Expression of the major surface glycoprotein of Leishmania donovani chagasi in virulent and attenuated promastigotes.

Using both hamster and mouse models of infection, we documented that the virulence of Leishmania donovani chagasi promastigotes decreases over time, when parasites are maintained in long term culture after isolation from an infected animal. Concomitant with this loss of virulence is a marked decrease in amount of the major promastigote surface glycoprotein, gp63, present in promastigotes. The latter was shown by a decrease in binding of polyclonal anti-gp63 serum to attenuated (cultivated long term) as compared to virulent (recently isolated) promastigotes, using immunofluorescence and Western blot assays. Binding of Con A to promastigote glycoproteins, separated by SDS-PAGE, documented a similar decrease. An alteration in the mechanism of promastigote attachment to macrophages was also noted: purified gp63 inhibited attachment of virulent promastigotes to human monocyte-derived macrophages, but it did not affect the attachment of attenuated promastigotes. Northern blot analysis showed that, despite marked differences in the amount of gp63 protein, the quantity of gp63 RNA was comparable in attenuated and virulent promastigotes. However, virulent promastigotes contained two major gp63 RNA species of 3.0 and 2.7 kb, whereas attenuated promastigotes had one predominant gp63 RNA of 2.7 kb and only minor amounts of 3.0 kb RNA. Thus, the decrease in gp63 expression in attenuated, contrasted to virulent, promastigotes is associated with qualitative, but not quantitative, differences in the gp63 messenger RNA.