No abstract

A trypanosome identified as Trypanosoma (Schizotrypanum) dionisii Bettencourt & França, 1905, has been isolated from Pipistrellus pipistrellus (Chiroptera) in England. At least five out of eight P. pipistrelhis were infected. 2. In the blood of P. pipistrelhis, the parasite closely resembled T. (S.) cruzi. When grown in vitro in monophasic or diphasic media at 28°C, epimastigotes and trypomastigotes developed. The latter were of two types — very long, thin forms and less numerous shorter individuals. 3. The trypanosomes multiplied as amastigotes within HeLa and mouse L cells in vitro. After 6–9 days in HeLa cells at 37°C, they transformed into small trypomastigotes and emerged from the cells. Higher infection rates (up to about 4%) were obtained in cell cultures inoculated with flagellates from older monophasic cultures, which contained more of the long, slender trypomastigotes. 4. Trypanosomes also entered (or were phagocytosed by) up to 60% or more of mouse peritoneal macrophages in vitro. Multiplication occurred by binary and multiple fission of amastigotes in at least some of the parasitized macrophages and transformation into trypomastigotes was seen after 7 or more days at 37°C. Development in macrophages was less synchronous than in HeLa cells.

SummaryIsolation of blood and intracellular forms of Trypanosoma cruzi was made mainly from rats (90–110 g) which had received 580 rad of whole-body γ-irradiation not more than 24 h before subcutaneous inoculation with 107 trypomastigotes of the Sonya strain of T. cruzi. Unirradiated chinchillas (250–350 g) were, however, used for some experiments. Blood forms were isolated using a technique involving differential centrifugation to remove most of the erythrocytes and DEAE–cellulose chromatography to remove the remaining blood cells. Overall recoveries were usually in the range 30–70%. Parasites were mainly (approximately 98%) broad forms and were motile, metabolically active (as judged by respiratory and radio-tracer incorporation studies) and had lost none of their infectivity for mice. Intracellular forms were isolated from hind-limb muscle tissue. This was disrupted in an MSE tissue homogenizer and the homogenate incubated with DNase, collagenase and trypsin. Parasites, contaminated only by a few blood cells, were then obtained by differential centrifugation. For purer preparations, a terminal sucrose gradient step was used. Recoveries ranged between 40 and 70%. About 1–3% of the parasites isolated were epimastigotes and trypomastigotes; the remainder are probably best collectively termed ‘amastigotes’, though they were pointed and most had a short, free flagellum. They were undamaged as judged by light and electron microscopy and metabolically active as judged by respiratory and radio-tracer incorporation studies. However, the infectivity for mice of both these purified preparations and the initial cell homogenates could be accounted for by the epimastigotes and trypomastigotes present in them. Preliminary biochemical studies with isolated parasites have shown that blood, intracellular and culture forms of T. cruzi have a respiratory system which is in part sensitive to CN- and that all forms synthesize nucleic acids and proteins when incubated in vitro. There appears, however, to be a lack of DNA synthesis in blood stages, and thus it is not surprising that these forms do not divide.

No abstract

No abstract