Insights from Melipona bicolor hybrid genome assembly: a stingless bee genome with chromosome-level scaffold.

BACKGROUND: The highly eusocial stingless bees are crucial pollinators of native and agricultural ecosystems. Nevertheless, genomic studies within this bee tribe remain scarce. We present the genome assembly of the stingless bee Melipona bicolor. This bee is a remarkable exception to the typical single-queen colony structure, since in this species, multiple queens may coexist and share reproductive duties, resulting in genetically diverse colonies with weak kinship connections. As the only known genuinely polygynous bee, M. bicolor's genome provides a valuable resource for investigating sociality beyond kin selection. RESULTS: The genome was assembled employing a hybrid approach combining short and long reads, resulting in 241 contigs spanning 259 Mb (N50 of 6.2 Mb and 97.5% complete BUSCOs). Comparative analyses shed light on some evolutionary aspects of stingless bee genomics, including multiple chromosomal rearrangements in Melipona. Additionally, we explored the evolution of venom genes in M. bicolor and other stingless bees, revealing that, apart from two genes, the conserved repertoire of venom components remains under purifying selection in this clade. CONCLUSION: This study advances our understanding of stingless bee genomics, contributing to the conservation efforts of these vital pollinators and offering insights into the evolutionary mechanisms driving their unique adaptations.

A versatile recording device for the analysis of continuous daily external activity in colonies of highly eusocial bees.

As pollinators, bees are key to maintaining the biodiversity of angiosperm plants, and for agriculture they provide a billion-dollar ecosystem service. But they also compete for resources (primarily nectar and pollen), especially the highly social bees that live in perennial colonies. So, how do they organize their daily temporal activities? Here, we present a versatile, low-cost device for the continuous, automatic recording and data analysis of the locomotor activity in the colony-entrance tube of highly eusocial bees. Consisting of an in-house built block containing an infrared detector, the passage of bees in the colony entrance tunnel is registered and automatically recorded in an Arduino environment, together with concomitant recordings of temperature and relative humidity. With a focus on the highly diverse Neotropical stingless bees (Meliponini), we obtained 10-day consecutive recordings for two colonies each of the species Melipona quadrifasciata and Frieseomelitta varia, and also for the honey bee. The Lomb-Scargle periodogram analysis identified a predominant circadian rhythmicity for all three species, but also indications of ultradian rhythms. For M. quadrifasciata, which is comparable in size to the honey bee, we found evidence for a possibly anticipatory activity already before sunrise. As all three species also presented activity at night in the colony entrance tube, this also raises questions about sleep organization in social insects. The cost and versatility of the device and the open-source options for data analysis make this an attractive system for conducting studies on circadian rhythms in social bees under natural conditions, complementing studies on flower visits by these important pollinators.

Bee Species, Botanical Sources and the Chemical Composition of Propolis from Yucatan, Mexico.

Propolis is used by corbiculated bees to protect the bee hive; it is mostly used to seal cracks, to reduce or prevent microbial growth and to embalm invaders. Different factors have been reported to influence the chemical composition of propolis, including bee species and the flora surrounding the hive. Nevertheless, the majority of the studies are focused on propolis produced by Apis mellifera, while studies on the chemical composition of propolis produced by stingless bees are still limited. In this investigation, the chemical composition of 27 propolis samples collected in the Yucatan Peninsula from A. mellifera beehives, together with 18 propolis samples from six different species of stingless bees, were analyzed by GC-MS. Results showed that lupeol acetate and ß-amyrin were the characteristic triterpenes in propolis samples from A. mellifera, while grandiflorenic acid and its methyl ester were the main metabolites present in samples from stingless bees. Multivariate analyses were used to explore the relationship between bee species and botanical sources on the chemical composition of the propolis samples. Differences in body size and, therefore, foraging abilities, as well as preferences for specific botanical sources among bee species, could explain the observed variation in propolis chemical composition. This is the first report on the composition of propolis samples from the stingless bees Trigona nigra, Scaptotrigona pectoralis, Nannotrigona perilampoides, Plebeia frontalis and Partamona bilineata.

Functional orthologs of honeybee CYP6AQ1 in stingless bees degrade the butenolide insecticide flupyradifurone.

Flupyradifurone (FPF), a novel butenolide insecticide binding to nicotinic acetylcholine receptors (nAChRs), has been shown to be less acutely toxic to western honey bees (Apis mellifera) than other insecticides such as neonicotinoids sharing the same target-site. A previous study revealed that this is due to enhanced oxidative metabolism of FPF, mediated by three cytochrome P450 monooxygenases (P450s), including CYP6AQ1. Therefore, we followed a toxicogenomics approach and investigated the potential role of functional CYP6AQ1 orthologs in FPF metabolism from eight different bee species, including stingless bees (Tribe: Meliponini). We conducted a phylogenetic analysis on four stingless bee species, including Frieseomelitta varia, Heterotrigona itama, Melipona quadrifasciata and Tetragonula carbonaria to identify CYP6AQ1-like functional orthologs. Three non-Meliponini, but tropical bee species, i.e., Ammobates syriacus, Euglossa dilemma and Megalopta genalis were analyzed as well. We identified candidate P450s in all (neo)tropical species with greater than 61% and 67% predicted protein sequence identities when compared to A. mellifera CYP6AQ1 and Bombus terrestris CYP6AQ26, respectively. Heterologous expression in High Five insect cells of these functional orthologs revealed a common coumarin substrate profile and a preference for the O-debenzylation of bulkier substrates. Competition assays using the fluorescent probe substrate 7-benzyloxymethoxy-4-trifluoromethylcoumarin (BOMFC) with these enzymes indicated inhibition of BOMFC metabolism by increasing concentrations of FPF. Furthermore, UPLC-MS/MS analysis revealed the capacity of all CYP6AQ1-like orthologs to metabolize FPF by hydroxylation in vitro at various levels, indicating a conserved FPF detoxification potential in different (neo)tropical bee species including Meliponini. This research, employing a toxicogenomics approach, provides important insights into the potential of stingless and other tropical bee species to detoxify FPF, and highlights the significance of investigating the detoxification mechanisms of insecticides in non-Apis bee species by molecular tools to inform risk assessment and conservation efforts.

Bioactive Attributes and Analysis of Electronic Nose Feature Signals of Colombian Stingless Bees Propolis.

The volatile and non-volatile chemical composition and bioactivity of propolis from the species Apis mellifera has been widely studied, but there is very little knowledge regarding propolis of other bee species, which ultimately hinders their differentiation and potential use. In this work, 53 propolis samples of A. mellifera and four stingless bee species (Frieseomielitta sp., Melipona eburnea, Melipona sp., and Trigona sp.) were collected in Colombia. An electronic nose with 10 metal oxide semiconductor sensors (MOS) was used to generate a pattern of the representative volatile compounds of the samples. Ethanolic extracts were obtained to assess their antioxidant activity towards DPPH radical and ABTS radical cation, total phenolics, and color (CIELAB space). The results showed an overall similarity of the aromatic profiles between species. The antioxidant activity of Frieseomielitta sp. propolis was higher than that of A. mellifera and the other species, in correspondence with a higher phenolic content. CIELAB color parameter b* was the most differentiating variable among samples, indicating a variation of propolis colors between red and yellow. By combining the data from physico-chemical analysis and aromatic profile, it was possible to differentiate the propolis from each bee species, with the exception of those from Melipona sp. and Trigona sp., indicating their similarity. These results have practical significance since they are a starting point to recognizing and valuing native stingless bee propolis and their bioactive potential, which, in addition to geographical differentiation and further quality parameters evaluation, will enhance their commercial exploitation.

Heat stress survival and thermal tolerance of Australian stingless bees.

Stingless bees (Meliponini) are important pollinators throughout the world's tropical and subtropical regions. Understanding their thermal tolerance is key to predicting their resilience to changing climates and increasingly frequent extreme heat events. We examined critical thermal maxima (CTmax), survival during 1-8 h heat periods, chill coma recovery and thermal preference for Australian meliponine species that occupy different climates across their ranges: Tetragonula carbonaria (tropical to temperate regions), T. hockingsi (tropical and subtropical regions only) and Austroplebeia australis (widely distributed including arid regions). We found interspecific differences in thermal tolerance consistent with differences in the climate variability observed in each species' range. Foragers of A. australis had a faster chill coma recovery (288 s) than foragers of T. hockingsi (1059 s) and T. carbonaria (872 s). Austroplebeia australis also had the highest CTmax of 44.5 °C, while the CTmax of the two Tetragonula species was ∼43.1 °C. After a 1-h heat exposure, T. carbonaria foragers experienced 95% mortality at 42 °C, and 100% at 45 °C. Surprisingly, larvae and pupae of both Tetragonula species were more resistant to heat exposure than foragers. Within an enclosed temperature gradient apparatus (17-38 °C), no clear preference was found for foragers; however, they were most frequently observed at ∼18 °C. Results indicate that in some regions of Australia, meliponines already experience periodic heat events exceeding their thermal maxima. Employing effective management strategies (such as nest site insulation and habitat preservation) may be crucial to colony survival under continued climate change.

Evidence of floral rewards in Brasiliorchis supports the convergent evolution of food-hairs in Maxillariinae.

PREMISE: Angiosperms distributed over a large geographical area may display considerable phenotypic variation that can be recognized at morphological and micromorphological levels. Here, we investigate the pollination biology and the presence of floral rewards in Brazilian populations of the widely distributed orchid, Brasiliorchis picta. Based on the new data presented here this study investigates the evolution of floral rewards in Maxillariinae, and tests for the occurrence of convergent evolution of food-hairs in this subtribe. METHODS: Micromorphological and histochemical analyses of the labellar tissues were conducted, together with chemical analysis of fragrance and experiments involving the use of chemical baits. The evolution of floral rewards in Maxillariinae were addressed. RESULTS: Microscopy revealed that B. picta offers food-hairs as a reward. Fragrance is produced by abaxially located labellar epidermal papillae. The main compound present in our samples (2-phenylethanol) also occurs in the aggregation pheromone produced by the mandible glands of pollinators, Meliponini bees. Our analyses indicate a high diversity of flower rewards and pollinators displayed by members of Maxillariinae, and support that edible trichomes evolved independently five times in the subtribe. CONCLUSIONS: The high diversity of floral rewards and pollinators displayed by members of Maxillariinae suggests that different pollinator pressures are involved in the evolution of this neotropical subtribe. In addition, the offering of food-hairs, which are generally infrequently encountered in Orchidaceae, arose by convergent evolution in Maxillariinae.

Nestmate recognition in the Amazonian stingless bee Melipona paraensis.

Meliponine bees use chemical-based nestmate discrimination to protect their colonies from unrelated intruders. However, species from the Amazon basin are relatively poorly known from this perspective. Here, we investigated Melipona paraensis nestmate discrimination in different contexts (nests vs. neutral arenas), testing aggression in bees facing other bees varying in age and origin (same/different colony or a different kleptoparasitic meliponine species) or experimentally treated with odors from unrelated colonies. As expected, M. paraensis did not discriminate against callow non-nestmate workers with weak/undifferentiated chemical signatures. Workers specialized in nest defense aggressed intruders more often than non-specialized workers, but were less aggressive in neutral arenas than in the nest. Our study provides novel behavioral information relevant for social insect research and meliponiculture.

Sensory and physicochemical analysis of honeys of nine stingless bee species of Mexico and Guatemala.

Honeys produced by species other than Apis mellifera are little known. The objective of this study was to carry out sensory and physicochemical analyses of 62 samples to characterize honeys from nine species of stingless bees from Mexico and Guatemala. Analyses were performed at the sensory level and at the physicochemical level. Qualitative and quantitative data were studied using the multivariate non-metric multidimensional scaling (NMDS) statistical technique, with the Gower metric. According to the stress value (p > 0.2), the most significant parameters were at the sensory level, for visual evaluation, physical state, color, and appearance; for olfactory-taste evaluation, elemental sweet and acid tastes; for tactile evaluation, consistency, and crystallization, and for physicochemical evaluation, pH, electrical conductivity and reducing sugars. Data analysis showed that Melipona beecheii honeys formed a defined group, with similar sensory characteristics and physicochemical values, even taking into account their place of origin. Frieseomelitta nigra honeys also showed significant differences with the other honeys analyzed. Our work suggests that the entomological origin is the main factor that determines the characteristics of honey, and that the floral origin is the secondary factor, a hypothesis that should be verified in future works.

Potential source of ecofriendly insecticides: Essential oil induces avoidance and cause lower impairment on the activity of a stingless bee than organosynthetic insecticides, in laboratory.

The negative effect of insecticides on bees has been reported as one of the factors associated with the decline in population of these pollinators. Thus, the aim of this study was to evaluate the response of the stingless bee Nannotrigona aff. testaceicornis (Lepeletier, 1836) to a promising source of new insecticide molecules obtained from Lippia sidoides (rosemary pepper) essential oil (EO) and its major compounds (thymol, ρ-cymene, and (E)-caryophyllene), comparing them to commercial insecticides (organosynthetic: imidacloprid, deltamethrin and semisynthetic: spinetoram). For this, stingless bees were exposed by contact with these compounds to evaluate the lethal and sublethal (locomotion and flight orientation) toxicity. The L. sidoides EO and its major compounds have low lethal toxicity to forager worker bees (N. aff. testaceicornis). The organosynthetics imidacloprid (LD50 =0.00146 µgbee-1) and deltamethrin (LD50 =0.0096 µg bee-1) were about 209,589 and 31,875 times more toxic, respectively, than the least toxic natural compound, (E)-caryophyllene (LD50 =306 µgbee-1). Locomotion ability and flight orientation were little affected by spinetoram and by L. sidoides EO and its major compounds, however, were greatly reduced by the imidacloprid and deltamethrin insecticides. Besides shows low lethal and sublethal toxicity, the bioinsecticides were also avoided by the forager bees. Individuals treated with the L. sidoides EO and thymol were avoided by the untreated bees. Therefore, the natural products studied here were promising due to their recognized effectiveness against pest insects and greater safety to bees N. aff. testaceicornis.

The nuclear and mitochondrial genomes of Frieseomelitta varia - a highly eusocial stingless bee (Meliponini) with a permanently sterile worker caste.

BACKGROUND: Most of our understanding on the social behavior and genomics of bees and other social insects is centered on the Western honey bee, Apis mellifera. The genus Apis, however, is a highly derived branch comprising less than a dozen species, four of which genomically characterized. In contrast, for the equally highly eusocial, yet taxonomically and biologically more diverse Meliponini, a full genome sequence was so far available for a single Melipona species only. We present here the genome sequence of Frieseomelitta varia, a stingless bee that has, as a peculiarity, a completely sterile worker caste. RESULTS: The assembly of 243,974,526 high quality Illumina reads resulted in a predicted assembled genome size of 275 Mb composed of 2173 scaffolds. A BUSCO analysis for the 10,526 predicted genes showed that these represent 96.6% of the expected hymenopteran orthologs. We also predicted 169,371 repetitive genomic components, 2083 putative transposable elements, and 1946 genes for non-coding RNAs, largely long non-coding RNAs. The mitochondrial genome comprises 15,144 bp, encoding 13 proteins, 22 tRNAs and 2 rRNAs. We observed considerable rearrangement in the mitochondrial gene order compared to other bees. For an in-depth analysis of genes related to social biology, we manually checked the annotations for 533 automatically predicted gene models, including 127 genes related to reproductive processes, 104 to development, and 174 immunity-related genes. We also performed specific searches for genes containing transcription factor domains and genes related to neurogenesis and chemosensory communication. CONCLUSIONS: The total genome size for F. varia is similar to the sequenced genomes of other bees. Using specific prediction methods, we identified a large number of repetitive genome components and long non-coding RNAs, which could provide the molecular basis for gene regulatory plasticity, including worker reproduction. The remarkable reshuffling in gene order in the mitochondrial genome suggests that stingless bees may be a hotspot for mtDNA evolution. Hence, while being just the second stingless bee genome sequenced, we expect that subsequent targeting of a selected set of species from this diverse clade of highly eusocial bees will reveal relevant evolutionary signals and trends related to eusociality in these important pollinators.

Fungicide pyraclostrobin affects midgut morphophysiology and reduces survival of Brazilian native stingless bee Melipona scutellaris.

Native stingless bees are key pollinators of native flora and important for many crops. However, the loss of natural fragments and exposure to pesticides can hinder the development of colonies and represent a high risk for them. Nevertheless, most studies are conducted with honeybees and there are not many studies on native species, especially in relation to the effects of fungicides on them. Therefore, the objective of this paper is to evaluate the effects of sublethal concentrations of pyraclostrobin, on Melipona scutellaris forager workers. These Brazilian native stingless bees were submitted to continuous oral exposure to three concentrations of pyraclostrobin in sirup: 0.125 ng a.i./µL (P1), 0.025 ng a.i./µL (P2), and 0.005 ng a.i./µL (P3). Histopathological and histochemical parameters of midgut, as well as survival rate were evaluated. All concentrations of fungicide showed an increase in the midgut lesion index and morphological signs of cell death, such as cytoplasmic vacuolizations, presence of atypical nuclei or pyknotic nuclei. Histochemical analyzes revealed a decreased marking of polysaccharides and neutral glycoconjugates both in the villi and in peritrophic membrane in all exposed-groups in relation to control-groups. P1 and P2 groups presented a reduction in total protein marking in digestive cells in relation to control groups. As a consequence of alteration in the midgut, all groups exposed to fungicide showed a reduced survival rate. These findings demonstrate that sublethal concentrations of pyraclostrobin can lead to significant adverse effects in stingless bees. These effects on social native bees indicate the need for reassessment of the safety of fungicides to bees.

Looking beyond the flowers: associations of stingless bees with sap-sucking insects.

The main sources of food for stingless bees are the nectar and pollen harvested from flowers, whereas one important kind of nesting material (i.e. wax) is produced by their own abdominal glands. Stingless bees can, nonetheless, obtain alternative resources of food and wax from exudates released by sap-sucking insects as honeydew and waxy cover, respectively. To date, there are no comprehensive studies investigating how diversified and structured the network interactions between stingless bees and sap-sucking insects are. Here, we conducted a survey of the data on relationship between stingless bees and sap-sucking insects to evaluate: (1) which resources are collected by which stingless bee species; (2) how diverse the interaction network is, using species degree and specialisation index as a proxy; and if (3) there would be any phylogenetic signal in the species degree and specialisation indices. Our findings demonstrate that approximately 21 stingless bee species like Trigona spp. and Oxytrigona spp. have been observed interacting with 11 sap-sucking species, among which Aethalion reticulatum is the main partner. From ca. 50 records, Brazil is the country with most observations (n = 38) of this type of ecological interaction. We found also that stingless bees harvest fivefold more honeydew than waxy covers on sap-sucking insects. However, we did not find any phylogenetic signal for the occurrence of this interaction, considering species degree and specialisation indices, suggesting that both traits apparently evolved independently among stingless bee species. We suggest that specific ecological demands may drive this opportunistic behaviour exhibited by stingless bees, because major sources of food are obtained from flowers and these bees produce their own wax.

Repeated switches from cooperative to selfish worker oviposition during stingless bee evolution.

Reproductive division of labour is a defining feature of insect societies. Stingless bees (Meliponini) are an interesting exception among the highly eusocial insects in that workers of many species contribute significantly to the production of males. Since workers remain sterile in other species of this large tropical tribe, it has been hypothesized that, in the latter species, ancestral queens have won the conflict over who produces the males. The fact that sterile workers of some species lay trophic eggs to feed the queen and display ritualized behaviours towards her during oviposition has been interpreted as an evolutionary relic of this ancient conflict. Here, I used ancestral state estimation to test whether worker reproduction is indeed the ancestral condition and worker sterility a derived state in stingless bees. Contrary to this hypothesis, data suggest that trophic egg laying was the ancestral condition, whereas selfish worker reproduction in queenright colonies evolved subsequently during stingless bee diversification. The appearance of worker reproduction in queenright conditions was tightly linked to the laying of trophic eggs, which suggests that having activated ovaries in queen presence facilitates the evolution of worker reproduction. Worker reproduction is also linked to brood cell architecture, but surprisingly not to colony size or queen-worker dimorphism. The reason for this association between brood cell architecture and worker oviposition is currently unknown. These results suggest that trophic eggs are not a relic of an ancient conflict, but a sign of overlapping interests between the queen and workers about who produces the males.

Impact of environmental temperatures on mortality, sex and caste ratios in Melipona interrupta Latreille (Hymenoptera, Apidae).

Temperature is a major driver of biological phenomena, from metabolism to ecological interactions and rates of evolutionary diversification. However, species vary greatly in their thermal tolerance, as well as the temperature under which they perform best. This study aimed to investigate the effect of experimental manipulation of environmental temperatures on the individual mortality and phenotypic composition of colonies of Melipona interrupta. To fulfill these objectives, 30 colonies in equivalent developmental conditions were artificially subjected to different temperatures. Temperatures were monitored by thermo-hygrometers, and immature mortality and sex and caste ratios were observed in brood combs during 14 months. A strong effect of external temperature on immatures was detected on deviations from 28 to 30 °C (the natural average temperature inside the colony), causing an increase in mortality. Likewise, a significant effect of temperature on sex ratio was detected, with male:female ratio decreasing at temperatures below and above 28-30 °C. Lastly, there was no clear evidence for an effect of temperature on caste ratio, although queens appeared to become relatively more frequent at warmer temperatures. The results of this study allow us to conclude that anthropogenic changes, whose effect can be extrapolated to the similar natural changes, that modify the environmental temperatures to which M. interrupta colonies are exposed are likely to compromise their survival, mainly through individual mortality.

Low-Cost Electronic Tagging System for Bee Monitoring.

This paper introduces both a hardware and a software system designed to allow low-cost electronic monitoring of social insects using RFID tags. Data formats for individual insect identification and their associated experiment are proposed to facilitate data sharing from experiments conducted with this system. The antennas' configuration and their duty cycle ensure a high degree of detection rates. Other advantages and limitations of this system are discussed in detail in the paper.

Foraging traits modulate stingless bee community disassembly under forest loss.

Anthropogenic land use change is an important driver of impacts to biological communities and the ecosystem services they provide. Pollination is one ecosystem service that may be threatened by community disassembly. Relatively little is known about changes in bee community composition in the tropics, where pollination limitation is most severe and land use change is rapid. Understanding how anthropogenic changes alter community composition and functioning has been hampered by high variability in responses of individual species. Trait-based approaches, however, are emerging as a potential method for understanding responses of ecologically similar species to global change. We studied how communities of tropical, eusocial stingless bees (Apidae: Meliponini) disassemble when forest is lost. These bees are vital tropical pollinators that exhibit high trait diversity, but are under considerable threat from human activities. We compared functional traits of stingless bee species found in pastures surrounded by differing amounts of forest in an extensively deforested landscape in southern Costa Rica. Our results suggest that foraging traits modulate competitive interactions that underlie community disassembly patterns. In contrast to both theoretical predictions and temperate bee communities, we found that stingless bee species with the widest diet breadths were less likely to persist in sites with less forest. These wide-diet-breadth species also tend to be solitary foragers, and are competitively subordinate to group-foraging stingless bee species. Thus, displacement by dominant, group-foraging species may make subordinate species more dependent on the larger or more diversified resource pool that natural habitats offer. We also found that traits that may reduce reliance on trees-nesting in the ground or inside nests of other species-correlated with persistence in highly deforested landscapes. The functional trait perspective we employed enabled capturing community processes in analyses and suggests that land use change may disassemble bee communities via different mechanisms in temperate and tropical areas. Our results further suggest that community processes, such as competition, can be important regulators of community disassembly under land use change. A better understanding of community disassembly processes is critical for conserving and restoring pollinator communities and the ecosystem services and functions they provide.

Enemy recognition is linked to soldier size in a polymorphic stingless bee.

Many ant and termite colonies are defended by soldiers with powerful mandibles or chemical weaponry. Recently, it was reported that several stingless bee species also have soldiers for colony defence. These soldiers are larger than foragers, but otherwise lack obvious morphological adaptations for defence. Thus, how these soldiers improve colony fitness is not well understood. Robbing is common in stingless bees and we hypothesized that increased body size improves the ability to recognize intruders based on chemosensory cues. We studied the Neotropical species Tetragonisca angustula and found that large soldiers were better than small soldiers at recognizing potential intruders. Larger soldiers also had more olfactory pore plates on their antennae, which is likely to increase their chemosensory sensitivity. Our results suggest that improved enemy recognition might select for increased guard size in stingless bees.

The Stingless Bee Melipona solani Deposits a Signature Mixture and Methyl Oleate to Mark Valuable Food Sources.

Stingless bees foraging for food improve recruitment by depositing chemical cues on valuable food sites or pheromone marks on vegetation. Using gas chromatography/mass spectrometry and bioassays, we showed that Melipona solani foragers leave a mixture composed mostly of long chain hydrocarbons from their abdominal cuticle plus methyl oleate from the labial gland as a scent mark on rich food sites. The composition of hydrocarbons was highly variable among individuals and varied in proportions, depending on the body part. A wide ratio of compounds present in different body parts of the bees elicited electroantennogram responses from foragers and these responses were dose dependent. Generally, in bioassays, these bees prefer to visit previously visited feeders and feeders marked with extracts from any body part of conspecifics. The mean number of visits to a feeder was enhanced when synthetic methyl oleate was added. We propose that this could be a case of multi-source odor marking, in which hydrocarbons, found in large abundance, act as a signature mixture with attraction enhanced through deposition of methyl oleate, which may indicate a rich food source.

Soldiers in a Stingless Bee.

The differentiation of workers into morphological subcastes (e.g., soldiers) represents an important evolutionary transition and is thought to improve division of labor in social insects. Soldiers occur in many ant and termite species, where they make up a small proportion of the workforce. A common assumption of worker caste evolution is that soldiers are behavioral specialists. Here, we report the first test of the "rare specialist" hypothesis in a eusocial bee. Colonies of the stingless bee Tetragonisca angustula are defended by a small group of morphologically differentiated soldiers. Contrary to the rare specialist hypothesis, we found that soldiers worked more (+34%-41%) and performed a greater variety of tasks (+23%-34%) than other workers, particularly early in life. Our results suggest a "rare elite" function of soldiers in T. angustula, that is, that they perform a disproportionately large amount of the work. Division of labor was based on a combination of temporal and physical castes, but soldiers transitioned faster from one task to the next. We discuss why the rare specialist assumption might not hold in species with a moderate degree of worker differentiation.