Anti-coagulase-negative Staphylococcus activity of ethanolic extracts of propolis from two Brazilian regions and synergism with antimicrobial drugs by the E-test method

Propolis is a natural resinous substance collected by bees from vegetal sources and its therapeutic properties have been investigated. In this work, we evaluated the inhibitory activity of ethanolic extracts of propolis (EEP) from the Southeast and South of Brazil on coagulase-negative Staphylococcus (CNS) growth as well as the EEP in vitro synergism with antimicrobial drugs by using the diffusion method (E-test). The EEP chemical characteristics (dry weight, pH, flavonoid and phenolic compounds) were determined. Seven drugs were tested, and synergism was observed between three drugs and Southeast EEP, six drugs and South EEP, and one drug and ethanol control. Ethanolic extracts of propolis from the South of Brazil presented the greatest flavonoid content and synergism rate, while EEP from the Southeast presented the greatest anti-CNS activity and phenolic compound content. Results showed the correlation among anti-CNS activity, synergism rate and chemical characteristics of propolis.

Microdissection: a tool for bee chromosome studies

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Changes in the morphology and ultrastructure of the Dufour's gland during the life cycle of the bumble bee queen, Bombus terrestris L-(Hymenoptera : Bombini)

The Dufour's gland is found closely associated with the sting apparatus of all female hymenopterans, playing multiple roles among bees. In some species of Bombus the gland may be involved in production of nestmate recognition pheromones, but in B. terrestris its function is not certain yet. The morphology of the :Dufour's gland of B. terrestris queens and the ultrastructural features of its cells were studied in different ages and behavioural stages using routine transmission electron microscopy. Measurements of the length and the diameter of the gland in the same conditions were also made. The Dufour's gland of the queen increases significantly in size (both in length and in diameter) with age and reproductive activity the ultrastructural features of the gland show electrondense material that comes from the haemolymph. This material is also present in the intercellular spaces, and is conducted to the subcuticular space, to be released directly into the glandular lumen. Hence at least part of the secretion is probably taken up directly from the haemolymph. The ultrastructural features indicate a more active phase of the gland corresponding to the period of egg-laying of the queen, and a decrease in activity when the queen is in hibernation as well as after the competition point. In conclusion, the gland is probably involved in reproduction, more specifically, in the marking off eggs.

Toxic effects of methanolic and dichloromethane extracts of flowers and peduncles of Stryphnodendron adstringens (Leguminosae : Mimosoideae) on Apis mellifera and Scaptotrigona postica workers

Laboratory bioassays were conducted to evaluate toxicity of methanolic and dichloromethane extracts obtained from Stryphnodendron adstringens to Apis mellifera and Scaptotrigona postica workers. The extracts were incorporated into the diet of the bees for evaluation of mortality rates. The ingestion bioassays were made with three concentrations (0.002mg/g, 0.005mg/g and 0.01 mg/g) for each bee species. The workers were kept in cages, with twenty workers per cage for each concentration tested. All bioassays had sixty workers in three cages that where maintained in a biological oxygen demand incubator with controlled temperature and humidity. The data obtained in the toxicity bioassays were analyzed statistically by Log Rank test and all methanolic and clichloromethane extracts showed significant (P < 0.0001) toxic effects in all tested concentrations.

Insect Venom Peptides

The insects of the order Hymenoptera ( bees, wasps, and ants) are classified in two groups, based on their life history: social and solitary. The venoms of the social Hymenoptera evolved to be used as defensive tools to protect the colonies of these insects from the attacks of predators. Generally they do not cause lethal effects but cause mainly inflammatory and/or immunological reactions in the victims of their stings. However, sometimes it is also possible to observe the occurrence of systemic effects like respiratory and/or kidney failure. Meanwhile, the venoms of solitary Hymenoptera evolved mainly to cause paralysis of the preys in order to permit egg laying on/within the prey's body; thus, some components of these venoms cause permanent/transient paralysis in the preys, while other components seem to act preventing infections of the food and future progenies. The peptide components of venoms from Hymenoptera are spread over the molar mass range of 1400 to 7000 da and together comprise up to 70% of the weight of freeze-dried venoms. Most of these toxins are linear polycationic amphipatic peptides with a high content of alpha-helices in their secondary structures. These peptides generally account for cell lysis, hemolysis, antibiosis, and sometimes promote the delivery of cellular activators/mediators through interaction with the G-protein receptor, and perhaps some of them are even immunogenic components. In addition to these peptides, the Hymenopteran venoms also may contain a few neurotoxins that target Na+ and/or Ca+2 channels or even the nicotinic ACh receptor. This review summarizes current knowledge of the biologically active Hymenoptera venoms.

Ultrastructure of the midgut epithelium of Meliponinae larvae with different developmental stages and diets

The comparative study of the ultrastructure of the midgut epithelium of stingless bee larvae that eat plant protein (pollen) and animal protein (carrion) throughout the larval phase, shows variations in the digestive cells that are only relative to larval aging and not to the type of larval diet. The cells of older larvae present a cytoplasm with empty spaces that result from emptying of lipid and glycogen stocks, and the presence of autophagic vacuoles. These results are discussed in relation to the hypothesis that variations in the digestive tract of insects may be associated with different diets or phylogeny. We conclude that different diets do not determine cell morphology adaptations in the studied species. As the variations in the ultrastructure of the midgut epithelium are the same in all studied species, including the necrophagous species Trigona hypogea, throughout the larval stage, this sequence of changes seems to be due to different physiological state during larval development.

Behavioral responses evoked in honey bee workers by Dufour gland extracts (Hymenoptera : Apidae)

A series of experiments was performed to assess possible roles of the honey bee Dufour gland secretion. Bioassays with extracts of queen and worker glands from two colonies were made under artificial conditions, in which nestmate and non-nestmate forager workers were tested. The results demonstrate that forager workers display behavioral responses when exposed to Dufour gland extracts of nestmates, but remain indifferent when exposed to non-nestmate extracts. Also, the results demonstrate that forager workers are attracted by virgin queen gland extracts, and repelled by forager worker extracts. The data demonstrate that the Dufour gland secretion is colony- and caste-specific. The attractant remains in the nest. The repellent effect of forager worker extracts is interpreted as an alarm-like pheromone. The attractant effect of virgin queen extracts could be useful in the swarming process to attract scout bees to the eventual founder virgin queen or to attract old foragers to the virgin queen that remains in the nest.

Morphological modifications induced by an artificial diet on the hypopharyngeal glands of Apis mellifera (Hymenoptera, apidae) during their degenerative process

The morphological aspects of the hypopharyngeal glands were analyzed in worker bees of Apis mellifera of 15 and 30 days of age. The individuals were kept in a room with controlled temperature at 32degreesC where they received water and either a protein or a high energy food. Nurse and foraging workers were used as a control for the experiment. The morphological results showed that the different diets modified the cell death characteristics and intensified its occurrence. Both diets caused precocious glandular degeneration. However, this anticipation of cell death was more pronounced in the glandular tissue of the workers who received the high energy diet when compared to the glands of the bees fed with the protein meal.The degenerative signs observed were an intense cytoplasmic vacuolization, with a loss of cytoplasm and of the cell boundaries, dilation or condensation of the cells and nuclei, and nuclear fragmentation. At the end of the degenerative process, we observed the extrusion of nuclei and, finally, the dissolution of the glands. The hypopharyngeal glands' remains were found in the haemolymph.

Toxicity of barbatimao to Apis mellifera and Scaptotrigona postica, under laboratory conditions

Two species of plants commonly known as barbatim (a) over tildeo occur in Brazil, Stryphnodendron adstringens (Fabaceae) (true barbatim (a) over tildeo) and Dimorphandra mollis (Caesalpiniaceae) (false barbatim (a) over tildeo). These two species have a similar flowering period and are considered by beekeepers to cause bee mortality during this period. Flowers were collected from both species, dehydrated, ground and incorporated into an experimental diet for bees of two different species, Apis mellifera and Scaptotrigona postica. Both plant species were toxic to A. mellifera, reducing their median survival. D. mollis was toxic to S. postica, and Stryphnodendron adstringens reduced median survival of this bee species even when used at a concentration of 2.5%. In a choice experiment carried out with A. mellifera and the two plant species, the honey bees could choose not to feed on the diets containing the flowers, and feed on sugar and honey instead, but they did not. This shows us that the flowers of S. adstringens were not repellent to the bees. The plants were more toxic to A. mellifera than to Scaptotrigona postica, a result that can be explained by the fact that A. mellifera was introduced into Brazil whereas S. postica is a native stingless bee.

Morphological features and occurrence of degenerative characteristics in the hypopharyngeal glands of the paper wasp Polistes versicolor (Olivier) (Hymenoptera : Vespidae)

Different histochemical techniques were applied to examine the morphological features of the secretory cells of hypopharyngeal glands in the wasp Polistes versicolor. The results showed that most analyzed individuals present active glands with secretion stored in the cytoplasm. In some glands, morphological analyses revealed the presence of degenerative characteristics. Analyses of cellular integrity, however, did not detect dead cells. The results showed that, in P. versicolor, the development and regression of the hypopharyngeal glands were not age related, unlike glands of social bees. (c) 2006 Elsevier Ltd. All rights reserved.

Modes of cell death in the hypopharyngeal gland of the honey bee (Apis mellifera L)

Different modes of cell death have been revealed in the regressing hypopharyngeal glands of worker honey bees. The hypopharyngeal gland, which is well developed in young nursing bees to produce protein for larval food, was seen to regress naturally in foraging adult worker bees. A range of techniques including histology, cytochemistry, in situ TUNEL, Annexin V and Comet assays indicated that cells within the gland demonstrate progressive symptoms of apoptosis, necrosis and a vacuolar form of programmed cell death. The latter mode of cell death did not display chromatin margination, but was accompanied by an enhanced level of autophagic and hydrolytic activity in which a cytosolic source of acid phosphatase became manifest in the extra-cisternal spaces. Normal and annexin-positive cells were found to occur in the younger nursing bees, whilst necrosis and an aberrant vacuolar type of apoptosis predominated in the older foraging bees. The relevance of these results to the classification of programmed cell death is discussed. (C) 2000 Academic Press.

Differences in the number of some antennal sensilla of four honey bee (Apis mellifera) types and comparisons with the defensive behaviour

The number of sensilla coeloconica, ampullacea and campaniformia of flagellomeres 3 to 10 of the antennae of workers of four honey bee types (Italian, Caucasian, African and Africanized) was studied by scanning electron microscopy. Comparisons of the four bee types showed that only African and Africanized honey bees did not differ from one another with respect to the number of sensilla coeloconica and ampullacea of flagellomere 10. African and Africanized honey bees and Caucasian and Italian honey bees also did not differ from one another in terms of flagellomere 9. In the other flagellomeres there were no differences among bee types. Italian and Caucasian honey bees differed from Africanized honey bees in terms of number of sensilla campaniformia on flagellomere 6, and Caucasian honey bees differed from African and Africanized honey bees in terms of flagellomere 3. Five significant but random correlation values were obtained between number of antennal sensilla and defensive behaviour in Africanized honey bees. Thus,there is no relationship between antennal structures and defence behaviour.

Enzymatic activity of hypopharyngeal gland extractsfrom workers of Apis mellifera (Hymenoptera, Apidae, Apinae)

This research presents a comparative study of enzymatic activity of the hypopharyngeal gland extracts from workers of Apis mellifera in three physiologic stages: newly emerged, nurse and forager workers, with the objective of contributing to the comprehension of the gland function. In order to determinate the enzymes present in the extracts, the Api Zym kit (Bio Merieux) was used to test the activity of 19 different enzymes. The enzymes found in larger amounts only in the hypopharyngeal glands from certain individuals were the following: in newly emerged workers, the N-acetyl-double down arrow-glucosaminidase that may be digesting the chitin of some food ingested by the bee; in forager workers, the acid phosphatase that is likely acting in authophagic processes, the a-glucosidase, in the processing of nectar into honey, and the double down arrow-glucosidases, in the pollen digestion.

Ocorrência e morfologia de glândulas tegumentares no abdome de algumas abelhas (Hymenoptera: Apidae): um estudo comparado

No presente trabalho foi estudada a ocorrência, distribuição e morfologia de glândulas tegumentares do abdome em Exomalopsis auropilosa Spinola, Centris fuscata Lepeletier, Epicharis flava Friese e Xylocopa (Neoxylocopa) suspecta Moure & Camargo. Os resultados mostraram a ocorrência de glândulas epiteliais (classe I) e unicelulares (classe III) com distribuição dorsal (tergais) e ventral (esternais) sem que se pudesse caracterizar um padrão relacionável com a posição filogenética ou grau de sociabilidade. No entanto, verificou-se uma tendência para maior número de glândulas em espécies com algum grau de sociabilidade como E. auropilosa e X. suspecta.

Análise morfológica da glândula de veneno de Apis mellifera L. (Hymenoptera: Apidae) em populações de Mato Grosso do Sul

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)