A new extinct genus and species of Pristocerinae, †Archeonesia eocena Tribull, Pankowski & Colombo, gen. et. sp. nov., are described from upper Eocene Baltic amber from the Yantarny amber mine in the Kaliningrad region, Russia. Descriptions, remarks, illustrations, and comparisons to all extinct and extant Pristocerinae are provided. †Archeonesia is described as a new genus because neither the male nor the female can be placed in any previously described genera, although the female is most similar to Acrenesia and the male is most similar to Cleistepyris. Rare for Bethylidae, and Hymenoptera in general, this fossil contains both a male and female specimen that we are describing as conspecifics. A brief review from the paleoentomological literature is provided to describe how insect fossils containing evidence of reproductive behavior have been used to associate conspecifics in extinct species.

Amber, Bethylidae, Eocene, fossil, Pristocerinae

Bethylidae are a family of parasitoid wasps within the aculeate superfamily Chrysidoidea that have a cosmopolitan distribution and are known for attacking lepidopteran and coleopteran hosts, including agricultural pests like the navel orangeworm, pink bollworm, and coffee berry borer (Gordh et al. 1983; Abraham et al. 1990; Azevedo et al. 2018). Currently, the family contains nearly 3,000 species in about 100 genera within five extant subfamilies (Bethylinae, Epyrinae, Mesitiinae, Pristocerinae, and Scleroderminae) and four extinct subfamilies (†Elektroepyrinae, †Lancepyrinae, †Holopsenellinae, and †Protopristocerinae) (Azevedo et al. 2018; Colombo et al. 2020a, b). There are about 90 flat wasps fossil species described, with the oldest known extinct Bethylidae coming from early Cretaceous amber deposits (Engel et al. 2016). However, the greatest number of fossil bethylids comes from the Eocene, specifically Oise, Rovno, and Baltic amber deposits (Colombo et al. 2020a, 2021).

With over 1,000 species in 26 genera, Pristocerinae are the most speciose subfamily within Bethylidae and are found worldwide (Azevedo et al. 2018; Colombo et al. 2020a, 2021). The subfamily is known for its remarkable sexual dimorphism, with males possessing robust bodies, wings, and conspicuous eyes and ocelli while females are wingless, lack ocelli, and have eyes that are extremely reduced or missing (Alencar et al. 2018). The vast majority of Pristocerinae species are known from ‘males-only’ or ‘females-only’ and conspecific associations between males and females are rare, typically the result of collecting specimens copulating or rearing them from the same host (Azevedo et al. 2016; Alencar et al. 2018; Chen and Azevedo 2020).

There are 15 extinct species of Pristocerinae in 10 genera, with the oldest species, †Foenobethylus electriphilus (Cockerell) known from mid-Cretaceous Burmese amber (Cockerell 1920; Falières and Nel 2019, 2020; Colombo et al. 2020a). Of these species, 11 are known from males only, three are known from females only, and only one species, Pristocera skwarrae (Brues), was described from a fossil that contained both a male and female. Additionally, Brues (1933) does not include any illustrations of the species and it was presumed to be lost during World War II (Colombo et al. 2021).

About 50% of extinct Bethylidae come from Baltic amber deposits dated to the late Eocene, including four of the 15 extinct Pristocerinae from the genera Pseudisobrachium Kieffer, Cleistepyris Kieffer, and Pristocera Klug (Colombo and Azevedo 2019; Colombo et al. 2021). Here we present a new extinct genus of Pristocerinae from Baltic amber, †Archeonesia Tribull, Pankowski & Colombo, gen. nov. with the type species †A. eocena Tribull, Pankowski & Colombo, sp. nov. Like the lost Pristocera skwarrae, this amber fossil shows both a male and female together, granting us the rare opportunity to describe both sexes for a new extinct bethylid genus.

The specimens are embedded in Baltic amber sourced from the Yantarny mine in the Kaliningrad region. The piece was acquired from, trimmed, and polished by Marius Veta and has clear dorsal and ventral views, although the lateral views are obscured by bubbles. The type material is deposited in the American Museum of Natural History, USA (AMNH, curator: David Grimaldi) with the specimen catalogue number AMNH_IZC 00361788.

The specimens were studied with an Olympus SZX-10 stereomicroscope and photomicrographs were acquired with a DP27 digital camera, using Olympus’s Cellsens software. Multiple Z-stacks were compiled using Helicon Focus. The drawings were scanned and vectorized into Adobe Illustrator CS6 version 23.0.3, and images were edited and combined into a single plate using Adobe Photoshop CC.

Terminology for the integument and sculpturing follows Harris (1979) and general terms follow Lanes et al. (2020).

The specimens considered here were assigned to the subfamily Pristocerinae based on the following character states, in the key from Azevedo et al. (2018), for males: (1) the forewings without Rs+M veins, (2) the metanotum is well-developed medially; and for females: (1) the wings are absent, (2) the eyes are very reduced, and (3) the ocelli are absent.

The male specimen studied here does not have visible important morphological characters, mainly from the genitalia, which does not allow the correct identification of the genus. We performed comparisons with males of all 23 known pristocerine genera, excluding Anisobrachium Kieffer, †Ekaterina Colombo & Azevedo (in Colombo et al. 2020a) and Scaphepyris Kieffer, because these genera are known only from females.

The male of this species has the ninth abdominal segment undivided and with the posterior margin weakly concave. For this reason, this species does not belong to †Eopristocera Falières & Nel, Pristocera Klug, Prosapenesia Kieffer, Propristocera Kieffer and Pristonesia Alencar & Azevedo (in Alencar et al. 2018), because the males of Pristocera have the ninth abdominal segment divided into two lobes and the males of †Eopristocera, Prosapenesia, Propristocera and Pristonesia have the ninth abdominal segment with the posterior margin strongly concave.

The male of this species does not belong to Dissomphalus Ashmead, Protisobrachium Benoit and Trichiscus Benoit, because those males have the clypeus with median and lateral lobes not well outlined, whereas †A. eocena sp. nov. has the clypeus with median lobe clearly defined and lateral lobes reduced.

Additionally, the median clypeal lobe of †A. eocena sp. nov. is projected forward, triangular, and the median clypeal carina is lower than frons, and for this reason, this species does not belong to Acrenesia Alencar & Azevedo (in Alencar et al. 2018), Dracunesia Alencar & Azevedo (in Alencar et al. 2018), Eleganesia Alencar & Azevedo (in Alencar et al. 2018), Epynesia Alencar & Azevedo (in Alencar et al. 2018), Pristepyris Kieffer and †Uniceratops Colombo & Azevedo (in Colombo et al. 2020), because the males of Pristepyris have the median clypeal lobe short and truncate; the males of Eleganesia also have the median clypeal lobe short and with apical margin outcurved; males of Acrenesia and Dracunesia have the median clypeal lobe trapezoidal; males of Epynesia have the median clypeal lobe very projected and sharply angulated, and the male of †Uniceratops has the median clypeal carina higher than frons, horn-shaped.

†A. eocena sp. nov. has the dorsal pronotal area with the lateral and anterior margins distinct and for this reason, this species does not belong to Apenesia Westwood, Afgoiogfa Argaman, Foenobethylus Kieffer and Parascleroderma Kieffer, because the males of these genera have the dorsal pronotal area with the lateral and anterior margins barely distinct. Additionally, the males of the last three genera, together with Austranesia Alencar & Azevedo (in Alencar et al. 2018), have the prepectus very large, about 0.25× mesopectus length, whereas the male of this species has the prepectus reduced, about ~0.15× mesopectus length.

The male of this species does not belong to Caloapenesia Terayama, Calobrachium Gobbi & Azevedo and Pseudisobrachium Kieffer, because these males have the eyes very setose, and the median clypeal lobe is usually trapezoidal, whereas †A. eocena sp. nov. has the eyes glabrous and the median clypeal lobe triangular, as aforementioned.

The last possible genus is Cleistepyris Kieffer. However, males of Cleistepyris have the forewings with the junction of 2r-rs and Rs veins almost indistinct, pterostigma triangular, and the R cell as wide as 1Cu cell, and for this reason, this species does not belong to Cleistepyris, because the male of this species has the forewings with the junction of 2r-rs and Rs veins distinct, pterostigma lanceolate, and the R cell wider than 1Cu cell. Finally, due to the male morphological differences aforementioned, we are allocating this species to a new genus.

We concluded that the female is the same species as the male (see below), and we studied the female morphology to support the allocation of this species in a new genus. The female specimen studied here does have important morphological characters visible and because of this, we used the key proposed by Azevedo et al. (2018, p. 57–58). Following the key, it would go to the genus Acrenesia, because of the following characters: (1) the head is longer than wide; (2) the clypeus with the median lobe trapezoidal; (3) the eyes with more than one facet; (4) the mesonotum is triangular, short, transverse, and much wider than long; (5) the mesopleuron is quite large and is reaching at least the anterior third of metapectal-propodeal complex, in dorsal view; (6) the anterior portion of the metapectal-propodeal complex is slightly narrower than posterior one; (7) the metapectal-propodeal complex with inconspicuous constriction near the spiracles; and (8) the second abdominal segment without flap expanded laterally. However, †A. eocena sp. nov. has the pronotum as wide as the metapectal-propodeal complex, in dorsal view; and anteromesoscutum with anterior margin straight; whereas Acrenesia has the pronotum wider than the metapectal-propodeal complex, in dorsal view; and anteromesoscutum with anterior margin incurved. Only †Ekaterina is not included in the pristocerine key of Azevedo et al. (2018, p. 57–58), because this genus was published later by Colombo et al. (2020a). †Ekaterina has the mesonotum somewhat diamond-shaped and metapectal-propodeal disc with strongly evident constriction; however, †A. eocena sp. nov. has the mesonotum triangular and metapectal-propodeal disc with weakly evident constriction. So, this female does not belong to any known pristocerine genera, and we are describing it as a new genus.

The amber fossil contains both a male and female specimen of Pristocerinae, an occurrence of conspecifics in the subfamily that has only previously been recovered in Pristocera skwarrae, which was never photographed or drawn, and was lost in World War II (see Colombo et al. 2021). While the two specimens were not preserved copulating, we have chosen to assign them to the same species as conspecifics. To justify this decision, we reviewed the paleoentomological literature concerning the association of conspecifics in fossils and found three main reasons why authors choose to associate specimens in the same fossil as conspecifics: 1) The specimens were preserved during copulation, 2) the specimens present evidence that copulation just occurred or was about to occur, and 3) knowledge of reproductive and/or social behavior of extant relatives was used to justify the association.

When previously undescribed insects are preserved during copulation, authors describe them as the same species – examples have been found in both rock impressions and amber fossils from a variety of different terrestrial arthropods. For example, in Li et al. (2013), a rock compression fossil from the middle Jurassic of China shows two froghoppers copulating with the male’s aedeagus inserted into the female. In Klimov and Sidorchuk (2011) and Sidorchuk and Klimov (2011), a pair of copulating mites were redescribed from upper Eocene Baltic amber. Although there are other records of insects (especially Diptera) being preserved while mating in amber or as rock compressions, these publications broadly focus on describing behavior instead of acting as new species records (Grimaldi and Engel 2005; Takahashi et al. 2017).

Taxonomic records written from insects preserved while unequivocally copulating are rare, and more common are examples in which specimens are presumed to have already mated or were engaged in mating behavior. Evidence provided by the authors typically relies on the close proximity of supposed conspecifics, evidence of reproductive behaviors (like grasping), or evidence of exposed genitalia. For example, in Fischer and Hörnig (2019), the authors describe a new species of Tineidae from a male and female preserved in Baltic amber, arguing that the orientation of abdomens and genitalia, plus the rarity of moths in Baltic amber (less than 1% of all inclusions), means that it is unlikely the fossil represents a random co-occurrence. In Andersen and Poinar (1998), a new species of water strider was described from a Dominican amber piece that contained a male grasping a female, a position that suggests that they were trapped just before, during, or after mating. From Burmese amber, Chen and Su (2019) described a new species of Zoraptera in which copulation was suspected based on the identification of the male’s intromittent organ.

Finally, authors also rely on knowledge of extant relatives’ behavior to suggest that male and female conspecifics were preserved before, during, or just after mating. From Mexican amber, Macadam and Ross (2016) describe two mayfly imagos (a male and female) as the same species, suggesting that it’s unlikely the pair would have been caught in the same amber if not during a mating swarm given the short lifespan of extant adult Ephemeroptera. From mid-Cretaceous Burmese amber, a new species of dance fly was described from possible conspecifics with the justification that extant dance flies only form aggregations during courtship and mating. The authors also rely on possible evidence of the consumption of a nuptial meal by the female and sexual dimorphism in the inflated antennae of the male (Zhang et al. 2021). Gregarious mating behavior is also used as a justification for conspecific association of a whitefly species, especially given that all previous extinct Aleyrodidae were only known from individual specimens (Szwedo and Drohojowska 2016). From Upper Cretaceous Burmese amber, Cockx and Mckellar (2018) described a new genus and species of Crabronidae from two males and a female, noting that the co-occurrence of the three could be due to either social behavior (as seen in extant relatives) or evidence of mating. It should also be noted that there are examples in the taxonomic literature in which no justification is given to support the association of conspecifics beyond the preservation of a male and female together (Szadziewski and Grogan 1998; Fanti and Kupryjanowicz 2017). Similarly, there are instances where authors do not assign potential conspecifics to the same species, but instead place them in the same genus (Wichard et al. 2020). Within Bethylidae, there have only been two other instances of sexual association entirely from fossils – the aforementioned †Pristocera skwarrae and †Lytopsenella maritima Ramos and Azevedo (from the subfamily Bethylinae). Beyond being found in the same amber piece, Brues (1933) does not provide justification for the former, and Ramos et al. (2014) cite shared anatomical features (sexual dimorphism is low in the species) and preservation in the same piece.

Herein, we associate the male and female syninclusions as conspecifics based on our knowledge of the reproductive behavior of extant Pristocerinae. Female Pristocerinae are apterous and rarely collected by the same methods that commonly capture winged males. Occasionally, males and females will be captured attached (in copula) from Malaise traps and yellow pan traps as a result of phoretic copulation, in which the male transports the female while mating (Azevedo et al. 2016; Chen and Azevedo 2020). Phoretic copulation is observed in several Hymenopteran families with extreme sexual dimorphism, such as Tiphiidae, Mutillidae and Bethylidae, and is hypothesized as a method to aid in the dispersal of apterous females in need of hosts or food sources (Azevedo et al. 2016; Vivallo 2021). The male and female described here might have been trapped in resin millions of years ago, much in the same way that an extant male Pristocerinae occasionally transports his mate to her demise.

In addition to †A. eocena sp. nov., four other species of Pristocerinae are known from Baltic amber deposits – †Pristocera skwarrae, †Cleistepyris baryonyx Colombo, Gobbi & Azevedo, †Pseudisobrachium elatus (Brues), and †Pseudisobrachium inhabilis (Brues) (Colombo et al. 2020a, 2021). Besides Baltic amber, Rovno amber deposits are also linked to the upper Eocene and contain extinct Pristocerinae – †Pseudisobrachium megalosaurus Colombo, Gobbi & Azevedo, †Pseudisobrachium stegosaurus Colombo, Gobbi & Azevedo, and †Cleistepyris allosaurus Colombo, Gobbi & Azevedo (Colombo et al. 2020a). Given that Baltic amber from the Kaliningrad region has been extracted for more than a century and averages much greater yields than Rovno amber from Ukranian mines in recent years, it’s possible that †A. eocena sp. nov., might also be recovered from Rovno amber (Colombo et al. 2021). This possibility is reinforced by the finding that 50% of Rovno amber Hymenoptera fauna are also found in Baltic amber (Perkovsky 2018).

Of the upper Eocene Pristocerinae, Cleistepyris, Pristocera, and Pseudisobrachium are known from both extant and extinct species, while †Archeonesia is only known from the extinct species described here. From lower Eocene Oise amber, there is also the extinct monotypic genus Eopristocera. While extant Pristocera are known from the Palaearctic (as well as other parts of the world), extant Cleistepyris are known only from Nearctic and Neotropical regions, and extant Pseudisobrachium are primarily found in southern distributions and common in the neotropics, which demonstrates how the biogeography of Pristocerinae has changed since the Eocene (Azevedo et al. 2018; Colombo et al. 2020a). The end of the Eocene marked the start of a cooling period, which likely drove the extinction of thermophile bethylids like †Archeonesia and †Eopristocera, as well as the local extinction of Cleistepyris in Europe (Bogri et al. 2017).

Compared to other Bethylidae families like Epyrinae (30 species) and Bethylinae (20 species), there are fewer records of extinct Pristocerinae (Ramos et al. 2014; Azevedo et al. 2018; Colombo et al. 2021). With the description of †Archeonesia eocena, there are now 16 extinct species of Pristocerinae. Given that Pristocerinae are the most speciose and abundant subfamily of Bethylidae, it is interesting that it would rank third in extinct diversity. However, this could be due to the extreme sexual dimorphism within the subfamily reducing the number of individuals that were captured in resin – unlike Pristocerinae, most females of Bethylinae and Epyrinae can fly. In Epyrinae and Bethylinae, more than 65% of fossil records are from females, but in Pristocerinae, the opposite is seen with more than 70% of fossil records from males, suggesting that when the female is winged in Bethylidae, the chances are greater that a specimen will be preserved as a fossil.

Finally, extinct Bethylidae known from both sexes are extremely rare. In addition to †Pristocera skwarrae, one other extinct species of Bethylidae is known from both sexes. In Bethylinae, one species (representing 5% of extinct bethylines), †Lytopsenella maritima, is known and the association was justified by the weak sexual dimorphism and proximity of the specimens in the amber. In Epyrinae, one species (representing 3% of all extinct epyrines), Epyris staphylinoides (Hope), is known for only one extinct male and other extant males and females, so the association was not performed by representatives in the same fossil. While we hope for the discovery of more fossils that have preserved both sexes of a species together, it is likely that their occurrence will continue to be rare in Bethylidae.

We thank the subject editor Dr. Michael Ohl, Dr. Evgeny Perkovsky and one anonymous reviewer for their valuable suggestions. The authors are thankful for the resources and support from their home institutions and research departments. CMT thanks the biology department and office of the provost at Farmingdale State College (State University of New York), as well as the American Museum of Natural History for accessioning the fossil. WDC thanks CAPES – Demanda Social for its Ph.D. scholarship. Additionally, we are thankful for the generosity of the Pankowski family in supporting paleontological research through their donation of fossils to research institutions.