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PubMed2004

Tyramine functions as a toxin in honey bee larvae during Varroa-transmitted infection by Melissococcus pluton.

From wounds of honey bee pupae, caused by the mite Varroa destructor, coccoid bacteria were isolated and identified as Melissococcus pluton. The bacterial isolate was grown anaerobically in sorbitol medium to produce a toxic compound that was purified on XAD columns, gelfiltration and preparative HPLC. The toxic agent was identified by GC-MS and FTICR-MS as tyramine. The toxicity of the isolated tyramine was tested by a novel mobility test using the protozoon Stylonychia lemnae. A concentration of 0.2 mg/ml led to immediate inhibition of mobility. In addition the toxicity was studied on honey bee larvae by feeding tyramine/water mixtures added to the larval jelly. The lethal dosis of tyramine on 4-5 days old bee larvae was determined as 0.3 mg/larvae when added as a volume of 20 microl to the larval food in brood cells. Several other biogenic amines, such as phenylethylamine, histamine, spermine, cadaverine, putrescine and trimethylamine, were tested as their hydrochloric salts for comparison and were found to be inhibitory in the Stylonychia mobility test at similar concentrations. A quantitative hemolysis test with human red blood cells revealed that tyramine and histamine showed the highest membranolytic activity, followed by the phenylethylamine, trimethylamine and spermine, while the linear diamines, cadaverine and putrescine, showed a significantly lower hemolysis when calculated on a molar amine basis. The results indicate that tyramine which is a characteristic amine produced by M. pluton in culture, is the causative agent of the observed toxic symptoms in bee larvae. Thus this disease, known as European foulbrood, is possibly an infection transmitted by the Varroa destructor mite.

Animals↗
PubMed2004

Transcriptional immune responses by honey bee larvae during invasion by the bacterial pathogen, Paenibacillus larvae.

Honey bee larvae are highly susceptible to the bacterial pathogen Paenibacillus larvae only during the first instar of larval development. Transcript levels were measured for genes encoding two antimicrobial peptides, abaecin and defensin, as well as for two candidates in the immune response cascade (PGRP-LD and masquerade) in control larvae and larvae exposed to the pathogen. Transcripts for all four are present throughout development. This suggests that other physiological or dietary factors may better explain the age-based change in vulnerability to this pathogen. One of these genes, abaecin, shows significant up-regulation 24 h following oral inoculation with P. larvae, precisely when the bacterium surmounts the midgut epithelia of bees. Expression of both antimicrobial peptides varied by 1000-fold across different nestmate bees, indicating an allelic component to their expression. The implications of these results for current hypotheses related to disease tolerance in social insects are discussed, along with implications for breeding bees resistant to this important disease.

Animals↗
PubMed2006

Horizontal and vertical transmission of viruses in the honey bee, Apis mellifera.

The most crucial stage in the dynamics of virus infections is the mode of virus transmission. In general, transmission of viruses can occur through two pathways: horizontal and vertical transmission. In horizontal transmission, viruses are transmitted among individuals of the same generation, while vertical transmission occurs from mothers to their offspring. Because of its highly organized social structure and crowded population density, the honey bee colony represents a risky environment for the spread of disease infection. Like other plant and animal viruses, bee viruses use different survival strategies, including utilization of both horizontal and vertical routes, to transmit and maintain levels in a host population. In this review, we explore the current knowledge about the honey bee viruses and transmission routes of bee viruses. In addition, different transmission strategies on the persistence and dynamics of host-pathogen interactions are also discussed.

Animals↗
PubMed2006

Beepath: an ordered quantitative-PCR array for exploring honey bee immunity and disease.

Honey bees and other insects face many important parasites and pathogens against which they have evolved behavioral, morphological, physiological, and immune-based defenses. To help validate honey bee immune-gene candidates and determine their responsiveness to pathogens, a quantitative-PCR array was developed to measure transcript levels for 48 honey bee and pathogen genes in parallel. It is shown that this array can accurately measure host and pathogen transcript abundance, providing a new tool for assessing the environmental and genetic components behind honey bee immunity. By using common platforms and chemistries (e.g., SYBR-Green intercalating dye), quantitative-PCR arrays such as the one described should have diverse uses in invertebrate pathology.

Animals↗
PubMed2006

Nutritional content of fresh, bee-collected and stored pollen of Aloe greatheadii var. davyana (Asphodelaceae).

Aloe greatheadii var. davyana is the most important indigenous South African bee plant. Fresh, bee-collected and stored pollen of this aloe was collected and analysed for its nutritional content, including amino acid and fatty acid composition. Highly significant differences were found between the three types of pollen. Collection and storage by the bees resulted in increased water (13-21% wet weight) and carbohydrate content (35-61% dry weight), with a resultant decrease in crude protein (51-28% dry weight) and lipid content (10-8% dry weight). Essential amino acids were present in equal or higher amounts than the required minimum levels for honeybee development, with the exception of tryptophan. Fatty acids comprised a higher proportion of total lipid in fresh pollen than in bee-collected and stored pollen. This study is the first to compare the changes that occur in pollen of a single species after collection by honeybees.

Aloe↗
PubMed2006

A review of neurohormone GPCRs present in the fruitfly Drosophila melanogaster and the honey bee Apis mellifera.

G protein-coupled receptor (GPCR) genes are large gene families in every animal, sometimes making up to 1-2% of the animal's genome. Of all insect GPCRs, the neurohormone (neuropeptide, protein hormone, biogenic amine) GPCRs are especially important, because they, together with their ligands, occupy a high hierarchic position in the physiology of insects and steer crucial processes such as development, reproduction, and behavior. In this paper, we give a review of our current knowledge on Drosophila melanogaster GPCRs and use this information to annotate the neurohormone GPCR genes present in the recently sequenced genome from the honey bee Apis mellifera. We found 35 neuropeptide receptor genes in the honey bee (44 in Drosophila) and two genes, coding for leucine-rich repeats-containing protein hormone GPCRs (4 in Drosophila). In addition, the honey bee has 19 biogenic amine receptor genes (21 in Drosophila). The larger numbers of neurohormone receptors in Drosophila are probably due to gene duplications that occurred during recent evolution of the fly. Our analyses also yielded the likely ligands for 40 of the 56 honey bee neurohormone GPCRs identified in this study. In addition, we made some interesting observations on neurohormone GPCR evolution and the evolution and co-evolution of their ligands. For neuropeptide and protein hormone GPCRs, there appears to be a general co-evolution between receptors and their ligands. This is in contrast to biogenic amine GPCRs, where evolutionarily unrelated GPCRs often bind to the same biogenic amine, suggesting frequent ligand exchanges ("ligand hops") during GPCR evolution.

Animals↗
PubMed2006

Mechanisms of bee venom-induced acute renal failure.

The spread of Africanized bees in the American continent has increased the number of severe envenomation after swarm attacks. Acute renal failure (ARF) is one of the major hazards in surviving patients. To assess the mechanisms of bee venom-induced ARF, rats were evaluated before, up to 70 min and 24h after 0.5mg/kg of venom injection. Control rats received saline. Bee venom caused an early and significant reduction in glomerular filtration rate (GFR, inulin clearance, 0.84+/-0.05 to 0.40+/-0.08 ml/min/100g, p<0.0001) and renal blood flow (RBF, laser Doppler flowmetry), which was more severe in the cortical (-72%) than in the medullary area (-48%), without systemic blood pressure decrease. Creatine phosphokinase, lactic dehydrogenase (LDH) and serum glutamic oxaloacetic transaminase increased significantly, pointing to rhabdomyolysis, whereas serum glutamic pyruvic transaminase and hematocrit remained stable. Twenty-four hours after venom, RBF recovered but GFR remained significantly impaired. Renal histology showed acute tubular injury and a massive tubular deposition of myoglobin. Venom was added to isolated rat proximal tubules (PT) suspension subjected to normoxia and hypoxia/reoxygenation (H/R) for direct nephrotoxicity evaluation. After 60 min of incubation, 0.1, 2 and 10 microg of venom induced significant increases in LDH release: 47%, 64% and 86%, respectively, vs. 21% in control PT while 2 microg of venom enhanced H/R injury (85% vs. 55%, p<0.01). These results indicate that vasoconstriction, direct nephrotoxicity and rhabdomyolysis are important mechanisms in the installation of bee venom-induced ARF that may occur even without hemolysis or hypotension.

Acute Kidney Injury↗
PubMed2006

Vertical transmission of American foulbrood (Paenibacillus larvae) in honey bees (Apis mellifera).

The mode of transmission between hosts (horizontal versus vertical) of disease agents is important for determination of the evolution of virulence in pathogens. For disease management, it is imperative that the epidemiology of the disease is understood and pathogen transmission rates between hosts is a key factor for this understanding. Surprisingly little is known about transmission rates in honey bee pathology. We have studied the rate of vertical transmission of Paenibacillus larvae, the causative agent of American foulbrood (AFB) in honey bee colonies, as colonies reproduce by colony fission (swarming), by culturing for the spores from repetitive samples of adult bees. The results demonstrate vertical pathogen transmission to daughter swarms. The spore density declines over time in both mother colonies and daughter swarms if mother colonies do not exhibit clinical disease symptoms. Occasional positive samples more than a year post swarming, also in daughter swarms, indicate production of infectious spores from diseased larvae, without clinical disease observable by beekeepers, and/or maintenance of infective spores in the hive environment, allowing both horizontal and vertical transmission to be maintained. The results suggest that the virulence of AFB, being lethal at colony level in contrast to other bee diseases shaped by evolution, could be dependent on apicultural practices and that the pathogen probably would be maintained without causing frequent colony mortality in a natural system.

Animals↗
PubMed2006

Phylogenetic analysis of honey bee behavioral evolution.

DNA sequences from three mitochondrial (rrnL, cox2, nad2) and one nuclear gene (itpr) from all 9 known honey bee species (Apis), a 10th possible species, Apis dorsata binghami, and three outgroup species (Bombus terrestris, Melipona bicolor and Trigona fimbriata) were used to infer Apis phylogenetic relationships using Bayesian analysis. The dwarf honey bees were confirmed as basal, and the giant and cavity-nesting species to be monophyletic. All nodes were strongly supported except that grouping Apis cerana with A. nigrocincta. Two thousand post-burnin trees from the phylogenetic analysis were used in a Bayesian comparative analysis to explore the evolution of dance type, nest structure, comb structure and dance sound within Apis. The ancestral honey bee species was inferred with high support to have nested in the open, and to have more likely than not had a silent vertical waggle dance and a single comb. The common ancestor of the giant and cavity-dwelling bees is strongly inferred to have had a buzzing vertical directional dance. All pairwise combinations of characters showed strong association, but the multiple comparisons problem reduces the ability to infer associations between states between characters. Nevertheless, a buzzing dance is significantly associated with cavity-nesting, several vertical combs, and dancing vertically, a horizontal dance is significantly associated with a nest with a single comb wrapped around the support, and open nesting with a single pendant comb and a silent waggle dance.

Animals↗
PubMed1996

Food allergy to honey: pollen or bee products? Characterization of allergenic proteins in honey by means of immunoblotting.

OBJECTIVE: To characterize the allergenic components of honey, 23 patients allergic to honey were investigated. All displayed allergic symptoms after ingestion of honey or honey-containing products, ranging from itching in the oral mucosa to severe systemic symptoms to anaphylactic shock. METHODS AND RESULTS: Immunoblot analyses of the patients' sera revealed IgE binding to proteins at a molecular mass of 54 kd, 60 kd, 72 kd, or to a 30 kd/33 kd double band, or to both in sunflower honey extracts. The three bands corresponding to higher molecular mass proteins could also be detected in the three other kinds of honey (locust tree, European chestnut and forest honey) that were tested and represented bee products because IgE binding to these proteins was inhibited by extracts of honeybee heads and extracts of isolated bee venom sacs. The 30 kd/33 kd bands could be identified as sunflower honey-specific. When testing sera from patients allergic to bee venom with honey extracts, in seven of 10 cases IgE binding to bee-specific components could be observed. CONCLUSION: Both proteins derived from secretions of pharyngeal and salivary glands of honeybee heads and pollen proteins contained in the honey cause allergic reactions to honey.

Adolescent↗
PubMed1999

Viability assessment of honey bee, Apis mellifera, sperm using dual fluorescent staining.

Since the development of instrumental insemination of honey bee (Apis mellifera) queens in the 1930s, there has been interest in the evaluation and in vitro storage of semen. Several fluorescent stains, when used in combination, have been effectively used to assess sperm viability in mammalian and avian species. Our objectives were to test two combinations of living:dead fluorescent stains, SYBR-14 with propidium iodide (PI), or Calcein-AM with PI, and validate the use of these probes with honey bee sperm. SYBR-14 is a nuclear stain producing green fluorescence of the DNA in living sperm, Calcein-AM is a membrane-permeant esterase substrate staining entire sperm green, and PI is a traditional dead cell stain giving a contrasting red color. Both living stains fluoresced bee sperm, but the SYBR-14:PI produced a clearer distinction between the living and dead sperm. A graduated series of known living:dead sperm proportions was used to validate the accuracy of the stains for determining sperm viability in honey bees.

Animals↗
PubMed2001

Responsiveness to sucrose affects tactile and olfactory learning in preforaging honey bees of two genetic strains.

Using sucrose solution as reward, we conditioned preforaging honey bees of two genetic strains to tactile stimuli or to odours. Acquisition, extinction of conditioned responses and discrimination between the conditioned stimuli and alternative tactile or olfactory stimuli were measured. Bees of the two genetic strains were selected for their foraging behaviour. In addition, they differ in their responsiveness to sucrose. To analyse the effects of sucrose perception on learning parameters independent of foraging behaviour, we conditioned preforaging bees of the two strains and compared their performance in tactile and olfactory learning paradigms. In both strains, acquisition in tactile and olfactory conditioning is determined by responsiveness to sucrose. There is no genetic effect on the relationship between responsiveness to sucrose and acquisition. Bees responding to low sucrose concentrations perform better than ones that only respond to higher concentrations. Extinction of conditioned responses correlates with acquisition. Responses to alternative stimuli are independent of responsiveness to sucrose.

Animals↗
PubMed1997

Expansion of the neuropil of the mushroom bodies in male honey bees is coincident with initiation of flight.

The mushroom bodies (MB), the insect brain structures most often associated with learning, have previously been shown to exhibit structural plasticity during the adult behavioral development of female worker and queen honey bees. We now show that comparable morphological changes occur in the brains of male honey bees (drones). The volume of the MB in the brains of drones was estimated from tissue sections using the Cavalieri method. Brains were obtained from six groups of drones that differed in age and flight experience. Circulating levels of juvenile hormone (JH) in these drones were determined by radioimmunoassay (RIA). There was an expansion of the neuropil of the MB that was temporally associated with drone behavioral development, as in female queens and workers. The observed changes in drones were maintained in the presence of low levels of JH, also as in females. These results suggest that expansion of the neuropil of the MB in honey bees is associated with learning the location of the nest, because this learning is the most prominent aspect of behavioral development common to all members (workers, drones, queen) of the honey bee colony.

Age Factors↗
PubMed2001

Eicosanoids act in nodulation reactions to bacterial infections in newly emerged adult honey bees, Apis mellifera, but not in older foragers.

Nodulation is the first, and qualitatively predominant, cellular defense reaction to bacterial infections in insects. We tested the hypothesis that eicosanoids also mediate nodulation reactions to bacterial challenge in adults of a social insect, the honey bee, Apis mellifera. Treating newly-emerged experimental bees with the eicosanoid biosynthesis inhibitor, dexamethasone, impaired nodulation reactions to bacterial infections, and the influence of dexamethasone was reversed by treating infected insects with arachidonic acid, an eicosanoid precursor. Several other eicosanoid biosynthesis inhibitors, including the cyclooxygenase inhibitor, indomethacin, and the dual cyclooxygenase/lipoxygenase inhibitor, phenidone, also impaired the ability of experimental honeybees to form nodules in reaction to bacterial challenge. The influence of phenidone on nodulation was expressed in a dose-dependent manner. However, in experiments with older honey bees foragers, similar bacterial challenge did not evoke nodulation reactions. We infer from our results that while eicosanoids mediate cellular immune responses to bacterial infections in newly emerged honey bees, and more broadly, in most insect species, nodulation reactions to bacterial challenge probably do not occur in all phases of insect life cycles.

Abdomen↗
PubMed2005

The impact of host starvation on parasite development and population dynamics in an intestinal trypanosome parasite of bumble bees.

Host nutrition plays an important role in determining the development and success of parasitic infections. While studies of vertebrate hosts are accumulating, little is known about how host nutrition affects parasites of invertebrate hosts. Crithidia bombi is a gut trypanosome parasite of the bumble bee, Bombus terrestris and here we use it as a model system to determine the impact of host nutrition on the population dynamics and development of micro-parasites in invertebrates. Pollen-starved bees supported significantly smaller populations of the parasite. In pollen-fed bees the parasite showed a temporal pattern in development, with promastigote transmission stages appearing at the start of the infection and gradually being replaced by choanomastigote and amastigote forms. In pollen-starved bees this developmental process was disrupted, and there was no pattern in the appearance of these three forms. We discuss the implications of these results for parasite transmission, and speculate about the mechanisms behind these changes.

Animals↗
PubMed1999

Sulfated sialic acid-polymers inhibit the cytotoxic action of bee and snake venom.

Colominic acid is an alpha2,8-linked sialic acid polymer produced by Escherichia coli. We found that synthetic sulfated-colominic acids (SC) remarkably inhibited the cytotoxicity of bee and snake venom toward mouse fibroblast cells, but colominic acids showed no inhibition themselves, indicating the important role of sulfate groups in the inhibitory activity of SC. Other sulfated carbohydrates such as chondroitin sulfates, heparin and heparan sulfate showed no inhibition. SC also exhibited potent inhibition of melittin, a highly basic peptide, which is a major cytotoxic component of bee venom. SC did not inhibit phospholipase A2 activity in bee venom. This suggests that the inhibition of bee and snake venom by SC is due to inhibition of melittin and cardiotoxin, which is a cytolytic peptide in snake venom, respectively. SC with a higher sulfur content and a larger molecular mass showed more potent activity. The interaction between SC and melittin basically seems an ionic one, however, the conformation of SC is also likely important. For the binding of SC to melittin leading loss of its cytotoxic activity, the sulfate groups of SC must be properly arranged to interact with lysine and arginine residues of melittin molecules, which play an important role in the cytolytic activity. A higher molecular mass of SC substituted with more sulfate groups is required for more obvious inhibition of the cytotoxic activity.

Animals↗
PubMed2002

Reproduction of Varroa destructor in worker brood of Africanized honey bees (Apis mellifera).

Reproduction and population growth of Varroa destructor was studied in ten naturally infested, Africanized honey bee (AHB) (Apis mellifera) colonies in Yucatan, Mexico. Between February 1997 and January 1998 monthly records of the amount of pollen, honey, sealed worker and drone brood were recorded. In addition, mite infestation levels of adult bees and worker brood and the fecundity of the mites reproducing in worker cells were determined. The mean number of sealed worker brood cells (10,070 +/- 1,790) remained fairly constant over the experimental period in each colony. However, the presence and amount of sealed drone brood was very variable. One colony had drone brood for 10 months and another for only 1 month. Both the mean infestation level of worker brood (18.1 +/- 8.4%) and adult bees (3.5 +/- 1.3%) remained fairly constant over the study period and did not increase rapidly as is normally observed in European honey bees. In fact, the estimated mean number of mites fell from 3,500 in February 1997 to 2,380 in January 1998. In May 2000 the mean mite population in the study colonies was still only 1,821 mites. The fertility level of mites in this study was much higher (83-96%) than in AHB in Brazil (25-57%). and similar to that found in EHB (76-94%). Mite fertility remained high throughout the entire study and was not influenced by the amount of pollen, honey or worker brood in the colonies.

Animals↗
PubMed2004

A stingless bee (Melipona seminigra) marks food sources with a pheromone from its claw retractor tendons.

By depositing scent marks on flowers, bees reduce both the search time and the time spent with the handling of nonrewarding flowers. They thereby improve the efficiency of foraging. Whereas in honey bees the source of these scent marks is unknown, it is assumed to be the tarsal glands in bumble bees. According to histological studies, however, the tarsal glands lack any openings to the outside. Foragers of the stingless bee Melipona seminigra have previously been shown to deposit an attractant pheromone at sugar solution feeders, which is secreted at the tips of their tarsi. Here we show that the claw retractor tendons have specialized glandular epithelia within the femur and tibia of all legs that produce this pheromone. The secretion accumulates within the hollow tendon, which also serves as the duct to the outside, and is released from an opening at the base of the unguitractor plate. In choice experiments, M. seminigra was attracted by feeders baited with pentane extracts of the claw retractor tendons in the same way as it was attracted by feeders previously scent marked by foragers. Our results resolve the seeming contradiction between the importance of foot print secretions and the lack of openings of the tarsal glands.

Animals↗