Interconnected marine habitats form a single continental-scale reef system in South America.

Large gaps in reef distribution may hinder the dispersal of marine organisms, interrupting processes vital to the maintenance of biodiversity. Here we show the presence and location of extensive reef habitats on the continental shelf between the Amazon Reef System (ARS) and the Eastern Brazilian Reef System (ERS), two reef complexes off eastern South America. Formations located 20-50 m deep include both biogenic and geogenic structures. The presence of diverse reef assemblages suggests the widespread occurrence of rocky substrates below 50 m. These habitats represent an expansion of both the ARS and ERS and the closure of the only remaining large-scale gap (~ 1000 km) among West Atlantic reef environments. This indicates that the SW Atlantic harbors a single, yet heterogeneous, reef system that stretches for about 4000 km, and thus, represents one of the largest semi-continuous tropical marine ecosystems in the world.

Heterogeneity in the synthesis of alpha-macroglobulins in outbred Swiss albino mice acutely infected with Trypanosoma cruzi.

Alpha-Macroglobulins (AM) are protease inhibitors with important roles in inflammation and in immunomodulation that behave as acute-phase proteins in many experimental models. In the present work the levels of AM in the plasma of outbred Swiss albino mice acutely infected with Trypanosoma cruzi were studied. The results showed that increased levels of AM were present in the majority of the infected mice and that AM levels increased independently of the rise in parasitaemia. There was a high degree of heterogeneity in the intensity of the modulation of AM levels as well as in the kinetics of AM synthesis. This heterogeneity was related neither with the intensity of infection nor with the sex of the host. No correlation between AM levels and survival to the acute phase could be observed in the outbred mice. The consequence of such a heterogeneity is unclear, although AM as immunoregulatory molecules could play a role in the development of the symptoms of the chronic phase of Chagas' disease.

Trypanosoma cruzi recognition by macrophages and muscle cells: perspectives after a 15-year study.

Macrophages and muscle cells are the main targets for invasion of Trypanosoma cruzi. Ultrastructural studies of this phenomenon in vitro showed that invasion occurs by endocytosis, with attachment and internalization being mediated by different components capable of recognizing epi- or trypomastigotes (TRY). A parasitophorus vacuole was formed in both cell types, thereafter fusing with lysosomes. Then, the mechanism of T. cruzi invasion of host cells (HC) is essentially similar (during a primary infection in the absence of a specific immune response), regardless of whether the target cell is a professional or a non-professional phagocytic cell. Using sugars, lectins, glycosidases, proteinases and proteinase inhibitors, we observed that the relative balance between exposed sialic acid and galactose/N-acetyl galactosamine (GAL) residues on the TRY surface, determines the parasite's capacity to invade HC, and that lectin-mediated phagocytosis with GAL specificity is important for internalization of T. cruzi into macrophages. On the other hand, GAL on the surface of heart muscle cells participate on TRY adhesion. TRY need to process proteolytically both the HC and their own surface, to expose the necessary ligands and receptors that allow binding to, and internalization in the host cell. The diverse range of molecular mechanisms which the parasite could use to invade the host cell may correspond to differences in the available "receptors" on the surface of each specific cell type. Acute phase components, with lectin or proteinase inhibitory activities (alpha-macroglobulins), may also be involved in T. cruzi-host cell interaction.

Trypanosoma cruzi: phagolysosomal fusion after invasion into non professional phagocytic cells.

Parasite-containing endocytic vacuoles are formed during the process of in vitro interiorization of the trypomastigote forms of Trypanosoma cruzi by primary culture of mouse fibroblasts, heart and skeletal muscle cells. Fusion of these vacuoles with host cell lysosomes takes place. The process of T. cruzi-muscle cell interaction was analysed by ultrastructural cytochemistry. Two lysosomal enzymes, acid phosphatase and aryl sulphatase and the fusion of peroxidase-labeled secondary lysosomes with the parasitophorus vacuoles were studied. These finding indicate that the basic mechanism of interaction of T. cruzi with the so called non phagocytic cells is similar to that which occurs with phagocytic cells.

Interaction of Trypanosoma cruzi with heart muscle cells: ultrastructural and cytochemical analysis of endocytic vacuole formation and effect upon myogenesis in vitro.

The process of interaction of bloodstream trypomastigotes of three different strains of Trypanosoma cruzi with heart mouse muscle cells in primary cultures, was analyzed. Differences were found in the ability of the parasites to infect the cells. Those from the Colombiana strain were more infective than those from the Y and CL strains. Infection of the cells with parasites of the Colombiana strain, but not with those of the Y strain, interfered with the normal myogenic process. Transmission electron microscopy of thin sections of heart muscle cells kept in contact with parasites for 18 h showed that many parasites are found within membrane-bounded endocytic vacuoles. Cytochemical localization of Ca2+-Mg2+-ATPase, adenylate cyclase and anionic sites (labelled with cationized ferritin) indicate that these components of the plasma membrane are not found in the membrane which lines the endocytic vacuole.

Trypanosoma cruzi: inhibition of host cell uptake of infective bloodstream forms by alpha-2-macroglobulin.

The infection of murine macrophages and fibroblasts by recently isolated infective bloodstream trypomastigotes of Trypanosoma cruzi is inhibited by the addition of human plasma protease inhibitor alpha-2-macroglobulin (alpha 2M) or of soybean trypsin inhibitor. The ingestion of the non-infective epimastigotes by macrophages is not affected by the physiological protease inhibitor. Incubation of bloodstream trypomastigotes for 20 h in a serum-free axenic medium enhances their ability to infect macrophages in a process influenced by the temperature and sensitive to alpha 2M. After this period the infectivity of the parasites to cells was not sensitive to alpha 2M. These observations suggest that proteases located on the surface and/or secreted by the bloodstream trypomastigote form of T. cruzi may modulate its ability to infect host cells.

Interaction of Trypanosoma cruzi with macrophages: effect of previous incubation of the parasites or the host cells with lectins.

The effect of incubation with lectins of the macrophages or two evolutive stages of Trypanosoma cruzi (noninfective epimastigotes and infective trypomastigotes) on the ingestion of the parasites by mouse peritoneal macrophages was studied. Lectins which bind to residues of mannose (Lens culinaris, LCA), N-acetyl-D-glucosamine or N-acetylneuraminic acid (Triticum vulgaris, WGA), beta-D-galactose (Ricinus communis, RCA), N-acetyl-D-galactosamine (Phaseolus vulgaris, PHA; Dolichos biflorus, DBA; and Wistaria floribunda, WFA), fucose (Lotus tetragonolobus, LTA), and N-acetylneuraminic acid (Limulus polyphemus, LPA) were used. By lectin blockage we concluded that, alpha-D-mannose-like, beta-D-galactose and N-acetyl-D-galactosamine (PHA, reagent) residues, located on the macrophage's surface are required for both epi- and trypomastigote uptake, while N-acetylneuraminic acid and fucose residues, impede trypomastigote ingestion but do not interfere with epimastigote interiorization. Macrophages' N-acetyl-D-glucosamine residues are required for epimastigote uptake. On the other hand, from the T. cruzi surface, mannose residues prevent ingestion of epi- and trypomastigotes. Galactose residues participate in endocytosis of trypomastigotes, but hinder epimastigote interiorization. Exposed N-acetyl-D-glucosamine residues are required for uptake of the two evolutive forms. N-acetylneuraminic acid residues on the trypomastigote membrane prevent their endocytosis by macrophages. These results together with those reported previously showing the effect of monosaccharides on the T. cruzi-macrophage interaction, indicate that (a) sugar residues located on the parasite and on macrophage surface play some role in the process of recognition of T. cruzi, (b) different macrophage carbohydrate-containing receptors are involved in the recognition of epimastigotes and trypomastigotes forms of T. cruzi, (c) N-acetylneuraminic acid residues located on the surface of trypomastigotes or macrophages impede the interaction of the parasite with these host cells, and suggest that (d) sugar-binding proteins located on the macrophage surface participate in the recognition of beta-D-galactose and N-acetyl-D-galactosamine residues located on the surface of trypomastigotes and exposed after blockage or splitting off of N-acetylneuraminic acid residues. Some lectins which bind to macrophages and block the ingestion of parasites did not interfere with their adhesion.

Effect of carbohydrates, periodate and enzymes in the process of endocytosis of Trypanosoma cruzi by macrophages.

The effect of mild enzyme (trypsin, neuraminidase) treatment, periodate treatment and addition of carbohydrates (mono, di-, and polysaccharides) on the ingestion of Trypanosoma cruzi epimastigotes and trypomastigotes by mouse macrophages was studied. Trypsin treatment did not interfere with the ingestion of epimastigotes but did, however, increase the ingestion of trypomastigotes by mouse peritoneal macrophages. Neuraminidase and periodate treatment of the parasites increased the uptake of epi- and trypomastigote forms. The neuraminidase effect was partially blocked by galactose or N-acetylgalactosamine. Galactose, mannose, fucose, N-acetylglucosamine, and N-acetylgalactosamine had an influence on the ingestion of T. cruzi by macrophages. This effect was dependent on the strain of parasite tested, and the medium used to cultivate the epimastigotes. The results obtained, in conjunction with the work of others, suggest that glycoproteins and/or glycolipids on the parasite and/or macrophage surface are involved in the T. cruzi-macrophage interaction.

Interaction of Trypanosoma cruzi with macrophages in vitro: dissociation of the attachment and internalization phases by low temperature and cytochalasin B.

Chicken macrophages, obtained by cultivation of blood monocytes, were infected with epimastigote and bloodstream trypomastigote forms of Trypanosoma cruzi strain Y. The percentage of macrophages containing parasites within parasitophorous vacuoles and of flagellates attached to cell surfaces was determined. By incubation of the macrophages at 4 degrees C or in the presence of cytochalasin B it was possible to dissociate the attachment from the internalization phases in the process of infection of macrophages. Both treatments had a marked effect on the internalization of epimastigote and trypomastigote forms. Cytochalasin B treatment and placement of the macrophages at 4 degrees C before infection inhibited this process by about 99 and 96%, respectively. These results suggest that endocytosis is the principal mechanism of internalization of T. cruzi by macrophages. They show also that epimastigote and trypomastigote forms of T. cruzi have a different rate of adhesion to the macrophage surface.

Interaction of epimastigote and trypomastigote forms of Trypanosoma cruzi with chicken macrophages in vitro.

Chicken macrophages, obtained by cultivation of blood monocytes, were infected with epimastigote and bloodstream trypomastigotes of the Y and the CL strains of Trypanosoma cruzi. The percentage of infected cells and the mean number of parasites per cell were determined after 2, 6, 12 and 24 h of parasite-cell contact. After 6 h of contact about 80% and 40% of the macrophages were infected by trypomastigotes of the Y and CL strains respectively. After longer periods of contact almost all macrophages were infected by Y trypomastigotes while only about 60% were infected by those of the CL strain. After 2 h of contact almost all macrophages were infected by CL epimastigotes while only about 50% were infected by Y epimastigotes. After 6 h of contact almost all macrophages were infected by epimastigotes of both strains. These results are discussed taking into consideration differences between parasites of the two strains and between the two developmental stages of the T. cruzi lifecycle.

Ultrastructural localization of calcium-binding sites in the electrocyte of Electrophorus electricus (L.).

Calcium-binding sites were detected in the electrocyte of Electrophorus electricus (L.) using the Oschman & Wall technique, in which CaCl2 was added to the fixative and washing solutions. Deposits were seen scattered along the plasma membrane of the electrocyte, inside mitochondria, associated with the post-synaptic membrane and the membrane of synaptic vesicles.