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.

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.

A contribution to the pathobiology of Biomphalaria glabrata hemocytes

This study attempts to investigate the relationship between the hemocytes in the two compartments: circulating peripheral lymph and the connective tissues. The hemocytes are compared with the vertebrate macrophages and constitute the principal line of defense against external aggression. The hemocytes were counted in circulating hemolymph and their phagocytic capability was evaluated in Schistosoma mansoni-infected Biomphalaria glabrata and the results were compared with those obtained from normal intact control snails. Although the number of circulating hemocytes revealed a mild increase in snails at the 6th week of infection, the overall findings were similar and pointed out that the cells in the two compartments are not functionally connected. However, the hemocytes found within the connective tissues of infected snails showed definite ultrastructural differences in the number and disposition of cytoplasmic prolongations and organelles in comparison with the hemocytes from non-infected snails. Histochemically, the staining for acid phosphatase activity served as a marker to hemocytes, sometimes being found in extracellular material at the foci of parasite-hemocyte interactions.

Changes induced in Biomphalaria glabrata (Say, 1818) following trials for artificial stimulation of its internal defense system

Biomphalaria glabrata can react through different pathways to Schistosoma mansoni miracidium penetration, according to the degree of resistance/susceptibility presented by different snail strains, which is a genetically determined character, resistance being the dominant feature. However, it has been observed that previous susceptible snail strain may change its reactive behavior along the course of infection, exhibiting later a pattern of cercarial shedding and histopatopathological picture compatible with high resistance. Such observation suggests the possibility of B. glabrata to develop a sort of adaptative immunity face a schistosome infection. To explore on this aspect, the present investigation looked for the behavior of S. mansoni infection in B. glabrata previously subjected to different means of artificial stimulation of its internal defense system. Snails previously inoculated with irradiated miracídia (Group I); treated with S. mansoni antigens (Group II) or with a non-related parasite antigen (Group III) were challenged with 20 viable S. mansoni miracidia, and later looked for cercarial shedding and histopathologic changes at different times from exposition. Nodules of hemocyte accumulations were found at the site of antigen injection. These nodules resembled solid granulomas, and were larger and more frequent in snails injected with S. mansoni products as compared to those injected with Capillaria hepatica. However, the presence of such granulomas did not avoid the S. mansoni challenge infection from developing in a similar way as that seen in controls. The data are indicative that hemocytes are able to proliferate locally when stimulated, such capacity also remaining localized, not being shared by the population of hemocytes located elsewhere within the snail body.

Effect of fixed epimastigote forms of Trypanosoma cruzi on the hemocytes and the prophenoloxidase-activating system of the crayfish Pacifastacus leniusculus

The effect of the parasite Trypanosoma cruzi on the hemocystes and the prophenoloxidase (proPO)-activating system of the crayfish Pacifastacus leniusculus was studied. Incubation of the crayfish hemocyte lysate with fixed epimastigote forms of the parasites (4 x 10(5) cells/ml) induced a marked activation of the crayfish proPO system, measured as phenoloxidase activity. The activation of proPO by the parasite was much stronger (7-fold) than that induced by beta-1,3-glucans (1 mg/ml) which are known to be efficient elicitors of the proPO system. The fixed parasites promoted the spreading and degranulation of different populations of the crayfish hemocytes isolated by Percoll gradients, and were often observed to be attached to the crayfish hemocytes in rosette-like fashion. The attachment of the epimastigote forms of T. cruzi to the crayfish blood cell surface was not dependent on the adhesive 76-kDa protein released by the crayfish hemocytes, since the exocytotic inhibitor SITS and monospecific antibodies to the 76-kDa protein did not prevent parasite adhesion. The crayfish hemocytes apparently are able to phagocytose the fixed epimastigote forms of T. cruzi in vitro.