Muscle potentials and temperature acclimation and acclimatization in flight muscles of workers and drones of Apis mellifera

1. 1.Muscle potentials in fibrillar flight muscles of worker and drone honeybees were recorded extracellularly at thoracic temperatures from 30 to 10°C.

2. 2.Extinction temperatures for muscle potentials were higher in drones for all treatments.

3. 3.Cold acclimation (15°C) lowered extinction temperatures significantly in workers and drones. Acclimitization changed extinction temperatures significantly only in drones.

4. 4.Cold acclimitization had a bigger effect on the rate of muscle potential amplitude decline with decreasing temperature than acclimation.

5. 5.Acclimation and acclimitization had no effect on the increase of muscle potential duration with falling temperature.

6. 6.Muscle potential frequency during shivering was not much different between cold and warm treated bees.

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     

Juvenile hormone and circadian locomotor activity in the honey bee Apis mellifera

Age-related division of labor in honeybees is associated with plasticity in circadian rhythms. Young nest bees care for brood around the clock with no circadian rhythms while older foragers have strong circadian rhythms that are used for sun compass navigation and for timing visits to flowers. Since juvenile hormone (JH) is involved in the coordination of physiological and behavioral processes underlying age-related division of labor in honey bees, we tested the hypothesis that JH influences the ontogeny of circadian rhythms and other clock parameters in young worker bees. Treatments with the JH analog methoprene or allatectomy did not influence the onset of rhythmicity, overall locomotor activity, or the free-running period of rhythmic locomotor behavior. There were, however, significant differences in the onset of rhythmicity, overall locomotor activity, and longevity between bees from different source colonies, suggesting that there is significant genetic variation for these traits. Our results suggest that JH does not coordinate all aspects of division of labor in bees and that coordination of task performance with circadian rhythms is probably mediated by other regulatory systems.     

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.     

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 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 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.     

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     

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.     

Honey bee (Apis mellifera jemenitica) colony performance and queen fecundity in response to different nutritional practices

Honey bee colony nutritional dynamics depend on the availability of floral resources throughout a countryside with varying forage circumstances. Few studies quantify the queen fecundity and colony performance about certain management approaches on a broad scale. The present study was conducted to investigate the queen bee fecundity and various colony performance parameters in response to different nutritional practices, i.e., Group-I, supplied with sucrose solution (1:1; w/v), Group-II, provided with locally available commercial pollen substitute, Group-III, supplied with both sucrose solution + locally available commercial pollen substitute, and Group-IV without any sugar solution and pollen substitute. Our results demonstrated that eggs laid by queen bees were significantly higher (1241.83 ± 46.24) in Group-III than in other groups over the time of observations. Similarly, a significant difference was noticed in the mean sealed worker brood area and honey store area between the different groups of management practices. Both, the max mean sealed worker brood area (2153.53 ± 29.18 cm²) and max mean honey store area (1713.33 ± 12.06 cm²) were observed in Group-III while, the mini mean sealed worker brood area (1066.53 ± 20.18 cm²) and mini mean honey store area (1058.86 ± 4.07 cm²) were observed in Group-IV. In contrast, a non-significant difference was observed in pollen stores between Group-II and Group-III (p > 0.005). Current findings add to our understanding of the mechanisms that underpin large-scale controlled colony performance when the natural pollens resources are insufficient.     

The effect of temperature on candidate gene expression in the brain of honey bee Apis mellifera (Hymenoptera: Apidae) workers exposed to neonicotinoid imidacloprid

Neonicotinoid insecticides are potent agonists of nicotinic acetylcholine receptors and are a major factor in the decline of pollinators worldwide. Several studies show that low doses of this neurotoxin influence honey bee physiology, however, little is known about how insecticides interact with other environmental variables. We studied the effects of two neonicotinoid Imidacloprid doses (IMD, 0, 2.5, and 10 ppb), and three temperatures (20, 28, and 36°C) on gene expression in the brains of worker honey bees (Apis mellifera). Using qRT-PCR we quantified the expression of eight key genes related to the nervous system, stress response, and motor and olfactory capacities. Gene expression tended to increase with the low IMD dose, which was further intensified in individuals maintained in the cold treatment (20°C). At 20°C the octopamine receptor gene (oa1) was underexpressed in bees that were not exposed to IMD, but overexpressed in individuals exposed to 2.5 ppb IMD. Also, heat shock proteins (hsp70 and hsp90) increased their expression at high temperatures (36°C), but not with IMD doses. These results suggest that despite the low insecticide concentrations used in this study (a field-realistic dose), changes in gene expression associated with honey bee physiological responses could be induced. This study contributes to the understanding of how neonicotinoid residual doses may alter honey bee physiology.     

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     

欧洲黑蜂和新疆黑蜂越冬期抗氧化酶活性及相关基因表达分析

以欧洲黑蜂和新疆黑蜂为研究对象,比较和分析两个蜂种在越冬条件下抗氧化酶活性和抗氧化能力相关基因表达的变化规律,探究二者在越冬期的抗氧化酶功能及其基因表达特征。本试验测定了蜂群损失率和饲料消耗量等越冬性能指标以及总抗氧化能力（T-AOC）、超氧化物歧化酶（T-SOD）和过氧化氢酶（CAT）等抗氧化酶的酶活力值,并采用实时荧光定量PCR方法检测SOD、CAT、Grx1、MsrA和Vg基因等抗氧化酶功能基因的表达情况。结果显示,欧洲黑蜂和新疆黑蜂的蜂群损失率和饲料消耗率分别为32.86%、32.91%和23.41%、23.91%。在抗氧化酶活性变化规律方面,两个蜂种的T-AOC酶活力呈逐步升高趋势;T-SOD酶活力呈先下降再升高的趋势,在越冬中期达到最低值;CAT酶活力呈先升高再降低又再升高的趋势;在抗氧化功能相关基因的mRNA表达方面,两个蜂种的SOD基因和CAT基因的表达在越冬中期达到最低值;Grx1基因的表达在越冬后期急剧升高,达到峰值;MsrA基因的表达呈先降低后升高的趋势;Vg基因的表达在越冬的中前期呈逐渐升高趋势。结果表明,在低温逆境条件下,两个蜂种体内的抗氧化酶系统可能发挥了重要的防御作用,存在一定的耐寒适应机制,其中越冬中期可能是酶活性变化和基因表达的发生显著变化的特殊阶段。本试验的研究为揭示蜜蜂体内抵御低温应答机制研究提供新线索,也为深入研究蜜蜂耐寒分子机制和抗寒蜂种的选育提供基础数据。     

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.     

Monthly changes in various drone characteristics of Apis mellifera ligustica and Apis mellifera syriaca

This study was conducted to investigate drone rearing activity and semen production of Apis mellifera ligustica and Apis mellifera syriaca . Tendency of worker bees of both subspecies towards egg laying under semiarid conditions were also monitored in the experiments. Differences were not observed in drone brood production between both honeybee subspecies throughout the investigation. Worker bees of both subspecies needed a significantly shorter time to start egg laying during February and March in comparison with the time those workers needed for laying eggs during the remaining months of the study. Syrian bee workers started egg laying earlier than Italian bee workers. Drones from laying workers were much smaller and produced less sperms with more abnormalities than normal drones. Drones produced from queens in May were heavier and produced more sperms with less abnormalities than those produced during the other months. The drone brood rearing of both subspecies tended to follow the same general cycle in 2005 and 2006. The study suggests that virgin queens have a better chance to receive adequate viable sperm amounts from drones in April and May in semiarid Mediterranean conditions.     

High-abundance mRNAs in Apis mellifera: comparison between nurses and foragers

A large volume of honey bee (Apis mellifera) tag-seq was obtained to identify differential gene expression via Solexa/lllumina Digital Gene Expression tag profiling (DGE) based on next generation sequencing. In total, 4,286,250 (foragers) and 3,422,327 (nurses) clean tags were sequenced, 24,568 (foragers) and 13,134 (nurses) distinct clean tags could not be match to the reference database, and 7508 and 6875 mapped genes were detected in foragers and nurses respectively. 7045 genes were found differentially expressed between foragers and nurses. Of those genes, 1621genes had significantly different expression, that is, they showed an expression ratio (foragers/nurses) of more than 2 and FDR (False Discovery Rate) of less than 0.001. We identified 101 genes that were uniquely expressed in foragers, and 9 genes that were only expressed in nurses. We performed the Gene Ontology (GO) category and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis, and found 415 genes with annotation terms linked to the GO cellular component category. 200 components of KEGG pathways were obtained, including 21 signaling pathways. The PPAR signaling pathway was the most highly enriched, with the lowest Q-value.     

Presence of chitinase in adult Varroa destructor,an ectoparasitic mite of Apis mellifera

The enzyme spectrum of an ectoparasitic mite of the honeybee,Varroa destructor (Anderson and Trueman) was studied usinga semi-quantitative method, especially designed for complex samples which havenot been purified. Exopeptidases and phosphatases are shown present. Achitinaseand enzymes able to transform carbohydrates are also present with a largerange in the intensity of the reaction. The role of the chitinase can berelatedto the supply of nutritional needs or/and the piercing and sucking behaviour ofthe adult parasite. Chitinase activity could be one factor influencing thebalance between the parasite and its host.     

The ontogeny of immunity in the honey bee, Apis mellifera L. following an immune challenge

The honey bee, Apis mellifera, is an ideal system for investigating ontogenetic changes in the immune system, because it combines holometabolous development within a eusocial caste system. As adults, male and female bees are subject to differing selective pressures: worker bees (females) exhibit temporal polyethism, while the male drones invest in mating. They are further influenced by changes in the threat of pathogen infection at different life stages. We investigated the immune response of workers and drones at all developmental phases, from larvae through to late stage adults, assaying both a constitutive (phenoloxidase, PO activity) and induced (antimicrobial peptide, AMP) immune response. We found that larval bees have low levels of PO activity. Adult workers produced stronger immune responses than drones, and a greater plasticity in immune investment. Immune challenge resulted in lower levels of PO activity in adult workers, which may be due to the rapid utilisation and a subsequent failure to replenish the constitutive phenoloxidase. Both adult workers and drones responded to an immune challenge by producing higher titres of AMPs, suggesting that the cost of this response prohibits its constant maintenance. Both castes showed signs of senescence in immune investment in the AMP response. Different sexes and life stages therefore alter their immune system management based on the combined factors of disease risk and life history.     

Nutritional effects of supplementary diets on brood development,biological activities and honey production of Apis mellifera L

The present research work was conducted to assess the impact of nutrient-enriched diet on the physiological activities and subsequently honey yield. Eighteen colonies of Apis mellifera L. were selected from Dera Ismail Khan region, KPK, Pakistan, during the winter and summer seasons, 2019–2020. Five pollen supplement diets were prepared and provided to screen out the palatable one to be fed as pollen alternative nutrition to bee bread. Results of diet consumption regarding mean data for consumption rate displayed that soybean flour enriched artificial diet was maximally consumed (74.34 g) by honey bees per week. Minimum consumption was observed for grinded groundnut enriched diet (64.62 g) which was relatively lesser than the other tested artificial diets. Results of area of worker brood disclosed that soybean flour fortified diet (1489.27 cm²/colony) statistically noteworthy than the other artificial diets whereas control (463.51 cm²/colony) was least effective. Highest bee strength (10.00 bee frames/colony) was noted in the bee colonies fed with soybean flour fortified diet, date paste (8.0 bee frames/colony) was the next effective one, among the tested pollen replacement diets whereas relatively least (5 bee frames/colony) was noticed in case of grinded groundnut. Highest body weight (12.41 g) of neonate bees was noted in case of soybean enriched diet while lowermost (5.31 g) was noted in control bees. Results of wax cell built up and foraging efficiency were also superior in artificial diets than respective control bees. Hence, artificial diets especially soybean-enriched pollen alternative diet can boost up the physiology of honey bee leading to increased honey yield and profit.     

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

Bees derived from artificially selected high- and low-pollen-hoarding strains were tested for their proboscis extension reflex response to water and varying sucrose concentrations. High-strain bees had a lower response threshold to sucrose than low-strain bees among pre-foragers, foragers, queens and drones. Pre-foraging low-strain workers showed ontogenetic changes in their response threshold to sucrose which was inversely related to age. High-strain foragers were more likely to return with loads of water compared to low-strain foragers. Whereas low-strain foragers were more likely to return with loads of nectar. Low-strain nectar foragers collected nectar with significantly higher sucrose concentrations than did the high-strain nectar foragers. Alternatively, low-strain foragers were more likely to return empty compared to high-strain foragers. These studies demonstrate how a genotypically varied sensory-physiological process, the perception of sucrose, are associated with a division of labor for foraging. Accepted: 27 October 1998