We contrasted the transcriptomic responses of females as they oviposit under conditions known to influence sex allocation: foundress number (a social cue) and the state of the host (parasitized or not). We found that when females encounter other females on a patch or assess host quality with their ovipositors, the resulting changes in sex allocation is not associated with significant changes in whole-body gene expression. We also found that the gene expression changes produced by females as they facultatively allocate sex in response to a host cue and a social cue are very closely correlated. We expanded the list of candidate genes associated with oviposition behavior in Nasonia, some of which may be involved in fundamental processes underlying the ability to facultatively allocate sex, including sperm storage and utilization.

Diploid males support a two-step mechanism of endosymbiont-induced thelytoky in a parasitoid wasp.

BACKGROUND: Haplodiploidy, where females develop from diploid, fertilized eggs and males from haploid, unfertilized eggs, is abundant in some insect lineages. Some species in these lineages reproduce by thelytoky that is caused by infection with endosymbionts: infected females lay haploid eggs that undergo diploidization and develop into females, while males are very rare or absent. It is generally assumed that in thelytokous wasps, endosymbionts merely diploidize the unfertilized eggs, which would then trigger female development. RESULTS: We found that females in the parasitoid wasp Asobara japonica infected with thelytoky-inducing Wolbachia produce 0.7-1.2% male offspring. Seven to 39% of these males are diploid, indicating that diploidization and female development can be uncoupled in A. japonica. Wolbachia titer in adults was correlated with their ploidy and sex: diploids carried much higher Wolbachia titers than haploids, and diploid females carried more Wolbachia than diploid males. Data from introgression lines indicated that the development of diploid individuals into males instead of females is not caused by malfunction-mutations in the host genome but that diploid males are most likely produced when the endosymbiont fails to activate the female sex determination pathway. Our data therefore support a two-step mechanism by which endosymbionts induce thelytoky in A. japonica: diploidization of the unfertilized egg is followed by feminization, whereby each step correlates with a threshold of endosymbiont titer during wasp development. CONCLUSIONS: Our new model of endosymbiont-induced thelytoky overthrows the view that certain sex determination mechanisms constrain the evolution of endosymbiont-induced thelytoky in hymenopteran insects. Endosymbionts can cause parthenogenesis through feminization, even in groups in which endosymbiont-diploidized eggs would develop into males following the hosts' sex determination mechanism. In addition, our model broadens our understanding of the mechanisms by which endosymbionts induce thelytoky to enhance their transmission to the next generation. Importantly, it also provides a novel window to study the yet-poorly known haplodiploid sex determination mechanisms in haplodiploid insects.

Population-level consequences of complementary sex determination in a solitary parasitoid.

BACKGROUND: Sex determination mechanisms are known to be evolutionarily labile but the factors driving transitions in sex determination mechanisms are poorly understood. All insects of the Hymenoptera are haplodiploid, with males normally developing from unfertilized haploid eggs. Under complementary sex determination (CSD), diploid males can be produced from fertilized eggs that are homozygous at the sex locus. Diploid males have near-zero fitness and thus represent a genetic load, which is especially severe under inbreeding. Here, we study mating structure and sex determination in the parasitoid Cotesia vestalis to investigate what may have driven the evolution of two complementary sex determination loci in this species. RESULTS: We genotyped Cotesia vestalis females collected from eight fields in four townships in Western Taiwan. 98 SNP markers were developed by aligning Illumina sequence reads of pooled DNA of eight different females against a de novo assembled genome of C. vestalis. This proved to be an efficient method for this non-model species and provides a resource for future use in related species. We found significant genetic differentiation within the sampled population but variation could not be attributed to sampling locations by AMOVA. Non-random mating was detected, with 8.1% of matings between siblings. Diploid males, detected by flow cytometry, were produced at a rate of 1.4% among diploids. CONCLUSIONS: We think that the low rate of diploid male production is best explained by a CSD system with two independent sex loci, supporting laboratory findings on the same species. Fitness costs of diploid males in C. vestalis are high because diploid males can mate with females and produce infertile triploid offspring. This severe fitness cost of diploid males combined with non-random mating may have resulted in evolution from single locus CSD to CSD with two independent loci.

Development of a Nasonia vitripennis outbred laboratory population for genetic analysis.

The parasitoid wasp genus Nasonia has rapidly become a genetic model system for developmental and evolutionary biology. The release of its genome sequence led to the development of high-resolution genomic tools, for both interspecific and intraspecific research, which has resulted in great advances in understanding Nasonia biology. To further advance the utility of Nasonia vitripennis as a genetic model system and to be able to fully exploit the advantages of its fully sequenced and annotated genome, we developed a genetically variable and well-characterized experimental population. In this study, we describe the establishment of the genetically diverse HVRx laboratory population from strains collected from the field in the Netherlands. We established a maintenance method that retains genetic variation over generations of culturing in the laboratory. As a characterization of its genetic composition, we provide data on the standing genetic variation and estimate the effective population size (N(e)) by microsatellite analysis. A genome-wide description of polymorphism is provided through pooled resequencing, which yielded 417,331 high-quality SNPs spanning all five Nasonia chromosomes. The HVRx population and its characterization are freely available as a community resource for investigators seeking to elucidate the genetic basis of complex trait variation using the Nasonia model system.