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There is evident variation in body size amongst Anthonomus santacruzi Hustache, 1924, weevils. The aims of this study were to assess if the variation in body size in A. santacruzi weevils is a result of sexual dimorphism and what features can be used to distinguish males from females. The weevils were collected from field sites in Mpumalanga, South Africa, where they were introduced as biocontrol agents of Solanum mauritianum Scopoli. Body structures and the presence/absence of the tergal notch was examined under an optical stereomicroscope and SEM to assess differences between sexes. The morphometric analysis of the body structures included rostrum length (base–apex and antennal insertion–apex), elytral length and width, pronotum length and width, first tarsus length, first tibia length, funiculus length and total body length. Rostrum length, elytra length and width and total body length were significantly larger in females than in males. A tergal notch in the 8th abdominal tergite was present in males and absent in females. The body structure; of rostrum length, elytra length and width and total body length overlapped between sexes in some specimens. The abdominal tergal notch was found to be the most useful body structure to distinguish males from females in A. santacruzi.

Liparus glabrirostris is one of the largest European weevils, and it has been recently proposed as the flagship species for threatened riparian habitats in the mountains. For effective conservation of its populations (and associated habitats), it is crucial to understand its history, contemporary distribution, genetic diversity and predict changes in the range, including its highly isolated populations on the Baltic coast. Here, we examined numerous populations of L. glabrirostris across almost the entire species range using phylogeography and species niche modeling (SNM) approaches. Analyses of mtDNA and nucDNA markers revealed the existence of 2 major mitochondrial lineages generally separated between 1) the Alpine region and 2) the Bohemian Massif, the Carpathians, and the Baltic coast areas. Genetic diversity in nuclear genes was more complicated with no clear division between populations. The origin of Baltic populations was derived from the Carpathians, but there were probably multiple expansion events to the north. SNM suggested the existence of glacial refugia for L. glabrirostris, mainly in the Alps and the Southern Carpathians. Current predictions of species range were found to be generally congruent with zoogeographic data; however, the Baltic coast was not really supported as a suitable area for L. glabrirostris. An important prediction of future distribution (2050–2070 CE) suggests a shrinkage of the L. glabrirostris range and extinction of some of its populations (particularly those isolated on lower altitudes). Based on the aforementioned data, proposals for the protection of this species are proposed, including the designation of several evolutionary units of conservation importance.

Wolbachia, Rickettsia, Spiroplasma and Cardinium are endosymbiotic and intracellular bacteria known to cause numerous disorders in host reproduction, reflected in their common name “male-killers”. In this study, 297 beetle species from various taxonomic groups were screened with the use of molecular markers for the presence of infection by any of these endosymbionts. Wolbachia was found to be the most common “male-killer” among beetle hosts as it infected approx. 27% of species. Rickettsia, Spiroplasma and Cardinium were much less prevalent as they infected: 8%, 3% and 2%, respectively, of the studied beetle species. This is the first report of Cardinium presence in beetle hosts. Incidences of co-infection of two bacteria taxa were very rare and only two weevil species were found to be infected by three different bacteria. These findings suggest that endosymbiotic bacteria inhabit their host at various levels of prevalence and that particular taxa usually infect different hosts, suggesting some competition among “male-killers”. This is the first study that simultaneously verifies infection status by all major endosymbiotic bacteria in hundreds of species (based on coleopterans).

Intracellular bacteria of the genus Wolbachia (alpha-Proteobacteria) are the most widespread endosymbionts of insects. Host infection is usually associated with alterations in reproduction, such as cytoplasmic incompatibility, the induction of parthenogenesis and offspring sex ratio bias: all phenomena that may influence host speciation. In the present study, by using well-established molecular tools, we investigated the presence of Wolbachia in leaf beetles of the genus Crioceris and their host plants, which are various species of Asparagus. Multilocus sequence typing of bacterial genes showed that despite their occurrence in the same habitat and feeding on the same plant, two species of Crioceris, C. quinquepunctata and C. quatuordecimpunctata, are infected by two different strains of Wolbachia. C. asparagi, C. paracenthesis and C. duodecimpunctata, which are sympatric with the infected species, do not harbour the bacterium. Interestingly, DNA of Wolbachia was detected in host plant tissues that are exploited by the beetles, providing evidence for the horizontal transmission of the bacterium between beetles and their host plants. Moreover, Wolbachia was detected in species of Crioceris that are not closely related.

In insects, morphological species identification is often challenging. The discrimination of closely related species may be hampered when only subtle differences in phenotypic characters or a continuum in their variability are present. This is exemplified in the Cryptocephalus flavipes species complex (Coleoptera; Chrysomelidae) where, until now, the species have been discriminated only by the yellow pattern on frons, pronotum, and epipleurae. In the present study, the phylogeny of the C. flavipes species complex was resolved through a multi-locus sequence approach, and the inclusion in the group of the phenotypically similar Cryptocephalus quadripustulatus Gyllenhal, 1813 was evaluated. Subsequently, the C. flavipes species complex was used to test the usefulness of DNA-based approaches for species recognition and identification. All previously described morphospecies were recovered by means of species delimitation approaches. In addition, lineages of Cryptocephalus flavipes Fabricius, 1781 and Cryptocephalus bameuli Duhaldeborde, 1999, which have not evolved clear morphological differences, were found to form genetically well-isolated groups. A new taxon from Turkey was recognized as a separate lineage by DNA-based methods, and morphological diagnostic characters are reported. Through this study, new morphological diagnostic characters were discovered and the suitability of the meta-episternal shape in species discrimination was tested by a geometric morphometric approach.

New data on the genus Rasilinus (Curculioninae, Eugnomini) are provided, including the description of a new species—Rasilinus bifasciatus sp. n., from New Caledonia. Illustrations and SEM photographs of male external morphology, terminalia, and a colour habitus of the adult of the new species are provided.

The genus Rasilinus gen. n. is described (type species Rasilinus tchambicus sp. n.). Nine new species: Rasilinus bicolor sp. n., Rasilinus bifurcatus sp. n., Rasilinus bimaculatus sp. n., Rasilinus grandidens sp. n., Rasilinus longulus sp. n., Rasilinus subgemellus sp. n., Rasilinus subnodulus sp. n., Rasilinus tchambicus sp. n., Rasilinus virgatus sp. n. are described from New Caledonia. Illustrations of the external morphology, male and female terminalia, dorsal habitus colour photographs of the adults, key to species and distribution map of the new genus Rasilinus are provided.

The alfalfa weevil (Hypera postica(Gyllenhal): Coleoptera: Curculionidae) is an invasive pest of alfalfa (Medicago sativaLinnaeus; Fabaceae) in the Holarctic region. Across the wide geographic distribution of the species different local adaptions have been observed. Further, several distinct mitochondrial lineages have been discovered, which have been treated as western and Egyptian/eastern strains. However, our knowledge of the biogeography ofH. posticais largely limited to North American and Japanese populations. We sampled the species from four locations in Iran and two countries in Europe (Poland and Czech Republic) and sequenced the mitochondrial COI gene to detect patterns of mitochondrial DNA divergence among Iranian and European strains. We detected two groups separated by a deep molecular split, even justifying the distinction of two molecular operational taxonomic units. Joint analyses with previously published sequences suggest that the European samples may represent the western strain, whereas both the western and Egyptian/eastern strains can be found in Iran. This distribution of genetic lineages may be the result of human-mediated translocations. The directionality, however, cannot be inferred. Our data suggest that translocations may have led to the coexistence of multipleH. posticalineages in some parts of the world,e.g., Iran. We discuss these findings against the background of the current taxonomy ofH. postica.

Crioceris quinquepunctata is a European leafbeetle, rare and strictly associated with steppe-like habitats in Central and Eastern Europe. We sampled suitable localities in Central East Europe to determine the current distribution and to verify whether populations isolated by the Carpathian Mountains (within Pannonian and Pontic area) show genetic differentiation. Sequences from the beetle COI and ITSl were amplified and compared. Furthermore, ftsZ and hcpA genes of the endosymbiont Wolbachia were analysed as additional genetic markers. We found only two populations of C. quinquepunctata (in Moravia and Podolian Upland). Unusually low genetic differences between these populations were revealed, which is in contrast to previous studies on other steppe beetles. The reasons for such low diversity are speculative and probably related to recent natural expansion or man-made translocation of C. quinquepunctata.

Fragmentation and isolation may have detrimental effects on the viability of populations. Xerothermic grasslands, which are extra-zonal analogues of steppes, are among Europe's most endangered natural environments. Information about connectivity between the remaining habitat patches is critical for effective conservation planning. However, very little is known about the actual levels of isolation of individual xerothermic patches at the inter-regional and local scale. In this study, 16 microsatellite loci were used to investigate genetic diversity, structuring, effective sizes (Ne), and connectivity among populations of the weevil Centricnemus leucogrammus at various geographic scales. Analysis of 22 populations from three geographic regions, differing in the time of origin and the degree of geographic isolation of the populations, showed that the isolated populations from northern Poland showed the least variability, whereas those from central Poland and Ukraine sustained a similar level of microsatellite variability and were moderately connected within regions. Very low effective sizes (Ne = 14–44) were estimated, and signatures of bottlenecks were detected in all populations. Effective sizes were generally higher in larger xerothermic patches, which constitute a good quality habitat, and in populations sampled in nature reserves, as opposed to unprotected turfs. Such information is useful for the design of conservation and management plans. As C. leucogrammus is characteristic of xerothermic and steppic habitats in central and eastern Europe, it may be considered an excellent target species for assessing the quality of xerothermic habitats as well as estimating and monitoring their connectivity.

Among Crioceris leaf-beetles, the two most widespread species (Crioceris asparagi and C. duodecimpunctata) are serious invasive plant pests, while another two (C. quatuordecimpunctata and C. quinquepunctata) are rare species restricted to steppe-like habitats in Eurasia. The aim of the research was to check the genetic distinctiveness of these four species and develop barcodes for their molecular identification using the mitochondrial Cytochrome Oxidase I (COI) gene and two nuclear markers: Elongation Factor 1-α (EF1-α) and Internal Transcribed Spacer 1 (ITS1). The identification of each species was possible and reliable with the use of COI and ITS1 markers. EF1-α was omitted in analyses due to its high level of heterozygosity (presence of multiple PCR products). C. duodecimpunctata and C. quatuordecimpunctata were shown to be sister taxa, but the similar genetic distances between all of the species indicate that these species originated almost simultaneously from a common ancestor. Identification of two separate clades in populations of C. quatuordecimpunctata suggested that the clades are isolated and can be considered as separate conservation units.

DNA barcoding facilitates many evolutionary and ecological studies, including the examination of the dietary diversity of herbivores. In this study, we present a survey of ecological associations between herbivorous beetles and host plants from seriously threatened European steppic grasslands. We determined host plants for the majority (65%) of steppic leaf beetles (55 species) and weevils (59) known from central Europe using two barcodes (trnL and rbcL) and two sequencing strategies (Sanger for mono/oligophagous species and Illumina for polyphagous taxa). To better understand the ecological associations between steppic beetles and their host plants, we tested the hypothesis that leaf beetles and weevils differ in food selection as a result of their phylogenetic relations (within genera and between families) and interactions with host plants. We found 224 links between the beetles and the plants. Beetles belonging to seven genera feed on the same or related plants. Their preferences were probably inherited from common ancestors and/or resulted from the host plant's chemistry. Beetles from four genera feed on different plants, possibly reducing intrageneric competition and possibly due to an adaptation to different plant chemical defences. We found significant correlations between the numbers of leaf beetle and weevil species feeding on particular plants for polyphagous taxa, but not for nonpolyphagous beetles. Finally, we found that the previous identifications of host plants based on direct observations are generally concordant with host plant barcoding from insect gut. Our results expand basic knowledge about the trophic relations of steppic beetles and plants and are immediately useful for conservation purposes.