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FMRFamide-like immunoreactivity in the brain of the honeybee (Apis mellifera). A light-and electron microscopical study.

Peptide-FMRFamide-like immunoreactivity in the brain and suboesophageal ganglion of the honeybee Apis mellifera L. is demonstrated with the peroxidase-antiperoxidase technique. Immunoreactivity is found in about 120 perikarya of the brain and in about 30 of the suboesophageal ganglion. These cells are distributed in 13 paired clusters representing neurons of different types including neurosecretory neurons projecting to neurohemal organs. Immunoreactivity of different intensity is found in the non-glomerular neuropil around the mushroom bodies, in the lateral protocerebrum, the central body, the optic tubercles, the lobula and medulla of optic lobe, the ocellar neuropil, in multiglomerular elements of the antennal lobes and in the dorsal deuterocerebrum. In the mushroom bodies, immunoreactivity is located in layers of the lobes and stalks, corresponding to intrinsic fibre bundles of some Kenyon cell types. The somata of these intrinsic cells did not show FMRFamide-like immunoreactivity. Electron microscopy of immunostained somata and nerve fibres was performed employing a pre-embedding peroxidase-antiperoxidase technique. Fibres of optic lobes and the non-glomerular neuropil contain immunoreactive dense core vesicles (diameter 50-165 nm) accumulated in boutons besides small synaptic vesicles and synaptic membrane specializations. Immunoreactive layers of the mushroom body neuropil were analysed at the ultrastructural level. Axon profiles with dense-core vesicles of a small type (diameter 35-75 nm) show only faint immunoreactive products. Immunoreactivity of intrinsic mushroom body neurons does not appear to be specifically correlated with synaptic organelles. Our results indicate that FMRFamide or related peptides peptides may be neuroactive compounds in different classes of nerve cells in the bee brain.

Animals

Phylogenetic relationships in the honeybee (genus Apis) as determined by the sequence of the cytochrome oxidase II region of mitochondrial DNA.

The complete nucleotide sequence of the mitochondrial cytochrome oxidase II (COII) gene was determined for five species of the honeybee (Genus: Apis): A. andreniformis, A. cerana, A. dorsata, A. florea, and A. koschevnikovi; these were then compared to the known sequence of the A. millifera gene from Crozier et al. (1989, Mol. Biol. Evol., 6: 399-411) and the wasp Excristes roborator (Liu and Beckenbach, 1992, Mol. Phylogenet. Evol., 1:41-52). Phylogenetic relationships were derived using the parasimony methods DNAPARS and PROTPARS of Felsenstein ("PHYLIP Manual Version 3.4, "University Herbarium, Univ. of California, Berkeley). The results suggest that A. dorsata is the most ancestral species, followed by the branching of A. florea/A. andreniformis and A. koschevnikovi, and then A. mellifera and A. cerana. This inference differs from the currently accepted view that considers the A. florea/A. andreniformis line to be the most ancestral.

Amino Acid Sequence

Comparative in silico analysis of Apis mellifera immune responses to Varroa destructor and Tropilaelaps mercedesae: Common and mite-specific molecular signatures.

Parasitic mites Varroa destructor and Tropilaelaps mercedesae represent major threats to global honey bee (Apis mellifera) health and productivity, yet comparative molecular insights into host responses remain limited. To address this, we systematically compiled published studies (2015-2025) reporting genes associated with honey bee interactions with V. destructor (11 studies, 87 genes), T. mercedesae (4 studies, 35 genes), and hygienic behavior (6 studies, 44 genes). Gene identifiers were harmonized to the Amel_HAv3.1 genome assembly, yielding three non-redundant sets: 64 Varroa-associated, 34 Tropilaelaps-associated, and 44 hygienic behavior-associated genes. Venn analysis identified 10 overlapping genes (including A0A088A8D5, A0A088ADL8, ABAE_APIME, Def1, Def2, Gapdh, HYTA_APIME, Imd, LOC726783, and Vg), suggesting conserved defense mechanisms, while 41 and 24 genes were uniquely associated with Varroa and Tropilaelaps, respectively. Enrichment analyses revealed Varroa-responsive genes were enriched in immune processes, chitin catabolism, and signaling pathways (Toll/Imd, MAPK, Wnt). Tropilaelaps-associated genes were enriched for antibacterial defense and stress response, with Toll/Imd signaling as the sole significantly enriched pathway. Overlapping genes reinforced core innate immunity activation. Protein-protein interaction network centrality analysis identified key hub genes: Def1, HYTA_APIME, ABAE_APIME, PPO, Imd, PGRP-LC, Vg for Varroa; and ACPH1_APIME, MRJP1, Vg, LOC726783 for Tropilaelaps. Results demonstrate that, despite differences in mite biology, honey bees show a conserved immune response against both parasites, centered on antibacterial defense, humoral immunity, and activation of the Toll/Imd pathway. Although limited by the in-silico nature and research asymmetries reflecting Tropilaelaps' emergence, this curated resource establishes a comprehensive framework for elucidating shared and distinct molecular defense mechanisms. Ultimately, this approach prioritizes diagnostic markers and candidate genes for functional validation and breeding strategies to enhance colony resilience against mite‑driven disease globally.

Animals

Phospholipase A2 in venom extracts from honey bees (Apis mellifera L.) of different ages.

We measured phospholipase A2 activity in the venom of worker honey bees (Apis mellifera L.) of known ages using chemical (titrimetric) and radioallergosorbent methods. The two techniques give similar results. Low levels of phospholipase A2 are present in the venom system at the time of eclosion. Phospholipase A2 activity in the venom increases steadily through the 10 days after eclosion. Maximal phospholipase A2 levels (about 40 micrograms phospholipase A2/venom sac) are maintained through the rest of the life of a worker bee in summer.

Aging

Enzyme activities of the strains belonging to family Leptospiraceae detected by the API ZYM system.

A total of 32 strains of the family Leptospiraceae (23 strains of Leptospira interrogans, 6 strains of Leptospira biflexa, 2 strains of Leptonema and 1 strain of Leptospira parva) were examined for enzyme activities using 89 substrates (API ZYM system). More than 90% of the strains belonging to the family Leptospiraceae possessed strong activities of beta-D-galactosidase, beta-D-glucosidase and 5 esterases (C5, C6, C8, C9 and C10). More than 90% of the strains belonging to the genus Leptospira, except L. parva, had strong activities of L-lysine arylamidase and alpha-L-glutamate arylamidase. L. biflexa strains, except serovar andamana, were different from the other strains examined in that they possessed glycyl-glycine arylamidase, glycyl-phenylalanine arylamidase and L-tryptophan arylamidase. L. biflexa strains, except andamana, L. parva and Leptonema strains possessed strong activities of glycine arylamidase and leucyl-glycine arylamidase. Two strains of the genus Leptonema were different from the strains belonging to the genus Leptospira in that they possessed strong activities of beta-D-lactosidase. L. parva lacked alpha-D-galactosidase which other strains belonging to the family Leptospiraceae possessed. Dendrogram analysis revealed that strains belonging to the family Leptospiraceae were divided into 4 groups. The first group consisted of all strains belonging to L. interrogans and serovar andamana of L. biflexa; the second group consisted of the remaining 5 serovars of L. biflexa; the third group consisted of the genus Leptonema; and the fourth group consisted of only L. parva.

Aminopeptidases

Transfer of the methyl group of methionine to choline and to tRNA in the honeybee Apis mellifica L.

Contrary to some previous reports on the absence of biological transmethylation reactions in some insect species, the transfer of the methyl group of methionine-methyl 14C leading to choline and to methylated bases in tRNA was shown in the honeybee Apis mellifica. The addition of antibiotics to the food of the insect does not diminish the incorporation of radioactivity, proving that intestinal bacteria are not responsible for the methylation reactions observed.

Animals

Purification and properties of a very high density lipoprotein from the hemolymph of the honeybee Apis mellifera.

A larval-specific very high density lipoprotein (VHDL) has been isolated from the hemolymph of the honeybee Apis mellifera. VHDL was isolated by a combination of density gradient ultracentrifugation and gel filtration. The purified protein is a dimer of Mr 160,000 apoproteins as shown by chemical cross-linking with dimethyl suberimidate. N-Terminal sequence analysis indicates that the two polypeptide chains are identical. The holoprotein contains 10% lipid by weight and 2.6% covalently bound carbohydrate. A native Mr 330,000 species was obtained by gel permeation chromatography. Antiserum directed against VHDL was used to show that VHDL is distinct from other hemolymph proteins and appears to constitute a novel lipoprotein of unknown function. However, the lipoprotein is present in high amounts in hemolymph only at the end of larval life, suggesting a potential role in lipid transport and/or storage protein metabolism during metamorphosis.

Amino Acid Sequence

Learning by honeybees (Apis mellifera) on arrival at and departure from a feeding place.

The question of when in the course of a visit to a feeding place foraging honeybees (Apis mellifera) learn about its location was studied in Experiment 1 by moving the animals a short distance after they arrived and began to feed. A preference for the arrival place developed, although less rapidly than in control animals for which the arrival and departure places were the same. In Experiments 2-5, a distinctive object was used to define the location of the feeding place. When the object was removed after arrival or introduced only after arrival, the animals learned less about its color and shape than did control animals for which it was present throughout each visit. The results contradict the claim that honeybees learn about certain characteristics of a feeding place only on arrival and about others only on departure.

Animals

Learning in honeybees (Apis mellifera) as a function of sucrose concentration.

Foraging honeybees (Apis mellifera) were trained with 2 successively presented targets differing in color or odor, one of which always contained a 5-microliters drop of 50% sucrose solution and the other, a 5-microliters drop of 20% sucrose solution. Latency of response to each target was measured during the training, and at the conclusion, preference was measured in an unrewarded choice test. Analysis of the latencies showed both a prospective effect (faster response to the 50% target than to the 20% target) and a nonassociative retrospective effect (faster response after leaving the 20% target than after leaving the 50% target) reminiscent of the frustration effect in rats. The results both for prospective latency and for choice can be understood on the simple theory that the attractiveness of a target depends on the strength of its association with sucrose and that the effect of concentration is on asymptotic strength.

Animals

Simultaneous conditioning in honeybees (Apis mellifera).

Honeybees (Apis mellifera) were classically conditioned with odor as conditioned stimulus (CS), sucrose as unconditioned stimulus (US), and proboscis extension as response. The purpose of Experiment 1 (Ns = 26 and 27) was to look for facilitation of forward conditioning by CS-US overlap, but rapid conditioning without overlap left little room for improvement. In 2 further experiments, CS and US were simultaneous, and response to odor alone was measured in subsequent tests. In Experiment 2, a Simultaneous group (N = 25) responded more to the training odor than did an Unpaired control group (N = 25). In Experiment 3, a differentially conditioned Simultaneous group (N = 29) responded more to an odor paired with sucrose in training (S+) than to an odor presented alone (S-). The implications of the results for the problem of the role of amount of reward in honeybee learning are considered.

Animals

The antigenicity of the carbohydrate moiety of an insect glycoprotein, honey-bee (Apis mellifera) venom phospholipase A2. The role of alpha 1,3-fucosylation of the asparagine-bound N-acetylglucosamine.

A rabbit polyclonal antiserum raised against honey-bee (Apis mellifera) venom phospholipase A2 (PLA2) contains antibodies that react exclusively with its glycosylated variants and cross-react with plant glycoproteins. The interaction of anti-(horseradish peroxidase) antiserum with PLA2 suggests the existence of a carbohydrate determinant common to both glycoproteins. E.l.i.s.a. binding and inhibition experiments, employing glycoproteins and glycopeptides of plant and animal origin with known N-glycan structures, in combination with chemical and enzymic deglycosylation, identified alpha 1,3-fucosylation of the asparagine-bound N-acetylglucosamine as the antigenic determinant. This fucose residue is present in the N-glycan of PLA2 and is frequently found in plant glycoproteins, whereas mammalian glycoproteins lack this modification.

Acetylglucosamine

Isolation and partial characterization of insulin of the honeybee (Apis mellifica).

In the honeybee (Apis mellifica), insulin-like material was partially purified with acid ethanol extractions by a classic method for recovering insulin and following gel filtration on a Sephadex G-50 column. The preparations were characterized by their ability to cross-react with porcine insulin antibodies. Insulin-like biological activity was demonstrated using the insulin bioassay. Stimulation of glucose oxidation or lipogenesis was measured by isolated rat adipocytes. Insulin seems to be more widespread in invertebrates than was previously assumed.

Adipose Tissue

Estimation of the number of sex alleles and queen matings from diploid male frequencies in a population of Apis mellifera.

The distribution of diploid males in a population of Apis mellifera was obtained by direct examination of the sexual phenotypes of the larvae. Using these data, estimates are derived for the number of sex alleles and the number or matings undergone by the queen. The number of sex alleles is estimated to be 18.9. The estimate is larger than previous ones, which have ranged between 10 and 12. However, the increase in the number of sex alleles can be explained by the large effective population number for our data. The best estimator of the number of matings by a queen is a maximum likelihood type that assumes a prior distribution on the number of matings. For the data presented here, this estimate is 17.3. This estimate is compared to others in the literature obtained by different approaches.

Alleles

Complete amino acid sequence of cytochrome c from the honeybee, Apis mellifera, and evolutionary relationship of the honeybee to other insects on the basis of the amino acid sequence.

The complete amino acid sequence of cytochrome c purified from the honeybee, Apis mellifera was determined. Only one molecular species of cytochrome c was found in the honeybee throughout its metamorphic stages. On the basis of a comparison of the amino acid sequence of honeybee cytochrome c with those of cytochromes c from other insects, it seems that the bee has evolutionarily appeared earlier than would be expected from the morphological and fossil evidence. If the classical phylogenetic relationships of the honeybee are correct, the evolutionary rate of cytochrome c must have been more rapid in the honeybee than in other insects.

Amino Acid Sequence

Geographical overlap of two mitochondrial genomes in Spanish honeybees (Apis mellifera iberica).

Restriction enzyme cleavage maps of mitochondrial DNA from the Spanish honeybee, Apis mellifera iberica (Hymenoptera: Apidae), were compared with those from the European subspecies A. m. mellifera, A. m. ligustica, and A. m. carnica, and the African subspecies A. m. intermissa and A. m. scutellata. The mitochondrial DNA (mtDNA) of the two African subspecies can be distinguished by restriction fragment polymorphisms revealed by Hinf I digests. Two distinct mtDNA types were found among Spanish honeybees: a west European mellifera-like type, which predominates in the north of Spain, and an African intermissa-like type, which predominates in the south. Spain appears to be a region of contact and hybridization between the two subspecies A. m. intermissa and A. m. mellifera, which respectively represent African and west European honeybee lineages. This natural boundary between European and African honeybee populations in the Old World may provide a model for predicting the eventual outcome of the colonization of North America by introduced African honeybees.

Africa

Differential response of Apis mellifera acetylcholinesterase towards pirimicarb.

The kinetic analysis of Apis mellifera acetylcholinesterase inhibition by the carbamate pirimicarb showed that native and detergent-solubilized membrane enzyme exhibited slightly different carbamylation kinetics. The acetylcholinesterase form sensitive to phosphatidylinositol-specific phospholipase C (PI-PLC) was carbamylated more rapidly (kapp = 36.4 X 10(-3) min-1) than the PI-PLC-resistant counterpart (kapp = 10.13 X 10(-3) min-1) which had a behavior close to that of the soluble tryptic enzyme (kapp = 11.89 X 10(-3) min-1). A difference in acetylcholinesterase sensitivity towards pirimicarb was also observed between foraging and emerging bees. These results show that the molecular structure, the mode of preparation and the source of acetylcholinesterase from the bee head should be taken into account in accurate toxicological studies.

Acetylcholinesterase

Biochemical typing of urinary Escherichia coli strains by means of the API 20 E enterobacteriaceae system.

With the API 20 E Enterobacteriaceae system of biochemical testing, a biotype, coded numerically, was determined for each of 574 strains of Escherichia coli isolated from patients with urinary tract infection. The serotypes of the strains were also determined. Fifty-five different biotypes were identified, two accounting together for 42% of the strains examined and seven others each accounting for between 8.4 and 1.9%. There was little correlation between biotype and serotype. Fifty pairs of strains were isolated from patients before treatment. In 43 the biotype and serotype of both strains of each pair were the same. In six pairs the biotype, but not the serotypes, differed, the difference being limited to the results of the tests for lysine decarboxylase. The biotypes of the strains of the remaining pair differed widely although their serotypes were the same. It is suggested that this method of biotyping offers a simple but accurate way of discriminating between recrudescent urinary tract infection caused by E. coli and that due to reinfection.

Bacteriuria

Evolutionary history of the honey bee Apis mellifera inferred from mitochondrial DNA analysis.

Variability of mitochondrial DNA (mtDNA) of the honey bee Apis mellifera L. has been investigated by restriction and sequence analyses on a sample of 68 colonies from ten different subspecies. The 19 mtDNA types detected are clustered in three major phylogenetic lineages. These clades correspond well to three groups of populations with distinct geographical distributions: branch A for African subspecies (intermissa, monticola, scutellata, andansonii and capensis), branch C for North Mediterranean subspecies (caucasica, carnica and ligustica) and branch M for the West European populations (mellifera subspecies). These results partially confirm previous hypotheses based on morphometrical and allozymic studies, the main difference concerning North African populations, now assigned to branch A instead of branch M. The pattern of spatial structuring suggests the Middle East as the centre of dispersion of the species, in accordance with the geographic areas of the other species of the same genus. Based on a conservative 2% divergence rate per Myr, the separation of the three branches has been dated at about 1 Myr BP.

Animals