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A variety of animals have been found to interact with and manipulate inanimate objects ‘just for fun’, that is, to play. Most clear examples of object play come from mammals and birds. However, whether insects interact with inanimate objects as a form of play has never been systematically examined. Here, we show that rolling of wooden balls by bumble bees, Bombus terrestris, fulfils behavioural criteria for animal play and is akin to play in other animals. We found that ball rolling (1) did not contribute to immediate survival strategies, (2) was intrinsically rewarding, (3) differed from functional behaviour in form, (4) was repeated but not stereotyped, and (5) was initiated under stress-free conditions. Through the design of the experiment and with the support of behavioural observations, we excluded the possibilities that ball rolling was driven by exploration for food, clutter clearing or mating. Similar to vertebrate play, we also found age and sex differences for ball rolling by bumble bees: younger bees rolled more balls than older bees and male bees rolled individual balls for longer durations than females. We explicitly show that ball rolling is itself a rewarding activity. After being trained to find freely movable balls in one of two differently coloured chambers, bees showed a preference for the colour of the chamber where they had rolled balls. Our results contribute to the question of sentience in insects and lend further support for the existence of positive affective states in these animals.

Vicia faba is a legume crop valued for its high protein content and ability to fix nitrogen. The UK is a key producer of faba beans, which are likely to be increasingly important as a less environmentally harmful protein source. However, without insect pollination, V. faba yield declines by 32.9% on average. When faced with continuing bee declines, it is imperative that more is done to understand how floral traits of V. faba may be used to improve crop yield through pollinator attraction. In this project, novel variation has been identified between commercial lines of V. faba in reward, attraction, and access traits. Using this variation, a hypothesis was formulated that V. faba lines possessing floral traits considered to be more attractive to bees would receive more bee visits. Data from two years of field trials supported this hypothesis, with bees showing preference for lines with more flowers and superior nectar sugar content. Field trials also supported the hypotheses that open pollination would have a positive effect on V. faba yield, and that lines with more attractive floral traits would receive a larger yield benefit with open pollination. These results have shown for the first time that V. faba floral trait variation affects bee attraction in the field and has significant effects on the pollination of the crop. Following field trials, preferences experiments using Bombus terrestris foragers in controlled condition were used to identify V. faba floral traits likely to be most important for bee attraction. Together, field and controlled condition experiments suggested that floral colour, nectar concentration, display size, and scent are traits most likely to increase pollinator attraction to the crop. These experiments have also contributed to the wider understanding of pollinator attraction, adding to evidence that bees show preference for purple colours. The evidence presented in this thesis suggests that by growing existing Vicia faba lines with floral traits including purple flowers with more concentrated nectar, farmers can ensure better crop yield in environments where pollinators are present. In addition, floral traits should be considered as breeding targets by V. faba breeders, as they have great potential to increase crop pollination and support wild pollinator populations.

Gray mold caused by Botrytis cinerea is a common postharvest disease in strawberries, reducing shelf life considerably. We investigated the potential of the yeast-like biocontrol fungus Aureobasidium pullulans (AP-SLU6) vectored by bumblebees (Bombus terrestris) in the Flying Doctors® system to inhibit the pathogen and increase the shelf life of harvested strawberries (cultivar Sonata). Using bumblebees as vectors of various biocontrol agents is becoming increasingly popular, but any potentially negative effects on bee performance have been understudied. Our results show that, over the 4-week period of the trial, the performance and activity of the bees were not negatively affected by A. pullulans. The bees successfully picked up the powder formulation; then, they carried and deposited it on the flowers. The vectoring of the biocontrol agent significantly reduced gray mold development on the harvested fruits by 45% and increased shelf life by 100% in comparison with control treatments. This suggests that the biocontrol fungus applied during flowering successfully reduced Botrytis infection and thus, effectively protected the fruits from gray mold. In addition, the bee-vectored application of the biocontrol agent was found to be significantly more effective than spray application because the latter may temporarily increase humidity around the flower, thereby creating a suitable environment for the pathogen to thrive. In summary, our study demonstrates that A. pullulans vectored by bumblebees can decrease gray mold infection and improve the shelf life of strawberries without adversely affecting the bees, thus providing a basis for the sustainable and efficient control of gray mold on strawberry. [Formula: see text] Copyright © 2022 The Author(s). This is an open access article distributed under the CC BY 4.0 International license .

Graphical abstract, Highlights • There is consensus that neonicotinoids can impact non-target animal fertility. • Thiamethoxam reduced both mating success and sperm physiology in bumblebees. • Queens mated by exposed males had 50% less total living sperm in their spermatheca. • Thiamethoxam may act as anti-aphrodisiac, thereby limiting conservation efforts., Sexual reproduction is common to almost all multi-cellular organisms and can be compromised by environmental pollution, thereby affecting entire populations. Even though there is consensus that neonicotinoid insecticides can impact non-target animal fertility, their possible impact on male mating success is currently unknown in bees. Here, we show that sublethal exposure to a neonicotinoid significantly reduces both mating success and sperm traits of male bumblebees. Sexually mature male Bombus terrestris exposed to a field-realistic concentration of thiamethoxam (20 ng g−1) or not (controls) were mated with virgin gynes in the laboratory. The results confirm sublethal negative effects of thiamethoxam on sperm quantity and viability. While the latency to mate was reduced, mating success was significantly impaired in thiamethoxam-exposed males by 32% probably due to female choice. Gynes mated by exposed males revealed impaired sperm traits compared to their respective controls, which may lead to severe constraints for colony fitness. Our laboratory findings demonstrate for the first time that neonicotinoid insecticides can negatively affect male mating success in bees. Given that holds true for the field, this provides a plausible mechanism contributing to declines of wild bee populations globally. The widespread prophylactic use of neonicotinoids may therefore have previously overlooked inadvertent anti-aphrodisiac effects on non-target animals, thereby limiting conservation efforts.

Wildflower plantings are an important mitigation tool within agri-environmental schemes to counter insect decline in resource-scarce agricultural landscapes. Effectiveness of wildflower plantings for insect conservation is typically studied at the community or species level. It is the individual, however, that is subject to changing abiotic and biotic conditions, not the species per se. Accordingly, functional traits of individuals, i.e., the intraspecific functional diversity within species, likely mediate responses to wildflower resources and landscape context. Here we focused on the ecologically and economically important wild insect pollinator Bombus terrestris to study its intraspecific functional diversity and plant-pollinator individual interactions in wildflower plantings. We found considerable trait variation among flower-visiting B. terrestris workers. Locally, this variation could be attributed to flowering plant traits, with larger workers visiting larger inflorescences and individuals with longer tongues preferentially feeding on zygomorphic but not radial symmetric flowers. In addition, wildflower plantings with high floral abundance attracted individuals with larger pollen baskets. At the landscape scale, increasing proportion of arable land resulted in smaller B. terrestris individuals in wildflower plantings, and a decrease in the overall size diversity of workers. These findings highlight the so far little considered role of intraspecific variation in functional traits of wild pollinators, which can mediate the trait-matching between plants and pollinator individuals. Landscape simplification from agriculture threatens intraspecific pollinator diversity, with potential harmful effects for pollinator fitness and plant reproduction. Tailored wildflower plantings can thus serve as an important tool to increase intraspecific variation in simplified landscapes. When designing seed mixtures for these plantings, high complementarity in plant traits is key for promoting high intraspecific trait diversity of bumblebees and potentially of other associated insect species.

The performance ofBombus terrestrismicro-colonies fed five diets differing in pollen species composition and level of nine essential amino acids (EAA; leucine, lysine, valine, arginine, isoleucine, phenylalanine, threonine, histidine, methionine) was assessed for 37 days by recording total biomass gain, nest building initiation, brood production (eggs, small and large larvae, pupae, drones), nectar, and pollen collection. Stronger colony performance was linked to higher amino acid levels but no consistent differences in biomass gain were recorded between mono- and poly-species diets. Poorest performance occurred in micro-colonies offered pure oilseed rape (OSR) pollen which contained the lowest EAA levels. Reduced micro-colony development (delayed nest initiation and lower brood production) was related to OSR proportion in the diet and lower EAA levels. Results are discussed in relation to selection of plant species in the design of habitats to promote bee populations.

In many communication systems, signal receivers profit from honest signals that indicate the signaller's quality, whereas low-quality signallers should profit from cheating. Under such a conflict of interests between signallers and signal receivers, the maintenance of honest signals presents a puzzle. In theory, honesty can represent an evolutionarily stable strategy, but the actual mechanisms have been studied in few systems only. Here, we investigate honest signalling in a plant species, Brassica rapa, that advertises nectar volumes to pollinators by two honest floral signals; corolla size and the floral volatile phenylacetaldehyde. In a series of experiments we tested for physiological constraints and pollinator behaviours related to honest floral signals and nectar volume, incorporated the result into a mathematical model, and verified its predictions experimentally. While honest floral signals attracted pollinators, the bees’ flower visitation time depended on nectar volume and was associated with the number of seeds that flowers developed. Furthermore, honest floral signals and seed set without pollen limitation both increased after soil fertilisation, indicating nutrient limitation in these traits. The mathematical model which incorporates these findings showed that honest signalling in B. rapa can be maintained by a combination of pollinator behaviour and resource limitation causing differential benefits of nectar production. The study demonstrates how honest floral signalling can evolve as a stable strategy in a plant species.

Given that pollinators usually visit flowers for hidden rewards, they need to rely on floral traits that indicate reward status (“honest signals”). However, the relationship between pollination, honest signals, and floral rewards is little documented in natural conditions. The Scotch broom (Cytisus scoparius) is an invasive shrub with polymorphism in the color of its flowers that can be yellow, orange, or red. In three areas dominated by the Scotch broom, we described the abundance of the floral morphs and estimated bumblebee (Bombus terrestris) visitation rate. We examined whether bumblebee visitation to the floral morphs was related to pollen reward. We collected flowers and classified their stamens according to their function: reward or pollen export. Then, we measured anther size and estimated pollen quantity. The yellow morph was more abundant and more visited by bumblebees than the orange and red morphs. The yellow flowers did indeed offer more pollen than the other morphs and this occurred only for rewarding anthers, suggesting that bumblebees could use yellow color as an honest signal to visit the most rewarding flowers. We discuss whether innate and/or learned preferences of bumblebees can explain why the yellow morph is more visited, pollinated, and abundant, while the other morphs are maintained at a lower frequency. This is one of the few field works that shows that variation in intra-specific floral traits is associated with variation in floral reward and pollinator visitation rate, helping to understand the foraging preferences of pollinators and the coexistence of floral morphs in nature. Clinical trials registration: Not applicable.

A primary approach to establishing hierarchy is through contests, in which conflicts are settled using various behavioural patterns. Although dominance ranks in a hierarchy are determined by quality-related innate factors, how the inequality of competitors is assessed and triggers the formation of a hierarchy by means of behavioural interactions remains unclear. Here, we report the sequence of aggressive behaviours that mediates the onset of hierarchy formation in eusocial bumble bee workers by continuously monitoring pairwise contests. Sequential analysis characterized the dynamics of the behavioural repertoire during hierarchy formation in Bombus terrestris; this was organized in phases associated with a divergence in dominance rank and included mutual pumping (a dorsoventral abdominal movement), bilateral physical contact and unilateral strikes. Escalated bilateral aggression served as a mutual assessment strategy and caused a rapid divergence in rank in evenly matched contests. Head butting was subsequently performed by dominants, suggesting it is a badge of higher rank. The behavioural progression was also similar in two other Bombus species despite interspecific variation in behavioural patterns. Size asymmetry reduced the duration of bilateral aggression, while age advantages over callow workers eliminated bilateral aggression. These results suggested potential roles of mutual assessment by ritualized fighting in hierarchy formation in queenless bumble bee societies.

Insect CAPA neuropeptides are considered to affect water and ion balance by mediating the physiological metabolism activities of the Malpighian tubules. In previous studies, the CAPA-PK analogue 1895 (2Abf-Suc-FGPRLamide) was reported to decrease aphid fitness when administered through micro-injection or via topical application. However, a further statistically significant decrease in the fitness of aphids and an increased mortality could not be established with pairwise combinations of 1895 with other CAPA analogues. In this study, we assessed the topical application of new combinations of 1895 with five CAPA-PVK analogues on the fitness of aphids. We found that 1895 and CAPA-PVK analogue 2315 (ASG-[β3 L]-VAFPRVamide) was statistically the most effective combination to control the peach potato aphid Myzus persicae nymphs via topical application, leading to 72% mortality. Additionally, the combination (1895+2315) was evaluated against a selection of beneficial insects, i.e. a pollinator (Bombus terrestris) and three natural enemies (Chrysoperla carnea, Nasonia vitripennis and Adalia bipunctata). We found no significant influence on food intake, weight increase and survival for the pollinator and the three representative natural enemies. These results could facilitate to further establish, and generate CAPA analogues as alternatives to broad spectrum and less friendly insecticides. This article is protected by copyright. All rights reserved.

Diploid males represent a cost for the viability of populations in haplodiploid species with a single-locus complementary sex determination system. In such social species as bumble bees, their pres...

Epitranscriptomics is an emerging field of investigation dedicated to the study of post-transcriptional RNA modifications. RNA methylations regulate RNA metabolism and processing, including changes in response to environmental cues. Although RNA modifications are conserved from bacteria to eukaryotes, there is little evidence of a epitranscriptomic pathway in insects. Here we identified genes related to RNA m6 A (N6-methyladenine) and m5 C (5-methylcytosine) methylation machinery in seven bee genomes (Apis mellifera, Melipona quadrifasciata, Frieseomelitta varia, Eufriesea mexicana, Bombus terrestris, Megachile rotundata and Dufourea novaeangliae). In A. mellifera, we validated the expression of methyltransferase genes and found that the global levels of m6 A and m5 C measured in the fat body and brain of adult workers differ significantly. Also, m6 A levels were differed significantly mainly between the fourth larval instar of queens and workers. Moreover, we found a conserved m5 C site in the honeybee 28S rRNA. Taken together, we confirm the existence of epitranscriptomic machinery acting in bees and open avenues for future investigations on RNA epigenetics in a wide spectrum of hymenopteran species. This article is protected by copyright. All rights reserved.

During buzz pollination, bees use vibrations to remove pollen from flowers. Vibrations at the natural frequency of pollen-carrying stamens are amplified through resonance, resulting in higher amplitude vibrations. Because pollen release depends on vibration amplitude, bees could increase pollen removal by vibrating at the natural frequency of stamens. However, few studies have characterized the natural frequencies of stamens and compared them to the frequencies of buzz-pollinating bees. We use laser Doppler vibrometry to characterize natural frequencies of stamens of six morphologically diverse, buzz-pollinated, heterantherous Solanum taxa and compare the frequency of bumblebee buzzes produced on two Solanum spp. with different natural frequencies. We found that stamen morphology and plant identity explain variation in their natural frequency. The natural frequencies of the stamens in the studied Solanum taxa fell between 45 and 295 Hz; in five out of six taxa the frequencies were < 190 Hz, which only partly overlaps floral vibrations of buzz-pollinating bees. We show that captive bumblebees produce vibrations at a frequency of 345 Hz and do not change their floral vibrations to match the natural frequency of the visited flowers. Our results suggest that pollen release induced by vibrating stamens at their natural frequencies might only play a role in a subset of buzz pollination interactions.

Pollinating insects are an inherent part of most terrestrial ecosystems as they provide a crucial service for most angiosperms, including numerous important crops. A decrease in pollinator populations can therefore have severe consequences for both natural ecosystems and agricultural yields. Pesticide usage has been pointed out as one of the drivers behind pollinator declines. Globally, neonicotinoids are one of the most commonly used insecticides and studies have shown that exposure at sub-lethal levels can alter foraging behaviour, ultimately negatively affecting survival.Using a custom-made bumblebee colony monitoring system, we examined how the number and duration of foraging bouts of bumblebees (Bombus terrestris) on an individual level, and hive growth rate, was affected by exposure to low (6.5 µg/L) and high (10.6 µg/L) sub-lethal concentrations of the neonicotinoid clothianidin via nectar. We also examined possible interaction between clothianidin exposure and abiotic factors (temperature and precipitation), and its impact on foraging bout number and duration.Exposure to sublethal concentrations of clothianidin increased foraging bout duration in bumblebees. Furthermore, the foraging bout duration decreased with increasing temperature at both exposure concentrations, whereas the unexposed control group was not affected by temperature. Neither number of foraging bouts nor the daily rhythm of foraging bout duration was affected by clothianidin exposure or temperature. The foraging bout duration decreased with increasing precipitation in both exposed and non-exposed groups. However, we did not find any interaction between precipitation and exposure, suggesting that precipitation does not affect toxicity.Our study shows the importance of semi-natural experiments and accounting for ambient factors when assessing the risk that pesticide exposure may present to pollinators. We conclude that the effect of clothianidin exposure on bumblebee foraging behaviour is temperature sensitive and that local climatic conditions and future climate change scenarios should be considered in risk assessments of clothianidin and other insecticides.

Ornamental plants are appreciated by humans for their colorfulness, beauty, abundant flowering and long blooming periods. Many ornamental plants can also constitute an additional foraging resource for flower-visiting insects. However, the ability of the popular ornamental plant Calibrachoa to support urban insect communities is not well documented. In this study, 20 different Calibrachoa cultivars were selected and tested in regard to their insect friendliness based on standardized observations (I) in flight tents using the large earth bumble bee Bombus terrestris as a model species and (II) in open field trials. To investigate what floral characteristics might constitute attractiveness to bumble bees, various floral traits were recorded and compared across all tested Calibrachoa cultivars. Over a two-year period, a total of 6,327 foraging bumble bees were recorded in the tent observations. In the open field observations, we counted 4,188 flower-visiting insects. Our results revealed that (I) all Calibrachoa cultivars were visited by insects for foraging, (II) the number of insect visitors varied significantly among the 20 tested cultivars and (III) the cultivars displayed different floral traits. For the morphometric floral traits and the aroma profiles of Calibrachoa, only the mean nectar quantity and a few identified compounds could be correlated with attractiveness to the model species B. terrestris. We also found that the petal color of the tested cultivars had a significant impact on the number of visitors. Therefore, B. terrestris clearly preferred red or blue Calibrachoa cultivars over those with other petal colors. However, as the cultivar preferences in the different insect groups differed, it is highly recommended to use various cultivars in urban plantings. Nevertheless, efforts must be made to explain what additional floral traits make Calibrachoa and other ornamental plants generally attractive to flower visitors. This information can then be used for breeding purposes to increase the insect friendliness of ornamental plants.

Bu çalışmada, üç farklı entomopatojen fungusun (% 1,5 Beauveria bassiana strain Bb-1, % 1,5 Lecanicillium lecani strain V1-1 ve % 1,5 Paecilomyces fumosoreus strain PFs-1) Bombus terrestris işçi arılarına etkisi kontrollü laboratuvar koşullarda araştırılmıştır. İşçi arılar topikal uygulama ve kuru film tekniği ile entomopatojen fungusların önerilen arazi dozlarına maruz bırakılmıştır. Her kolonideki 30 adet işçi arının 15 adeti mavi, 15 adeti ise kırmızı olacak şekilde göğüs bölümlerinden markalanmıştır. Mavi boyalı işçi arılar herhangi bir uygulama yapılmadan, kırmızı boyalı işçi arılar ise entomopatojen funguslara maruz bırakıldıktan sonra kendi yuvalarına geri konulmuştur. Böylece, B. terrestris kolonilerdeki işçi arıların bazılarının entomopatojen funguslar ile temas etmesinin (i) temas eden işçi arılarda (ii) aynı kolonideki diğer işçi arılarda ve (iii) aynı kolonide temas sonrasında ergin hale gelen işçi arılarda ölüme yol açıp açmadığı belirlenmiştir. Sonuçlar, her üç durum için gözlemlenen işçi arı ölümlerinin kolonilerin gelişim sürecini olumsuz etkilemeyecek kadar düşük düzeyde gerçekleştiğini göstermiştir.

Many endangered plants are susceptible to pollination failure due to their rareness. We studied the threatened Chilean tree Legrandia concinna (Myrtaceae) with melittophilous pollen-only flowers and the pollen loads of Manuelia postica and asked: Do bees guarantee fruit set? What is the role of the endangered plant as a pollen source for M. postica? Only bees of the relictual endemic Manuelia postica and introduced Bombus terrestris and honeybees visited the flowers. Overall flower visitation was very low and so was fruit set. Exotic B. terrestris was the only effective pollinator. The small native M. postica scarcely contributed to pollination and mainly used pollen of L. concinna to rear offspring in the resource-poor Nothofagus forest: pollen of 19 flowers was sufficient to feed a single larvae. Reintroduction of trees of L. concinna as a measure of species protection would also benefit the maintenance of native populations of M. postica. We assume that the introduced bumblebees displaced native Bombus dahlbomii as pollinators of this red list tree species.

While many bumblebee species decline due to climate and land-use changes, others cope well with contemporary conditions. One example is Bombus terrestris, which is common in intensively managed agricultural landscapes. During the 20th century its subgenus, which includes the B. lucorum complex (B. lucorum, B. cryptarum and B. magnus) came to dominate Scandinavian bumblebee communities, but the specific contribution of B. terrestris remains to be understood. Using historical data on males, we assessed how the relative abundances of B. terrestris and the B. lucorum complex changed over the past 150 years in southernmost Sweden. We tested if these changes differed between simplified and mixed landscapes and whether the relative abundance of B. terrestris was related to annual mean temperatures. Because floral availability has advanced as a response to climate change, we also tested if the activity period of males (estimated as catching date) has advanced and whether the advancement differs between taxa. The relative abundance of B. terrestris increased similarly in both landscapes, from 21% to 79% over the period, and this was largely explained by increasing temperature. Male activity period has advanced similarly in the two taxa, with 41 days between 1900 and 2015. Although the dominance of B. terrestris correlates clearly with annual mean temperature, it remains to disentangle why. It also remains to understand whether the success of B. terrestris occurs at the expense of other species or simply reflects that this species copes better with contemporary conditions.

Bumblebees are important pollinators for many wild plants and crops. To explore gene expression characteristics in depth, stable reference genes will be essential across different developmental stages. In this context, we evaluated the stability of eight common reference genes at ten developmental stages in the bumblebee Bombus terrestris (B. terrestris) with four algorithms and two rank approaches. The results showed that all the primers for the reference genes exhibited high specificity, linear correlation (R2-value 0.9917~0.9978) and a good PCR amplification efficiency (E-value 95.4~108.6%). The combined use of RPL13 and RP49 was suggested to be the most suitable reference genes. Our candidate reference genes will help to improve the precision of the detection of gene expression at different developmental stages of bumblebee species.

Bombus (Pyrobombus) taiwanensis Williams, Sung, Lin and Lu sp. nov. (Figures 1, 2) urn:lsid:zoobank.org:act: 726E8A64-28A2-4DD9-8801-651DED8BD495 [Bombus sp. non descripta Chiu 1948: 71, 1&female; (worker) (TARI)] [Bombus (Pyrobombus) nr. hypnorum Starr, 1992: 149, 3&female; (workers) 2&male; (TARI)] +Inference of being conspecific supported by published (Huang et al. 2015; Williams et al. 2020) or ++unpublished (Thanoosing, in prep.) GMYC- or PTP-coalescent analyses for COI barcodes. *Inference of being separate species supported by PTP-coalescent analyses for COI barcodes (Williams et al. 2020) or **by morphology (Williams et al. 2009). Bombus (Megabombus) trifasciatus Smith, 1852 Bombus wilemani Cockerell, 1911 Bombus (Megabombus) bicoloratus Smith, 1879 Bombus (Psithyrus) turneri (Richards, 1929 [Psithyrus]) Psithyrus monozonus (Friese, 1931) not of Friese 1909 [= B. lucorum (Linnaeus)] Bombus (Pyrobombus) taiwanensis sp. nov. * Bombus (Pyrobombus) flavescens Smith, 1852 Bombus (Pyrobombus) sonani (Frison, 1934 [Bremus])* Bombus (Alpigenobombus) angustus Chiu, 1948 * [Bombus (Bombus) terrestris (Linnaeus, 1758) – introduced from Europe only in glass houses] Bombus (Melanobombus) eximius Smith, 1852 Bombus latissimus Friese, 1910 Bombus (Melanobombus) formosellus (Frison, 1934 [Bremus])* Material examined. Holotype: 1&female; (queen), labels (1): white, printed in black ‘ Mt. Hehuan, South peak/(3100 m), Nantou Co.,/ Taiwan, VI-15-2019,/ I. H. Sung’; (2) white, printed in black ‘ Bombus taiwanensis /Williams, Sung, Lin & Lu sp. nov./det. Paul H. Williams’; (3) red, printed in black ‘ HOLOTYPE / &female; / Bombus taiwanensis /Williams, Sung, Lin & Lu’; (4) white, printed in black ‘[barcode]/00214314’ (TFRI). Paratypes: 1&female; (worker), labels: (1) white, printed in black ‘Mt. Hehuan, South peak/ (3100 m), Nantou Co,/ Taiwan, VI-15-2019,/I. H. Sung’; (2) white, printed in black ‘ Bombus taiwanensis /Williams, Sung, Lin & Lu sp. nov./det. Paul H. Williams’; (3) red, printed in black ‘ PARATYPE / &female; / Bombus taiwanensis / Williams, Sung, Lin & Lu’; (4) white, printed in black ‘[barcode]/00214315’ (TFRI). 1&female; (worker), labels (1): white, printed in black ‘ Mt. Hehuan, South peak/(3100 m), Nantou Co.,/ Taiwan, VI-15-2019 /I. H. Sung’; (2) white, printed in black ‘ Bombus taiwanensis /Williams, Sung, Lin & Lu sp. nov./det. Paul H. Williams’; (3) red, printed in black ‘ PARATYPE / &female; / Bombus taiwanensis / Williams, Sung, Lin & Lu’; (4) white, printed in black ‘[barcode]/00214316’ (TFRI). 1&female; (worker), labels (1): white, printed in black ‘Mt. Hehuan, South peak/(3100 m), NantouCo.,/ Taiwan, VI-15-2019,/I. H. Sung’; (2) white, printed in black ‘ Bombus taiwanensis / Williams, Sung, Lin & Lu sp. nov./det. Paul H. Williams’; (3) red, printed in black ‘ PARATYPE / &female; / Bombus taiwanensis / Williams, Sung, Lin & Lu’; (4) white, printed in black ‘[barcode]/00214317’ (TFRI). 1&female; (worker), labels (1): white, printed in black ‘ Mt. Hehuan, South peak/(3100 m), Nantou Co.,/ Taiwan, VI-15- 2019,/I. H. Sung’; (2) white, printed in black ‘ Bombus taiwanensis /Williams, Sung, Lin & Lu sp. nov./det. Paul H. Williams’; (3) red, printed in black ‘ PARATYPE / &female; / Bombus taiwanensis / Williams, Sung, Lin & Lu’; (4) white, printed in black ‘[barcode]/00214318’ (TFRI). Compared material. 1&female; (worker), labels: (1) orange, printed in black ‘C. TAIWAN: Tsuifeng/ 2300 m. Nantou Hsien / 23‒25.VI.1983 /K.S. Lin & S.C. Lin’; (2) white, handwritten ‘nr. ardens ’; (3) white, handwritten ‘[worker] Bombus /(Pyrobombus)/ hypnorum /(Linnaeus)/ det. P.H. Williams /1991’; (4) green, printed in black ‘BOLD# (BEE-BOL /BBW/ PHW)/ 1550A06-TWN’; (5) white, printed in black ‘[worker] Bombus /(Pyrobombus)/ taiwanensis / det. P.H. Williams /2021’ (PHW). 1&male;, labels: (1) orange, printed in black ‘C. TAIWAN: Tayuling/ 2560 m. Hualien Hsien / 9‒16.VI.1980 /K.S. Lin & B.H. Chen’; (2) white, handwritten ‘ &male; Bombus /(Pyrobombus)/ hypnorum /(Linnaeus)/det. P.H. Williams /1991’; (3) green, printed in black ‘BOLD# (BEE-BOL /BBW/ PHW)/1550A07-TWN’; (4) white, printed in black ‘ &male; Bombus /(Pyrobombus)/ taiwanensis /det. P.H. Williams /2021’ (PHW). Etymology Named for its occurrence in Taiwan, avoiding homonymy with earlier names in Bombus of formosanus, formosellus, formosulus and formosus. Diagnosis A predominantly black, orange-tailed bumblebee from Taiwan (Figure 1). Within the hypnorum complex distinguished by (Figure 1): female and male with the front, middle and hind leg basitarsi all with the integument lighter brown than the tibiae (vs darker like the rest of the body for B. hypnorum); hair (pubescence) of the metasoma with all of terga 4‒6 a pale but dull sandy brown or very pale orange (vs white for B. hypnorum). Female clypeus in the half nearest to the labrum smooth and shining with only a few large punctures, mostly spaced by much more than their own widths (Figure 2) (vs large punctures often separated by only their own widths for B. hypnorum). Male genitalia with the gonostylus inner anterior (basal) projection separated from the gonocoxa by less than the breadth of the recurved hook of the penis-valve head (vs separated by much more than the breadth of the recurved hook of the penis-valve head for B. hypnorum). Description Female habitus illustrated in Figure 1, body size small (worker body length 10‒12 mm), hair (pubescence) moderately long, wings nearly clear. Mandible with the distal notch anterior to the posterior tooth (incisura) very shallow and hardly marked. Oculo-malar area (‘cheek’ sensu Williams et al. 2014; not the gena) of medium length, 1.0× longer (length measured between the ventral edge of the compound eye and the edge of the malar area at the articulation of the mandible midway between the mandibular condyles) than the breadth of the mandible at its base (breadth between and including the mandibular condyles). Clypeus weakly swollen (Figure 2), its raised area nearly flat, the central area with few widely scattered large and medium punctures, few punctures especially medially and ventrally adjacent to the labrum. The area between the inner edge of the compound eye and the outer edge of the lateral ocellus occupied in just over its outer third by a broad band with a few mostly large punctures, spaced by more than their own widths, the smaller punctures between the larger punctures very few. Mid basitarsus with the distal posterior corner broadly rounded; hind tibia outer surface with a corbicula, the surface sculpturing weakly reticulate so that the surface appears slightly matt; hind basitarsus in the distal three-quarters covered with short branched decumbent and weakly overlapping hairs with golden reflections; tergum 6 posteriorly rounded and not divided medially, with a small subapical dorsal boss. Colour pattern of the hair of the body predominantly brownish black. Head entirely black except for orange hairs anteriorly on the labrum and laterally on the mandibles. Thoracic dorsum anterior to the wing bases with paler but dark sandy brown hair intermixed, this paler hair extending down the side of the thorax anteriorly, and on the scutellum posteriorly; the front, middle and hind leg tibiae all with some hairs orange, the basitarsi all with the integument lighter brown than the tibiae and with more orange hairs. Hair of the metasoma with tergum 3 posteriorly and all of terga 4‒6 a pale but dull sandy brown or very pale orange. Male body size small (body length 12 mm), hair (pubescence) moderately long, wings nearly clear. Colour pattern of the hair of the body predominantly dark brownish, almost black. Head and the thoracic dorsum anterior to the wing bases with paler but dull sandy brown hair intermixed, this pale hair covering the side of the thorax (mesepisternum), and on most of the scutellum; the front, middle and hind leg tibiae all with the long hairs orange, the basitarsi all with the integument lighter brown than the tibiae and with the long hairs orange. Hair of the metasoma with tergum 1 and all of terga 4‒7 a pale but dull sandy-brown or very pale orange. Male genitalia with the gonostylus nearly triangular, the inner anterior (basal) projection separated from the gonocoxa by less than the breadth of the recurved hook of the penis-valve head; the volsella scarcely projecting beyond the gonostylus; the penis valve with the head recurved as a flattened sickle-shaped hook that is not strongly tapered. Distribution Endemic to the mountains of Hsinchu, Nantou and Hualien counties, central Taiwan, at elevations of 2300‒3100 m., Published as part of Williams, Paul H., Sung, I-Hsin, Lin, Yi-Jing & Lu, Sheng-Shan, 2022, Discovering endemic species among the bumblebees of Taiwan (Apidae, genus Bombus), pp. 435-447 in Journal of Natural History 56 (5 - 8) on pages 438-442, DOI: 10.1080/00222933.2022.2052991, http://zenodo.org/record/6809722, {"references":["Chiu SC. 1948. Revisional notes on the Formosan bombid-fauna (Hymenoptera). Notes D'entomologie Chinoise. 12: 57 - 81.","Starr CK. 1992. The bumble bees (Hymenoptera: Apidae) of Taiwan. Bulletin National Museum of Nat Sci. 3: 139 - 157.","Huang J-X, An J-D, Wu J, Williams PH. 2015. Extreme food-plant specialisation in Megabombus bumblebees as a product of long tongues combined with short nesting seasons. PLoSONE. 10 (8): 1 - 15. doi: 10.1371 / journal. pone. 0132358.","Williams PH, Altanchimeg D, Byvaltsev A, De Jonghe R, Jaffar S, Japoshvili G, Kahono S, Liang H, Mei M, Monfared A, et al. 2020. Widespread polytypic species or complexes of local species? Revising bumblebees of the subgenus Melanobombus world-wide (Hymenoptera, Apidae, Bombus). Eur J Taxon. 719: 1 - 120.","Frison TH. 1934. Records and descriptions of Bremus and Psithyrus from Formosa and the asiatic mainland. Trans Nat Hist Soc Formosa. 24: 150 - 185.","Williams PH, Thorp RW, Richardson LL, Colla SR. 2014. Bumble bees of North America. An identification guide. Princeton, New Jersey: Princeton University Press."]}

Bumblebees (Bombus spp.) are important pollinators of agricultural crops and wild plants, especially tomato and sweet pepper species requiring large-sized and buzzing pollinators. The endoparasitic mite Locustacarus buchneri Stammer 1951 (Acari; Podapolipidae) was identified from commercial populations of the introduced European bumblebee Bombus terrestris. We report the genus and species of this mite for the first time in Koran fauna. Identification was based on morphological and molecular characteristics. Morphological details were illustrated and compared with those to the original description. We analyzed partial cytochrome oxidase I gene to understand the phylogenetic relationship and possible source of invention. The maximum likelihood tree and haplotype network revealed that the Korean population is closely related to Japanese population than European population. The ecological significance of this finding and possible consequences of the invasive species to the native species are discussed.

The dataset is presented as an Excel spreadsheet whose worksheets contain data on ingresses and egresses by worker bees at colonies of the bumble bee, Bombus terrestris. The overall aim was to determine the number of bees outside the colony in relation to time of day. In all worksheets, time of day is recorded in columns A and B, where A shows the hour on a 24-hour clock and B shows minutes after the hour. Each line indicates the time of an event (either worker ingress or egress). For data recorded in 2018, each line indicates the time of an event (either worker ingress or egress), column D (EFFECT) reports a positive value for ingress and negative for egress. For data recorded in 2019, each line indicates the events (ingress or egress) during the time interval up to the next recording and columns D and E report the frequencies of ingress and egress. For all worksheets, the estimated number of bees outside the nest is reported in column G.

Microsporidia are spore-forming intracellular parasites of various invertebrates and vertebrates. Vairimorpha bombi negatively affects the fitness of bumblebees and its prevalence correlates with declining bumblebee populations. The invasive alien species Bombus terrestris colonized Japan and possibly introduced new parasites. To assess the infection prevalence of V. bombi in Japanese bumblebees and B. terrestris, we investigated V. bombi infections using PCR and microscopy. The prevalence of sporulating V. bombi infections in three Bombus s. str. species/subspecies was low, whereas that of non/low-sporulating Vairimorpha sp. infections in three Diversobombus species/subspecies was high. Invasive B. terrestris showed low prevalence of non/low-sporulating V. bombi infections and shared the same V. bombi haplotype with B. hypocrita found in Hokkaido, where B. terrestris is present, and in Honshu, where B. terrestris is absent. Although V. bombi may have been introduced with B. terrestris colonies imported from Europe, it seems to be originally distributed in Japan. Furthermore, a new Vairimorpha sp. was found in Japanese bumblebee species. V. bombi and Vairimorpha sp. showed different organ and host specificities in bumblebees. There are no reports on the specific effects of other Vairimorpha spp. on bumblebees; further studies are needed to clarify the individual characteristics of Vairimorpha spp.

Bumble bees (Bombus sp.) are important pollinators for agricultural systems and natural landscapes and have faced population declines globally in recent decades. Neonicotinoid pesticides have been implicated as one of the reasons for the population reductions in bumble bees and other pollinators due to their widespread use, specificity to the invertebrate nervous system, and toxicity to bees. Adverse outcome pathways (AOPs) are used to describe the mechanism of action of a toxicant through sequential levels of biological organization to understand the key events that occur for a given adverse outcome. We used the AOP framework to organize and present the current literature available on the impacts of neonicotinoids on bumble bees. The present review focuses on Bombus terrestris and B. impatiens, the 2 most commonly studied bumble bees due to their commercial availability. Our review does not seek to describe an AOP for the molecular initiating event shared by neonicotinoids, but rather aims to summarize the current literature and determine data gaps for the Bombus research community to address. Overall, we highlight a great need for additional studies, especially those examining cellular and organ responses in bumble bees exposed to neonicotinoids. Environ Toxicol Chem 2021;40:309-322. © 2020 SETAC. This article has been contributed to by US Government employees and their work is in the public domain in the USA.

The invasion of non-native bees to new ecological territories could spread novel pathogens causing emerging infectious diseases (EIDs) in native species. We provide novel information on the prevalence, load, and co-infection network of honey bee viruses, trypanosoma, microsporidia and neogregarinorida pathogens in native Bombus dahlbomii and non-native Bombus terrestris and Bombus ruderatus. Apicystis bombi and Crithidia bombi were highly prevalent (> 78%) in three bumble bee species, with high loads of these pathogens. Nosema bombi was detected only in B. terrestris (37%) and B. ruderatus (15%). Lotmaria passim was detected in low prevalence (

International audience; Climate change is related to an increase in frequency and intensity of extreme events such as heatwaves. It is well established that such events may worsen the current world-wide biodiversity decline. In many organisms, heat stress is associated with direct physiological perturbations and could lead to a decrease of fitness. In contrast to endotherms, heat stress resistance has been poorly investigated in heterotherms; especially in insects, in which the internal physiological mechanisms available to regulate body temperature are almost negligible making them sensitive to extreme temperature variations.Wild bees are crucial pollinators for wild plants and crops. Among them, bumblebees are experiencing a strong decline across the world. Therefore, the ongoing global decline of these insect pollinators partly due to climate change could cause major economic issues.Here we assess how simulated heatwaves impact fertility and attractiveness (key parameters of sustainability) of bumblebee males. We used three model species: Bombus terrestris, a widespread and warm-adapted species, B. magnus and B. jonellus, two declining and cold-adapted species.We highlight that heat shock (40 degrees C) negatively affects sperm viability and sperm DNA integrity only in the two cold-adapted species. Heat shock can also impact the structure of cephalic labial glands and the production of pheromones only in the declining species.The specific disruption in key reproductive traits we identify following simulated heatwave conditions could provide one important mechanistic explanation for why some pollinators are in decline through climate change.A free Plain Language Summary can be found within the Supporting Information of this article.

Assessing the extent of parasite diversity requires the application of appropriate molecular tools, especially given the growing evidence of multiple parasite co-occurrence. Here, we compared the performance of a next-generation sequencing technology (Ion PGM ™ System) in 12 Bombus terrestris specimens that were PCR-identified as positive for trypanosomatids (Leishmaniinae) in a previous study. These bumblebees were also screened for the occurrence of Nosematidae and Neogregarinorida parasites using both classical protocols (either specific PCR amplification or amplification with broad-range primers plus Sanger sequencing) and Ion PGM sequencing. The latter revealed higher parasite diversity within individuals, especially among Leishmaniinae (which were present as a combination of Lotmaria passim, Crithidia mellificae and Crithidia bombi), and the occurrence of taxa never reported in these hosts: Crithidia acanthocephali and a novel neogregarinorida species. Furthermore, the complementary results produced by the different sets of primers highlighted the convenience of using multiple markers to minimize the chance of some target organisms going unnoticed. Altogether, the deep sequencing methodology offered a more comprehensive way to investigate parasite diversity than the usual identification methods and provided new insights whose importance for bumblebee health should be further analysed.

Bees efficiently learn asocial and social cues to optimize foraging from fluctuating floral resources. However, it remains unclear how bees respond to divergent sources of social information, and whether such social cues might modify bees' natural preferences for nonsocial cues (e.g. flower colour), hence affecting foraging decisions. Here, we investigated honey bees', Apis mellifera, inspection and choices of unfamiliar flowers based on both natural colour preferences and simultaneous foraging information from conspecifics and heterospecifics. Individual honey bees' preferences for flowers were recorded when the reward levels of a learned flower type had declined and novel-coloured flowers were available where they would find either no social information or one conspecific and one heterospecific bumble bee, Bombus terrestris, each foraging from a different coloured flower (magenta or yellow). Honey bees showed a natural preference for magenta flowers. They modified their inspection of both types of flowers in response to conspecific and heterospecific social information. The presence of a conspecific demonstrator on the less-preferred yellow flower increased honey bees' inspection of yellow flowers and the likelihood of foraging on them, thus outweighing the original preference for magenta flowers. The presence of a heterospecific on a magenta flower increased honey bees' inspection of magenta flowers, but this effect was not observed when bumble bees fed on yellow flowers. Our results indicate that flower colour preferences of honey bees are rapidly adjusted in response to both conspecific and heterospecific presence, in different ways, with a preference for conspecific information possibly favouring the transmission of adaptive foraging information within species.

Bumblebees are constantly exposed to a wide range of biotic and abiotic stresses which they must defend themselves against to survive. Pathogens and pesticides represent important stressors that influence bumblebee health, both when acting alone or in combination. To better understand bumblebee health, we need to investigate how these factors interact, yet experimental studies to date generally focus on only one or two stressors. The aim of this study is to evaluate how combined effects of four important stressors (the gut parasite Nosema ceranae, the neonicotinoid insecticide thiamethoxam, the pyrethroid insecticide cypermethrin and the EBI fungicide tebuconazole) interact to affect bumblebees at the individual and colony levels. We established seven treatment groups of colonies that we pulse exposed to different combinations of these stressors for 2 weeks under laboratory conditions. Colonies were subsequently placed in the field for 7 weeks to evaluate the effect of treatments on the prevalence of N. ceranae in inoculated bumblebees, expression levels of immunity and detoxification-related genes, food collection, weight gain, worker and male numbers, and production of worker brood and reproductives. Exposure to pesticide mixtures reduced food collection by bumblebees. All immunity-related genes were upregulated in the bumblebees inoculated with N. ceranae when they had not been exposed to pesticide mixtures, and bumblebees exposed to the fungicide and the pyrethroid were less likely to have N. ceranae. Combined exposure to the three-pesticide mixture and N. ceranae reduced bumblebee colony growth, and all treatments had detrimental effects on brood production. The groups exposed to the neonicotinoid insecticide produced 40%-76% fewer queens than control colonies. Our findings show that exposure to combinations of stressors that bumblebees frequently come into contact with have detrimental effects on colony health and performance and could therefore have an impact at the population level. These results also have significant implications for current practices and policies for pesticide risk assessment and use as the combinations tested here are frequently applied simultaneously in the field. Understanding the interactions between different stressors will be crucial for improving our ability to manage bee populations and for ensuring pollination services into the future.Los abejorros están constantemente expuestos a una amplia gama de agentes estresantes bióticos y abióticos de los que deben defenderse para sobrevivir. Los patógenos y los pesticidas son importantes factores estresantes que influyen en la salud de los abejorros, tanto cuando actúan solos como en combinación. Para tener un mejor conocimiento sobre la salud de los abejorros, debemos investigar cómo interactúan estos factores estresantes, pero los estudios experimentales hasta la fecha generalmente se centran en estudiar solo uno o dos factores. El objetivo de nuestro estudio es evaluar cómo los efectos combinados de cuatro importantes factores estresantes (el parásito intestinal Nosema ceranae, el insecticida neonicotinoide tiametoxam, el insecticida piretroide cipermetrina y el fungicida EBI tebuconazol) interactúan para afectar a los abejorros a nivel individual y de colonia. Establecimos siete grupos de tratamiento de colonias de abejorros que expusimos a diferentes combinaciones de estos factores estresantes durante dos semanas en condiciones de laboratorio, y posteriormente se colocaron en el campo durante siete semanas, para evaluar el efecto de los tratamientos sobre la prevalencia de N. ceranae en abejorros inoculados, los niveles de expresión de genes relacionados con la inmunidad y la desintoxicación, la recolección de alimentos, el aumento de peso, el número de obreras y machos, y la producción de cría de obreras, machos y reinas. La exposición a mezclas de pesticidas redujo la recolección de alimentos por parte de los abejorros. Todos los genes relacionados con la inmunidad se sobre-expresaron en los abejorros inoculados con N. ceranae cuando no habían estado expuestos a mezclas de pesticidas, y los abejorros expuestos al fungicida y al piretroide presentaron menos probabilidades de tener N. ceranae. La exposición combinada a la mezcla de tres pesticidas y N. ceranae redujo el crecimiento de la colonia de abejorros y todos los tratamientos tuvieron efectos perjudiciales en la producción de crías. Los grupos expuestos al insecticida neonicotinoide produjeron entre un 40 y un 76% menos de reinas que las colonias control. Nuestros hallazgos muestran que la exposición a combinaciones de factores estresantes con los que los abejorros entran frecuentemente en contacto tiene efectos perjudiciales sobre la salud y el rendimiento de la colonia y, por lo tanto, podría tener un impacto a nivel poblacional. Estos resultados también tienen importantes implicaciones para las prácticas y políticas actuales de evaluación de riesgos y uso de plaguicidas, ya que las combinaciones probadas aquí se aplican con frecuencia simultáneamente en el campo. Comprender las interacciones entre los diferentes factores de estrés es fundamental para mejorar nuestra capacidad de gestión de las poblaciones de abejas y así garantizar los servicios de polinización en el futuro.

Worldwide pollinator declines lead to pollination deficits in crops and wild plants, and managed bees are frequently used to meet the increasing demand for pollination. However, their foraging can be affected by flower availability and colony size. We investigated how mass-flowering oilseed rape (OSR) can influence the pollen resource use of small and large honey bee (Apis mellifera L.) and bumble bee (Bombus terrestris L.) colonies. Colonies were placed adjacent to strawberry fields along a gradient of OSR availability in the landscapes. We used ITS2 metabarcoding to identify the pollen richness based on ITS2 amplicon sequencing and microscopy for quantification of target pollen. Bumble bees collected pollen from more different plant genera than honey bees. In both species, strawberry pollen collection decreased with high OSR availability but was facilitated by increasing strawberry flower cover. Colony size had no effect. The relationship between next-generation sequencing-generated ITS2 amplicon reads and microscopic pollen counts was positive but pollen type-specific. Bumble bees and, to a lesser degree, honey bees collected pollen from a wide variety of plants. Therefore, in order to support pollinators and associated pollination services, future conservation schemes should sustain and promote pollen plant richness in agricultural landscapes. Both bee species responded to the availability of flower resources in the landscape. Although honey bees collected slightly more strawberry pollen than bumble bees, both can be considered as crop pollinators. Metabarcoding could provide similar quantitative information to microscopy, taking into account the pollen types, but there remains high potential to improve the methodological weaknesses.

Insect‐mediated pollination is fundamental for terrestrial ecosystems as well as for crop production. However, bee populations are declining as a consequence of different global change pressures. Even though these pressures do not act in isolation, their interactive effects have received less attention. Through a cross‐treatment field experiment with Bombus terrestris colonies, we analysed the joint effect of climate warming, exposure to pesticides and landscape transformation on bumblebee development variables related to individual and colony fitness. Our results show that an average increase of 1.62°C in the presence of high resource availability in the landscape increases the number of individuals of all colony castes. However, temperature increase is only favourable when foraging resources are not limiting. This is partly because under elevated temperatures, colonies can develop faster but also need to invest more in wax and silk material to protect the offspring and to cool the colony. We also found a decrease in the number of male production with exposure to pesticides. At the individual level, colonies exposed to increased temperatures produced larger queens and workers, and at the same time, all castes presented less variability in body size. In addition, we observed a reduction in queen size with increasing resource availability. Our experiment indicates that the interaction between global change pressures can be non‐additive and that colony‐level emergent properties of social species could buffer some of the individual impacts of these pressures. Hence, it is necessary to consider not only the joint effects of global change pressures but also the plasticity of the organisms' responses. This would help us to predict how populations will respond to these stressors in a complex real world and how species life‐history traits could modulate their adverse effects through complex phenotypic plastic responses. A free Plain Language Summary can be found within the Supporting Information of this article.

Negative patterning discrimination is considered a nonelemental form of learning, and has been extensively investigated across taxa. Several insect species have also been examined with this task, but only honey bees, Apis mellifera, have so far demonstrated this capacity. Recent empirical and theoretical studies suggest that negative patterning can be accounted for by the emergent property of the mushroom body circuitry in the honey bee brain. Given the neurobiological similarity between bumble bees and honey bees, we hypothesized that bumble bees should also have this cognitive capacity. Here we showed that free-flying bumble bees, Bombus terrestris, learned chromatic, achromatic and visual-olfactory bimodal negative patterning discrimination. Further analyses showed that individual bumble bees adopted distinct strategies in approaching negative patterning discrimination, some solving the task in a configural manner, while others solved it via a simpler strategy. Our findings are in contrast to previous work using harnessed bumble bees and highlight the importance of designing experiments around the requirements of specific species. Our results add to our knowledge on insect nonelemental learning and open up new opportunities for examining cognitive limitations and individual decision-making strategies.

The buff-tailed bumblebee Bombus terrestris presents two distinct colony phenotypes in which some colonies already start producing males very early in the season, while others switch to producing sexuals much later in the season, and specialize mainly in the production of virgin queens. Despite having been extensively investigated in the past, the mechanisms underlying these two distinct phenotypes remain poorly understood. Here we use a combination of behavioral and genetic data to demonstrate that early switchers are in fact diploid male producing colonies, while late switchers produced normal, haploid males. Hence, at a proximate level, early switching to male production was caused by queens mating with males that by chance carried the same sex allele. These patterns were not the result of strong inbreeding within the study population, as the average inbreeding coefficient was very low and in fact slightly negative (−0.11). Moreover, an effect of inbreeding was further excluded because hybrids produced by crossing two distinct outbred populations also produced diploid males in early switching colonies. Finally, we found that diploid males only emerged after a first cohort of workers was produced, while we would have expected diploid males to be produced simultaneously with workers in a 50:50 ratio. We demonstrate that adult diploid males had a distinct cuticular hydrocarbon profile and propose that queens likely cull diploid males during early colony development to reduce the cost of diploid male production.

Bees provide a good model to investigate the evolution of lateralization. So far, most studies focused on olfactory learning and memories in tethered bees. This study investigated possible behavioural biases in free-flying buff-tailed bumblebees (Bombus terrestris) by analysing their turning decisions in a T-maze. Bees of various size were trained to associate a syrup reward with a blue target placed at the centre of the T-maze. The bees were then tested over 16 trials by presenting them with blue targets at the end of the maze’s arms. The maze was rotated 180° after the first 8 trials to control for environmental factors. The number of turnings to the left and right arms were analysed. The bees sampled exhibited a population-level rightward turning bias. As bumblebees vary significantly in size with large bees being better learners than smaller ones, we measured the thorax width to identify a possible relationship between size and bias. No significant correlation was identified. This study shows that bees present lateralization in a visuo-motor task that mimics their foraging behaviour, indicating a possible specialization of the right side of the nervous system in routine tasks.

Double-stranded RNA (dsRNA) designed to target genes in a sequence-specific manner can be valuable in pest control. However, in scenarios where a nontarget could be susceptible and is predicted to be exposed to insecticidal dsRNA in the environment, it is useful to understand possible gene silencing effects in the nontarget. In this study, a sequence complementarity-based approach was used to evaluate potential gene silencing effects in the bumblebee, Bombus terrestris, following oral exposure to a fusion dsRNA product (Ba-dsCOP-vg) specifically designed to target both alpha-coatomer (alphaCOP) and vitellogenin (vg) in the pest pollen beetle, Brassicogethes aeneus. The susceptibility of bumblebees to ingested dsRNA was first evaluated by feeding them with bumblebee-specific dsRNA (Bt-dsCOP-vg) (1 µg/µl) targeting its alphaCOP and vg. A respective 50% and 47% reduction in the transcripts of alphaCOP and vg at 96 h post exposure confirmed its susceptibility to the ingested dsRNA. Using computational tools, a pool of all possible continuous 22-nucleotide stretches from Ba-dsCOP-vg was generated and mapped to all potential off-target genes in the B. terrestris genome, with at least a 13-nt match within the 22-nt stretch. A set of criteria based on sequence complementarity was then used to select 24 potential off-targets for transcript analysis in bumblebees. Following ingestion of Ba-dsCOP-vg (1 µg/µl), no reduction was found in the transcript level for all off-targets, including an off-target with 20-continuous-nt matches. Our findings provide guidance to future risk analysis of dsRNA products and for risk assessments frameworks that incorporate sequence complementarity-based analysis for off-target predictions.

In bumblebees, dominance behaviour contributes to the regulation of the reproductive division of labour between queens and workers. Towards the end of the colony cycle, at the onset of the competition phase, reproductive workers will establish a dominance hierarchy and challenge the reproductive monopoly of the queen by laying unfertilized, male-destined eggs. However, it has been suggested that the dominance and reproductive hierarchies in bumblebee workers are not completely aligned. By performing manipulative experiments and observing both aggressive behaviour and ovarian activation in Bombus terrestris workers, we aimed to elucidate how these two hierarchies interact and are modulated by the presence of the queen and other reproductive workers. We found that the reintroduction of a queen in a queenless colony not only caused worker ovary regression, but also decreased aggressive interactions between workers, thereby restoring the harmony in the colony. Furthermore, transplanting dominant reproductive workers from queenless colonies into queenright colonies induced the resident workers to activate their ovaries and aggressively compete over reproduction, thereby implying that positive feedback drives the onset of the competition phase. However, we did not see this result when subordinate reproductive workers from queenless colonies were added to queenright colonies, suggesting that ovary activation is mainly initiated in reaction to displays of dominance. Additionally, we observed that workers were attacked irrespective of their reproductive state, questioning the importance of signals that advertise either fertility or sterility in bumblebees. Finally, larger workers were significantly more aggressive and significantly more likely to be attacked, even though smaller workers were equally likely to have activated ovaries. This suggests that large workers resort to aggression to increase their chances of successfully reproducing due to their inherent fighting advantage, whereas small cheater workers could possibly reproduce by staying under the radar to evade nestmate aggression.

Bumble bees are ecologically and economically important insect pollinators. Three abundant and widespread species in western North America, Bombus bifarius, Bombus vancouverensis, and Bombus vosnesenskii, have been the focus of substantial research relating to diverse aspects of bumble bee ecology and evolutionary biology. We present de novo genome assemblies for each of the three species using hybrid assembly of Illumina and Oxford Nanopore Technologies sequences. All three assemblies are of high quality with large N50s (> 2.2 Mb), BUSCO scores indicating > 98% complete genes, and annotations producing 13,325 – 13,687 genes, comparing favorably with other bee genomes. Analysis of synteny against the most complete bumble bee genome, Bombus terrestris, reveals a high degree of collinearity. These genomes should provide a valuable resource for addressing questions relating to functional genomics and evolutionary biology in these species.

Robber flies (Asilidae) are the main predatory fly family feeding on beetles, butterflies, other flies for true flies, and even spiders; however, Hymenoptera is the most common prey. Invasive Hymenoptera species are common in central and southern Chile; however, few predators of these are known. The hunting behavior and prey of Chilean robber fly species are also poorly known. The aim of this study is to provide the first hunting behavior records of five Chilean giant robber fly species on invasive Hymenoptera. In addition, an updated distribution of these species is provided. Records of hunting behavior were based on fieldwork collections and citizen science observations. The historical distribution was compared with citizen science observations using chi-square analyzes. Twelve predation events were recorded. Obelophorus terebratus was the most common predator. Bombus terrestris was the invasive Hymenoptera most preyed upon. Both the extension of occurrence of Lycomya germainii as Obelophorus species showed changes in his distribution. Only O. landbecki shown changes in area of occupancy. Citizen science is playing a key role in the knowledge of biological interactions and distribution of endemic and native Chilean robber fly species.

International audience; AbstractIn agricultural landscapes, differences in floral resources provided by crops compared with adjacent habitats promote the spillover of pollinators seeking to fulfil their feeding needs. These foraging patterns play an important role in both crop production and wild plant fitness. However, in classical observational studies, pollinator spillover patterns may be confounded by differences in pollinator phenologies and population sizes. To avoid these confounding effects, we quantified the combined effect of relative floral availability and season on pollinator spillover between strawberry crops and adjacent pinewoods by using commercial bumblebee colonies. We used 60 colonies that were placed in 6 open polytunnel strawberry crops and in 8 pinewoods adjacent to crops in landscapes with either low or high berry (mostly strawberry) crop cover. We repeated the experiment in winter and in spring to account for differences in flower resource availability. While strawberries were in bloom during the entire study period, wild flowering plants were scarce and abundant in winter and in spring, respectively. Spillover in crops was quantified as the percentage of bumblebee individuals that carried pollen from non-berry flowers, while spillover in pinewoods was quantified as the percentage of individuals that carried pollen from berry flowers. Overall, 526 bumblebees were collected. We found them carrying the pollen of 15 plant taxa in winter and 39 in spring, in accordance with seasonal floral availability. In crop colonies, around 30% of bumblebees spilled over into non-berry habitats in winter and 78% in spring, regardless of crop cover in the surrounding landscape. However, in pinewood colonies, we found an interaction effect between season and crop cover: even though on average 34% of bumblebees spilled over into berry crops, in winter it was almost twice in landscapes with high crop cover. Spillover patterns mainly mirrored seasonal changes in floral availability between habitats.

Biocontrol agents (BCAs) are commonly sprayed on flowering pipfruit trees to prevent them from getting infected by various pathogens. By entomovectoring, BCAs can be directly delivered onto the flowers. However, we currently lack knowledge on the distribution dynamics of BCAs by pollinators. Here, managed bees, both bumble bees (Bombus terrestris) and mason bees (Osmia bicornisandOsmia cornuta), were placed in the vicinity of flowering pipfruit trees (pear -'Conference', and apple-'Svatava' and 'Jonagold'), and this allowed us to investigate the distribution of a model BCA, namely, Serenade (R) MAX, from spray-inoculated flowers of a centralized tree to non-inoculated flowers of surrounding receiver trees by bees in an experimental setup in outdoor conditions. One hour after inoculation, we detected an enrichment of BCA in the flowers of the receiver trees and this for each tested pipfruit. The distribution of BCA from treated to untreated flowers was homogenous between the receiver trees for 'Svatava', while significantly different loads were detected for both 'Conference' and 'Jonagold', which might be due to differences in environmental factors, and/or bee characteristics. More research is needed to understand the distribution dynamics of BCAs by pollinators in field conditions, such as in commercial orchards or crop fields, and how this could result in an efficient control.

In bumble bee colonies, pollen is the only protein source for larval feeding and its shortage causes a distress in larval development. Adult bumble bees need pollen for the development of glands and the reproductive system. In bumble bee rearing, honey bee collected pollen is used as the main protein source, either as fresh-frozen or dried pellets, and pollen provisioning is the most problematic and expensive aspect of mass rearing. In honey bee breeding, pollen substitutes are used during the period of food shortage or to stimulate colony strength. We tested different protein diets (five commercial pollen substitutes and two natural protein sources) for the maintenance of bumble bee colonies in captivity. We further mixed Feedbee®, one of the substitutes that gave the best results, with different amounts of pollen to evaluate the optimal amount needed for the whole colony development. Although none of the pure protein diets alone were adequate, diets with a 1 to 1 and 1 to 3 ratio of Feedbee to pollen were both suitable for colony development and queen production. The colony consumed between 2 and 4 g per day of the Feedbee mixed diets, corresponding to a protein consumption of 0.75-0.85 g day-1. Nevertheless, the consumption rate of the pure pollen showed that a mean amount of protein between 0.4 and 0.5 g day-1 was enough to allow colony development indicating the suitability of Feedbee mixed diets. Fil: Bortolotti, Laura. Centro di Ricerca Agricoltura e Ambiente; Argentina Fil: Pošćić, Filip. Institute For Adriatic Crops And Karst Reclamation; Croacia Fil: Bogo, Gherardo. Centro di Ricerca Agricoltura e Ambiente; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Patagonia Norte. Instituto de Investigaciones en Biodiversidad y Medioambiente. Universidad Nacional del Comahue. Centro Regional Universidad Bariloche. Instituto de Investigaciones en Biodiversidad y Medioambiente; Argentina

Oil peony (Paeonia spp.) is a new type of woody oil crop in China with a large cultivation area. Inadequate pollination is one of the main reasons for low seed yield. A pollination net room was built at an oil tree peony base, the numbers of honeybees (Apis mellifera L.) and ground bumblebees (Bombus terrestris L.) were artificially increased and the foraging behaviors and daily activities of the two bees on the plants were observed. Four different pollination methods (honeybee pollination, ground bumblebee pollination, natural field pollination and pollination without insects) were applied. The visit interval, single-flower visit time, number of single-flower visits, number of flowers visited per minute and number of stigma contacts were compared. Meanwhile, the effects of honeybee and bumblebee pollination on the oil yield and quality of peony seeds were compared. There were noticeable differences in daily activity between honeybees and ground bumblebees. Significant differences in the single-flower visit time, visit interval and visit frequency were also observed; honeybee and ground bumblebee pollination increased the seed yield of oil tree peony by 78.74% and 31.88%, respectively. Therefore, both honeybees and ground bumblebees are effective pollinators of oil tree peony. These results provide a theoretical basis for further utilization of bee resources for oil tree peony pollination.