The Chalcidoidea bush of life: evolutionary history of a massive radiation of minute wasps.

Chalcidoidea are mostly parasitoid wasps that include as many as 500 000 estimated species. Capturing phylogenetic signal from such a massive radiation can be daunting. Chalcidoidea is an excellent example of a hyperdiverse group that has remained recalcitrant to phylogenetic resolution. We combined 1007 exons obtained with Anchored Hybrid Enrichment with 1048 ultra-conserved elements (UCEs) for 433 taxa including all extant families, >95% of all subfamilies, and 356 genera chosen to represent the vast diversity of the superfamily. Going back and forth between the molecular results and our collective knowledge of morphology and biology, we detected bias in the analyses that was driven by the saturation of nucleotide data. Our final results are based on a concatenated analysis of the least saturated exons and UCE datasets (2054 loci, 284 106 sites). Our analyses support an expected sister relationship with Mymarommatoidea. Seven previously recognized families were not monophyletic, so support for a new classification is discussed. Natural history in some cases would appear to be more informative than morphology, as illustrated by the elucidation of a clade of plant gall associates and a clade of taxa with planidial first-instar larvae. The phylogeny suggests a transition from smaller soft-bodied wasps to larger and more heavily sclerotized wasps, with egg parasitism as potentially ancestral for the entire superfamily. Deep divergences in Chalcidoidea coincide with an increase in insect families in the fossil record, and an early shift to phytophagy corresponds with the beginning of the "Angiosperm Terrestrial Revolution". Our dating analyses suggest a middle Jurassic origin of 174 Ma (167.3-180.5 Ma) and a crown age of 162.2 Ma (153.9-169.8 Ma) for Chalcidoidea. During the Cretaceous, Chalcidoidea may have undergone a rapid radiation in southern Gondwana with subsequent dispersals to the Northern Hemisphere. This scenario is discussed with regard to knowledge about the host taxa of chalcid wasps, their fossil record and Earth's palaeogeographic history.

Black gold rush - Evaluating the efficiency of the Fractionator in separating Hymenoptera families in a meadow ecosystem over a two week period.

In the face of insect decline, monitoring projects are launched widely to assess trends of insect populations. Collecting over long time periods results in large numbers of samples with thousands of individuals that are often just stored in freezers waiting to be further processed. As the time-consuming process of sorting and identifying specimens prevents taxonomists from working on mass samples, important information on species composition remains unknown and taxonomically neglected species remain undiagnosed. Size fractioning of bulk samples can improve sample handling and, thus, can help to overcome the taxonomic impediment. In this paper, we evaluate the efficiency of the fractionator in separating Hymenoptera families from a Malaise trap sample of a meadow ecosystem over a two week interval to make them available for further morphological identification. The fractionator system by Buffington and Gates (2008) was used to separate the sample in two size classes - a large (macro) and a small (micro) fraction - and Hymenoptera specimens were then counted and identified on family level. In total, 2,449 Hymenoptera specimens were found in the macro fraction and 3,016 in the micro fraction (5,465 specimens in total). For 24 out of 34 Hymenoptera families (71%), separation was significant. This study illustrates the efficiency of the fractionator and its potential to improve workflows dealing with specimen-rich Malaise trap samples.

Illuminating a Dark Taxon: Revision of European Trichacis Förster (Hymenoptera: Platygastridae) reveals a glut of synonyms.

The parasitoid wasp genus Trichacis Förster is revised for Europe. Examination of historical and modern collections combined with DNA barcoding revealed the presence of only a single species in Europe, Trichacis tristis (Nees, 1834), redescribed here. Fourteen new synonymies are proposed for T. tristis: T. abdominalis Thomson, 1859 syn.nov.; T. bidentiscutum Szabó, 1981 syn.nov.; T. didas (Walker, 1835) syn.nov.; T. fusciala Szabó, 1981 syn.nov.; T. hajduica Szabó, 1981 syn.nov.; T. illusor Kieffer, 1916 syn.nov.; T. nosferatus Buhl, 1997 syn.nov.; T. pisis (Walker, 1835) syn.nov.; T. persicus Asadi & Buhl, 2021 syn.nov.; T. pulchricornis Szelényi, 1953 syn.nov.; T. quadriclava Szabó, 1981 syn.nov.; T. remulus (Walker, 1835) syn.nov.; T. vitreus Buhl, 1997 syn.nov.; T. weiperti Buhl, 2019 syn.nov.. Four species are transferred to Amblyaspis Förster: A. afurcata (Szabó, 1977) comb. nov., A. hungarica (Szabó, 1977), comb. nov., A. pannonica (Szabó, 1977) comb. nov., and A. tatika (Szabó, 1977) comb. nov. Intraspecific variation, biological associations, and taxonomic history are discussed. DNA barcodes are provided and analyzed in the context of worldwide Trichacis and its sister genus Isocybus Förster.

Evolution of flexible biting in hyperdiverse parasitoid wasps.

One key event in insect evolution was the development of mandibles with two joints, which allowed powerful biting but restricted their movement to a single degree of freedom. These mandibles define the Dicondylia, which constitute over 99% of all extant insect species. It was common doctrine that the dicondylic articulation of chewing mandibles remained unaltered for more than 400 million years. We report highly modified mandibles overcoming the restrictions of a single degree of freedom and hypothesize their major role in insect diversification. These mandibles are defining features of parasitoid chalcid wasps, one of the most species-rich lineages of insects. The shift from powerful chewing to precise cutting likely facilitated adaptations to parasitize hosts hidden in hard substrates, which pose challenges to the emerging wasps. We reveal a crucial step in insect evolution and highlight the importance of comprehensive studies even of putatively well-known systems.

Mission possible: an optimised protocol for the unbarcodable Ceraphronoidea (Hymenoptera).

DNA barcodes provide a reliable and efficient solution to resolving cryptic species complexes and accelerate species discoveries. The superfamily Ceraphronoidea (Hymenoptera) is a group of parasitoid wasps for which a barcoding approach could be of great help, if it were not for the very poor results. The inability to obtain barcodes for the majority of treated ceraphronoids halts progress on the taxonomy of this hyperdiverse parasitoid group. We here present a working protocol for the barcoding of ceraphronoid wasps which yields a first-time over 90% success rate.

Tiny wasps, huge diversity - A review of German Pteromalidae with new generic and species records (Hymenoptera: Chalcidoidea).

BACKGROUND: Despite their ecological and economic importance, hymenopteran parasitoids are severely understudied. Even in countries with a long taxonomic history such as Germany, dating back to the 18th century and including prolific figures like Christian Gottfired Nees von Esenbeck and Otto Schmiedeknecht, those species-rich groups are seldom the subject of comprehensive research efforts, leaving their true diversity unknown. This is often due to their small size of a few millimetres on average, leading to difficulties in their identification and examination. The chalcidoid family Pteromalidae is no exception to this neglect. So far, 735 species have been reported from Germany. Estimating the diversity of this group is not possible, but it has to be assumed that many more species are still to be discovered in Germany. NEW INFORMATION: With this study, we improve the knowledge on pteromalid diversity and present new records of 17 genera and 41 species, previously unknown to occur in Germany. We also match and describe previously unknown sexes of two species, based on DNA barcode data. The results of this study were generated as part of the German Barcode of Life Project. The newly-recorded species are illustrated and notes on the biology and distribution are given. The ecological significance of Pteromalidae and potential value as indicators for nature conservation efforts are briefly discussed.

New records of German Scelionidae (Hymenoptera: Platygastroidea) from the collection of the State Museum of Natural History Stuttgart.

BACKGROUND: Scelionid wasps are arthropod egg parasitoids, many of which are relevant to global biosecurity. However, the scelionid fauna of Germany has not received much attention from professional taxonomists. NEW INFORMATION: Eleven species and four genera are recorded for the first time from Germany, including species of interest to agriculture and biological control. First genus records include Baryconus Förster, Macroteleia Westwood, Paratelenomus Dodd and Probaryconus Kieffer. First species records include B.europaeus (Kieffer), Idrisnigroclavatus (Kieffer), Idrissemiflavus (Kieffer), M.bicolora Kieffer, M.pannonica Szabo, Paratelenomussaccharalis (Dodd), Trimorusvaricornis (Walker), Trissolcusbasalis (Wollaston), Trissolcusbelenus (Walker), Trissolcuscolemani (Crawford) and Trissolcusflavipes (Thompson). COI barcodes are identified for the first time from B.europaeus and M.bicolora. Each species is illustrated and updated world distributions are provided. Implications for agriculture are discussed.

Taxonomic description and phylogenetic placement of two new species of Spalangiopelta (Hymenoptera: Pteromalidae: Ceinae) from Eocene Baltic amber.

Spalangiopelta is a small genus of chalcid wasps that has received little attention despite the widespread distribution of its extant species. The fossil record of the genus is restricted to a single species from Miocene Dominican amber. We describe two new fossil species, Spalangiopelta darlingi sp. n. and Spalangiopelta semialba sp. n. from Baltic amber. The species can be placed within the extant genus Spalangiopelta based on the distinctly raised hind margin of the mesopleuron. 3D models reconstructed from µCT data were utilized to assist in the descriptions. Furthermore, we provide a key for the females of all currently known Spalangiopelta species. The phylogenetic placement of the fossils within the genus is analyzed using parsimony analysis based on morphological characters. Phylogenetic and functional relevance of two wing characters, admarginal setae and the hyaline break, are discussed. The newly described Baltic amber fossils significantly extend the minimum age of Spalangiopelta to the Upper Eocene.

Integrative taxonomic review of the genus Synopsia Hübner, 1825 in the Middle East (Lepidoptera: Geometridae: Ennominae).

The geometrid genera Synopsia Hübner, 1825 and Synopsidia Djakonov, 1935 are revised, both being earlier validated at genus rank. Type specimens, original descriptions and additional specimens from different localities were examined. The revision is based on morphological characters, molecular data and distribution records. As a result, Synopsidia syn. nov. is regarded as junior synonym of the genus Synopsia. The synonymies of the species Scodonia tekkearia Christoph, 1883 and Synopsia znojkoi Djakonov, 1935 with Synopsia phasidaria phasidaria (Rogenhofer, 1873) comb. nov. are confirmed. Furthermore, Synopsidia phasidaria alvandi Wiltshire, 1966 syn. nov., Synopsidia phasidaria ardschira Brandt, 1938 syn. nov., Synopsidia phasidaria chiraza Brandt, 1938 syn. nov., Hashtaresia jodes Wehrli, 1936 syn. nov. and Synopsidia phasidaria mirabica Wehrli, 1941 syn. nov. are regarded as synonyms of Synopsia phasidaria phasidaria (Rogenhofer, 1873) comb. nov.. Synopsia phasidaria afghana (Wiltshire, 1966) comb. nov. is tentatively validated at subspecific rank. Synopsia centralis (Wiltshire, 1966) comb. nov., bona sp. is upgraded from subspecies to species level. Wing pattern, as well as male and female genitalia and diagnostic characters of examined genera and species are illustrated and analyzed. The distribution patterns of Synopsia phasidaria comb. nov. and Synopsia centralis bona sp., as well as the type localities of all discussed taxa, are provided. A complete checklist of the genus is given.

Taxonomic revision of the genus Nychiodes Lederer, 1853 (Geometridae: Ennominae: Boarmiini) with description of three new species-an integrative approach.

The non-European taxa of the genus Nychiodes Lederer, 1853 are revised. Type specimens of all described species and a large series of about 800 additional specimens were morphologically examined. More than 400 genitalia preparations were made and analyzed along with distributional and DNA barcode data. As a result of our integrative taxonomic approach, Nychiodes waltheri saerdabica Wehrli, 1938 syn. nov., is synonymized with N. waltheri Wagner, 1919; N. palaestinensis libanotica Zerny, 1933 syn. nov. is synonymized with N. palaestinensis Wagner, 1919 and the synonymy of N. persuavis Wehrli, 1929 syn. rev. with N. palaestinensis is confirmed; N. admirabila safidaria Wiltshire, 1943 syn. nov. is synonymized with N. admirabila Brandt, 1938; N. agatcha Brandt, 1938 syn. nov., N. subvirida disjuncta Wehrli, 1941 syn. nov. and N. subvirida taftana Brandt, 1941 syn. nov. are synonymized with N. subvirida Brandt, 1938. Also, N. variabila variabila Brandt, 1938 syn. nov., N. variabila opulenta Brandt, 1941 syn. nov., N. divergaria elbursica Wehrli, 1937 syn. nov., N. divergaria fallax Wehrli, 1939 syn. nov. and N. divergaria achtyca Wehrli, 1939 syn. nov. are synonymized with N. divergaria Staudinger, 1892. Nychiodes convergata sp. nov. from Israel, N. mirzayansi sp. nov. from the Iran and N. eberti sp. nov. from Turkey are described. Lecto- and paralectotypes are designated for N. palaestinensis, N. antiquaria, N. divergaria. Furthermore, N. antiquaria is reported as a new species for Pakistan, N. rayatica is reported as a new species for Iran and the hypothetical occurrence of N. amygdalaria in Iran is confirmed. Additionally, N. tyttha needs to be excluded from the genus. Wing pattern, male and female genitalia and diagnostic characters of all examined species are illustrated and distribution maps are provided. Illustrated keys based on genitalia, as well as a complete checklist of the genus is given here.

Biodiversi-TEA-A quick and easy handling method for arthropod trap material in ethanol.

A quick and easy handling method for ethanol-preserved arthropods is presented, based on tea filter bags and holders. The method is especially suitable for short term storage of specimens resulting from Malaise traps, yellow pan traps and pitfall traps and can be used directly in the field, for subsequent transport and processing of specimens in the laboratory.

The Fixator-A simple method for mounting of arthropod specimens and photography of complex structures in liquid.

Mounting and preparing arthropods in liquids for photography and further investigations is a challenging task and may lead to unsatisfactory results and, in the worst case, to damage to specimens. A new method is presented here, which allows the fixation of specimens of different sizes under various degrees of pressure. The method is illustrated by three case studies from different groups of insects and arachnids.

Microdissection and whole chromosome painting confirm karyotype transformation in cryptic species of the Lariophagus distinguendus (Förster, 1841) complex (Hymenoptera: Pteromalidae).

Karyotypes of two cryptic species of parasitoid Hymenoptera with n = 5 and 6 belonging to the Lariophagus distinguendus (Förster, 1841) complex, which includes cosmopolitan parasitoids of coleopteran stored-product pests, were studied using glass-needle based microdissection, reverse and cross-species fluorescence in situ hybridisation (FISH). This experiment strongly indicates that the largest metacentric chromosome in the karyotype with n = 5 originated from a particular fusion between the only acrocentric and a smaller metacentric chromosome of the set with n = 6, therefore confirming our previous hypothesis based on the karyotypic analysis using chromosome morphometrics. This study represents the first successful application of both microdissection and whole chromosome painting for the reconstruction of karyotypic rearrangements in closely related species of parasitoids, as well as in the order Hymenoptera in general.

Molecular and cytogenetic differentiation within the Lariophagusdistinguendus (Förster, 1841) species complex (Hymenoptera, Pteromalidae).

Several strains of the apparently well-known cosmopolitan synanthropic parasitoid of coleopteran stored-product pests, Lariophagusdistinguendus (Förster, 1841) from Western Europe, were studied using DNA sequencing and chromosomal analysis. The presence of at least two cryptic species with different COI sequences and chromosome numbers (n = 5 and 6) was supported. The species with n = 6 is associated with the drugstore beetle Stegobiumpaniceum (Linnaeus, 1758), whereas the other one with n = 5 mostly develops on the granary weevil Sitophilusgranarius (Linnaeus, 1758). A phylogenetic study revealed that the karyotype with n = 6 represents an ancestral character state in this complex. Consequently, the chromosome set with n = 5 which is characteristic of a particular internal clade, apparently originated via chromosomal fusion which was probably preceded by a pericentric inversion. If this is true, inverted chromosome segments could accumulate a number of genetic loci responsible for certain interspecific differences.

Parasitoid biology preserved in mineralized fossils.

About 50% of all animal species are considered parasites. The linkage of species diversity to a parasitic lifestyle is especially evident in the insect order Hymenoptera. However, fossil evidence for host-parasitoid interactions is extremely rare, rendering hypotheses on the evolution of parasitism assumptive. Here, using high-throughput synchrotron X-ray microtomography, we examine 1510 phosphatized fly pupae from the Paleogene of France and identify 55 parasitation events by four wasp species, providing morphological and ecological data. All species developed as solitary endoparasitoids inside their hosts and exhibit different morphological adaptations for exploiting the same hosts in one habitat. Our results allow systematic and ecological placement of four distinct endoparasitoids in the Paleogene and highlight the need to investigate ecological data preserved in the fossil record.

A new lineage of Cretaceous jewel wasps (Chalcidoidea: Diversinitidae).

Jewel wasps (Hymenoptera: Chalcidoidea) are extremely species-rich today, but have a sparse fossil record from the Cretaceous, the period of their early diversification. Three genera and three species, Diversinitus attenboroughi gen. & sp. n., Burminata caputaeria gen. & sp. n. and Glabiala barbata gen. & sp. n. are described in the family Diversinitidae fam. n., from Lower Cretaceous Burmese amber. Placement in Chalcidoidea is supported by the presence of multiporous plate sensilla on the antennal flagellum and a laterally exposed prepectus. The new taxa can be excluded from all extant family level chalcidoid lineages by the presence of multiporous plate sensilla on the first flagellomere in both sexes and lack of any synapomorphies. Accordingly, a new family is proposed for the fossils and its probable phylogenetic position within Chalcidoidea is discussed. Morphological cladistic analyses of the new fossils within the Heraty et al. (2013) dataset did not resolve the phylogenetic placement of Diversinitidae, but indicated its monophyly. Phylogenetically relevant morphological characters of the new fossils are discussed with reference to Cretaceous and extant chalcidoid taxa. Along with mymarid fossils and a few species of uncertain phylogenetic placement, the newly described members of Diversinitidae are among the earliest known chalcidoids and advance our knowledge of their Cretaceous diversity.

Transcriptome sequence-based phylogeny of chalcidoid wasps (Hymenoptera: Chalcidoidea) reveals a history of rapid radiations, convergence, and evolutionary success.

Chalcidoidea are a megadiverse group of mostly parasitoid wasps of major ecological and economical importance that are omnipresent in almost all extant terrestrial habitats. The timing and pattern of chalcidoid diversification is so far poorly understood and has left many important questions on the evolutionary history of Chalcidoidea unanswered. In this study, we infer the early divergence events within Chalcidoidea and address the question of whether or not ancestral chalcidoids were small egg parasitoids. We also trace the evolution of some key traits: jumping ability, development of enlarged hind femora, and associations with figs. Our phylogenetic inference is based on the analysis of 3,239 single-copy genes across 48 chalcidoid wasps and outgroups representatives. We applied an innovative a posteriori evaluation approach to molecular clock-dating based on nine carefully validated fossils, resulting in the first molecular clock-based estimation of deep Chalcidoidea divergence times. Our results suggest a late Jurassic origin of Chalcidoidea, with a first divergence of morphologically and biologically distinct groups in the early to mid Cretaceous, between 129 and 81 million years ago (mya). Diversification of most extant lineages happened rapidly after the Cretaceous in the early Paleogene, between 75 and 53 mya. The inferred Chalcidoidea tree suggests a transition from ancestral minute egg parasitoids to larger-bodied parasitoids of other host stages during the early history of chalcidoid evolution. The ability to jump evolved independently at least three times, namely in Eupelmidae, Encyrtidae, and Tanaostigmatidae. Furthermore, the large-bodied strongly sclerotized species with enlarged hind femora in Chalcididae and Leucospidae are not closely related. Finally, the close association of some chalcidoid wasps with figs, either as pollinators, or as inquilines/gallers or as parasitoids, likely evolved at least twice independently: in the Eocene, giving rise to fig pollinators, and in the Oligocene or Miocene, resulting in non-pollinating fig-wasps, including gallers and parasitoids. The origins of very speciose lineages (e.g., Mymaridae, Eulophidae, Pteromalinae) are evenly spread across the period of chalcidoid evolution from early Cretaceous to the late Eocene. Several shifts in biology and morphology (e.g., in host exploitation, body shape and size, life history), each followed by rapid radiations, have likely enabled the evolutionary success of Chalcidoidea.

Transcriptome and target DNA enrichment sequence data provide new insights into the phylogeny of vespid wasps (Hymenoptera: Aculeata: Vespidae).

The wasp family Vespidae comprises more than 5000 described species which represent life history strategies ranging from solitary and presocial to eusocial and socially parasitic. The phylogenetic relationships of the major vespid wasp lineages (i.e., subfamilies and tribes) have been investigated repeatedly by analyzing behavioral and morphological traits as well as nucleotide sequences of few selected genes with largely incongruent results. Here we reconstruct their phylogenetic relationships using a phylogenomic approach. We sequenced the transcriptomes of 24 vespid wasp and eight outgroup species and exploited the transcript sequences for design of probes for enriching 913 single-copy protein-coding genes to complement the transcriptome data with nucleotide sequence data from additional 25 ethanol-preserved vespid species. Results from phylogenetic analyses of the combined sequence data revealed the eusocial subfamily Stenogastrinae to be the sister group of all remaining Vespidae, while the subfamily Eumeninae turned out to be paraphyletic. Of the three currently recognized eumenine tribes, Odynerini is paraphyletic with respect to Eumenini, and Zethini is paraphyletic with respect to Polistinae and Vespinae. Our results are in conflict with the current tribal subdivision of Eumeninae and thus, we suggest granting subfamily rank to the two major clades of "Zethini": Raphiglossinae and Zethinae. Overall, our findings corroborate the hypothesis of two independent origins of eusociality in vespid wasps and suggest a single origin of using masticated and salivated plant material for building nests by Raphiglossinae, Zethinae, Polistinae, and Vespinae. The inferred phylogenetic relationships and the open access vespid wasp target DNA enrichment probes will provide a valuable tool for future comparative studies on species of the family Vespidae, including their genomes, life styles, evolution of sociality, and co-evolution with other organisms.

Evolutionary History of the Hymenoptera.

Hymenoptera (sawflies, wasps, ants, and bees) are one of four mega-diverse insect orders, comprising more than 153,000 described and possibly up to one million undescribed extant species [1, 2]. As parasitoids, predators, and pollinators, Hymenoptera play a fundamental role in virtually all terrestrial ecosystems and are of substantial economic importance [1, 3]. To understand the diversification and key evolutionary transitions of Hymenoptera, most notably from phytophagy to parasitoidism and predation (and vice versa) and from solitary to eusocial life, we inferred the phylogeny and divergence times of all major lineages of Hymenoptera by analyzing 3,256 protein-coding genes in 173 insect species. Our analyses suggest that extant Hymenoptera started to diversify around 281 million years ago (mya). The primarily ectophytophagous sawflies are found to be monophyletic. The species-rich lineages of parasitoid wasps constitute a monophyletic group as well. The little-known, species-poor Trigonaloidea are identified as the sister group of the stinging wasps (Aculeata). Finally, we located the evolutionary root of bees within the apoid wasp family "Crabronidae." Our results reveal that the extant sawfly diversity is largely the result of a previously unrecognized major radiation of phytophagous Hymenoptera that did not lead to wood-dwelling and parasitoidism. They also confirm that all primarily parasitoid wasps are descendants of a single endophytic parasitoid ancestor that lived around 247 mya. Our findings provide the basis for a natural classification of Hymenoptera and allow for future comparative analyses of Hymenoptera, including their genomes, morphology, venoms, and parasitoid and eusocial life styles.