Involvement of calyculin A-sensitive phosphatase(s) in the differentiation of Trypanosoma cruzi trypomastigotes to amastigotes.

Differentiation of the non-dividing trypomastigote form of Trypanosoma cruzi, the causative agent of Chagas disease, to the dividing amastigote form normally occurs in cytoplasm of infected cells. Here we show that calyculin A. a potent inhibitor of protein phosphatases 1 and 2A, induces at pH 7.5 extracellular transformation of long slender trypomastigotes to round amastigote-like forms which acquire characteristic features observed after the normal differentiation process: repositioning and structural changes of the kinetoplast, release of surface neuraminidase, and expression of amastigote-specific epitopes. Calyculin A inhibits parasite phosphatases and changes in the phosphorylation of specific proteins occur during the transformation process. As an exposure of trypomastigotes to calyculin A concentrations as low as 1 nM and for only 1-2 h is sufficient to induce transformation, the inhibition of calyculin A-sensitive phosphatase(s) appears to play a major role in initiating the trypomastigote differentiation.

Tardivesicula duplicata gen. et sp. nov. (microspora, duboscqiidae), a microsporidian parasite of the caddis fly Limnephilus centralis (trichoptera, limnephilidae) in Sweden.

The microsporidium Tardivesicula duplicata gen. et sp. nov., a parasite of the adipose tissue of larvae of the caddis fly Limnephilus centralis in Sweden, is described based on light microscopic and ultrastructural characteristics. All life cycle stages have isolated nuclei. The sporogony is polysporoblastic, yielding 16-32 spores. The spores are rod-shaped, 1.3-1.4 × 3.7-4.2 µm in living condition. The spore wall has a 34-40 nm thick, layered exospore, with a median layer resembling a unit membrane. The endospore is thicker than the exospore. The polaroplast has two lamellar parts, where the posterior lamellae are the widest. The isofilar, 128-149 nm thick polar filament is arranged in 9-10 coils in a single layer close to the spore wall in the posterior 2/3 of the spore. The angle of tilt is c. 45°. The transversely sectioned polar filament has distinct concentrical layers of variable thickness and electron density. Sporophorous vesicles are ellipsoid and fragile. The thin electron-dense envelope is initiated as narrow tubulus-like promordia, appearing when the nucleus of the sporont has divided once. The episporontal space of vesicles with sporogonial plasmodia is devoid of inclusions. Vesicles with sporoblasts and spores have tubular exospore-derived inclusions of two dimensions. The new genus is compared to genera with polysporoblastic sporogony in sporophorous vesicles, primarily to Cystosporogenes and Flabelliforma. The placing of the new genus in the family Duboscqiidae is discussed.

Selective and reversible effects of vinca alkaloids on Trypanosoma cruzi epimastigote forms: blockage of cytokinesis without inhibition of the organelle duplication.

Vinca alkaloids, vincristine and vinblastine, produce differential effects on the cell division of Trypanosoma cruzi epimastigote forms depending on drug concentrations. These effects are related to different microtubule-based mechanisms. For 15 microM vinblastine and 50 microM vincristine, the drugs inhibit both nuclear division and cytokinesis, and affect cell shape. At 3 microM vinblastine and 10 microM vincristine, however, cytokinesis is inhibited without major effect on the progression of the cell cycle; this yields giant cells having multiple nuclei, kinetoplasts and flagella. Cultures maintained over 1 week with daily drug replacement produced cells with more than 16 nuclei and 24 kinetoplasts, indicating that an equivalent of a fifth cell cycle was initiated. The ultrastructure of the multinucleate cells showed a basic organization closely similar to that of trypanosomes. Cytokinesis inhibition by vinca alkaloids seems to result from modulations of interactions between microtubules and associated proteins, rather than from an inhibition of microtubule dynamics as is usually proposed for vinca alkaloids. Cytokinesis inhibition is reversible: after removing the drug, epimastigotes emerge from the multinucleate cells. The emerging process follows a precise axis and polarity which are determined by the position of the flagellum/kinetoplast complex. This region could play an essential role in cell morphogenesis since zoids (cells without a nucleus) are frequently observed.