Functional morphology of adult female Culex quinquefasciatus midgut during blood digestion

The adult female Culex quinquefasciatus midgut comprises a narrow anterior and a dilated posterior region, with epithelia composed of a monolayer of adjacent epithelial cells joined at the apical portion by septate junctions. Densely packed apical microvilli and an intricate basal labyrinth characterise each cell pole. Our morphological studies suggest that, during blood digestion, the anterior midgut region also participates in an initial absorptive stage which is probably related to the intake of water, salts and other small molecules. This activity peaked by 6 h after bloodmeal feeding (ABF) and ended approximately 18 h ABF, when the peritrophic membrane was already formed. After this time, absorption only occurred in the posterior region, with morphologic and biochemical evidence of high synthetic activity related to the secretion of proteases. Chymotrypsin, elastase, aminopeptidase, and trypsin reached their maximum activity at around 36 h ABF. Digestion products were apparently absorbed and transported to the basal labyrinth, from where they should be released to the hemolymph. At 72 h ABF, proteolysis had already ended and protein levels had returned to those observed before blood meal. The epithelium of the posterior region, however, did not return to its initial morphology, appearing quite disorganised. Additionally, from 48 h ABF onwards some epithelial cells showed morphological signals of apoptosis. (C) 2002 Elsevier B.V. Ltd. All rights reserved.

Identification and characterization of excretory/secretory products of Giardia duodenalis trophozoites

One of the major questions concerning Giardia is the understanding of pathophysiological processes associated with small intestine abnormalities. There are evidences that Giardia trophozoites contain and/or release proteolytic enzymes that may be implicated in the host intestinal epithelium. The present investigation was undertaken to examine the protease activity in excretory/secretory (E/S) products of Giardia duodenalis trophozoites of an axenic Brazilian strain (BTU-11) and the reference strain Portland 1 (P1). E/S products from trophozoites of each strain in conditioned medium were tested with sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) for the protein profiles, and the protease activity was analyzed using substrate-impregnated SDS-PAGE (gelatin and collagen) and hemoglobin assay. The proteases characterization was based on inhibition assays including synthetic inhibitors. Electrophoresis analysis of E/S products revealed a banding pattern composed by few bands (4 to 6 bands) in the migration region of 123 to 28 kDa. Proteolytic products were detected in the conditioned medium by trophozoites of both assayed strains. In the gels containing copolymerized gelatin and collagen, E/S products promoted substrate degradation and the most evident proteolysis zones were distributed in the migration regions of 77 to 18 kDa and 145 to 18 kDa, respectively, in the patterns of gelatinolytic and collagenolytic activities. Degradation of hemoglobin was also observed, and the pattern of hydrolysis was similar in both E/S products assayed. Inhibitor assays showed that the main proteolytic activity in both E/S products is due to cysteine proteases, although the presence of serine proteases was also indicated. Degradation of substrates including collagen and hemoglobin could lead us to speculate different functions of Giardia excreted/secreted proteases in vivo, but to confirm this possibility and to elucidate its implication on host-parasite interactions, further experiments applying protocols for the purification of proteases are necessary. Even so, our observations are relevant and hold the perspective for the understanding about protease activity in Giardia trophozoites of axenic strain isolated in an endemic area.

Morphology, ultramorphology and histology of ovaries of workers and mated queens of Pachycondyla striata ants (Hymenoptera : Ponerinae)

The aim of the present work is to perform morphological and histological studies of the ovaries of workers and mated queens of Pachycondyla striata ants, which belong to the subfamily Ponerinae. The ovaries, after being removed, were schematized. Next, historesin and electronic scanning microscopy techniques were applied, making it possible to note that the left ovary owns a greater number of ovarioles when compared to the right one (workers - 7 to the right and 8 to the left; queens - 6 to the right and 7 to the left) and that the ovarioles of workers present a rather wrinkled aspect due to the fact that they are not developed. The same situation does not occur in mated queens once they always present oocytes in distinct development phases in their ovarioles. Histologically it was observed that the ovarioles are of the meroistic polytrophic kind. Inside the ovarioles of workers, germinative cells were observed in their distal portion, but their lumen were empty. on the other hand, ovarioles of mated queens presented the germinative cells as well as oocytes in different degrees of development, although more than one developing oocyte was never observed in the interior of each ovariole. It was possible to note the presence of follicular epithelium, chorion and vitellin membrane in oocytes of mated queens, which change morphologically according the oocyte development stage.

Histology and histochemistry of the ventriculus of Dolichoderus (=Monacis) bispinosus (OLIVIER, 1792) (Hymenoptera : Formicidae)

In this study we histologically and histochemically describe the ventriculus of Dolichoderus bispinosus. The epithelium consists of two basic cell types, highly basophilic generative cells, and digestive cells. The latter present several cytoplasmic vesicles. rich in acidic and neutral polysaccharides, and basic proteins. Also, these. cells exhibit an apocrine secretion pattern. A mass of fibrous material is observed on the surface of the epithelium. Finally, we discuss the results obtained. (c) 2005 Elsevier Ltd. All rights reserved.

Venom gland of Pachycondyla striata worker ants (Hymenoptera : Ponerinae). Ultrastructural characterization

Morphological data concerning the venom gland of worker ants of Pachycondyla striata revealed that this gland consists of three distinct regions: an external secretory portion, composed by a secretory filament that bifurcates in order to give rise to other two filaments; an internal secretory portion, represented by the convoluted gland; and a storage portion, represented by a sac-shaped reservoir. The ultrastructural analysis showed that the reservoir is enveloped by a simple pavementous epithelium, coated internally with a cuticle. The external secretory portion is composed by cells forming a simple cubic epithelium, in which the apical portion presents numerous microvilli while the basal portion of the cells shows infoldings of the plasma membrane containing numerous mitochondria. The convoluted gland possesses cells of irregular morphology with nuclei containing condensed chromatin, suggesting inactivity. However, these cells are in fact undergoing secretory activity, which is probably added to the final secretion produced by the gland. The cytoplasm of these cells contains several elements distributed therein, such as ribosomes and polyribosomes, lipid droplets, and protein inclusions in the form of crystals, thus Suggestive of protein storage, which would be used by the insect when metabolically required. (c) 2005 Elsevier Ltd. All rights reserved.

Morphohistological study of the venom gland in workers of the ant Pachycondyla striata F. Smith (Hymenoptera : Ponerinae)

Themorphology of the venom gland in workers of the ant Pachycondyla striata (F. Smith) (Hymenoptera: Ponerinae) consists of an elongated sac that is directly connected to the sting apparatus. Three distinct regions compose this gland: the external secretory portion, composed by a secretory filament that bifurcates to originate another two; the internal secretory portion, which is represented by the convoluted gland from which rises the excretory duct that liberates the venom; and the storage portion, consisting of a large sac-shaped reservoir. The histology showed that the gland possesses a strong musculature on its distal third. Underneath these muscle layers, we noted the presence of an epithelium that envelops the internal wall of the reservoir. The presence of a convoluted gland as well as secretion inside the reservoir was also noted.

Ultrastructure of the ducts of the reproductive tract of males of Melipona bicolor bicolor Lepeletier (Hymenoptera, Apinae, Meliponini)

The present paper describes the ultrastructural features of seminal vesicle, post-vesicular vas deferens and ejaculatory duct of Melipona bicolor bicolor from newly emerged and mature males. Although the results do not show very consistent morphological signs of secretory activity by the epithelium of these organs, lipidic droplets and lamellar granules present in mature males' seminal vesicles and the vacuoles present in post-vesicular vas deferens are probably secretion. Besides, the spermatozoa in the lumen are immersed in a material of characteristic structure, which must be produced in superior regions of the reproductive system of immature males, not studied here. The presence of sperm cells, apparently in cytoplasm vesicles of seminal vesicle and post-vesicular vas deferens, suggests spermiophagy by their epithelium.

Ultrastructural studies of the mandibular glands of the minima, media and soldier ants of Atta sexdens rubropilosa (Forel 1908)

The mandibular glands of Hymenoptera are structures associated with the mandibles and constitute part of the salivary glands system. Histological studies in workers of Atta sexdens rubropilosa revealed that this gland contains two portions: a secretory and a storage portion or reservoir. Both portions are connected by means of canaliculi. The object of the present work was the study of the ultratructure of the mandibular glands of minima, media and soldier ant of A. s. rubropilosa by TEM techniques. The glands, in the three castes studied, possess a reservoir, constituted by a simple pavementous epithelium surrounded by the cuticular intima and the secretory portion is constituted by cells of rounded shape. The secretory cells, mainly of minima and soldier, were rich in smooth endoplasmic reticulum. The media worker and soldier presented a large number of mitochondria, of varying shape. Well-developed Golgi complexes were also present in the soldiers. The secretory cells in minima, media and soldier were provided with collecting intracellular canaliculi, which were linked to the reservoir through the extracellular portion. The cytoplasm of the canaliculi-forming cell was poor in organelles. In the individuals of the three castes of A. s. rubropilosa, the presence of lipid secretion granules suggested, beyond the other functions, also a possible pheromonal action. The different roles executed by the different insect castes are directly dependent on the glandular products and, consequently, on the secretory cellular characteristics. (c) 2005 Elsevier Ltd. All rights reserved.

The intramandibular gland of leaf-cutting ants (Atta sexdens rubropilosa Forel 1908)

The eusociality developed in Hymenoptera and Isoptera is driven by an efficient interaction between exocrine glands and jointed appendages, both in close interaction with the environment. In this context, the mandible of ants plays an important role, since, in addition to being the main jointed appendage, it possess glandular functions. As an example we might name the two glands associated with the mandible: the mandibular and the intramandibular glands. The intramandibular gland is found inside the mandible and consists of a hypertrophied secretory epithelium and secretory cells in the mandible's lumen. The secretion of the secretory epithelium is liberated through intracuticular ducts that open at the base of hairs at the mandible's surface. The secretion of the intramandibular gland (epithelium and secretory cells) reacted positively to tests for the detection of polysaccharides and proteins, thus suggesting that it consists of glycoproteins. The ultrastructure of the secretor epithelium presents variations related to the developmental stage of the individual, showing a large number of ribosomes and microvilli close to the cuticle in young individuals, while in the older specimens it was possible to note the formation of ail intracellular reservoir. These variations of secretory epithelium, as also the interaction between the cellular groups inside the mandible, are important information about this gland in leaf-cutting ants. (c) 2005 Elsevier Ltd. All rights reserved.

Ultrastructural features of the regenerative cells of the bees (Hymenoptera, Apidae) midguts

The epithelium of the bee ventriculus is formed by two cell types: the principal or digestive cells and the regenerative cells. In this article the ultrastructure of the regenerative cells is described, as well as the features of their differentiation into digestive cells during epithelium renewal.

Lipid distribution in salivary glands of larvae and adult bees (Hymenoptera : Apidae)

Cytochemical studies were carried out to establish lipid distribution in the salivary glands of larvae and adult bees, using the imidazole buffer technique. In the duct cells of the larval salivary gland, the reaction was positive in the epicuticle and negative in the glandular lumen. The absence of smooth endoplasmic reticulum and the presence of lipids in the intercellular space suggest that lipids absorbed from the haemolymph could be used in the constitution of the epicuticle, after having been conveyed through the epithelium. In adult workers (new-emerged, nurse and forager workers), the head salivary glands presented a positive reaction in the secretion in glandular lumen, identifying its lipidic nature.

Programmed cell death in the larval salivary glands of Apis mellifera (Hymenoptera, Apidae)

The morphological and histochemical features of degeneration in honeybee (Apis mellifera) salivary glands were investigated in 5th instar larvae and in the pre-pupal period. The distribution and activity patterns of acid phosphatase enzyme were also analysed. As a routine, the larval salivary glands were fixed and processed for light microscopy and transmission electron microscopy. Tissue sections were subsequently stained with haematoxylin-eosin, bromophenol blue, silver, or a variant of the critical electrolyte concentration (CEC) method. Ultrathin sections were contrasted with uranyl acetate and lead citrate. Glands were processed for the histochemical and cytochemical localization of acid phosphatase, as well as biochemical assay to detect its activity pattern. Acid phosphatase activity was histochemically detected in all the salivary glands analysed. The cytochemical results showed acid phosphatase in vesicles, Golgi apparatus and lysosomes during the secretory phase and, additionally, in autophagic structures and luminal secretion during the degenerative phase. These findings were in agreement with the biochemical assay. At the end of the 5th instar, the glandular cells had a vacuolated cytoplasm and pyknotic nuclei, and epithelial cells were shed into the glandular lumen. The transition phase from the 5th instar to the pre-pupal period was characterized by intense vacuolation of the basal cytoplasm and release of parts of the cytoplasm into the lumen by apical blebbing; these blebs contained cytoplasmic RNA, rough endoplasmic reticule and, occasionally, nuclear material. In the pre-pupal phase, the glandular epithelium showed progressive degeneration so that at the end of this phase only nuclei and remnants of the cytoplasm were observed. The nuclei were pyknotic, with peripheral chromatin and blebs. The gland remained in the haemolymph and was recycled during metamorphosis. The programmed cell death in this gland represented a morphological form intermediate between apoptosis and autophagy.

Ultramorphology and histology of the larval salivary gland of Pachycondyla villosa (Fabricius) (Hymenoptera: Formicidae, Ponerinae)

A glândula salivar apresenta-se com um duto anterior único, formado por um epitélio colunar, dois dutos laterais curtos, os quais apresentam-se com epitélio cúbico simples e que na sua porção mais proximal torna-se colunar. Posterior a estes, encontram-se os dois reservatórios, os quais possuem o epitélio bastante delgado e é neste reservatório que a região secretora da glândula se abre. Os ramos dorsal e ventral da região secretora da glândula conectam-se por meio de comissuras transversais, sendo que, posteriormente, a região secretora termina em forma de alça. A região secretora é uniforme, não apresenta tipos celulares distintos e é formada por um epitélio cúbico simples. Neste trabalho é apresentada, também, a revisão sobre a morfologia da glândula salivar larval em insetos, principalmente com relação aos Hymenoptera-Aculeata.

Tegumentary epithelial glands in the abdomen of virgin and physogastric queens of the stingless bee Scaptotrigona postica Latreille (Meliponini: Trigonina)

Nas operárias das abelhas eussociais, regiões do epitélio tegumentar do abdome podem hipertrofiar-se e tornarem-se glandulares, sendo responsáveis pela produção de cera usada na contrução dos alvéolos de cria. Nos meliponíneos, epitélio com essas características também foi observado nas rainhas, porém sua função ainda continua desconhecida. Teria esse epitélio função homóloga nas operárias? As rainhas, além da função reprodutiva, desempenhariam funções na colônia até então consideradas exclusivas das operárias? Para tentar colaborar no esclarecimento dessas questões realizou-se um estudo histológico e ultra-estrutural das glândulas epiteliais tegumentares do terceiro tergito em rainhas virgens e fisogástricas de Scaptotrigona postica Latreille. Os resultados morfológicos mostraram que o epitélio glandular existe e é facilmente discernível no terceiro tergito das rainhas, sendo muito mais desenvolvido nas fisogástricas do que nas virgens. A ultra-estrutura mostrou que há diferenças no tipo de organização do retículo endoplasmático liso entre as rainhas, o que, juntamente com os resultados da histologia e morfometria, indicam poder haver diferenças funcionais desse epitélio entre elas. Como já observado na literatura, nas rainhas virgens tal epitélio pode estar envolvido na produção de cera e confecção de alvéolos de cria, porém sua função nas rainhas fisogástricas é totalmente desconhecida. A hipótese lançada no presente estudo é de que esse epitélio pode estar envolvido na produção de feromônios que irão atuar nas interações sociais da rainha poedeira.

Occurrence, morphology and ultrastructure of the Dufour gland in Melipona bicolor Lepeletier (Hymenoptera, Meliponini)

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