Biological aspects of the DM28C clone of Trypanosoma cruzi after metacylogenesis in chemically defined media

The biological characterization of the Trypanosoma cruzi clone Dm 28c in terms of its growth in LIT medium, cell-cycle, infectivity to mice and interaction with professional and non-professional phagocytic cells shows that it behaves as a bona fide T. cruzi representant. The biological properties of this myotropic clone do not change according to the origin of the trypomastigote forms (i. e., from triatomines, infected mice, cell-culture or from the chemically defined TAUP and TAU3AAG media). In addition Dm 28c metacyclic trypomastigotes from TAU3AAG medium display a high infectivity level to fibroblasts and muscle cells. Experiments on binding of cationized ferritin to trypomastigotes surface show the existence of cap-like structures of ferritin in regions near the kinetoplast. However the nature and role of these anionic sites remain to be determined. The results indicate that metacyclic trypomastigotes from Dm 28c clone obtained under chemically defined conditions reproduce the biological behaviour of T. cruzi, rendering this system very suitable for the study of cell-parasite interactions and for the isolation of trypanosome relevant macromolecules.

Caracterização biológica de clones das cepas Y, CL e MR de Trypanosoma cruzi em camundongos C3H isogênicos

Dez clones isolados das cepas Y, CL e MR foram caracterizados segundo infectividade das culturas, curvas de parasitemia, polimorfismo e mortalidade em camundongos C3H isogênicos. Entra os clones das cepas Y e CL foram encontradas diferenças intragrupos bastante significativas. Os clones da cepa MR apresentaram maior homogeneidade. Estes resultados indicam que as cepas do T. cruzi podem apresentar diferentes graus de heterogeneidade. Também sugerem que as condições utilizadas para a manutenção de cepas de T. cruzi podem resultar em vantagens seletivas para algumas subpopulações, podendo uma cepa ser o resultado da interação destas subpopulações (clones) selecionadas após alguns anos de manutenção em laboratório.

El ensayo inmunoenzimatico en microgotas sobre nitrocelulosa (Dot-ELISA) en el diagnostico de la enfermedad de Chagas: I. Estudio comparativo de dos preparaciones antigenicos de Trypanosoma cruzi

Se estudia el Ensayo Inmunoenzimático en Microgotas sobre Nitrocelulosa (Dot-ELISA)comparando dos preparados antigénicos de formas epimastigotas de cultivo de T. cruzi: 1) la fracción citoplasmática (antígeno citoplasmático y 2) el parásito total fijado previamente con formaldehido (antígeno integral). Se usaron sueros de: 95 pacientes chagásicos con serología convencional positiva, cardiopatía crónica y algunos con xenodiagnóstico positivo; 42 personas sanas y 32 con miocardipatía crónica con serología negativa y 74 pacientes con diferentes patologías incluyendo: sífilis, toxoplasmosis, lupus eritematoso diseminado, con factor reumatoide, leishmaniasis visceral, y leishmaniasis cutánea. Definidos los títulos diagnósticos (cut-off) de 1:512 con antígeno citoplasmático y de 1: 128 con antígeno integral, la especificidad fue 96% para el primero y de 100% para el segundo; mientras que la sensibilidad fue de 100% para ambas. En el estudio comparativo con las pruebas serológicas convencionales examinando 147 sueros tomados de personas referidas al laboratório, Dot-ELISA con antígeno citoplasmático presentó índices deco-positividad de 1,0, co-negatividad de 0,989 y eficiencia 0,993. Dot-ELIS con antígeno integral dió 1,0, 0,979 y 0,986 respectivamente. De acuerdo con esta evaluación, la técnica Dot-ELISA con antígeno integral se presenta como una alternativa práctica para el diagnóstico serológico de la enfermedad de Chagas.

Effect of cAMP on macromolecule synthesis in the pathogenic protozoa Trypanosoma cruzi

Macromolecule synthesis of Trypanosoma cruzi in culture was monitored using radioactive tracers. Cells of different days in culture displayed a preferential incorporation of precursors as follows: 1 day for (³H)-thymidine cells; 3 days for (³H)-uridine cells, and 4 days for (³H)-leucine cells. Autoradiographic studies showed that (³H)-thymidine was incorporated in the DNA of both kinetoplast and nucleus in this order. Shifts in the intracellular content of cAMP either by addition of dibutyryl-cAMP or by stimulation of the adenylcyclase by isoproterenol, caused an inhibition in the synthesis of DNA, RNA and proteins. Addition to the T. cruzi cultures of these agents which elevate the intracellular content ofcAMP provoked an interruption of cell proliferation as a result of the impairment of macromolecule synthesis. A discrimination was observed among the stereoisomers of isoproterenol, the L configuration showing to be most active.

Morphological evidence by scanning electron microscopy of excretion of metacyclic forms of Trypanosoma cruzi in vector's urine

Comparision by scanning electron microscopy (SEM) of Trypanosoma cruzi flagellates attached to the cuticle of the rectal gland of infected Dipetalogaster maxima nymphs, showed marked differences before amd after feeding. Before feeding numerous metacyclic trypomastigotes were observed among the abundant epimastigotes that formed the carpet of flagellates. On the other hand, in insects that were allowed to urinate for 24 hours after a meal, the metacyclics were scarce,indicating that they had been detached by the urine flow. An asymetric type of cell division, probably originating both an epi-and a trypomastigote, was occasionally observed. The occurrence of swellings at different levels of the flagella of epimastigotes suggests that secondary sites of attachment may be common.