Genetic variance of mating frequency in the honeybee (Apis mellifera L.)

Summary. The genetic variance of queen mating frequency was studied in honeybees (Apis mellifera carnica). Worker offspring (N = 966) of 28 naturally mated half sister-queens (r = 0.25) from seven unrelated breeding lines were genotyped at four DNA microsatellites. The mating frequencies of the queens were derived from the offspring genotypes. The number of observed matings per queen ranged from 10 to 28 with an average of 17.32 ± 1.10 (number of estimated matings: 24.94 ± 2.51; number of effective matings: 20.09 ± 1.73). Half-sib analyses of the breeding lines were used to estimate heritability. Heritability was h² = 0.449 ± 0.135 for the estimated number of matings and h² = 0.262 ± 0.103 for the number of effective males, which are both significantly different from zero. We conclude that a high genetic variance for polyandry in honeybees can be favored by balanced selection between individual queen and colony level.Received 16 October 2003; revised 4 May 2004; accepted 4 May 2004.     

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

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

Foraging behaviour of honey bees (Apis mellifera L.) on transgenic oilseed rape (Brassica napus L.var. oleifera)

The impact of genetically modified oilseed rape (Brassica napus L.) on the foraging behaviour of honey bees (Apis mellifera L.) was evaluated on two different lines transformed to express constitutively heterologous chitinase in somatic tissue for enhanced disease resistance. Experiments were conducted in confinement in an indoor flight room with controlled conditions and in an outdoor flight cage with conditions more representative of the open environment. Foraging behaviour was analysed by observations of general bee behaviour (total number of visits) and of individual bee behaviour (using a video camera coupled with a special software program to process the data). The plants were analysed in terms of nectar quantity and quality (nectar volume and sugar content). The results showed no effects on bee foraging behaviour due to the modification of the genome of these plants by the introduction of a chitinase gene even though some differences between lines were found in the nectar. The methods applied in this original approach for the evaluation of the impact of genetically modified oilseed rape were shown to be sufficiently sensitive to detect changes in bee behaviour resulting from differences between plants.     

Adult honeybees (Apis mellifera L.) abandon hemocytic, but not phenoloxidase-based immunity

Hemocytes and the (prophenol-) phenoloxidase system constitute the immediate innate immune system in insects. These components of insect immunity are present at any post-embryonic life stage without previous infection. Differences between individuals and species in these immune parameters can reflect differences in infection risk, life expectancy, and biological function. In honeybees which show an age-related division of labor within the worker caste, previous studies demonstrated that foragers show a strongly reduced number of hemoctyes compared to the younger nurse bees. This loss of immune competence has been regarded advantageous with respect to an already high mortality rate due to foraging and to redistribution of energy costs at the colony level. Based on the idea that abandoning hemocytes in all adults would be a reasonably direct regulatory mechanism, we posed the hypothesis that abandoning hemocytic immunity is not restricted to worker honeybees. We tested our hypotheses by performing a comprehensive analysis of hemocyte number and phenoloxidase (PO)-activity levels in immunologically naive workers, queens, and drones. We found that in all three adult phenotypes hemocyte number is dramatically reduced in early adult life. In contrast, we found that the dynamics of PO-activity levels have sex and caste-specific characteristics. In workers, PO activity reached a plateau within the first week of adult life, and in queens enzyme levels continuously increased with age and reached levels twice as high as those found in workers. PO-activity levels slightly declined with age in drones. These data support our hypothesis, from which we infer that the previously reported reduction of hemocyte in foragers is not worker specific but represents a general phenomenon occurring in all honeybee adult phenotypes.     

Modelling the role of intracolonial genetic diversity on regulation of brood temperature in honey bee (Apis mellifera L.) colonies

In polyandrous social insects such as honey bees, a worker’s affinity for a particular task may be genetically infl uenced and so some patrilines may have lower stimulus thresholds for commencing a task than others. We used simulation models to investigate the effects of intracolonial diversity in the task thresholds that stimulate workers to engage in heating and cooling during nest thermoregulation. First, we simulated colonies comprised of one or 15 patrilines that were engaged in heating the brood nest, and observed that single patriline colonies maintained, on average, less stable brood nest temperatures than multiple patriline colonies. Second we simulated colonies with five patrilines that were engaged in cooling their nest, recording the proportions of bees of different patrilines that engaged in nest cooling in response to changing temperatures. Both of our simulations show remarkably similar qualitative patterns to those that we have previously observed empirically. This provides further support for the hypothesis that geneticallybased variability in task thresholds among patrilines within honey bee colonies is an important contributor to the ability of colonies to precisely thermoregulate their nests, and we suggest that diversity is important for optimal expression of a range of other colony-level phenotypes. Received 17 June 2005; revised 27 October 2005; accepted 23 December 2005.     

Super and half-sister discrimination by honey bee workers (Apis mellifera L.) in a trophallactic bioassay

Summary Kin recognition and nepotism between honeybee workers (Apis mellifera L.) was analysed in a trophallactic bio-assay. Donor workers were fed dyed sugar syrup and introduced into a recipient group consisting of 12 to 15 workers of the same colony. After allowing for 1 hour of trophallaxis, the distribution of the dyed food was analysed with spectrophotometry. The subfamily composition in the recipient group was varied such that the donor bees had to discriminate between workers of 2 to 7 different patrilines. Donor bees preferentially fed super sisters if few patrilines were present in the recipient group. However, preferential feeding was not observed if the recipient group consisted of workers of more than three subfamilies. Since the natural degree of polyandry causes intracolonial genetic variance to exceed the genetic variability in the experiments, nepotistic behaviour among workers may not reveal intranidal subfamily recognition in honeybees.     

The fine structure of the tarsal glands of the honeybee Apis mellifera L. (Hymenoptera)

Summary Tarsal glands are located in the 6th tarsomere of adult honeybee queens, workers and drones. Their structural features are not cast or sex specific. The glandular epithelium is lined by a thin endocuticular layer. A cuticular pocket is formed from a postimaginal delamination of the cuticle secreted by the glandular epithelium. The apical plasma membrane of the glandular cells shows numerous cristae and microvilli lining large crypts that communicate with the subcuticular space. Pinocytotic vesicles, multivesicular bodies and residual dense bodies are present in the apical part of the glandular cells. The RER is well developed in perinuclear and basal parts of the glandular cells, but the Golgi apparatus is a discrete organelle without secretory granules. No exocytotic secretory structures were observed. To reach the glandular pocket, the non-proteinaceous secretory product must pass across the subcuticular space, the cuticular intima, the space between the intima and the cuticular wall, and the cuticular wall of the glandular pocket.     

Variation in worker response to honey bee (Apis mellifera L.) queen mandibular pheromone (Hymenoptera: Apidae)

Genetic and environmental influences on the worker honey bee retinue response to queen mandibular gland pheromone (QMP) were investigated. Worker progeny were reared from queens originating from four sources: Australia, New Zealand, and two locations in British Columbia, Canada (Simon Fraser University and Vancouver Island). Progeny from New Zealand queens responded significantly higher (P < 0.05) than progeny from Australia in a QMP retinue bioassay. Retinue response was not related to queen production of pheromone or colony environment, and the strain-dependent differences in retinue bioassay responses were maintained over a wide range of dosages. Selected high- and low-responding colonies were bioassayed over the course of 1 year. High-responding colonies contacted QMP lures more frequently than low-responding colonies (P < 0.05) throughout the year except in late summer. We conclude that there is a strong genetic component to QMP response by worker honey bees, as well as a seasonal effect on response.     

Queen attendance behavior of worker honey bees (Apis mellifera L.) that are high and low responding to queen mandibular pheromone

Summary Queen attendance behavior of workers from selected honey bee colonies with high and low worker retinue response to synthetic queen mandibular gland pheromone (QMP) was investigated. Antennating, licking, grooming, and feeding of the queen by workers from high and low responding colonies were examined. High and low QMP responding workers did not attend the queen differently. However, workers originating from different colonies antennated and licked the queen more frequently than others, suggesting there may be a genetic basis for queen attendance behavior not necessarily associated with response to QMP. The median age of queen attendance was independent of strain.     

Spatial memory, navigation and dance behaviour in Apis mellifera

Navigation and dance communication in Apis mellifera have been extensively studied on the level of sensory processing, but the structure and content of the spatial memory underlying such phenomena have yet to be addressed. Here we survey new findings indicating that the memory used by bees to navigate within the range of their orientation flights is much more complex than hitherto thought. It appears to allow them to decide between at least two goals in the field, and to steer towards them over considerable distances. Two models concerning the structure of bees’ spatial memory are developed from new empirical evidence. The first one relies on the integration of at least two flight vectors, while the second assumes the existence of a ‘functional’ map based on the information available on-site. These findings also raise questions about the process of encoding and decoding information in the context of the waggle dance. We review published data and recent evidence indicating that memories of topographical features might also be involved in dance communication, and point out what needs to be addressed to elucidate the corresponding memory demands. The flight paths of recruited bees can now be traced by means of radar techniques, and thus tools are available to tackle these questions.     

Estimation of intracolonial worker relationship in a honey bee colony (Apis mellifera L.) using DNA fingerprinting

Summary The number and frequencies of subfamilies in a honey bee colony were determined by DNA fingerprinting. Queen and brood samples were taken from three colonies with artificially inseminated queens and from one colony with a naturally mated queen. UsingHae III restriction enzyme and (GATA)₄ oligonucleotide, the number of subfamilies in the colonies with artificially inseminated queens corresponded with the number of drones used for insemination. In the colony with the naturally mated queen, 12 subfamilies were found in a random sample of 104 workers. Considering that subfamily frequencies range from 1 to 26%, introcolonial worker relationship was estimated to be 0.328, corresponding to a genetical effective number of 6.4 matings.     

Flower choice by honey bees (Apis mellifera L.): sex-phase of flowers and preferences among nectar and pollen foragers

Bees foraging for nectar should choose different inflorescences from those foraging for both pollen and nectar, if inflorescences consist of differing proportions of male and female flowers, particularly if the sex phases of the flowers differ in nectar content as well as the occurrence of pollen. This study tested this prediction using worker honey bees (Apis mellifera L.) foraging on inflorescences of Lavandula stoechas. Female flowers contained about twice the volume of nectar of male flowers. As one would predict, bees foraging for nectar only chose inflorescences with disproportionately more female flowers: time spent on the inflorescence was correlated with the number of female flowers, but not with the number of male flowers. Inflorescence size was inversely correlated with the number of female flowers, and could be used as a morphological cue by these bees. Also as predicted, workers foraging for both pollen and nectar chose inflorescences with relatively greater numbers of both male and female flowers: time spent on these inflorescences was correlated with the number of male flowers, but not with the number of females flowers. A morphological cue inversely associated with such inflorescences is the size of the bract display. Choice of flowers within inflorescences was also influenced predictably, but preferences appeared to be based upon corolla size rather than directly on sex phase.     

Nest-mate recognition template of guard honeybees (Apis mellifera) is modified by wax comb transfer

In recognition, discriminators use sensory information to make decisions. For example, honeybee (Apis mellifera) entrance guards discriminate between nest-mates and intruders by comparing their odours with a template of the colony odour. Comb wax plays a major role in honeybee recognition. We measured the rejection rates of nest-mate and non-nest-mate worker bees by entrance guards before and after a unidirectional transfer of wax comb from a 'comb donor' hive to a 'comb receiver' hive. Our results showed a significant effect that occurred in one direction. Guards in the comb receiver hive became more accepting of non-nest-mates from the comb donor hive (rejection decreased from 70 to 47%); however, guards in the comb donor hive did not become more accepting of bees from the comb receiver hive. These data strongly support the hypothesis that the transfer of wax comb increases the acceptance of non-nest-mates not by changing the odour of the bees, but by changing the template used by guards.     

Experimental infection of Apis mellifera honeybees with Nosema ceranae (Microsporidia)

In this report, an experimental infection of Apis mellifera by Nosema ceranae, a newly reported microsporidian in this host is described. Nosema free honeybees were inoculated with 125,000 N. ceranae spores, isolated from heavily infected bees. The parasite species was identified by amplification and sequencing the SSUrRNA gene of the administered spores. Three replicate cages of 20 honeybees each were prepared, along with one control cage (n=20) supplied with sugar syrup only. The infection rate was 100% at the dosage administered. The presence of Nosema inside ventricular cells was confirmed in the samples using ultrathin sectioning and transmission electron microscopy. By day 3 p.i. a few cells (4.4%+/-1.2) were observed to be parasitized, whereas by 6 days p.i. more than half of the counted cells (66.4%+/-6) showed different parasite stages, this value increasing on day 7 p.i. (81.5%+/-14.8). Only one control bee died on day 7 p.i. In the infected groups, mortality was not observed until day 6 p.i. (66.7%+/-5.6). Total mortality on day 7 p.i. was 94.1% in the three infected replicates and by day 8 p.i. no infected bee was alive. After the infection, the parasites invaded both the tip of folds and the basal cells of the epithelium and the autoinfective capacity of the spores seemed to spread the infection rapidly between epithelial cells. On day 3 p.i., mature spores could be seen inside host cell tissue implying that the developmental cycle had been completed. The large number of parasitized cells, even the regenerative ones, the presence of autoinfective spores and the high mortality rate demonstrate that N. ceranae is highly pathogenic to Apis mellifera. Possible relation with bee depopulation syndrome is discussed by authors.     

Influence of carbon dioxide on Nosema apis infection of honeybees (Apis mellifera)

Young workers of the honeybee Apis mellifera carnica were individually inoculated with Nosema apis spores subjected to carbon dioxide (CO(2)) treatment. The spores were kept in a CO(2) atmosphere for 30, 35 and 40 h. The course of the infection was evaluated on the basis of the survival rate of bee workers and the number of N. apis spores in their digestive tracts. CO(2) treatment of N. apis spores resulted in faster proliferation of the parasite as well as higher mortality among workers infected with spores kept in CO(2) for 30 and 35 h.     

Kin recognition of worker brood by worker honey bees,Apis mellifera L.

Experimental colonies of honey bees consisting of two patrilines were observed as they reared worker brood. Seven behavior patterns that relate to brood care were recorded. Worker bees biased the care they provided to eggs and larvae destined to become workers on the basis of brood patrilines. Both patrilineal and antipatrilineal preferences in various behavioral patterns were observed. There was variation among colonies that may have been the result of the frequencies of brood of each patriline and the total amount of brood available to be reared. In addition, there were some differences between workers of the two patrilines in the way that they cared for the two patrilines of brood.     

Mitochondrial DNA variability in the Canary Islands honeybees (Apis mellifera L.)

The mitochondrial DNA (mtDNA) of individuals from 79 colonies of Apis mellifera from five Canary Islands was studied using the Dra I test based on the restriction of PCR products of the tRNAleu–COII intergenic region. Five haplotypes of the African (A) lineage and one of the west European (C) lineage were found. The haplotypes A14 and A15 are described for the first time. These haplotypes have a new P sequence named P₁. The wide distribution and high frequency of haplotype A15 suggest that it is characteristic of the Canarian Archipelago. Sources of haplotype variability of honeybee mtDNA in the Canary Islands (waves of colonization from Africa, queen importations, habitat diversification) are discussed.     

Number of reproductive cycles of Varroa jacobsoni in honey-bee (Apis mellifera) colonies

Very little data exists concerning the number of reproductive cycles performed by individual Varroa mites. To understand the population dynamics of the Varroa mite it is necessary to know the number of fertile female offspring each Varroa female produces during her lifetime. The lifetime reproduction capacity of the mite consists of the mean number of fertile female offspring produced during each reproductive cycle multiplied by the mean number of cell passages. This paper describes an experimental design to estimate the number of reproductive cycles where mites are transferred to new mite-free colonies for reproduction in sealed brood cells. The data presented suggests that the mean number of reproductive cycles performed by the individual female mite is larger than previously accepted. Under optimal conditions, the mean number of reproductive cycles by Varroa females is probably greater than 1.5 but less than 2. Furthermore, the results show that the reproductive success of Varroa females going into cells to reproduce is not influenced by previous brood cycles.