The Fat Body of the Hematophagous Insect, Panstrongylus megistus (Hemiptera: Reduviidae): Histological Features and Participation of the ß-Chain of ATP Synthase in the Lipophorin-Mediated Lipid Transfer.

In insects, lipid transfer to the tissues is mediated by lipophorin, the major circulating lipoprotein, mainly through a nonendocytic pathway involving docking receptors. Currently, the role of such receptors in lipid metabolism remains poorly understood. In this work, we performed a histological characterization of the fat body of the Chagas' disease vector, Panstrongylus megistus (Burmeister), subjected to different nutritional conditions. In addition, we addressed the role of the ß-chain of ATP synthase (ß-ATPase) in the process of lipid transfer from lipophorin to the fat body. Fifth-instar nymphs in either fasting or fed condition were employed in the assays. Histological examination revealed that the fat body was composed by diverse trophocyte phenotypes. In the fasting condition, the cells were smaller and presented a homogeneous cytoplasmic content. The fat body of fed insects increased in size mainly due to the enlargement of lipid stores. In this condition, trophocytes contained abundant lipid droplets, and the rough endoplasmic reticulum was highly developed and mitochondria appeared elongated. Immunofluorescence assays showed that the ß-ATPase, a putative lipophorin receptor, was located on the surface of fat body cells colocalizing partially with lipophorin, which suggests their interaction. No changes in ß-ATPase expression were found in fasting and fed insects. Blocking the lipophorin-ß-ATPase interaction impaired the lipophorin-mediated lipid transfer to the fat body. The results showed that the nutritional status of the insect influenced the morphohistological features of the tissue. Besides, these findings suggest that ß-ATPase functions as a lipophorin docking receptor in the fat body.

The modern morphometric approach to identify eggs of Triatominae.

BACKGROUND: Egg morphometrics in the Triatominae has proved to be informative for distinguishing tribes or genera, and has been based generally on traditional morphometrics. However, more resolution is required, allowing species or even population recognition, because the presence of eggs in the domicile could be related to the species ability to colonize human dwellings, suggesting its importance as a vector. RESULTS: We explored the resolution of modern morphometric methods to distinguish not only tribes and genera, but also species or geographic populations in some important Triatominae. Four species were considered, representing two tribes and three genera: Panstrongylus chinai and P. howardi, Triatoma carrioni and Rhodnius ecuadoriensis. Within R. ecuadoriensis, two geographical populations of Ecuador were compared. For these comparisons, we selected the most suitable day of egg development, as well as the possible best position of the egg for data capture. The shape of the eggs in the Triatominae does not offer true anatomical landmarks as the ones used in landmark-based morphometrics, except for the egg cap, especially in eggs with an evident "neck", such as those of the Rhodniini. To capture the operculum shape variation, we used the landmark- and semilandmark-based method. The results obtained from the metric properties of the operculum were compared with the ones provided by the simple contour of the whole egg, as analyzed by the Elliptic Fourier Analysis. Clear differences could be disclosed between the genera, between the species - among which two very close species (P. chinai and P. howardi), as well as between two allopatric, conspecific populations. The whole egg contour (including the operculum) produced reclassification scores much more satisfactory than the ones obtained using the operculum only. CONCLUSIONS: We propose the outline-based approach as the most convenient characterization tool to identify unknown eggs at the species or population levels.

Contribution of AT-, GC-, and methylated cytidine-rich DNA to chromatin composition in Malpighian tubule cell nuclei of Panstrongylus megistus (Hemiptera, Reduviidae).

The Malpighian tubule cell nuclei of male Panstrongylus megistus, a vector of Chagas disease, contain one chromocenter, which is composed solely of the Y chromosome. Considering that different chromosomes contribute to the composition of chromocenters in different triatomini species, the aim of this study was to determine the contribution of AT-, GC-, and methylated cytidine-rich DNA in the chromocenter as well as in euchromatin of Malpighian tubule cell nuclei of P. megistus in comparison with published data for Triatoma infestans. Staining with 4',6-diamidino-2-phenylindole/actinomycin D and chromomycin A(3)/distamycin, immunodetection of 5-methylcytidine and AgNOR test were used. The results revealed AT-rich/GC-poor DNA in the male chromocenter, but equally distributed AT and GC DNA sequences in male and female euchromatin, like in T. infestans. Accumulation of argyrophilic proteins encircling the chromocenter did not always correlate with that of GC-rich DNA. Methylated DNA identified by immunodetection was found sparsely distributed in the euchromatin of both sexes and at some points around the chromocenter edge, but it could not be considered responsible for chromatin condensation in the chromocenter, like in T. infestans. However, unlike in T. infestans, no correlation between the chromocenter AT-rich DNA and nucleolus organizing region (NOR) DNA was found in P. megistus.

Prevalence and intensity of infection, metacyclogenesis and nuclear phenotypes in Panstrongylus megistus (Burmeister, 1835) after ingestion of Trypanosoma cruzi (Chagas, 1909) II and subjection to heat shock.

This study aimed to contribute to our knowledge of the parasite-vector interaction associated with Trypanosoma cruzi (Chagas, 1909) infection in Panstrongylus megistus (Burmeister, 1835), an important vector of Chagas' disease in Brazil. The prevalence and intensity of T. cruzi infection, the incidence of metacyclogenesis and the frequency of nuclear phenotypes in Malpighian tubules were investigated in nymphs of P. megistus, reared at 28 °C and subjected to heat shock (40 °C, 1 hour) two days after infection with T. cruzi II (Y strain). Following the 45-day post-infection period, the frequency of epimastigotes was much higher than that of trypomastigotes in both heat-shocked and non-shocked insects, and the prevalence of infection was not altered by heat shock. Fewer epimastigotes and trypomastigotes were found in the infected insects subjected to the heat shock, indicating that the multiplication and metacyclogenesis of the parasites were affected by the stress. In infected specimens heat shock promoted an increased frequency of cell nuclei with heterochromatin decondensation, a cell survival response to stress, and did not affect insect survival. The effects of infection and heat shock, especially on the multiplication and metacyclogenesis of T. cruzi, and the observed resistance to heat shock developed by P. megistus nymphs are suggestive that they should be considered when adequate conditions for rearing these infected insects in the laboratory are pursued.

Prevalence and intensity of infection, metacyclogenesis and nuclear phenotypes in Panstrongylus megistus (Burmeister, 1835) after ingestion of Trypanosoma cruzi (Chagas, 1909) II and subjection to heat shock / Prevalência e intensidade de infecção, metaciclogênese e fenótipos nucleares em Panstrongylus megistus (Burmeister, 1835) após ingestão de Trypanosoma cruzi (Chagas, 1909) II e choque de temperatura

This study aimed to contribute to our knowledge of the parasite-vector interaction associated with Trypanosoma cruzi (Chagas, 1909) infection in Panstrongylus megistus (Burmeister, 1835), an important vector of Chagas' disease in Brazil. The prevalence and intensity of T. cruzi infection, the incidence of metacyclogenesis and the frequency of nuclear phenotypes in Malpighian tubules were investigated in nymphs of P. megistus, reared at 28 °C and subjected to heat shock (40 °C, 1 hour) two days after infection with T. cruzi II (Y strain). Following the 45-day post-infection period, the frequency of epimastigotes was much higher than that of trypomastigotes in both heat-shocked and non-shocked insects, and the prevalence of infection was not altered by heat shock. Fewer epimastigotes and trypomastigotes were found in the infected insects subjected to the heat shock, indicating that the multiplication and metacyclogenesis of the parasites were affected by the stress. In infected specimens heat shock promoted an increased frequency of cell nuclei with heterochromatin decondensation, a cell survival response to stress, and did not affect insect survival. The effects of infection and heat shock, especially on the multiplication and metacyclogenesis of T. cruzi, and the observed resistance to heat shock developed by P. megistus nymphs are suggestive that they should be considered when adequate conditions for rearing these infected insects in the laboratory are pursued.

O presente estudo teve como objetivo contribuir para o conhecimento sobre a interação parasita-vetor associada à infecção por Trypanosoma cruzi (Chagas, 1909) em Panstrongylus megistus (Burmeister, 1835), importante vetor da doença de Chagas no Brasil. A prevalência e a intensidade da infecção por T. cruzi, a incidência de metaciclogênese e a frequência de fenótipos nucleares presentes em túbulos de Malpighi foram investigadas em ninfas de P. megistus criadas à temperatura de 28 °C e submetidas a choque térmico por 1 hora a 40 °C, dois dias após alimentação em camundongos infectados com T. cruzi II (linhagem Y). A análise realizada 45 dias pós-infecção revelou que tanto nos insetos submetidos ao choque térmico como nos respectivos controles a frequência de epimastigotos se apresentava muito maior do que a dos tripomastigotos; a prevalência da infecção não foi alterada pelo choque térmico. Menos epimastigotos e tripomastigotos foram encontrados nos insetos submetidos ao choque térmico, indicando que a multiplicação e a metaciclogênese dos parasitas possam ser afetadas com o estresse. Nos espécimes infectados, o choque térmico induziu aumento na frequência de núcleos com descompactação da heterocromatina, uma resposta de sobrevivência celular ao estresse, e não afetou a sobrevivência propriamente dita do inseto. Os efeitos da infecção e do choque térmico, especialmente sobre a multiplicação e a metaciclogênese de T. cruzi, e a resistência ao choque térmico desenvolvidos pelas ninfas de P. megistus são indicativos de que devam ser considerados quando se buscam condições adequadas de criação em laboratório de insetos infectados.

Estudo morfológico e ultraestrutural da glândula salivar de Rhodnius prolixus, Panstrongylus megistus e Triatoma infestans (Hemiptera, Triatominae)

Os triatomíneos são vetores do Trypanosoma cruzi e do Trypanosoma rangeli. Apresentam hábito hematófago durante todo o ciclo de vida. Durante o repasto, secretam uma saliva que antagoniza os sistemas hemostático, inflamatório e imunológico do hospedeiro vertebrado. A fim de caracterizar os aspectos morfológicos da glândula salivar, foi desenvolvido um estudo do crescimento desse órgão em triatomíneos de três espécies: Rhodnius prolixus, Panstrongylus megistus e Triatoma infestans. Os insetos foram cedidos pelo insetário do Laboratório de Triatomíneos e Epidemiologia da Doença de Chagas (CPqRR - FIOCRUZ). As glândulas salivares foram dissecadas e processadas para microscopia de luz, de fluorescência, laser confocal e eletrônica de varredura. Os resultados indicaram que a glândula é morfologicamente semelhante durante todo o ciclo de vida do inseto, sendo formada por um par de lobos, cada qual formado por dois (R. prolixus) ou três lóbulos (P. megistus e T. infestans). Durante o ciclo de vida, a glândula quadruplicou o seu tamanho inicial, devido ao crescimento celular, visto que o número de células permaneceu constante.

Os lóbulos se mostraram formados por um epitélio simples que delimita o lúmen (onde a saliva é armazenada), revestidos por fibras musculares (cuja contração permite a liberação da saliva), traquéias e nervos. Serotonina apresentou intensa atividade na glândula das três espécies analisadas durante alimentação. Em R. prolixus, atividade dopaminérgia e de tirosina-hidroxilase (enzima da via das catecolaminas) também foram observadas. Dobras de membrana foram visualizadas na região apical das células e associadas ao aumento da área de secreção (tipo merócrina). Além disso, a presença de vesículas de secreção e células inteiras (estas observadas apenas em R. prolixus) no lúmen indicaram secreções do tipo apócrina e holócrina, respectivamente. Bactérias Gram-negativas e Gram-positivas foram observadas dentro das células e do lúmen do lóbulo posterior de T. infestans

Estudo morfológico e ultraestrutural da glândula salivar de Rhodnius prolixus, Panstrongylus megistus e Triatoma infestans (Hemiptera, Triatominae) / Morphological and ultrastructural study of salivary gland of Rhodnius prolixus and Triatoma infestans Panstrongylus megistus (Hemiptera, Triatominae)

Os triatomíneos são vetores do Trypanosoma cruzi e do Trypanosoma rangeli. Apresentam hábito hematófago durante todo o ciclo de vida. Durante o repasto, secretam uma saliva que antagoniza os sistemas hemostático, inflamatório e imunológico do hospedeiro vertebrado. A fim de caracterizar os aspectos morfológicos da glândula salivar, foi desenvolvido um estudo do crescimento desse órgão em triatomíneos de três espécies: Rhodnius prolixus, Panstrongylus megistus e Triatoma infestans. Os insetos foram cedidos pelo insetário do Laboratório de Triatomíneos e Epidemiologia da Doença de Chagas (CPqRR - FIOCRUZ). As glândulas salivares foram dissecadas e processadas para microscopia de luz, de fluorescência, laser confocal e eletrônica de varredura. Os resultados indicaram que a glândula é morfologicamente semelhante durante todo o ciclo de vida do inseto, sendo formada por um par de lobos, cada qual formado por dois (R. prolixus) ou três lóbulos (P. megistus e T. infestans). Durante o ciclo de vida, a glândula quadruplicou o seu tamanho inicial, devido ao crescimento celular, visto que o número de células permaneceu constante.

Os lóbulos se mostraram formados por um epitélio simples que delimita o lúmen (onde a saliva é armazenada), revestidos por fibras musculares (cuja contração permite a liberação da saliva), traquéias e nervos. Serotonina apresentou intensa atividade na glândula das três espécies analisadas durante alimentação. Em R. prolixus, atividade dopaminérgia e de tirosina-hidroxilase (enzima da via das catecolaminas) também foram observadas. Dobras de membrana foram visualizadas na região apical das células e associadas ao aumento da área de secreção (tipo merócrina). Além disso, a presença de vesículas de secreção e células inteiras (estas observadas apenas em R. prolixus) no lúmen indicaram secreções do tipo apócrina e holócrina, respectivamente. Bactérias Gram-negativas e Gram-positivas foram observadas dentro das células e do lúmen do lóbulo posterior de T. infestans

[Histophysiology of the neurosecretory cells of the pars intercerebralis of larval female Panstrongylus megistus (Heteroptera: Reduviidae)]. / Etude histophysiologique des cellules neurosécrétrices de la pars intercerebralis de larves femelles de Panstrongylus megistus (Heteroptera: Reduviidae)

Important changes in the amount of neurosecretory material in the A and A' cells of the parsintercerebralis during the fifth instar intermoult of P. megistus occur. The release of neurosecretory material at different moments of intermoult, suggests that the A cells induce mitosis and that meiosis is induced by the release of A' cells material.