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Joint actions of deltamethrin and azole fungicides on honey bee thermoregulation.

The effects of sublethal doses of deltamethrin, a pyrethroid insecticide, and prochloraz and difenoconazole, two azole fungicides, on honey bee thermoregulation were investigated by infrared thermography of honey bees kept at 22 degrees C. Deltamethrin at doses of 0.5 and 1.5 ng/bee did not elicit any significant effect on bee thermogenesis whereas doses of 2.5 and 4.5 ng/bee caused a severe hypothermia. Similarly, prochloraz and difenoconazole did not elicit any significant effect on thermogenesis at doses of up to 850 ng/bee whereas they triggered hypothermia at 1250 ng/bee. When associated with prochloraz or difenoconazole at 850 ng/bee, deltamethrin elicited a joint hypothermia at doses that did not induce a significant effect on thermoregulation when used alone.

Animals↗

Modulatory roles of the adenosine triphosphate P2x-purinoceptor in generation of the persistent nociception induced by subcutaneous bee venom injection in the conscious rat.

To study the role of adenosine triphosphate (ATP) P2x-purinoceptor in the persistent nociceptive response induced by subcutaneous (s.c.) bee venom injection, we used a selective P2x receptor antagonist, pyridoxal-phosphate-6-azophenyl-2',4'-disulfonic acid (PPADS), to evaluate whether spinal P2x receptor play a role in development of spontaneous persistent pain. Injection s.c. of bee venom into the plantar surface of one hindpaw in the conscious rat produces a monophasic, prolonged persistent nociception characterized by continuously flinching reflex of the injected paw for 1-2 h. Intrathecal (i.t.) pretreatment with PPADS at two lower doses of 5 and 10 microg resulted in suppression of the flinching reflex in a dose dependent manner with the inhibitory rate 37 and 44%, respectively, when compared with the control group; whereas i.t. PPADS at a higher dose of 30 microg failed to produce any inhibitory effect. This result suggests that activation of P2x-purinoceptor in the spinal cord contributes to the induction of bee venom-induced prolonged persistent pain. However, the antinociceptive effect of ATP P2x-purinoceptor antagonist such as PPADS on clinical pathological pain seems to be limited due to its lack of effectiveness at higher dose.

Adenosine Triphosphate↗

Arginine kinase is highly expressed in the compound eye of the honey bee, Apis mellifera.

We have cloned and sequenced a 1.68-kb cDNA encoding arginine kinase in the honey bee, Apis mellifera. The predicted protein shows a high level of identity to known arginine kinases in invertebrates and to other proteins belonging to the conserved family of ATP: guanidino phospho-transferases. The pattern of expression of arginine kinase has been investigated for the first time in various tissues including the brain, antennae and compound eye. Our results show that three isoforms of arginine kinase, transcribed from a single gene, are expressed in a characteristic pattern in major tissues of the honey bee. Arginine kinase mRNA is relatively abundant in the central nervous system and in the antennae. However, the highest level of expression, that is at least two to three times higher than in the brain, is found in the compound eye of the bee. By contrast, the levels of mRNAs encoding another metabolically important enzyme, alpha-glycerolphosphate dehydrogenase (alpha-GPDH), are low in the eye. These findings suggest that arginine kinase is an important component of the energy releasing mechanism in the visual system that has high and fluctuating energy demands. Furthermore, our results support the role of phosphagen kinases in energy transport in polarised cells and are consistent with the role of arginine kinase as an energy shuttle that delivers ATP generated by mitochondria to high energy-requiring processes, such as massive membrane turnover and pigment regeneration in the retina.

Amino Acid Sequence↗

Honey bee mortality due to tracheal mite parasitism.

We demonstrate, by truncating the expected negative binomial distribution, that the tracheal mite parasite, Acarapis woodi, causes mortality in the European honey bee, Apis mellifera, but, that this mortality can be documented only during periods of low mite densities (mites per bee). At high mite densities, this technique no longer reveals mite-induced mortality. We suggest that this paradox results from a reduction in the mortality threshold at high mite densities, concealing mortality from our truncation procedures. Longitudinal comparisons of hive and forager bee cohorts, nevertheless, suggests mortality at all levels of infestation. We propose that persistent parasitism overcomes a colony's ability to compensate for losses, leading to its sudden decline and death.

Animals↗

PCR amplification of a locus with RFLP alleles specific to African honey bees.

An anonymous honey bee locus, detected previously with a cloned probe, has HhaI RFLP alleles specific to African bees or common to both African and European bees. To facilitate identification of these alleles, this region, 1231, was made analyzable with the PCR. The two halves of the region, excluding the termini, were amplified as two overlapping segments. Restriction sites were mapped, and the site differences responsible for the allelic RFLP patterns were determined. In the first half of the region, two polymorphic HhaI sites are present in the common alleles, whereas one, the other, or both of the sites are absent in the African alleles. In the second half, a third polymorphic HhaI site is present or absent in both common and African alleles. A short part of the second half of the region, including more of the terminus, was amplified as a third segment. Within this segment, close to this terminus, a fourth polymorphic HhaI site is absent in some African alleles.

Alleles↗

Distribution of deformed wing virus within honey bee (Apis mellifera) brood cells infested with the ectoparasitic mite Varroa destructor.

The distribution of deformed wing virus (DWV) in adult female Varroa destructor and in their progeny in relation to the pupal host bee was investigated to evaluate acquisition and transfer of DWV by the mites. The results clearly show that adult female mites regularly act as competent vectors of DWV, however, they do not acquire or transfer virus on all possible occasions. Mother mites may contain DWV while the pupal host remains free from overt infection and both mother mites and mite progeny may not acquire detectable amounts of DWV from an infected host bee. However, a majority of mites feeding on pupae that emerge with deformed wings will contain DWV. The data also demonstrates that both adult and immature mite progeny most likely acquire DWV from DWV-infected host bees and not from their mother mites. Possible explanations for the obtained results are discussed.

Animals↗

Neotropical Africanized honey bees have African mitochondrial DNA.

Non-indigenous African honey bees have invaded most of South and Central America in just over 30 years. The genetic composition of this population and the means by which it rapidly colonizes new territory remain controversial. In particular, it has been unclear whether this 'Africanized' population has resulted from interbreeding between African and domestic European bees, or is an essentially pure African population. Also, it has not been known whether this population expanded primarily by female or by male migration. Restriction site mapping of 62 mitochondrial DNAs of African bees from Brazil, Venezuela and Mexico reveals that 97% were of African (Apis mellifera scutellata) type. Although neotropical European apiary populations are rapidly Africanized by mating with neotropical African males, there is little reciprocal gene flow to the neotropical African population through European females. These are the first genetic data to indicate that the neotropical African population could be expanding its range by female migration.

Animals↗

Chromosome-level genome assembly of the large carpenter bee Xylocopa dejeanii Lepeletier, 1841 (Hymenoptera: Apidae).

Xylocopinae, a diverse bee subfamily comprising over 1,000 bee species, and also a major model system for studying the pollination and evolution of sociality. The lack of chromosome-level genome assembly resources for the Xylocopinae limits our research of their biology and evolution. Here, we provided the first pseudo-chromosomes genome assembly of the Xylocopa dejeanii combined PacBio CLR long reads, Illumina sequences, and Hi-C data. The final genome is 194.44 Mb located in 16 chromosomes. Our assembly includes 141 scaffolds, with a scaffold N50 length of 13.15 Mb. BUSCO analysis revealed 99.00% completeness. Genome annotation identified 28.27 Mb of repetitive elements, 10,970 protein-coding genes, and 432 ncRNAs. This high-quality X. dejeanii assembly advances our understanding of Xylocopinae genomics and provides new insights into bee evolution.

Animals↗

Bioenergetic actions of beta-bungarotoxin, dendrotoxin and bee-venom phospholipase A2 on guinea-pig synaptosomes.

Low concentrations of beta-bungarotoxin or bee-venom phospholipase A2 cause a progressive Ca2+-dependent increase in the proton permeability of the mitochondria within the synaptosomal cytosol, manifested as an increase in oligomycin-insensitive respiration and a partial depolarization of the mitochondrial membrane potential. This uncoupling appears to be a consequence of fatty acids liberated by phospholipase A2 activity at the plasma membrane, since it can be mimicked by the addition of oleate-albumin complexes, in which case there is no requirement for external Ca2+. Dendrotoxin does not affect the mitochondrial proton permeability in situ, but protects partially against the uncoupling action of beta-bungarotoxin. In contrast, this effect of bee-venom phospholipase A2 is unaffected by dendrotoxin. beta-Bungarotoxin, but not bee-venom phospholipase A2, induces a slow progressive depolarization of the plasma membrane. The action of beta-bungarotoxin at the plasma membrane appears not to be related to fatty acid production, since it is augmented rather than inhibited by raising albumin concentrations in the medium. It is concluded that beta-bungarotoxin has at least two actions on intact synaptosomes, both of which may involve interaction at the plasma membrane with a site common to dendrotoxin: first, a mitochondrial uncoupling mediated by fatty acids and, secondly, a depolarization at the plasma membrane.

Animals↗

Protection by chlorpromazine, albumin and bivalent cations against haemolysis induced by melittin, [Ala-14]melittin and whole bee venom.

The ability of the peptides melittin, [Ala-14]melittin (P14A) and whole bee venom to lyse red blood cells (RBC) and to cause shape transformation, binding, partitioning and changes in volume of the cells during haemolysis, as well as the action of the bivalent cations Zn2+ and Ca2+, chlorpromazine, albumin and plasma on the peptide-induced haemolysis of RBC in high ionic-strength solution, have been investigated. The protective effect of all inhibitors depends on whether they have been added to the media before or after the cells. When added before the cells they reduced significantly the rate of peptide-induced haemolysis and shape transformation. The effect was maximal when agents acted simultaneously after introduction of the cells into the media containing both inhibitors and peptides. Incubation of the cells in isotonic solution before the addition of peptides enhanced 2-3-fold the RBC susceptibility (i.e. rate of haemolysis) to lytic action of the same amount of peptides, and increased the order of the haemolytic reaction, although the power law coefficient did not exceed a value of 2 for all peptides, suggesting that haemolysis is attributable to the monomeric or dimeric forms of the peptides. Partition coefficients were of the order of approximately 10(6) M-1, and P14A possessed a value 3-fold larger compared with melittin and bee venom, which correlated with its enhanced haemolytic activity. The protective action of inhibitors against peptide-induced haemolysis has been explained on the basis of their ability to compete with peptide binding at an early stage of peptide-membrane interaction, and not as a result of inhibition of a pre-existing peptide-induced pore. Whereas melittin increased the volume of RBC during haemolysis, P14A, melittin in the presence of phospholipase A2 or bee venom, reduced the volume in a concentration-dependent manner. The present data reveal the significant role of the initial stage of peptide-membrane interaction and peptide structure in the mechanism of haemolysis. These data are not consistent with a lipid-based mechanism of peptide-induced haemolysis, indicating that the mode of peptide-protein interaction is an important and decisive step in the haemolytic mechanism. It should be noted that data (in the form of three additional Tables) on the ability of inhibitors to protect cells from haemolysis when inhibitor and peptide act simultaneously are available. They are reported in Supplementary Publication SUP 50178, which has been deposited at the British Library Document Supply Centre, Boston Spa, Wetherby, West Yorkshire LS23 7BQ, U.K., from whom copies can be obtained on the terms indicated in Biochem. J. (1996) 313, 9.

Albumins↗

Isolation of seven unique biogenic amine receptor clones from the honey bee by library scanning.

The biogenic amine receptor genes constitute an ancient and highly divergent family within the larger superfamily of G-protein-coupled receptors. These receptors play a central role in modulating nerve cell activity and thus behaviour. Because the honey bee offers numerous advantages for behavioural studies we endeavoured to isolate as many members of this gene family as possible from the bee. We compared numerous approaches to gene isolation and found that PCR amplification from small subfractions of cDNA or genomic DNA libraries enabled us to isolate clones that are otherwise undetectable. In total we isolated seven biogenic amine receptor clones and identified five additional related sequences by low-stringency Southern hybridization. Two clones, AmBAR4 and AmBAR6, are 84% and 72% identical to the Drosophila 5-HT2 and D1b receptors, respectively, and probably represent orthologous genes. Phylogenetic analysis indicates that AmBAR5 clusters loosely with a variety of tyramine and octopamine receptors with which it shares <66% identity. The other four clones, AmBAR1, AmBAR2, AmBAR3 and AmBAR7, are weakly to moderately related (28-45% identical) to Drosophila dopaminergic or mammalian adrenergic receptors and probably represent receptors of these classes whose orthologues have not previously been isolated from any insect. The honey bee clones expand the size of the known insect biogenic amine receptor gene family to sixteen members. Therefore the size of the biogenic amine receptor gene family of insects approaches that of vertebrates. This is true despite the reduced behavioural and genetic complexity of the insects relative to vertebrate animals.

Amino Acid Sequence↗

Chloride-dependent transport of NH4+ into bee retinal glial cells.

Mammalian astrocytes convert glutamate to glutamine and bee retinal glial cells convert pyruvate to alanine. To maintain such amination reactions these glial cells may take up NH4+/NH3. We have studied the entry of NH4+/NH3 into bundles of glial cells isolated from bee retina by using the fluorescent dye BCECF to measure pH. Ammonium caused intracellular pH to decrease by a saturable process: the rate of change of pH was maximal for an ammonium concentration of about 5 mM. This acidifying response to ammonium was abolished by the loop diuretic bumetanide (100 microM) and by removal of extracellular Cl-. These results strongly suggest that ammonium enters the cell by contransport of NH4+ with Cl-. Removal of extracellular Na+ did not abolish the NH(4+)-induced acidification. The NH(4+)-induced pH change was unaffected when nearly all K+ conductance was blocked with 5 mM Ba2+ showing that NH4+ did not enter through Ba(2+)-sensitive ion channels. Application of 2 mM NH4+ led to a large increase in total intracellular proton concentration estimated to exceed 13.5 mEq/L. As the cell membrane appeared to be permeable to NH3, we suggest that when NH4+ entered the cells, NH3 left, so that protons were shuttled into the cell. This shuttle, which was strongly dependent on internal and external pH, was quantitatively modelled. In retinal slices, 2 mM NH4+ alkalinized the extracellular space: this alkalinization was reduced in the absence of bath Cl-. We conclude that NH4+ enters the glial cells in bee retina on a cotransporter with functional similarities to the NH4+(K+)-Cl- cotransporter described in kidney cells.

Animals↗

[Hyposensitisation treatment with pure bee venom (author's transl)].

Hyposensitisation with pure bee venom was undertaken in eleven persons with proven allergy to bee stings. Rapid hyposensitisation led to side effects in seven of them during dosage increase. Serial determination of total IgA, IgG, IgM and alpha1-antitrypsin revealed no changes, while allergen-specific IgG and IgE and total IgE increased maximally after 30 days. All persons were exposed to one bee sting during the treatment: all tolerated it well without the allergic reactions which had previously been noted.

Adolescent↗

Evolution of sociality in a primitively eusocial lineage of bees.

Eusociality is a major evolutionary innovation involving alterations in life history, morphology, and behavior. Advanced eusocial insects, such as ants, termites, and corbiculate bees, cannot provide insights into the earliest stages of eusocial evolution because eusociality in these taxa evolved long ago (in the Cretaceous) and close solitary relatives are no longer extant. In contrast, primitively eusocial insects, such as halictid bees, provide insights into the early stages of eusocial evolution because eusociality has arisen recently and repeatedly. By mapping social behavior onto well-corroborated phylogenies, I show that eusociality has arisen only three times within halictid bees (contrary to earlier estimates of six or more origins). Reversals from eusocial to solitary behavior have occurred as many as 12 times, indicating that social reversals are common in the earliest stages of eusocial evolution. Important attributes of social complexity (e.g., colony size, queen/worker dimorphism) show no obvious association with phylogeny, and some reversals to solitary nesting are related to host-plant switches (from polylecty to oligolecty). These results provide a glimpse of social evolution in its earliest stages and provide insights into the early evolution of advanced eusocial organisms.

Animals↗

Signatures of selection among sex-determining alleles of the honey bee.

Patterns of DNA polymorphisms are a primary tool for dissecting signatures of selection; however, the underlying selective forces are poorly understood for most genes. A classical example of diversifying selection is the complementary sex-determining locus that is found in the very large insect order Hymenoptera (bees, wasps, ants, and sawflies). The gene responsible for sex determination, the complementary sex determiner (csd), has been most recently identified in the honey bee. Females are heterozygous at this locus. Males result when there is only one functional allele present, as a result of either homozygosity (fertilized eggs) or, more commonly, hemizygosity (unfertilized eggs). The homozygotes, diploid males, do not reproduce and have zero fitness, which implies positive selection in favor of rare alleles. Large differences in csd cDNA sequences within and between four populations were found that fall into two major groups, types I and II. Type I consists of several allelic lineages that were maintained over an extended period, an indication of balancing selection. Diversifying selection has operated on several confined parts of the protein, as shown by an excess of nonsynonymous differences. Elevated sequence differences indicate another selected part near a repeat region. These findings have general implications about the understanding of both the function of the multiallelic mechanism and the adaptive processes on the level of nucleotide sequences. Moreover, the first csd sequence data are a notable basis for the avoidance of diploid males in bee selection programs by allele-assisted breeding.

Alleles↗

Genomic dissection of behavioral maturation in the honey bee.

Honey bees undergo an age-related, socially regulated transition from working in the hive to foraging that has been previously associated with changes in the expression of thousands of genes in the brain. To understand the meaning of these changes, we conducted microarray analyses to examine the following: (i) the ontogeny of gene expression preceding the onset of foraging, (ii) the effects of physiological and genetic factors that influence this behavioral transition, and (iii) the effects of foraging experience. Although >85% of approximately 5,500 genes showed brain differences, principal component analysis revealed discrete influences of age, behavior, genotype, environment, and experience. Young bees not yet competent to forage showed extensive, age-related expression changes, essentially complete by 8 days of age, coinciding with previously described structural brain changes. Subsequent changes were not age-related but were largely related to effects of juvenile hormone (JH), suggesting that the increase in JH that influences the hive bee-forager transition may cause many of these changes. Other treatments that also influence the onset age of foraging induced many changes but with little overlap, suggesting that multiple pathways affect behavioral maturation. Subspecies differences in onset age of foraging were correlated with differences in JH and JH-target gene expression, suggesting that this endocrine system mediates the genetic differences. We also used this multifactorial approach to identify candidate genes for behavioral maturation. This successful dissection of gene expression indicates that, for social behavior, gene expression in the brain can provide a robust indicator of the interaction between hereditary and environmental information.

Aging↗

New components of the honey bee (Apis mellifera L.) queen retinue pheromone.

The honey bee queen produces pheromones that function in both releaser and primer roles such as attracting a retinue of workers around her, attracting drones on mating flights, preventing workers from reproducing at the individual (worker egg-laying) and colony (swarming) level, and regulating several other aspects of colony functioning. The queen mandibular pheromone (QMP), consisting of five synergistic components, is the only pheromone chemically identified in the honey bee (Apis mellifera L.) queen, but this pheromone does not fully duplicate the pheromonal activity of a full queen extract. To identify the remaining unknown compounds for retinue attraction, honey bee colonies were selectively bred to have low response to synthetic QMP and high response to a queen extract in a laboratory retinue bioassay. Workers from these colonies were then used in the bioassay to guide the isolation and identification of the remaining active components. Four new compounds were identified from several glandular sources that account for the majority of the difference in retinue attraction between synthetic QMP and queen extract: methyl (Z)-octadec-9-enoate (methyl oleate), (E)-3-(4-hydroxy-3-methoxyphenyl)-prop-2-en-1-ol (coniferyl alcohol), hexadecan-1-ol, and (Z9,Z12,Z15)-octadeca-9,12,15-trienoic acid (linolenic acid). These compounds were inactive alone or in combination, and they only elicited attraction in the presence of QMP. There was still unidentified activity remaining in the queen extract. The queen therefore produces a synergistic, multiglandular pheromone blend of at least nine compounds for retinue attraction, the most complex pheromone blend known for inducing a single behavior in any organism.

Animals↗

Pheromone-mediated gene expression in the honey bee brain.

We tested the hypothesis that queen mandibular pheromone (QMP) causes changes in gene expression in the brain of the adult worker honey bee, and that these changes can be correlated to the downstream behavioral responses induced by QMP. In support of the first hypothesis, cage experiments revealed that QMP transiently regulated expression of several hundred genes and chronically regulated the expression of 19 genes. Several of these genes were also affected by QMP in experiments with bee colonies in the field, demonstrating robust gene regulation by pheromone. To evaluate the second hypothesis, we focused on one function of QMP: delaying the transition from working in the hive (e.g., brood care, or "nursing") to foraging. We compared the list of QMP-regulated genes with the lists of genes differentially regulated in nurse and forager brains generated in a separate study. QMP consistently activated "nursing genes" and repressed "foraging genes," suggesting that QMP may delay behavioral maturation by regulating genes in the brain that produce these behavioral states. We also report here on an ortholog of the Drosophila transcription factor kruppel homolog 1 that was strongly regulated by QMP, especially in the mushroom bodies of the bee brain. These results demonstrate chronic gene regulation by a primer pheromone and illustrate the potential of genomics to trace the actions of a pheromone from perception to action, and thereby provide insights into how pheromones regulate social life.

Animals↗