Natural enemies impose a selective pressure on solitary insects that may favour the evolution of sociality. In the socially polymorphic orchid bee Euglossa viridissima, females found nests solitarily and provision a first batch of brood. After brood maturity, a nest can remain solitary (all offspring disperse) or become social, when one or more subordinate daughters forage for nesting material and brood provisions for the dominant mother. Solitary females leave their nest unguarded when foraging whilst a female in a social nest can guard the nest while nestmates are foraging. By observing solitary and social nests, we found that subordinate foragers in social nests undertook longer provisioning trips than solitary females. The presence of a guarding female in a social nest protected the nest against intrusion, possibly favouring longer provisioning trips. Moreover, the frequency of successful attempts by intruders to enter nests was significantly lower in social nests. Our results provide strong support for the parasite defence hypothesis for the evolution of social behaviour.

Global warming may have a significant negative impact on insects in the tropics due to an increase in the frequency and intensity of heat waves. Heat waves may have a particularly harsh impact on the relatively sessile juvenile stages of holometabolous insects, such as larvae and pupae. The honey bee ( Apis mellifera L.) is an important tropical holometabolous insect whose queens and drones may also be negatively impacted by heat waves during their development, leading to reduced reproductive capacity. However, we have little information on the thermal tolerance of honey bee subspecies in tropical regions. To address this knowledge gap, we evaluated the effect of simulated heat waves during development on drones and queens of Africanized A. mellifera in terms of the size, shape, and symmetry of their forewings, as well as their reproductive traits such as sperm concentration and viability in drones, and ovariole number and spermatheca volume in queens. Drones raised at high temperatures were more asymmetric in forewing size but not in shape, and had a smaller body size and reduced sperm concentration and viability compared to those raised at normal hive temperatures. In contrast, no effect of an elevated temperature regime during development was seen in queens in terms of the size and symmetry of the forewing, the number of ovarioles, and the volume of the spermatheca. Our results support the notion that males of insects are more susceptible to high temperatures compared to females. We discuss the implications of our findings for the reproductive fitness of honey bee colonies in tropical regions.

Body size has frequently been discussed as related to reproductive success. Stingless bees show remarkable variation in body size among and within sexes and castes, but very little is known about the proximate causes and the consequences of such variation in these highly eusocial bees. In this study, we evaluated the effect of different amounts of food provided to the larvae on developmental time, body size, abdominal mass, and vitellogenin (Vg) hemolymph titers of gynes of the stingless bee Scaptotrigona pectoralis, a species with trophic caste determination. Our results provide empirical evidence of a significant effect of the quantity of larval food on the duration of development, body size, and traits of reproductive fitness. We discuss proximate causes that may explain gyne size variation in stingless bees. We see this as an initial step towards understanding the adaptive significance and ultimate causes of this phenomenon in highly eusocial bees.

Studies on the expression of genes in different contexts are essential to our understanding of the functioning of organisms and their adaptations to the environment. Gene expression studies require steps of normalization, which are done using the stable expression pattern of reference genes. For many different eusocial bees reference genes have been discovered, but not for the primitively eusocial euglossine bees.We used available genomic resources of euglossine species and the gene information of Apis melliferato develop a set of reference genes for the primitive eusocial bee Euglossaviridissima. We tested nine genes in distinct developmental stages three different algorithms to infer the stability of gene expression. The Tata binding protein(Tbp) and 14-3-3epsilon were the most stable genes across all different stages. The strongest deviation in gene expression pattern occurred in pupae, which require a different set of genes for normalizing gene expression.

The relative contributions of heritable and environmentally acquired components of colony odor towards individual recognition are scarcely known in social insects. Larval food may affect cuticular profiles which in turn may serve as cues in the process of elimination of excess gynes characteristic of the eusocial stingless bees. In this study, we evaluated the contribution of larval food to cuticular profiles of stingless bee gynes and quantitatively tested if recognition (latency) from workers may be related to gyne chemotype and origin in the species Scaptotrigona pectoralis. Our results showed that the origin of food did not significantly affect the cuticular profiles of gynes, as larvae of the same origin reared on food from different colonies showed similar cuticular profiles at emergence. We suggest that overlapping over floral resources may account for the similarity in cuticular cues derived from food across experimental colonies. Additionally, workers showed similar latency time to first aggression towards gynes irrespectively of their chemotype and origin. Gyne’s mass had no effect on the aggressive response from workers either. We observed that gynes threatened aggressive workers which counteracted further aggression. Our results indicate that in stingless bees, cuticular hydrocarbons at emergence seem to have genetic origin and that gyne tolerance seems not related to cuticular chemical profiles. We suggest that cuticular cues may serve as caste labels for the identification of newly emerged gynes after which worker aggression towards them would elicit behavioral indicators of their fitness.

Male neotropical orchid bees collect volatile substances from their environment and compose complex, species-specific odour blends in leg pouches. These perfumes are accumulated by the males over time and are exposed during pre-mating display. It has been hypothesized that perfumes are indicators of male genotypic quality and that females chose mates by the quality or intensity of their odour. Because direct experimental proof is lacking, we investigated whether the amount or complexity of male perfumes is related to male (1) size, (2) fluctuating (left–right) asymmetry, or (3) age, traits that are known to be sexually selected in other animals. We measured left and right forewings (cell node distances for 1 and 2, wing wear for 3) of Euglossa dilemma and Euglossa viridissima collected during dry and rainy seasons on the Yucatán peninsula, Mexico (total N = 768). Wing size was not related to the quantity or the complexity of perfume extracts in either species after excluding the effect of season, which positively affected both size and perfume load in E. dilemma. Wing asymmetry had also no effect, except in rainy season E. viridissima, where it was positively (not negatively) correlated with the quantity of perfume. Wing wear, an established age indicator of orchid bees, had the only consistent overall effect, being positively correlated with perfume amount and complexity in both species. This is in agreement with the idea that perfumes are an honest indicator of male survival capacity.

Defense castes are know from highly eusocial insects yet have rarely been described in social species with a small colony size. In nests of Euglossa viridissima , an orchid bee exhibiting primitively eusocial behavior, we recorded one subordinate female per nest to specialize in guarding in the presence of a dominant and a second subordinate who specialized in foraging. Guarding may have arisen as a response to cleptobiosis by conspecifics, as nests with a guard more successfully avoided intrusion and resin theft.

Males of sibling orchid bees Euglossa viridissima and Euglossa dilemma are morphologically cryptic, except for the number and shape of mandibular teeth. An alternative morph of E. viridissima has a third tooth similar to males of E. dilemma . We used this model system to evaluate the potential of wing morphometrics for the resolution of these groups. We found differences in the size characters of forewings of E. viridissima and E. dilemma albeit with substantial overlapping amongst them. However, geometric morphometrics of forewing vein intersections separated both species and, to a lesser extent, morphotypes. A discriminant analysis of the shape of the radial cell showed separation between all three groups, too, albeit with higher misclassification between E. viridissima and E. dilemma . We show that sibling cryptic species and morphotypes can be identified by geometric morphometrics, supporting its application with other methods as powerful aids to infrageneric taxonomy in bees.

Isometric worker size variation has been found in various species of stingless bees but the adaptive value of this phenomenon is little understood. We studied intra-colony worker size variation in Melipona in relation to individual foraging preference and colony growth in periods of resource abundance and scarcity. We found significant forager size differences across colonies. In spite of this, intra-colony size of foragers collecting different resources was highly similar, suggesting that foraging preferences and flexibility are size constrained. On the other hand, inter-colony forager size was associated with parameters of colony growth, albeit differently, depending on the season. Our results suggest that isometric forager size may reflect the state of food storage, and thus, colony development in stingless bees. It seems that stingless bee communities respond to seasonal abundance and scarcity with variable cycles of individual colony growth and reduction of reserves that may explain the asynchronous reproduction of colonies, spanning years in these insects.

The internal transcribed spacers of the ribosomal RNA gene have been recently proposed as an appropriate marker for genetic analysis of the molecular variation of stingless bees. Herein we report the characterization of the complete ITS1 region in two populations (from Mexico and Guatemala) of the endangered Melipona yucatanica Camargo, Moure and Roubik. Phylogenetic analyses showed low genetic variation between populations but defined a geographic structure with Mexican and Guatemalan specimens forming two well supported clades. Low ITS1 genetic variation found between populations contrasts with high genetic variation found in other markers. Phylogenetic analysis corroborates the inclusion of M. yucatanica within the subgenus Melipona sensu stricto based on previous morphological studies. The results highlight the utility of the ITS1 for the characterization of stingless bee populations.

Melipona colimana is endemic to the temperate forests of western Mexico and may be in conservation risk due to forest exploitation. Differences between the density of nests, nesting sites and spatial distribution in two places with different levels of human disturbance were established. A preserved (C) and a disturbed area (P) were identified and it was determined that nest density, nesting sites and the number of potential nest sites were predominant in the C zone. In total, 27 of 30 colonies were found on oak trees (Quercus laurina) with a diameter at breast height of 183.4 ± 34.21 cm which shows a close relationship of this bee species with this type of tree. A positive correlation between the DBH of the nesting sites in relation to the trees with nests and the presence of cavities was found. It was determined that the nests are distributed in the form of aggregates in both zones (R = 0.31 and 0.39) with a density of 0.17 hˉ¹ and 0.04 hˉ¹ colonies. Forestry exploitation seems to be affecting wild populations since the tree that bees use as nesting sites are destroyed; that is why it is recommended to implement a preservation program and a sustainable management of this species.

Euglossines have long been regarded as largely solitary, though some species are known to exhibit social behavior. We studied the nesting behavior of Euglossa viridissima over an annual cycle, comparing sociality and offspring production across the rainy (RS) and dry seasons (DS) in the Yucatan Peninsula of Mexico. Nests were built in both the RS and the DS, but with greater nest initiation and brood provisioning in the RS, presumably as a consequence of more floral resources at this time of year. Across the year, numerical sex ratios were female biased (0.7 as females/total); sex ratios varied across individual nests from 0.3 to 1.00, though without a clear relationship to sociality. Egg-to-adult development was quicker in females than males and, within a sex, quicker when ambient temperatures were higher. Multi-female (social) nests were only founded at the end of the RS and the beginning of the DS, coincided with the presence of Hymenopteran and Dipteran parasites in nests headed by solitary females. Reduced floral resources and a higher risk of parasitism, possibly coupled with higher female density or reduced nesting sites, may be factors favoring the formation of multi-female associations in this euglossine. Better nourishment of foundress females in the RS may improve lifespan and permit overlapping generations which, coupled with the kin structure of their nests, may favor social nesting in E. viridissima.

In this study, variability of microsatellite loci has been for the first time analyzed in the wild stingless bee Melipona colimana. Data have been compared with those obtained from an insular population of the managed species M. beecheii. Genotyping results in M. colimana demonstrate that microsatellite loci sequences are conserved within the genus Melipona. Population parameters such as number of alleles and heterocigosity values, were not significantly different between the two studied species, both being within the observed range in other Melipona species. Values of the insular M. beecheii population were lower than in previously studied continental populations of the same species, in agreement with the results obtained in other island organisms. Wild M. colimana colonies are susceptible to deforestation effects, so that, we have compared the genetic parameters of colonies from a deforested area with those of a conserved area. Population diversity was not significantly different, may be due to recent environmental perturbations that are not yet reflected in the genetic diversity of these insects

Melipona colimana is endemic to the temperate forests of western Mexico and may be in conservation risk due to forest exploitation. Differences between the density of nests, nesting sites and spatial distribution in two places with different levels of human disturbance were established. A preserved (C) and a disturbed area (P) were identified and it was determined that nest density, nesting sites and the number of potential nest sites were predominant in the C zone. In total, 27 of 30 colonies were found on oak trees (Quercus laurina) with a diameter at breast height of 183.4 ± 34.21 cm which shows a close relationship of this bee species with this type of tree. A positive correlation between the DBH of the nesting sites in relation to the trees with nests and the presence of cavities was found. It was determined that the nests are distributed in the form of aggregates in both zones (R = 0.31 and 0.39) with a density of 0.17 hˉ¹ and 0.04 hˉ¹ colonies. Forestry exploitation seems to be affecting wild populations since the tree that bees use as nesting sites are destroyed; that is why it is recommended to implement a preservation program and a sustainable management of this species.

Transferred copies of mitochondrial DNA (mtDNA) into the nuclear genome (numts) have been reported in several Hymenoptera species, even at a high density in the honey bee nuclear genome. The accidental amplification of numts in phylogenetic studies focused on mtDNA highlights the importance of a correct determination of numts and their related mtDNA sequences. We report here the presence of numts derived from a mitochondrial rDNA 16S gene in the genome of the stingless bee species Melipona colimana and M.fasciata (tribe Meliponini) from Western Mexico. PCR products were cloned in both species obtaining thirty paralogous numts. Numts were identified by the presence of insertions and deletions and the disruption of the 16S secondary structure. Further phylogenetic analyses including alternative mitochondrial cox1 and nuclear ITS1 genes have revealed the presence of another numt (cox1) in the nuclear genome of these two species, and place both as sister lineages within the subgenus Michmelia. This is one of the first studies reporting the presence of numts in Meliponini species, and supports previous studies suggesting frequent transfer of mtDNA to the nuclear genome in Hymenoptera.

Male orchid bees (Euglossini, Apidae, Hymenoptera) expose species-specific blends of volatile chemicals (perfume bouquets) during their courtship display. The perfumes are acquired by collecting fragrant substances from environmental sources, which are then accumulated in specialised hind leg pouches. To balance the perfume composition, the males need to find and collect the required substances in specific relative amounts while facing seasonal, local or habitat-dependent differences in compound availability. Experience-dependent choice of odours, i.e. ‘learned avoidance’ of recently collected components, has been proposed as the mechanism that mediates the accumulation of the stereotypical compound ratios. In the present study, we used the presence of certain compounds in male hind leg pouches as proxy for the respective local compound availability, and investigated whether differences in content are correlated with differences in chemical choice assays. Our results suggest that volatile availability differs between localities ( n = 16) as well as habitats ( n = 2; coastal vs. inland) across the Yucatán peninsula, Mexico, for both studied species. Male Euglossa dilemma showed a pronounced preference for benzyl benzoate and eugenol at locations where those compounds were rare in hind leg extracts, as predicted by the learned avoidance model. No equivalent correlations were found for Euglossa viridissima . This is the first study to combine chemical analyses of perfumes with bioassays of odour choice. It strengthens the view that negative feedback from collected odours modifies future chemical choice and helps males to acquire specific perfume blends.