Outbreeding and lack of temporal genetic structure in a drone congregation of the neotropical stingless bee Scaptotrigona mexicana

Drone aggregations are a widespread phenomenon in many stingless bee species (Meliponini), but the ultimate and proximate causes for their formation are still not well understood. One adaptive explanation for this phenomenon is the avoidance of inbreeding, which is especially detrimental for stingless bees due to the combined effects of the complementary sex-determining system and the small effective population size caused by eusociality and monandry. We analyzed the temporal genetic dynamics of a drone aggregation of the stingless bee Scaptotrigona mexicana with microsatellite markers over a time window of four weeks. We estimated the drones of the aggregation to originate from a total of 55 colonies using sibship re-construction. There was no detectable temporal genetic differentiation or sub-structuring in the aggregation. Most important, we could exclude all colonies in close proximity of the aggregation as origin of the drones in the aggregation, implicating that they originate from more distant colonies. We conclude that the diverse genetic composition and the distant origin of the drones of the S. mexicana drone congregation provides an effective mechanism to avoid mating among close relatives.     

Genetic structure of nest aggregations and drone congregations of the southeast Asian stingless bee Trigona collina

In stingless bees, sex is determined by a single complementary sex-determining locus. This method of sex determination imposes a severe cost of inbreeding because an egg fertilized by sperm carrying the same sex allele as the egg results in a sterile diploid male. To explore how reproductive strategies may be used to avoid inbreeding in stingless bees, we studied the genetic structure of a population of 27 colonies and three drone congregations of Trigona collina in Chanthaburi, Thailand. The colonies were distributed across six nest aggregations, each aggregation located in the base of a different fig tree. Genetic analysis at eight microsatellite loci showed that colonies within aggregations were not related. Samples taken from three drone congregations showed that the males were drawn from a large number of colonies (estimated to be 132 different colonies in our largest swarm). No drone had a genotype indicating that it could have originated from the colony that it was directly outside. Combined, these results suggest that movements of drones and possibly movements of reproductive swarms among colony aggregations provide two mechanisms of inbreeding avoidance.     

Chemical basis for inter-colonial aggression in the stingless bee Scaptotrigona bipunctata (Hymenoptera: Apidae)

Inter-colonial aggression was tested using three colonies of Scaptotrigona bipunctata in a natural setting when their nests were moved and by artificial contact between individuals. Examination of the cuticular lipids of individuals from two colonies kept under identical conditions showed clear differences in their cuticular hydrocarbon profiles. The cuticular lipids were a mixture of hydrocarbons (saturated and unsaturated alkanes and alkenes) within the range of C23-C29. The use of multivariate analysis (PCA and discriminant analysis) showed that seven of the identified surface compounds are enough to separate workers from colonies A and B from each other.     

Paternity frequency and maternity of males in some stingless bee species

In monogynous hymenopteran societies, the number of mates of a queen strongly influences the potential for conflict between workers and queens over the maternity of males. Queens always 'prefer' their own sons to sons of workers, regardless of queen mating frequency. When a queen mates once, workers are more closely related to, and therefore are expected to prefer, their own sons and then sons of sisters to sons of the queen. However, if effective paternity frequency exceeds 2, workers on average should prefer queen-produced males to males produced by their sisters. We studied the queen mating frequency of seven stingless bee species: the Mexican species Scaptotrigona mexicana, S. pectoralis and the Australian species Austroplebeia symei, Trigona clypearis, T. hockingsi, T. mellipes and T. sapiens. We then determined whether males arise from eggs laid by workers or queens in A. symei, T. clypearis, T. hockingsi and T. mellipes. We show that all seven species investigated are most likely singly mated and that the queen dominates reproduction. This indicates that the queen's mating frequency alone does not determine whether workers or the queen produces the males.     

Allometric and multivariate analysis of sex and caste polymorphism in the neotropical stingless bee,Scaptotrigona postica

Summary Morphometric data obtained from natural andin vitro-rared queens, workers, and drones of the stingless bee.Scaptotrigona postica were subjected to allometric and multivariate tests to study sex- and caste-related growth patterns. As imaginai phenotype variation appears to be restricted in members of a normal colony, the inclusion ofin vitro-reared individuals permitted an analysis of a considerably wider size range for each morph, and proved important for the calculation of reliable regression statistics in the allometric analyses. Our results show that essentially no individuals with a mixture or transition of sex or caste traits are produced, and, consequently, that for most morphometric characters, queens, workers, and drones lie on different regression lines. Hence, workers cannot be considered as masculinized females, as has been suggested in studies based exclusively on multivariate analyses. Instead, the various morphs may result from a stepwise programming and reprogramming of a few growth rule parameters during critical stages of preimaginal development, and complex hypothetical shifts in the balance of maleness and femaleness genes need not be assumed. A multivariate discriminant analysis, on the other hand, proved to be an efficient tool for the selection of diagnostic size characters.Dedicated to Warwick Estevam Kerr, the Brazilian pioneer of stingless bee genetics, on the occasion of his 70th birthday, September 9, 1992.     

Microsatellite loci for the stingless bee Melipona rufiventris (Hymenoptera: Apidae)

Eight microsatellite primers were developed from ISSR (intersimple sequence repeats) markers for the stingless bee Melipona rufiventris. These primers were tested in 20 M. rufiventris workers, representing a single population from Minas Gerais state. The number of alleles per locus ranged from 2 to 5 (mean = 2.63) and the observed and expected heterozygosity values ranged from 0.00 to 0.44 (mean = 0.20) and from 0.05 to 0.68 (mean = 0.31), respectively. Several loci were also polymorphic in M. quadrifasciata, M. bicolor, M. mandacaia and Partamona helleri and should prove useful in population studies of other stingless bees.     

Caste-specific modulation of juvenile hormone III content and ecdysteroid titer in postembryonic development of the stingless bee,Scaptotrigona postica depilis

Summary Juvenile hormone III content and ecdysteroid titer were analyzed for larval and pupal development of the stingless bee,Scaptotrigona postica depilis. Castespecific differences in juvenile hormone III content were detected at three developmental phases: at the transition from the fourth to the fifth larval stadium, in the spinning phase of the fifth larval stadium, and shortly after the imaginal moult. During the fifth larval stadium, juvenile hormone content closely reflects corpora allata activity. Juvenile hormone synthesis may thus be responsible for the elevated hormone titer in spinning-phase queen larvae, a phase of known sensitivity for induction of queen characters by exogenous juvenile hormone. For ecdysteroids, two phases of caste-specific differences were found: in the pre-pupal phase, and shortly after the imaginal moult. In both periods the titer in queens is distinctly higher compared to workers.Abbreviations Im imago 1 day after eclosion - L3, L4, L5 larval instars 3, 4, and 5 - L5F1, L5F2 substages of feeding phase in fifth larval instar - L5S1, L5S2, L5S3 substages of spinning phase in fifth larval instar - PP1, PP2 substages of prepupal phase - Pw white eyed pupa - Pp pink eyed pupa - Pr red eyed pupa - Pd dark eyed pupa - Pdl, Pdm, Pdd dark eyed pupa with progressive tanning of cuticle - RIA radioimmunoassay     

Genetic divergence between Melipona quadrifasciata Lepeletier (Hymenoptera,Apidae) populations

Melipona quadrifasciata is a stingless bee widely found throughout the Brazilian territory, with two recognized subspecies, M. quadrifasciata anthidioides, that exhibits interrupted metasomal stripes, and M. quadrifasciata quadrifasciata, with continuous metasomal stripes. This study aimed to estimate the genetic variability of these subspecies. For this purpose, 127 colonies from 15 Brazilian localities were analyzed, using nine species-specific microsatellite primers. At these loci, the number of alleles ranged from three to 15 (mean: 7.2), and the observed heterozygosity (H_o) ranged from 0.03–0.21, while the expected heterozygosity (H_e) ranged from 0.23–0.47. The genetic distances among populations ranged from 0.03–0.45. The F_ST multilocus value (0.23) indicated that the populations sampled were structured, and the clustering analysis showed the formation of two subgroups and two more distant populations. The first group contained the subspecies M. quadrifasciata quadrifasciata, and the other, the subspecies M. quadrifasciata anthidioides and the two M. quadrifasciata populations with continuous metasomal stripes from northern Minas Gerais. These results confirmed that the yellow metasomal stripes alone are not a good means for correctly identifying the different subspecies of M. quadrifasciata.     

Foragers of the stingless bee Plebeia droryana inform nestmates about the direction,but not the distance to food sources

1. The tropical stingless bees have evolved intricate communication systems to recruit nestmates to food locations. Some species are able to accurately communicate the location of food, whereas others simply announce the presence of food in the environment.
2. Plebeia droryana is a tiny Neotropical stingless bee that, until recently, was thought to use a solitary foraging strategy, that is without the use of a recruitment communication system. However, recent research has indicated that P. droryana might be able to recruit nestmates to specific food source locations.
3. We tested this by studying whether foragers can guide nestmates in the direction and the distance of artificial feeders placed in the vicinity of the colony. We trained bees to a scented sucrose solution feeder at 10 m and placed different feeders either in different directions (experiment 1) or in different distances (experiment 2). We found that P. droryana directs newcomers in the right direction, but distance information does not seem to be communicated.
4. Moreover, we then tested whether newcomers use chemical and visual cues originating from nestmates foraging at the food source, but found no evidence for the use of these social cues provided by conspecifics.
5. The potential mechanism that P. droryana may use to orient recruits toward the food source, however, remains unknown and requires further study.

     

Genetic variability in five populations of Partamona helleri (Hymenoptera, Apidae) from Minas Gerais State, Brazil

Partamona is a Neotropical genus of stingless bees that comprises 33 species distributed from Mexico to southern Brazil. These bees are well-adapted to anthropic environments and build their nests in several substrates. In this study, 66 colonies of Partamona helleri from five localities in the Brazilian state of Minas Gerais (São Miguel do Anta, Teixeiras, Porto Firme, Viçosa and Rio Vermelho) were analyzed using nine microsatellite loci in order to assess their genetic variability. Low levels of observed (H_o = 0.099-0.137) and expected (H _e = 0.128-0.145) heterozygosity were encountered and revealed discrete genetic differentiation among the populations (F _ST = 0.025). AMOVA further showed that most of the total genetic variation (94.24%) in P. helleri was explained by the variability within local populations.     

Observations on Cymbella mexicana (Ehrenb.) Cleve var. mexicana (Bacillariophyceae) with special reference to the band structure

Cymbella mexicana (Ehrenb.) Cleve var. mexicana was collected from Tama-gawa (Tama River), Tokyo. The fine structure of the species was examined by scanning electron microscopy (SEM), with special reference to colony formation and band morphology. A mature cingulum of this species is usually composed of four open bands. The correlation between the position of the second band (B2) of the cingulum and the attaching pole of the frustule is discussed.     

Effect of group size on the aggression strategy of an extirpating stingless bee, Trigona spinipes

Summary. Group aggression influences communication and defense strategies in many social insect communities. Such aggression plays a particularly significant role in the lives of stingless bees, important native Neotropical pollinators, in which the battle for food resources can be deadly and critical to colony survival. However, the effects of group size on individual aggression levels and the spatio-temporal aggression strategy of communal aggressors have not been fully explored. We therefore investigated how group size affects the aggression levels and the spatio-temporal attack strategy (which body parts, and the amount of time spent in attacking each part) in close combats between Trigona spinipes foragers and a natural competitor, Melipona rufiventris. In all trials, T. spinipes foragers competitively excluded M. rufiventris foragers from nearby feeders, exhibiting four levels of aggressive behavior ranging from threat displays to prolonged grappling and decapitation. Surprisingly, aggression levels and spatial strategy corresponded to the size of group attacks. Larger groups of attackers used individually lower aggression levels than small groups of attackers. Smaller groups also attacked appendages linked to escape (legs and wings) with greater frequency than larger groups, which focused on vital central body areas (abdomen, thorax, and head). Increased aggression corresponded to increased risks for attackers and the attacked. All combatants engaging at the highest level of aggression died (100% mortality). Thus the dominance style of T. spinipes may minimize attack risk and maximize victim harm with finely tuned hostility.Received 22 May 2004; revised 18 August 2004; accepted 26 October 2004.     

Dispersal of Eucalyptus torelliana seeds by the resin-collecting stingless bee,Trigona carbonaria

Ants are the only group of invertebrates currently identified as significant dispersers of seeds, but we report here the dispersal of Eucalyptus torelliana seeds by bees. Fruits of E. torelliana produce resin which is collected by workers of the stingless bee Trigona carbonaria. Seeds adhere to resin in the workers' corbiculate and are transported to the nest. Workers transported seeds distances of more than 300 m from the parent tree and seeds at the nest were viable and capable of germination. Seeds were removed from the nests by workers and discarded away from the nest, and E. torelliana trees became established in the vicinity of colonies of T. carbonaria. Mellitochory may be a novel method of seed dispersal where bees are dispersers, and associated with fruits that produce resin as an attractant for bees.     

Genetic structure of drone congregation areas of Africanized honeybees in southern Brazil

As yet, certain aspects of the Africanization process are not well understood, for example, the reproductive behavior of African and European honeybees and how the first Africanized swarms were formed and spread. Drone congregation areas (DCAs) are the ideal place to study honeybee reproduction under natural conditions since hundreds of drones from various colonies gather together in the same geographical area for mating. In the present study, we assessed the genetic structure of seven drone congregations and four commercial European-derived and Africanized apiaries in southern Brazil, employing seven microsatellite loci for this purpose. We also estimated the number of mother-colonies that drones of a specific DCA originated from. Pairwise comparison failed to reveal any population sub-structuring among the DCAs, thus indicating low mutual genetic differentiation. We also observed high genetic similarity between colonies of commercial apiaries and DCAs, besides a slight contribution from a European-derived apiary to a DCA formed nearby. Africanized DCAs seem to have a somewhat different genetic structure when compared to the European.     

Ontogeny of cephalic volatile patterns in queens and mating biology of the neotropical stingless bee,Scaptotrigona postica

Summary

Chemical communication is an ancient yet still immensely important part of reproduction. Amongst all invertebrates, the most sophisticated “chemical languages” are used by social insects. Here the sex- and caste-specific pheromonal messages consist of multicomponent mixtures. In the neotropical stingless bee Scaptotrigona postica, an inhabitant of dense tropical rain forests, the cephalic volatiles of a queen transmit information on her reproductive status to males. A distinct ontogenetic pattern of the queen pheromone composition allows drones to discriminate receptive virgins which are then chased during the short nuptial flight through the forest understorey. By means of gas chromatographic/mass spectroscopic analyses, the numerous volatile compounds found in pentane extracts of individual bee heads could be identified. Qualitative as well as quantitative changes of these volatiles in the course of imaginal development could be determined, and bioassays with synthetic compounds were undertaken in order to decode the chemical signals used during the short encounter of a young queen and her mate.     

Ergonomics in stingless bees: changes in intranidal behavior after partial removal of storage pots and honey inMelipona favosa (Hym. Apidae,Meliponinae)

Summary The ergonomics of task allocation in stingless bees was examined in two laboratory colonies ofMelipona favosa containing individually marked workers. Performance of intranidal behavior patterns, brood cell production and the processing and storage of nectar, were studied during a control and a wax-deprived period. Experimental wax deprivation consisted of regular controlled removal of storage pots and their contents. Stored carbohydrate concentration and storage pot mass declined during the wax-deprived period. Behavior was measured by sampling for a complete ethogram at seven intranidal areas.In comparison withApis mellifera, M. favosa appeared to be more sensitive to stress, with a distinct reallocation of effort between tasks. The significant increase under stress of brood production and brood-rearing behavior patterns observed was previously also found inApis. Self-oriented behavior patterns declined under stressed circumstances.Involucrum construction declined in wax-deprivedMelipona colonies. Relatively low rates of walking behavior in general inMelipona suggest a low mean free distance between tasks. These results are of particular interest in relation to the basically different nest structure of stingless bees, e.g. very distinct areas for brood production and the processing and storage of food. This architecture of stingless bee nests appeared to influence strongly their specific responses to wax deprivation.