Pulcherrimasaponin, from the leaves of Calliandra pulcherrima, as adjuvant for immunization in the murine model of visceral leishmaniasis.

A novel triterpenoidal saponin, called pulcherrimasaponin (CP05), isolated from the leaves of Calliandra pulcherrima Benth. shows remarkable similarities to the previously described potent adjuvant, QS21 saponin (Quillaja saponaria Molina). On the basis of chemical and physicochemical evidence, its structure was established as [3beta,16alpha,28[2E,6S[2E,6S(2E,6S)]]]-olean-12-en-28-oic acid 3-[[O-alpha-l-arabinopyranosyl-(1-->2)-O-alpha-l-arabinopyranosyl-(1-->6)-2-(acetylamino)-2-deoxy-beta-d-glucopyranosyl]oxy]-16-hydroxy-O-beta-d-glucopyranosyl-(1-->3)-O-[O-beta-d-xylopyranosyl-(1-->3)-beta-d-xylopyranosyl-(1-->4)-O-6-deoxy-alpha-l-mannopyranosyl-(1-->2)-6-O-[6-[[2-O-2,6-dimethyl-1-oxo-6-(beta-d-xylopyranosyloxy)-2,7-octadienyl]-[(6-deoxy-beta-d-glucopyranosyl)oxy]-2,6-dimethyl-1-oxo-2,7-octadienyl]-beta-d-xylopyranosyl]oxy]-2,6-dimethyl-1-oxo-2,7-octadienyl]-beta-d-glucopyranosyl ester. In vivo toxicity assays disclosed similar and transitory local swelling and loss of hair but no lethality for mice. The haemolytic index was higher for QS21 (5 microg/ml) than for CP05 (13 microg/ml). Mouse vaccination with either CP05 or QS21 in combination with the fucose-mannose ligand (FML) antigen of Leishmania donovani showed anti-FML responses, significantly enhanced over the saponin and saline controls, in IgM, IgG, IgG1, IgG2a, IgG2b and IgG3. Antibody levels were similar for both vaccines in most subtypes. However, QS21-FML vaccine showed a 1.5 to 2.1 proportional increase over the CP05-FML vaccine in IgG, IgG2a and IgG3 responses. The delayed type of hypersensitivity against leishmanial antigen was impressively increased for CP05-FML and for QS21-FML-treated animals over controls (p<0.005). Enhancement was similar for both vaccines (p<0.05). The safety analysis and the effect on humoral and cellular immune responses demonstrated that the novel Calliandra pulcherrima Benth. CP05 saponin is a potential candidate for a vaccine adjuvant.

The FML (Fucose Mannose Ligand) of Leishmania donovani: a new tool in diagnosis, prognosis, transfusional control and vaccination against human kala-azar

The Fucose-Mannose Ligand (FML) of Leishmania donovani is a complex glycoproteic fraction. Its potential use as a tool for diagnosis of human visceral leishmaniasis was tested with human sera from Natal, Rio Grande do Norte, Brazil. The FML-ELISA test, showed 100 per cent sensitivity and 96 per cent specificity, identifying patients with overt kala-azar (p < 0.001, when compared to normal sera), and subjects with subclinical infection. More than 20 per cent apparently healthy subjects with positive reaction to FML developed overt kala-azar during the following 10 months. In the screening of human blood donnors, a prevalence of 5 per cent of sororeactive subjects was detected, attaining 17 per cent in a single day. The GP36 glycoprotein of FHL is specifically reconized by human kala-azar sera. The immunoprotective effect of FML on experimental L. donovani infection was tested in swiss albino mice. The protection scheemes included three weekly doses of FML, supplemented or not with saponin by the subcutaneous or intraperitoneal routes and challenge with 2 x 10(7) amastigotes of Leishmania donovani. An enhancement of 80.0 per cent in antibody response (p < 0.001) and reduction of 85.5 per cent parasite liver burden (p < 0.001) was detected in animals immunized with FML saponin, unrespectively of the immunization route.