Phylogeographic and Paleoclimatic Modelling Tools Improve Our Understanding of the Biogeographic History of Hierophis viridiflavus (Colubridae).

Phylogeographic and paleoclimatic modelling studies have been combined to infer the role of Pleistocene climatic oscillations as drivers of the genetic structure and distribution of Mediterranean taxa. For the European whip snake, Hierophis viridiflavus, previous studies based on paleoclimatic modelling have depicted a low reliability in the pattern of past climatic suitability across the central Mediterranean Basin, which barely fits the species' genetic structure. In this study, we combined phylogeographic and paleoclimatic modelling tools to improve our understanding of the biogeographic history of H. viridiflavus, particularly extending the sampling and phylogeographic inferences to previously under-sampled regions. Phylogeographic analyses recovered two major clades that diverged at the beginning of the Pleistocene and had diversified in different ways by the late Pleistocene: the east clade (composed of three subclades) and the west clade (with no further structure). Paleoclimatic models highlighted the temperate character of H. viridiflavus, indicating range contractions during both the last inter-glacial and last glacial maximum periods. Range expansions from southern-located climatic refugia likely occurred in the Bølling-Allerød and Middle Holocene periods, which are supported by signals of demographic growth in the west clade and South-East-North subclade. Overall, this work improves our understanding of the historical biogeography of H. viridiflavus, providing further insights into the evolutionary processes that occurred in the Mediterranean Basin hotspot.

Nomenclatural changes in a taxonomically complex group (Amphipoda: Gammaridae).

The family Gammaridae is a large monophyletic group with a complicated taxonomy. A nomenclatural review of the taxonomic status of Pectenogammarus Reid, 1940 was conducted based on Hou & Sket (2016) and Sket & Hou (2018) phylogenetic proposals. These authors used the name Homoeogammarus Schellenberg, 1937 for a set of morphologically diverse taxa previously included mainly within Echinogammarus Stebbing, 1899. Unfortunately, the use of the name Homoeogammarus Schellenberg, 1937 is problematic. As a result of a detailed bibliographic review, we propose the name Pectenogammarus Reid, 1940 (stat. nov.) for the "Homoeogammarus" clade (Sket & Hou 2018), which results in 20 new combinations. In addition, two genera are here formally synonymised as follows: Pectenogammarus Reid, 1940 = Homoeogammarus Barnard et Barnard, 1983, syn. nov. = Laurogammarus G.S. Karaman, 1984, syn. nov. A synonymic catalogue is provided.

Analyses of head and thorax in Eupomphini (Meloidae) suggest that complex behaviors are not associated to changes in general shape.

Our results reject an association between general shape of head and thorax with defensive and courtship behaviors in the studied Eupomphini blister beetles. Instead we found that shape of thorax and head radiated in all directions of the morphospace. We also identified a radical separation between two lineages that could represent future evolutionary constraints for thorax evolution.

Mitogenomics and hidden-trait models reveal the role of phoresy and host shifts in the diversification of parasitoid blister beetles (Coleoptera: Meloidae).

Changes in life history traits are often considered speciation triggers and can have dramatic effects on the evolutionary history of a lineage. Here, we examine the consequences of changes in two life history traits, host-type and phoresy, in the hypermetamorphic blister beetles, Meloidae. Subfamilies Nemognathinae and Meloinae exhibit a complex life cycle involving multiple metamorphoses and parasitoidism. Most genera and tribes are bee-parasitoids, and include phoretic or nonphoretic species, while two tribes feed on grasshopper eggs. These different life strategies are coupled with striking differences in species richness among clades. We generated a mitogenomic phylogeny for Nemognathinae and Meloinae, confirming the monophyly of these two clades, and used the dated phylogeny to explore the association between diversification rates and changes in host specificity and phoresy, using state-dependent speciation and extinction (SSE) models that include the effect of hidden traits. To account for the low taxon sampling, we implemented a phylogenetic-taxonomic approach based on birth-death simulations, and used a Bayesian framework to integrate parameter and phylogenetic uncertainty. Results show that the ancestral hypermetamorphic Meloidae was a nonphoretic bee-parasitoid, and that transitions towards a phoretic bee-parasitoid and grasshopper parasitoidism occurred multiple times. Nonphoretic bee-parasitoid lineages exhibit significantly higher relative extinction and lower diversification rates than phoretic bee-and grasshopper-parasitoids, but no significant differences were found between the latter two strategies. This suggests that Orthopteran host shifts and phoresy contributed jointly to the evolutionary success of the parasitoid meloidae. We also demonstrate that SSE models can be used to identify hidden traits coevolving with the focal trait in driving a lineage's diversification dynamics.

A new systematic arrangement for the blister beetle genus Eurymeloe (Meloini, Meloidae, Coleoptera) with the description of a new species from Spain.

The taxonomic status and subgeneric arrangement of the genus Eurymeloe have been debated for decades. In this work, the internal taxonomy of Eurymeloe is redefined by recognising three subgenera: Eurymeloe for the former Eurymeloebrevicollis species group, Coelomeloe for Eurymeloetuccia, and Bolognaia Ruiz, García-París, Sánchez-Vialas & Recuero, subgen. nov., to accommodate the species of the formerly recognised Eurymeloerugosus species group. Additionally, a new species of the newly described subgenus Bolognaia is described from the Iberian Peninsula based on molecular and morphological traits. The new species, Eurymeloe (Bolognaia) orobatessp. nov., can be distinguished from all other species of Eurymeloe by the following combination of morphological traits: dispersed brownish setae over the body that are arranged in small tufts on the abdominal terga; a small, very transverse pronotum that presents a unique macrosculpture; a deeply and densely punctured integument of the head and pronotum; and the very rugose elytra. The characters displayed by E.orobates suggest that the species groups that were previously defined and recognised for Eurymeloe, and that are now integrated within the newly erected subgenus Bolognaia, are non-monophyletic.

Antiparasitic Effects of Potentially Toxic Beetles (Tenebrionidae and Meloidae) from Steppe Zones.

Arthropods and specifically beetles can synthesize and/or sequester metabolites from dietary sources. In beetle families such as Tenebrionidae and Meloidae, a few studies have reported species with toxic defensive substances and antiparasitic properties that are consumed by birds. Here we have studied the antiparasitic activity of extracts from beetle species present in the habitat of the Great Bustard (Otis tarda) against four pathogen models (Aspergillus niger, Meloidogyne javanica, Hyalomma lusitanicum, and Trichomonas gallinae). The insect species extracted were Tentyria peiroleri, Scaurus uncinus, Blaps lethifera (Tenebrionidae), and Mylabris quadripunctata (Meloidae). M. quadripunctata exhibited potent activity against M. javanica and T. gallinae, while T. peiroleri exhibited moderate antiprotozoal activity. The chemical composition of the insect extracts was studied by gas chromatography coupled with mass spectrometry (GC-MS) analysis. The most abundant compounds in the four beetle extracts were hydrocarbons and fatty acids such as palmitic acid, myristic acid and methyl linoleate, which are characteristic of insect cuticles. The presence of cantharidin (CTD) in the M. quadripunctata meloid and ethyl oleate (EO) in T. peiroleri accounted for the bioactivity of their extracts.

Settling taxonomic and nomenclatural problems in brine shrimps, Artemia (Crustacea: Branchiopoda: Anostraca), by integrating mitogenomics, marker discordances and nomenclature rules.

High morphological plasticity in populations of brine shrimp subjected to different environmental conditions, mainly salinity, hindered for centuries the identification of the taxonomic entities encompassed within Artemia. In addition, the mismatch between molecular and morphological evolution rates complicates the characterization of evolutionary lineages, generating taxonomic problems. Here, we propose a phylogenetic hypothesis for Artemia based on two new complete mitogenomes, and determine levels of congruence in the definition of evolutionary units using nuclear and mtDNA data. We used a fossil of Artemia to calibrate the molecular clock and discuss divergence times within the genus. The hypothesis proposed herein suggests a more recent time frame for lineage splitting than previously considered. Phylogeographic analyses were performed using GenBank available mitochondrial and nuclear markers. Evidence of gen e flow, identified through discordances between nuclear and mtDNA markers, was used to reconsider the specific status of some taxa. As a result, we consider Artemia to be represented by five evolutionary units: Southern Cone, Mediterranean-South African, New World, Western Asian, and Eastern Asian Lineages. After an exhaustive bibliographical revision, unavailable names for nomenclatural purposes were discarded. The remaining available names have been assigned to their respective evolutionary lineage. The proper names for the evolutionary units in which brine shrimps are structured remain as follows: Artemia persimilis Piccinelli & Prosdocimi, 1968 for the Southern Cone Lineage, Artemia salina (Linnaeus, 1758) for the Mediterranean-SouthAfrican Lineage, Artemia urmiana Günther, 1899 for the Western Asian Lineage, and Artemia sinica Cai, 1989 for the Eastern Asian Lineage. The name Artemia monica Verrill, 1869 has nomenclatural priority over A. franciscana Kellogg, 1906 for naming the New World Lineage. New synonymies are proposed for A. salina (= C. dybowskii Grochowski, 1896 n. syn., and A. tunisiana Bowen & Sterling, 1978 n. syn.), A. monica (= A. franciscana Kellogg, 1906 n. syn., and A. salina var. pacifica Sars, 1904 n. syn.); A. urmiana (= B. milhausenii Fischer de Waldheim, 1834 n. syn., A. koeppeniana Fischer, 1851 n. syn., A. proxima King, 1855 n. syn., A. s. var. biloba Entz, 1886 n. syn., A. s. var. furcata Entz, 1886 n. syn., A. asiatica Walter, 1887 n. syn., A. parthenogenetica Bowen & Sterling, 1978 n. syn., A. ebinurica Qian & Wang, 1992 n. syn., A. murae Naganawa, 2017 n. syn., and A. frameshifta Naganawa & Mura, 2017 n. syn.). Internal deep nuclear structuring within the A. monica and A. salina clades, might suggest the existence of additional evolutionary units within these taxa.

Proctodeal extrusion as a defensive behavioral response in blister beetles (Coleoptera: Meloidae).

Defensive mechanisms in blister beetles (Coleoptera: Meloidae) include a wide variety of behavioral responses, chemical defense, and conspicuous external colorations. Although some of these mechanisms have been previously described, proctodeal extrusion, a defensive behavior involving the extrusion of inner abdominal membranes from the proctodeal region which appear intensely red or orange colored when the hemolymph is seen through them, has not been reported to date. Here, we tested the ability to display proctodeal extrusion in response to threat stimuli in wild populations of three blister beetle species inhabiting Central Spain: Berberomeloe majalis (Linnaeus, 1758), Berberomeloe comunero Sánchez-Vialas, García-París, Ruiz & Recuero, 2020, and Physomeloe corallifer (Germar, 1818). In addition, we observed and recorded various other defensive behaviors such as immobility, antennal threat display, autohemorrhage (reflex bleeding), defecation, and thanatosis (death feigning). The frequency at which proctodeal extrusion was observed differed among species, as did the stress intensity needed for extrusion and the probability of proctodeal extrusion in response to a particular threatening stimulus. Our findings indicate that, although proctodeal extrusion might be a widespread potential defensive mechanism in Meloidae, the ability to elicit it is not generalized across lineages. Physomeloe and Berberomeloe are endemic to the semi-arid Mediterranean region, and species adapted to such a climate would have developed strategies that limit hydric stress such as proctodeal extrusion, which mirrors the effect of autohemorrhage but without the fluid loss.

Taxonomy of the North Moroccan and Iberian species of the subgenus Amblypteraca/ (Coleoptera: Tenebrionidae: Pimeliinae: Pimelia/).

The subgenus Amblypteraca Mas-Peinado, Buckley, Ruiz García-París, 2018 of Pimelia Fabricius, 1775, is restricted to the southern Iberian Peninsula and western Maghreb (northern and western Morocco). The distribution of Amblypteraca throughout the African-European edges overlaps largely with the geographic range of the subgenus Amblyptera, which is sister to the clade grouping subgenera Amblypteraca and Ecphoroma Solier, 1836. Delimiting species boundaries in the speciose genus Pimelia is often challenging, and the taxonomic status of some groups within the aforementioned subgenera is still a matter of debate. Here, we aim to stabilize some of the available names in Amblypteraca, and to correct some previous misidentifications. For that purpose, we discuss the composition and taxonomic structure within Amblypteraca by (i) assessing the phylogenetic congruence between mitochondrial and nuclear markers, and (ii) examining external morphological traits in 568 Amblypteraca specimens under the light of the phylogenetic hypotheses proposed here. Based on our results, Amblypteraca consists of three species: P. rotundipennis Kraatz, 1865, P. fairmairii Kraatz, 1865 and P. chrysomeloides Pallas, 1781. Both molecular and morphological data revealed four lineages within P. chrysomeloides: P. chrysomeloides chrysomeloides, distributed on both sides of the Strait of Gibraltar; P. chrysomeloides fornicata Herbst, 1799 from Portugal (Troia region); P. chrysomeloides bathyglypta Antoine, 1949, restricted to a narrow strip between Larache and Arbaoua (northern atlantic Moroccan coast), and P. chrysomeloides subris Koch, 1941 from Kenitra-La Mamora forest (Morocco). We designate a neotype of Tenebrio chrysomeloides Pallas, 1781 and propose the synonymy of P. chrysomeloides (Pallas, 1781) = P. obesa Solier, 1836 syn. n. Pimelia tristis Haag-Rutenberg, 1875, previously misidentified and included in Amblypteraca, is now transferred back to Amblyptera. Further studies with ad hoc sampling designs and analytical tools would be in need to delimit the exact geographic ranges of these taxa, and to analyse the patterns of diversity within and among species and subspecies.

Catalogue, distribution, taxonomic notes, and conservation of the Western Palearctic endemic hunchback beetles (Tenebrionidae, Misolampus).

Hunchback darkling beetles of the Ibero-Maghrebian genus Misolampus Latreille, 1807 (Tenebrionidae, Stenochiinae) encompass six species: M. gibbulus (Herbst, 1799), M. goudotii Guérin-Méneville, 1834, M. lusitanicus Brême, 1842, M. ramburii Brême, 1842, M. scabricollis Graells, 1849, and M. subglaber Rosenhauer, 1856. Previously known distribution ranges of the species were delineated using many old records, the persistence of such populations being questionable under the current situation of global biodiversity loss. Additionally, the status of geographically isolated populations of the genus have been the subject of taxonomic controversy. An exhaustive bibliographical revision and field search was undertaken, and the Misolampus collection of the Museo Nacional de Ciencias Naturales (MNCN-CSIC) was revised. The aims are to (i) provide an updated geographic distribution range for the species of Misolampus; (ii) to determine the taxonomic status of controversial populations; (iii) to provide a catalogue for Misolampus; and (iv) to discuss the conservation status of these saproxylic beetles. As a result, a catalogue including synonymies and type localities, geographical records, diagnoses, and information on natural history for all species of Misolampus is presented. The results reveal that the distribution ranges of the species of Misolampus have not undergone a reduction in the last century, and indicate the presence of the genus in areas where it had never been recorded before. The morphological variability of M. goudotii drove the proposal of different taxa that are here formally synonymised as follows: M. goudotii Guérin-Méneville, 1834 = M. erichsoni Vauloger de Beaupré, 1900, syn. nov. = M. peyerimhoffi Antoine, 1926, syn. nov.

Limited long-distance dispersal success in a Western European fairy shrimp evidenced by nuclear and mitochondrial lineage structuring.

Anostraca are known by their ability for long-distance dispersal, but the existence in several species of deep, geographically structured mtDNA lineages suggests their populations are subjected to allopatric differentiation, isolation, and prevalence of local scale dispersion. Tanymastix stagnalis is one of the most widespread species of Anostraca and previous studies revealed an unclear geographical pattern of mtDNA genetic diversity. Here, we analyze populations from the Iberian and Italian Peninsulas, Central Europe, and Scandinavia, with the aim to characterize the patterns of genetic diversity in a spatio-temporal framework using mtDNA and nuclear markers to test gene flow among close populations. For these aims we built a time-calibrated phylogeny and carried out Bayesian phylogeographic analyses using a continuous diffusion model. Our results indicated that T. stagnalis presents a deeply structured genetic diversity, including 7 ancient lineages, some of them even predating the Pleistocene. The Iberian Peninsula harbors high diversity of lineages, with strong isolation and recent absence of gene flow between populations. Dispersal at local scale seems to be the prevailing dispersal mode of T. stagnalis, which exhibits a pattern of isolation-by-distance in the Iberian Peninsula. We remark the vulnerability of most of these lineages, given the limited known geographic distribution of some of them, and the high risk of losing important evolutionary potential for the species.

A new singular species of Croscherichia Pardo Alcaide, 1950 (Coleoptera, Meloidae, Mylabrini) from arid zones of eastern Morocco.

A new species of blister beetle (Coleoptera, Meloidae, Mylabrini), Croscherichia armass Ruiz, François & García-París, sp. nov., is described from the arid steppes of eastern Morocco (Missour, Boulemane Province). The new species presents traits shared with both Croscherichia and desert species of the genus Ammabris, making it morphologically singular. Conspicuous external similarities (coloration pattern, shape of the mandibles, setation) between C. armass sp. nov. and Ammabris allow the two to be easily confused. However, C. armass sp. nov. can be readily distinguished from all other Croscherichia species by the following traits: reddish-orange legs with dark tarsi; relatively short black antennae with the proximal-most three to four antennomeres of each antenna having a reddish-brown coloration; dense and silvery body setation that lies over most of the body integument; straight and pointed outer mandible margins that protrude from the labrum; a mesosternum with an angulate anterior margin; a short, subcylindrical, and weakly spatulate external metatibial spur that is truncated obliquely at the apex. Croscherichia armass sp. nov. is only known from three localities in the arid Hammada steppes, which are located within the Quaternary alluvial plains of the Muluya river valley. Live specimens of C. armass sp. nov. were found in flight and actively feeding on Atriplex halimus (Chenopodiaceae) flowers at the end of summer (mid-September). The phenology of C. armass sp. nov. is exceptional as no other Mylabrini species known from eastern areas of Morocco are active in late summer.

High extinction rates and non-adaptive radiation explains patterns of low diversity and extreme morphological disparity in North American blister beetles (Coleoptera, Meloidae).

Untangling the relationship between morphological evolution and lineage diversification is key to explain global patterns of phenotypic disparity across the Tree of Life. Few studies have examined the relationship between high morphological disparity and extinction. In this study, we infer phylogenetic relationships and lineage divergence times within Eupomphini (Meloidae), a tribe of blister beetles endemic to the arid zone of North America, which exhibits a puzzling pattern of very low species richness but wild variation in morphological diversity across extant taxa. Using Bayesian and maximum likelihood inference, we estimate diversification and phenotypic evolutionary rates and infer the time and magnitude of extinction rate shifts and mass extinction events. Our results suggest that Eupomphini underwent an event of ancient radiation coupled with rapid morphological change, possibly linked to the loss of the evolutionary constraint in the elytral shape. A high extinction background associated to the Miocene-Pliocene transition decimated the diversity within each major clade, resulting in the species-poor genera observed today. Our study supports a connection between high extinction rates and patterns of decoupled phenotypic evolution and lineage diversification, and the possibility of a radiation in the absence of ecological release.

Mitochondrial phylogeography of the Iberian endemic frog Rana iberica, with implications for its conservation.

Genetic characterization of species using phylogeographic approaches represents a basic reference to understand their evolutionary history as well as to identify conservation priorities to protect areas of particular interest regarding evolutionary potential. Even in well-studied regions such information is lacking for the majority of species, including many endemic species with reduced distribution ranges. We investigate the phylogeographic pattern of the Iberian frog Rana iberica, an endemic amphibian restricted to Central and North-Western Iberian Peninsula. Using mitochondrial sequences, we reconstruct the phylogeographic history of the species to test the effect of Quaternary climate changes on the evolutionary diversification of lineages, that is, the differentiation of mitochondrial lineages and the formation of genetic diversity melting pots, and integrate phylogeographic evidence for future conservation planning. Our results indicate the existence of 3 main mitochondrial lineages differentiated during the Upper Pleistocene. Both historical demographic analyses and climatic niche modeling show a strong effect of glacial climate changes, suggesting recurrent range contractions and expansions. Under such circumstances, differentiation took place most likely by isolation in allopatric interglacial refugia. Secondary lineage admixture in northern Portugal generated a broad mixed zone with highest nucleotide diversity. Given its particular evolutionary potential, its reduced distribution and eventual threats under current climate change scenario, conservation priorities should focus on the isolated lineage from Sierra de Guadalupe.

Biology of tiny animals: three new species of minute salamanders (Plethodontidae: Thorius) from Oaxaca, Mexico.

We describe three new species of minute salamanders, genus Thorius, from the Sierra Madre del Sur of Oaxaca, Mexico. Until now only a single species, T. minutissimus, has been reported from this region, although molecular data have long shown extensive genetic differentiation among geographically disjunct populations. Adult Thorius pinicola sp. nov., T. longicaudus sp. nov., and T. tlaxiacus sp. nov. are larger than T. minutissimus and possess elliptical rather than oval nostrils; T. pinicola and T. longicaudus also have longer tails. All three new species occur west of the range of T. minutissimus, which has the easternmost distribution of any member of the genus. The new species are distinguished from each other and from other named Thorius in Oaxaca by a combination of adult body size, external morphology and osteology, and by protein characters (allozymes) and differences in DNA sequences. In addition, we redescribe T. minutissimus and a related species, T. narisovalis, to further clarify the taxonomic status of Oaxacan populations and to facilitate future studies of the remaining genetically differentiated Thorius that cannot be satisfactorily assigned to any named species. Populations of all five species considered here appear to have declined dramatically over the last one or two decades and live specimens are difficult to find in nature. Thorius may be the most endangered genus of amphibians in the world. All species may go extinct before the end of this century.

Salt lakes of La Mancha (Central Spain): A hot spot for tiger beetle (Carabidae, Cicindelinae) species diversity.

The tiger beetle assemblage of the wetlands of La Mancha (central Spain) comprises nine species: Calomera littoralis littoralis, Cephalota maura maura, Cephalota circumdata imperialis, Cephalota dulcinea, Cicindela campestris campestris, Cicindela maroccana, Cylindera paludosa, Lophyra flexuosa flexuosa, and Myriochila melancholica melancholica. This assemblage represents the largest concentration of tiger beetles in a single 1º latitude / longitude square in Europe. General patterns of spatial and temporal segregation among species are discussed based on observations of 1462 specimens registered during an observation period of one year, from April to August. The different species of Cicindelini appear to be distributed over space and time, with little overlapping among them. Three sets of species replace each other phenologically as the season goes on. Most of the species occupy drying or dried salt lakes and salt marshes, with sparse vegetation cover. Spatial segregation is marked in terms of substrate and vegetation use. Calomera littoralis and Myriochila melancholica have been observed mainly on wet soils; Cephalota circumdata on dry open saline flats; Cephalota dulcinea and Cylindera paludosa in granulated substrates with typical halophytic vegetation; Cephalota maura is often present in man-modified areas. Cephalota circumdata and Cephalota dulcinea are included as species of special interest in the list of protected species in Castilla-La Mancha. Conservation problems for the Cicindelini assemblage arise from agricultural activities and inadequate use of sport vehicles. Attempts at restoring the original habitat, supressing old semi-industrial structures, may affect the spatial heterogeneity of the lakes, and have an effect on Cicindelinae diversity.

Speciation history and widespread introgression in the European short-call tree frogs (Hyla arborea sensu lato, H. intermedia and H. sarda).

European tree frogs (Hyla) characterized by short temporal parameters of the advertisement call form six genetically differentiated but morphologically cryptic taxa, H. arborea sensu stricto, H. orientalis and H. molleri from across Europe to western Asia (together referred to as H. arborea sensu lato), two putative taxa within H. intermedia (Northern and Southern) from the Italian Peninsula and Sicily, and H. sarda from Sardinia and Corsica. Here, we assess species limits and phylogenetic relationships within these 'short-call tree frogs' based on mitochondrial DNA and nuclear protein-coding markers. The mitochondrial and nuclear genes show partly incongruent phylogeographic patterns, which point to a complex history of gene flow across taxa, particularly in the Balkans. To test the species limits in the short-call tree frogs and to infer the species tree, we used coalescent-based approaches. The monophyly of H. arborea sensu lato is supported by the mtDNA as well as by the all-gene species tree. The Northern and Southern lineages of H. intermedia have been connected by nuclear gene flow (despite their deep mtDNA divergence) and should be treated as conspecific. On the contrary, the parapatric taxa within H. arborea sensu lato should be considered distinct species (H. arborea, H. orientalis, H. molleri) based on the coalescent analysis, although signs of hybridization were detected between them (H. arborea×H. orientalis; H. arborea×H. molleri). A mitochondrial capture upon secondary contact appears to explain the close mtDNA relationship between the geographically remote Iberian H. molleri and H. orientalis from around the Black Sea. Introgressive hybridization occurred also between the Balkan H. arborea and northern Italian H. intermedia, and between the Minor Asiatic H. orientalis and Arabian H. felix arabica (the latter belonging to a different acoustic group/clade). Our results shed light on the species limits in the European short-call tree frogs and show that introgression played an important role in the evolutionary history of the short-call tree frogs and occurred even between taxa supported as distinct species.

Males of a strongly polygynous species consume more poisonous food than females.

We present evidence of a possible case of self-medication in a lekking bird, the great bustard Otis tarda. Great bustards consumed blister beetles (Meloidae), in spite of the fact that they contain cantharidin, a highly toxic compound that is lethal in moderate doses. In addition to anthelminthic properties, cantharidin was effective against gastrointestinal bacteria that cause sexually-transmitted diseases. Although both sexes consumed blister beetles during the mating season, only males selected them among all available insects, and ingested more and larger beetles than females. The male-biased consumption suggests that males could use cantharidin to reduce their parasite load and increase their sexual attractiveness. This plausibly explains the intense cloaca display males perform to approaching females, and the meticulous inspection females conduct of the male's cloaca, a behaviour only observed in this and another similar species of the bustard family. A white, clean cloaca with no infection symptoms (e.g., diarrhoea) is an honest signal of both, resistance to cantharidin and absence of parasites, and represents a reliable indicator of the male quality to the extremely choosy females. Our results do not definitely prove, but certainly strongly suggest that cantharidin, obtained by consumption of blister beetles, acts in great bustards as an oral anti-microbial and pathogen-limiting compound, and that males ingest these poisonous insects to increase their mating success, pointing out that self-medication might have been overlooked as a sexually-selected mechanism enhancing male fitness.

Evolutionary history of Ichthyosaura alpestris (Caudata, Salamandridae) inferred from the combined analysis of nuclear and mitochondrial markers.

Widespread species with morphologically and ecologically differentiated populations are key to understand speciation because they allow investigating the different stages of the continuous process of population divergence. The alpine newt, Ichthyosaura alpestris, with a range that covers a large part of Central Europe as well as isolated regions in all three European Mediterranean peninsulas, and with strong ecological and life-history differences among populations, is an excellent system for such studies. We sampled individuals across most of the range of the species, and analyzed mitochondrial (1442 bp) and nuclear (two nuclear genes -1554 bp- and 35 allozyme loci) markers to produce a time-calibrated phylogeny and reconstruct the historical biogeography of the species. Phylogenetic analyses of mtDNA data produced a fully resolved topology, with an endemic, Balkan clade (Vlasina) which is sister to a clade comprising an eastern and a western group. Within the former, one clade (subspecies I. a. veluchiensis) is sister to a clade containing subspecies I. a. montenegrina and I. a. serdara as well as samples from southern Romania, Bosnia-Herzegovina, Serbia and Bulgaria (subspecies I. a. reiseri and part of I. a. alpestris). Within the western group, populations from the Italian peninsula (subspecies I. a. apuana and I. a. inexpectata) are sister to a clade containing samples from the Iberian Peninsula (subspecies I. a. cyreni) and the remainder of the samples from subspecies I. a. alpestris (populations from Hungary, Austria, Poland, France, Germany and the larger part of Romania). Results of (∗)BEAST analyses on a combined mtDNA and nDNA dataset consistently recovered with high statistical support four lineages with unresolved inter-relationships: (1) subspecies I. a. veluchiensis; (2) subspecies I. a. apuana+I. a. inexpectata; (3) subspecies I. a. cyreni+part of subspecies I. a. alpestris (the westernmost populations, plus most Romanian populations); and (4) the remaining populations, including subspecies I. a. serdara, I. a. reiseri and I. a. montenegrina and part of subspecies I. a. alpestris, plus samples from Vlasina. Our time estimates are consistent with ages based on the fossil record and suggest a widespread distribution for the I. alpestris ancestor, with the split of the major eastern and western lineages during the Miocene, in the Tortonian. Our study provides a solid, comprehensive background on the evolutionary history of the species based on the most complete combined (mtDNA+nDNA+allozymes) dataset to date. The combination of the historical perspective provided by coalescent-based analyses of mitochondrial and nuclear DNA variation with individual-based multilocus assignment methods based on multiple nuclear markers (allozymes) also allowed identification of instances of discordance across markers that highlight the complexity and dynamism of past and ongoing evolutionary processes in the species.

Pharmacological properties of blister beetles (Coleoptera: Meloidae) promoted their integration into the cultural heritage of native rural Spain as inferred by vernacular names diversity, traditions, and mitochondrial DNA.

ETHNOPHARMACOLOGICAL RELEVANCE: Beetles of the family Meloidae (blister beetles) are often reported in pharmacological literature because of their content of cantharidin. Cantharidin has a long history in human medicine and was commonly applied in the 19th and the early 20th centuries, although its use has been progressively abandoned since then. Contrary to most, even common, large species of Coleoptera, blister beetles of the genera Berberomeloe, Physomeloe and to a lesser extent Meloe, are usually recognized and often incorporated into local folk taxonomy by inhabitants of rural areas in Spain. AIM OF THE STUDY: To demonstrate the role that pharmacological properties of blister beetles must have played in their integration in the culture of early Iberian human societies, but also in the preservation of their identity until today, a rare case for Spanish insects. To achieve this purpose we document the diversity of vernacular names applied in rural areas of Spain, and we determine, using molecular data, the antiquity of the presence of two species of the better-known blister beetle in rural Spain, Berberomeloe majalis and Berberomeloe insignis. MATERIALS AND METHODS: We try to document the extent of traditional knowledge of meloid beetles in rural areas by interviewing about 120 people from villages in central and southern Spain. We also use mitochondrial DNA sequences (Cytochrome Oxidase I and 16SrRNA) obtained from several populations of two species of the better known blister beetle in rural Spain, Berberomeloe majalis and Berberomeloe insignis, to determine whether these beetles were already present in the Iberian Peninsula when earlier ancient cultures were developing. RESULTS: Our results show that, based on mitochondrial DNA, blister beetles of the genus Berberomeloe were present in the Iberian Peninsula long before humans arrived, so ancient Iberian cultures were in contact with the same beetle species occurring now in rural areas. On the other hand, people interviewed in rural communities provided us with more than 28 different vernacular names, a few short songs incorporated to local folklore, and some therapeutic uses. CONCLUSIONS: Current knowledge of blister beetles of the family Meloidae in rural Spain was likely developed as a consequence of their pharmacological properties; we hypothesize this knowledge was inherited from ancient pre-Christian Iberian native cultures as part of their traditional therapeutic traditions. It is possible then, that current vernacular names and traditional songs are the only remnants of an ancient knowledge of pharmacological uses of meloid beetles, verbally transmitted from the ancestral cultures to modern day rural Spain. Our work suggests that this legacy, part of the European Cultural Heritage, is disappearing fast, in parallel to the loss of traditional agricultural techniques.