Brazilian Theraphosidae: a toxicological point of view

The Theraphosidae family includes the largest number of species of the Mygalomorphae infraorder, with hundreds of species currently catalogued. However, there is a huge lack on physiologic and even ecologic information available, especially in Brazil, which is the most biodiverse country in the world. Over the years, spiders have been presented as a source of multiple biologically active compounds with basic roles, such as primary defense against pathogenic microorganisms or modulation of metabolic pathways and as specialized hunters. Spider venoms also evolved in order to enable the capture of prey by interaction with a diversity of molecular targets of interest, raising their pharmaceutical potential for the development of new drugs. Among the activities found in compounds isolated from venoms and hemocytes of Brazilian Theraphosidae there are antimicrobial, antifungal, antiparasitic and antitumoral, as well as properties related to proteinase action and neuromuscular blockage modulated by ionic voltage-gated channel interaction. These characteristics are present in different species from multiple genera, which is strong evidence of the important role in spider survival. The present review aims to compile the main results of studies from the last decades on Brazilian Theraphosidae with special focus on results obtained with the crude venom or compounds isolated from both venom and hemocytes, and their physiological and chemical characterization.(AU)

Advancements in the study of the classification and immune function of shrimp hemocytes / 生物工程学报

Hemocytes play an important role in the immune defense system of animals, especially for invertebrates that have no adaptive immune system. In those animals, hemocytes not only participate in the cellular immunity including phagocytosis, encapsulation, and nodules formation, but also the humoral immunity via storage and release of immune factors. Identification of the components of hemocytes is the basis for understanding the immune mechanism and the function of hemocytes. Despite various researches have been done on distinguishing the composition and function of shrimp hemocytes, no standard is used uniformly until now. So, we analyze and summarize the results on shrimp hemocytes research and offer a three subgroups category in this review. We also introduce the morphological characters and immune function of three subgroups in detail. We hope this work will be beneficial for understanding the function and molecular mechanism of hemocytes in invertebrate, bringing ideas for new separation technology development.

Dynamics of haemocytes from Pseudosuccinea columella circulating infected by Fasciola hepatica / Dinâmica de hemócitos circulantes de Pseudosuccinea columella infectados por Fasciola hepatica

Abstract The lymnaeids are important in the epidemiology of Fasciola hepatica, a neglected and endemic zoonosis. The interaction between the internal defense system of Pseudosuccinea columella and F. hepatica has been little studied. In the present study the effect of infection by F. hepatica on P. columella circulating haemocytes was investigated. Changes in the average number of total circulating haemocytes have been observed at 30 minutes post-infection and 1, 7, 10, 14, 21, 28 and 50 days post-infection (dpi). Miracidia were observed head-foot and mantle at 30 minutes post-infection. Miracidia/Sporocysts in the mantle skirt 1 dpi, and fully formed sporocysts were observed in the head-foot at 7 dpi. Rediae became evident at 10 dpi and were located between the haemocoel and the muscles from 14 dpi; 50 dpi, the rediae in the digestive gland contained cercariae. The statistical analysis of the total haemocytes of P. columella infected by F. hepatica showed significant differences on the 30 minutes post-infection and 1, 14, 21, and 28 dpi in comparison to uninfected molluscs (0 dpi). Therefore, the interference observed on the internal defence system of P. columella may have direct association with the development of F. hepatica.

Resumo Os limnaeideos são importantes na epidemiologia de Fasciola hepatica, uma zoonose negligenciada e endêmica. A interação entre o sistema interno de defesa de Pseudosuccinea columella e F. hepatica tem sido pouco estudada. No presente estudo, investigou-se o efeito da infecção por F. hepatica nos hemócitos circulantes de P. columella. Alterações no número médio de hemócitos circulantes foram observadas aos 30 minutos e 1, 7, 10, 14, 21, 28 e 50 dias após a infecção (dpi). Miracídios foram observados na região cefalopodal e manto aos 30 minutos após a infecção. Miracídio/esporocistos foram observados no colar do manto ao 1 dpi, e esporocistos totalmente formados na região cefalopodal aos 7 dpi. Rédias tornam-se evidentes aos 10 dpi entre a hemocele e músculos a partir de 14 dpi; e rédias com cercárias próximas a glândula digestiva aos 50 dpi. A análise estatística dos hemócitos totais de P. columella infectados por F. hepatica demonstrou diferenças significativas nos 30 minutos pós-infecção e 1, 14, 21 e 28 dpi em comparação aos moluscos não infectados (0 dpi). Portanto, a interferência observada no sistema de defesa interna de P. columella pode ter associação direta com o desenvolvimento de F. hepatica.

Estudio de componentes hemolinfáticos de Rhodnius prolixus y Triatoma maculata alimentados con diferentes fuentes sanguíneas / Study of hemolymphatic components of Rhodnius prolixus and Triatoma maculata fed on different blood sources

RESUMEN Introducción: Tradicionalmente, Rhodnius prolixus ha sido considerado el principal vector, mientras que Triatoma maculata resulta ser un vector secundario de Trypanosoma cruzi en el medio rural venezolano. En este trabajo se provee información de interés acerca de los componentes bioquímicos y del sistema inmune, humoral y celular, de la hemolinfa de R. prolixus y T. maculata alimentados sobre gallina y rata. Metodología: los insectos provenían de una colonia del laboratorio, mantenida a una temperatura de 27-29ºC, 50% de humedad relativa y ocho horas luz/día. Se analizó la hemolinfa de adultos alimentados sobre gallina y rata. Se observaron los hemocitos mediante microscopía óptica y electrónica de transmisión. Resultados: los parámetros bioquímicos de glucosa, lípidos y proteínas fueron afectados de forma distinta en R. prolixus y T. maculata según la fuente alimenticia. T. maculata presentó mayores niveles de actividad lítica de lisozima. Se observaron cuatro poblaciones de hemocitos (prohemocitos, plasmatocitos, granulocitos y oenocitos), cuyas características y medidas están acorde a lo reportado por otros autores sobre la subfamilia Triatominae. T. maculata presentó mayor número de prohemocitos y oenocitos que R. prolixus. Conclusiones: En su hemolinfa, R. prolixus y T. maculata son afectados distintamente en los componentes bioquímicos (glucosa, lípidos y proteínas) e inmunes, tanto humorales (lizosima) como celulares (prohemocitos, oenocitos), según sean alimentados sobre gallina y rata. Esto pone de manifiesto de la alimentación sobre uno de los componentes de la competencia vectorial, como lo es el sistema inmune propio de la hemolinfa de los insectos.

ABSTRACT Introduction: Traditionally, Rhodnius prolixus has been considered the main vector, while Triatoma maculata remains to be a secondary vector of Trypanosoma cruzi in the Venezuelan rural habitat. In this work, interesting information about the biochemical and immune (humoral and cellular) aspects of the hemolymph of R. prolixus and T. maculate, feeding on hen and rat, are presented. Methodology: Hemolymph was extracted from adult insects, maintained at 2729 °C with 50% of relative humidity and 8/16 (Light/Dark) hours, and fed on hen and rat. Hemocytes were observed by optical and transmission electron microscopy. Results: Biochemical parameters (glucose, lipids and proteins) varied in both species according to the food source. T. maculata presented higher levels of lysozyme lytic activity. Four hemocytes populations were observed by optical and transmission electron microscopy (prohemocytes, plasmocytes, granulocytes and oenocytes), exhibiting characteristics and measurements in accordance with previous literature related to the Triatominae subfamily. T maculata presented more prohemocytes and oenocytes than R. prolixus. Conclusions: On their hemolymph, R. prolixus and T. maculata are distinctly affected in their biochemical (glucose, lipids and proteins) and immune components, both humoral (lysozyme) and cellular (prohemocytes, oenocytes), depending on whether they were fed on hens or rats. Our results show that the food source affects the immune system of triatomines, and subsequently, their vectorial capacity may be compromised as well.

Comparación del recuento celular entre un método manual y un contador automatizado en líquidos de derrame / Comparison of manual microscope method and an automated cell count in serous fluids / Comparação da recontagem celular entre um método manual e um contador automatizado em líquidos de derrame

El objetivo del trabajo consistió en comparar el recuento celular total en los líquidos de derrame de cavidades serosas entre el método automatizado empleado en hematología y el método manual en hemocitómetro. Se procesaron 107 muestras: 45 líquidos ascíticos (LA) y 62 líquidos pleurales (LP) a los que se les realizó el recuento celular en cámara de Neubauer y en contador hematológico Sysmex XT 1800i. Se obtuvieron los siguientes resultados: 1) Regresión lineal: los coeficientes de correlación indicaron una alta correlación entre ambos métodos (LA r2: 0,999; p<0,0001 y LP r2: 0,997; p<0,0001). 2) Bland-Altman: El análisis de las figuras muestra una excelente concordancia entre ambos métodos. El error sistemático fue 51 para los LA y 97 para los LP, por lo que estos valores son despreciables dado el valor diagnóstico de los datos. Los resultados demuestran que los métodos son comparables entre sí y, por ende, se puede remplazar el recuento manual por el automatizado, de demostrada eficiencia y exactitud. Sin embargo, todos los líquidos requieren una observación al microscopio óptico previa al procesamiento por el contador hematológico, donde se apreciará la presencia de agrupamientos celulares como, por ejemplo, células neoplásicas en disposición glandular que dificultan el análisis por parte del equipo o la interpretación del resultado.

The purpose of this work was to compare the total cell count in liquids serous cavities between the automated method used in hematology and the manual method hemocytometer. A total of 107 samples were processed: 45 ascites fluids (LA for its name in Spanish) and 62 pleural fluids (LP for its name in Spanish). The cells were counted in improved Neubauer counting chamber and hematology analyzer Sysmex XT 1800i. The following results were obtained: 1) Linear Regression correlation coefficients indicated a high correlation between the two methods (LA r2: 0.999; p<0.0001 LP r2: 0.997; p<0.0001). 2) Bland-Altman analysis graphics showed excellent agreement between both methods. The systematic error was 51 for LA and 97 for LP; these values are insignificant considering the diagnostic value of the data. he results demonstrate that the methods are comparable and therefore can replace the manual counting by the automated method with proven efficiency and accuracy. However, all fluids require observation by optical microscope before being processed by the hematology analyzer, where the presence of cell clusters such as neoplastic cells in glandular disposition will be appreciated, which hinder the analysis by the equipment or interpretation of results.

O objetivo do trabalho consistiu em comparar a contagem total de células em líquidos de derrame de cavidades serosas entre o método automatizado utilizado em hematologia e o método manual em hemocitômetro. Foram processadas 107 amostras: 45 líquidos ascíticos (LA) e 62 líquidos pleurais (LP) nos quais se realizou a recontagem celular na câmara de Neubauer e no contador hematológico Sysmex XT 1800i. Foram obtidos os seguintes resultados: 1) Regressão linear: os coeficientes de correlação indicaram uma alta correlação entre ambos os métodos (LA r2: 0,999; p<0,0001 e LP r2: 0,997; p<0,0001). 2) Bland-Altman: A análise dos Figuras mostra uma excelente concordância entre ambos os métodos. O erro sistemático foi 51 para os LA e 97 para os LP, resultando estes valores desprezáveis dado o valor diagnóstico dos dados. Os resultados demonstram que os métodos são comparáveis entre si e, portanto, pode ser substituída a contagem manual pela automatizada, de eficiência e exatidão demonstradas. Entretanto, todos os líquidos requerem observação no microscópio óptico prévia ao processamento pelo contador hematológico. Nesse momento se apreciará a presença de agrupamentos celulares como, por exemplo, células neoplásicas em disposição glandular que dificultam a análise por parte da equipe ou a interpretação do resultado.

Hypoxia increases susceptibility of Pacific white shrimp to whitespot syndrome virus (WSSV) / Condição de hipóxia aumenta a susceptibilidade do camarão branco do Pacíficoao vírus da mancha branca (WSSV)

The present study aimed to evaluate the mortality, reactive oxygen species production (ROS) and total hemocyte counts (THC) of the marine shrimp Litopenaeus vannamei infected with the white spot syndrome virus (WSSV) at three levels of oxygen saturation. For this, 360 shrimp (20±2g) were distributed in 24 tanks (60L), divided in two groups (infected and non-infected), which were subjected to 30, 60 and 100% of dissolved oxygen saturation (in quadruplicate). During 96 hours after infection, daily hemolymph samples were collected for hemato-immunological parameter evaluation (THC and ROS) and dead animals were removed and computed to assess cumulative mortality rates. In the infected group, animals subjected to 100% saturation showed higher ROS production (P<0.05) after 48 hours, while THC was significantly reduced (P<0.05), regardless of oxygen saturation. The hypoxia resulted in high mortality when compared to 100% saturation condition. In the uninfected group, no significant differences were observed in all evaluated parameters. Thus, the hypoxia condition increased the susceptibility of shrimp to the infection of WSSV, which may be partly related to the low ROS production showed by the animals subjected to 30% oxygen saturation.

O presente estudo teve por finalidade avaliar a mortalidade e a contagem total de hemócitos (CTH) e espécies reativas de oxigênio (EROs) de camarão Litopenaeus vannamei infectados com o vírus da mancha branca (WSSV) e submetidos a três níveis de saturação de oxigênio. Para tanto, 360 camarões (20±2g) foram distribuídos em 24 tanques (60L), divididos em dois grupos, infectados e não infectados e submetidos a 30, 60 e 100% de saturação de oxigênio (em quadruplicata). Após a infecção, diariamente foram coletadas amostras de hemolinfa dos animais para avaliação dos parâmetros hematoimunológicos (CTH e EROs) e foi estimada a mortalidade, por 96 horas. No grupo com infecção, os animais submetidos à saturação de 100% apresentaram um aumento na produção de EROs (P<0,05) após 48 horas, ao mesmo tempo em que a CTH demonstrou uma redução (P<0,05) independentemente da saturação do oxigênio, e a condição de hipóxia acarretou maiores mortalidades quando comparada à do grupo com 100% de saturação. No grupo sem infecção, não foram observadas diferenças significativas nos parâmetros avaliados nem mortalidade. Dessa forma, pode-se concluir que a hipóxia aumentou a susceptibilidade do camarão à infecção com o vírus da mancha branca, que pode estar, em parte, relacionada com a baixa contagem de hemócitos e produção de EROs observadas nos animais submetidos a essa condição.

Caracterização hemocitária de uma linhagem resistente de Biomphalaria straminea (Dunker, 1848) exposta a schistosoma mansoni sambon, 1907 / Haemocyte characterization of a resistant lineage of biomphalaria straminea (Dunker, 1848) exposed to schistosoma mansoni sambon, 1907

A esquistossomose mansônica ainda constitui um grave problema de saúde pública no Brasil e, portanto, o conhecimento dos diversos aspectos da interação Schistosoma mansoni-Biomphalaria, são pertinentes e relevantes como subsídios à medidas de controle e prevenção desta parasitose. Estudos in vitro têm mostrado que a suscetibilidade dos hospedeiros intermediários de S. mansoni, Biomphalaria straminea e Biomphalaria glabrata está relacionado à presença de lectinas. Importante transmissor da Esquistossomose no Brasil, B. straminea apresenta baixos índices de infecção em condições experimentais. Apesar disto, existem poucos estudos a respeito das características morfológicas e do comportamento in vitro dos hemócitos deste molusco frente à infecção por S. mansoni. Com o objetivo de caracterizar a resposta hemocitária de B. straminea (cepa Souza-PB) expostas a miracídios de S. mansoni, os moluscos foram expostos em massa e expostos individualmente a cinco miracídios. Exemplares de B. glabrata foram utilizadas como controle positivo. A hemolinfa foi coletada, corada em Azul de Tripan e os hemócitos foram contados em câmara de Neubauer. Alguns hemócitos foram colocados em placa de cultura para observação da interação das células com os parasitos e outros foram submetidos à marcação por lectinas fluoresceinadas de Griffonia simplicifolia e Lens culinaris conjugadas a FITC. As imagens foram obtidas e gravadas através de microscópio AxioObserver e câmera McR5 Zeiss. Os moluscos expostos em massa foram sacrificados logo após a penetração inicial do parasito (0 minuto), 24, 48, 72 horas e 30 dias após exposição (dpe)...

Schistosomiasis still remain as an important parasitic disease under the public health point of view.Thus, studies that involve the interaction between Schistosoma mansoni-Biomphalaria are pertinentand useful to the disease prevention and control. In vitro studies have shown that the susceptibility ofBiomphalaria straminea and Biomphalaria glabrata, intermediate hosts of Schistosoma mansoni, isrelated to the presence of lectins. As an important carrier of schistosomiasis in Brazil, B. stramineapresents low infection rates under laboratory conditions. Despite of that, there are few studiesregarding the morphological features and in vitro behavior of this mollusc's haemocytes challenged byS. mansoni miracidia. For this purpose, we analyzed the haemocyte response of B. straminea (Sousa– PB strain) challenged by the parasite, so the molluscs were put under mass exposure and individualexposure to five miracidia. B. glabrata molluscs were used as positive controls. The molluscs had theirhaemolymph drawn, the haemocytes were stained in Trypan Blue and counted in Neubauer chamber.Some haemocytes were put into cell culture plates in order to observe the interaction between thecells and the parasites and other cells were stained by FITC conjugated lectins from Griffoniasimplicifolia and Lens culinaris. The images and films were made at the microscope AxioObserver withMcR5 camera from Zeiss. The mass exposed molluscs were killed at 0 minutes (first penetration of themiracidia), 24, 48, 72 hours and 30 days after exposure (dae)...

Perfil proteico da hemolinfa de Biomphalaria glabrata e linhagens de B. straminea, frente à exposição ao Schistosoma mansoni / Protein profile of hemolymph of Biomphalaria glabrata and strains of B. straminea, against the exposure to the Schistosoma mansoni

O gênero Biomphalaria possui espécies de grande relevância médica uma vez que atuam como hospedeiros intermediários naturais do parasita Schistosoma mansoni, causador da esquistossomose. Dentro desse gênero de moluscos, três espécies são tidas como hospedeiros naturais do parasita, Biomphalaria glabrata, B. straminea e B. tenagophila. O perfil de suscetibilidade à infecção por S. mansoni dentro do gênero é muito variado e muitas pesquisas buscam elucidar a dinâmica da relação parasita-hospedeiro intermediário na finalidade de criar novas medidas de controle da doença. Por isso, esse estudo tem como objetivo determinar o perfil bidimensional de proteínas que podem estar envolvidas na resposta imune contra o S. mansoni comparando duas espécies com diferentes perfis de susceptibilidade B. glabrata, B. straminea além de uma refratária ao S. mansoni, a B. straminea R3. Para isso, os caramujos de cada espécie foram divididos em dois grupos: Infectado, expostos aos miracídios do S. mansoni; e Controle, submetidos ao estresse do processo de infecção livre de miracídios. A hemolinfa foi retirada 24 horas após a exposição. Foi feito o extrato proteico total e determinada a concentração das proteínas totais para cada grupo investigado. As proteínas foram separadas por eletroforese bidimensional onde foi obtido o ponto isoelétrico e peso molecular de todos os spots nos géis...

The Biomphalaria has species of great medical relevance since that act as natural intermediate hosts of the parasite Schistosoma mansoni, which causes schistosomiasis. Within this kind of mollusks, three species are considered natural hosts of the parasite, Biomphalaria glabrata, B. stramineaand B. tenagophila. The profile of usceptibility to S. mansoni infection within the genre is very varied and many studies seek to elucidate the dynamics of host-parasite relationship intermediary in order to create new disease control easures. Therefore, this study aims to determine the two-dimensional profile of proteins that may be involved in the immune response against S. mansonicomparing two species with different susceptibility profiles B. glabrata, B. straminea and a refractory to S. mansoni, B. straminea R3. For that, the snails of each species were divided into two groups: Infected exposed to iracidia of S. mansoni; and control, subjected to stress the miracidia free infection process. The hemolymph was removed 24 hours after exposure. It was made the total protein extract and determined the concentration of total protein for each group investigated. Proteins were separated by two-dimensional electrophoresis was obtained where the isoelectric point and molecular weight of all the spots in the gels...

Identification and characterization of a novel hemocyte specific gene Bm04862 in silkworm, Bombyx mori / 生物工程学报

Bm04862 is a novel hemocyte-specific gene, which has been identified and cloned through the microarray data in silkworm, Bombyx mori. Initially, we successfully obtained the full-length cDNA sequence of Bm04862 via the rapid amplification of cDNA ends (RACE). The sequence consists of an open reading frame (ORF) with 819 bp, which could encode 273 amino acid residues. The bioinformatics analysis predicted that Bm04862 was a transmembrane protein. Furthermore, qRT-PCR analysis revealed that the expression of Bm04862 was specifically detected in hemocyte and reached a peak at L4M and PP2 stage. Bm04862 was overexpressed in Sf9 insect cells, and the cellular localization indicated that Bm04862 was specifically located in cytoplasm and nuclei membrane. Interestingly, the expression of Bm04862 increas'd dramatically after challenged with Escherichia coli for 24 hours, which predicted its potential role in the innate immune system. Overall, this study could provide the fundamental knowledge for further research.

Análise da variabilidade imunológica e da expressão dos genes FREPs entre as espécies Biomphalaria glabrata e B. straminea com diferentes perfis de suscetibilidade ao parasito Schistosoma mansoni / Analysis of immune variability and expression of FREPS genes between the species Biomphalaria glabrata and B. straminea with different susceptibility profiles to infection to by Schistosoma mansoni

Algumas espécies do gênero Biomphalaria se apresentam como potenciais hospedeiras ao parasito Schistosoma mansoni, estando a suscetibilidade a este parasito, neste gênero, ligada ao sistema interno de defesa de cada espécie de Biomphalaria. Um dos componentes importantes no sistema imune de invertebrados é a enzima fenoloxidase, que ainda apresenta muitos aspectos desconhecidos no sistema de defesa do gênero Biomphalaria. Foi relatado também que os genes de proteínas relacionadas ao fibrinogênio (FREPs) possuem importância na resposta imune de Biomphalaria glabrata, entre esses, as subfamílias dos FREPs 3 e 4 são diferencialmente expressas em linhagens susceptíveis e resistentes frente a infecção com trematódeos. No entanto os trabalhos existentes em sua maioria estudam a espécie Biomphalaria glabrata, excluindo a espécie Biomphalaria straminea, amplamente distribuída no Brasil e principal responsável pela disseminação da esquistossomose. Tendo em vista a falta de conhecimento sobre a resposta imune destes moluscos hospedeiros, principalmente em relação à expressão de genes imune relevantes e ao tipo de resposta, o presente trabalho se propôs a estudar a variação do número de hemócitos, da produção de fenoloxidase e da expressão dos genes dos FREP 3 e FREP 4 envolvidos com a ligação a antígenos de trematódeos, nas espécies Biomphalaria glabrata, Biomphalaria straminea pós-infecção com S. mansoni, bem como em caramujos pré-expostos a antígenos de S. mansoni...

Caracterização da resposta imunológica de Aedes aegypti e Culex quinquefasciatus à infecção pelo sorotipo 1 do Dengue vírus / Characterization of immune response of Aedes aegypti and Culex quinquefasciatus to infection by Dengue virus serotype 1

Os insetos podem atuar como pragas agrícolas e vetores de patógenos causadores de doenças ao homem e outros animais. Investigações a respeito do sistema imunológico de Ae. aegypti e Cx. quinquefasciatus poderão contribuir para o desenvolvimento de métodos de controle das doenças veiculadas por estes insetos, principalmente a dengue, enfermidade causadora de sério problema de saúde pública no mundo. Apesar de Ae. aegypti ser a única espécie vetora confirmada na transmissão do vírus Dengue no Brasil, considera-se também importante um melhor entendimento dos mecanismos imunológicos de Cx. quinquefasciatus tido como refratário ao vírus. Neste estudo foram utilizadas linhagens de Ae. aegypti e Cx. quinquefasciatus mantidas no Insetário do Departamento de Entomologia do CPqAM/FIOCRUZ. Três grupos experimentais de fêmeas com 10 dias de idade foram formados para cada espécie. Grupo I, composto por fêmeas alimentadas com solução sacarose (10 por cento); grupo II, fêmeas alimentadas com sangue limpo e grupo III, fêmeas alimentadas com sangue infectado com o sorotipo DENV-1. De cada grupo foram obtidos hemolinfa, glândula salivar, intestino médio e corpo gorduroso para avaliação da expressão dos antimicrobianos defensina e transferrina. Essa avaliação foi realizada através de PCR em Tempo Real utilizando o kit QuantiFast SYBR Green - One-Step qRT-PCR. A avaliação da hemodinâmica foi realizada utilizando 10 microlitros de hemolinfa de cada grupo, através da contagem das células em câmara de Neubauer...

The significance of the amoebocyte-producing organ in Biomphalaria glabrata

In molluscs, internal defence against microorganisms is performed by a single cell type, i.e., the haemocyte or amoebocyte. The origin of these cells in Biomphalaria glabrata was initially thought to be localised within the vasculo-connective tissue. More recently, origin from a single organ, termed the amoebocyte-producing organ (APO), has been postulated based on the occurrence of hyperplasia and mitoses during Schistosoma mansoni infection. The present investigation represents a histological, immuno-histochemical and ultra-structural study of the B. glabrata APO, whereby histological identification was facilitated by means of collecting epithelial basophilic cells. These cells were comprised of single-cell layers that cover a portion of the stroma, which contains many small, round cells and haemolymph sinuses, as well as a small area of the pericardial surface of the reno-pericardial region. On occasion, this epithelial component vaguely resembled the vertebrate juxtaglomerular apparatus, which reinforces its presumed relationship to the kidney. Both in normal and infected molluscs, mitoses were only occasionally found. The present quantitative studies failed to demonstrate the presence of APO cellular hyperplasia, either in normal or schistosome-infected B. glabrata. Conversely, several structural details from the APO region in B. glabrata were found to be consistent with the hypothesis that the APO is a filtration organ, i.e., it is more closely related to the kidney rather than the bone marrow, as has been suggested in the literature.

Pancytopenia Associated with Hypopituitarism in an Acromegaly Patient: A Case Report and a Review of the Literature

We present the case of a patient with acromegaly who had pancytopenia with hypopituitarism secondary to the excision of a pituitary macroadenoma and radiation therapy. A 28-year-old man presented with pancytopenia and serum electrolyte abnormalities. He was diagnosed with acromegaly and underwent surgery and gamma-knife radiotherapy for a pituitary macroadenoma at the age of 22 years. A recent brain magnetic resonance imaging showed an empty sella, and the basal hormonal profile demonstrated deficiencies of pituitary hormones except thyrotropin. As presenting pancytopenia, his bone marrow biopsy showed hypocellular marrow. The total number of hemocytes increased after hydrocortisone replacement. Hypopituitarism was a possible cause of pancytopenia, and glucocorticoids had crucial effects on converting pancytopenia to normal in this case.

Participation of N-acetyl-D-glucosamine carbohydrate moieties in the recognition of Schistosoma mansoni sporocysts by haemocytes of Biomphalaria tenagophila

Lectin-carbohydrate binding may be involved in the recognition of Schistosoma mansoni sporocysts by haemocytes of Biomphalaria; therefore, we tested if this interaction is associated with snail resistance against Schistosoma infection. In vitro data showed that most of the S. mansoni sporocysts cultured with haemocytes from Biomphalaria glabrata BH, a highly susceptible snail strain, had a low number of cells that adhered to their tegument and a low mortality rate. Moreover, the addition of N-acetyl-D-glucosamine (GlcNAc) did not alter this pattern of adherence and mortality. Using haemocytes and haemolymph of Biomphalaria tenagophila Cabo Frio, we observed a high percentage of sporocysts with adherent cells, but complete encapsulation was not detected. Low concentrations of GlcNAc increased haemocyte binding to the sporocysts and mortality, which returned to basal levels with high concentrations of the carbohydrate. In contrast, haemocytes plus haemolymph from B. tenagophila Taim encapsulated cellular adhesion index of level 3 and destroyed over 30 percent of the S. mansoni sporocysts in culture. Interestingly, the addition of GlcNAc, but not mannose, to the culture medium resulted in the significant inhibition of cellular adhesion to the parasite tegument and the reduction of parasite mortality, suggesting that GlcNAc carbohydrate moieties are important to the recognition of S. mansoni by B. tenagophila Taim.

Interaction between primary and secondary sporocysts of Schistosoma mansoni and the internal defence system of Biomphalaria resistant and susceptible to the parasite

The outcome of the interaction between Biomphalaria and Schistosoma mansoni depends on the response of the host internal defence system (IDS) and the escape mechanisms of the parasite. The aim of this study was to evaluate the responsiveness of the IDS (haemocytes and soluble haemolymph factors) of resistant and susceptible Biomphalaria tenagophila lineages and Biomphalaria glabrata lineages in the presence of in vitro-transformed primary sporocysts and secondary sporocysts obtained from infected B. glabrata. To do this, we assayed the cellular adhesion index (CAI), analysed viability/mortality, used fluorescent markers to evaluate the tegumental damage and transplanted secondary sporocysts. B. tenagophila Taim was more effective against primary and secondary sporocystes than the susceptible lineage and B. glabrata. Compared with secondary sporocysts exposed to B. tenagophila, primary sporocysts showed a higher CAI, a greater percentage of dead sporocysts and were labelled by lectin from Glycine max and Alexa-Fluor 488 fluorescent probes at a higher rate than the secondary sporocysts. However, the two B. tenagophila lineages showed no cercarial shedding after inoculation with secondary sporocysts. Our hypothesis that secondary sporocysts can escape the B. tenagophila IDS cannot be confirmed by the transplantation experiments. These data suggest that there are additional mechanisms involved in the lower susceptibilty of B. tenagophila to S. mansoni infection.

Interactions of spodoptera littoralis haemocytes following injection with the entomopathogenic fungi: Beauveria Bassiana and Nomuraea Rileyi

This study compared the cellular interactions of Spodopteralittoralis haemocytes with two virulence-different entomopathogenic fungi: Beauveriabassiana and Nomuraearileyi. Using light and transmission microscopy, five types of haemocytes namely, prohaemocytes [PRs], plasmatocytes [PLs], granulocytes [GRs], spherule cells [SPs] and oenocytoids [OEs] were identified in the 6[th] instar larvae. PRs and PLs were found in the haemopoietic tissue.Intra-haemocoelic injection of blastospores induced ultrastructural alterations in the cytoplasm and nucleiof circulating haemocytes of treated larvae. Different responses were observed in the populations of haemocyte types following injection with the tested fungi. The most important changes were the decrease of the numbers of GRs aiccompanied with increase inSPs at 12-48h following injection with B. bassiana, whereas,a decrease of PLs with a commitment increase inSPs and OEs were observed at most time intervals after injection with N. rileyi. Both fungi provoked a decrease of the total number of haemocytes at 48h followed by an increase at 72h post-injection. In vivo assay showed that the GRs and PLs actively phagocytised fungal blastospores. There was a time-dependent decrease and increase in the phagocytosis activity after injection of B. bassiana and N. Rileyi, respectively .In B. bassiana-injected insects, the numbers nodules increased significantly at 6-48h in comparison with the controls post-injection. In N. rileyi-injected insects, nodules increased significantly only at 72h post-injection.No cellular encapsulation was observed in any of the examined insects

Non-specific Defensive Factors of the Pacific Oyster Crassostrea gigas against Infection with Marteilioides chungmuensis: A Flow-Cytometric Study

In order to assess changes in the activity of immunecompetency present in Crassostrea gigas infected with Marteilioides chungmuensis (Protozoa), the total hemocyte counts (THC), hemocyte populations, hemocyte viability, and phagocytosis rate were measured in oysters using flow cytometry. THC were increased significantly in oysters infected with M. chungmuensis relative to the healthy appearing oysters (HAO) (P<0.05). Among the total hemocyte composition, granulocyte levels were significantly increased in infected oysters as compared with HAO (P<0.05). In addition, the hyalinocyte was reduced significantly (P<0.05). The hemocyte viability did not differ between infected oysters and HAO. However, the phagocytosis rate was significantly higher in infected oysters relative to HAO (P<0.05). The measurement of alterations in the activity of immunecompetency in oysters, which was conducted via flow cytometry in this study, might be a useful biomarker of the defense system for evaluating the effects of ovarian parasites of C. gigas.

Caracterização dos hemócitos de operários de Nasutitermes coxipoensis(Holmgren) (Isoptera: Termitidae) e avaliação hemocitária após parasitismo por Metarhizium anisopliae / Hemocyte characterization of Nasutitermes coxipoensis (Holmgren) (Isoptera: Termitidae) workers and hemocyte evaluation after parasitism by Metarhizium anisopliae

O objetivo desta pesquisa foi caracterizar morfológica e ultraestruturalmente os hemócitos de operários de Nasutitermes coxipoensis (Holmgren) e quantificar os tipos celulares 24h, 48h e 72h após inoculação com Metarhizium anisopliae. Seis tipos de hemócitos foram identifi cados: plasmatócito, granulócito, esferulócito, prohemócito, adipohemócitos e oenocitóide. Nenhuma alteração na morfologia dessas células foi evidenciada durante os intervalos. Entretanto, houve variações na proporção dos hemócitos em relação à testemunha para esferulócitos, adipohemócitos e oenocitóides, nos três intervalos de avaliação, e para plasmatócitos e granulócitos no intervalo de 48h. As causas potenciais dessa variação e suas implicações são apresentadas e discutidas neste artigo.

We aimed to characterize the morphology and ultrastructure of hemocytes of Nasutitermes coxipoensis (Holmgren) workers and to quantify the cell types 24h, 48h and 72h after inoculation with Metarhizium anisopliae. Six hemocytes types were identified, plasmatocyte, granulocyte, spherulocyte, prohemocyte, adipohemocyte and eonocytoid Hemocytes did not present any morphological alteration at the several observation periods, but they did have a change in their abundance, as observed for spherulocytes, adipohemocytes and eonocytoids at all intervals, and for plasmatocytes and granulocytes at 48h after host inoculation. We argue on the possible reasons and implications of the observed changes.

Ultra-estrutura dos hemócitos de Aedes aegypti (Linnaeus, 1762) (Diptera, Culicidae) / Hemocytes ultrastructure of Aedes aegypti

O sucesso dos insetos em explorar diversos ambientes é devido, em grande parte, à habilidade em se defender contra patógenos e parasitas. Nos insetos, os principais mecanismos de defesa são desempenhados pelos hemócitos. A classificação dos tipos de hemócitos presentes na hemolinfa ainda é bastante controversa. A biodiversidade desses organismos tem proporcionado modelos importantes para o estudo de suas estratégias antimicrobianas, as quais podem fornecer informações relevantes para o combate a diversas doenças, bem como para o estudo da imunologia geral. O objetivo do presente estudo foi caracterizar a resposta imune celular de Aedes aegypti sob o ponto de vista morfológico, funcional e ultra-estrutural através da microscopia óptica e eletrônica de transmissão. Em nossos estudos identificamos seis tipos morfológicos de hemócitos circulantes na hemolinfa de Ae. aegypti através da microscopia de luz, eletrônica de transmissão e contraste de interferência diferencial, são eles: prohemócitos, adipohemócitos, granulócitos, plasmatócitos, oenocitóides e trombocitóides. Os prohemócitos foram as menores células encontradas na hemolinfa. Sua principal característica é a presença de um citoplasma ocupando uma pequena área em torno do núcleo. Os adipohemócitos foram os mais abundantes tipos celulares presentes e exibiam grandes inclusões lipídicas preenchendo praticamente todo o citoplasma. Os granulócitos possuem um citoplasma contendo diversos grânulos elétron-densos. Os plasmatócitos exibiram morfologia bastante polimórfica e diversos filopódios e pseudópodes. Os oenocitóides possuem citoplasma homogêneo com poucas organelas. Os trombocitóides são raros e possuem características similares aos oenocitóides com organelas pouco desenvolvidas. Os hemócitos responsáveis pela resposta imune contra partículas de látex conjugadas a FITC foram identificados através da microscopia laser confocal, assim como as lectinas que marcam os hemócitos. Os granulócitos foram as únicas células envolvidas na fagocitose de alvos abióticos in vitro e in vivo. As lectinas BSI, ConA, HPA, LCA, PNA, UEA e WGA marcaram os hemócitos com variações na intensidade. A WFA e LPL não marcaram hemócitos de Ae. aegypti .

Ki-67 is expressed in multiplying forms of Schistosoma mansoni, but not in snail host tissues

Ki-67 is a protein expressed in the nucleus of several species during cell-division, being absent during the GO resting phase of the cellular cycle. During attempts to disclose mitosis in the so-called " amebocyte-producing organ " in Biomphalaria glabrata infected with Schistosoma mansoni, the parasite multiplying forms appeared strongly marked for Ki-67, while the snail tissues were completely negative. These data are worth registering to complement general data on Ki-67, and to help future studies on the relationship of the parasite and of its intermediate host.