Social Reinforcement Delays in Free-Flying Honey Bees (Apis mellifera L.)

Free-flying honey bees (Apis mellifera L.) reactions were observed when presented with varying schedules of post-reinforcement delays of 0 s, 300 s, or 600 s. We measured inter-visit-interval, response length, inter-response-time, and response rate. Honey bees exposed to these post-reinforcement delay intervals exhibit one of several patterns compared to groups not encountering delays, and had longer inter-visit-intervals. We observed no group differences in inter-response time. Honey bees with higher response rates tended to not finish the experiment. The removal of the delay intervals increased response rates for those subjects that completed the trials.     

Sex-Specific Differences in Pathogen Susceptibility in Honey Bees (Apis mellifera)

Sex-related differences in susceptibility to pathogens are a common phenomenon in animals. In the eusocial Hymenoptera the two female castes, workers and queens, are diploid and males are haploid. The haploid susceptibility hypothesis predicts that haploid males are more susceptible to pathogen infections compared to females. Here we test this hypothesis using adult male (drone) and female (worker) honey bees (Apis mellifera), inoculated with the gut endoparasite Nosema ceranae and/or black queen cell virus (BQCV). These pathogens were chosen due to previously reported synergistic interactions between Nosema apis and BQCV. Our data do not support synergistic interactions between N. ceranae and BQCV and also suggest that BQCV has limited effect on both drone and worker health, regardless of the infection level. However, the data clearly show that, despite lower levels of N. ceranae spores in drones than in workers, Nosema-infected drones had both a higher mortality and a lower body mass than non-infected drones, across all treatment groups, while the mortality and body mass of worker bees were largely unaffected by N. ceranae infection, suggesting that drones are more susceptible to this pathogen than workers. In conclusion, the data reveal considerable sex-specific differences in pathogen susceptibility in honey bees and highlight the importance of ultimate measures for determining susceptibility, such as mortality and body quality, rather than mere infection levels.     

Colonization history and population differentiation of the Honey Bees (Apis mellifera L.) in Puerto Rico

Honey bees (Apis mellifera L.) are the primary commercial pollinators across the world. The subspecies A. m. scutellata originated in Africa and was introduced to the Americas in 1956. For the last 60 years, it hybridized successfully with European subspecies, previous residents in the area. The result of this hybridization was called Africanized honey bee (AHB). AHB has spread since then, arriving to Puerto Rico (PR) in 1994. The honey bee population on the island acquired a mosaic of features from AHB or the European honey bee (EHB). AHB in Puerto Rico shows a major distinctive characteristic, docile behavior, and is called gentle Africanized honey bees (gAHB). We used 917 SNPs to examine the population structure, genetic differentiation, origin, and history of range expansion and colonization of gAHB in PR. We compared gAHB to populations that span the current distribution of A. mellifera worldwide. The gAHB population is shown to be a single population that differs genetically from the examined populations of AHB. Texas and PR groups are the closest genetically. Our results support the hypothesis that the Texas AHB population is the source of gAHB in Puerto Rico.     

Vibration Signal Modulates the Behavior of House-hunting Honey Bees (Apis mellifera)

Honey bees ( Apis mellifera ) in house-hunting swarms perform vibration signals (dorsoventral abdominal vibration (DVAV)) of 18.05 ± 0.45 Hz for 1.36 ± 0.23 s throughout the house selection process. These signals are performed by a specialized subset of bees, most of whom never perform recruitment dances to nest sites. Individuals repeatedly vibrate others. The patterns of vibration signal performance are consistent with the hypothesis that it serves to activate bees for take-off, but may also activate bees to scout for nest sites.     

Acaricide,Fungicide and Drug Interactions in Honey Bees (Apis mellifera)

Background

Chemical analysis shows that honey bees (Apis mellifera) and hive products contain many pesticides derived from various sources. The most abundant pesticides are acaricides applied by beekeepers to control Varroa destructor. Beekeepers also apply antimicrobial drugs to control bacterial and microsporidial diseases. Fungicides may enter the hive when applied to nearby flowering crops. Acaricides, antimicrobial drugs and fungicides are not highly toxic to bees alone, but in combination there is potential for heightened toxicity due to interactive effects.

Methodology/Principal Findings

Laboratory bioassays based on mortality rates in adult worker bees demonstrated interactive effects among acaricides, as well as between acaricides and antimicrobial drugs and between acaricides and fungicides. Toxicity of the acaricide tau-fluvalinate increased in combination with other acaricides and most other compounds tested (15 of 17) while amitraz toxicity was mostly unchanged (1 of 15). The sterol biosynthesis inhibiting (SBI) fungicide prochloraz elevated the toxicity of the acaricides tau-fluvalinate, coumaphos and fenpyroximate, likely through inhibition of detoxicative cytochrome P450 monooxygenase activity. Four other SBI fungicides increased the toxicity of tau-fluvalinate in a dose-dependent manner, although possible evidence of P450 induction was observed at the lowest fungicide doses. Non-transitive interactions between some acaricides were observed. Sublethal amitraz pre-treatment increased the toxicity of the three P450-detoxified acaricides, but amitraz toxicity was not changed by sublethal treatment with the same three acaricides. A two-fold change in the toxicity of tau-fluvalinate was observed between years, suggesting a possible change in the genetic composition of the bees tested.

Conclusions/Significance

Interactions with acaricides in honey bees are similar to drug interactions in other animals in that P450-mediated detoxication appears to play an important role. Evidence of non-transivity, year-to-year variation and induction of detoxication enzymes indicates that pesticide interactions in bees may be as complex as drug interactions in mammals.     

Tactile Conditioning And Movement Analysis Of Antennal Sampling Strategies In Honey Bees (Apis mellifera L.)

Honey bees (Apis mellifera L.) are eusocial insects and well known for their complex division of labor and associative learning capability^{1, 2}. The worker bees spend the first half of their life inside the dark hive, where they are nursing the larvae or building the regular hexagonal combs for food (e.g. pollen or nectar) and brood³. The antennae are extraordinary multisensory feelers and play a pivotal role in various tactile mediated tasks⁴, including hive building⁵ and pattern recognition⁶. Later in life, each single bee leaves the hive to forage for food. Then a bee has to learn to discriminate profitable food sources, memorize their location, and communicate it to its nest mates⁷. Bees use different floral signals like colors or odors^{7, 8}, but also tactile cues from the petal surface⁹ to form multisensory memories of the food source. Under laboratory conditions, bees can be trained in an appetitive learning paradigm to discriminate tactile object features, such as edges or grooves with their antennae^{10, 11, 12, 13}. This learning paradigm is closely related to the classical olfactory conditioning of the proboscis extension response (PER) in harnessed bees¹⁴. The advantage of the tactile learning paradigm in the laboratory is the possibility of combining behavioral experiments on learning with various physiological measurements, including the analysis of the antennal movement pattern.     

The Behavior of Honey Bees (Apis mellifera ligustica) during Queen Duels

Conflict is rare among the members of a highly cooperative society such as a honey bee colony. However, conflict within a colony increases drastically during colony reproduction ('swarming') when newly produced queens fight each other until only one queen remains in the nest. This study describes the behavior of queens and workers during naturally occurring queen combat. The duels of five pairs of queens were observed in three observation colonies. A typical duel is described qualitatively and the events of all five duels are described quantitatively. Several aspects of duels that are of particular interest are examined in detail, including the behavior of queens near capped queen cells, worker aggression toward queens, queen tooting, and the relation of queen and worker behavior to the outcome of the duel. The results of this investigation serve as a foundation for rigorous tests of hypotheses regarding the adaptive significance of queen and worker behavior during queen combat. The results presented suggest that: young queens patrol queen cells to kill rival queens while they are vulnerable; workers aggress queens to prevent them from destroying queen cells; queens toot to inhibit worker aggression; workers immobilize queens to make them easy targets for rival queens; and queens eject hind-gut contents to cause their rival to be immobilized by the workers.     

Caste-Specific Differences in Hindgut Microbial Communities of Honey Bees (Apis mellifera)

Host-symbiont dynamics are known to influence host phenotype, but their role in social behavior has yet to be investigated. Variation in life history across honey bee (Apis mellifera) castes may influence community composition of gut symbionts, which may in turn influence caste phenotypes. We investigated the relationship between host-symbiont dynamics and social behavior by characterizing the hindgut microbiome among distinct honey bee castes: queens, males and two types of workers, nurses and foragers. Despite a shared hive environment and mouth-to-mouth food transfer among nestmates, we detected separation among gut microbiomes of queens, workers, and males. Gut microbiomes of nurses and foragers were similar to previously characterized honey bee worker microbiomes and to each other, despite differences in diet, activity, and exposure to the external environment. Queen microbiomes were enriched for bacteria that may enhance metabolic conversion of energy from food to egg production. We propose that the two types of workers, which have the highest diversity of operational taxonomic units (OTUs) of bacteria, are central to the maintenance of the colony microbiome. Foragers may introduce new strains of bacteria to the colony from the environment and transfer them to nurses, who filter and distribute them to the rest of the colony. Our results support the idea that host-symbiont dynamics influence microbiome composition and, reciprocally, host social behavior.     

Brood Odor Discrimination Abilities in Hygienic Honey Bees (Apis mellifera L.) Using Proboscis Extension Reflex Conditioning

To understand the effect of abnormal brood odors on the initiation or control of hygienic behavior in honey bees, we employed the associative learning paradigm, proboscis extension reflex conditioning. Bees from two genetic lines(hygienic and non-hygienic) were able to discriminate between high concentrations of two floral odors equally well. Differential discrimination abilities were observed between the two lines when healthy and diseased brood odors were used, with the bees from the hygienic line discriminating between the pair of brood odors better than the non-hygienic bees. These results suggest that hygienic behavior in individual bees is associated with the bees' responses to olfactory stimuli emanating from diseased brood.     

A Causal Analysis of Observed Declines in Managed Honey Bees (Apis mellifera)

The European honey bee (Apis mellifera) is a highly valuable, semi-free-ranging managed agricultural species. While the number of managed hives has been increasing, declines in overwinter survival, and the onset of colony collapse disorder in 2006, precipitated a large amount of research on bees’ health in an effort to isolate the causative factors. A workshop was convened during which bee experts were introduced to a formal causal analysis approach to compare 39 candidate causes against specified criteria to evaluate their relationship to the reduced overwinter survivability observed since 2006 of commercial bees used in the California almond industry. Candidate causes were categorized as probable, possible, or unlikely; several candidate causes were categorized as indeterminate due to lack of information. Due to time limitations, a full causal analysis was not completed at the workshop. In this article, examples are provided to illustrate the process and provide preliminary findings, using three candidate causes. Varroa mites plus viruses were judged to be a “probable cause” of the reduced survival, while nutrient deficiency was judged to be a “possible cause.” Neonicotinoid pesticides were judged to be “unlikely” as the sole cause of this reduced survival, although they could possibly be a contributing factor.     

RNAi-mediated Double Gene Knockdown and Gustatory Perception Measurement in Honey Bees (Apis mellifera)

This video demonstrates novel techniques of RNA interference (RNAi) which downregulate two genes simultaneously in honey bees using double-stranded RNA (dsRNA) injections. It also presents a protocol of proboscis extension response (PER) assay for measuring gustatory perception.RNAi-mediated gene knockdown is an effective technique downregulating target gene expression. This technique is usually used for single gene manipulation, but it has limitations to detect interactions and joint effects between genes. In the first part of this video, we present two strategies to simultaneously knock down two genes (called double gene knockdown). We show both strategies are able to effectively suppress two genes, vitellogenin (vg) and ultraspiracle (usp), which are in a regulatory feedback loop. This double gene knockdown approach can be used to dissect interrelationships between genes and can be readily applied in different insect species.The second part of this video is a demonstration of proboscis extension response (PER) assay in honey bees after the treatment of double gene knockdown. The PER assay is a standard test for measuring gustatory perception in honey bees, which is a key predictor for how fast a honey bee''s behavioral maturation is. Greater gustatory perception of nest bees indicates increased behavioral development which is often associated with an earlier age at onset of foraging and foraging specialization in pollen. In addition, PER assay can be applied to identify metabolic states of satiation or hunger in honey bees. Finally, PER assay combined with pairing different odor stimuli for conditioning the bees is also widely used for learning and memory studies in honey bees.     

Field-Level Sublethal Effects of Approved Bee Hive Chemicals on Honey Bees (Apis mellifera L)

In a study replicated across two states and two years, we tested the sublethal effects on honey bees of the miticides Apistan (tau fluvalinate) and Check Mite+ (coumaphos) and the wood preservative copper naphthenate applied at label rates in field conditions. A continuous covariate, a colony Varroa mite index, helped us disambiguate the effects of the chemicals on bees while adjusting for a presumed benefit of controlling mites. Mite levels in colonies treated with Apistan or Check Mite+ were not different from levels in non-treated controls. Experimental chemicals significantly decreased 3-day brood survivorship and increased construction of queen supercedure cells compared to non-treated controls. Bees exposed to Check Mite+ as immatures had higher legacy mortality as adults relative to non-treated controls, whereas bees exposed to Apistan had improved legacy mortality relative to non-treated controls. Relative to non-treated controls, Check Mite+ increased adult emergence weight. Although there was a treatment effect on a test of associative learning, it was not possible to statistically separate the treatment means, but bees treated with Apistan performed comparatively well. And finally, there were no detected effects of bee hive chemical on colony bee population, amount of brood, amount of honey, foraging rate, time required for marked released bees to return to their nest, percentage of released bees that return to the nest, and colony Nosema spore loads. To our knowledge, this is the first study to examine sublethal effects of bee hive chemicals applied at label rates under field conditions while disambiguating the results from mite control benefits realized from the chemicals. Given the poor performance of the miticides at reducing mites and their inconsistent effects on the host, these results defend the use of bee health management practices that minimize use of exotic hive chemicals.     

Mating Frequencies of Honey Bee Queens (Apis mellifera L.) in a Population of Feral Colonies in the Northeastern United States

Across their introduced range in North America, populations of feral honey bee (Apis mellifera L.) colonies have supposedly declined in recent decades as a result of exotic parasites, most notably the ectoparasitic mite Varroa destructor. Nonetheless, recent studies have documented several wild populations of colonies that have persisted. The extreme polyandry of honey bee queens—and the increased intracolony genetic diversity it confers—has been attributed, in part, to improved disease resistance and may be a factor in the survival of these populations of feral colonies. We estimated the mating frequencies of queens in feral colonies in the Arnot Forest in New York State to determine if the level of polyandry of these queens is especially high and so might contribute to their survival success. We genotyped the worker offspring from 10 feral colonies in the Arnot Forest of upstate New York, as well as those from 20 managed colonies closest to this forest. We found no significant differences in mean mating frequency between the feral and managed queens, suggesting that queens in the remote, low-density population of colonies in the Arnot Forest are neither mate-limited nor adapted to mate at an especially high frequency. These findings support the hypothesis that the hyperpolyandry of honey bees has been shaped on an evolutionary timescale rather than on an ecological one.     

Drone and Worker Brood Microclimates Are Regulated Differentially in Honey Bees,Apis mellifera

Background

Honey bee (Apis mellifera) drones and workers show differences in morphology, physiology, and behavior. Because the functions of drones are more related to colony reproduction, and those of workers relate to both survival and reproduction, we hypothesize that the microclimate for worker brood is more precisely regulated than that of drone brood.

Methodology/Principal Findings

We assessed temperature and relative humidity (RH) inside honey bee colonies for both drone and worker brood throughout the three-stage development period, using digital HOBO^® Data Loggers. The major findings of this study are that 1) both drone and worker castes show the highest temperature for eggs, followed by larvae and then pupae; 2) temperature in drones are maintained at higher precision (smaller variance) in drone eggs and larvae, but at a lower precision in pupae than the corresponding stages of workers; 3) RH regulation showed higher variance in drone than workers across all brood stages; and 4) RH regulation seems largely due to regulation by workers, as the contribution from empty honey combs are much smaller compared to that from adult workers.

Conclusions/Significance

We conclude that honey bee colonies maintain both temperature and humidity actively; that the microclimate for sealed drone brood is less precisely regulated than worker brood; and that combs with honey contribute very little to the increase of RH in honey bee colonies. These findings increase our understanding of microclimate regulation in honey bees and may have implications for beekeeping practices.     

Nest Defense Behavior in Colonies from Crosses Between Africanized and European Honey Bees (Apis mellifera L.) (Hymenoptera: Apidae)

Honey bee (Apis mellifera L.) colonies with either European or Africanized queens mated to European or Africanized drones alone or in combination were tested for defensive behavior using a breath test. The most defensive colonies were those with European or Africanized queens mated to Africanized drones. In colonies where both European and Africanized patrilines existed, most of the workers participating in nest defense behavior for the first 30 s after a disturbance were of African patrilines. Nest defense behavior appears to be genetically dominant in honey bees.     

Urbanization Increases Pathogen Pressure on Feral and Managed Honey Bees

Given the role of infectious disease in global pollinator decline, there is a need to understand factors that shape pathogen susceptibility and transmission in bees. Here we ask how urbanization affects the immune response and pathogen load of feral and managed colonies of honey bees (Apis mellifera Linnaeus), the predominant economically important pollinator worldwide. Using quantitative real-time PCR, we measured expression of 4 immune genes and relative abundance of 10 honey bee pathogens. We also measured worker survival in a laboratory bioassay. We found that pathogen pressure on honey bees increased with urbanization and management, and the probability of worker survival declined 3-fold along our urbanization gradient. The effect of management on pathogens appears to be mediated by immunity, with feral bees expressing immune genes at nearly twice the levels of managed bees following an immune challenge. The effect of urbanization, however, was not linked with immunity; instead, urbanization may favor viability and transmission of some disease agents. Feral colonies, with lower disease burdens and stronger immune responses, may illuminate ways to improve honey bee management. The previously unexamined effects of urbanization on honey-bee disease are concerning, suggesting that urban areas may favor problematic diseases of pollinators.     

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

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