Mandibular Gland Volatiles and Their Ontogenetic Patterns in Queen Honey Bees,Apis mellifera carnica

Extracts of mandibular glands taken from adult queens of the honey bee, Apis mellifera carnica, were analysed by gas chromatography-mass spectroscopy. More than 100 compounds could be identified among which oxygenated fatty acids with six, eight, 10 and 12 carbon atoms are particularly interesting since they show structural relationships to the queen substance, (E)-9-oxo-2-decenoic acid. Changes in the patterns of volatiles were followed up from emergence until the full dominant status of an egg-laying queen in a strong colony. Generally, the amount of volatiles per gland was found to increase with age. The final level of queen substance (9-ODA) content is reached at the postmating stage about 10 days after emergence. Ontogenetic patterns of concentrations were determined for those components regarded to predominantly contribute to the royal pheromone. Characteristic compositions of signals, possibly involved in the premating, mating and postmating dominance status of a honey bee queen are discussed. Copyright 1997 Elsevier Science Ltd. All rights reserved     

Primer effect of queen pheromone on juvenile hormone biosynthesis in adult worker honey bees

Juvenile hormone synthesis in adult worker honey bees was measured by an in vitro corpora allata bioassay. Adult queenless workers exhibit higher rates of juvenile hormone biosynthesis than queenright workers. Hormone synthesis is not correlated with the volume of the glands. Extract of queen mandibular glands, applied to a dummy, reduces juvenile hormone biosynthesis in caged queenless workers to the level of queenright workers. The same result was obtained with synthetic (E)-9-oxo-2-decenoic acid, the principal component of the queen mandibular gland secretion. This pheromonal primer effect may function as a key regulating element in maintaining eusocial colony homeostasis. The presence of brood does not affect the hormone production of the corpora allata.Abbreviations BSA bovine serum albumin - CA Corpora allata - JH juvenile hormone - 9-ODA (E)-9-oxo-2-decnoic acid     

Suppression of queen rearing in European and Africanized honey beesApis mellifera L. by synthetic queen mandibular gland pheromone

Summary Queen rearing is suppressed in honey bees (Apis mellifera L.) by pheromones, particularly the queen's mandibular gland pheromone. In this study we compared this pheromonally-based inhibition between temperate and tropically-evolved honey bees. Colonies of European and Africanized bees were exposed to synthetic queen mandibular gland pheromone (QMP) for ten days following removal of resident queens, and their queen rearing responses were examined. Queen rearing was suppressed similarly in both European and Africanized honey bees with the addition of synthetic QMP, indicating that QMP acts on workers of both races in a comparable fashion. QMP completely suppressed queen cell production for two days, but by day six, cells containing queen larvae were present in all treated colonies, indicating that other signals play a role in the suppression of queen rearing. In queenless control colonies not treated with QMP, Africanized bees reared 30% fewer queens than Europeans, possibly due to racial differences in response to feedback from developing queens and/or their cells. Queen development rate was faster in Africanized colonies, or they selected older larvae to initiate cells, as only 1 % of queen cells were unsealed after 10 days compared with 12% unsealed cells in European colonies.     

Colony insularity through queen control on worker social motivation in ants

We investigated the relative contribution of the queen and workers to colony nestmate recognition cues and on colony insularity in the Carpenter ant Camponotus fellah. Workers were either individually isolated, preventing contact with both queen and workers (colonial deprived, CD), kept in queenless groups, allowing only worker-worker interactions (queen deprived, QD) or in queenright (QR) groups. Two weeks post-separation QD and QR workers were amicable towards each other but both rejected their CD nestmates, which suggests that the queen does not measurably influence the colony recognition cues. By contrast, aggression between QD and QR workers from the same original colony was apparent only after six months of separation. This clearly demonstrates the power of the Gestalt and indicates that the queen is not a dominant contributor to the nestmate recognition cues in this species. Aggression between nestmates was correlated with a greater hydrocarbon (HC) profile divergence for CD than for QD and QR workers, supporting the importance of worker-worker interactions in maintaining the colony Gestalt odour. While the queen does not significantly influence nestmate recognition cues, she does influence colony insularity since within 3 days QD (queenless for six months) workers from different colony origins merged to form a single queenless colony. By contrast, the corresponding QR colonies maintained their territoriality and did not merge. The originally divergent cuticular and postpharyngeal gland HC profiles became congruent following the merger. Therefore, while workers supply and blend the recognition signal, the queen affects worker-worker interaction by reducing social motivation and tolerance of alien conspecifics.     

Egg marking pheromones of anarchistic worker honeybees (Apis mellifera)

In honeybees, worker policing via egg eating enforces functionalworker sterility in colonies with a queen and brood. It is thoughtthat queens mark their eggs with a chemical signal, indicatingthat their eggs are queen-laid. Worker-laid eggs lack this signaland are, therefore, eaten by policing workers. Anarchistic workerhoneybees have been hypothesized to circumvent worker policingby mimicking the queen egg-marking signal. We investigated thisphenomenon by relating chemical profiles of workers and theireggs to egg acceptability. We found that the ability of someworkers (anarchistic workers in queenright colonies and deviantworkers from a queenless colony) to lay more acceptable eggsis due to them producing significant amounts of queen-like estersfrom their Dufour's gland. These esters appear to be transferredto eggs during laying and increase egg survival. However, theseesters cannot be the normal queen egg-marking signal, as theyare generally absent from queen-laid eggs and only increasethe short-term persistence of worker-laid eggs, because only7–30% of anarchistic worker-laid eggs persisted to hatchingversus 91–92% of queen-laid eggs. All workers can producesome esters, but only workers that greatly increase their esterproduction lay more acceptable eggs. The production of estersappears to be a flexible response, as anarchistic workers rearedin queenless colonies did not increase their ester production,while some deviant workers in queenless colonies did increasetheir ester production.     

Queen pheromones affecting the production of queen-like secretion in workers

The honeybee queen pheromones promote both worker sterility and worker-like pheromone composition; in their absence workers become fertile and express the queen pheromones. Which of the queen pheromones regulate worker pheromone expression and how, is still elusive. Here we investigated how two queen pheromones, the mandibular and Dufour’s, singly or combined, affect worker ovarian activation and occurrence of queen-like Dufour’s esters. Although queen mandibular pheromone (QMP) alone, or combined with Dufour’s secretion, inhibited to some extent worker reproduction, neither was as effective as the queen. The effect of the queen pheromones on worker pheromone expression was limited to workers with developed ovaries. Here too, QMP and Dufour’s combined had the greatest inhibitory effect. In contrast, treatment with Dufour’s alone resulted in augmentation of esters in the workers. This is another demonstration that a pheromone emitted by one individual affects the rates of its production in another individual. Ester production was tightly coupled to ovarian development. However fertile workers from queenright or QMP-treated colonies had significantly higher amounts of esters in their Dufour’s gland than untreated queenless colonies. The fact that the queen or QMP exert greater suppression on signal production than on ovary activation, suggests disparate regulatory pathways, and presents a challenging ultimate as well as proximate questions.     

Juvenile hormone control of oögenesis in bumblebee workers, Bombus terrestris

In queenright workers of Bombus terrestris oögenesis is inhibited by the queen, the activity of the corpora allata is suppressed, and the resulting JH titer in the haemolymph remains low. In contrast, in queenless workers the JH production is stimulated on the first day of adult emergence, the JH titer increases, and eggs are rapidly formed. After injection of JH I in newly emerged workers oögenesis can also be induced in the presence of a queen in the same way as in queenless workers. The induced oögenesis is JH dosage dependent, dosages of less than 8 μg stimulate the production of vitellogenins, whereas a complete oögenesis can be induced by high dosages of about 50 μg. From studies on the rate of JH excretion it can be concluded that such high dosages must be injected to obtain the required JH titer during the whole egg maturation of 5 days. It is evident, therefore, that the initiation and the maintenance of oögenesis depends on JH.The injected JH ester is completely degraded to JH acid and JH diol mainly in the hindgut. The excretion starts quickly after the injection, but only traces of unchanged JH are excreted.From these results it can be suggested that a queen inhibits egg maturation in workers by suppressing the JH production in the corpora allata and thus lowering the JH titer. This influence also enables a queen to block oögenesis once stimulated, for instance in queenless workers. Breakdown of JH and excretion are not under the control of the queen and therefore neither of them play any rôle in regulating egg maturation in the worker caste.     

Identification of vitellogenin in the ant, Camponotus festinatus: changes in hemolymph proteins and fat body development in workers.

Vitellogenin has been identified in the ant Camponotus festinatus, both in queens and workers. In the workers, it is already present before adult eclosion in low concentrations (less than 1 microgram/microliter hemolymph). Vitellogenin and vitellin are immunologically identical and are composed of a single type of apoprotein with an apparent Mr = 185,000. The molecular weight of the native molecules was estimated as approximately 460,000 by pore limiting gradient electrophoresis. Vitellogenin was detected as a major protein in the hemolymph of young workers, both under queenright and queenless conditions. Thus, in spite of their sterility in the presence of the queen, C. festinatus workers are able to synthetize vitellogenin which is identical both in size and immunologically to the queen vitellogenin. About 6-7 weeks after adult eclosion, however, vitellogenin was usually undetectable in the hemolymph of queenright workers, particularly the minor workers, while it constituted about 30% of total protein in queenless workers. Protein concentration in the hemolymph of queenless insects increased up to 20-fold as compared to 1-day-old insects. Queenless workers also developed large amounts of perivisceral fat body, while queenright workers, particularly the minor workers, showed a dramatic fat body regression about 6 weeks after emergence.     

The queen in relation to wax secretion and comb building in honeybees

Summary The role of the queen in relation to wax secretion and comb building in honeybees was analyzed with respect to queen status (mated, virgin and dead queens and queenlessness), and pheromones of the head and abdominal tergite of queens. Worker variables considered were colony size, percentage of bees bearing wax scales, wax scale weight, and weight of constructed combs.The amount of wax recovered from festoon bees and the percentage of festoon bees bearing wax were independent of queen status, the pheromones of queens and access to the queen. Colonies with full access to freely moving mated queens always constructed significantly more comb than those headed by virgin or dead queens as well as all permutations of caged and division board queens whose mandibular glands and/or abdominal tergite glands were operative or not.Despite pheromonal similarity of virgin queens to mated ones, colonies headed by virgin queens constructed as little comb as did queenless colonies. The bouquets of the mandibular glands did not differ significantly among queens nor was the amount of comb constructed correlated with pheromonal bouquet. Comb building is greatest among colonies having full access to freely moving queens but the stimulus for such building is not attributable to the 90DA, 9HDA and 10HDA components of the queen's mandibular gland secretions.     

蜜蜂上颚腺及其分泌物研究进展

上颚腺是蜜蜂重要的外分泌腺体,其分泌物是维系蜂群社会性结构的重要物质。蜂王和工蜂上颚腺分泌物合成均以硬脂酸为合成前体,但在脂肪酸的β-氧化过程中表现出级型差异性,导致分泌物组分比例不同。蜂王上颚腺分泌物以9-羰基-2癸烯酸(9-ODA)为主,有吸引工蜂和雄蜂、抑制工蜂卵巢发育等作用;工蜂上颚腺分泌物以10-羟基-2癸烯酸(10-HDA)和10-羟基癸酸(10-HDAA)为主,是蜂王浆的重要组成部分。同时,这种具备典型级型差异的分泌物组成又具有级型间可塑性,在不同蜂种间也存在区别。近年来在转录水平和蛋白水平的一些研究进一步揭示了级型间差异的分子基础。针对蜜蜂上颚腺及其分泌物的研究在蜜蜂生物学、行为学和蜂产品质量控制等方面具有重要的意义。本文通过总结国内外相关研究进展,旨在为上颚腺分泌物的作用机制、生物合成机制等领域的进一步深入研究提供借鉴。     

Dose-dependent inhibition of emergency queen rearing by synthetic 9-ODA in the honey bee,Apis mellifera carnica

In the honey bee colony queen rearing is usually suppressed by releaser effects of the queen's pheromone. This is part of the dominance hierarchy maintaining the monogynous homeostasis. Under queenless conditions, the queen's control over the construction of emergency queen cells by the workers can be substitued by exposure to only one component of the mandibular pheromone secretion of a queen, the main compound (E)-9-oxo-2-decenoic acid. A novel and simple synthesis of (E)-9-oxo-2-decenoic acid is described, and a bioassay was developed by which a dose-dependent effect of synthetic (E)-9-oxo-2-decenoic acid presented on a dummy bee was evaluated.Abbreviation 9-ODA (E)-9-oxo-2-decenoic acid In memoriam Viktor Schwartz (1907–1992), Professor of Zoology and Developmental Biology, University of Tübingen, who introduced smoothened bee stings into microsurgery     

Movement of honey bee (Apis mellifera L.) queen mandibular gland pheromone in populous and unpopulous colonies

The mode of intranest transfer of the honey bee queen mandibular gland pheromone complex (QMP) was investigated in unpopulous and populous, slightly congested colonies, using synthetic QMP containing tritiated 9-keto-2(E)-decenoic acid, one of the QMP components. Radiolabel was rapidly transported from the center to the peripheral regions of the nest, and in a manner consistent with worker to worker transport. Population size and congestion had no effect on the relative rates of movement from the center to the periphery of the nest or on the mean amounts of radiolabel on individual bees. However, a significantly smaller proportion of the workers in the populous colonies received detectable amounts of radiolabel than in the uncongested colonies, and workers carrying especially large amounts of radiolabel were less numerous in the crowded colonies. It is suggested that, at the stage of colony development that the colonies were in, population size has more of an effect on intranest pheromone transmission than does crowding. Interference with pheromone transfer may occur only at higher levels of congestion than were created, and nearer to the reproductive phase of colony development. An alternative hypothesis is that colony crowding does not significantly affect QMP transport and that the onset of reproductive queen rearing may be associated more with changes in worker thresholds of response to QMP.     

Effect of queen excluders on ovary activation in workers of the Eastern honeybee Apis cerana

Honeybee workers generally refrain from personal reproduction when a queen is present. Workers discern the presence and fecundity of the queen via volatile pheromones that permeate throughout the colony. Pheromones are emitted both by the queen herself and by the brood that she produces. If pheromone production is disrupted, some workers initiate egg laying. The Eastern honeybee Apis cerana is unusual in that workers have high levels of ovary activation even in the presence of a queen. To investigate the effect of disruption to pheromone dispersal, we fitted three A. cerana colonies with vertical queen excluders, thus splitting the colonies into a half containing a queen and a half without a queen. We regularly sampled adult workers from both sides of the excluder for 3 weeks. We also sampled workers from three control colonies that did not contain excluders. We found a significant increase in worker ovary activation 3 days after addition of excluders, suggesting that the reduced dispersal of pheromones allowed some workers to become reproductively active. Workers attempted to rear queen cells on the queenless halves of all three colonies. Queen-rearing ceased on day 9, at which time no queen-laid brood remained on the queenless halves of the colonies. Ovary activation rates continued to climb until day 9 and then gradually began to decline. With the exception of one egg, we did not observe worker-laid brood on the queenless side of the colonies, suggesting that workers continued policing eggs laid by workers. We conclude that if the distribution of brood pheromone is impeded, workers prepare to supersede their queen, accompanied by high levels of worker ovary activation. However, because workers continue to police each other, high ovary activation does not result in worker-produced drones.     

Chemical Profiles of Two Pheromone Glands Are Differentially Regulated by Distinct Mating Factors in Honey Bee Queens (Apis mellifera L.)

Pheromones mediate social interactions among individuals in a wide variety of species, from yeast to mammals. In social insects such as honey bees, pheromone communication systems can be extraordinarily complex and serve to coordinate behaviors among many individuals. One of the primary mediators of social behavior and organization in honey bee colonies is queen pheromone, which is produced by multiple glands. The types and quantities of chemicals produced differ significantly between virgin and mated queens, and recent studies have suggested that, in newly mated queens, insemination volume or quantity can affect pheromone production. Here, we examine the long-term impact of different factors involved during queen insemination on the chemical composition of the mandibular and Dufour''s glands, two of the major sources of queen pheromone. Our results demonstrate that carbon dioxide (an anesthetic used in instrumental insemination), physical manipulation of genital tract (presumably mimicking the act of copulation), insemination substance (saline vs. semen), and insemination volume (1 vs. 8 µl) all have long-term effects on mandibular gland chemical profiles. In contrast, Dufour''s gland chemical profiles were changed only upon insemination and were not influenced by exposure to carbon dioxide, manipulation, insemination substance or volume. These results suggest that the chemical contents of these two glands are regulated by different neuro-physiological mechanisms. Furthermore, workers responded differently to the different mandibular gland extracts in a choice assay. Although these studies must be validated in naturally mated queens of varying mating quality, our results suggest that while the chemical composition of Dufour''s gland is associated with mating status, that of the mandibular glands is associated with both mating status and insemination success. Thus, the queen appears to be signaling both status and reproductive quality to the workers, which may impact worker behavior and physiology as well as social organization and productivity of the colony.     

意大利蜜蜂工蜂卵巢发育的研究

蜜蜂Apis mellifera L.蜂群中的工蜂卵巢发育和工蜂产卵现象受多种因素控制,了解其影响因素对养蜂生产具有重要意义。本研究将意大利蜜蜂蜂群设置为囚王群(tg1)、无王有子群(tg2)、无王无子群(tg3)以及正常有王蜂群(CK)4个试验组,通过对工蜂卵巢管的显微观察,确定不同处理组工蜂在不同时间段内卵巢的发育情况。结果表明:随着时间的延长,与CK处理组相比,tg3处理组中的工蜂卵巢发育水平最高,tg2次之,tg1最低;在31d时,4个处理组两两之间差异均达到显著水平(P<0.05)。tg1、tg2和tg3处理组中工蜂产卵前期时间分别为35、22和17d,而CK蜂群在试验期内未出现工蜂产卵现象;tg2和tg3处理组的工蜂产卵的封盖前期时间分别为8和6d,而tg1和CK组在试验期内未出现子房封盖现象。蜂群失王时间过长会刺激工蜂卵巢发育,并导致其产卵;蜂群的短期失王和蜂王老化也会刺激工蜂卵巢发育,但是刺激程度较低;蜂群中的蜂子能抑制工蜂卵巢管的发育,因此在蜂群短时间失王时可以适当地补充子脾延缓工蜂卵巢发育。     

Age at Which Larvae Are Orphaned Determines Their Development into Typical or Rebel Workers in the Honeybee (Apis mellifera L.)

In the honeybee, diploid larvae fed with royal jelly develop into reproductive queens, whereas larvae fed with royal jelly for three days only and subsequently with honey and pollen develop into facultatively sterile workers. A recent study showed that worker larvae fed in a queenless colony develop into another female polyphenic form: rebel workers. These rebel workers are more queenlike and have greater reproductive potential than normal workers. However, it was unclear whether larvae orphaned at any time during their feeding period can develop into rebels. To answer this question, the anatomical features of newly emerged workers reared in queenless conditions at different ages during the larval period were evaluated. Our results showed that larvae orphaned during the final four or more days of their feeding life develop into rebel workers with more ovarioles in their ovaries, smaller hypopharyngeal glands, and larger mandibular and Dufour’s glands compared with typical workers with low reproductive potential that were reared with a queen or orphaned at the third to last or a later day of feeding life.     

The pheromones of laying workers in two honeybee sister species: <Emphasis Type="Italic">Apis cerana</Emphasis> and <Emphasis Type="Italic">Apis mellifera</Emphasis>

When a honeybee colony loses its queen, workers activate their ovaries and begin to lay eggs. This is accompanied by a shift in their pheromonal bouquet, which becomes more queen like. Workers of the Asian hive bee Apis cerana show unusually high levels of ovary activation and this can be interpreted as evidence for a recent evolutionary arms race between queens and workers over worker reproduction in this species. To further explore this, we compared the rate of pheromonal bouquet change between two honeybee sister species of Apis cerana and Apis mellifera under queenright and queenless conditions. We show that in both species, the pheromonal components HOB, 9-ODA, HVA, 9-HDA, 10-HDAA and 10-HDA have significantly higher amounts in laying workers than in non-laying workers. In the queenright colonies of A. mellifera and A. cerana, the ratios (9-ODA)/(9-ODA + 9-HDA + 10-HDAA + 10-HDA) are not significantly different between the two species, but in queenless A. cerana colonies the ratio is significant higher than in A. mellifera, suggesting that in A. cerana, the workers’ pheromonal bouquet is dominated by the queen compound, 9-ODA. The amount of 9-ODA in laying A. cerana workers increased by over 585% compared with the non-laying workers, that is 6.75 times higher than in A. mellifera where laying workers only had 86% more 9-ODA compared with non-laying workers.     

Individual versus social pathway to honeybee worker reproduction (Apis mellifera): pollen or jelly as protein source for oogenesis?

Honeybee workers, Apis mellifera, can reproduce in queenless colonies. The production of queen-like pheromones may be associated with their reproductive activity and induce nestmates to respond by feeding them. Such frequent trophallaxis could supply their protein needs for oogenesis, constituting a social pathway to worker reproduction. However, some individuals can develop ovaries without producing queen pheromones. The consumption of protein-rich pollen could be an alternative solitary pathway for them to satisfy this dietary requirement. In order to investigate the way in which workers obtain proteins for oogenesis, we created orphaned worker groups and determined ovarian and pheromonal development in relation to pollen consumption of selected workers. Individuals that did not consume pollen had significantly more developed ovaries and produced significantly more queen mandibular pheromone than workers that fed directly on pollen. Our results suggest that workers producing queen-like secretions are fed trophallactically. However, reproductive workers that lacked queen pheromones had consumed little or no pollen, suggesting that they also obtained trophallaxis. Although pollen consumption might contribute to sustaining oogenesis, it does not appear to be sufficient. Trophallaxis as a means of obtaining proteins seems to be necessary to attain reproductive status in queenless honeybee colonies.     

First record of emergency queen rearing in stingless bees (Hymenoptera, Apinae, Meliponini)

Summary: This paper presents the first record of emergency queen rearing in stingless bees. The main steps of the event in Trigona (Frieseomelitta) varia (Lepeletier) queenless colonies are: 1) construction and provisioning of an auxiliary cell in close contact to that housing a late feeding-stage larva; 2) construction of a connection between the two cells; 3) ingestion of food stored in the auxiliary cell by the larva via this connection; 4) dismantling of the empty auxiliary cell by the workers; 5) gradual restructuring of the cell containing the extra-fed larva into a royal cell; 6) emergence of the imago after 25 days. In one case, the new gyne started ovipositing at 19 days old.     

Plasticity in caste-related exocrine secretion biosynthesis in the honey bee (Apis mellifera)

Plasticity of Dufour's gland secretion in the honey bee is correlated with the individual's plasticity. Queens and queenless (QL) egg-laying workers possess a bouquet of esters and hydrocarbons, whereas queenright (QR) workers produce exclusively hydrocarbons. The effects of social environment (QR vs. QL conditions) and possible physiological constraints on the gland were studied by following the biosynthesis of these classes of compounds in vivo and in vitro. Biosynthesis in vivo followed the prediction based on glandular chemistry. Queens and QL egg-laying workers, but not QR workers or QL foragers, showed incorporation of sodium acetate into both hydrocarbons and esters. In contrast, the in vitro studies revealed that, in addition to queens and QL egg-laying workers, QR nurses retained their ability to produce the queen characteristic esters. Although there was some ester production in foragers, it occurred to a lesser extent. It is possible that the glands in the older foragers undergo irreversible changes. The in vitro incubation also revealed a temporal activation of ester biosynthesis in QR workers. In these glands alcohols, corresponding to the alcohol moiety of the esters, predominated in short-term incubations but decreased as the amount of newly synthesized esters increased. In contrast, queens and QL egg-laying workers showed predominant incorporation into esters from the onset of incubation. Thus, expression within the workers' Dufour's gland is regulated. In the presence of a queen, ester production is inhibited. Once the queen is removed the physiologically unconstrained gland starts to biosynthesize the queen-specific esters after a certain lag needed for the build-up of precursors and the enzymatic machinery.