The effect of genotype on response thresholds to sucrose and foraging behavior of honey bees (Apis mellifera L.)

Honey bee foragers were tested for their proboscis extension response (PER) to water and varying solutions of sucrose. Returning pollen and nectar foragers were collected at the entrance of a colony and were assayed in the laboratory. Pollen foragers had a significantly higher probability of responding to water and to lower concentrations of sucrose. Bees derived from artificially selected high- and low-pollen-hoarding strains were also tested using the proboscis extension assay. Returning foragers were captured and tested for PERs to 30% sucrose. Results demonstrated a genotypic effect on PERs of returning foragers. The PERs of departing high- and low-strain foragers were consistent with those of returning foragers. The PERs were related to nectar and water reward perception of foragers. High strain bees were more likely to return with loads of water and lower concentrations of sucrose than foragers from the low pollen strain. Low-strain bees were more likely to return empty. We identified a previously mapped genomic region that contains a variable quantitative trait locus that appears to influence sucrose response thresholds. These studies demonstrate a gene-brain-behavior pathway that can be altered as a consequence of colony-level selection for quantities of stored food. Accepted: 3 September 1997     

Modulation of sucrose response thresholds in honey bees (Apis mellifera L.): influence of genotype, feeding, and foraging experience

The perception of sugar is important to honey bees for making foraging decisions. We measured bees' perception by determining what concentration of sucrose touched to the antennae elicited the proboscis extension response (response threshold). A low response threshold (extension at low concentration) suggests a high perceptual value of sucrose. and vice versa. Perception of sucrose solutions differed between two artificially selected genotypic strains and was modulated by the bees' recent feeding experiences. Bees offered 10%, 30%, or 50% sucrose solutions in small cages overnight, and in large flight-cages or free-flying in the field for several days, had subsequent response thresholds positively correlated to the concentration offered. Empty bees, whether they were nectar, water or pollen foragers, dancers or non-dancers, had a significantly lower threshold than loaded bees. Crop volume affected response thresholds directly and independently of sucrose concentration. We interpret these findings as multiple mechanisms that operate in different time scales, modulating perception of sucrose. Changes occurred in the time scale of evolutionary processes as demonstrated by genotypic differences. Changes with foraging experience occur in hours or minutes while effects of crop filling are instantaneous.     

The relationship between population size,amount of brood,and individual foraging behaviour in the honey bee,Apis mellifera L.

This study experimentally examines the relationship between colony state and the behaviour of individual pollen and nectar foragers in the honey bee, Apis mellifera L. In the first experiment we test the prediction that individual pollen foragers from colonies with higher brood quantities should exhibit a greater work effort for pollen resources than individual pollen foragers from colonies with low brood quantities. Eight colonies were assigned into two treatment groups; HIGH brood colonies were manipulated to contain 9600±480 cm² brood area; LOW brood colonies were manipulated to contain 1600±80 cm² brood area. We measured colony brood levels over the course of the experiment and collected individual pollen loads from returning pollen foragers. We found that, while colonies remained significantly different in brood levels, individual pollen foragers from HIGH brood colonies collected larger loads than individuals from LOW brood colonies. In the second experiment we investigated the influence of colony size on the behaviour of individual nectar foragers. We assigned eight colonies to two treatment groups; LARGE colonies were manipulated to contain 35000±1700 adult workers with 3500±175 cm² brood area, and SMALL colonies were manipulated to contain 10000±500 adult workers with 1000±50 cm² brood area. We observed foraging trips of individually marked workers and found that individuals from LARGE colonies made longer foraging trips than those from SMALL colonies (LARGE: 1666.7±126.4 seconds, SMALL: 1210.8±157.6 seconds), and collected larter nectar loads (LARGE: 19.2±1.0 l, SMALL: 14.6±0.8 l). These results indicate that individual nectar foragers from LARGE colonies tend to work harder than individuals from SMALL colonies. Both experiments indicate that the values of nectar and pollen resources to a colony change depend on colony state, and that individual foragers modify their behaviour accordingly.     

The accuracy of morphometric characteristic analysis depends on the type of the assessed traits of honey bees (Apis cerana F. and Apis mellifera L.)

Conservation of honey bees as pollinators and honey producers requires their assessment using convenient general methods. One of the most commonly used methods involves assessment of morphometric characteristics using random traits. The purpose of the study was to determine a set of morphometric characteristics for identifying the local (R, H: Apis cerana koreana) and adapted (A, C, F: Apis melifera ligustica; D: Apis mellifera carpathica; V: Apis mellifera) lines of honey bees bred in Korea. We found traits with significant differences (p < 0.001), high discriminant coefficient (>100), and range of variation (<50 %). The analysis demonstrated that the widths of the abdomen, forewing, and head, and the lengths of the antenna, body, proboscis, tergites 4 and 3, and forewing can be used as the nine main morphological characteristics for distinguishing the bred lines of honey bees. Furthermore, the last two traits and the length of the head can be used for distinguishing the adapted subspecies A. m. ligustica and A. m. carpathica. Our observations can be used for improving the morphometric method for the conservation of honey bees.     

The neonicotinoids thiacloprid,imidacloprid, and clothianidin affect the immunocompetence of honey bees (Apis mellifera L.)

A strong immune defense is vital for honey bee health and colony survival. This defense can be weakened by environmental factors that may render honey bees more vulnerable to parasites and pathogens. Honey bees are frequently exposed to neonicotinoid pesticides, which are being discussed as one of the stress factors that may lead to colony failure. We investigated the sublethal effects of the neonicotinoids thiacloprid, imidacloprid, and clothianidin on individual immunity, by studying three major aspects of immunocompetence in worker bees: total hemocyte number, encapsulation response, and antimicrobial activity of the hemolymph. In laboratory experiments, we found a strong impact of all three neonicotinoids. Thiacloprid (24 h oral exposure, 200 μg/l or 2000 μg/l) and imidacloprid (1 μg/l or 10 μg/l) reduced hemocyte density, encapsulation response, and antimicrobial activity even at field realistic concentrations. Clothianidin had an effect on these immune parameters only at higher than field realistic concentrations (50–200 μg/l). These results suggest that neonicotinoids affect the individual immunocompetence of honey bees, possibly leading to an impaired disease resistance capacity.     

Tactile learning and the individual evaluation of the reward in honey bees (Apis mellifera L.)

Using the proboscis extension response we conditioned pollen and nectar foragers of the honey bee (Apis mellifera L.) to tactile patterns under laboratory conditions. Pollen foragers demonstrated better acquisition, extinction, and reversal learning than nectar foragers. We tested whether the known differences in response thresholds to sucrose between pollen and nectar foragers could explain the observed differences in learning and found that nectar foragers with low response thresholds performed better during acquisition and extinction than ones with higher thresholds. Conditioning pollen and nectar foragers with similar response thresholds did not yield differences in their learning performance. These results suggest that differences in the learning performance of pollen and nectar foragers are a consequence of differences in their perception of sucrose. Furthermore, we analysed the effect which the perception of sucrose reward has on associative learning. Nectar foragers with uniform low response thresholds were conditioned using varying concentrations of sucrose. We found significant positive correlations between the concentrations of the sucrose rewards and the performance during acquisition and extinction. The results are summarised in a model which describes the relationships between learning performance, response threshold to sucrose, concentration of sucrose and the number of rewards. Accepted: 14 April 1999     

Effect of pheromones,hormones, and handling on sucrose response thresholds of honey bees (Apis mellifera L.)

The responsiveness of bees to sucrose is an important indicator of honey bee foraging decisions. Correlated with sucrose responsiveness is forage choice behavior, age of first foraging, and conditioned learning response. Pheromones and hormones are significant components in social insect systems associated with the regulation of colony-level and individual foraging behavior. Bees were treated to different exposure regimes of queen and brood pheromones and their sucrose responsiveness measured. Bees reared with queen or brood pheromone were less responsive than controls. Our results suggest responsiveness to sucrose is a physiologically, neuronally mediated response. Orally administered octopamine significantly reduced sucrose response thresholds. Change in response to octopamine was on a time scale of minutes. The greatest separation between octopamine treated and control bees occurred 30 min after feeding. There was no significant sucrose response difference to doses ranging from 0.2 g to 20 g of octopamine. Topically applied methoprene significantly increased sucrose responsiveness. Handling method significantly affected sucrose responsiveness. Bees that were anesthetized by chilling or CO₂ treatment were significantly more responsive than control bees 30 min after handling. Sixty minutes after handling there were no significant treatment differences. We concluded that putative stress effects of handling were blocked by anesthetic.Abbreviations BP brood pheromone - JH juvenile hormone - OA octopamine - PER proboscis extension response - PER-RT proboscis extension response threshold - QMP queen mandibular pheromone     

Antennal sucrose perception in the honey bee (Apis mellifera L.): behaviour and electrophysiology

Physiological mechanisms of antennal sucrose perception in the honey bee were analysed using behavioural and electrophysiological methods. Following sucrose stimulation of the tip of a freely moving antenna, the latency of proboscis extension was 320–340 ms, 80–100 ms after the first activity in muscle M17 controlling this response. When bees were allowed to actively touch a sucrose droplet with one antenna, contacts with the solution were frequent with durations of 10–20 ms and average intervals between contacts of approximately 40 ms. High sucrose concentrations led to short and frequent contacts. The proboscis response and M17 activity were largely independent of stimulus duration and temporal pattern. Taste hairs of the antennal tip displayed spike responses to sucrose concentrations down to at least 0.1%. The first 25 ms of the response were suitable for discrimination of sucrose concentrations. This time interval corresponds to the duration of naturally occurring gustatory stimuli. Sucrose responses between different hairs on the same antenna showed a high degree of variability, ranging from less than five to over 40 spikes per 0.5 s for a stimulus of 0.1% sucrose. This variability of receptor responses extends the dynamic range of sucrose perception over a large range of concentrations.     

Larval juvenile hormone treatment affects pre-adult development,but not adult age at onset of foraging in worker honey bees (Apis mellifera)

Previous research has shown that juvenile hormone (JH) titers increase as adult worker honey bees age and treatments with JH, JH analogs and JH mimics induce precocious foraging. Larvae from genotypes exhibiting faster adult behavioral development had significantly higher levels of juvenile hormone during the 2nd and 3rd larval instar. It is known that highly increased JH during this period causes the totipotent female larvae to differentiate into a queen. We treated third instar larvae with JH to test the hypothesis that this time period may be a developmental critical period for organizational effects of JH on brain and behavior also in the worker caste, such that JH treatment at a lower level than required to produce queens will speed adult behavioral development in workers. Larval JH treatment did not influence adult worker behavioral development. However, it made pre-adult development more queen-like in two ways: treated larvae were capped sooner by adult bees, and emerged from pupation earlier. These results suggest that some aspects of honey bee behavioral development may be relatively insensitive to pre-adult perturbation. These results also suggest JH titer may be connected to cues perceived by the adult bees indicating larval readiness for pupation resulting in adult bee cell capping behavior.     

The microRNA ame-miR-279a regulates sucrose responsiveness of forager honey bees (Apis mellifera)

Increasing evidence demonstrates that microRNAs (miRNA) play an important role in the regulation of animal behaviours. Honey bees (Apis mellifera) are eusocial insects, with honey bee workers displaying age-dependent behavioural maturation. Many different miRNAs have been implicated in the change of behaviours in honey bees and ame-miR-279a was previously shown to be more highly expressed in nurse bee heads than in those of foragers. However, it was not clear whether this difference in expression was associated with age or task performance. Here we show that ame-miR-279a shows significantly higher expression in the brains of nurse bees relative to forager bees regardless of their ages, and that ame-miR-279a is primarily localized in the Kenyon cells of the mushroom body in both foragers and nurses. Overexpression of ame-miR-279a attenuates the sucrose responsiveness of foragers, while its absence enhances their sucrose responsiveness. Lastly, we determined that ame-miR-279a directly target the mRNA of Mblk-1. These findings suggest that ame-miR-279a plays important roles in regulating honey bee division of labour.     

The relationship between comb age and performance of honey bee (Apis mellifera) colonies

A study on the relationship between the age of comb and the activity of the hybrid Carniolan honey bee colonies in collecting pollen activity, worker brood production, colony strength, and honey yield was conducted. In comparison to colonies with combs aged 4-years, colonies with combs aged 1, 2 and 3-years significantly exceeded in the number returning workers, number returning workers with pollen loads, rate of storing pollen, rate of worker brood production, and size of colony population. Colonies with combs aged 1, 2 and 3-years produced significantly more honey than colonies with combs aged 4-years (5.25, 4.90 and 4.65 kg/colony vs. 4.45 kg/colony, respectively). It can be concluded that the foraging rate, gathering and storing pollen, brood production, colony population size, and honey yield significantly depended on the age of combs. Beekeepers can replace old combs with new ones to increase brood and honey production.     

Calorimetric investigations of the different castes of honey bees,Apis mellifera carnica

Summary Honey bees of different age and castes were investigated calorimetrically at 20, 25 and 30 °C. Experiments were completed by endoscopic observation of the insects in the visible and the near infrared range and by acoustical monitoring and subsequent frequency analysis of various locomotor activities. Direct calorimetric results of this paper are compared with data of indirect calorimetry from the literature using a respiratory quotient of 1.00 and 21.13 J consumed. Agreements between both methods are generally good. The results show that weight-specific heat production rates increase with age of worker bees by a factor of 5.6 at 30 °C, 3.7 at 25 °C and 40.0 at 20 °C. In groups of foragers the heat production decreases with growing group size to around 6% of the value for an isolated bee. The presence of a fertile queen or of brood reduces the heat output of a small worker group significantly. Adult drones exhibit a much higher metabolic rate (up to 19.7-fold at 20 °C) than juveniles with strong fluctuations in the power-time curves. Fertile queens show a less pronounced heat production rate than virgin queens (54% at 30 °C, 87% at 25 °C and 77% at 20 °C). Calorimetric unrest is much higher for young than for adult queens. Heat production is very low in both uncapped and capped brood and less than 30% of that of a newly emerged worker. In most cases temperature showed a significant influence on the metabolic level, although its sign was not homogeneous between the castes or even within them. Locomotor activities are easily recorded by the acoustic frequency spectrum (0–7.5 kHz) and in good agreement with endoscopic observations and calorimetric traces.Abbreviations RQ respiratory quotient - ww wet weight This paper is part of the PhD thesis of L.F.     

Operant conditioning of antennal muscle activity in the honey bee (Apis mellifera L.)

Antennal movements of the honey bee can be conditioned operantly under laboratory conditions. Using this behavioural paradigm we have developed a preparation in which the activity of a single antennal muscle has been operantly conditioned. This muscle, the fast flagellum flexor muscle, is innervated by an identified motoneuron whose action potentials correlate 1:1 with the muscle potentials. The activity of the fast flagellum flexor muscle was recorded extracellularly from the scapus of the antenna. The animal was rewarded with a drop of sucrose solution whenever the muscle activity exceeded a defined reward threshold. The reward threshold was one standard deviation above the mean spontaneous frequency prior to conditioning. After ten conditioning trials, the frequency of the muscle potentials had increased significantly compared to the spontaneous frequency. The conditioned changes of frequency were observed for 30 min after conditioning. No significant changes of the frequency were found in the yoke control group. The firing pattern of the muscle potentials did not change significantly after conditioning or feeding. Fixing the antennal joints reduces or abolishes associative operant conditioning. The conditioned changes of the frequency of muscle potentials in the freely moving antenna are directly comparable to the behavioural changes during operant conditioning. Accepted: 29 March 2000     

Lipid-enhanced pollen and lipid-reduced flour diets and their effect on the longevity of honey bees (Apis mellifera L.)

Abstract  As eucalypt pollens contain low concentrations of lipid, enhancing pollen diets with fatty acids was hypothesised to improve honey bee longevity. Different concentrations of linoleic and oleic acid added to eucalypt pollen were trialled in small cages containing approximately 1400 bees each. Bees fed diets of redgum ( Corymbia calophylla (Lindl.) Hill & Johnson, formerly Eucalyptus calophylla ) pollen had the lowest mortality of 22 diets tested for 6 weeks and had life spans (50%) greater than 42 days. Linoleic acid mixed with a redgum diet in concentrations >6% corresponded to life spans of 24–25 days. Bee longevity appeared to be more sensitive to oleic acid as life spans decreased to 15–21 days when diets had concentrations >2%. The life spans of bees fed soya bean flour were 26 days on low (0.6% lipid) fat, 19 days on defatted and 20 days on full-fat diets. Bees fed lupin flour had a life span of 23 days. Adding redgum pollen to lupin flour caused increased mortality, but addition of pollen to soya bean flour was beneficial. Thus, beekeepers who choose to utilise soya bean or lupin flours as protein substitutes to pollen will have bees with reduced longevity. Bees fed redgum pollen that had been dried, crushed, irradiated and hermetically stored in a cool room for several years had similar longevity to bees fed fresh-collected and frozen redgum pollen.     

Antennal hygroreceptors of the honey bee,Apis mellifera L.

Summary Antennal hygroreceptors of the honey bee, Apis mellifera L., have been investigated electrophysiologically and the sensillum containing these receptors with SEM. Moist and dry hygroreceptors have been identified along with a thermal receptor in a specialized coeloconic sensillum. This sensillum comprises a cuticular, shallow depression (diameter; 4 ) having a central opening (1.4–1.5 m) and a mushroom-shaped protrusion (1.4–1.5 m) from the opening. The head of the protrusion is irregular in shape and is not perforated. This sensillum has been thus far referred to as a sensillum campaniformium (Dietz and Humphreys 1971), henceforth, it is referred to as a coelocapitular sensillum.The responses of both moist and dry hygroreceptors are of a phasic-tonic manner. Both receptors are antagonistic with respect to their responses to humidity; one responds with an increase in impulse frequency to rising humidity, the other to falling humidity. The humidity-response relationship is independent of stimulus flux.     

Sucrose response thresholds of honey bee (Apis mellifera) foragers are not modulated by brood ester pheromone

Division of labor is a hallmark of eusocial insects and their ecological success can be attributed to it. Honey bee division of labor proceeds along a stereotypical ontogenetic path based on age, modulated by various internal and external stimuli. Brood pheromone is a major social pheromone of the honey bee that has been shown to affect honey bee division of labor. It elicits several physiological and behavioral responses; notably, regulating the timing of the switch from performing in-hive tasks to the initiation of foraging. Additionally, brood pheromone affects future foraging choice. In honey bees, sucrose response threshold is a physiological correlate of age of first foraging and foraging choice. Brood pheromone has been shown to modulate sucrose response threshold in young bees, but its effects on sucrose response thresholds of bees in advanced behavioral states (foragers) are not known. In this study we examined the sucrose response thresholds of two different task groups, foragers (pollen and non-pollen) and non-foraging bees, in response to honey bee brood pheromone. Sucrose response thresholds were not significantly different between brood pheromone treatment and controls among both non-pollen and pollen foragers. However, the sucrose response threshold of non-foraging bees was significantly higher in the brood pheromone treatment group than in the control group. The switch to foraging task is considered a terminal one, with honey bee lifespan being determined at least partially by risks and stress accompanying foraging. Our results indicate that foragers are physiologically resistant to brood pheromone priming of sucrose response thresholds.     

Choice of flowers by foraging honey bees (Apis mellifera): possible morphological cues

Abstract.

• 1 Honey bees foraging for nectar on lavender (Lavandula stoechas) chose inflorescences with more of their flowers open. The number of open flowers predicted whether an inflorescence was visited by bees, inspected but rejected, or ignored. Inflorescences chosen arbitrarily by observers had numbers of open flowers intermediate between those of visited and ignored inflorescences.
• 2 Differences in morphological characters between types of inflorescence correlated with nectar volume and sugar weight per flower so that visited inflorescences had a disproportionately greater volume of nectar and weight of sugar per flower and greater variance in nectar volume.
• 3 Although there were significant associations between nectar content and the morphological characters of inflorescences, discriminant function analysis revealed discrimination on the basis of morphology rather than nectar content.
• 4 Visited inflorescences tended to have smaller than average flowers but bees tended to probe the largest flowers on visited inflorescences.
• 5 Choice of flowers within inflorescences is explicable in terms of the relationship between flower size and nectar content.

     

The interplay between dancing and trophallactic behavior in the honey bee Apis mellifera

The interplay between the recruitment dance and food-giving trophallactic contacts of returning Apis mellifera foragers was analyzed. Dancing and trophallactic events were recorded for bees returning from a rate feeder that provided 50% weight on weight sucrose solution at a constant flow rate of 5 μl min⁻¹. Bees that had danced immediately before their trophallactic contact had more recipients per trophallaxis compared with bees that did not dance before. Thus, besides information coded in dancing behavior, dance maneuvers could serve as a stimulus to increase attention of bees located on the dance floor to receive nectar. In addition, the number of bees receiving food during a trophallaxis showed a positive correlation with the probability of dancing immediately after contacting. The time from arrival at the hive to when the first or the subsequent contacts took place presented no correlation with the probability of dancing after trophallaxis. Also, the duration of a trophallaxis was positively correlated with the number of recipients per trophallaxis. These results suggest that returning foragers could receive information during a trophallactic contact with their hive mates that modify thresholds for dancing. Dance maneuvers and trophallactic contacts performed by foraging bees seem to be “mutually” affected. Accepted: 29 November 1999     

The coelocapitular sensillum,an antennal hygro- and thermoreceptive sensillum of the honey bee,Apis mellifera L.

Summary The sensillum coelocapitulum, a hygro- and thermoreceptive sensillum of the honey bee, Apis mellifera, was investigated by electron microscopy. The cuticular apparatus of the sensillum is a mushroomshaped protrusion, devoid of pores, set in a narrow cylindrical pit positioned centrally within a cuticular, shallow depression. There may be three or four receptor cells. Three receptor cells have unbranched sensory cilia, containing densely packed microtubules, which extend distally into the cuticular apparatus and completely fill its cavity. These connecting cilia are of the usual 9+0 type. The fourth receptor, if present, has a thin sensory cilium which terminates beneath the cuticular apparatus. Its connecting cilium has armed outer doublets. The outer cavity is formed by two enveloping cells and is completely sealed off. Lipid deposits are present within the cavity and the tormogen cell. The thecogen cell has scolopale rod-like structures around the inner cavity. Features common to the insect hygro- and thermoreceptive sensilla are discussed in comparison with those of other insects.     

Responsiveness to sucrose and habituation of the proboscis extension response in honey bees

In honey bees, complex behaviours such as associative learning correlate with responsiveness to sucrose. In these behaviours, the subjective evaluation of a sucrose stimulus influences the behavioural performance. Habituation is a well-known form of non-associative learning. In bees, the proboscis extension response can be habituated by repeatedly stimulating the antennae with a low sucrose concentration. A high sucrose concentration can dishabituate the response. This study tests whether habituation correlates with responsiveness to sucrose in bees of different behavioural states and in bees which are habituated with different sucrose concentrations. Habituation and dishabituation in newly emerged bees, 5-day-old bees and foragers strongly correlated with responsiveness to sucrose. Bees with high responsiveness to sucrose displayed a lower degree of habituation and showed greater dishabituation than bees with low responsiveness. The degree of habituation and dishabituation also depended on the concentration of the habituation stimulus. These experiments demonstrate for the first time in a non-associative learning paradigm that the subjective strength of a sucrose stimulus determines the behavioural performance. Non-associative learning shares this property with associative learning, which suggests that the two processes might rely on similar neural mechanisms.Abbreviations: GRS Gustatory response score - PER Proboscis extension response