K Vickerman

ABSTRACT Pathogenic trypanosomes in their bloodstream phase have a smooth and compact coat 12–15 nm thick enveloping the entire surface membrane of the body and flagellum. In the sleeping-sickness trypanosome Trypanosoma rhodesiense this coat is absent from the stages of development in the midgut of the tsetse-fly vector and from their counterparts obtained by cultivation of the trypanosome in vitro. In the salivary glands of the vector, however, the coat is reacquired as the trypanosomes transform from epimastigote forms into the metacyclic stage which is infective to the mammalian host. This loss and acquisition of the surface coat can be correlated with the cyclical changes in net surface charge on the trypanosome which have been observed by other workers. The trypanosome populations of successive relapses in- the blood are known to differ in their surface antigens (agglutinogens) and the loss of antigenic identity detected when any of these populations are put into culture indicates that these variable antigens are located in the surface coat. It is suggested that the coat in bloodstream trypanosomes constitutes a replaceable surface which, after being replaced, enables the trypanosome to escape the effects of host antibodies. The coat is therefore an adaptation to life in the bloodstream. Reacquisition of the surface coat by the metacyclic trypanosome after development in the vector may reflect reversion to a ‘basic’ antigenic type at this stage, preparatory to invading the blood of the mammalian host. The surface coat may be removed by the wide-spectrum proteolytic enzyme pronase, and this fact together with evidence from pH/mobility relationships and chemical analysis of the variable antigens suggest that the coat is basically proteinaceous. The coat may facilitate pinocytosis by binding proteins at sites within the pocket surrounding the base of the flagellum. In the non-pathogenic trypanosome T. lewisi a more diffuse filamentous coat is present in bloodstream forms and absent from culture forms. This trypanosome is said to carry a negative charge in both bloodstream and culture phases, so it seems likely that the nature of the coat in T. lewisi is different from that found in the pathogenic trypanosomes. In all these trypanosomes the flagellar membrane adheres to the surface membrane of the body throughout the life-cycle. Along the zone of adhesion lies a regular row of junctional complexes of the macula adherens type which, it is argued, serve in attachment. These attachments persist regardless of changes in the intervening cell surfaces.

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A rapid method for the bulk isolation of purified Leishmania mexicana mexicana amastigotes from parasite-induced lesions in experimentally infected mice is described. The procedure includes purification steps based on differences in net cell charge, lysis susceptibility and buoyant density between parasite and host cells. Yields of up to 2 x 10(10) untransformed amastigotes with minimal contamination with host cells and cell debris can be obtained. At least 90% of the purified amastigotes are viable as judged by light and electron microscopy, the staining of their lysosomes with acridine orange, their ability to transform to promastigotes and their infectivity to macrophages in vivo and in vitro.

An agglutination reaction for trypanosomes is described, using whole blood from infected mice as antigen and antisera from infected rabbits. By using antigens frozen down to −79°C. and stored at that temperature, the comparison of large numbers of strains is facilitated. The antigenic analysis of 10 strains of recent isolation has been attempted together with two relapses of one of them. At least six different antigens were recognized, the number of antigens identified in a particular strain varying from one to five. The results, although by no means exhaustive, indicate that not only may strains from man and animals have antigenic components in common, but also successive relapses of a strain in the same host. However, differences have been discerned in these common antigens between animal and human strains and between relapses. Their interpretation is discussed.