1. Evolutionary Biology

A Mesozoic clown beetle myrmecophile (Coleoptera: Histeridae)

  1. Yu-Lingzi Zhou
  2. Adam Ślipiński
  3. Dong Ren
  4. Joseph Parker  Is a corresponding author
  1. Institute of Zoology, Chinese Academy of Sciences, China
  2. CSIRO, Australia
  3. Capital Normal University, China
  4. California Institute of Technology, United States
https://doi.org/10.7554/eLife.44985
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  1. Yu-Lingzi Zhou
  2. Adam Ślipiński
  3. Dong Ren
  4. Joseph Parker
(2019)
A Mesozoic clown beetle myrmecophile (Coleoptera: Histeridae)
eLife 8:e44985.
https://doi.org/10.7554/eLife.44985
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Abstract

Complex interspecies relationships are widespread among metazoans, but the evolutionary history of these lifestyles is poorly understood. We describe a fossil beetle in 99-million-year-old Burmese amber that we infer to have been a social impostor of the earliest-known ant colonies. Promyrmister kistneri gen. et sp. nov. belongs to the haeteriine clown beetles (Coleoptera: Histeridae), a major clade of ‘myrmecophiles’—specialized nest intruders with dramatic anatomical, chemical and behavioral adaptations for colony infiltration. Promyrmister reveals that myrmecophiles evolved close to the emergence of ant eusociality, in colonies of stem-group ants that predominate Burmese amber, or with cryptic crown-group ants that remain largely unknown at this time. The clown beetle-ant relationship has been maintained ever since by the beetles host-switching to numerous modern ant genera, ultimately diversifying into one of the largest radiations of symbiotic animals. We infer that obligate behavioral symbioses can evolve relatively rapidly, and be sustained over deep time.

https://doi.org/10.7554/eLife.44985.001

eLife digest

Many animals live lives that are closely intertwined with those of other species. While a clown fish sheltering within the tentacles of a sea anemone may be a textbook example, ‘symbiotic’ interactions that occur inside ant nests are among some of the most dramatic.

Known as myrmecophiles – after the Greek for ‘ant lovers’, many insects, spiders and mites have evolved to live alongside ants in one way or another. Some of these animals display elaborate behaviors – like mouth-to-mouth feeding or grooming of worker ants – which assimilates them into the nest society; some even release chemicals that mimic the ants’ own scents to avoid being detected as an intruder.

The earliest examples of ancestral ants are found encapsulated in 99-million-year-old amber from a mine in northern Myanmar (Burma). Zhou et al. have now discovered an ancient beetle, perfectly preserved in the same amber deposits, that may have also lived within the colonies of those earliest-known ants. Based on its appearance, the beetle – named Promyrmister kistneri – belongs within a subfamily of clown beetles (called the Haeteriinae) that are all specialized nest intruders with dramatic behavioral and chemical adaptations that help them to infiltrate ant colonies.

The ancient clown beetle shares several of features with its modern relatives – including thick, spiked legs and well-protected head and antennae – which are believed to help the beetles withstand handling by the ants’ jaws. The specimen also has glands near the base of its legs, implying that it also released chemical signals that may have helped it to deceive or pacify the ancient ants.

The fact that this extinct clown beetle is as old as the earliest-known ants implies that the close relationship between these insects has been sustained for an exceptionally long time. It is potentially the oldest known example of a symbiotic interaction in the animal kingdom that depends on social interactions between the two organisms. However, the host ants of Promyrmister are believed to be long-extinct, suggesting that symbiotic clown beetles had to switch to living inside colonies of modern ants to circumvent their own extinction. This flexibility to adapt to new partner species may be a critical feature that allows some symbiotic organisms to persist throughout evolution.

https://doi.org/10.7554/eLife.44985.002

Introduction

A pervasive feature of colony-forming insect societies is the profusion of intruder arthropods that have evolved to exploit their rich resources (Kistner, 1979; Kistner, 1982; Hölldobler and Wilson, 1990; Parker, 2016). The diversity of such organisms is impressive, with ~10,000 species hypothesized to target or profit from ant nests alone (Elmes, 1996). Hostility of ant workers to virtually all non-nestmate organisms has selected for defensive or host-deceptive adaptations in myrmecophiles which are often phenotypically remarkable, involving changes in anatomy, chemical ecology and behavior (Kistner, 1979; Kistner, 1982; Hölldobler and Wilson, 1990; Parker, 2016). In a number of cases, traits have arisen that enable the myrmecophile to manipulate worker behavior, circumventing aggression and enabling social interactions to evolve that assimilate the symbiont into colony life. Such relationships rank among the most behaviorally intimate interactions known between animal species (Kistner, 1979; Hölldobler and Wilson, 1990; Parker, 2016), and are typically achieved by the myrmecophile’s capacity to mimic the chemical and/or tactile cues involved in nestmate recognition (Kistner, 1979; Hölldobler and Wilson, 1990; Parker, 2016). The clown beetle family Histeridae includes multiple lineages that have independently evolved myrmecophily (Parker, 2016; Kovarik and Caterino, 2005), including Haeteriinae, a subfamily of ~335 described species comprising possibly the single largest radiation of myrmecophiles known within the Coleoptera (Parker, 2016; Kovarik and Caterino, 2005; Helava et al., 1985). We report the discovery of a crown-group haeteriine in Upper Cretaceous Burmese amber, revealing that the clown beetle-ant interaction has an exceptionally deep evolutionary history. To our knowledge, the relationship constitutes the most ancient behavioral symbiosis known in the Metazoa.

Results and discussion

Systematic palaeontology

  • Order Coleoptera Linnaeus, 1758

  • Superfamily Histeroidea Gyllenhal, 1808

  • Family Histeridae Gyllenhal, 1808

  • Subfamily Haeteriinae Marseul, 1857

  • Promyrmister kistneri Zhou, Ślipiński and Parker gen. et sp. nov. 

Holotype

Sex unknown. CNU-008021, deposited in Key Laboratory of Insect Evolution and Environmental Changes, Capital Normal University, Beijing. The holotype is well preserved in a small, transparent amber piece, 5.5 mm length ×3.5 mm width (Figure 1—figure supplement 1A). The entire external anatomy is observable (Figure 1A‒C), but the left region of the dorsal side is partially covered by white exudate (Figure 1A,G) emanating from the ventral side of the pronotal margin (arrow in Figure 1A,G).

Figure 1 with 2 supplements see all
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Promyrmister kistneri gen.et sp. nov.

(A) Dorsal habitus of holotype CNU-008021 with origin of exudate globule (black arrow) and elytral striae (white arrows) indicated; the three lateral striae are complete (top three arrows), the three medial striae appear incomplete. (B) Right lateral habitus with flight wing indicated. (C) Ventral habitus, boxed region expanded in panel H. (D) Head, dorsal and (inset) frontal views, with antennal scapes and frontoclypeal carina (FC) indicated. (E) Right foreleg, laterally expanded femur and tibia indicated. (F) Proventral-mesoventral boundary showing proventral keel with posterior incision. (G) Left pronotal margin, lateral view, showing possible origin of putative glandular exudate; arrow in G corresponds to that in panel A. (H) Ventrite one showing proximal leg segments (Trc: trochanter) and postcoxal gland openings (white arrows), with globule of putative exudate emanating from left postcoxal gland opening.

https://doi.org/10.7554/eLife.44985.003

Diagnosis of new genus and species

Haeteriine histerid that is distinguished from all other genera and species of Haeteriinae by possession of the following combination of characters: (1) deep depression behind meso- and metacoxae (Figure 1C,H); (2) metaventral postcoxal line recurved and extending laterally to metanepisternum (Figure 1B,C; (3) three complete striae on each elytron (Figure 1A); (4) lack of dorsal furrows on pronotum demarcating glandular lobe (Figure 1A,B); (5) strongly developed apical spur on protibia (Figure 1D,E); (6) frontoclypeus carinate medially (Figure 1D, Figure 1—figure supplement 1B,C); (7) triangular-shaped cavities to receive scapes of antennae (Figure 1—figure supplement 1B,C); (8) glandular opening in postcoxal cavity behind metacoxae (Figure 1H; Figure 1—figure supplement 2A,B,D). Promyrmister specifically differs from the closely related Haeterius in having deep epistomal depressions (Figure 1—figure supplement 1B,C; compare to Figure 2—figure supplement 1D), carinated epistomal striae convergent in the middle (Figure 1—figure supplement 1B,C) and paddle-shaped protibia with large apical spur (Figure 1D,E).

Locality and age

The holotype inclusion is derived from an amber mine located near Noije Bum, Tanaing, Kachin, Myanmar. The U-Pb dating of zircons from the volcanoclastic matrix yielded an age of 98.79 ± 0.62 million years (Shi et al., 2012).

Etymology

The generic name Promyrmister is a combination of the Greek πϱó (pro) meaning ‘before’ or ‘early’, μûρμηξ (myrmex) meaning ‘ant’, and Hister Linnaeus, type genus of Histeridae. The name refers to the likely symbiotic habits of the fossil taxon inside early ant colonies. The gender is masculine. The specific epithet recognizes the lifetime contribution of Dr. David H. Kistner, a global authority on social insect symbionts.

Description

Length 3.2 mm, width 2.3 mm. Body elongate oval (Figure 1A,C), moderately convex (Figure 1B); black or dark brown with dorsal surfaces bearing short and somewhat squamiform setae, visible along pronotal and elytral edges but on dorsal side often obscured by accumulation of water/dirt and appearing as tiny granules; interstices between setae moderately to distinctly shiny.

Head only partially visible, deeply inserted into prothorax (Figure 1D; Figure 1—figure supplement 1B). Frons with distinct frontoclypeal carina (FC in Figure 1D), and widely interrupted frontal stria, the lateral parts of which extend to the frontoclypeal carina, connecting to inwardly-arching epistomal striae (Figure 1—figure supplement 1C). Clypeus bordered by deep epistomal depressions to receive antennal scapes in repose (Figure 1—figure supplement 1B). Clypeus and labrum apparently fused but with distinct transverse ridge above the base of labrum. Mandibles strongly arcuate apically. Antennal scape large and triangular (Figure 1—figure supplement 1B), covering eye, and densely rugose dorsally.

Prothorax (length 1.0 mm and width 1.8 mm) widest at base, sides weakly rounded, converging anteriorly, anterior angles distinctly projecting and rounded; posterior angles weakly obtuse (Figure 1A). Lateral margins crenulate, each projection bearing weakly squamiform setae. Pronotum with marginal stria complete anteriorly and along lateral margins; sides without impressions or obvious gland openings along lateral carina; disc weakly convex, setose. Proventral lobe strongly prominent medially, covering most of ventral head surface and extending laterally to antennal cavities without visible marginal stria. Prosternal process (proventrite) narrowly elevated with apex about 0.1 times as broad as prothorax, expanding apically and deeply emarginate at apex (Figure 1F); prosternal carinae converging anteriorly but not apparently joined; junction between proventrite and prosternal lobe deeply depressed. Antennal cavity present on anterior angles of hypomeron, deep and completely closed from below via proventral alae (Figure 1—figure supplement 1B). Procoxae not clearly visible. Trochanter large, triangular and bearing several long setae; profemur very broad, width nearly 0.45 mm, and flat, covering most of the ventral side of prothorax (Figure 1E); protibia flat and expanded, width about 0.38 mm, bearing row of strong spines along external edge and an apical spur (Figure 1E); protarsi short and thin, sitting in straight groove on dorsal side of protibia (Figure 1D). Scutellum obscured dorsally by secretion (Figure 1A). Elytra (1.8 mm length ×2.3 mm width), with relatively complete dorsal striae 1‒three and reduced striae 4–6 (white arrows in Figure 1A); outer subhumeral stria complete, sutural stria very fine and visible apically.

Mesoventrite between mesocoxae very broad, about 1/3 of body width at the same position (Figure 1C); anterior margin projecting medially fitting into prosternal process (Figure 1F); Margin between metaventrite and visible abdominal ventrite one shallowly grooved. Ventrite one weakly convex medially, deeply concave laterally (Figure 1C) to accommodate strongly flattened legs. Postcoxal lines behind meso- and metacoxae completely recurved dorsally; discrimen complete. Hind coxae triangular (Figure 1C,H; Figure 1—figure supplement 2A–C), large and broadly separated from each other, with numerous regular rows of oblique striae on inner surface (Figure 1—figure supplement 2A–C); hind femur oval-shaped, distinctly large and flat, enveloping small and setose trochanter (Figure 1C,H). Hind tibia flat, paddle-like, about as broad as femur (Figure 1C), with a row of strong spines along external edge, double rows of stiff and apically modified setae along inner edge, inner surface with transverse ridges and row of pointed spines before; apical spur small. Tarsal formula 5-5-5. Hind tarsi slim, locating on inner side of tibia, tarsomere one longest, length subequal to tarsomeres 2‒four combined. Wings visible apically (Figure 1B), presumed functional.

Abdomen with median part of ventrite one delimited to a flat and polished central plate by inner abdominal stria, much longer than the remaining ventrites combined (Figure 1C); postcoxal line (outer abdominal stria) recurved and strongly diverging laterally (Figure 1B,C); also with deeply depressed rest for hind tibia outside the postcoxal line. Large abdominal gland opening behind hind coxa located between inner and outer abdominal striae (Figure 1H), on right side with exudate flowing out (paired arrows in Figure 1H; Figure 1—figure supplement 2B,D, note that laser reflectance indicates this is solidified material and not a gas/air bubble, which would appear dark). Ventrites 2‒four equal in length, without posterior marginal striae.

Systematic position

The fossil beetle is placed in Histeridae based on its possession of the following characters (Kovarik and Caterino, 2000): (i) a broad and compact body shape (Figure 1A,C); (ii) striate elytra that expose the two posterior abdominal tergites (Figure 1A); ii) five visible abdominal sternites (Figure 1C); (iv) short legs with broad, flattened tibiae (Figure 1C,E); (v) antenna short and geniculate with compact, 3-segmented club (Figure 1—figure supplement 1B); (vi) antenna retracting into cavity underneath the pronotum (Figure 1—figure supplement 1B); (vii) tarsal formula 5-5-5. Of the 11 histerid subfamilies (Bouchard et al., 2011), Promyrmister can be placed unequivocally in the subfamily Haeteriinae based on the following characters: head deflexed, with clypeus arched downwards in a different plane to the vertex (Figure 1D); antenna with enlarged, triangular scape received in repose in frontal groove and hiding the eye (Figure 1D; Figure 1—figure supplement 1B,C); antennal cavities located on hypomeron (Figure 1—figure supplement 1B), covered from below by proventral alae; proventral lobe strongly developed anteriorly, covering head from below, and extending laterally to form proventral alae (without lateral notch); proventral keel narrowly elevated between coxae and distinctly emarginate posteriorly to receive the projecting mesoventral process (Figure 1F). Additionally, the front, mid, and hind legs are extremely broad (Figure 1C,E), which is a feature of the clade Yarmister + Haeteriinae (Caterino and Tishechkin, 2015), in which Yarmister lacks the distinctly elevated proventral keel with posterior incision, which is present in Promyrmister and is an autapomorphy of the Haetaeriinae (Helava et al., 1985). The labrum of the fossil specimen may also be fused, but fossilization position precludes definitive assessment.

Within Haeteriinae, the new taxon appears to bear a close relationship to the genus Haeterius Erichson and some closely allied genera that share several morphological characters supporting their monophyly (Yélamos, 1997), principally the broad and externally rounded tibiae, the deep depressions behind meso- and metacoxae to accommodate retracted legs (Figure 1C,H) (Caterino and Tishechkin, 2015), the metaventral postcoxal line being recurved and extending laterally to the metanepisternum (Figure 1B,C), and the presence of three complete striae on each elytron (Figure 1A). Morphological features of the extant Haeterius, with key character states shared with Promyrmister, are shown in Figure 2—figure supplement 1 (see Diagnosis for separation of Promyrmister and Haeterius). Consistent with our evaluation of Promyrmister’s likely phylogenetic placement, both cladistic and Bayesian analysis of a set of morphological characters (Caterino and Tishechkin, 2015) from the fossil specimen and a selection of Recent histerid taxa places the new taxon within Haeteriinae as sister to Haeterius (Figure 2A,B; Figure 2—figure supplements 2 and 3). We infer that Promyrmister represents an extinct Cretaceous lineage that belongs within the crown-group of Haeteriinae.

Figure 2 with 3 supplements see all
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Phylogenetic relationships of Promyrmister.

(A) Consensus Bayesian Inference (BI) tree of representative histerid taxa including Haeteriinae, and Promyrmister. Posterior probabilities above 0.8 are shown on branches. (B) The consensus parsimony tree (MP) using TNT under the Traditional Search; bootstrap percentages above 80 are shown on branches. (C) Habitus photograph of Haeterius (H. ferrugineus), an inferred extant close relative of Promyrmister (photo credit: C. Fägerström) (D, E) Living Haeterius ferrugineus beetles interacting with Formica (D) and Lasius (E) host ants (photo credit: P. Krásenský). (F) Reconstruction of Promyrmister with stem-group host ant and larva (ant based on Gerontoformica).

https://doi.org/10.7554/eLife.44985.006

Promyrmister and deep-time persistence of a social symbiosis

Symbiotic relationships in which different animal species interact socially with each other have arisen sporadically across the metazoan tree of life. Such relationships encompass a spectrum of dependency, from transient, facultative associations seen in mixed-species groups of insectivorous birds (Sridhar et al., 2009), cetaceans and ungulates (Stensland et al., 2003; Goodale et al., 2017), to obligate symbiotic lifestyles typified by brood parasitic cuckoos, cowbirds (Johnsgard, 1997), mutualistic cleaner fish (Grutter, 1999) and oxpeckers (Nunn et al., 2011). Of all animal groups, however, the complex societies of ants play host to the greatest diversity of behavioral symbionts. Several major radiations of myrmecophiles are known, each containing hundreds of symbiotic species, including the lycaenid butterflies (Pierce et al., 2002), eucharitid wasps (Murray et al., 2013), paussine ground beetles (Moore and Robertson, 2014) and multiple lineages of rove beetles (Kistner, 1979; Kistner, 1982; Parker, 2016; Parker and Grimaldi, 2014; Seevers, 1965; Maruyama and Parker, 2017). The diversity and often-broad geographic ranges of these clades imply that their relationships with ants are evolutionarily ancient (Parker and Grimaldi, 2014; Yamamoto et al., 2016). Although fossil myrmecophiles are known from as far back as the Eocene (Parker and Grimaldi, 2014; Wasmann, 1929), ant eusociality is known to be at least twice as old, with the earliest definitively social ants occurring in Upper Cretaceous Burmese amber (Barden and Grimaldi, 2016). Whether their colonies were targeted by myrmecophiles has, however, been unclear: ants are comparatively scarce in Cretaceous ambers (Grimaldi and Agosti, 2000; LaPolla et al., 2013; Barden, 2016; Barden, 2018), and myrmecophilous invertebrates typically live at densities orders of magnitude lower than their hosts (Kistner, 1979). The unlikely discovery of a myrmecophile clown beetle in Burmese amber reveals that a major radiation of ant symbionts has its origins in Mesozoic ant societies.

Analysis of Promyrmister’s morphology and phylogenetic position indicates the new genus represents an extinct lineage within the crown-group of Haeteriinae, a clade of obligate myrmecophiles (Figure 2A,B; Figure 2—figure supplements 2 and 3; see Materials and methods). In haeteriine taxa for which detailed behavioral observations exist, the beetles have been shown to engage in intimate behaviors with ants, involving stomodeal trophallaxis (mouth-to-mouth feeding) (Wheeler, 1908; Henderson and Jeanne, 1990; Akre, 1968), grooming workers with their appendages (and being groomed or licked by hosts in return) (Akre, 1968), physically grasping onto ants (phoresis) (Akre, 1968; von Beeren and Tishechkin, 2017), or being carried around nests by workers (Kistner, 1982). Mimicry of colony cuticular hydrocarbons occurs (Lenoir et al., 2012), as well as chemical manipulation of host ants via ‘appeasement’ substances exuded from gland openings on the margins of the prothorax (Kistner, 1982; Seyfried, 1928) or in the postcoxal regions of the beetle’s underside (Figure 2—figure supplement 1C). Promyrmister appears to be closely allied to the extant genus Haeterius (Figure 2A–C; Figure 2—figure supplement 1). This genus and a handful of closely related taxa including Eretmotus, Sternocoelis and Satrapes comprise the only group of Haeteriinae known to occur in the Palaearctic, consistent with the Eurasian palaeolocality of Promyrmister in Burmese amber. Like all of these genera, Promyrmister exhibits classical haeteriine attributes that are thought to be true adaptations for myrmecophily, including broad expansions of the tibiae with spines on the outer margin (Figure 2C; Figure 2—figure supplement 1E), short tarsi received on the outer face of each tibia (Figure 2—figure supplement 1E), a triangular antennal scape (Figure 2—figure supplement 1D), pronounced antennal cavities on the prothoracic hypomeron (Figure 2—figure supplement 1D) and a broad proventral lobe to fully embrace the retracted head (Figure 2—figure supplement 1B–D). Those features are thought to be protective modifications that enable myrmecophile beetles to withstand handling by ant mandibles (Parker, 2016).

We and others have previously described rove beetles (Staphylinidae) in the Burmese palaeofauna that were putative symbionts of termite colonies (Yamamoto et al., 2016; Cai et al., 2017). These specimens exhibit a defensive ecomorphology and are thought to have been persecuted intruders that were not behaviorally integrated into their host’s societies (Yamamoto et al., 2016; Cai et al., 2017). In contrast, Haeteriinae embody a form of true behavioral symbiosis, where the relationship with host ants can involve social interactions (Figure 2D,E; Figure 4). The prothoracic glandular openings of Haeteriinae that secrete putative appeasement compounds are challenging to demonstrate even in extant taxa, but in the Promyrmister holotype, a large globule of possible exudate originates from the left margin of the prothorax, consistent with the position of such glands (Figure 1A,G). Additionally, Promyrmister possesses clear postcoxal secretory glands (Figure 1H; Figure 1—figure supplement 2A,B,D), with a globule of possible exudate emanating from the postcoxal gland opening on the right side of the body (Figure 1H; Figure 1—figure supplement 2B,D). Beyond Promyrmister’s phylogenetic position within the Haeteriinae clade, the fossil’s anatomy implies a chemical strategy to become accepted or at least tolerated inside colonies (hypothetical reconstruction in Figure 2F), akin to modern haeteriine species that have so far been examined (Akre, 1968; Lenoir et al., 2012; Seyfried, 1928).

What were the Cretaceous host ants of Promyrmister? All ants thus far described from Burmese amber belong to stem-group Formicidae, including members of the extinct subfamily Sphecomyrminae and three other genera, Gerontoformica, Myanmyrma and Camelomecia that similarly lack crown-group features but are placed incertae sedis within Formicidae (Barden, 2016; Barden, 2018). In contrast, fossils of definitive crown-group ant subfamilies are absent, or vanishingly rare, among the thousands of ant inclusions now recovered from this amber deposit (Barden, 2016; McKellar et al., 2013) (P. Barden, personal communication). Crown-group ants are also unknown from contemporaneous Charentese amber (Barden, 2016). We posit that the overwhelming prevalence of stem-group ants in Burmese amber implies that they were potential hosts of Promyrmister (Figure 2F). Such a scenario entails that haeteriines may not have originated with the modern ant groups that host them today; instead myrmecophily evolved first in stem-group ant colonies, with the beetles later switching to crown-group ants. We cannot, however, rule out an alternative scenario, that an as-yet undiscovered diversity of crown-group ants were, in fact, present in the Burmese palaeofauna, and it was these that selected for the early evolution of myrmecophily. Molecular dating indicates that crown-group ants had originated by this time (Brady et al., 2006; Moreau and Bell, 2013; Borowiec et al., 2017a) (see dotted lines in Figure 3). If present in this ancient ecosystem, perhaps their cryptic biologies limited their entrapment in amber.

Figure 3
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Antiquity of Promymister implies pervasive host switching of Haeteriinae from Cretaceous to Recent.

Age of Promyrmister is shown (red circle). The inferred window of occurrence of stem-group ants is indicated by the top orange bar. The ages of crown-group ants as a whole, New World (NW) army ants, and other specific subfamilies and genera that are known hosts of Haeteriinae are also presented. Orange bars extend back from the Recent to the age of the earliest-known fossil; dotted lines extend back to molecularly-inferred origins of crown groups. Molecular dating implies crown-group ants existed at the same time as Promyrmister, but stem-group ants are the only ants so far known in Burmese and other contemporaneous ambers. All modern host ant genera are inferred to have Cenozoic origins, implying extensive host switching between the inferred Early Cretaceous origin of Haeteriinae and the present day.

https://doi.org/10.7554/eLife.44985.010

Whether haeteriines evolved in stem- or crown-group ant colonies, their original hosts are presumably long-extinct. The present-day host associations of haeteriines imply that these myrmecophiles have host-switched between many modern ant lineages (Figure 3). The beetles have been recorded in colonies of ant species scattered across the subfamilies Dolichoderinae, Dorylinae, Formicinae, Myrmicinae and Ponerinae (Helava et al., 1985; Tishechkin, 2007) (Figure 3). We suggest that it is this capacity for host switching that may explain the great longevity of the clown beetle-ant symbiosis. Through host switching, the clade as a whole has circumvented potential coextinction with host ant lineages that disappeared from the Cretaceous to the present (Barden and Grimaldi, 2016; Barden, 2016). Moreover, in some cases, the beetles have radiated dramatically with certain ant groups: the vast majority of the contemporary species richness of Haeteriinae is found in taxa that have adapted to colonies of Neotropical army ants (Ecitonini), including at least 30 genera associated with Eciton army ants alone (Parker, 2016; Helava et al., 1985; von Beeren and Tishechkin, 2017; Tishechkin, 2007). Some of these haeteriines have remarkable adaptations for life in colonies of those nomadic ants (Figure 4). Neotropical army ants are thought to have begun diversifying approximately in the Oligocene (Brady et al., 2014; Borowiec et al., 2017b), implying that the bulk of haeteriine cladogenesis occurred within this window too, long after the beetles originated in the Cretaceous.

Figure 4
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Diversity of modern Haeteriinae associated with Neotropical army ants (Ecitonini).

(A) Colonides beetle walks in an emigration of Eciton burchellii army ants. Note the color mimicry of the host ant (Peru; photo: Taku Shimada). (B) Euxenister beetle walking alongside an Eciton hamatum army ant worker. The long legs facilitate grooming ants to obtain the colony odor, as well as clinging to emigrating workers (Peru; photo credit: Takashi Komatsu). (C) Nymphister kronaueri beetle phoretically attached to the petiole of an Eciton mexicanum army ant worker. The beetles bite onto this part of the ant’s body, enabling them to migrate with the host. The beetle resembles the ant’s gaster from above, potentially camouflaging the beetles to avoid predation (Costa Rica; photo credit: Daniel Kronauer).

https://doi.org/10.7554/eLife.44985.011

An ancient association between histerids and ants is further suggested by inquiline-like morphology in two other Cretaceous clown beetle fossils (Caterino et al., 2015; Caterino and Maddison, 2018), although unlike Promyrmister, the taxonomic affinities of these specimens are ambiguous and they are not definitive members of wholly symbiotic lineages. We infer that Haeteriinae was a relatively diverse clade by at least the beginning of the Upper Cretaceous, and likely originated and began undergoing basal cladogenesis very soon after the inferred Early Cretaceous emergence of ant eusociality (Barden and Grimaldi, 2016; Grimaldi and Agosti, 2000; Barden, 2016; Brady et al., 2006; Moreau and Bell, 2013; Borowiec et al., 2017a). A time-calibrated molecular phylogeny of Haeteriinae may provide a more precise estimate of this temporal window. However, based on such analyses for other myrmecophile taxa, rapid evolution of specialized symbiotic phenotypes appears to be a common feature to clades of social insect symbionts (Moore and Robertson, 2014; Parker and Grimaldi, 2014), and presumably results from intense selection pressures inside colonies (Kistner, 1979; Parker, 2016). Promyrmister adds further support to the view that the earliest-known ants were socially complex (Barden and Grimaldi, 2016). Evidently, their colonies were also resource rich enough for exploitation by impostor myrmecophiles, which we conclude have been an unremitting part of ant biology. Despite their phenotypic intricacy and obligate dependency on other species, complex behavioral relationships between animals can be extraordinarily ancient, and persist over deep evolutionary time.

Materials and methods

Material and photography

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This study is based on a single specimen of Burmese amber (CNU-008021) collected from Noije Bum, Tanaing, Kachin, Myanmar. The specimen is housed at Key Laboratory of Insect Evolution and Environmental Changes, Capital Normal University, Beijing. The holotype of the new genus and species is embedded in a cuboid amber piece. The holotype was examined under a Leica M205C dissecting microscope and photographed using a Visionary Digital BK Lab Plus system (Austin, Texas). The source images were aligned and stacked in Helicon Focus (Ukraine). Fluorescence images of the fossil were made on a Zeiss LSM 880 (with Airyscan) confocal microscope (Germany) with a 488 nm laser. Scanning electron microscopic images of Haeterius were obtained using a Tabletop Hitachi Microscope TM3030Plus (Japan). Morphological terminology follows Ślipiński and Mazur (1999), Zhou et al. (2018), and Caterino and Tishechkin (2015).

Taxon sampling, morphological characters and phylogenetic analysis

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We scored Promyrmister for 259 external morphological characters used by Caterino and Tishechkin (2015) in a study investigating relationships among the tribe Exosternini, which is closely related to Haeteriinae. From the original matrix, we selected all taxa from the nearest sister clades of Haeteriinae, including 35 taxa belonging to Exosternini, including all species of Yarmister (apparently the closest genus to Haeteriinae; Caterino and Tishechkin, 2015). We also included representatives of four other tribes: Omalodini, Histerini, Hololeptini and Platysomatini, and assigned Hister unicolor as the primary outgroup, following Caterino and Tishechkin (2015). The final taxon list is presented in Supplementary file 2A.

We also enlarged our data matrix by adding one more taxon (Haeterius ferrugineus) two more characters (260 and 261), and one more state for Character 14:

260: Epistoma: (1) without depressions receiving scapes in repose, occasionally with small depressions but without sharp arched-inwards epistomal striae; (2) with large depressions receiving scapes in repose, often defined by sharp arched-inwards epistomal striae.

261: Arched-inwards epistomal striae: (1) convergent, but separated from each other in the middle; (2) convergent, and meeting each other in the middle; (3) inapplicable.

14: Epistoma, surface: 6) deeply depressed, with lateral ridges (=raised epistomal striae) aligned with frontal stria.

The complete matrix of 46 taxa, 261 characters was constructed in Mesquite v. 3.20 (Maddison and Maddison, 2016); the matrix is provided in the nexus file (Supplementary file 1). Bayesian analysis was carried out using MrBayes 3.2.6 (Ronquist et al., 2012) accessed via the CIPRES Science Gateway Version 3 (Miller et al., 2010) (phylo.org). The Mkv model of character evolution was used with a gamma distribution, and two MCMC were executed with four chains for 100 million generations. Convergence was judged to have occurred when the standard deviation of split frequencies dropped below 0.005, and by ESS values higher than 200 in Tracer v1.7.0 (Rambaut et al., 2018), indicating adequate estimation of the posterior. The first 25% of trees were discarded as burn-in. We used Treeannotator (Bouckaert et al., 2014) to obtain the maximum clade credibility tree from post burn-in trees (ESS > 200) (Figure 2A), and added the estimated nodal Bayesian posterior probability (BPP) in FigTree v1.4.3 (https://github.com/rambaut/figtree/). Parsimony analysis was conducted in TNT Version 1.5 (Goloboff and Catalano, 2016) using Traditional Search without, and with implied weighting setting (function K = 13 in Figure 2—figure supplements 2 and 3). A consensus tree (Figure 2B; L = 1604, CI = 25, RI = 42) was obtained from four shortest-length trees (L = 1483, CI = 28, RI = 48) and the branch support was also calculated using 10,000 bootstrap replicates. Mapping character state changes onto the tree was performed in WinClada (Nixon, 2002).

Host ant ages

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A list of haeteriine host ant genera was obtained from the literature (Helava et al., 1985; Yélamos, 1997; Tishechkin, 2007; Lapeva-Gjonova, 2013; Mazur, 1981). To estimate ages of stem-group and Recent host ant taxa in Figure 3a, data for earliest-known fossils were obtained from Barden (2016) (Barden, 2016; Barden, 2018), and molecular age estimates of crown-groups were taken from recent taxon-specific phylogenetic studies (Borowiec et al., 2017a; Borowiec et al., 2017b; Blaimer et al., 2015; Ward et al., 2015; Ferguson-Gow et al., 2014; Ward et al., 2010; Schmidt, 2013). Data are presented in Supplementary file 2B.

Nomenclatural acts

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This published work and the nomenclatural acts it contains have been registered in ZooBank, the online registration system for the International Code of Zoological Nomenclature. The ZooBank LSIDs (Life Science Identifiers) can be resolved and the associated information viewed through any standard web browser by appending the LSID to the prefix ‘http://zoobank.org/’. The LSIDs for this publication are to be found at:

  • urn:lsid:zoobank.org:pub:4AE2E535-B2B7-4A9A-829F-FA17CB98AD9C.

The specific LSIDs for new nomenclatural acts:

  • Genus: urn:lsid:zoobank.org:act:8125C3AB-A6C3-41AF-A9D7-0B67BBA2ACAD

  • Species: urn:lsid:zoobank.org:act:56DE873C-0163–4 F94-8D09-196F20B84C57

Data availability

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All data generated or analyzed during this study are included in this published article (and its Supplementary Information files). The holotype specimen of Promyrmister kistneri is housed at Key Laboratory of Insect Evolution and Environmental Changes, Capital Normal University, Beijing (accession number CNU-008021).

Data availability

All data generated or analyzed during this study are included in the manuscript and supporting files. Source data for Figure 2, Figure 2 figure supplement 2 and Figure 2 figure supplement 3 are provided in Supplementary File 1.

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Decision letter

  1. John A Long
    Reviewing Editor; Flinders University, Australia
  2. Diethard Tautz
    Senior Editor; Max-Planck Institute for Evolutionary Biology, Germany
  3. Margaret Thayer
    Reviewer; Field Museum, United States

In the interests of transparency, eLife includes the editorial decision letter and accompanying author responses. A lightly edited version of the letter sent to the authors after peer review is shown, indicating the most substantive concerns; minor comments are not usually included.

[Editors’ note: a previous version of this study was rejected after peer review, but the authors submitted for reconsideration. The first decision letter after peer review is shown below.]

Thank you for submitting your work entitled "A Mesozoic Clown Beetle Myrmecophile" for consideration by eLife. Your article has been reviewed by three peer reviewers, one of whom is a member of our Board of Reviewing Editors, and the evaluation has been overseen by a Senior Editor. The following individual involved in review of your submission has agreed to reveal their identity: Margaret Thayer (Reviewer #3).

Our decision has been reached after consultation between the reviewers. Based on these discussions and the individual reviews below, we regret to inform you that your work will not be considered further for publication in eLife.

The paper is a significant contribution to our understanding of the evolution of social relationships of these insect groups in its findings, and is generally well-written and well-illustrated. However some of the findings are inferred and stated very strongly, as highlighted by reviewer 2, and there are also some major issues raised by reviewer 3, concerning the diagnostic characters of the new genus, and the characters used in phylogenetic analysis.

Reviewer #1:

This is a well-written and beautifully illustrated paper describing a new taxon of fossil clown beetle Mymrecophile from the Burmese amber lagerstatte of Late Cretaceous age. The significance of the discovery lies in the fact that clown beetles are nest intruders into ant colonies, so this fossil gives a lower end date for the beginning of this type of complex inter-social adaptation in these two insect groups. From the fossil evidence the authors deduce that such obligate relationships must have evolved rapidly, and began earlier in the Cretaceous.

The perfect 3D preservation of the fossil beetle allows it to be describe in detail, and shows clear taxonomic features so taxonomy is not disputed here. It is so rare in the fossil record to get this kind of preservation that the vast amount of information coming from the Burmese amber insects makes it difficult to determine precisely when such complex social behaviours first originated (we would need other amber lagerstatten in the Jurassic and /or the Triassic to pin-point such events).

I expect the arthropod taxonomists to offer more detailed comments about the taxonomy etc. My only comments for the authors to consider are minor ones:

Results and Discussion: 'mid-Cretaceous' as anon formal date is here confusing as the age of the deposit make sit Cenomanian (technically "Upper Cretaceous"), and absolute age place sits in the top 1/3 of the Cretaceous age span, so Upper cretaceous is more preferable than using' mid-Cretaceous'.

Please consider my comments above about the 'lagerstate effect' -such well-preserved fossils can suggest biases in the fossil record, so estimating dates of origin of social behaviour may rest upon a combination of molecular dates of when each group might first evolved calibrated with the dated fossil record.

Reviewer #2:

This study reports a myrmecophilous beetle of the histerid subfamily Haeteriinae from ca. 100 million year old amber from Myanmar. This fossil is of particular interest, because it coincides with early ant evolution, suggesting that social parasitism in histerids evolved early on and has persisted until today. The manuscript is extremely well written and beautifully illustrated. However, I couldn't escape the impression that the discovery is being oversold.

1) Throughout the manuscript, the authors emphasize the point that the described beetle must have been a symbiont of stem-group ants, i.e. ants that are represented in the fossil record but only distantly related to extant ants. Their main argument is that extant crown-group ants are not represented in Burmese amber. They also argue that molecular dating analyses suggest that extant host taxa of Haeteriinae are of more recent age. But I think there are a few issues with this argument. First, the fossil record is incomplete, and absence of extant ants in the fossil record therefore doesn't mean that they weren't there. Second, there is in fact at least one record of crown-group ants in Burmese amber (the extant subfamily Aneuretinae), showing that stem-group and crown-group ants coexisted during that time (see e.g. Barden, 2017, cited by the authors). Furthermore, the most recent estimates based on molecular data date the origin of crown-group ants to ca. 103-124 MYA, i.e. several million years prior to the Burmese amber deposits (Borowiec et al., 2017; not cited by the authors). This all suggests that crown-group ants in fact co-occurred with the described beetle. The evolutionary scenario described (subsection “Sphecomyrmister and deep time persistence of a social symbiosis", fourth paragraph) posits that these beetles originally evolved with stem-group ants and then switched to crown-group ants later. But there is really no evidence for this. In fact, from a purely parsimony principle point of view, it seems more parsimonious that the beetles evolved to parasitize early crown-group ants, and that they can still be found associated with a subset of lineages today. If you allow for host switching anyway, then the fact that extant hosts of Haeteriinae are of younger age than the fossil beetle (Figure 3) also doesn't strengthen the argument that they would have originally come from stem-group ants. Unfortunately, the stem-group ant angle is the main selling point of the manuscript – it's reflected in Figure 2F (which I find misleading) and even the proposed genus name of the beetle (Sphecomyrmister), and it takes up large proportions of the Introduction and Discussion. But I would say that the support for this is rather weak.

2) A couple of other recent papers have hinted at associations between histerids and ants of similar age (Caterino, Wolf-Schwenninger and Bechly, 2015, and Maddison and Caterino, 2008; discussed in the last paragraph of the subsection “Sphecomyrmister and deep time persistence of a social symbiosis”), and I think that takes away from the novelty of the current discovery. Beetle fossils could in principle be associated with ants in three ways I guess. The most convincing would be inclusions that contain both the beetles and the host ants which, as far as I know, have not been found. The second most compelling evidence comes from morphological adaptations that are restricted to inquilines – this is the case both for this fossil and the previous fossils. The third and, by itself, arguably most circumstantial is phylogenetic placement of the fossil. The fossil described here can be phylogenetically placed within a group of obligately myrmecophilous beetles, which of course is very nice. But I don't think that the evidence for myrmecophily is so much stronger here compared to the previous fossils. So even though I must admit that the packaging is much more compelling here, I therefore fail to see the major advancement in our knowledge of myrmecophile evolution.

Reviewer #3:

The new fossil taxon described in this paper represents an exciting addition to our knowledge of both the beetle family Histeridae and, more broadly, evolution of myrmecophily in beetles. The authors describe and illustrate the new fossil well and provide convincing justification for placing the new taxon within the haeteriine Histeridae. Since all known extant Haeteriinae are obligate myrmecophiles, the morphology-based placement supports the inference that Sphecomyrmister was also a myrmecophile, although in a few places the authors tend toward presenting this as a fact rather than an inference.

Nevertheless, I have some concerns about the phylogenetic analysis. Although the analysis itself was carried out in reasonable fashion, simple adoption of a data matrix from a paper (Caterino and Tishechkin, 2015) that focused on a different part of the Histeridae (Histerinae: Exosternini) is problematic. The original matrix included a few representatives of Haeteriinae, along with hundreds of other taxa belonging to the focal group of that paper and an assortment of other Histeridae as outgroups. The taxon sub-sampling in the current manuscript from the larger Caterino and Tishechkin, 2015 matrix seems reasonable at first glance, but it is not clear how or why they chose particular Exosternini genera (aside from Yarmister), and on further consideration I am concerned by two fundamental aspects of their analysis.

First, although understanding the relationships of Sphecomyrmister within Haeteriinae is presumably a major focus of the paper, the authors did not add to the matrix any characters that might be suitable or necessary for resolving those relationships. Especially considering the specialized morphology of Haeteriinae, surely there are relevant characters that were not included in Caterino and Tishechkin, 2015's analysis of other Histeridae, such as those cited in the current manuscript to (in combination) separate Sphecomyrmister from all other Haeteriinae. (I admit I did not examine Caterino and Tishechkin, 2015's long character list in detail to search for those.)

Second, the authors did not include representatives of any of the three largest and most heavily sampled genera in Caterino and Tishechkin, 2015's paper (Phelister, Operclipygus, and Baconia), and for some reason all but one of the Exosternini genera included are Old World rather than New World taxa (a point not mentioned), even though the focus of Caterino and Tishechkin, 2015's paper was Neotropical Exosternini and nearly all Haeteriinae are Neotropical. Although selecting single species from the large genera might have been challenging (there appears to be substantial variation within each genus), I found that completely leaving them out led to partly spurious results. Specifically, Figure 2—figure supplement 2 seems to show as unique apomorphies for Sphecomyrmister (I think-the character state numbers are blurry) the states 12-2, 112-2, 113-2, and 114-2; this is puzzling since the (new) genus obviously was not included in Caterino and Tishechkin, 2015's data but the characters were. For the latter three characters, Caterino and Tishechkin, 2015, said the modified gland openings involved occur only in some species groups of Operclipygus (Exosternini). Clearly, then, they are not globally unique to Sphecomyrmister, but appear as such in the present analysis because no Operclipygus were included! Similarly, Caterino and Tishechkin, 2015, illustrated state 12-2 with figures of two species of Baconia (Exosternini), so again this is not truly a unique apomorphy of Sphecomyrmister. Figure 2—figure supplement 2 does not show any unique apomorphies for Haeterius, so reciprocal monophyly of that and Sphecomyrmister do not seem to be supported. Mirroring that problem in the text, although the authors list a combination of characters to separate Sphecomyrmister from all other Haeteriinae, and others to show its close relationship to Haeterius, I could not find a clear statement of what separates those two genera. This is unsatisfactory from two standpoints: 1) compliance with the [ICZN] Code requirement for a statement purporting to distinguish the new taxon and 2) supporting the authors' evolutionary contention that Sphecomyrmister represents an extinct Cretaceous lineage.

https://doi.org/10.7554/eLife.44985.016

Author response

[Editors’ note: the author responses to the first round of peer review follow.]

Thank you so much for your excellent feedback on our paper. We have taken every effort to improve the new version based on your constructive comments. We have briefly summarized your reviews, and our responses to them:

Reviewer 1 recognizes the novelty and valuable contribution of our paper. Reviewer 1 notes that due to the incompleteness of the fossil record, it is “difficult to determine precisely when such complex social behaviours first originated”. Our fossil provides a minimum age, but future molecular dating of groups like Haeteriinae will be illuminating. We agree, and have included this suggestion in the revised manuscript, along with citations to papers that have done exactly this for other myrmecophile clades.

Reviewer 2 agrees that our fossil discovery is “of particular interest, because it coincides with early ant evolution” but questions whether stem-group ants were likely hosts. In the revised paper, we present our original scenario together with its tantalizing alternative: that an as-yet undiscovered diversity of crown-group ants could have lived in the Burmese palaeofauna, and been potential hosts. Unlike reviewers 1 and 3, reviewer 2 also argues that previously-described ‘myrmecophile-looking’ histerids reduce our paper’s novelty. We respectfully disagree: prior speculation cannot match the discovery of hard evidence. Our paper uncovers the earliest definitive evidence of Cretaceous myrmecophily, contemporaneous with the earliest-known eusocial ants.

Reviewer 3 recognizes the novelty and valuable contribution of our paper. Reviewer 3 has thoroughly studied our phylogenetic analysis and raised several legitimate criticisms. In the revised paper, we have addressed these criticisms for a more solid phylogenetic analysis and taxonomic placement of the fossil. We have also attended to numerous minor errors that reviewer 3 highlighted.

The paper is a significant contribution to our understanding of the evolution of social relationships of these insect groups in its findings, and is generally well-written and well-illustrated. However some of the findings are inferred and stated very strongly, as highlighted by reviewer 2, and there are also some major issues raised by reviewer 3, concerning the diagnostic characters of the new genus, and the characters used in phylogenetic analysis.

Reviewer #1:

[…] The perfect 3D preservation of the fossil beetle allows it to be describe in detail, and shows clear taxonomic features so taxonomy is not disputed here. It is so rare in the fossil record to get this kind of preservation that the vast amount of information coming from the Burmese amber insects makes it difficult to determine precisely when such complex social behaviours first originated (we would need other amber lagerstatten in the Jurassic and /or the Triassic to pin-point such events).

Such well-preserved fossils can suggest biases in the fossil record, so estimating dates of origin of social behaviour may rest upon a combination of molecular dates of when each group might first evolved calibrated with the dated fossil record.

Thank you for your wonderful comments. We agree with your major point that inferring more precisely when myrmecophily in haeteriines evolved is still murky. Our fossil implies the Early Cretaceous at the latest, but molecular dating of a comprehensive phylogeny of these beetles would provide more resolution, or at least a more precise inference. Such efforts have been used for certain clades of myrmecophile staphylinids, as well as paussine carabids. In the new paper, we make mention of this, and argue that creating a dated phylogeny of haeteriinae, (and more broadly, Histeridae) is particularly timely given this discovery of our fossil and its implications for symbiotic relationships of early ants.

Reviewer #2:

This study reports a myrmecophilous beetle of the histerid subfamily Haeteriinae from ca. 100 million year old amber from Myanmar. This fossil is of particular interest, because it coincides with early ant evolution, suggesting that social parasitism in histerids evolved early on and has persisted until today. The manuscript is extremely well written and beautifully illustrated. However, I couldn't escape the impression that the discovery is being oversold.

1) Throughout the manuscript, the authors emphasize the point that the described beetle must have been a symbiont of stem-group ants, i.e. ants that are represented in the fossil record but only distantly related to extant ants. Their main argument is that extant crown-group ants are not represented in Burmese amber. They also argue that molecular dating analyses suggest that extant host taxa of Haeteriinae are of more recent age. But I think there are a few issues with this argument. First, the fossil record is incomplete, and absence of extant ants in the fossil record therefore doesn't mean that they weren't there. Second, there is in fact at least one record of crown-group ants in Burmese amber (the extant subfamily Aneuretinae), showing that stem-group and crown-group ants coexisted during that time (see e.g. Barden, 2017, cited by the authors). Furthermore, the most recent estimates based on molecular data date the origin of crown-group ants to ca. 103-124 MYA, i.e. several million years prior to the Burmese amber deposits (Borowiec et al., 2017; not cited by the authors). This all suggests that crown-group ants in fact co-occurred with the described beetle. The evolutionary scenario described (subsection “Sphecomyrmister and deep time persistence of a social symbiosis", fourth paragraph) posits that these beetles originally evolved with stem-group ants and then switched to crown-group ants later. But there is really no evidence for this. In fact, from a purely parsimony principle point of view, it seems more parsimonious that the beetles evolved to parasitize early crown-group ants, and that they can still be found associated with a subset of lineages today. If you allow for host switching anyway, then the fact that extant hosts of Haeteriinae are of younger age than the fossil beetle (Figure 3) also doesn't strengthen the argument that they would have originally come from stem-group ants.

Thank you for raising this point. In our original manuscript we felt compelled to point out that stem-group subfamilies seemed the most likely hosts, because these are the only ones known for definite in Burmese amber (see separate comment below about the aneuretine). We are aware that molecular dating studies imply that crown-group ants may have existed at this time, but because they are simply not known in thousands of ant inclusions in Burmese amber, we felt it highly unlikely that their myrmecophiles would make an appearance but they would not.

However, we acknowledge that the likelihood of whether stem- or crown-group ants were the beetle’s hosts could be judged equivocal, depending on one’s preference for data type. In our view, the overwhelming predominance of stem-group ant fossils in Burmese amber would imply that these were the hosts – it is not a stretch to think that stem group ant colonies were resource rich enough to select for the initial evolution of myrmecophily. Conversely, based on reviewer 2’s entirely reasonable suggestion, molecular dating implies that crown-group ants had evolved by 99MYA, so could also legitimately be hosts. In the revised paper, we present both hypotheses as valid, although convey our preference for the former scenario, which we think is not unreasonable based on the lack of crown-group ant subfamilies in Burmite. Accordingly, we have modified the taxon’s name to accommodate both hypotheses (we have adopted Promyrmister – “early ant hister”). We have kept Figure 3, but modified it with molecular dates from Borowiec et al., 2017, and now use this figure to explain the two alternative hypotheses. Modifying the paper in this way does not detract from the impact or novelty of the story – quite the opposite – it raises two, equally fascinating possibilities: either stem-group ants were the beetle’s hosts, or the beetle alludes to an as-yet undiscovered diversity of crown-group ants in the Burmese palaeofauna.

Second, there is in fact at least one record of crown-group ants in Burmese amber (the extant subfamily Aneuretinae), showing that stem-group and crown-group ants coexisted during that time (see e.g. Barden, 2017, cited by the authors).

Please note that the identity of this specimen, Burmomyrma rossi, is highly contentious. Even Dlussky, the taxon’s author, expressed doubt that this specimen was an aneuretine. According to Barden, 2017:

“Because the sole specimen (an alate female) is missing the entire head and portions of the mesosoma, Dlussky, 1996, was initially equivocal in his assignment of Burmomyrma to Aneuretinae, stating the “systematic position could not be determined reliably due to poor preservation of the only specimen known”. Tentative aneuretine placement was based on highly reduced forewing venation, curved sting, a single segmented petiole, and a gaster without constrictions. This particular assortment of characters cannot be used to assign Burmomyrma rossi to any subfamily with confidence, particularly as the wing venation described is not shared by other known aneuretines (Boudinot, 2015).”

Moreover, a recent work concluded that Burmomyrma rossi is probably not an ant at all, but a chrysidid wasp (See: Lucena, and Melo, Cretaceous Research 89, 279–291). Consequently, there are no definitive crown-group ants in Burmese amber.

2) A couple of other recent papers have hinted at associations between histerids and ants of similar age (Caterino, Wolf-Schwenninger and Bechly, 2015 and Maddison and Caterino, 2008; discussed in the last paragraph of the subsection “Sphecomyrmister and deep time persistence of a social symbiosis”), and I think that takes away from the novelty of the current discovery. Beetle fossils could in principle be associated with ants in three ways I guess. The most convincing would be inclusions that contain both the beetles and the host ants which, as far as I know, have not been found. The second most compelling evidence comes from morphological adaptations that are restricted to inquilines – this is the case both for this fossil and the previous fossils. The third and, by itself, arguably most circumstantial is phylogenetic placement of the fossil. The fossil described here can be phylogenetically placed within a group of obligately myrmecophilous beetles, which of course is very nice. But I don't think that the evidence for myrmecophily is so much stronger here compared to the previous fossils. So even though I must admit that the packaging is much more compelling here, I therefore fail to see the major advancement in our knowledge of myrmecophile evolution.

We respectfully disagree. Previously-described fossil histerids with myrmecophile-like morphology cannot compete with the impact of our story. Such a suggestion equates the discovery of scientific evidence with the mere speculation that preceded the discovery. Morphology alone does not meet the burden of proof for myrmecophily – there are numerous extant insect groups that have a somewhat myrmecophile-like appearance but are not myrmecophiles (e.g. in Carabidae: the subfamily Rhysodinae and the scaritine genera Solenogenys and Salcedia; in Staphylinidae: Falagriini such as Myrmecocephalus, many Pselaphinae such as Brachygluta abdominalis). In contrast, a species with myrmecophilous adaptations that additionally belongs to a clade composed entirely of obligate myrmecophiles leaves little doubt.

Prior to our paper, definitive myrmecophiles were unknown from the entire Cretaceous, let alone from the same deposit as the earliest-known ants. The literature is sprinkled with speculative examples of Cretaceous insects claimed to have morphology suggestive of myrmecophily or termitophily (e.g. a supposed myrmecophiline cricket in the Crato Formation (Martins-Neto, 1991; Parker and Grimaldi, 2014); a Lebanese amber scarabaeoid (Crowson, 1981)). The two histerids mentioned are but two more such examples. Yet, the whole notion of Cretaceous myrmecophily, and early ant colony infiltration, has remained entirely speculative due to lack of any hard evidence in the form of a specimen belonging to wholly myrmecophilous group.

Our paper now uncovers a bona fide, anatomically specialized myrmecophile from the crown-group of a modern clade composed entirely of such creatures. The fossil satisfies the essential burden of proof of myrmecophily. That such an organism could have existed in the earliest-known ant societies in the Cretaceous is remarkable. The fossil is a member of one of the largest-known modern radiations of myrmecophiles, illuminating the deep ancestry of the social symbiosis between clown beetles and ants. The implications are broad for the evolution of social interactions, early colony formation in ants, and symbiotic relationships between animals more generally. Our paper is thus of special value compared to the speculative works that preceded it.

Crowson, R. A. (1981). The Biology of the Coleoptera. London: Academic Press.

Martins-Neto, R. G. (1991). Sistemática dos Ensifera (Insecta, Orthopteroida) da formação Santana, Cretáceo Inferior do Nordeste do Brasil. Acta Geologica Leopoldensia 32, 5–160.

Reviewer #3:

The new fossil taxon described in this paper represents an exciting addition to our knowledge of both the beetle family Histeridae and, more broadly, evolution of myrmecophily in beetles. The authors describe and illustrate the new fossil well and provide convincing justification for placing the new taxon within the haeteriine Histeridae. Since all known extant Haeteriinae are obligate myrmecophiles, the morphology-based placement supports the inference that Sphecomyrmister was also a myrmecophile, although in a few places the authors tend toward presenting this as a fact rather than an inference.

In the new paper we have changed the language in places to avoid appearing to do this.

Nevertheless, I have some concerns about the phylogenetic analysis. Although the analysis itself was carried out in reasonable fashion, simple adoption of a data matrix from a paper (Caterino and Tishechkin 2015) that focused on a different part of the Histeridae (Histerinae: Exosternini) is problematic. The original matrix included a few representatives of Haeteriinae, along with hundreds of other taxa belonging to the focal group of that paper and an assortment of other Histeridae as outgroups. The taxon sub-sampling in the current manuscript from the larger Caterino and Tishechkin, 2015 matrix seems reasonable at first glance, but it is not clear how or why they chose particular Exosternini genera (aside from Yarmister), and on further consideration I am concerned by two fundamental aspects of their analysis.

We believe the adoption of the Caterino and Tishechkin, 2015 data matrix was reasonable and justifiable. The most recent molecular phylogenies of Histeridae (Caterino and Vogler, 2002, McKenna et al., 2015) have recovered Haeteriinae as a sister group to Histerinae. Both Caterino and Tishechkin, 2015, and McKenna et al., 2015, recovered a monophyletic Haeteriinae, which is represented in our taxon sampling by genera belonging to all three recognized tribes of this subfamily. Our a priori assessment of the fossil was that it was a haeteriine, so we evaluated this placement phylogenetically, adopting the Caterino and Tishechkin, 2015 morphological data set due to its empirically demonstrated ability to resolve Haeteriinae monophyly. The original taxon sampling in Caterino and Tishechkin, 2015 is vast and exceeded what we needed for our intended purpose. So, the haeteriines and a subset of outgroups were used. The taxonomic scope of our analysis was, we felt, sufficient for just seeing whether the fossil was recovered in Haeteriinae, which our a priori assessment had implied. However, we agree that a more objective judgment of the placement of the fossil taxon could be achieved, so we have performed a revised analysis. We scored one more Haeterius species ourselves, and included at least one species (type species of the genus if available) of the genera of Exosternini belonging to the sister clades of Haeteriinae, and added one species of Baconia, three species of Operclipygus, all species of Yarmister, and New and Old World representatives of Hypobletus from Caterino and Tishechkin, 2015. We believe this represents a sufficient sampling of histerid outgroups for the sole purpose of testing the fossil’s placement within the Haeteriinae clade.

First, although understanding the relationships of Sphecomyrmister within Haeteriinae is presumably a major focus of the paper, the authors did not add to the matrix any characters that might be suitable or necessary for resolving those relationships. Especially considering the specialized morphology of Haeteriinae, surely there are relevant characters that were not included in Caterino and Tishechkin, 2015's analysis of other Histeridae, such as those cited in the current manuscript to (in combination) separate Sphecomyrmister from all other Haeteriinae. (I admit I did not examine Caterino and Tishechkin, 2015's long character list in detail to search for those.)

The morphological treatment in the Caterino and Tishechkin, 2015 paper is amazingly detailed and comprehensive. It is hard to find structures or features of adult beetles that are not covered there. Most of the characters from the Caterino and Tishechkin, 2015 paper can be observed and coded from our amber fossil, and the character set was evidently comprehensive enough to resolve a monophyletic Haeteriinae, both in our analysis in the original Caterino and Tishechkin, 2015 paper. Furthermore, to address your comment more precisely, while some haeteriines are extremely anatomically specialized for myrmecophily, many are not so dramatically modified. For example, the external morphology of many Exosternini and Haeteriinae is very similar, which has led to misplacement of several genera in the past (e.g., Kaszabister, Yarmister, Tarsilister). This means that potential new, haeteriine-relevant characters are quite hard to find. Despite this, we have found and added two more characters (no. 260, 261) and one more state (6) to character 14 to distinguish Haeteriinae from outgroups. We hope this addresses your comment adequately.

Second, the authors did not include representatives of any of the three largest and most heavily sampled genera in Caterino and Tishechkin, 2015's paper (Phelister, Operclipygus, and Baconia), and for some reason all but one of the Exosternini genera included are Old World rather than New World taxa (a point not mentioned), even though the focus of Caterino and Tishechkin, 2015's paper was Neotropical Exosternini and nearly all Haeteriinae are Neotropical. Although selecting single species from the large genera might have been challenging (there appears to be substantial variation within each genus), I found that completely leaving them out led to partly spurious results.

As mentioned above, in the revised analysis we have included at least one species (type species of the genus if available) of the genera of Exosternini belonging to the sister clades of Haeteriinae, and added several species of the variable genera Operclipygus and Yarmister, as well as New and Old World representatives of Hypobletus, and one species of Baconia examined by Caterino and Tishechkin, 2015. For a more robust representation of the putuative sister genus of Promyrmister, we have added a Palaearctic species of Haeterius (H. ferrugineus) the type species of the genus, and scored it for all the characters.

Specifically, Figure 2—figure supplement 2 seems to show as unique apomorphies for Sphecomyrmister (I think-the character state numbers are blurry) the states 12-2, 112-2, 113-2, and 114-2; this is puzzling since the (new) genus obviously was not included in Caterino and Tishechkin, 2015's data but the characters were. For the latter three characters, Caterino and Tishechkin, 2015, said the modified gland openings involved occur only in some species groups of Operclipygus (Exosternini). Clearly, then, they are not globally unique to Sphecomyrmister, but appear as such in the present analysis because no Operclipygus were included! Similarly, Caterino and Tishechkin, 2015, illustrated state 12-2 with figures of two species of Baconia (Exosternini), so again this is not truly a unique apomorphy of Sphecomyrmister. Figure 2—figure supplement 2 does not show any unique apomorphies for Haeterius, so reciprocal monophyly of that and Sphecomyrmister do not seem to be supported.

We apologize, but the specific apomorphies shown in Figure 2—figure supplement 2, were not easily visible, e.g. (112-2) and (113-2) and were misread by the reviewer because of the low quality of the file. We have supplied a higher resolution image of this tree in the revised paper. To be clear, currently fifteen unambiguous apomophies have been found for Promyrmister: 2-2, 5-2, 11-3, 12-2, 13-2, 14-6, 56-4, 109-2, 111-3, 112-3, 112-3, 124-1, 125-1, 135-2, and 261-2. Most of them appear as homoplasious states but two of them: (14-6) (epistoma, surface deeply depressed, with epistomal striae joined basally with frontal stria), and (261-2) epistomal striae anteriorly arching-inwards, meeting each other at the middle, only appear in Promyrmister.

Mirroring that problem in the text, although the authors list a combination of characters to separate Sphecomyrmister from all other Haeteriinae, and others to show its close relationship to Haeterius, I could not find a clear statement of what separates those two genera. This is unsatisfactory from two standpoints: 1) compliance with the [ICZN] Code requirement for a statement purporting to distinguish the new taxon and 2) supporting the authors' evolutionary contention that Sphecomyrmister represents an extinct Cretaceous lineage.

We have now clarified in the diagnosis section that Promyrmister specifically differs from the putatively closely related Haeterius in having deep epistomal depressions (compare Figure 1—figure supplement 1B, C to Figure 2—figure supplement 1B), epistomal striae carinate and convergent medially (Figure 1—figure supplement 1B, C) and paddle-shaped protibia with large apical spur (Figure 1E, F). See Results and Discussion for a complete description of the new genus and species, as well as a discussion of the taxon’s systematic placement in Histeridae.

https://doi.org/10.7554/eLife.44985.017

Article and author information

Author details

  1. Yu-Lingzi Zhou

    1. Key Laboratory of Zoological Systematics and Evolution, Institute of Zoology, Chinese Academy of Sciences, Beijing, China
    2. Australian National Insect Collection, CSIRO, Canberra, Australia
    Contribution
    Conceptualization, Data curation, Formal analysis, Investigation, Visualization, Methodology, Writing—original draft, Writing—review and editing
    Competing interests
    No competing interests declared

  2. Adam Ślipiński

    Australian National Insect Collection, CSIRO, Canberra, Australia
    Contribution
    Conceptualization, Resources, Data curation, Formal analysis, Funding acquisition, Investigation, Visualization, Methodology, Writing—original draft, Project administration, Writing—review and editing
    Competing interests
    No competing interests declared

  3. Dong Ren

    College of Life Sciences, Capital Normal University, Beijing, China
    Contribution
    Resources, Funding acquisition, Project administration
    Competing interests
    No competing interests declared

  4. Joseph Parker

    Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, United States
    Contribution
    Conceptualization, Resources, Data curation, Formal analysis, Validation, Investigation, Visualization, Methodology, Writing—original draft, Project administration, Writing—review and editing
    For correspondence
    joep@caltech.edu
    Competing interests
    No competing interests declared
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-9598-2454

Funding

National Natural Science Foundation of China (31402008)

  • Yu-Lingzi Zhou

International Postdoctoral Exchange Fellowship (20150064)

  • Yu-Lingzi Zhou

Rita Allen Foundation

  • Joseph Parker

Esther A. and Joseph Klingenstein Fund

  • Joseph Parker

Shurl & Kay Curci Foundation

  • Joseph Parker

Simons Foundation

  • Joseph Parker

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Acknowledgements

We are grateful to Alexey K Tishechkin (USDA), Michael S Caterino (Clemson University) and Alfred Newton (Field Museum) for their helpful advice on the placement of the fossil and to Margaret K Thayer (Field Museum) for a very thorough review of the paper. Phil Barden (New Jersey Insitute of Technology) provided invaluable insight into the ant fossil record and possible hosts of Promyrmister. This research was supported by a Shurl and Kay Curci Foundation Research Grant, a Rita Allen Scholars Award and a Klingenstein-Simons Fellowship Award in the Neurosciences to JP, and the National Natural Science Foundation of China Grant no. 31402008 and International Postdoctoral Exchange Fellowship no. 20150064 to YLZ. Rolf Oberprieler (ANIC) and Hong Pang (Sun Yat-Sen University) helped with the fossil preparation, Lauren Ashman (ANIC) provided advice on improving the manuscript and Cate Lemann (CSIRO) provided technical assistance.

Senior Editor

  1. Diethard Tautz, Max-Planck Institute for Evolutionary Biology, Germany

Reviewing Editor

  1. John A Long, Flinders University, Australia

Reviewer

  1. Margaret Thayer, Field Museum, United States

Publication history

  1. Received: January 9, 2019
  2. Accepted: March 11, 2019
  3. Version of Record published: April 16, 2019 (version 1)

Copyright

© 2019, Zhou et al.

This article is distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use and redistribution provided that the original author and source are credited.

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  1. Yu-Lingzi Zhou
  2. Adam Ślipiński
  3. Dong Ren
  4. Joseph Parker
(2019)
A Mesozoic clown beetle myrmecophile (Coleoptera: Histeridae)
eLife 8:e44985.
https://doi.org/10.7554/eLife.44985

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