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Among terrestrial orchids, and particularly among the subtribe Orchidinae, flies are underrepresented as pollinators. The European Neotinea ustulata, which developed specialized pollination by tachinid flies, is known to produce high relative concentrations of the floral cuticular alkenes (Z)-11-tricosene and (Z)-11-pentacosene (referred to as (Z)-11-C23/C25enes), which seem to be uncommon among orchid flowers. If the evolution of tachinid pollination is related to that of (Z)-11-C23/C25enes, we can expect that closely related species have a different floral chemical pattern and significantly small or no production of (Z)-11-C23/C25enes, independently of their pollinator guild identity (e.g., bees, flies, moths). We chemically compared the floral cuticular composition among Neotinea species, performed electrophysiological analyses, reconstructed the phylogenetic Orchidinae tree, and identified the evolutionary history of pollinator guild and (Z)-11-C23/C25enes production within the Orchidinae. Neotinea ustulata has evolved a markedly different floral cuticular composition compared to other Neotinea and produces both compounds ((Z)-11-C23/C25enes) in high relative quantities (i.e., above 8% in combination), which are detectable by tachinid antennae. Moreover, most Orchidinae taxa have minimal or no production of these alkenes, independently of the identity of their pollinator guild. Our ancestral reconstruction suggested that (Z)-11-C23/C25enes production was an evolutionary exaptation in Neotinea, whereas tachinid pollination was a unique evolutionary innovation for N. ustulata. Floral cuticular composition and, in particular, the combined production of (Z)-11-C23/C25enes at relatively high concentrations is intimately linked to the evolution of tachinid pollination within the Orchidinae.

Schall et al. (2020) assessed how a combination of different forest management systems in managed forest landscapes dominated by European beech may affect the biodiversity (alpha, beta and gamma) of 14 taxonomic groups. Current forest policy and nature conservation often demand for combining uneven-aged managed and unmanaged, set-aside for nature conservation, beech forests in order to promote biodiversity. In contrast to this, Schall et al. (2020) found even-aged shelterwood forests, represented by different developmental phases, to support highest regional (gamma) diversity. By pointing out that unmanaged forests included in our study are not old-growth forests, Bruun and Heilmann-Clausen (2021) challenge our conclusion as not providing sound scientific advice to societies. It is true that the studied unmanaged forests are not representing old-growth forests as defined in the literature. However, we demonstrate the representativeness of our unmanaged forests for current beech forest landscapes of Central Europe, where managed forests were more or less recently set-aside in order to develop old-growth structures. We also show that the managed and recently unmanaged forests in our study already differ distinctively in their forest structures. We use this response to stress the role of forest reserves for promoting certain species groups, and to emphasise their importance as valuable research sites today and in the future. Synthesis and applications. We see two main conclusions from our study. First, unmanaged forests still matter. We agree with Bruun and Heilmann-Clausen (2021) on the general importance of unmanaged, old-growth or long-untouched forests, and we do not question the importance of set-aside forests for biodiversity conservation. However, a complete complementarity to managed systems may only reveal after many decades of natural development. Second, safeguarding biodiversity in largely managed forest landscapes should focus on providing a landscape matrix of different developmental phases with varying environmental conditions rather than on maximising the vertical structure within stands. Such landscapes can partly compensate for structures that are still missing in vital, dense and closed forests recently set-aside or for unsuitable phases that may occur due to a cyclic synchronisation of forest structures in unmanaged forests. © 2021 The Authors. Journal of Applied Ecology published by John Wiley & Sons Ltd on behalf of British Ecological Society

In eusocial insects, chemical communication is crucial for mediating many aspects of social activities, especially the regulation of reproduction. Though queen signals are known to decrease ovarian activation of workers in highly eusocial species, little is known about their evolution. In contrast, some primitively eusocial species are thought to control worker reproduction through physical aggression by the queen rather than via pheromones, suggesting the evolutionary establishment of chemical signals with more derived sociality. However, studies supporting this hypothesis are largely missing. Socially polymorphic halictid bees, such as Halictus rubicundus, with social and solitary populations in both Europe and North America, offer excellent opportunities to illuminate the evolution of caste-specific signals. Here we compared the chemical profiles of social and solitary populations from both continents and tested whether (i) population or social level affect chemical dissimilarity and whether (ii) caste-specific patterns reflect a conserved queen signal. Our results demonstrate unique odor profiles of European and North American populations, mainly due to different isomers of n-alkenes and macrocyclic lactones; chemical differences may be indicative of phylogeographic drift in odor profiles. We also found common compounds overproduced in queens compared to workers in both populations, indicating a potential conserved queen signal. However, North American populations have a lower caste-specific chemical dissimilarity than European populations which raises the question if both use different mechanisms of regulating reproductive division of labor. Therefore, our study gives new insights into the evolution of eusocial behavior and the role of chemical communication in the inhibition of reproduction., publishedVersion

Floral innovations are key for pollinator specialization and play a significant role in plant diversification. Orchidaceae present many examples of floral innovations that allow its high degrees of pollinator specialization and promoted speciation. The rich neotropical genus Telipogon evolved an uncinate (=hook-like) viscidium on the pollinarium and setae-bearing flowers, which are uncommon in Orchidaceae; however, the importance of them on pollination success and whether they are floral innovations or exaptations in Telipogon have not been investigated. Here we investigate the morphology of the viscidium and floral setae within the Telipogon alliance (including the genera Hofmeisterella, Trichoceros and Telipogon), test their significance in pollination, and their occurrence and evolution across the Oncidiinae. We used Telipogon peruvianus as a model species to test whether uncinate viscidium and floral setae increased pollination success compared with a cochleariform (=spoon-like) viscidium and lack of floral setae condition. We show that the uncinate viscidium is a synapomorphy for Telipogon; setae-bearing flowers are not universally found among all species of Telipogon and evolved once in the Telipogon alliance. Furthermore, Telipogon peruvianus flowers with an uncinate viscidium have achieved higher pollinia export than those with cochleariform viscidium (ancestral condition), whereas flowers with setae have both higher success in pollinia removal, although not significant, and pollinia deposition than those lacking of setae (ancestral condition). We demonstrate that uncinate viscidia and floral setae in Telipogon are a key innovation and exaptation, respectively, that enhance pollination success and they might act as drivers of diversification and pollinator specialization in this genus.

Telipogon peruvianus is a highly restricted, sexually deceptive, Neotropical orchid species, endemic to the southern Peruvian Andes. It is only known from two localities, which are affected by anthropogenic disturbance. Here, we investigated whether the restricted distribution of T. peruvianus has led to low genetic diversity and inbreeding, thus threatening its survival. We isolated 10 novel microsatellite loci specific for T. peruvianus (and that also cross-amplified in related species) from two adjacent populations. We found that genetic diversity within populations was only moderately reduced, along with some evidence of inbreeding. We found low levels of genetic differentiation, suggesting connectivity by pollen/seed flow between the two populations. Effective population size was comparable to the real number of flowering individuals in the populations and we did not detect the signature of recent bottlenecks. Taken together, these results show that, despite increasing anthropogenic pressure, the two investigated populations of T. peruvianus still host valuable genetic diversity that should be preserved through appropriate conservation strategies.

Chemical communication plays a fundamental role in many aspects of an animal’s life from assessing habitat quality to finding mating partners. Behavioural observations show that chemical communication likewise plays an important role in spiders, but the contexts and the substances involved are little explored. Here, we investigate the chemical communication in the garden cross spider Araneus diadematus (Clerck, 1757) between and within the sexes. Using choice trials, we demonstrate that males are attracted to odours of adult females, but not to those of subadult females. Our data further suggest that adult females avoid odours of conspecific adult females, possibly in order to reduce reproductive competition with other females. Cuticle and silk extracts as well as headspace samples of subadult and adult virgin females were analysed via GC–MS. Available candidate compounds for the female sex pheromone were tested via electroantennography on palps (electropalpography) of adult virgin females and on females in behavioural trials. We propose sulcatone (6-methyl-5-hepten-2-one) as a candidate substance for the female volatile pheromone and several long-chained alkanes and alcohols as candidates for contact pheromones. Apart from demonstrating that attraction of males to females depends on the latter’s developmental stage, our study suggests that pheromones can also play an important role between females, an aspect that requires further attention. Supplementary Information The online version contains supplementary material available at 10.1007/s00114-021-01747-9.

Visual and olfactory floral signals play key roles in plant-pollinator interactions. In recent decades, studies investigating the evolution of either of these signals have increased considerably. However, there are large gaps in our understanding of whether or not these two cue modalities evolve in a concerted manner. Here, we characterized the visual (i.e., color) and olfactory (scent) floral cues in bee-pollinated Campanula species by spectrophotometric and chemical methods, respectively, with the aim of tracing their evolutionary paths. We found a species-specific pattern in color reflectance and scent chemistry. Multivariate phylogenetic statistics revealed no influence of phylogeny on floral color and scent bouquet. However, univariate phylogenetic statistics revealed a phylogenetic signal in some of the constituents of the scent bouquet. Our results suggest unequal evolutionary pathways of visual and olfactory floral cues in the genus Campanula. While the lack of phylogenetic signal on both color and scent bouquet points to external agents (e.g., pollinators, herbivores) as evolutionary drivers, the presence of phylogenetic signal in at least some floral scent constituents point to an influence of phylogeny on trait evolution. We discuss why external agents and phylogeny differently shape the evolutionary paths in floral color and scent of closely related angiosperms., publishedVersion

Despite increased focus on elucidating the various reproductive strategies employed by orchids, we still have only a rather limited understanding of deceptive pollination systems that are not bee- or wasp-mediated. In Europe, the orchid Neotinea ustulata has been known to consist of two phenologically divergent varieties, neither of which provide rewards to its pollinators. However, detailed studies of their reproductive biology have been lacking. Our study aimed to characterize and understand the floral traits (i.e., morphology, color, and scent chemistry) and reproductive biology of N. ustulata. We found that the two varieties differ in all their floral traits; furthermore, while Neotinea ustulata var. ustulata appears to be pollinated by both bees (e.g., Anthophora, Bombus) and flies (e.g., Dilophus, Tachina), var. aestivalis is pollinated almost entirely by flies (i.e., Nowickia, Tachina). Tachinids were also found to be much more effective than bees in removing pollinaria, and we show experimentally that they use the characteristic dark inflorescence top as a cue for approaching inflorescences. Our results thus suggest that while both N. ustulata varieties rely on tachinids for pollination, they differ in their degree of specialization. Further studies are, however, needed to fully understand the reproductive strategy of N. ustulata varieties., publishedVersion

Different enantiomers of chiral compounds within floral perfumes usually trigger distinct responses in insects; however, this has frequently been neglected in studies investigating semiochemicals in plant-pollinator interactions. Approximately 1000 neotropical plants produce floral perfumes as the only reward for pollinators, i.e. male euglossine bees. The chiral compound carvone epoxide is a key component of the scent bouquet of many perfume-rewarding plants that are pollinated by males of Eulaema. Here, we tested the biological activity of the four carvone epoxide stereoisomers to four Eulaema species occurring in the Atlantic Rainforest of NE-Brazil. We determined the stereochemistry of carvone epoxide in the floral scent of several Catasetum species, tested whether the antennae of bees respond differentially to these stereoisomers and investigated if there is a behavioural preference for any of the stereoisomers. We found that 1) Catasetum species emit only the (-)-trans-stereoisomer of carvone epoxide, 2) for E. atleticana and E. niveofasciata antennal responses to the (-)-trans-carvone epoxide were significantly stronger than those to (-)-cis-carvone epoxide, 3) the strength and pattern of antennal responses to all 4 stereoisomers (separately tested) did not differ among Eulaema species, and 4) there were significant differences in attractiveness of the four stereoisomers to the bees species with the (-)-trans-stereoisomer being particularly attractive. We assume (-)-trans-carvone epoxide to be the dominant isomer in perfume-rewarding plants pollinated by Eulaema. The universal occurrence of carvone epoxide in Catasetum species pollinated by Eulaema, suggests that this compound has evolved in perfume-rewarding as a specific attractant for Eulaema bees as pollinators.

Although common among orchids, pollination by perfume-gathering male euglossine bees is quite rare in other Neotropical families. In Gesneriaceae, for example, it is reported in two genera only, Drymonia and Gloxinia. Flowers of G. perennis are known to emit perfume, thereby attracting male euglossine bees as pollinators. However, detailed reports on the pollination ecology, as well as on chemistry of floral perfume of individuals in natural populations, are still missing. In this study, we report on the pollination ecology of G. perennis, focusing on the ecological significance of its floral perfume. In natural populations in Peru, we documented the floral biology and breeding system of G. perennis, as well as its interaction with flower visitors. We also characterised the chemical composition of floral perfume, as well as its timing of emission. Gloxinia perennis is self-compatible and natural pollination success is high. Spontaneous self-pollination occurs as a 'just in case strategy' when pollinators are scarce. Perfume-collecting males of Eulaema cingulata and El. meriana were identified as pollinators. The perfume bouquet of G. perennis consists of 16 compounds. (E)-Carvone epoxide (41%) and limonene (23%) are the major constituents. Perfume emission is higher at 09:00 h, matching the activity peak of Eulaema pollinators. Flowers of G. perennis have evolved a mixed strategy to ensure pollination (i.e. self- and cross-pollination), but cross-pollination is favoured. The size and behaviour of Eulaema males enables only these bees to successfully cross-pollinate G. perennis. Furthermore, G. perennis floral perfume traits (i.e. chemistry and timing of emission) have evolved to optimise the attraction of these bees.

Research into vibrational communication is becoming increasingly important as we discover more species using vibrational signals in different types of behavior. Males of the solitary red mason bee, Osmia bicornis, are known to use vibrations in female choice, where the females not only evaluate a male’s fitness through their vibrational signal but also use them to distinguish between males of different origins. This was shown conclusively via bioassays, in which females from Germany rejected English males unless they were imbued with an artificial German signal and vice versa. However, an investigation into which parameters of the signal might differ between species and populations has been lacking so far. We therefore recorded O. bicornis as well as O. cornuta males from Germany, England, and Denmark using a laser vibrometer and analyzed the recordings using the software Spike to measure frequency, modulation range, and average pulse duration in each signal. Our results clearly showed significant differences in the signal between O. bicornis and O. cornuta males in all three parameters measured. O. bicornis populations from Germany and Denmark surprisingly also differed significantly in frequency and modulation range, with England lying in between the two. We believe that the females probably use another signal parameter that we have not evaluated yet to make their mate choice. This leaves us with the question of why the observed signal parameters differ between Germany and Denmark. From our knowledge about the system to date, we believe that we are looking at the first steps of speciation in this system and further study could help us with new insights into evolutionary processes in the future.

Rove beetles (Coleoptera: Staphylinidae) are one of the most important colonizers of vertebrate cadavers. We have previously investigated carrion-associated rove beetle communities across various forests and demonstrated that the study regions are the main drivers of the local rove beetle species pool that is, in turn, able to colonize a carcass. Nevertheless, little is known about their temporal variation in community composition during the decomposition process. The aim of our study has been to analyze the temporal changes of the composition of the rove beetle community and to identify new, potentially useful candidate species for forensic entomological evaluations. We determined 80 rove beetle species that were attracted to 60 piglet cadavers across various forest stands in Germany. Both the abundance and the community composition changed over the decomposition process, independently of the species-specific geographical variation across study regions. In the region Schorfheide-Chorin, species from the genus Philonthus proved to be a suitable group for future forensic entomological investigations. They appeared in markedly high numbers at piglet cadavers from the bloated stage until the advanced decay stage. For the study region Schwabische Alb, we newly describe the species Omalium septentrionis as a member of the carrion-associated rove beetle fauna. Based on the geographical variation in rove beetle community compositions, we have filtered out several species that might contribute to advances in postmortem interval estimations or other applied fields in forensic entomological contexts.

Reduced water availability can cause physiological stress in plants that affects floral development leading to changes in floral morphology and traits that mediate interactions with pollinators. As pollinators can detect small changes in trait expressions, drought-induced changes in floral traits could affect pollinator visitations. However, the linkage between changes in floral traits and pollinator visitations under drought conditions is not well explored. We, therefore, tested whether drought-induced changes in floral morphology and abundance of flowers are linked to changes in pollinator visitations. We conducted flight cage experiments with a radio frequency identification system for automated visitation recordings with bumble bees (Bombus terrestris) and common charlock (Sinapis arvensis) as the model system. In total, we recorded interactions for 31 foraging bumble bees and 6569 flower visitations. We found that decreasing soil moisture content correlated with decreasing size of all measured morphological traits except stamen length and nectar tube width. The reductions in floral size, petal width and length, nectar tube depth and number of flowers resulted in decreasing visitation rates by bumble bees. These decreasing visitations under lower soil moisture availability might be explained by lower numbers of flowers and thus a reduced attractiveness and/or by increased difficulties experienced by bumble bees in handling smaller flowers. Whether these effects act additively or synergistically on pollinator behaviour and whether this leads to changes in pollen transfer and to different selectionp ressures require further investigation., Under dry conditions, plants can reduce their flower number and size to save water. Such changes, however may lead to reduced pollinator visitations as these traits play an important role in pollinator attraction. In this study, we explored how drought affects pollinator visitation via changes in floral traits. Therefore, we conducted a flight cage experiment with an automated visitation recording system observing bumblebee interactions with drought stressed and well-watered plants. Our results show that decreasing flower number and size under drought conditions resulted in decreasing bumblebee visits. This indicates that drought-stressed plants become less attractive for pollinators.

Intensification of anthropogenic land use is a major threat to biodiversity and thus to essential ecosystem services provided by insects. Rove beetles (Coleoptera: Staphylinidae), which react sensitively to habitat changes, are species-rich colonizers of vertebrate cadavers and contribute to the important ecosystem service of carrion decomposition. The unveiling of anthropogenic and environmental drivers that modify carrion-associated rove beetle communities should improve our understanding of the plasticity of cadaver decay. We report the presence of 80 rove beetle species on 65 decomposing piglet cadavers at forest sites characterized by a gradient of management intensity across three geographic regions in Germany. Local and landscape drivers were revealed that shape beetle abundance, diversity, and community composition. Forest management and regions affect rove beetle abundance, whereas diversity is influenced by local habitat parameters (soil pH, litter cover) and regions. The community composition of rove beetles changes with management intensification by promoting generalist species. Regarding single species, Philonthus decorus and Anotylus mutator are linked to unmanaged forests and Ontholestes tessellatus to highly used forest stands. The spatial information provided about carrion-associated rove beetle communities in German forests is not only of carrion-ecological but also of forensic entomological interest., publishedVersion

Some mixotrophic plants from temperate forests use the mycorrhizal fungi colonizing their roots as a carbon source to supplement their photosynthesis. These fungi are also mycorrhizal on surrounding trees, from which they transfer carbon to mixotrophic plants. These plants are thus reputed difficult to transplant, even when their protection requires it. Here, we take profit of a successful ex situ pot cultivation over 1 to 3 years of the mixotrophic orchid Epipacis helleborine to investigate its mycorrhizal and nutrition status. Firstly, compared with surrounding autotrophic plants, it did not display the higher N content and higher isotopic (13C and 15N) abundance that normally feature mixotrophic orchids because they incorporate N-, 13C-, and 15N-rich fungal biomass. Second, fungal barcoding by next-generation sequencing revealed that the proportion of ectomycorrhizal fungi (expressed as percentage of the total number of either reads or operational taxonomic units) was unusually low compared with E. helleborine growing in situ: instead, we found a high percentage of rhizoctonias, the usual mycorrhizal partners of autotrophic orchids. Altogether, this supports autotrophic survival. Added to the recently published evidence that plastid genomes of mixotrophic orchids have intact photosynthetic genes, this suggests that at least some of them have abilities for autotrophy. This adds to the ecological plasticity of mixotrophic plants, and may allow some reversion to autotrophy in their evolution.

Although some spiders are globally invasive, found at high densities, and may be considered pests (particularly those that are toxic to humans), there are few pest management methods based on experimental data. 'Common wisdom' and advertisements on internet websites assert that a number of natural substances repel spiders. We tested whether the three substances cited most frequently (lemon oil, peppermint oil, and chestnut-fruits) effectively repelled female spiders or whether these were myths. We presented each of the putative repellents versus a control in a two-choice assay and tested responses of females of three invasive spider species in two different families: theridiids, Latrodectus geometricus C. L. Koch (Araneae: Theridiidae) and Steatoda grossa C. L. Koch (Araneae: Theridiidae) and the araneid, Araneus diadematus Clerck . Chestnuts (Araneae: Araneidae) and mint oil strongly repelled L. geometricus and A. diadematus. S. grossa was less sensitive to these chemicals but had a slight tendency to avoid chestnuts. However, lemon oil, the substance most likely to be cited as a repellent (over 1,000,000 hits on Google), had no effect on any of these spiders. We conclude that volatiles released by mint oil and chestnuts may be effective in deterring spider settlement in two different families of spiders, but lemon oil as a repellent is a myth.

The potential high pollinator specificity and the mosaic of highly differentiated potential habitats in the Mediterranean have probably fueled species diversification in the sexually deceptive genus Ophrys. Several endemic species have been described whose taxonomic status remains questionable. By using a combination of morphological, chemical, behavioural and genetic analyses, we have investigated the taxonomic status of O. vallesiana, a presumed endemic species of the O. iricolor/eleonorae group in Tunisia. We have found that O. eleonorae and O. vallesiana produce similar odour bouquets, are attractive to the same pollinators, have overlapping flower morphology and are genetically indistinct. Thus, all experimental evidence supports the conclusion that, in spite of the observed morphological differences in plant height and flower number, the local endemic O. vallesiana clusters together with the widespread and co-occurring O. eleonorae. Concomitantly, our results exclude a putative origin of O. vallesiana as a hybrid taxon between O. eleonorae and O. gazella, even if hybridization between these two species can occur locally. Our study demonstrates the need for accurate and multidisciplinary investigations for the proper taxonomic delimitation of local endemics and highlights the complexity of the evolutionary dynamics in Ophrys species complexes.

Many species of the sexually deceptive genus Ophrys are characterized by insect-like flowers. Their form has been traditionally considered to play an important role in pollinator attraction and manipulation. Yet, the evolution of the floral form remains insufficiently understood. We hypothesize that pollinator-mediated selection is essential for driving floral form evolution in Ophrys, but that form components are being subjected to varying selection pressures depending on their role in mediating interactions with pollinators. By using the Eucera-pollinated Ophrys leochroma as a model, our aim has been to assess whether and in what manner pollination effectiveness is altered by experimental manipulation of the flower form. Our results show that floral form plays an essential and, so far, underestimated role in ensuring effective pollination by mechanically guiding pollinators towards the reproductive structures of the flower. Pollinators are significantly less effective in interacting with flowers having forms altered to resemble those of species pollinated by different hymenopteran genera. Further, those components used by pollinators as gripping points were found to be more effective in ensuring pollinia transfer than those with which pollinators do not directly interact. Thus, mechanically active and inactive components appear to be under different selection pressures. As a consequence, mechanically active components of the flower form could reflect adaptations to the interaction with particular pollinator groups, whereas mechanically inactive components can vary more freely. Disentangling selection patterns between the functionally different components of flower form may provide valuable insights into the mechanisms driving the morphological diversification of sexually deceptive pollination systems.

Pheromones play an important role in the mating behavior of many insects, especially in bees, and it has recently been shown that female Osmia bicornis use the odor bouquet found on male antennae in female choice. Nevertheless, the glandular origin of the substances found on the male antennae is so far unknown. In this study, we investigated if antennal glands exist in males and if they are the glandular source of substances on the antennae’s cuticle, as has been suggested by previous studies. Since electron and light microscopy revealed no such glands, we investigated further possible places of origin for the antennal compounds by conducting chemical analyses of solvent extracts of the male abdominal surface and the antennae, as well as of two glands found in the bee’s head, the labial and mandibular gland. Our results clearly show that the substances found on the abdominal surface and in solvent extracts of the antennae differ significantly in relative amounts. Therefore, the compounds on the antennae must have additional sources. Since the relative amounts of compounds on the surface of antennae are different from the blends on the mandibular gland or the labial gland, neither one of them can be the sole source of the antennal compounds. Hence we believe another glandular source is also involved in the production of the bouquet on the antennae. Especially interesting were high amounts of sterols on the antennae, which also play a role in population differences. This sheds new light on the importance of the antennal compounds and also the importance sterols might play in communication.

Climate change and the resulting changes in air temperature are known to have a major influence on most animals, especially poikilothermic insects, because they depend on the high enough temperatures to function. Previous studies have shown that various signals can be affected by changes in temperature. However, research into the effect of temperature on mating signals and subsequently communication between mates and on female choice is still rare. In the red mason bee, Osmia bicornis, which emerges early in spring and is therefore subject to extensive temperature changes, females choose suitable males based on, among other criteria, their thorax vibrations and odor. So far there has been no research into the effect of temperature changes on these signals. We therefore investigated whether the environmental temperature has an influence on the male's mating signals by measuring vibrations using a laser vibrometer and collecting male and female odor at different temperatures. Furthermore, we performed behavioral experiments in order to show whether there is an effect of temperature‐induced changes and female choice. Our results showed that rejected males differed in their vibrations between the two temperature settings but accepted ones did not. Temperature changes therefore seem to have a stronger effect on those males that are rejected by the females, whereas the accepted males are the ones that can produce desirable signals despite temperature fluctuations. Furthermore, we found that the differences in odor profiles were greater between temperature settings than between the sexes and that females change their preference for odor with temperature. We conclude that temperature strongly influences the male mating signals and therefore may have a major impact on sexual selection in this species. This is an important aspect to consider, not only in future studies on mating behavior, but also in view of our ever raising temperatures.

Floral traits may function as attractants for some flower visitors and also as mechanisms that filter out non-beneficial flower visitors. Our current understanding of floral filter mechanisms is limited as this particular aspect of floral function has received less attention than floral attractants. Here we report on a case of two closely related Gomphocarpus species, G. physocarpus and G. fruticosus, that are similar in their general floral shape and architecture but differ in their main pollinators, which are vespid wasps and honeybees, respectively. To identify the floral traits that best explain the observed pollinators we analysed and compared the floral scent composition, the flower colour, flower morphology, and nectar sugar composition of the two species. Furthermore, the response to floral scent was tested in behavioural experiments with Belanogaster and Polistes vespid wasps. The floral scent composition differed significantly between the plant species. Acetic acid was a main component in the wasp-pollinated G. physocarpus, which is highly unusual for a floral scent bouquet. Choice experiments suggest that the species-specific odour might be responsible for the selective attraction of either wasps or bees. Furthermore, wasp-pollinated G. physocarpus flowers offer relatively large amounts of nectar that is freely accessible to insect visitors, whereas in G. fruticosus the site of nectar accumulation is covered by a specific morphological structure that makes it difficult for short-tongued insects such as wasps to access the nectar. Altogether, pollinator specialisation in the two Gomphocarpus species seem to be due to a combination of floral filter mechanisms and olfactory cues that cause flower visitors to learn to differentiate between the two species.

Ophrys flowers mimic sex pheromones of attractive females of their pollinators and attract males, which attempt to copulate with the flower and thereby pollinate it. Virgin females and orchid flowers are known to use the same chemical compounds in order to attract males. The composition of the sex pheromone and its floral analogue, however, vary between pollinator genera. Wasp-pollinated Ophrys species attract their pollinators by using polar hydroxy acids, whereas Andrena-pollinated species use a mixture of non-polar hydrocarbons. The phylogeny of Ophrys shows that its evolution was marked by episodes of rapid diversification coinciding with shifts to different pollinator groups: from wasps to Eucera and consequently to Andrena and other bees. To gain further insights, we studied pollinator attraction in O. leochroma in the context of intra- and inter-generic pollinator shifts, radiation, and diversification in the genus Ophrys. Our model species, O. leochroma, is pollinated by Eucera kullenbergi males and lies in the phylogeny between the wasp and Andrena-pollinated species; therefore, it is a remarkable point to understand pollinator shifts. We collected surface extracts of attractive E. kullenbergi females and labellum extracts of O. leochroma and analyzed them by using gas chromatography with electroantennographic detection (GC-EAD) and gas chromatography coupled with mass spectrometry (GC-MS). We also performed field bioassays. Our results show that O. leochroma mimics the sex pheromone of its pollinator's female by using aldehydes, alcohols, fatty acids, and non-polar compounds (hydrocarbons). Therefore, in terms of the chemistry of pollinator attraction, Eucera-pollinated Ophrys species might represent an intermediate stage between wasp- and Andrena-pollinated orchid species.

Studying the relationship between parental and mating effort helps us to understand the evolution of parental care and, consequently, has been the subject of many theoretical and empirical investigations. Using burying beetles as a model, we found no correlation between the intensity of a sexual signal (sex pheromone quantity) and the amount of care provided by males. However, males that were given the opportunity to breed and care for young went on to produce a higher amount of their sexual signal and attracted three times more females in the field than control males that were not given the opportunity to breed. The likely explanation for our finding is that specific aspects of care in burying beetles, that is the defense and preservation of a nutrient rich breeding resource, a small vertebrate cadaver, is not only beneficial for the offspring but also for the adults themselves. Obtaining a good carrion meal possibly enables males to store resources that they can subsequently allocate toward sexual signaling. Collectively, our results highlight that conditions can exist where male participation in brood care has a positive effect on its sexual attractiveness. This in turn might have facilitated the evolution of male assistance in parental care.

The pollen diet provided by adult bees to their offspring varies immensely. While some species collect pollen on several plants irrespective of their phylogenetic relatedness (polyleges), others collect only on plants within a genus or family (oligoleges). Floral scents play a central role in bee-plant interactions. To locate flowers, polyleges are assumed to rely on compounds commonly found as floral scent constituents, whereas oligoleges rely on unusual compounds to recognize host flowers unambiguously. Campanula flowers are visited by both polylectic and oligolectic species, and their scent bouquets consist of common and unusual (e.g., spiroacetals) volatiles. In a comparative approach, we performed electroantennographic analyses to investigate the antennal responses of three polyleges and three oligoleges to three common volatiles and four spiroacetals. We hypothesized that: 1) oligoleges and polyleges should respond similarly to common flower volatiles, and 2) Campanula oligoleges should be more sensitive to spiroacetals than are polyleges. In corroboration, we found that antennal sensitivity to common volatiles was similar among bees irrespective of pollen diet, whereas oligoleges of Campanula were more sensitive to spiroacetals than polyleges. Newly emerged bees of the Campanula oligolege Chelostoma rapunculi rely on spiroacetals for recognizing host-flowers, and our results suggest that this might also be true for other Campanula oligoleges, since Chelostoma campanularum and Hoplitis mitis also were able to perceive these specific volatiles at very low concentrations. Together, our results provide interesting insights into the significance of olfactory adaptations in oligolectic and polylectic bee species.

1. Forest management greatly influences biodiversity across spatial scales. At the landscape scale, combining management systems that create different stand properties might promote biodiversity due to complementary species assemblages. In European beech forests, nature conservation and policy advocate a mixture of unmanaged (UNM) forests and uneven-aged (UEA) forests managed at fine spatial grain at the expense of traditionally managed even-aged shelterwood forests (EA). Evidence that such a landscape composition enhances forest biodiversity is still missing. 2. We studied the biodiversity (species richness 0D, Shannon diversity 1D, Simpson diversity 2D) of 14 taxonomic groups from bacteria to vertebrates in ‘virtual’ beech forest landscapes composed of varying shares of EA, UEA and UNM and investigated how γ-diversity responds to landscape composition. Groups were sampled in the largest contiguous beech forest in Germany, where EA and UEA management date back nearly two centuries, while management was abandoned 20–70 years ago (UNM). We used a novel resampling approach that created all compositional combinations of management systems. 3. Pure EA landscapes preserved a maximum of 97.5% γ-multidiversity (0D, 1D) across all taxa. Pure and mixed UEA/UNM landscapes reduced γ-multidiversity by up to 12.8% (1D). This effect was consistent for forest specialists (1D: −15.3%). We found only weak complementarity among management systems. 4. Landscape composition significantly affected γ-diversity of 6–9 individual taxa, depending on the weighting of species frequencies with strongest responses for spiders, beetles, vascular plants and birds. Most showed maximum diversity in pure EA landscapes. Birds benefited from UNM in EA-dominated landscapes. Deadwood fungi showed highest diversity in UNM. 5. Synthesis and applications. Our study shows that combining fine-grained forest management and management abandonment at the landscape scale will reduce, rather than enhance, regional forest biodiversity. We found an even-aged shelterwood management system alone operating at intermediate spatial scales and providing stands with high environmental heterogeneity was able to support regional biodiversity. However, some taxa require certain shares of uneven-aged and unmanaged forests, emphasizing their general importance. We encourage using the here presented resampling approach to verify our results in forest landscapes of different composition and configuration across the temperate zone., publishedVersion

Summary Eusociality is characterized by the reproductive division of labor between two castes: fertile queens and largely sterile workers. Queen pheromones are known to influence worker behavior and reproductive physiology and are therefore key components in regulating complex eusocial behavior [ 1 ]. Recent studies indicate that cuticular hydrocarbons (CHCs) act as queen pheromones in various eusocial hymenopteran species [ 2 , 3 , 4 , 5 , 6 , 7 , 8 ]. However, almost all species investigated to date are highly eusocial and do not include extant transitory stages from solitary to eusocial behavior [ 9 ]. Indeed, primitively eusocial species, which largely lack morphologically distinct castes, are thought to control worker reproduction through the physical aggression of the queen rather than via pheromones [ 10 , 11 , 12 ]. Halictid or sweat bees exhibit a high variability of eusociality including solitary and facultatively eusocial species [ 9 , 13 , 14 , 15 , 16 ]. However, the mechanisms controlling worker reproduction in these transitory species are unknown. The results of a recent correlative study based on caste-specific chemical profiles in various halictid bees of different social levels have revealed an overproduction of macrocyclic lactones in queens compared with workers [ 17 ]. Using chemical analyses and behavioral experiments in which we simulated below-ground nests of the primitively eusocial sweat bee Lasioglossum malachurum, we identified a queen pheromone and found that macrocyclic lactones, not CHCs, influence worker behavior and decrease ovarian activation in this species. Our data suggest that the evolution of queen pheromones is more complex than previously inferred from highly eusocial species and shed new light on the complexity of the evolution of queen pheromones.

The plant genus Ficus is a keystone resource in tropical ecoystems. One of the unique features of figs is the diversity of fruit traits, which in many cases match their various dispersers, the so-called fruit syndromes. The classic example of this is the strong phenotypic differences found between figs with bat and bird dispersers (color, size, and presentation). The ‘bird-fig’ Ficus colubrinae represents an exception to this trend since it attracts the small frugivorous bat species Ectophylla alba at night, but during the day attracts bird visitors. Here we investigate the mechanism by which this ‘bird-fig’ attracts bats despite its fruit traits, which should appeal solely to birds. We performed feeding experiments with Ectophylla alba to assess the role of fruit scent in the detection of ripe fruits. Ectophylla alba was capable of finding ripe figs by scent alone under exclusion of other sensory cues. This suggests that scent is the main foraging cue for Ectophylla alba. Analyses of odor bouquets from the bat- and bird-dispersal phases (i.e. day and night) differed significantly in their composition of volatiles. The combination of these two findings raises the question whether E. alba and F. colubrinae resemble a co-adaptation that enables a phenotypically classic ‘bird-fig’ to attract bat dispersers by an olfactory signal at night thus maximizing dispersal.

In sexually deceptive orchids, pollinator specificity is the main factor responsible for species isolation. The two sexually deceptive orchids Ophrys chestermanii and Ophrys normanii occur sympatrically on Sardinia and attract the same pollinator, males of Bombus vestalis, by producing the same odour bouquets. Since gene flow between these two species has been shown to be almost absent, their genetic distinctness seems to be preserved by other forms of reproductive barrier than pollinator specificity. The aim of this study is to investigate the nature and strength of these isolation barriers which apparently halt gene flow between these two orchids that share the same pollinator. Morphological measurements, combined with observations of cross-pseudocopulations, revealed that, in sympatry, the longer caudicles of O. normanii and the smaller and differently shaped stigmatic cavity in O. chestermanii are the main factors preventing gene flow between these two sympatric species. In contrast, other pre- or post-mating barriers seem to be weak or absent, as the two species have overlapping flowering time and we were able to rear hybrid offspring from interspecific manual crosses. At the same time, genetic analysis of orchid plantlets collected at sympatric sites detected the presence of only two putative hybrid individuals. Thus, the probability of natural cross-pollination between O. normanii and O. chestermanii seems to be strongly limited by divergence in their floral morphology. For some traits, this divergence was found increased between sympatric populations.

Deception of floral visitors in pollination systems is widely distributed among flowering plants. In deceptive systems, the flower (or part of it) or inflorescence mimics either a specific or an unspecific model to attract pollinators. A previous study showed that Telipogon peruvianus flowers developed sexual deception for pollination. However, it was unknown which stimuli were playing a role in pollination. Therefore, we aim to throw some light onto these questions using colour and chemical analysis and biotests. Interestingly, using spectral reflectance, we show here that the flowers present high contrast similar to that produced by a female tachinid fly sitting on a daisy inflorescence, which is used as food resource. We also tested the role of chemical signals in pollinator attraction by collecting floral and female extracts for chemical and electrophysiological analyses, and carried out behavioural tests. For biotests, various treatments, including synthetic mixtures of the electrophysiologically active compounds found in common in females and flowers, have demonstrated that T. peruvianus flowers mimic the sexual pheromone of their pollinator's females. Thus, we give evidence that T. peruvianus flowers mimic a model composed of two organisms. Our study contributes to the understanding of the evolution of deceptive pollination.

In the necrophagous burying beetle Nicrophorus vespilloides (Coleoptera: Silphidae), cadaver preference appears to depend on cadaver size and on the maturity of the beetle. We previously showed that newly emerged females with immature ovaries prefer later stages of decomposition of large cadavers. Our present aim is the determination of specific chemical compounds involved in the discrimination of cadaveric odor bouquets and the recognition of specific stages of decomposition. We used headspace samples of maggot-infested piglet cadavers at various decomposition stages and performed gas chromatography coupled with electroantennography (GC-EAD) to record 45 EAD-active compounds. Using GC coupled with mass spectrometry, we identified 13 of the EAD-active compounds. The headspace of the fresh decomposition stage was characterized mainly by high relative amounts of trimethylpyrazine. High relative amounts of dimethyl trisulfide were characteristic of bloated, post-bloating, and advanced decay stages. The advanced decay and dry remains stages were dominated by high relative amounts of phenol. Statistically, this compound had the highest impact on discrimination between the fresh decomposition stage, which is important for mature burying beetles for reproduction, and the advanced decay stage, which is nutritionally more attractive for newly emerged beetles. Phenol might, therefore, function as a key substance for newly emerged female burying beetles, so that they can locate suitable cadavers for feeding to maturation.

Chemical communication is crucial for the maintenance of colony organization in eusocial insects and chemical signals are known to mediate important aspects of their social life, including the regulation of reproduction. Sociality is therefore hypothesized to be accompanied by an increase in the complexity of chemical communication. However, little is known about the evolution of odor signals at the transition from solitary living to eusociality. Halictid bees are especially suitable models to study this question as they exhibit considerable variability in social behavior. Here we investigated whether the dissimilarities in cuticle chemical signals in females of different castes and life stages reflect the level of social complexity across halictid bee species. Our hypothesis was that species with a higher social behavior ergo obligate eusocial species possess a more distinct chemical profile between castes or female life stages. We analyzed cuticular chemical profiles of foundresses, breeding females and workers of ancestrally solitary species, facultative and obligate eusocial halictid species. We also tested whether social complexity was associated with a higher investment in chemical signals. Our results revealed higher chemical dissimilarity between castes in obligate than in facultative eusocial species, especially regarding macrocyclic lactones, which were the single common compound class overproduced in queens compared with workers. Chemical dissimilarities were independent of differences in ovarian status in obligate eusocial species but were dependent on ovarian status in facultative eusocial species, which we discuss in an evolutionary framework.

SummaryDifferences in female preference for certain male characteristics can be a driving force for population divergence and speciation [1–4]. During precopulation, females of the red mason bee, Osmia bicornis, choose suitable males based on, among other criteria, their thoracic vibrations [5]. These vibrations are thought to be a signal of a male’s fitness with females choosing the strongest males that can vibrate for the longest time [5]. The precise role of such vibrational signals, however, has not been determined by bioassays, and the vibrations might also play a role in species recognition [6]. There are two main subspecies of O. bicornis in Europe distinguishable only by a single morphological trait [7] (Figure S1). We therefore developed a new bioassay allowing us to impose the vibrations of one live male onto another in order to discern possible selective mate choice by females from O. bicornis originating from different regions of Europe. Females showed strong preference for males from their own region, and male vibrations were the main signal involved in this choice. Thus, vibrational signals encode not only fitness but also information about the region of origin indicating that divergence exists between the different European O. bicornis populations, which might ultimately lead to speciation. These results provide new insights into the scope of vibrational communication in bees, a group previously considered to rely predominantly on chemical signals [8, 9]. Our newly developed method should shed further light on many exciting questions concerning vibrational communication in bees and other animal taxa.

Catasetum is a neotropical orchid genus that comprises about 160 dioecious species with a remarkable sexual dimorphism in floral morphology. Flowers of Catasetum produce perfumes as rewards, which are collected only by male euglossine bees. Currently, floral scents are known to be involved in the selective attraction of specific euglossine species. However, sexual dimorphism in floral scent and its eventual role in the pollination of Catasetum species have never been investigated. Here, we have investigated the pollination of Catasetum uncatum and asked: (1) Is floral scent a sexual dimorphic trait? (2) Does pollinarium removal/deposition affect scent emission? (3) Does sexual dimorphism in floral scent and changed scent emission have implications with regard to the behaviour of the pollinators? The frequency and behaviour of floral visitors were observed in non-manipulated flowers (both flower sexes) and in manipulated flowers (pistillate only) in which pollinaria were deposited. Scents of staminate and pistillate flowers (both manipulated and non-manipulated) were collected by using dynamic headspace methods and analysed chemically. Electrophysiological analyses were performed to detect compounds triggering antennal depolarisation in the euglossine species. C. uncatum is pollinated mainly by males of Euglossa nanomelanotricha. Pollinators were more frequent in pistillate than in staminate inflorescences. Bees approaching staminate flowers frequently flew away without visiting them, a behavioural pattern not observed in pistillate flowers. In the chemical analyses, we recorded 99 compounds, 31 of which triggered antennal depolarisation in pollinators. Multivariate analyses with the electrophysiological-active compounds did not detect differences between the scent composition of staminate and pistillate flowers. Pollinarium removal or deposition resulted in diminished scent emission within 24 h in staminate and pistillate flowers, respectively. Surprisingly, bees discriminated pollinated from non-pollinated pistillate flowers as early as 2 h after pollination. The rapid loss in the attractiveness of flowers following pollinarium removal/deposition can be interpreted as a strategy to direct pollinators to non-pollinated flowers. We have found no evidence that euglossine males discriminate staminate from pistillate flowers by means of floral scent. Instead, we speculate that bees use visual cues, such as sex dimorphic traits, to discriminate flowers of different sexes. Together, our results provide interesting insights into the evolution of floral signals in gender-dimorphic species and into its significance in plant reproductive biology.

Life history theory predicts a trade-off between male sexual trait expression and immunocompetence. Using burying beetles, Nicrophorus vespilloides, as a model, we investigated the relationship between male immune function, sex pheromone emission, and attractiveness under field conditions. In the first experiment, we tested whether there is a positive correlation between immune capacity, sex pheromone characteristics (quantity, relative composition, and time invested in pheromone emission), and male attractiveness. As a measurement of immune capacity, we used an individual's encapsulation ability against a novel antigen. In the second experiment, we specifically examined whether a trade-off between chemical trait expression and immune function existed. To this end, we challenged the immune system and measured the subsequent investment in sex pheromone emission and the attractiveness of the male under field conditions. We found that a male's immunocompetence was neither related to the emission of the male's sex pheromone nor to its attractiveness in the field. Furthermore, none of the immune-challenge treatments affected the subsequent investment in pheromone emission or number of females attracted. However, we showed that the same males that emitted a high quantity of their sex pheromone in the laboratory were able to attract more females in the field. Our data suggest that the chemical signal is not a reliable predictor of a male's immunocompetence but rather is a general important fitness-related trait, with a higher emission of the sex pheromone measured in the laboratory directly affecting the attractiveness of a male under field conditions.

The decomposition of dead mammalian tissue involves a complex temporal succession of epinecrotic bacteria. Microbial activity may release different cadaveric volatile organic compounds which in turn attract other key players of carcass decomposition such as scavenger insects. To elucidate the dynamics and potential functions of epinecrotic bacteria on carcasses, we monitored bacterial communities developing on still-born piglets incubated in different forest ecosystems by combining high-throughput Illumina 16S rRNA sequencing with gas chromatography-mass spectrometry of volatiles. Our results show that the community structure of epinecrotic bacteria and the types of cadaveric volatile compounds released over the time course of decomposition are driven by deterministic rather than stochastic processes. Individual cadaveric volatile organic compounds were correlated with specific taxa during the first stages of decomposition which are dominated by bacteria. Through best-fitting multiple linear regression models, the synthesis of acetic acid, indole and phenol could be linked to the activity of Enterobacteriaceae, Tissierellaceae and Xanthomonadaceae, respectively. These conclusions are also commensurate with the metabolism described for the dominant taxa identified for these families. The predictable nature of in situ synthesis of cadaveric volatile organic compounds by epinecrotic bacteria provides a new basis for future chemical ecology and forensic studies.

Bumble bees are of major importance, ecologically and economically as pollinators in cool and temperate biomes and as model organisms for scientific research. Chemical signals and cues have been shown to play an outstanding role in intraspecific and interspecific communication systems within and outside of a bumble bee colony. In the present review we compile and critically assess the literature on the chemical ecology of bumble bees, including cuckoo bumble bees. The development of new and more sensitive analytical tools and improvements in sociogenetic methods significantly enhanced our knowledge about chemical compounds that mediate the regulation of reproduction in the social phase of colony development, about the interactions between host bumble bees and their social parasites, about pheromones involved in mating behavior, as well as about the importance of signals, cues and context-dependent learning in foraging behavior. Our review intends to stimulate new studies on the many unresolved questions concerning the chemical ecology of these fascinating insects.

Summary Frugivory among bats (Chiroptera) has evolved independently in the New and Old World tropics: within the families Phyllostomidae and Pteropodidae, respectively. Bats from both families rely primarily on olfaction for the location of fruits. However, the influence of bats on the evolution of fruit scent is almost completely unknown. Using the genus Ficus as a model, the aims of this study were to explore the chemical composition of fruit scent in relation to two contrasting seed dispersal syndromes in Panama and Malaysia and to assess the influence of fruit scent on the foraging behaviour of neo- and palaeotropical fruit-eating bats (Artibeus jamaicensis and Cynopterus brachyotis, respectively). Two hypotheses were tested: (i) variation in fruit scent, between bat- and bird-dispersed figs, is independent of phylogeny and (ii) Old and New World fruit bats, which have evolved independently in each hemisphere, share the same olfactory preferences with respect to fruit scent. The fruit scents of bat- and bird-dispersed fig species were sampled in the field, using dynamic headspace adsorption techniques. New and Old World fruit bats were then captured and tested on natural fig fruit scents from both hemispheres. Chemical analyses, using gas chromatography (GC) and GC/mass spectrometry (MS), revealed a broad overlap in scent compounds between bat-dispersed fig species from both hemispheres. Their fruit scents were dominated by monoterpenes, which contrary to phylogenetic predictions, were completely absent from bird-dispersed species from both regions. The fruit scents of bat-dispersed figs were highly attractive to neotropical bats (A. jamaicensis) in behavioural experiments, whereas those of bird-dispersed figs were completely rejected. Neotropical bats (A. jamaicensis) exhibited a significant preference for fig fruit scents dominated by monoterpenes, independent of the geographical origin of the scent. Palaeotropical bats (C. brachyotis), by contrast, rejected monoterpene-rich fruit scents from the Neotropics. In a cluster analysis (which included additional, published data from the literature), the fruit scents of bat-dispersed figs were clumped by subgenus, with the exception of palaeotropical figs of the subgenus Sycomorus. C. brachyotis, from Malaysia, was the only fruit bat species that significantly preferred the fruit scents of Sycomorus figs that contained no monoterpenes.

Dispersal is a central life-history trait for most animals and plants: it allows to colonize new habitats, escape from competition or avoid inbreeding. Yet, not all species are mobile enough to perform sufficient dispersal. Such passive dispersers may use more mobile animals as dispersal vectors. If multiple potential vectors are available, an active choice can allow to optimize the dispersal process and to determine the distribution of dispersal distances, i.e. an optimal dispersal kernel. We explore dispersal and vector choice in the neotropical flower mite Spadiseius calyptrogynae using a dual approach which combines experiments with an individual-based simulation model. Spadiseius calyptrogynae is found in lowland rainforests in Costa Rica. It inhabits inflorescences of the understorey palm Calyptrogyne ghiesbreghtiana and is phoretic on a number of flower visitors including bats, beetles and stingless bees. We hypothesised that the mites should optimise their dispersal kernel by actively choosing a specific mix of potential phoretic vectors. In a simple olfactometer setup we showed that the flower mites do indeed discriminate between potential vectors. Subsequently we used an individual-based model to analyse the evolutionary forces responsible for the observed patterns of vector choice. The mites combine vectors exhibiting long-distance dispersal with those allowing for more localized dispersal. This results in a fat-tailed dispersal kernel that guarantees the occasional colonization of new host plant patches (long distance) while optimizing the exploitation of clumped resources (local dispersal). Additionally, kin competition results in a preference for small vectors that transport only few individuals at a time. At the same time, these vectors lead to directed dispersal towards suitable habitat, which increases the stability of this very specialized interaction. Our findings can be applied to other phoretic systems but also to vector-based seed dispersal, for example.

Specificity in biological signalling systems is often important to keep information private. Foragers of several species of stingless bees deposit chemical marking signals to guide nestmates to food sources. The markings show species- and colony-specific compositions and primarily attract a bee’s nestmates. An interesting question is whether the bees innately recognize specific trail markings or learn their particular composition from nestmates. To investigate this question, we tested whether Scaptotrigona pectoralis and Scaptotrigona subobscuripennis workers taken from their mother colonies and workers that emerged from combs transferred to foster colonies of the congeneric species are attracted to the marking compounds of workers from their natal colony or from the foster colony. A significant majority of workers were attracted to extracts prepared from foragers of the nest they inhabited, regardless of whether this was the original mother or the congeneric foster colony. Thus, the preference of stingless bee workers for specific food-marking scent mixtures is not innate, but is influenced by the odour they experience within their colony. Despite marked differences in the chemical composition of the scent marks in labial gland secretions of the two investigated species they also shared some main components. We hypothesize that recruitment trail information in stingless bees is composed of one or a few key pheromone compounds acting in conjunction with an additional signature mixture that is species and colony specific and must be learnt by recruited workers. La especificidad en los sistemas de señalización biológica suele ser importante para mantener la información privada. Los forrajeadores de varias especies de abejas sin aguijón depositan señales químicas de marcado para guiar a sus compañeras de nido hacia las fuentes de alimento. Las marcas muestran composiciones específicas de la especie -y de la colonia- y atraen principalmente a los compañeros de nido de la abeja. Una cuestión interesante es si las abejas reconocen de forma innata las marcas de rastro específicas o si aprenden su composición particular de sus compañeras de nido. Para investigar esta cuestión, probamos si las obreras de Scaptotrigona pectoralis y Scaptotrigona subobscuripennis tomadas de sus colonias madre y las obreras surgidas de panales transferidos a colonias de acogida de la especie congénere son atraídas por los compuestos de marcado de las obreras de su colonia natal o de la colonia de acogida. Una mayoría significativa de las obreras se sintieron atraídas por los extractos preparados a partir de las forrajeras del nido que habitaban, independientemente de que éste fuera la colonia madre original o la colonia de acogida congénere. Así pues, la preferencia de las obreras de las abejas sin aguijón por mezclas específicas de olores que marcan el alimento no es innata, sino que está influida por el olor que experimentan dentro de su colonia. A pesar de las marcadas diferencias en la composición química de las marcas de olor en las secreciones de las glándulas labiales de las dos especies investigadas, también comparten algunos componentes principales. Nuestra hipótesis es que la información del rastro de reclutamiento en las abejas sin aguijón se compone de uno o unos pocos compuestos clave de feromonas que actúan junto con una mezcla adicional de firma que es específica de la especie y de la colonia y que debe ser aprendida por las obreras reclutadas. Universidad Nacional, Costa Rica. Centro de Investigaciones Apícolas Tropicales

Burying beetles have fascinated scientists for centuries due to their elaborate form of biparental care that includes the burial and defense of a vertebrate carcass, as well as the subsequent feeding of the larvae. However, besides extensive research on burying beetles, one fundamental question has yet to be answered: what cues do males use to discriminate between the sexes? Here, we show in the burying beetle Nicrophorus vespilloides that cuticular lipids trigger male mating behavior. Previous chemical analyses have revealed sex differences in cuticular hydrocarbon (CHC) composition; however, in the current study, fractionated-guided bioassay showed that cuticular lipids, other than CHCs, elicit copulation. Chemical analyses of the behaviorally active fraction revealed 17 compounds, mainly aldehydes and fatty acid esters, with small quantitative but no qualitative differences between the sexes. Supplementation of males with hexadecanal, the compound contributing most to the statistical separation of the chemical profiles of males and females, did not trigger copulation attempts by males. Therefore, a possible explanation is that the whole profile of polar lipids mediates sex recognition in N. vespilloides.

Studies on the evolution of parental care have focused primarily on the costs and benefits of parental care and the life-history attributes that favour it. However, once care evolves, offspring in some taxa appear to become increasingly dependent on their parents. Although offspring dependency is a central theme in family life, the evolutionary dynamics leading to it are not fully understood. Beetles of the genus Nicrophorus are well known for their elaborate biparental care, including provisioning of their young. By manipulating the occurrence of pre- or post-hatching care, we show that the offspring of three burying beetle species, N. orbicollis, N. pustulatus, and N. vespilloides, show striking variation in their reliance on parental care. Our results demonstrate that this variation within one genus arises through a differential dependency of larvae on parental feeding, but not on pre-hatching care. In N. pustulatus, larvae appear to be nutritionally independent of their parents, but in N. orbicollis, larvae do not survive in the absence of parental feeding. We consider evolutionary scenarios by which nutritional dependency may have evolved, highlighting the role of brood size regulation via infanticide in this genus.

Apple proliferation (AP) and pear decline (PD) are the most severe diseases in pome fruit growing areas. AP-infected trees show typical symptoms such as witches' broom, enlarged stipules, tasteless, and dwarf fruits. PD-infected pears show a progressive weakening, reduced terminal growth, smaller fruits, and die within weeks (quick decline) or years (slow decline). The diseases are caused by the cell-wall lacking bacteria Candidatus Phytoplasma mali (AP phytoplasma) and Ca. P. pyri (PD phytoplasma), respectively. In previous studies it has been shown that AP-infected apple trees emitted higher amounts of the sesquiterpene β-caryophyllene, an attractant of the insect vector Cacopsylla picta (Hemiptera: Psyllidae), thereby facilitating the dispersal of AP phytoplasma. In the present study, volatile organic compounds (VOCs) occurring in the headspace of plants infected with Ca. P. mali strains causing different severity of symptoms in apple plants were collected, analyzed, and identified. Headspace samples from healthy and AP-infected model plant tobacco (Nicotiana occidentalis) and apple (Malus domestica) as well as from healthy and PD-infected pear (Pyrus communis) were investigated via thermodesorption and GC-MS analysis. Significantly higher concentrations of ethyl benzoate were produced in all phytoplasma-infected plants compared to healthy ones and an as yet unidentified sesquiterpene differed between the odor bouquets of healthy and by Ca. P. mali infected tobacco plants. Additionally, statistically significant higher amounts of both compounds were measured in the headspace of plants infected by the virulent AP strain. In apple, significantly higher concentrations of ethyl benzoate and methyl salicylate were observed for trees infected with strains of Ca. P. mali. Ethyl benzoate was also detected in the headspace of pear trees infected with Ca. P. pyri.

Social insects have evolved highly developed communication systems, enabling them to coordinate complex interactions in their colonies. Pheromones play a major role in the coordination of many tasks. In Trigona corvina, a stingless bee that occurs in Central America, foragers use pheromones produced in their labial glands to scent mark solid substrates between a food source and their nest. Newly recruited bees subsequently follow these scent marks until they reach the food source. A recent study has revealed nestspecific differences in the composition of these trail pheromones in colonies of T. corvina, suggesting that pheromone specificity may serve to avoid competition between foragers from different nests. However, the nests used in this study came from different populations and their foragers certainly never met in the field (Jarau et al., 2010). The aim of the present study was to investigate whether differences in the trail pheromones of foragers from different nests can also be found between neighbouring colonies within populations. We analysed the composition of trail pheromones from labial gland secretions extracted from workers from nine colonies collected at three different populations in Costa Rica. The differences in pheromone composition were even more distinct between neighbouring nests within a population than between nests of different populations. This finding corroborates the hypothesis that nest specificity of trail pheromones serves to communicate the location of a food source exclusively to nestmates, thereby avoiding intraspecific competition at resources. Resource partitioning by avoiding conspecific non-nestmates is particularly adaptive for aggressive bee species, such as T. corvina. Los insectos sociales han desarrollado sistemas de comunicación muy desarrollados que les permiten coordinar complejas interacciones en sus colonias. Las feromonas desempeñan un papel importante en la coordinación de muchas tareas. Trigona corvina, una abeja sin aguijón que habita en América Central, las recolectoras utilizan feromonas producidas en sus glándulas labiales para marcar con su olor los sustratos sólidos entre una fuente de alimento y su nido. Las abejas recién reclutadas siguen posteriormente estas marcas de olor hasta llegar a la fuente de alimento. Un estudio reciente ha revelado diferencias en la composición de estas feromonas de rastro en colonias de T. corvina, lo que sugiere que la especificidad de las feromonas puede servir para evitar la competencia entre las forrajeadoras de diferentes nidos. Sin embargo, los nidos utilizados en este estudio procedían de poblaciones diferentes y sus forrajeadores seguramente nunca se encontraron en el campo (Jarau et al., 2010). El objetivo del presente estudio fue investigar si las diferencias en las feromonas de rastro de los forrajeadores de diferentes nidos pueden encontrarse también entre colonias vecinas dentro de las poblaciones. Analizamos la composición de las feromonas de rastro de las secreciones de las glándulas labiales extraídas de las obreras de nueve colonias recogidas en tres poblaciones diferentes de Costa Rica. Las diferencias en la composición de las feromonas fueron aún más marcadas entre nidos vecinos dentro de una población que entre nidos de poblaciones diferentes. Este hallazgo corrobora la hipótesis de que la especificidad de nido de las feromonas de rastro sirve para comunicar la ubicación de una fuente de alimento exclusivamente a los compañeros de nido, evitando así la competencia intraespecífica por los recursos. La partición de recursos evitando a los no compañeros de nido coespecíficos es particularmente adaptativa para las especies de abejas agresivas, como T. corvina. Universidad Nacional, Costa Rica. University of Ulm, Alemania. Division of Nutritional Medicine and General Medicine, Alemania. Centro de Investigaciones Apícolas Tropicales

International audience; To investigate whether flower selection in polylectic solitary bees is modulated by chemical imprinting to nest provisions, larvae of Osmia bicornis (L.) were reared on either Brassica napus L. (Brassicaceae) or Onobrychis viciifolia Scop. (Fabaceae). Flower preferences by adults were evaluated in multiple-choice behavioral tests based on visit number and duration, and flowers selected in the first three visits; data were compared to control bees from the wild. Females reared on B. napus showed only subtle increases in selection for this species, which was highly attractive to both control and experimental bees, masking any effects of imprinting; however, in the first three visits, experimental bees tended to select B. napus more frequently and consistently than controls. Bees reared on O. viciifolia were few and mostly males, which tended to visit this species more than controls. Rearing larvae on either plant affected bee attraction to other plant species. Overall, the data do not provide clear evidence of imprinting, but suggest that rearing bees on a single plant can both directly and indirectly affect flower selection by adults.

International audience; In social insects, the high cost of parental investment has promoted the evolution of traits that reduce or avoid care for unrelated offspring. They have evolved a sophisticated recognition system which enables them to reject foreign individuals, and therefore prevent their nest's exploitation by parasites and predators. Nonetheless, many social parasites are able to overcome social insect recognition and parasitize their parental care behaviour by laying their eggs in host nests. Although several studies have already been conducted on the strategies used by social parasites to successfully invade host nests, the strategy used by its offspring to evade host rejection is still poorly understood. In bumblebees, previous investigations indicated that male offspring of Bombus vestalis (Geoffroy) (Hymenoptera: Apidae), the specific obligate social parasite of Bombus terrestris (L.), may use allomones to evade host worker attacks. Following these studies, we performed electrophysiological studies and behavioural experiments to identify the potential repellents. Six compounds elicited electroantennographic responses in host worker antennae. The corresponding mixture of synthetic volatiles as well as pure tetradecyl acetate, and to a lesser extent pure (Z)-11-hexadecenyl acetate, showed a repellent effect on starved host workers in a feeding bioassay. We hypothesize that tetradecyl acetate has a key function in the repellent effect of the secretions of B. vestalis male offspring towards B. terrestris host workers.

Certain species of Scrophularia (Scrophulariaceae), such as S. nodosa and S. umbrosa, are mainly pollinated by social wasps and are consequently described as wasp-flowers. Because plants attract their pollinators with the help of various floral cues, such as floral odour and/or optical cues, we have investigated the role of olfactory and visual floral signals responsible for wasp attraction in S. umbrosa. Using a combination of chemical (GC, GC-MS) and electrophysiological analyses (GC-EAD), we identified ten compounds in the complex floral odour bouquet that are detectable by the wasps' antennae. As in the wasp-flower Epipactis helleborine, we found so-called 'green leaf volatiles' (GLVs) in the floral odour; these GLVs are highly attractive to the wasps. GLVs, mostly six-carbon aldehydes, alcohols and acetates, and other volatile organic compounds (VOCs), are emitted by many plants infested with herbivores, e.g. caterpillars. In contrast to other investigated wasp-flowers, behavioural experiments have demonstrated that, in addition to the floral odour of S. umbrosa, visual cues are involved in pollinator attraction.

Social parasitism is widespread in many groups of social living hymenopteran species and has also evolved in the genus Bombus. Cuckoo bumblebees (subgenus Psithyrus) are obligate brood parasites in nests of other bumblebee species. After nest usurpation and the killing of the host queen, the parasite female has to control worker reproduction in order to accomplish and maintain reproductive dominance and to ensure her reproductive success. The aim of our study was to examine whether the generalist parasitic bumblebee Bombus bohemicus monopolizes and prevents worker reproduction by physical or chemical means and to identify possible odor compounds involved therein. We performed bioassays with callow workers of the host Bombus terrestris and have shown that B. bohemicus females are able to suppress host worker ovarian development, when these host workers are under the direct influence of the parasite female. Furthermore, by chemical analyses, we have demonstrated that the parasite females adjust to the odor profiles of their host queens in order to maintain the level of fertility signaling inside the host colony although the host queen is absent. We also found that host workers change their odor profile after nest usurpation by the parasite female and consequently, we suggest that the host and parasite are caught up in a chemical arms race.

Many bee species are adapted to just a few specific plants in order to collect pollen (oligolecty). To reproduce successfully, it is important for oligolectic bees to find and recognise the specific host flowers. In this study, we investigated the role of floral volatiles used by an oligolectic bee to recognise its host plants. We compared the attractiveness of natural and synthetic scent samples of host flowers to foraging-naïve and -experienced Hoplitis adunca (Megachilidae) bees that are specialised on Echium and Pontechium (Boraginaceae) plants. The investigations showed that naïve H. adunca females are attracted to 1,4-benzoquinone. During their lifetime, bees learn additional floral cues while foraging on host flowers. In contrast to naïve ones, experienced H. adunca females use, in addition to 1,4-benzoquinone, other compounds to recognise their host plants. 1,4-Benzoquinone is an uncommon floral compound only known from the host plants of H. adunca, and is therefore ideally suited to be used as a plant-specific recognition cue. Several arthropods use this compound to deter insect predators. Therefore, 1,4-benzoquinone as an attractant in Echium flowers may have evolved from a primary function as a defensive compound against insect herbivores.

A decaying cadaver emits volatile organic compounds that are used by necrophilous and necrophagous insects in order to find their brood substrate. Although volatile organic compounds (VOCs) that are released by carcasses have been identified, little is known about the specific compounds that are used by these insects while searching for a brood substrate. Therefore, we have investigated the chemical ecology involved in the attraction of the necrophagous hide beetle Dermestes maculatus , which feeds as an adult and larva upon decomposing carcasses. Our aims have been to identify the responsible compounds in the odours of the carcass that are important for the attraction of the beetles. Furthermore, we have studied sex- and age-related differences in beetle attraction and tested whether the hide beetle can distinguish between various stages of decomposition by means of the emitted odours. Headspace collection of volatiles released from piglet carcasses (bloated stage, post-bloating stage, advanced decay and dry remains), coupled gas chromatography–mass spectrometry (GC–MS), gas chromatography with electroantennographic detection (GC–EAD) and bioassays were conducted to identify the volatiles responsible for the attraction of the beetles. Freshly emerged male beetles were attracted by the odour of piglets in the post-bloating stage (9 days after death; T mean = 27 °C) and the EAD-active compound benzyl butyrate. Statistical analysis revealed a higher relative proportion of benzyl butyrate in the odour bouquet of the post-bloating stage in comparison with the other stages. We therefore conclude that this compound plays an important role in the attraction of hide beetles to carcass odour. This underlines the potential use of D. maculatus for the estimation of the post mortem interval. The decomposition stage at which the female beetles are attracted to the odour of a cadaver remains unknown, as does the nature of this attraction. Pheromones (sexual or aggregation pheromones) might play an essential role correlated with their attraction to carrion and consequently with their attraction to the substrate for mating and ovipositioning.

Sexually deceptive orchids of the genus Ophrys attract male insects for pollination. Pollinator attraction is achieved by mimicking sex pheromones of virgin females of their pollinators, mostly bee species. In earlier investigations, we showed that the phylogenetically distinct Ophrys species O. chestermanii and O. normanii on Sardinia attract their pollinator, males of the cuckoo bumblebee B. vestalis, with the same bouquets of relatively polar volatile compounds. In this investigation, we studied the sex pheromone of virgin females of B. vestalis with the aim of identifying male-attracting compounds and of comparing them with labellum extracts of the two orchids, which were found to release male-attracting compounds in earlier investigations (Gogler et al. 2009). In bioassays, shock-frozen females, cuticle extracts and polar fractions of cuticle extracts of virgin females stimulated mating behaviour in the males. Using gas chromatography coupled with electroantennography (GC-EAD) and gas chromatography coupled with mass spectrometry (GC-MS), we detected in polar fractions of cuticle extracts of B. vestalis females the same electrophysiologically active compounds as in labellum extracts of both orchid species, including aldehydes, esters, fatty acids and alcohols. Since statistical comparisons of the relative proportions of esters showed strong similarities between virgin females and orchids, our results support the hypotheses that this highly specialized Ophrys–pollinator relationship represents another case of chemical mimicry and that esters play a key role in male attraction.