Phylogenomics of Ichneumonoidea (Hymenoptera) and implications for evolution of mode of parasitism and viral endogenization.

Ichneumonoidea is one of the most diverse lineages of animals on the planet with >48,000 described species and many more undescribed. Parasitoid wasps of this superfamily are mostly beneficial insects that attack and kill other arthropods and are important for understanding diversification and the evolution of life history strategies related to parasitoidism. Further, some lineages of parasitoids within Ichneumonoidea have acquired endogenous virus elements (EVEs) that are permanently a part of the wasp's genome and benefit the wasp through host immune disruption and behavioral control. Unfortunately, understanding the evolution of viral acquisition, parasitism strategies, diversification, and host immune disruption mechanisms, is deeply limited by the lack of a robust phylogenetic framework for Ichneumonoidea. Here we design probes targeting 541 genes across 91 taxa to test phylogenetic relationships, the evolution of parasitoid strategies, and the utility of probes to capture polydnavirus genes across a diverse array of taxa. Phylogenetic relationships among Ichneumonoidea were largely well resolved with most higher-level relationships maximally supported. We noted codon use biases between the outgroups, Braconidae, and Ichneumonidae and within Pimplinae, which were largely solved through analyses of amino acids rather than nucleotide data. These biases may impact phylogenetic reconstruction and caution for outgroup selection is recommended. Ancestral state reconstructions were variable for Braconidae across analyses, but consistent for reconstruction of idiobiosis/koinobiosis in Ichneumonidae. The data suggest many transitions between parasitoid life history traits across the whole superfamily. The two subfamilies within Ichneumonidae that have polydnaviruses are supported as distantly related, providing strong evidence for two independent acquisitions of ichnoviruses. Polydnavirus capture using our designed probes was only partially successful and suggests that more targeted approaches would be needed for this strategy to be effective for surveying taxa for these viral genes. In total, these data provide a robust framework for the evolution of Ichneumonoidea.

Systematics, Biology, and Evolution of Microgastrine Parasitoid Wasps.

The braconid parasitoid wasp subfamily Microgastrinae is perhaps the most species-rich subfamily of animals on Earth. Despite their small size, they are familiar to agriculturalists and field ecologists alike as one of the principal groups of natural enemies of caterpillars feeding on plants. Their abundance and nearly ubiquitous terrestrial distribution, their intricate interactions with host insects, and their historical association with mutualistic polydnaviruses have all contributed to Microgastrinae becoming a key group of organisms for studying parasitism, parasitoid genomics, and mating biology. However, these rich sources of data have not yet led to a robust genus-level classification of the group, and some taxonomic confusion persists as a result. We present the current status of understanding of the general biology, taxonomic history, diversity, geographical patterns, host relationships, and phylogeny of Microgastrinae as a stimulus and foundation for further study. Current progress in elucidating the biology and taxonomy of this important group is rapid and promises a revolution in the classification of these wasps in the near future.

Fragmentary Gene Sequences Negatively Impact Gene Tree and Species Tree Reconstruction.

Species tree reconstruction from genome-wide data is increasingly being attempted, in most cases using a two-step approach of first estimating individual gene trees and then summarizing them to obtain a species tree. The accuracy of this approach, which promises to account for gene tree discordance, depends on the quality of the inferred gene trees. At the same time, phylogenomic and phylotranscriptomic analyses typically use involved bioinformatics pipelines for data preparation. Errors and shortcomings resulting from these preprocessing steps may impact the species tree analyses at the other end of the pipeline. In this article, we first show that the presence of fragmentary data for some species in a gene alignment, as often seen on real data, can result in substantial deterioration of gene trees, and as a result, the species tree. We then investigate a simple filtering strategy where individual fragmentary sequences are removed from individual genes but the rest of the gene is retained. Both in simulations and by reanalyzing a large insect phylotranscriptomic data set, we show the effectiveness of this simple filtering strategy.

DiscoVista: Interpretable visualizations of gene tree discordance.

Phylogenomics has ushered in an age of discordance. Analyses often reveal abundant discordances among phylogenies of different parts of genomes, as well as incongruences between species trees obtained using different methods or data partitions. Researchers are often left trying to make sense of such incongruences. Interpretive ways of measuring and visualizing discordance are needed, both among alternative species trees and gene trees, especially for specific focal branches of a tree. Here, we introduce DiscoVista, a publicly available tool that creates a suite of simple but interpretable visualizations. DiscoVista helps quantify the amount of discordance and some of its potential causes.

Widespread genome reorganization of an obligate virus mutualist.

The family Polydnaviridae is of interest because it provides the best example of viruses that have evolved a mutualistic association with their animal hosts. Polydnaviruses in the genus Bracovirus are strictly associated with parasitoid wasps in the family Braconidae, and evolved ∼100 million years ago from a nudivirus. Each wasp species relies on its associated bracovirus to parasitize hosts, while each bracovirus relies on its wasp for vertical transmission. Prior studies establish that bracovirus genomes consist of proviral segments and nudivirus-like replication genes, but how these components are organized in the genomes of wasps is unknown. Here, we sequenced the genome of the wasp Microplitis demolitor to characterize the proviral genome of M. demolitor bracovirus (MdBV). Unlike nudiviruses, bracoviruses produce virions that package multiple circular, double-stranded DNAs. DNA segments packaged into MdBV virions resided in eight dispersed loci in the M. demolitor genome. Each proviral segment was bounded by homologous motifs that guide processing to form mature viral DNAs. Rapid evolution of proviral segments obscured homology between other bracovirus-carrying wasps and MdBV. However, some domains flanking MdBV proviral loci were shared with other species. All MdBV genes previously identified to encode proteins required for replication were identified. Some of these genes resided in a multigene cluster but others, including subunits of the RNA polymerase that transcribes structural genes and integrases that process proviral segments, were widely dispersed in the M. demolitor genome. Overall, our results indicate that genome dispersal is a key feature in the evolution of bracoviruses into mutualists.

Parasitism rate, parasitoid community composition and host specificity on exposed and semi-concealed caterpillars from a tropical rainforest.

The processes maintaining the enormous diversity of herbivore-parasitoid food webs depend on parasitism rate and parasitoid host specificity. The two parameters have to be evaluated in concert to make conclusions about the importance of parasitoids as natural enemies and guide biological control. We document parasitism rate and host specificity in a highly diverse caterpillar-parasitoid food web encompassing 266 species of lepidopteran hosts and 172 species of hymenopteran or dipteran parasitoids from a lowland tropical forest in Papua New Guinea. We found that semi-concealed hosts (leaf rollers and leaf tiers) represented 84% of all caterpillars, suffered a higher parasitism rate than exposed caterpillars (12 vs. 5%) and their parasitoids were also more host specific. Semi-concealed hosts may therefore be generally more amenable to biological control by parasitoids than exposed ones. Parasitoid host specificity was highest in Braconidae, lower in Diptera: Tachinidae, and, unexpectedly, the lowest in Ichneumonidae. This result challenges the long-standing view of low host specificity in caterpillar-attacking Tachinidae and suggests higher suitability of Braconidae and lower suitability of Ichneumonidae for biological control of caterpillars. Semi-concealed hosts and their parasitoids are the largest, yet understudied component of caterpillar-parasitoid food webs. However, they still remain much closer in parasitism patterns to exposed hosts than to what literature reports on fully concealed leaf miners. Specifically, semi-concealed hosts keep an equally low share of idiobionts (2%) as exposed caterpillars.

A revision of the parasitoid wasp genus Alphomelon Mason with the description of 30 new species (Hymenoptera, Braconidae).

The parasitoid wasp genus Alphomelon Mason, 1981 is revised, based on a combination of basic morphology (dichotomous key and brief diagnostic descriptions), DNA barcoding, biology (host data and wasp cocoons), and distribution data. A total of 49 species is considered; the genus is almost entirely Neotropical (48 species recorded from that region), but three species reach the Nearctic, with one of them extending as far north as 45° N in Canada. Alphomelon parasitizes exclusively Hesperiinae caterpillars (Lepidoptera: Hesperiidae), mostly feeding on monocots in the families Arecaceae, Bromeliaceae, Cannaceae, Commelinaceae, Heliconiaceae, and Poaceae. Most wasp species parasitize either on one or very few (2-4) host species, usually within one or two hesperiine genera; but some species can parasitize several hosts from up to nine different hesperiine genera. Among species with available data for their cocoons, roughly half weave solitary cocoons (16) and half are gregarious (17); cocoons tend to be surrounded by a rather distinctive, coarse silk (especially in solitary species, but also distinguishable in some gregarious species). Neither morphology nor DNA barcoding alone was sufficient on its own to delimit all species properly; by integrating all available evidence (even if incomplete, as available data for every species is different) a foundation is provided for future studies incorporating more specimens, especially from South America. The following 30 new species are described: cruzi, itatiaiensis, and palomae, authored by Shimbori & Fernandez-Triana; and adrianguadamuzi, amazonas, andydeansi, calixtomoragai, carolinacanoae, christerhanssoni, diniamartinezae, duvalierbricenoi, eldaarayae, eliethcantillanoae, gloriasihezarae, guillermopereirai, hazelcambroneroae, josecortesi, keineraragoni, luciarosae, manuelriosi, mikesharkeyi, osvaldoespinozai, paramelanoscelis, paranigriceps, petronariosae, ricardocaleroi, rigoi, rostermoragai, sergioriosi, and yanayacu, authored by Fernandez-Triana & Shimbori.

Many evolutionary roads led to virus domestication in ichneumonoid parasitoid wasps.

The investigation of endogenous viral elements (EVEs) has historically focused on only a few lineages of parasitoid wasps, with negative results consistently underreported. Recent studies show that multiple viral lineages were integrated in at least seven instances in Ichneumonoidea and may be much more widespread than previously thought. Increasingly affordable genomic and bioinformatic approaches have made it feasible to search for viral sequences within wasp genomes, opening an extremely promising research avenue. Advances in wasp phylogenetics have shed light on the evolutionary history of EVE integration, although many questions remain. Phylogenetic proximity can be used as a guide to facilitate targeted screening, to estimate the number and age of integration events and to identify taxa involved in major host switches.

Extreme diversity of tropical parasitoid wasps exposed by iterative integration of natural history, DNA barcoding, morphology, and collections.

We DNA barcoded 2,597 parasitoid wasps belonging to 6 microgastrine braconid genera reared from parapatric tropical dry forest, cloud forest, and rain forest in Area de Conservación Guanacaste (ACG) in northwestern Costa Rica and combined these data with records of caterpillar hosts and morphological analyses. We asked whether barcoding and morphology discover the same provisional species and whether the biological entities revealed by our analysis are congruent with wasp host specificity. Morphological analysis revealed 171 provisional species, but barcoding exposed an additional 142 provisional species; 95% of the total is likely to be undescribed. These 313 provisional species are extraordinarily host specific; more than 90% attack only 1 or 2 species of caterpillars out of more than 3,500 species sampled. The most extreme case of overlooked diversity is the morphospecies Apanteles leucostigmus. This minute black wasp with a distinctive white wing stigma was thought to parasitize 32 species of ACG hesperiid caterpillars, but barcoding revealed 36 provisional species, each attacking one or a very few closely related species of caterpillars. When host records and/or within-ACG distributions suggested that DNA barcoding had missed a species-pair, or when provisional species were separated only by slight differences in their barcodes, we examined nuclear sequences to test hypotheses of presumptive species boundaries and to further probe host specificity. Our iterative process of combining morphological analysis, ecology, and DNA barcoding and reiteratively using specimens maintained in permanent collections has resulted in a much more fine-scaled understanding of parasitoid diversity and host specificity than any one of these elements could have produced on its own.

A new genus of Cedriini (Hymenoptera: Braconidae, Hormiinae) from the Brazilian cerrado.

A new genus of the braconid wasp tribe Cedriini, Sagarana n. gen., (Hymenoptera, Braconidae) is described and illustrated from the Brazilian cerrado based on a newly discovered species, Sagarana cerradensis n. sp. The genus is clearly distinct from the other genera of the Cedriini, with unusual fringed depressions on the second metasomal sternite that are of unknown function. The confusing taxonomy of this tribe is briefly discussed.

Cotesiacassina sp. nov. from southwestern Colombia: a new gregarious microgastrine wasp (Hymenoptera, Braconidae) reared from the pest species Opsiphanescassina Felder & Felder (Lepidoptera, Nymphalidae) feeding on Elaeis oil palm trees (Arecaceae).

A new species of microgastrine wasp, Cotesiacassina Salgado-Neto, Vásquez & Whitfield, sp. nov., is described from southwestern Colombia in Tumaco, Nariño. This species is a koinobiont gregarious larval endoparasitoid, and spins a common mass of cocoons underneath the host caterpillars of Opsiphanescassina (Felder & Felder) (Lepidoptera, Nymphalidae), feeding on oil palm trees (interspecific hybrid Elaeisoleifera × E.guineensis) (Arecaceae). While superficially similar, both morphologically and biologically, to C.invirae Salgado-Neto & Whitfield from southern Brazil, the two species are distinct based on DNA barcodes, host species, geographical range and morphological characters.

Chromosomal scale assembly of parasitic wasp genome reveals symbiotic virus colonization.

Endogenous viruses form an important proportion of eukaryote genomes and a source of novel functions. How large DNA viruses integrated into a genome evolve when they confer a benefit to their host, however, remains unknown. Bracoviruses are essential for the parasitism success of parasitoid wasps, into whose genomes they integrated ~103 million years ago. Here we show, from the assembly of a parasitoid wasp genome at a chromosomal scale, that bracovirus genes colonized all ten chromosomes of Cotesia congregata. Most form clusters of genes involved in particle production or parasitism success. Genomic comparison with another wasp, Microplitis demolitor, revealed that these clusters were already established ~53 mya and thus belong to remarkably stable genomic structures, the architectures of which are evolutionary constrained. Transcriptomic analyses highlight temporal synchronization of viral gene expression without resulting in immune gene induction, suggesting that no conflicts remain between ancient symbiotic partners when benefits to them converge.

Orientocardiochiles, a new genus of Cardiochilinae (Hymenoptera, Braconidae), with descriptions of two new species from Malaysia and Vietnam.

For the first time in 21 years, a new genus of cardiochiline braconid wasp, Orientocardiochiles Kang & Long, gen. nov. (type species Orientocardiochiles joeburrowi Kang, sp. nov.), is discovered and described. This genus represents the ninth genus in the Oriental region. Two new species (O. joeburrowi Kang, sp. nov. and O. nigrofasciatus Long, sp. nov.) are described and illustrated, and a key to species of the genus, with detailed images, is added. Diagnostic characters of the new genus are analyzed and compared with several other cardiochiline genera to allow the genus to key out properly using an existing generic treatment. The scientific names validated by this paper and morphological data obtained from this project will be utilized and tested in the upcoming genus-level revision of the subfamily based on combined morphological and molecular data.

Experimental support for multiple-locus complementary sex determination in the parasitoid Cotesia vestalis.

Despite its fundamental role in development, sex determination is highly diverse among animals. Approximately 20% of all animals are haplodiploid, with haploid males and diploid females. Haplodiploid species exhibit diverse but poorly understood mechanisms of sex determination. Some hymenopteran insect species exhibit single-locus complementary sex determination (sl-CSD), where heterozygosity at a polymorphic sex locus initiates female development. Diploid males are homozygous at the sex locus and represent a genetic load because they are inviable or sterile. Inbreeding depression associated with CSD is therefore expected to select for other modes of sex determination resulting in fewer or no diploid males. Here, we investigate an alternative, heretofore hypothetical, mode of sex determination: multiple-locus CSD (ml-CSD). Under ml-CSD, diploid males are predicted to develop only from zygotes that are homozygous at all sex loci. We show that inbreeding for eight generations in the parasitoid wasp Cotesia vestalis leads to increasing proportions of diploid males, a pattern that is consistent with ml-CSD but not sl-CSD. The proportion of diploid males (0.27 +/- 0.036) produced in the first generation of inbreeding (mother-son cross) suggests that two loci are likely involved. We also modeled diploid male production under CSD with three linked loci. Our data visually resemble CSD with linked loci because diploid male production in the second generation was lower than that in the first. To our knowledge, our data provide the first experimental support for ml-CSD.

Reared microgastrine wasps (Hymenoptera: Braconidae) from Yanayacu Biological Station and environs (Napo Province, Ecuador): diversity and host specialization.

The microgastrine braconid wasps recovered up through 2007 by the NSF-sponsored rearing project "Caterpillars and Parasitoids of the Eastern Andes in Ecuador" are summarized in terms of their host specialization and faunistic uniqueness. Two hundred fifty eight rearings of caterpillars resulted in records of Microgastrinae, distributed among 14 genera (Apanteles Förster, Choeras Mason, Cotesia Cameron, Diolcogaster Ashmead, Distatrix Mason, Dolichogenidea Viereck, Exix Mason, Glyptapanteles Ashmead, Hypomicrogaster Ashmead, Papanteles Mason, Parapanteles Ashmead, Protapanteles Ashmead, Sathon Mason and Venanus Mason). Eleven records of hyperparasitoids of Microgastrinae are also summarized; Mesochorus Gravenhorst (Ichneumonidae) and Perilampidae are both recorded. The results are compared to those recovered by surveys in other parts of the world, especially by the Janzen-Hallwachs survey of the Area de Conservación Guanacaste (ACG) in Costa Rica. An annotated list of microgastrine genera not yet recorded at Yanayacu, but which we expect to eventually find there based on extrapolation from their known geographic distributions, is provided.

A key to new world distatrix mason (Hymenoptera: Braconidae), with descriptions of six new reared neotropical species.

Six new species of the genus Distatrix Mason from Central and South America, D. loretta, D. xanadon, D. vigilis, D. pitillaensis, D. pandora Grinter, n. sp., and D. antirrheae Whitfield & Grinter, n. sp., are described from large-scale caterpillar inventory endeavors, mostly from the larvae of geometrid moths. Biological information and diagnostic features that distinguish these species from other previously described Distatrix, especially those from the Neotropical region, are provided; and the first key to New World species is presented. The new discoveries expand our knowledge of the World's widespread Distatrix fauna by about a third, suggesting that similar survey efforts in other poorly sampled regions will reveal numerous additional undescribed species.

A species-level taxonomic review and host associations of Glyptapanteles (Hymenoptera, Braconidae, Microgastrinae) with an emphasis on 136 new reared species from Costa Rica and Ecuador.

The descriptive taxonomic study reported here is focused on Glyptapanteles, a species-rich genus of hymenopteran parasitoid wasps. The species were found within the framework of two independent long-term Neotropical caterpillar rearing projects: northwestern Costa Rica (Área de Conservación Guanacaste, ACG) and eastern Andes, Ecuador (centered on Yanayacu Biological Station, YBS). One hundred thirty-six new species of Glyptapanteles Ashmead are described and all of them are authored by Arias-Penna. None of them was recorded in both countries; thus, 78 are from Costa Rica and the remaining 58 from Ecuador. Before this revision, the number of Neotropical described Glyptapanteles did not reach double digits. Reasonable boundaries among species were generated by integrating three datasets: Cytochrome Oxidase I (COI) gene sequencing data, natural history (host records), and external morphological characters. Each species description is accompanied by images and known geographical distribution. Characteristics such as shape, ornamentation, and location of spun Glyptapanteles cocoons were imaged as well. Host-parasitoid associations and food plants are also here published for the first time. A total of 88 species within 84 genera in 15 Lepidoptera families was encountered as hosts in the field. With respect to food plants, these wild-caught parasitized caterpillars were reared on leaves of 147 species within 118 genera in 60 families. The majority of Glyptapanteles species appeared to be relatively specialized on one family of Lepidoptera or even on some much lower level of taxonomic refinement. Those herbivores in turn are highly food-plant specialized, and once caterpillars were collected, early instars (1-3) yielded more parasitoids than later instars. Glyptapanteles jimmilleri Arias-Penna, sp. nov. is the first egg-larval parasitoid recorded within the genus, though there may be many more since such natural history requires a more focused collection of eggs. The rate of hyperparasitoidism within the genus was approximately 4% and was represented by Mesochorus spp. (Ichneumonidae). A single case of multiparasitoidism was reported, Copidosoma floridanum Ashmead (Encyrtidae) and Glyptapanteles ilarisaaksjarvi Arias-Penna, sp. nov. both parasitoid species emerged from the caterpillar of Noctuidae: Condica cupienta (Cramer). Bodyguard behavior was observed in two Glyptapanteles species: G. howelldalyi Arias-Penna, sp. nov. and G. paulhansoni Arias-Penna, sp. nov. A dichotomous key for all the new species is provided. The numerous species described here, and an equal number already reared but not formally described, signal a far greater Glyptapanteles species richness in the Neotropics than suggested by the few described previously.

A new species of Cotesia Cameron (Hymenoptera, Braconidae, Microgastrinae) reared from the hickory horned devil, Citheronia regalis, and luna moth, Actias luna, in east Texas.

The braconid wasp parasitoid Cotesia nuellorum Whitfield, new species, is described from specimens reared from a caterpillar of the hickory horned devil, Citheronia regalis (F.), and from a caterpillar of the luna moth, Actias luna (L.), in eastern Texas. The species is diagnosed with respect to other species of Cotesia recorded from North American Saturniidae, and details of its biology are provided.

Whole Genome Sequence of the Parasitoid Wasp Microplitis demolitor That Harbors an Endogenous Virus Mutualist.

Microplitis demolitor (Hymenoptera: Braconidae) is a parasitoid used as a biological control agent to control larval-stage Lepidoptera and serves as a model for studying the function and evolution of symbiotic viruses in the genus Bracovirus Here we present the M. demolitor genome (assembly version 2.0), with a genome size of 241 Mb, and a N50 scaffold and contig size of 1.1 Mb and 14 Kb, respectively. Using RNA-Seq data and manual annotation of genes of viral origin, we produced a high-quality gene set that includes 18,586 eukaryotic and 171 virus-derived protein-coding genes. Bracoviruses are dsDNA viruses with unusual genome architecture, in which the viral genome is integrated into the wasp genome and is comprised of two distinct components: proviral segments that are amplified, circularized, and packaged into virions for export into the wasp's host via oviposition; and replication genes. This genome assembly revealed that at least two scaffolds contain both nudivirus-like genes and proviral segments, demonstrating that at least some of these components are near each other in the genome on a single chromosome. The updated assembly and annotation are available in several publicly accessible databases; including the National Center for Biotechnology Information and the Ag Data Commons. In addition, all raw sequence data available for M. demolitor have been consolidated and are available for visualization at the i5k Workspace. This whole genome assembly and annotation represents the only genome-scale, annotated assembly from the lineage of parasitoid wasps that has associations with bracoviruses (the 'microgastroid complex'), providing important baseline knowledge about the architecture of co-opted virus symbiont genomes.