Light exposure leads to reorganization of microglomeruli in the mushroom bodies and influences juvenile hormone levels in the honeybee

Honeybees show a remarkable behavioral plasticity at the transition from nursing inside the hive to foraging for nectar and/or pollen outside. This plasticity is important for age‐related division of labor in honeybee colonies. The behavioral transition is associated with significant volume and synaptic changes in the mushroom bodies (MBs), brain centers for sensory integration, learning, and memory. We tested whether precocious sensory exposure to light leads to changes in the density of synaptic complexes [microglomeruli (MG)] in the MBs. The results show that exposure to light pulses over 3 days induces a significant decrease in the MG density in visual subregions (collar) of the MB. Earlier studies had shown that foragers have increased levels of juvenile hormone (JH) co‐occurring with a decrease of vitellogenin (Vg). Previous work further established that RNAi‐mediated knockdown of vg and ultraspiracle (usp) induced an upregulation of JH levels, which can lead to precocious foraging. By disturbing both Vg and JH pathways using gene knockdown of vg and usp, we tested whether the changes in the hormonal system directly affect MG densities. Our study shows that MG numbers remained unchanged when Vg and JH pathways were perturbed, suggesting no direct hormonal influences on MG densities. However, mass spectrometry detection of JH revealed that precocious light exposure triggered an increase in JH levels in the hemolymph (HL) of young bees. This suggests a dual effect following light exposure via direct effects on MG reorganization in the MB calyx and a possible positive feedback on HL JH levels. © 2014 Wiley Periodicals, Inc. Develop Neurobiol 74: 1141–1153, 2014     

Octopamine influences division of labor in honey bee colonies

Forager honey bees have higher brain levels of octopamine than do bees tending larvae in the hive. To test the hypothesis that octopamine influences honey bee division of labor we treated bees orally with octopamine or its immediate precursor tyramine and determined whether these treatments increased the probability of initiating foraging. Octopamine treatment significantly elevated levels of octopamine in the brain and caused a significant dose-dependent increase in the number of new foragers. This effect was seen for precocious foragers in single-cohort colonies and foragers in larger colonies with more typical age demographies. Tyramine treatment did not increase the number of new foragers, suggesting that octopamine was exerting a specific effect. Octopamine treatment was effective only when given to bees old enough to forage, i.e., older than 4 days of age. Treatment when bees were 1-3 days of age did not cause a significant increase in the number of new foragers when the bees reached the minimal foraging age. These results demonstrate that octopamine influences division of labor in honey bee colonies. We speculate that octopamine is acting in this context as a neuromodulator.     

Effects of experience and juvenile hormone on the organization of the mushroom bodies of honey bees

There is an age-related division of labor in the honey bee colony that is regulated by juvenile hormone. After completing metamorphosis, young workers have low titers of juvenile hormone and spend the first several weeks of their adult lives performing tasks within the hive. Older workers, approximately 3 weeks of age, have high titers of juvenile hormone and forage outside the hive for nectar and pollen. We have previously reported that changes in the volume of the mushroom bodies of the honey bee brain are temporally associated with the performance of foraging. The neuropil of the mushroom bodies is increased in volume, whereas the volume occupied by the somata of the Kenyon cells is significantly decreased in foragers relative to younger workers. To study the effect of flight experience and juvenile hormone on these changes within the mushroom bodies, young worker bees were treated with the juvenile hormone analog methoprene but a subset was prevented from foraging (big back bees). Stereological volume estimates revealed that, regardless of foraging experience, bees treated with methoprene had a significantly larger volume of neuropil in the mushroom bodies and a significantly smaller Kenyon cell somal region volume than did 1-day-old bees. The bees treated with methoprene did not differ on these volume estimates from untreated foragers (presumed to have high endogenous levels of juvenile hormone) of the same age sampled from the same colony. Bees prevented from flying and foraging nonetheless received visual stimulation as they gathered at the hive entrance. These results, coupled with a subregional analysis of the neuropil, suggest a potentially important role of visual stimulation, possibly interacting with juvenile hormone, as an organizer of the mushroom bodies. In an independent study, the brains of worker bees in which the transition to foraging was delayed (overaged nurse bees) were also studied. The mushroom bodies of overaged nurse bees had a Kenyon cell somal region volume typical of normal aged nurse bees. However, they displayed a significantly expanded neuropil relative to normal aged nurse bees. Analysis of the big back bees demonstrates that certain aspects of adult brain plasticity associated with foraging can be displayed by worker bees treated with methoprene independent of foraging experience. Analysis of the over-aged nurse bees suggests that the post-metamorphic expansion of the neuropil of the mushroom bodies of worker honey bees is not a result of foraging experience. © 1995 John Wiley & Sons, Inc.     

Bees get a head start on honey production

Nectar concentration is assumed to remain constant during transport by honeybees between flowers and hive. We sampled crop contents of nectar foragers on Aloe greatheadii var. davyana, a major winter bee plant in South Africa. The nectar is dilute (approx. 20% w/w), but the crop contents of bees captured on flowers are significantly more concentrated. In returning foragers, the concentration increases further to 38–40%, accompanied by a volume decrease. The doubling of sugar concentration suggests that nectar is regurgitated onto the tongue and evaporated during foraging and on the return flight. Processing of the dilute nectar into honey thus begins early, aided by low ambient humidities. This has implications for honeybee thermoregulation, water balance and energetics during foraging, and for the communication of nectar quality to recruits.     

Cover Caption

The Western honey bee, Apis mellifera (L.), is perhaps the most beneficial insect we know, mainly because of the pollination services it provides to fruits and vegetables. Honey bee workers show changes in behaviors as they age. Young bees typically are “nurses” and perform in‐hive tasks such as feeding larvae and take care of the queen, old bees become foragers and bring in food sources (nectar, pollen, and water) or propolis. Methoprene has been known to accelerate worker development so bees become foragers earlier, but its mechanisms were not known. In this study Huang et al. show that most likely methoprene works directly on hormone receptors to mimic juvenile hormone, in causing bees to forage early (pages 235–240). Photo by Zachary Y. Huang. [Correction added on 17 April 2018, after first online publication: Cover caption has been revised.]     

Experience-dependent plasticity in the mushroom bodies of the solitary bee Osmia lignaria (Megachilidae)

All members of the solitary bee species Osmia lignaria (the orchard bee) forage upon emergence from their natal nest cell. Conversely, in the honey bee, days-to-weeks of socially regulated behavioral development precede the onset of foraging. The social honey bee's behavioral transition to foraging is accompanied by neuroanatomical changes in the mushroom bodies, a region of the insect brain implicated in learning. If these changes were general adaptations to foraging, they should also occur in the solitary orchard bee. Using unbiased stereological methods, we estimated the volume of the major compartments of the mushroom bodies, the neuropil and Kenyon cell body region, in adult orchard bees. We compared the mushroom bodies of recently emerged bees with mature bees that had extensive foraging experience. To separate effects of general maturation from field foraging, some orchard bees were confined to a cage indoors. The mushroom body neuropil of experienced field foragers was significantly greater than that of both recently emerged and mature caged orchard bees, suggesting that, like the honey bee, this increase is driven by outdoor foraging experience. Unlike the honey bee, where increases in the ratio of neuropil to Kenyon cell region occur in the worker after emerging from the hive cell, the orchard bee emerged from the natal nest cell with a ratio that did not change with maturation and was comparable to honey-bee foragers. These results suggest that a common developmental endpoint may be reached via different development paths in social and solitary species of foraging bees.     

Competition between honey bees and wild bees and the role of nesting resources in a nature reserve

The European honey bee exploits floral resources efficiently and may therefore compete with solitary wild bees. Hence, conservationists and bee keepers are debating about the consequences of beekeeping for the conservation of wild bees in nature reserves. We observed flower-visiting bees on flowers of Calluna vulgaris in sites differing in the distance to the next honey-bee hive and in sites with hives present and absent in the Lüneburger Heath, Germany. Additionally, we counted wild bee ground nests in sites that differ in their distance to the next hive and wild bee stem nests and stem-nesting bee species in sites with hives present and absent. We did not observe fewer honey bees or higher wild bee flower visits in sites with different distances to the next hive (up to 1,229 m). However, wild bees visited fewer flowers and honey bee visits increased in sites containing honey-bee hives and in sites containing honey-bee hives we found fewer stem-nesting bee species. The reproductive success, measured as number of nests, was not affected by distance to honey-bee hives or their presence but by availability and characteristics of nesting resources. Our results suggest that beekeeping in the Lüneburg Heath can affect the conservation of stem-nesting bee species richness but not the overall reproduction either of stem-nesting or of ground-nesting bees. Future experiments need control sites with larger distances than 500 m to hives. Until more information is available, conservation efforts should forgo to enhance honey bee stocking rates but enhance the availability of nesting resources.     

Viral Infection Affects Sucrose Responsiveness and Homing Ability of Forager Honey Bees,Apis mellifera L.

Honey bee health is mainly affected by Varroa destructor, viruses, Nosema spp., pesticide residues and poor nutrition. Interactions between these proposed factors may be responsible for the colony losses reported worldwide in recent years. In the present study, the effects of a honey bee virus, Israeli acute paralysis virus (IAPV), on the foraging behaviors and homing ability of European honey bees (Apis mellifera L.) were investigated based on proboscis extension response (PER) assays and radio frequency identification (RFID) systems. The pollen forager honey bees originated from colonies that had no detectable level of honey bee viruses and were manually inoculated with IAPV to induce the viral infection. The results showed that IAPV-inoculated honey bees were more responsive to low sucrose solutions compared to that of non-infected foragers. After two days of infection, around 10⁷ copies of IAPV were detected in the heads of these honey bees. The homing ability of IAPV-infected foragers was depressed significantly in comparison to the homing ability of uninfected foragers. The data provided evidence that IAPV infection in the heads may enable the virus to disorder foraging roles of honey bees and to interfere with brain functions that are responsible for learning, navigation, and orientation in the honey bees, thus, making honey bees have a lower response threshold to sucrose and lose their way back to the hive.     

The ontogeny of immunity: development of innate immune strength in the honey bee (Apis mellifera)

Honey bees (Apis mellifera) are of vital economic and ecological importance. These eusocial animals display temporal polyethism, which is an age-driven division of labor. Younger adult bees remain in the hive and tend to developing brood, while older adult bees forage for pollen and nectar to feed the colony. As honey bees mature, the types of pathogens they experience also change. As such, pathogen pressure may affect bees differently throughout their lifespan. We provide the first direct tests of honey bee innate immune strength across developmental stages. We investigated immune strength across four developmental stages: larvae, pupae, nurses (1-day-old adults), and foragers (22-30 days old adults). The immune strength of honey bees was quantified using standard immunocompetence assays: total hemocyte count, encapsulation response, fat body quantification, and phenoloxidase activity. Larvae and pupae had the highest total hemocyte counts, while there was no difference in encapsulation response between developmental stages. Nurses had more fat body mass than foragers, while phenoloxidase activity increased directly with honey bee development. Immune strength was most vigorous in older, foraging bees and weakest in young bees. Importantly, we found that adult honey bees do not abandon cellular immunocompetence as has recently been proposed. Induced shifts in behavioral roles may increase a colony's susceptibility to disease if nurses begin foraging activity prematurely.     

The pollination efficiency of a pollinator depends on its foraging strategy,flowering phenology,and the flower characteristics of a plant species

Honey bees and stingless bees are generalist visitors of several wild and cultivated plants. They forage with a high degree of floral fidelity and thereby help in the pollination services of those plants. We hypothesized that pollination efficiency might be influenced by flowering phenology, floral characteristics, and resource collection modes of the worker bees. In this paper, we surveyed the foraging strategies of honey bees (Apis cerana, Apis dorsata, and Apis florea) and stingless bees (Tetragonula iridipennis) concerning their pollination efficiencies. Bees showed different resource gathering strategies, including legitimate (helping in pollination as mixed foragers and specialized foragers) and illegitimate (serving as nectar robbers and pollen thieves) types of flower visitation patterns. Foraging strategies are influenced by the shape of flowers, the timing of the visitation, floral richness, and bee species. Honey bees and stingless bees mainly acted as legitimate visitors in most plants studied. Sometimes honey bees served as nectar robbers in tubular flowers and stingless bees as pollen thieves in large-sized flowers. Among the legitimate categories, mixed foragers have a comparatively lower flower visitation rate than the specialized nectar and pollen foragers. However, mixed foragers have greater abundance and higher values of the single-visit pollination efficiency index (PEi) than nectar and pollen foragers. The value of the combined parameter ‘importance in pollination (PI)’ was thus higher in mixed foragers than in nectar and pollen foragers.     

Pollinator genetics and pollination: do honey bee colonies selected for pollen‐hoarding field better pollinators of cranberry Vaccinium macrocarpon?

1. Genetic polymorphisms of flowering plants can influence pollinator foraging but it is not known whether heritable foraging polymorphisms of pollinators influence their pollination efficacies. Honey bees Apis mellifera L. visit cranberry flowers for nectar but rarely for pollen when alternative preferred flowers grow nearby. 2. Cranberry flowers visited once by pollen‐foraging honey bees received four‐fold more stigmatic pollen than flowers visited by mere nectar‐foragers (excluding nectar thieves). Manual greenhouse pollinations with fixed numbers of pollen tetrads (0, 2, 4, 8, 16, 32) achieved maximal fruit set with just eight pollen tetrads. Pollen‐foraging honey bees yielded a calculated 63% more berries than equal numbers of non‐thieving nectar‐foragers, even though both classes of forager made stigmatic contact. 3. Colonies headed by queens of a pollen‐hoarding genotype fielded significantly more pollen‐foraging trips than standard commercial genotypes, as did hives fitted with permanently engaged pollen traps or colonies containing more larvae. Pollen‐hoarding colonies together brought back twice as many cranberry pollen loads as control colonies, which was marginally significant despite marked daily variation in the proportion of collected pollen that was cranberry. 4. Caloric supplementation of matched, paired colonies failed to enhance pollen foraging despite the meagre nectar yields of individual cranberry flowers. 5. Heritable behavioural polymorphisms of the honey bee, such as pollen‐hoarding, can enhance fruit and seed set by a floral host (e.g. cranberry), but only if more preferred pollen hosts are absent or rare. Otherwise, honey bees' broad polylecty, flight range, and daily idiosyncrasies in floral fidelity will obscure specific pollen‐foraging differences at a given floral host, even among paired colonies in a seemingly uniform agricultural setting.     

Hydrocarbons Emitted by Waggle-Dancing Honey Bees Increase Forager Recruitment by Stimulating Dancing

Hydrocarbons emitted by waggle-dancing honey bees are known to reactivate experienced foragers to visit known food sources. This study investigates whether these hydrocarbons also increase waggle-dance recruitment by observing recruitment and dancing behavior when the dance compounds are introduced into the hive. If the hydrocarbons emitted by waggle-dancing bees affect the recruitment of foragers to a food source, then the number of recruits arriving at a food source should be greater after introduction of dance compounds versus a pure-solvent control. This prediction was supported by the results of experiments in which recruits were captured at a feeder following introduction of dance-compounds into a hive. This study also tested two nonexclusive behavioral mechanism(s) by which the compounds might stimulate recruitment; 1) increased recruitment could occur by means of increasing the recruitment effectiveness of each dance and/or 2) increased recruitment could occur by increasing the intensity of waggle-dancing. These hypotheses were tested by examining video records of the dancing and recruitment behavior of individually marked bees following dance-compound introduction. Comparisons of numbers of dance followers and numbers of recruits per dance and waggle run showed no significant differences between dance-compound and solvent-control introduction, thus providing no support for the first hypothesis. Comparison of the number of waggle-dance bouts and the number of waggle runs revealed significantly more dancing during morning dance-compound introduction than morning solvent-control introduction, supporting the second hypothesis. These results suggest that the waggle-dance hydrocarbons play an important role in honey bee foraging recruitment by stimulating foragers to perform waggle dances following periods of inactivity.     

Red mason bees cannot compete with honey bees for floral resources in a cage experiment

Intensive beekeeping to mitigate crop pollination deficits and habitat loss may cause interspecific competition between bees. Studies show negative correlations between flower visitation of honey bees (Apis mellifera) and wild bees, but effects on the reproduction of wild bees were not proven. Likely reasons are that honey bees can hardly be excluded from controls and wild bee nests are generally difficult to detect in field experiments. The goal of this study was to investigate whether red mason bees (Osmia bicornis) compete with honey bees in cages in order to compare the reproduction of red mason bees under different honey bee densities. Three treatments were applied, each replicated in four cages of 18 m³ with 38 red mason bees in all treatments and 0, 100, and 300 honey bees per treatment with 10–20% being foragers. Within the cages, the flower visitation and interspecific displacements from flowers were observed. Niche breadths and resource overlaps of both bee species were calculated, and the reproduction of red mason bees was measured. Red mason bees visited fewer flowers when honey bees were present. Niche breadth of red mason bees decreased with increasing honey bee density while resource overlaps remained constant. The reproduction of red mason bees decreased in cages with honey bees. In conclusion, our experimental results show that in small and isolated flower patches, wild bees can temporarily suffer from competition with honey bees. Further research should aim to test for competition on small and isolated flower patches in real landscapes.     

Juvenile hormone levels in honey bee (Apis mellifera L.) foragers: foraging experience and diurnal variation

A rising blood titer of juvenile hormone (JH) in adult worker honey bees is associated with the shift from working in the hive to foraging. We determined whether the JH increase occurs in anticipation of foraging or whether it is a result of actual foraging experience and/or diurnal changes in exposure to sunlight. We recorded all foraging flights of tagged bees observed at a feeder in a large outdoor flight cage. We measured JH from bees that had taken 1, 3-5, or >100 foraging flights and foragers of indeterminate experience leaving or entering the hive. To study diurnal variation in JH, we sampled foragers every 6h over one day. Titers of JH in foragers were high relative to nurses as in previous studies, suggesting that conditions in the flight cage had no effect on the relationship between foraging behavior and JH. Titers of JH in foragers showed no significant effects of foraging experience, but did show significant diurnal variation. Our results indicate that the high titer of JH in foragers anticipates the onset of foraging and is not affected by foraging experience, but is modulated diurnally.     

Biogenic amines and division of labor in honey bee colonies

Brain levels of dopamine, serotonin, and octopamine were measured in relation to both age-related division of labor and inter-individual differences in task specialization independent of age in honey bee colonies. The only differences among similarly aged bees performing different tasks were significantly lower levels of dopamine in food storers than comb builders and significantly lower levels of octopamine in soldiers than foragers, but soldiers also were slightly younger than foragers. Differences associated with age-related division of labor were stronger. Older bees, notably foragers, had significantly higher levels of all three amines than did younger bees working in the hive. Using social manipulations to unlink chronological age and behavioral status, octopamine was found to exhibit the most robust association between behavior and amine level, independent of age. Octopamine levels were significantly lower in normal-age nurses versus precocious foragers and overage nurses versus normal-age foragers, but not different in reverted nurses versus reversion colony foragers. Dopamine levels were significantly lower in normal-age nurses versus precocious foragers, but higher in reverted nurses versus reversion colony foragers. Serotonin levels did not differ in any of these comparisons. These correlative results suggest that octopamine is involved in the regulation of age-related division of labor in honey bees. Accepted: 10 February 1999     

Neuronal plasticity in the mushroom body calyx during adult maturation in the honeybee and possible pheromonal influences

Honeybee workers express a pronounced age‐dependent polyethism switching from various indoor duties to foraging outside the hive. This transition is accompanied by tremendous changes in the sensory environment that sensory systems and higher brain centers have to cope with. Foraging and age have earlier been shown to be associated with volume changes in the mushroom bodies (MBs). Using age‐ and task‐controlled bees this study provides a detailed framework of neuronal maturation processes in the MB calyx during the course of natural behavioral maturation. We show that the MB calyx volume already increases during the first week of adult life. This process is mainly driven by broadening of the Kenyon cell dendritic branching pattern and then followed by pruning of projection neuron axonal boutons during the actual transition from indoor to outdoor duties. To further investigate the flexible regulation of division of labor and its neuronal correlates in a honeybee colony, we studied the modulation of the nurse‐forager transition via a chemical communication system, the primer pheromone ethyl oleate (EO). EO is found at high concentrations on foragers in contrast to nurse bees and was shown to delay the onset of foraging. In this study, EO effects on colony behavior were not as robust as expected, and we found no direct correlation between EO treatment and synaptic maturation in the MB calyx. In general, we assume that the primer pheromone EO rather acts in concert with other factors influencing the onset of foraging with its effect being highly adaptive. © 2015 Wiley Periodicals, Inc. Develop Neurobiol 75: 1368–1384, 2015     

In-Hive Behavior of Pollen Foragers (Apis mellifera) in Honey Bee Colonies Under Conditions of High and Low Pollen Need

Pollen collection in honey bees is regulated around a homeostatic set-point. How the control of pollen collection is achieved is still unclear. Different feedback mechanisms have been proposed but little is known about the experience of pollen foragers in the hive. A detailed documentation of the behavior of pollen foragers in the hive under different pollen need conditions is presented here. Taking a broad observational approach, we analyze the behavior of individual pollen foragers in the hive between collecting trips and quantify the different variables constituting the in-hive stay. Comparing data from two colonies and 143 individuals during experimentally induced times of low vs. times of high pollen need, we show that individual foragers modulate their in-hive working tempo according to the actual pollen need of the colony: pollen foragers slowed down and stayed in the hive longer when pollen need was low and spent less time in the hive between foraging trips when pollen need by their colony was high. Furthermore, our data show a significant change in the trophallactic experience of pollen foragers with changing pollen need conditions of their colony. Pollen foragers had more short (< 3 s) trophallactic contacts when pollen need was high, resulting in an increase of total number of trophallactic contacts. Thus, our results support the hypothesis that trophallactic experience is one of the various information pathways used by pollen foragers to assess their colony's pollen need.     

Netted crop covers reduce honeybee foraging activity and colony strength in a mass flowering crop

The widespread use of protective covers in horticulture represents a novel landscape‐level change, presenting the challenges for crop pollination. Honeybees (Apis mellifera L) are pollinators of many crops, but their behavior can be affected by conditions under covers. To determine how netting crop covers can affect honeybee foraging dynamics, colony health, and pollination services, we assessed the performance of 52 nucleus honeybee colonies in five covered and six uncovered kiwifruit orchards. Colony strength was estimated pre‐ and postintroduction, and the foraging of individual bees (including pollen, nectar, and naïve foragers) was monitored in a subset of the hives fitted with RFID readers. Simultaneously, we evaluated pollination effectiveness by measuring flower visitation rates and the number of seeds produced after single honeybee visits. Honeybee colonies under cover exhibited both an acute loss of foragers and changes in the behavior of successful foragers. Under cover, bees were roughly three times less likely to return after their first trip outside the hive. Consequently, the number of adult bees in hives declined at a faster rate in these orchards, with colonies losing on average 1,057 ± 274 of their bees in under two weeks. Bees that did forage under cover completed fewer trips provisioning their colony, failing to reenter after a few short‐duration trips. These effects are likely to have implications for colony health and productivity. We also found that bee density (bees/thousand flowers) and visitation rates to flowers were lower under cover; however, we did not detect a resultant change in pollination. Our findings highlight the need for environment‐specific management techniques for pollinators. Improving honeybee orientation under covers and increasing our understanding of the effects of covers on bee nutrition and brood rearing should be primary objectives for maintaining colonies and potentially improving pollination in these systems.     

Lifetime- and caste-specific changes in flight metabolic rate and muscle biochemistry of honeybees, Apis mellifera

Honeybees, Apis mellifera, who show temporal polyethism, begin their adult life performing tasks inside the hive (hive bees) and then switch to foraging when they are about 2–3 weeks old (foragers). Usually hive tasks require little or no flying, whereas foraging involves flying for several hours a day and carrying heavy loads of nectar and pollen. Flight muscles are particularly plastic organs that can respond to use and disuse, and accordingly it would be expected that adjustments in flight muscle metabolism occur throughout a bee’s life. We thus investigated changes in lifetime flight metabolic rate and flight muscle biochemistry of differently aged hive bees and of foragers with varying foraging experience. Rapid increases in flight metabolic rates early in life coincided with a switch in troponin T isoforms and increases in flight muscle maximal activities (V _max) of the enzymes citrate synthase, cytochrome c oxidase, hexokinase, phosphofructokinase, and pyruvate kinase. However, further increases in flight metabolic rate in experienced foragers occurred without additional changes in the in vitro V _max of these flight muscle metabolic enzymes. Estimates of in vivo flux (v) compared to maximum flux of each enzyme in vitro (fractional velocity, v/V _max) suggest that most enzymes operate at a higher fraction of V _max in mature foragers compared to young hive bees. Our results indicate that honeybees develop most of their flight muscle metabolic machinery early in life. Any further increases in flight metabolism with age or foraging experience are most likely achieved by operating metabolic enzymes closer to their maximal flux capacity.