Contribution of the Malpighian tubules for the maintenance of symbiotic microorganisms in cephalotes ants

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Morphological characterization of the nymphs rhipicephalus sanguineus ticks (Latreille, 1806) (Acari: Ixodidae). Description of the testes, integument, malpighian tubules, and midgut on the detachment day

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

Morphometric changes associated with sex and development in the Malpighian tubules of Aedes aegypti

The Malpighian tubules of Aedes aegypti showed significant differences in their diameters between male and female larvae, male and female pupae, male larvae and male adults and male pupae and male adults. in every case, female values were greater than in males. Measurements of mean nuclear areas of the principal and stellate cells from Malpighian tubules, taken in males and females during development, showed that this parameter in both types of cell was significantly greater in females than in male larvae, pupae and adult stages. In males, significant differences between developmental stages were observed only in comparison with the nuclear area of larvae and adults in the principal cells, but in females, every comparison between stages showed significant differences except between pupae and adults in stellate cells. The frequency distribution of nuclear area values, in development, for male stellate and principal cells, were mostly concentrated in the first seven classes among the 30 classes considered in every stage, while for females, the frequency dropped drastically in the same classes from larvae to pupae and adults, moving to classes of higher values. Considering the importance of Malpighian tubules in insect physiology, the meaning of the differences detected are discussed on the basis of different metabolic levels, between sexes and developmental stages.

Peroxidase activity in Malpighian tubules of Triatoma infestans Klug

Benzidine and diamino benzidine (DAB) oxidation, typically performed by peroxidases, was demonstrated by light and electron microscopy in peroxisomes, mitochondria and membranous structures which occurred in close contact with urate crystals in Malpighian tubules of nymphs and adults of Triatoma infestans. Peroxisomes were predominantly identified in cells of the distal region of the tubules, which is engaged in excretory mechanisms. DAB oxidation in mitochondria, even in the absence of hydrogen peroxide, may indicate the existence of a mitochondrial peroxidase and possibly a cytochrome c peroxidase. The localization of the extracellular membranous structures appeared restricted to the lumen of the proximal region of the tubules and they were assumed to be remnants of endoplasmic reticulum containing peroxidases.

Mg2+-dependent ATPase activity in Malpighian tubules of Triatoma infestans Klug

Mg2+-dependent ATPases were investigated in Malpighian tubules of the blood-sucking insect, Triatoma infestans, with cytochemical procedures for light and electron microscopy. The aim was to establish patterns of enzyme occurrence in the blood-sucking insect under control rearing conditions for further comparisons with animals subjected to the action of stress factors. Enzyme activity was found in laminated "concretions" present in distal cells, in edges of urate crystals at the lumen of the proximal region of tubules, in the basement membrane of proximal cells, and variously distributed in plasmalemma invaginations of both distal and proximal cells. Presence of ATPases in the "concretions" and urate crystals is presumed to be due to engulfment of other ATPase-containing components during formation of these structures. Cytochemical reactivity in the basement membrane and plasmalemma invaginations is assumed to be involved with active transport of waste molecules from and to hemolymph and differs as a function of the Malpighian tubule region. This paper provides a basic understanding of the enzyme occurrence in the blood sucking insects, and can be used as a pattern for comparative means of the staining patterns among Triatominae species. (C) 2011 Elsevier Ltd. All rights reserved.

Acid phosphatase activity in Malpighian tubules of Triatoma infestans Klug

Acid phosphatase activity was detected in the Malpighian tubules of the bloodsucking hemipteran, Triatoma infestans. The enzyme activity was especially prominent in the cytoplasmic globules which were assumed to be laminated 'concretions', which occur in the distal cells of the organ. It was also verified in the nuclei and in some cytoplasmic granules (lysosomes) of the proximal cells. The data indicated that lysosomes were involved with the nature or origin of the laminated concretions, but it is still questionable whether acid phosphatase activity exists in the nuclei.

Ultrastructural aspects of acid phosphatase activity in Malpighian tubules of Triatoma infestans Klug

Acid phosphatase activity was investigated ultrastructurally in Malpighian tubules of Triatoma infestans. Enzyme activity was demonstrated in laminated 'concretions' (distal cells) and in typical lysosomes, as well as in basal plasmalemma infoldings and basement membranes (especially in distal cells). This activity was assumed to be related to the excretory functions carried out mostly by the distal cells. Heterochromatin-nucleolus functional relationships involving RNA transcription may promote the nuclear reaction verified in the proximal cells and in some distal cells. A lead phosphate precipitate appeared free in the cytoplasm encircling the nuclei and was assumed to be a contamination from the nuclear precipitates.

Segmental specializations in the Malpighian tubules of the fire ant Solenopsis saevissima Forel 1904 (Myrmicinae): an electron microscopical study

The Malpighian tubules of workers of the fire ant Solenopsis saevissima (Myrmicinae) were analyzed by scanning and transmission electron microscopy in order to determine their functional organization and association with the hindgut epithelium. The ants showed six Malpighian tubules with three segments morphologically and structurally different. The proximal segment was long and its cells showed abundant smooth endoplasmic reticulum and lipid droplets, which suggest their role in lipid secretion. The mid segment was long and undulated and it was composed by the cells that showed the typical features of ion transporting epithelia. The distal segment, short and flattened, adheres to the rectum wall. The cells of this segment showed the basal lamina fused to that of the rectum, it is probable that this part of the tubule may play a role in ion and water uptake from the feces. (C) 2002 Elsevier B.V. Ltd. All rights reserved.

Nucleolar changes in malpighian tubules cells of Rhodnius prolixus during starvation

This study focused on nuclear and nucleolar area changes in Malpighian tubule cells of Rhodnius prolixus, an hematophagous insect, vector of Chagas' disease. Male and female adult insects were dissected after a 28-day starvation period, as well as insects which had been fed again after a 30-day starvation period. Malpighian tubules were fixed and silver stained. In both, males and females, nucleolar fusions and regions of nucleolar corpuscles were observed to be differentially impregnated by silver during feeding stress. In the males and females that were fed again, nucleolar corpuscles were partially fusioned, indicating a slight recovery upon the 30-day starvation period. The changes observed in the nucleolar phenotype and in both nuclear and nucleolar areas indicated that, as a result of stress, a more intense, compensatory activity occurred to supply the metabolic rate. The mechanism of this phenomenon may be associated to the decondensation and activation of chromatin that carries rDNA in order to increase rRNA transcription, and,consequently, protein synthesis.

Cellular responses in the Malpighian tubules of Scaptotrigona postica (Latreille, 1807) exposed to low doses of fipronil and boric acid

Studies of sub-lethal effects of pesticide residues on stingless bees are scarce and morphological analysis of organs would add information to toxicological analysis in order to clarify the continuous exposure of Scaptotrigona postica to insecticides. The aim of this study was to evaluate the morphology and histochemistry of the Malpighian tubules (excretory organ) of S. postica exposed to fipronil or boric acid to detect cellular responses that indicate toxicity or adaptative mechanisms to stress induced by exposure of worker bees to low doses of these chemical compounds. Newly emerged bees were submitted to toxicological bioassays and morphological analyses by optical microscopy and Transmission Electron Microscopy, as well as histochemical methods, were performed to detect proteins and glycoconjugates. Additionally, immunohistochemical detection of DNA fragmentation and HSP70 (70-kDa Heat shock protein) were performed to detect cell death and stress response, respectively. Statistical analysis, for the bioassays conducted with ingestion of contaminated diet with boric acid at 0.75% (w/w) or with fipronil at 0.1μg/kg of food, showed that the survival of bees that ingested the contaminated diets were significantly different to the survival rate presented by the control group (P<0.0001). Although some characteristics indicative of initiation of cell death were observed, the cells remained metabolically active in the processes of excretion and inactivation of chemical compounds. The data from this study reinforce the importance of research on sublethal effects of low doses of pesticides on bees in an attempt to assess a possible realistic dose and evaluate the risk assessment of stingless bee S. postica foraging in the vicinity of cultivated fields and/or in green urban areas. © 2013 Elsevier Ltd.

Effects of sublethal doses of imidacloprid in malpighian tubules of africanized Apis mellifera (Hymenoptera, Apidae)

In Brazil, imidacloprid is a widely used insecticide on agriculture and can harm bees, which are important pollinators. The active ingredient imidacloprid has action on the nervous system of the insects. However, little has been studied about the actions of the insecticide on nontarget organs of insects, such as the Malpighian tubules that make up the excretory and osmoregulatory system. Hence, in this study, we evaluated the effects of chronic exposure to sublethal doses of imidacloprid in Malpighian tubules of Africanized Apis mellifera. In the tubules of treated bees, we found an increase in the number of cells with picnotic nuclei, the lost of part of the cell into the lumen, and a homogenization of coloring cytoplasm. Furthermore, we observed the presence of cytoplasmic vacuolization. We confirmed the increased occurrence of picnotic nuclei by using the Feulgan reaction, which showed the chromatin compaction was more intense in the tubules of bees exposed to the insecticide. We observed an intensification of the staining of the nucleus with Xylidine Ponceau, further verifying the cytoplasmic negative regions that may indicate autophagic activity. Additionally, immunocytochemistry experiments showed TUNEL positive nuclei in exposed bees, implicating increased cell apoptosis after chronic imidacloprid exposure. In conclusion, our results indicate that very low concentrations of imidacloprid lead to cytotoxic activity in the Malpighian tubules of exposed bees at all tested times for exposure and imply that this insecticide can alter honey bee physiology. © 2013 Wiley Periodicals, Inc.

Cytotoxic Effects of Thiamethoxam in the Midgut and Malpighian Tubules of Africanized Apis mellifera (Hymenoptera: Apidae)

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

Metabolism of an insect diuretic hormone by Malpighian tubules studied by liquid chromatography coupled with electrospray ionization mass spectrometry

The larger of two diuretic hormones of the tobacco hornworm, Manduca sexta, (Mas-DH) is a peptide of 41 residues. It is one of a family of seven currently known insect diuretic hormones that are similar to the corticotropin-releasing factor–urotensin–sauvagine family of peptides. We investigated the possible inactivation of Mas-DH by incubating it in vitro with larval Malpighian tubules (Mt), the target organ of the hormone. The medium was analyzed, and degradation products were identified, using on-line microbore reversed-phase liquid chromatography coupled to electrospray ionization mass spectrometry (RPLC-ESI-MS). This sensitive technique allows identification of metabolites of Mas-DH (present at an initial level of ≈1 μM). An accurate Mr value for a metabolite is usually sufficient for unambiguous identification. Mas-DH is cleaved by Mt proteases initially at L29–R30 and R30–A31 under our assay conditions; some Mas-DH is also oxidized, apparently at M2 and M11. The proteolysis can be inhibited by 5 mM EDTA, suggesting that divalent metals are needed for peptide cleavage. The oxidation of the hormone can be inhibited by catalase or 1 mM methionine, indicating that H2O2 or related reactive oxygen species are responsible for the oxidative degradation observed. RPLC-ESI-MS is shown here to be an elegant and efficient method for studying peptide hormone metabolism resulting from unknown proteases and pathways.

Review: Malpighian Tubule, an essential organ for insects

We did an overhaul on morphophysiology characteristics, cryptonephric system and secondary specializations in the Malpighian tubules of the class Insecta. This review grouped the most important information about this important organ that has different functions essential for insects.

Avaliação da concentração de Cl, K e Ca na urina, hemolinfa e túbulos de Malpighi de Rhodnius prolixus usando a técnica de fluorescência de raios X por reflexão total por radiação síncrotron (SR-TXRF)

Neste trabalho utilizou-se a técnica fluorescência de raios X usando radiação síncrotron (SR-TXRF) para estudar, quantitativamente, o transporte de cloro, potássio e cálcio na hemolinfa, urina e túbulos de Malpighi (TM) em ninfas de quinto estágio do Rhodnius prolixus (R. prolixus), considerando a excreção destes elementos em diferentes dias após o repasto sanguíneo. R. prolixus é um dos principais vetores do Trypanosoma cruzi, agente causador da doença de Chagas. R. prolixus fornece um sistema modelo particularmente útil porque seus TMs tanto secretam quanto reabsorvem íons a taxas elevadas. Os TMs filtram a hemolinfa e secretam um líquido que é muitas vezes comparado com a urina primária em vertebrados. Os resultados obtidos mostram que a concentração de potássio na urina é substancialmente maior do que na hemolinfa. A concentração de cloro na hemolinfa é menor do que na urina, mas a diferença não é tão marcada como no caso do potássio. No caso do Rhodnius é razoável interpretar a elevada concentração de potássio na urina como adaptativo para o problema de excreção imediato do inseto. A concentração de cálcio nos TMs é substancialmente maior em comparação com os valores encontrados na hemolinfa e urina. Este resultado mostra que o cálcio é retido no corpo do R. prolixus e pouco eliminado. Os resultados obtidos estão coerentes com a literatura. Avaliou-se também o efeito no transporte de Cl, K e Ca após um repasto de sangue de coelho contaminado com HgCl2 de modo a avaliar o efeito da presença deste metal tóxico no balanço iônico nos fluidos de excreção urina e hemolinfa e também pelo principal órgão de transporte, os túbulos de Malpighi. As excreções de Cl e K pela urina são afetadas pela ingestão. Este resultado é esperado levando-se em consideração a ingestão de excesso de Cl através do HgCl2. O transporte de Cl, K e Ca na hemolinfa do Rhodnius prolixus não é afetada pela ingestão de HgCl2. Nos túbulos de Malpighi, as altas concentrações de Ca obtidas foram comparáveis àquelas encontradas nos insetos controle. Pode-se concluir que SR-TXRF é um método muito promissor para análises diretas, rápidas e confiáveis para a quantificação simultânea de elementos envolvidos na regulação do transporte e em todo o sistema excretor de insetos. Além disso, o estudo do transporte e a excreção de elementos no inseto Rhodnius prolixus abrem oportunidade para a maior compreensão de efeitos da poluição em espécies de invertebrados.