Social Modulation of Stress Reactivity and Learning in Young Worker Honey Bees

Alarm pheromone and its major component isopentylacetate induce stress-like responses in forager honey bees, impairing their ability to associate odors with a food reward. We investigated whether isopentylacetate exposure decreases appetitive learning also in young worker bees. While isopentylacetate-induced learning deficits were observed in guards and foragers collected from a queen-right colony, learning impairments resulting from exposure to this pheromone could not be detected in bees cleaning cells. As cell cleaners are generally among the youngest workers in the colony, effects of isopentylacetate on learning behavior were examined further using bees of known age. Adult workers were maintained under laboratory conditions from the time of adult emergence. Fifty percent of the bees were exposed to queen mandibular pheromone during this period, whereas control bees were not exposed to this pheromone. Isopentylacetate-induced learning impairments were apparent in young (less than one week old) controls, but not in bees of the same age exposed to queen mandibular pheromone. This study reveals young worker bees can exhibit a stress-like response to alarm pheromone, but isopentylacetate-induced learning impairments in young bees are suppressed by queen mandibular pheromone. While isopentylacetate exposure reduced responses during associative learning (acquisition), it did not affect one-hour memory retrieval.     

Worker-worker inhibition of honey bee behavioural development independent of queen and brood

Summary. Foragers inhibit the behavioural development of young adult worker honey bees, delaying the age at onset of foraging. But the similar effect caused by pheromones produced by both the queen and brood raised the possibility that some of the previously attributed forager effects might be due to queen, brood, or both. Here we studied whether physical contacts between young bees and old foragers can inhibit behavioural development while controlling for queen and brood effects. Results demonstrated that foragers inhibit the behavioural development of young adult worker bees independent of the queen and brood, via a mechanism that requires physical contact.Received 24 November 2003; revised 27 March 2004; accepted 21 April 2004.     

Age and Behavior of Honey Bees,Apis mellifera (Hymenoptera: Apidae), that Perform Vibration Signals on Queens and Queen Cells

The vibration signal of the honey bee (Apis mellifera) may play a central role in the regulation of queen behavior during reproductive swarming and supersedure. We examined honey bee workers that performed vibration signals on queens and developing queen cells in three observation hives, each containing a population of marked bees of known age. In all three colonies, workers of all ages greater than 2 d old could perform vibration signals on queens and queen cells. However, most signals were performed by a small proportion of the bees of greater than 10 d of age. Relatively few workers less than 10 d old vibrated queens and queen cells, even though this age-group is typically associated with queen care. Thus, the regulation of queen behavior by the vibration signal may occur primarily through a relatively small subset of older workers that, under most circumstances, have only limited involvement with queens. It is unclear what triggers the vibrating of queens. Workers producing vibration signals did not differ from same-age non-vibrating controls in rate of locomotion in the hive or in task performance, and they rarely engaged in foraging, even though the majority of observed bees were of foraging age; vibrators also did not spend more time with queens and queen cells compared with controls. Vibration signals performed on queens and queen cells therefore do not appear to be influenced by task performance or increased contact with queens.     

Chemical detection of the proteins and lipids in the fat body cells from workers of Attini ants (Hymenoptera: Formicidae)

The ultrastructural cytochemistry study and the chemical analysis carried out to examine the fat body of workers of the basal Attini Cyphomyrmex rimosus and Mycetarotes parallelus and the derived Acromyrmex disciger and Atta laevigata revealed very electrondense protein granules of varied sizes and shapes distributed throughout the cytoplasm of parietal and perivisceral trophocytes and oenocytes. Lipids were present in the cytoplasm of trophocytes as very electrondense granules. In M. parallelus, A. disciger, and A. laevigata, lipids were present as droplets. In parietal as well as perivisceral oenocytes, lipid granules were very electrondense. In A. laevigata, lipids were present as droplets. The chemical analysis using gas chromatography showed that the main lipid compounds present in trophocytes and oenocytes were saturated (myristic, palmitic, and stearic) and unsaturated fatty acids (oleic and linoleic). Also, in basal species, lipid compounds consisted of mainly saturated fatty acids, while in derived species saturated as well as unsaturated fatty acids were observed.     

Differential antennal proteome comparison of adult honeybee drone, worker and queen (Apis mellifera L.)

To understand the olfactory mechanism of honeybee antennae in detecting specific volatile compounds in the atmosphere, antennal proteome differences of drone, worker and queen were compared using 2-DE, mass spectrometry and bioinformatics. Therefore, 107 proteins were altered their expressions in the antennae of drone, worker and queen bees. There were 54, 21 and 32 up-regulated proteins in the antennae of drone, worker and queen, respectively. Proteins upregulated in the drone antennae were involved in fatty acid metabolism, antioxidation, carbohydrate metabolism and energy production, protein folding and cytoskeleton. Proteins upregulated in the antennae of worker and queen bees were related to carbohydrate metabolism and energy production while molecular transporters were upregulated in the queen antennae. Our results explain the role played by the antennae of drone is to aid in perceiving the queen sexual pheromones, in the worker antennae to assist for food search and social communication and in the queen antennae to help pheromone communication with the worker and the drone during the mating flight. This first proteomic study significantly extends our understanding of honeybee olfactory activities and the possible mechanisms played by the antennae in response to various environmental, social, biological and biochemical signals.     

The effects of honey bee (Apis mellifera L.) queen reproductive potential on colony growth

Reproduction in species of eusocial insects is monopolized by one or a few individuals, while the remaining colony tasks are performed by the worker caste. This reproductive division of labor is exemplified by honey bees (Apis mellifera L.), in which a single, polyandrous queen is the sole colony member that lays fertilized eggs. Previous work has revealed that the developmental fate of honey bee queens is highly plastic, with queens raised from younger worker larvae exhibiting higher measures in several aspects of reproductive potential compared to queens raised from older worker larvae. Here, we investigated the effects of queen reproductive potential (“quality”) on the growth and winter survival of newly established honey bee colonies. We did so by comparing the growth of colonies headed by “high-quality” queens (i.e., those raised from young worker larvae, which are more queen-like morphologically) to those headed by “low-quality” queens (i.e., those raised from older worker larvae, which are more worker-like morphologically). We confirmed that queens reared from young worker larvae were significantly larger in size than queens reared from old worker larvae. We also found a significant positive effect of queen grafting age on a colony’s production of worker comb, drone comb, and stored food (honey and pollen), although we did not find a statistically significant difference in the production of worker and drone brood, worker population, and colony weight. Our results provide evidence that in honey bees, queen developmental plasticity influences several important measures of colony fitness. Thus, the present study supports the idea that a honey bee colony can be viewed (at least in part) as the expanded phenotype of its queen, and thus selection acting predominantly at the colony level can be congruent with that at the individual level.     

Seasonal variation in the titers and biosynthesis of the primer pheromone ethyl oleate in honey bees

Honey bees allocate tasks along reproductive and non-reproductive lines: the queen mates and lays eggs, whereas the workers nurse the brood and forage for food. Among workers, tasks are distributed according to age: young workers nurse and old workers fly out and forage. This task distribution in the colony is further regulated by an increase in juvenile hormone III as workers age and by pheromones. One such compound is ethyl oleate (EO), a primer pheromone that delays the onset of foraging in young workers. EO is produced by foragers when they are exposed to ethanol (from fermented nectar) while gathering food. EO is perceived by younger bees via olfaction. We describe here the seasonal variation of EO production and the effects of Methoprene, a juvenile hormone analog. We found that honey bee workers biosynthesize more EO during the growing season than during the fall and winter months, reaching peak levels at late spring or summer. When caged workers were fed with syrup+d(6)-ethanol, labeled EO accumulated in the honey crop and large amounts exuded to the exoskeleton. Exuded levels were high for several hours after exposure to ethanol. Treatment with Methoprene increased the production of EO in worker bees, by speeding up its movement from biosynthetic sites to the exoskeleton, where EO evaporates. Crop fluid from bees collected monthly during the growing season showed a modest seasonal variation of in vitro EO biosynthetic activity that correlated with the dry and sunny periods during which bees could forage.     

Organization of the antennal lobe in the queen honey bee,Apis mellifera L. (Hymenoptera : Apidae)

Antennal afferent pathway topography was studied in the queen honey bee, Apis mellifera L. (Hymenoptera : Apidae) by staining with cobalt chloride applied directly to the cut antennal axons. Antennal lobe organization in the queen was compared with those in worker and drone bees. The organization is similar in queen and worker bees. For the first time in a female insect the possible existence of a macroglomerulus is shown, which may be involved in the processing of species-specific information. A comparative quantitative neuromorphological study of the glomeruli was performed between young queens (8-day old) and 1-yr-old queens. The mean volume of the glomeruli is 46% greater in the older queens.     

Fat body cells of female reproductive castes of Attini ants (Hymenoptera: Formicidae): An ultrastructural and chemical analysis

The ultrastructural study on the fat body of gynes (virgin queens) of the basal ant species Cyphomyrmex rimosus and Mycetarotes parallelus and the derived Acromyrmex disciger and Atta laevigata queens showed vesicular rough endoplasmic reticulum, Golgi complex, and mitochondria in trophocytes, suggesting the involvement of these cells in protein synthesis, in addition to digestive vacuoles associated with the digestion of endocytosed compounds or rejected cell organelles. Oenocytes, another cell type present in the fat body of these species exhibit mitochondria, digestive vacuoles, and vesicles, indicating a mobilization of compounds by these cells. In A. laevigata, oenocytes also exhibited large storage sites of glycogen, in addition to a well-developed vesicular rough endoplasmic reticulum, suggesting an intensive participation of these cells in protein synthesis. The ultrastructural cytochemistry study also revealed electrodense granules of basic proteins present throughout the cytoplasm of trophocytes. The same was observed in oenocytes, although with smaller amounts of proteins. In the cytoplasm of trophocytes and oenocytes were also found droplets or electrodense granules of lipids. In oenocytes of A. disciger and in trophocytes of A. laevigata, lipids were observed in mitochondria, suggesting that this organelle might be a site of synthesis of these compounds. The chemical analysis of lipids revealed that in gynes, the main compounds present in fat body cells were saturated fatty acids, while in queens, saturated as well as unsaturated fatty acids were found. In conclusion, the present study showed that the fat body cells of gynes and queens, in general, maintained the same compounds and original features through the evolution process of the Attini tribe.     

No intracolonial nepotism during colony fissioning in honey bees

Most species of social insects have singly mated queens, but in some species each queen mates with numerous males to create a colony whose workers belong to multiple patrilines. This colony genetic structure creates a potential for intracolonial nepotism. One context with great potential for such nepotism arises in species, like honey bees, whose colonies reproduce by fissioning. During fissioning, workers might nepotistically choose between serving a young (sister) queen or the old (mother) queen, preferring the former if she is a full-sister but the latter if the young queen is only a half-sister. We examined three honeybee colonies that swarmed, and performed paternity analyses on the young (immature) queens and samples of workers who either stayed with the young queens in the nest or left with the mother queen in the swarm. For each colony, we checked whether patrilines represented by immature queens had higher proportions of staying workers than patrilines not represented by immature queens. We found no evidence of this. The absence of intracolonial nepotism during colony fissioning could be because the workers cannot discriminate between full-sister and half-sister queens when they are immature, or because the costs of behaving nepotistically outweigh the benefits.     

A comparative study of fat body morphology in five mosquito species

The insect fat body plays major roles in the intermediary metabolism, in the storage and transport of haemolymph compounds and in the innate immunity. Here, the overall structure of the fat body of five species of mosquitoes (Aedes albopictus, Aedes fluviatilis, Culex quinquefasciatus, Anopheles aquasalis and Anopheles darlingi) was compared through light, scanning and transmission electron microscopy. Generally for mosquitoes, the fat body consists of lobes projecting into the haemocoel and is formed by great cell masses consisting of trophocytes and oenocytes. Trophocytes are rich in lipid droplets and protein granules. Interestingly, brown pigment granules, likely ommochromes, were found exclusively in the trophocytes located within the thorax and near the dorsal integument of Anopheles, which is suggestive of the role these cells play in detoxification via ommochrome storage. This study provides a detailed comparative analysis of the fat body in five different mosquito species and represents a significant contribution towards the understanding of the structural-functional relationships associated with this organ.     

Making a queen: an epigenetic analysis of the robustness of the honeybee (Apis mellifera) queen developmental pathway

Specialized castes are considered a key reason for the evolutionary and ecological success of the social insect lifestyle. The most essential caste distinction is between the fertile queen and the sterile workers. Honeybee (Apis mellifera) workers and queens are not genetically distinct, rather these different phenotypes are the result of epigenetically regulated divergent developmental pathways. This is an important phenomenon in understanding the evolution of social insect societies. Here, we studied the genomic regulation of the worker and queen developmental pathways, and the robustness of the pathways by transplanting eggs or young larvae to queen cells. Queens could be successfully reared from worker larvae transplanted up to 3 days age, but queens reared from older worker larvae had decreased queen body size and weight compared with queens from transplanted eggs. Gene expression analysis showed that queens raised from worker larvae differed from queens raised from eggs in the expression of genes involved in the immune system, caste differentiation, body development and longevity. DNA methylation levels were also higher in 3‐day‐old queen larvae raised from worker larvae compared with that raised from transplanted eggs identifying a possible mechanism stabilizing the two developmental paths. We propose that environmental (nutrition and space) changes induced by the commercial rearing practice result in a suboptimal queen phenotype via epigenetic processes, which may potentially contribute to the evolution of queen–worker dimorphism. This also has potentially contributed to the global increase in honeybee colony failure rates.     

Differential protein expression in honeybee (Apis mellifera L.) larvae: underlying caste differentiation

Honeybee (Apis mellifera) exhibits divisions in both morphology and reproduction. The queen is larger in size and fully developed sexually, while the worker bees are smaller in size and nearly infertile. To better understand the specific time and underlying molecular mechanisms of caste differentiation, the proteomic profiles of larvae intended to grow into queen and worker castes were compared at 72 and 120 hours using two dimensional electrophoresis (2-DE), network, enrichment and quantitative PCR analysis. There were significant differences in protein expression between the two larvae castes at 72 and 120 hours, suggesting the queen and the worker larvae have already decided their fate before 72 hours. Specifically, at 72 hours, queen intended larvae over-expressed transketolase, aldehyde reductase, and enolase proteins which are involved in carbohydrate metabolism and energy production, imaginal disc growth factor 4 which is a developmental related protein, long-chain-fatty-acid CoA ligase and proteasome subunit alpha type 5 which metabolize fatty and amino acids, while worker intended larvae over-expressed ATP synthase beta subunit, aldehyde dehydrogenase, thioredoxin peroxidase 1 and peroxiredoxin 2540, lethal (2) 37 and 14-3-3 protein epsilon, fatty acid binding protein, and translational controlled tumor protein. This differential protein expression between the two caste intended larvae was more pronounced at 120 hours, with particular significant differences in proteins associated with carbohydrate metabolism and energy production. Functional enrichment analysis suggests that carbohydrate metabolism and energy production and anti-oxidation proteins play major roles in the formation of caste divergence. The constructed network and validated gene expression identified target proteins for further functional study. This new finding is in contrast to the existing notion that 72 hour old larvae has bipotential and can develop into either queen or worker based on epigenetics and can help us to gain new insight into the time of departure as well as caste trajectory influencing elements at the molecular level.     

Physiological correlates of division of labor among similarly aged honey bees

Hormone analyses and exocrine gland measurements were made to probe for physiological correlates of division of labor among similarly aged adult worker honey bees (Apis mellifera L.). Middle-age bees (ca. 2 weeks old) performing different tasks showed significant differences in both juvenile hormone (JH) biosynthesis rates and hemolymph titers; guards and undertakers had high JH, and wax producers and food storers, low JH. Guards and undertakers had similar hormone levels to foragers, even though they were 10 days younger than foragers. No differences in JH were detected among young bees (1-week-old queen attendants and nurses) or older bees (3–4 week-old pollen foragers, non-pollen foragers, and soldiers). Hypopharyngeal gland size was inversely correlated with worker age and rate of JH biosynthesis, but soldiers had significantly larger hypopharyngeal glands than did foragers, despite their similar age and JH level. Results from soldiers indicate that exocrine gland development is not always linked with age-related behavior and endocrine development; they also support the recent claim that soldiers constitute a group of older bees that are distinct from foragers. Hormonal analyses indicate that the current model of JH's role in honey bee division of labor needs to be expanded because high levels of JH are associated with several other tasks besides foraging. JH may be involved in the regulation of division of labor among similarly aged workers in addition to its role in age-related division of labor.Abbreviations JH Juvenile hormone - RIA radioimmunoassay - CA corpora allata - HPLC high performance liquid chromatography - TLC thin layer chromatography     

家白蚁群体数增长的生殖生理特点

家白蚁Coptotermes formosanus Shiraki营大巢生活,由于雌虫生殖潜能很大,群体蚁数极多.群体蚁数的逐年增长和雌蚁卵巢内卵巢管的数目增长有密切关系.实验表明,分飞时的雌虫卵巢有8条卵巢管,初建群体,半年内一群体的蚁数仅为57个.经过四年后发育的群体,蚁数是1,767个,这时的蚁后卵巢管数是75条.二年群体的年幼蚁后腹部稍膨大,三年以上的群体,蚁后的1腹部膨大较显著.一年的群体,雌虫体长是0.78厘米,雄虫体长0.71厘米.九年后的群体,蚁后体长是2.1厘米,蚁王体长为1.2厘米.和卵巢相似,雄虫精巢叶的数目也随时间而增加.     

Effect of royal jelly on longevity and memory-related traits of Apis mellifera workers

Royal jelly (RJ) is a key factor for honey bee caste determination. The queen bee is fed with RJ by worker bees throughout her life, while the worker bees eat bee bread themselves. This study was designed to explore the effect of nutrient-rich RJ on longevity and learning and memory abilities of workers of the western honey bee Apis mellifera. The newly emerged worker bees were randomly divided into three groups and were fed 50% sucrose solution containing 0%, 10%, and 20% RJ. We found that worker bees fed with 10% and 20% RJ showed significantly improved longevity and higher proboscis extension response success rate compared to bees fed with 50% sucrose containing 0% RJ. Additionally, bees fed with 20% RJ showed significantly higher level of expression of memory related genes (GluRA and Nmdar1) compared to the control group. Furthermore, expression of the Nmdar1 gene of worker bees fed with 10% RJ was also significantly higher than in the control group. These results indicate that RJ has potential effects on the longevity and learning and memory abilities of A. mellifera.     

Metabolism and longevity: is there a role for membrane fatty acids?

More than 100 years ago, Max Rubner combined the fact that both metabolic rate and longevity of mammals varies with body size to calculate that "life energy potential" (lifetime energy turnover per kilogram) was relatively constant. This calculation linked longevity to aerobic metabolism which in turn led to the "rate-of-living" and ultimately the "oxidative stress" theories of aging. However, the link between metabolic rate and longevity is imperfect. Although unknown in Rubner's time, one aspect of body composition of mammals also varies with body size, namely the fatty acid composition of membranes. Fatty acids vary dramatically in their susceptibility to peroxidation and the products of lipid peroxidation are very powerful reactive molecules that damage other cellular molecules. The "membrane pacemaker" modification of the "oxidative stress" theory of aging proposes that fatty acid composition of membranes, via its influence on peroxidation of lipids, is an important determinant of lifespan (and a link between metabolism and longevity). The relationship between membrane fatty acid composition and longevity is discussed for (1) mammals of different body size, (2) birds of different body size, (3) mammals and birds that are exceptionally long-living for their size, (4) strains of mice that vary in longevity, (5) calorie-restriction extension of longevity in rodents, (6) differences in longevity between queen and worker honeybees, and (7) variation in longevity among humans. Most of these comparisons support an important role for membrane fatty acid composition in the determination of longevity. It is apparent that membrane composition is regulated for each species. Provided the diet is not deficient in polyunsaturated fat, it has minimal influence on a species' membrane fatty acid composition and likely also on it's maximum longevity. The exceptional longevity of Homo sapiens combined with the limited knowledge of the fatty acid composition of human tissues support the potential importance of mitochondrial membranes in determination of longevity.     

Interadult feeding of jelly in honeybee (Apis mellifera L.) colonies

Summary Honeybee nurses (8 days old) were injected with ¹⁴C-phenylalanine. These bees then dispensed the ¹⁴C-labelled protein-rich products of their hypopharyngeal glands to the queen and the brood, and also to young drones and workers of all age classes. In small colonies containing 400–800 bees, nearly one-quarter of the radioactivity which could not be recovered in the nurses was fed by them in a protein-bound form to other members of the worker caste. During one night, one nurse fed an average of 4–5 foragers with proteinaceous food. The role of nurses in the work allotment system of honeybee colonies therefore needs a new, extended definition. Nurses are largely responsible for preparing nutrients from pollen, which is difficult to digest. They then distribute the nutritionally valuable protein produced by their hypopharyngeal glands to practically all hive mates.Dedicated to Professor Dr. O. Kepka on the occasion of his 65th birthday     

Queen Dominance May Reduce Worker Mushroom Body Size in a Social Bee

The mushroom body (MB) is an area of the insect brain involved in learning, memory, and sensory integration. Here, we used the sweat bee Megalopta genalis (Halictidae) to test for differences between queens and workers in the volume of the MB calyces. We used confocal microscopy to measure the volume of the whole brain, MB calyces, optic lobes, and antennal lobes of queens and workers. Queens had larger brains, larger MB calyces, and a larger MB calyces:whole brain ratio than workers, suggesting an effect of social dominance in brain development. This could result from social interactions leading to smaller worker MBs, or larger queen MBs. It could also result from other factors, such as differences in age or sensory experience. To test these explanations, we next compared queens and workers to other groups. We compared newly emerged bees, bees reared in isolation for 10 days, bees initiating new observation nests, and bees initiating new natural nests collected from the field to queens and workers. Queens did not differ from these other groups. We suggest that the effects of queen dominance over workers, rather than differences in age, experience, or reproductive status, are responsible for the queen–worker differences we observed. Worker MB development may be affected by queen aggression directly and/or manipulation of larval nutrition, which is provisioned by the queen. We found no consistent differences in the size of antennal lobes or optic lobes associated with differences in age, experience, reproductive status, or social caste.