Phylogenomic and mitogenomic data can accelerate inventorying of tropical beetles during the current biodiversity crisis.

Conservation efforts must be evidence-based, so rapid and economically feasible methods should be used to quantify diversity and distribution patterns. We have attempted to overcome current impediments to the gathering of biodiversity data by using integrative phylogenomic and three mtDNA fragment analyses. As a model, we sequenced the Metriorrhynchini beetle fauna, sampled from ~700 localities in three continents. The species-rich dataset included ~6500 terminals, ~ 1850 putative species delimited at 5% uncorrected pairwise threshold, possibly ~1000 of them unknown to science. Neither type of data could alone answer our questions on biodiversity and phylogeny. The phylogenomic backbone enabled the integrative delimitation of robustly defined natural genus-group units that will inform future research. Using constrained mtDNA analysis, we identified the spatial structure of species diversity, very high species-level endemism, and a biodiversity hotspot in New Guinea. We suggest that focused field research and subsequent laboratory and bioinformatic workflow steps would substantially accelerate the inventorying of any hyperdiverse tropical group with several thousand species. The outcome would be a scaffold for the incorporation of further data from environmental sequencing and ecological studies. The database of sequences could set a benchmark for the spatiotemporal evaluation of biodiversity, would support evidence-based conservation planning, and would provide a robust framework for systematic, biogeographic, and evolutionary studies.

Conspicuousness, phylogenetic structure, and origins of Müllerian mimicry in 4000 lycid beetles from all zoogeographic regions.

Biologists have reported on the chemical defences and the phenetic similarity of net-winged beetles (Coleoptera: Lycidae) and their co-mimics. Nevertheless, our knowledge has remained fragmental, and the evolution of mimetic patterns has not been studied in the phylogenetic context. We illustrate the general appearance of ~ 600 lycid species and ~ 200 co-mimics and their distribution. Further, we assemble the phylogeny using the transcriptomic backbone and ~ 570 species. Using phylogenetic information, we closely scrutinise the relationships among aposematically coloured species, the worldwide diversity, and the distribution of aposematic patterns. The emitted visual signals differ in conspicuousness. The uniform coloured dorsum is ancestral and was followed by the evolution of bicoloured forms. The mottled patterns, i.e. fasciate, striate, punctate, and reticulate, originated later in the course of evolution. The highest number of sympatrically occurring patterns was recovered in New Guinea and the Andean mountain ecosystems (the areas of the highest abundance), and in continental South East Asia (an area of moderate abundance but high in phylogenetic diversity). Consequently, a large number of co-existing aposematic patterns in a single region and/or locality is the rule, in contrast with the theoretical prediction, and predators do not face a simple model-like choice but cope with complex mimetic communities. Lycids display an ancestral aposematic signal even though they sympatrically occur with differently coloured unprofitable relatives. We show that the highly conspicuous patterns evolve within communities predominantly formed by less conspicuous Müllerian mimics and, and often only a single species displays a novel pattern. Our work is a forerunner to the detailed research into the aposematic signalling of net-winged beetles.

Interactions in multi-pattern Müllerian communities support origins of new patterns, false structures, imperfect resemblance and mimetic sexual dimorphism.

Mimicry is a hot spot of evolutionary research, but de novo origins of aposematic patterns, the persistence of multiple patterns in Müllerian communities, and the persistence of imperfect mimics still need to be investigated. Local mimetic assemblages can contain up to a hundred of species, their structure can be a result of multiple dispersal events, and the gradual build-up of the communities. Here, we investigate the structure of lowland and mountain mimetic communities of net-winged beetles by sampling the Crocker Range in north-eastern Borneo and neighbouring regions. The local endemics evolved from the Bornean lowland fauna which is highly endemic at the species level. We inferred that metriorrhynchine net-winged beetles evolved in high elevations yellow/black and reticulate aposematic high-contrast signals from a widespread low-contrast brown/black pattern. As the mountain range is ~ 6 million years old, and these patterns do not occur elsewhere, we assume their in situ origins. We demonstrate that a signal with increased internal contrast can evolve de novo in a mimetic community and can persist despite its low frequency. Additionally, a similar aposematic signal evolves from different structures and its similarity is imperfect. The community with multiple patterns sets conditions for the evolution of aposematic sexual dimorphism as demonstrated by the yellow/black male and reticulate female pattern of Micronychus pardus. These insights elucidate the complex character of the evolution of mimetic signalling in the dynamically diversifying biota of high tropical mountains.

Drilidae Blanchard, 1845: multi-gene molecular phylogenies versus morphological similarity. An answer to Kovalev et al.

In a recent issue of Zootaxa, Kovalev et al. (2019) reported the description of a new drilid genus Drilorhinus Kovalev, Kirejtshuk et Shapovalov, 2019 and they discuss at length the relationships and systematic classification of the drilid lineage. Although they did not formally propose a new status for Drilidae Blanchard, 1845 [not Lacordaire, 1857, as cited in their study] in the sense of the Code (ICZN 1999), they in fact resurrected the traditional concept of Drilidae as a separate 'cantharoid' family and rejected Drilini in Elateridae: Agrypninae of Kundrata Bocak (2011). The authors strongly defended the historical classification of the genera Selasia Laporte, 1838, Malacogaster Bassi, 1834, Drilus Olivier, 1790, Drilorhinus, and putatively also their relatives (Kundrata et al. 2017), as a separate 'cantharoid' family in the sense proposed in 19th century and held until recently by some traditional morphologists (Blanchard 1845; Crowson 1955, 1972; Lawrence Newton 1982, 1995; Branham Wenzel 2003; Lawrence et al. 2011; Kazantsev 2013; Lawrence Slipinski 2013; Lawrence 2016). Although no new data were presented or methodological flaws of earlier analyses identified by Kovalev et al., the authors argued that the present evidence is insufficient for the placement of the Drilini in Elateridae, Agrypninae. Unfortunately, they did not take into account recent molecular analyses suggesting multiple origins of soft-bodied elateroid families, i.e., the polyphyly of the historical cantharoid group of families, and the relationships of drilids and agrypnine click beetles (Bocakova et al. 2007; Hunt et al., 2007; Timmermans et al. 2010, 2016; Kundrata et al. 2014; McKenna et al. 2015; Bocak et al. 2016; Kusy et al. 2018a, b; Linard et al. 2018; Zhang et al. 2018). They only discussed a single molecular analysis by Kundrata Bocak (2011).

New species of Moluccan trichaline net-winged beetles, with remarks on the phylogenetic position and distribution of Schizotrichalus (Coleoptera: Lycidae: Metriorrhynchinae).

The Moluccan net-winged beetle fauna remains poorly studied and here, new species of Schizotrichalus Kleine, 1926 and Eniclases Waterhouse, 1879 are reported from Halmahera. Using morphological traits and cox1 mitochondrial DNA sequences, we propose two new species, Eniclases kusyi sp. nov. and Schizotrichalus halmaherensis sp. nov., and redescribe E. moluccanus Kleine, 1930. New molecular data confirm morphology-based sister relationships between Schizotrichalus and Eniclases and the analysis identifies the combined area of the present-day Halmahera and New Guinea as an ancestral area of these genera. Now, Halmahera and New Guinea are quite similar in respect of the number of trichaline genera. Concerning the size of islands and the recent origin of the nowadays northern Moluccas, these results are unexpected and thus the general validity of this distribution pattern should be confirmed with other groups of beetles.

Persistence of multiple patterns and intraspecific polymorphism in multi-species Müllerian communities of net-winged beetles.

BACKGROUND: In contrast to traditional models of purifying selection and a single aposematic signal in Müllerian complexes, some communities of unprofitable prey contain members with multiple aposematic patterns. Processes responsible for diversity in aposematic signaling are poorly understood and large multi-species communities are seldom considered. RESULTS: We analyzed the phylogeny and aposematic patterns of closely related Eniclases net-winged beetles in New Guinea using mtDNA and nextRAD data. We suggest three clades of closely related and incompletely reproductively isolated lineages, detail the extent of polymorphism among Eniclases, and categorize their low-contrast aposematic patterns. The warning signal of Eniclases consists of body shape and color, with ambiguous color perception under some circumstances, i.e., when resting on the undersides of leaves. Field observations suggest that perception of the aposematic signal is affected by beetle behavior and environmental conditions. Local communities containing Eniclases consisted of 7-85 metriorrhynchine species assigned to 3-10 colour patterns. CONCLUSION: As a result, we suggest that under certain light conditions the aposematic colour signal is less apparent than the body shape in net-winged beetle communities. We document variable environmental factors in our study area and highly diverse multi-species communities of other net-winged beetles. Which implies dynamically changing community structure in space and time. Variable environmental conditions and diverse community composition are suggested to be favorable for the persistence of multiple aposematic patterns, imperfect mimics, and intraspecific polymorphism. Further research should identify the relative effect of these factors on purifying selection and the alleles which are responsible for phenotypic differences.

Genomic and Mitochondrial Data Identify Different Species Boundaries in Aposematically Polymorphic Eniclases Net-Winged Beetles (Coleoptera: Lycidae).

Species delineation is essential for any evolutionary and biodiversity research, and recent advances in genomic sequencing have made it possible to robustly define species boundaries and detect hidden diversity. Here, we studied 14 species of aposematically colored New Guinean Eniclases (Coleoptera: Lycidae) whose conventional morphology- and single-locus mtDNA-based taxonomy has been contentious. We analyzed mitochondrial and restriction site associated DNA fragments to obtain a phylogenetic hypothesis and compared relationships recovered by the RAD analysis with species limits based on other information. The results show the presence of cryptic diversity and common mitonuclear discordance when over 30% of individuals were incorrectly assigned to species if only mitogenomic markers were considered. Nuclear data falsified the species rank of one species and identified one earlier unrecognized lineage deserving species rank. Further, our analyses demonstrate a highly variable phenotypic differentiation, with several pairs of cryptic species standing in contrast with genetically close but phenotypically highly divergent lineages. We show that morphological and mitogenomic analyses produce reliable information for taxonomy in most cases. Nevertheless, the species boundaries among closely related species should be based on all lines of evidence, including nuclear markers.

The Malacca Strait separates distinct faunas of poorly-flying Cautires net-winged beetles.

We investigated the spatial and temporal patterns of Cautires diversification on the Malay Peninsula and Sumatra to understand if the narrow and frequently dry Malacca Strait separates different faunas. Moreover, we analyzed the origin of Cautires in Malayan and Sumatran mountains. We sampled 18 localities and present the mtDNA-based phylogeny of 76 species represented by 388 individuals. The phylogenetic tree was dated using mtDNA evolution rates and the ancestral ranges were estimated using the maximum likelihood approach. The phylogeny identified multiple lineages on the Malay Peninsula since the Upper Eocene (35 million years ago, mya) and a delayed evolution of diversity in Sumatra since the Upper Oligocene (26 mya). A limited number of colonization events across the Malacca Strait was identified up to the Pliocene and more intensive faunal exchange since the Pleistocene. The early colonization events were commonly followed by in situ diversification. As a result, the Malacca Strait now separates two faunas with a high species-level turnover. The montane fauna diversified in a limited space and seldom took part in colonization events across the Strait. Besides isolation by open sea or a savannah corridor, mimetic patterns could decrease the colonization capacity of Cautires. The Malay fauna is phylogenetically more diverse and has a higher value if conservation priorities should be defined.

Molecular Phylogeny, Diversity and Zoogeography of Net-Winged Beetles (Coleoptera: Lycidae).

We synthesize the evidence from molecular phylogenetics, extant distribution, and plate tectonics to present an insight in ancestral areas, dispersal routes and the effectiveness of geographic barriers for net-winged beetle tribes (Coleoptera: Lycidae). Samples from all zoogeographical realms were assembled and phylogenetic relationships for ~550 species and 25 tribes were inferred using nuclear rRNA and mtDNA markers. The analyses revealed well-supported clades at the rank of tribes as they have been defined using morphology, but a low support for relationships among them. Most tribes started their diversification in Southeast and East Asia or are endemic to this region. Slipinskiini and Dexorini are Afrotropical endemics and Calopterini, Eurrhacini, Thonalmini, and Leptolycini remained isolated in South America and the Caribbean after their separation from northern continents. Lycini, Calochromini, and Erotini support relationships between the Nearctic and eastern Palearctic faunas; Calochromini colonized the Afrotropical realm from East Asia and Metriorrhynchini Afrotropical and Oriental realms from the drifting Indian subcontinent. Most tribes occur in the Oriental and Sino-Japanese realms, the highest alpha-taxonomic diversity was identified in Malesian tropical rainforests. The turn-over at zoogeographical boundaries is discussed when only short distance over-sea colonization events were inferred. The lycid phylogeny shows that poor dispersers can be used for reconstruction of dispersal and vicariance history over a long time-span, but the current data are insufficient for reconstruction of the early phase of their diversification.

Genome sequences identify three families of Coleoptera as morphologically derived click beetles (Elateridae).

Plastoceridae Crowson, 1972, Drilidae Blanchard, 1845 and Omalisidae Lacordaire, 1857 (Elateroidea) are families of the Coleoptera with obscure phylogenetic relationships and modified morphology showing neotenic traits such as soft bodies, reduced wing cases and larviform females. We shotgun sequenced genomes of Plastocerus, Drilus and Omalisus and incorporated them into data matrices of 66 and 4202 single-copy nuclear genes representing Elateroidea. Phylogenetic analyses indicate their terminal positions within the broadly defined well-sclerotized and fully metamorphosed Elateridae and thus Omalisidae should now be considered as Omalisinae stat. nov. in Elateridae Leach, 1815. The results support multiple independent origins of incomplete metamorphosis in Elateridae and indicate the parallel evolution of morphological and ecological traits. Unlike other neotenic elateroids derived from the supposedly pre-adapted aposematically coloured and unpalatable soft-bodied elateroids, such as fireflies (Lampyridae) and net-winged beetles (Lycidae), omalisids and drilids evolved from well-sclerotized click beetles. These findings suggest sudden morphological shifts through incomplete metamorphosis, with important implications for macroevolution, including reduced speciation rate and high extinction risk in unstable habitats. Precise phylogenetic placement is necessary for studies of the molecular mechanisms of ontogenetic shifts leading to profoundly changed morphology.

Genome sequencing of Rhinorhipus Lawrence exposes an early branch of the Coleoptera.

BACKGROUND: Rhinorhipidae Lawrence, 1988 is an enigmatic beetle family represented by a single species, Rhinorhipus tamborinensis Lawrence, 1988, from Australia, with poorly established affinities near the superfamily Elateroidea (click beetles, soldier beetles and fireflies) or the more inclusive series (infraorder) Elateriformia. Its evolutionary position may inform the basal relationships of the suborder Polyphaga, the largest clade of Coleoptera. RESULTS: We analyzed four densely sampled DNA datasets of major coleopteran lineages for mitogenomes, rRNA genes and single copy nuclear genes. Additionally, genome sequencing was used for incorporation of R. tamborinensis into a set of 4220 orthologs for 24 terminals representing 12 polyphagan superfamilies. Topologies differed to various degrees, but all consistently refute the proposed placement of Rhinorhipidae in Elateroidea and instead indicate either sister relationships with other Elateriformia, frequently together with Nosodendridae, another divergent small family hitherto placed in Derodontoidea, or in an isolated position among the deepest lineages of Polyphaga. The phylogenomic analyses recovered Rhinorhipus in a sister position to all other Elateriformia composed of five superfamilies. Therefore, we erect the new superfamily Rhinorhipoidea Lawrence, 1988, stat. Nov., with the type-family Rhinorhipidae. The origins of the Rhinorhipidae were dated to the Upper Triassic/Lower Jurassic at the very early phase of polyphagan diversification. CONCLUSIONS: Thus, Rhinorhipidae adds another example to several recently recognized ancient relict lineages which are interspersed within contemporaneous hugely species-rich lineages of Coleoptera.

Incomplete sclerotization and phylogeny: The phylogenetic classification of Plastocerus (Coleoptera: Elateroidea).

The relationships of the monogeneric family Plastoceridae Crowson, 1972 (Coleoptera: Elateroidea) have remained contentious due to its modified morphology, incorrect information on incomplete metamorphosis of females and the absence of molecular data. We produced the sequences for P. angulosus (Germar, 1844) (the type-species of Plastocerus Schaum, 1852) and performed molecular phylogenetic analyses to estimate its position. The analyses of Elateroidea (186 spp.) and Elateridae (110 spp.) molecular datasets of two mitochondrial and two nuclear gene fragments repeatedly placed Plastocerus Schaum, 1852 in relationships with the elaterid genera Oxynopterus Hope, 1842 and Pectocera Hope, 1842. Alternative topologies were rejected by likelihood tests. Therefore, Plastoceridae Crowson, 1972 are down-ranked to the subfamily Plastocerinae in Elateridae Leach, 1815. We suggest that the morphology-based placement and high rank for some elateroid lineages were inferred from the presence of homoplasies which evolved due to incomplete sclerotization. Distantly related soft-bodied elateroids share freely movable and transverse coxae, a shortened prosternum, and a weakly sclerotized abdomen with free ventrites. Importantly, the apomorphic structures characteristic for their closest relatives, such as the prosternal process, mesoventral cavity, and intercoxal keel in the first abdominal ventrite are regularly absent. Consequently, morphology-based phylogenetic analyses suggest deeply rooted positions for lineages without expressed apomorphic character states. Molecular data represent an independent character system that is not affected by the convergent morphological evolution, and therefore molecular phylogenies can elucidate the relationships of incompletely sclerotized lineages.

The comparison of molecular and morphology-based phylogenies of trichaline net-winged beetles (Coleoptera: Lycidae: Metriorrhynchini) with description of a new subgenus.

Separate morphological and molecular phylogenetic analyses are presented and the classification of trichaline net-winged beetles is revised. The clade, earlier given a subfamily, tribe or subtribe rank, is a terminal lineage in Metriorrhynchina and contains Diatrichalus Kleine, 1926, Eniclases Waterhouse, 1879, Flabellotrichalus Pic, 1921, Lobatang Bocak, 1998, Microtrichalus Pic, 1921, Schizotrichalus Kleine, 1926, and Trichalus Waterhouse, 1877. Maibrius subgen. nov. is proposed in Flabellotrichalus with the type-species Flabellotrichalus (Maibrius) horaki sp. nov. Unlike previous studies, Lobatang is included in the trichaline clade. Further, Spinotrichalus Kazantsev, 2010, stat. nov. is down-ranked to the subgenus in Lobatang Bocak, 1998 and a new combination, Lobatang (Spinotrichalus) telnovi (Kazantsev, 2010) comb. nov., is proposed. The morphology does not provide a sufficient support for robust phylogeny due to the intrageneric variability of most phenotypic traits and the limited number of characters supporting deep relationships. Most morphological generic diagnoses must be based on the shape of male genitalia. Other characters, such as the shapes of pronotum and antennae are commonly variable within genera. The fronto-lateral pronotal ridges of Eniclases + Schizotrichalus resemble the ancestral condition in Metriorrhynchini and they re-evolved in the terminal clade and do not indicate the early split of Eniclases + Schizotrichalus from other trichaline genera. The evolution of morphological traits and the conflict in the morphological and molecular phylogenetic signal are discussed in details. We suggest that the general appearance is affected by the evolution of mimetic complexes, the patterns of elytral costae by their strengthening function, and the presence of flabellate antennae by their role in sexual communication. Then, similar phenotypic traits evolve in unrelated lineages. The results demonstrate that phylogenetic classification must be based on all available information because neither morphological traits nor DNA data robustly support all recovered relationships.

New species of Diatrichalus (Coleoptera: Lycidae) from New Guinea and the Moluccas.

The recently collected material of Diatrichalus Kleine, 1926 from New Guinea and the Moluccas was studied. Four new species are described: D. buruensis sp. n., D. manokwarensis sp. n., D. mindikensis sp. n., and D. robustus sp. n. The diagnostic morphological characters are described in detail and illustrated. New records of previously described species are reported from several localities in New Guinea and D. mancus (Kleine, 1926) is for the first time reported from the Australian continent. Some Diatrichalus have similar male genitalia and are considered as closely related. Despite supposed relationships, they differ in the presence of secondary elytral longitudinal costae, which have been traditionally used for the delimitation of genera in net-winged beetles.

Species limits in polymorphic mimetic Eniclases net-winged beetles from New Guinean mountains (Coleoptera, Lycidae).

Species delimitation was compared in a group of closely related lineages of aposematically colored Eniclases (Coleoptera, Lycidae) using morphology, genetic distances, and Bayesian implementation of the Poisson Tree Processes model. A high diversity of net-winged beetles was found in previously unsampled regions of New Guinea and ten new species are described: Eniclases bicolor sp. n., Eniclases bokondinensis sp. n., Eniclases brancuccii sp. n., Eniclases elelimensis sp. n., Eniclases infuscatus sp. n., Eniclases niger sp. n., Eniclases pseudoapertus sp. n., Eniclases pseudoluteolus sp. n., Eniclases tikapurensis sp. n., and Eniclases variabilis sp. n. Different levels of genetic and morphological diversification were identified in various sister-species pairs. As a result, both morphological and molecular analyses are used to delimit species. Sister-species with uncorrected pairwise genetic divergence as low as 0.45% were morphologically distinct not only in color pattern, but also in the relative size of eyes. Conversely, differences in color pattern regardless of their magnitude did not necessarily indicate genetic distance and intraspecific mimicry polymorphism was common. Additionally, genetic divergence without morphological differentiation was detected in one sister-species pair. Low dispersal propensity, diverse mimicry patterns, and mimetic polymorphism resulted in complex diversification of Eniclases and uncertain species delimitation in recently diversified lineages.