Fig. 3. Immunization with PdSP15 protects NHP against vector-transmitted CL. (A to C) Immunity in PdSP15- immunized (PdSP15) or sham-immunized (CTL) NHP 48 hours (A) or 2 weeks (B and C) after last immunization. (A) Skin induration after inoculation with bovine serum albumin (CTL) or rPdSP15 (P = 0.0067, t test; n = 10). (B) IFN-g SFC by ELISPOT (P = 0.0002, t test; n = 10). (C) Anti-saliva IgG levels before (Pre) or after (Post) immunization in controls (CTL), PdSP15immunized NHP producing IFN-g (PdSP15IFN+) or not (PdSP15-IFN−) (P < 0.0001, one-way ANOVA; n = 3 to 10). (D to L) Evaluation of disease (D to H) and Leishmania-specific immunity (I to L) in CTL, PdSP15-IFN+, or PdSP15-IFN− NHP after challenge with 50 infected sand flies. (D) Disease burden (P = 0.0490, one-way ANOVA; n = 3 to 11). (E) Maximum lesion size (P = 0.0465, one-way ANOVA; n = 3 to 11). (F) Kaplan-Meier plot of the healing time [P = 0.1770, log-rank (MantelCox) test; n = 3 to 11]. (G) Representative photographs 5 weeks after challenge. (H) Parasite number 5 weeks after challenge (P = 0.0034, one-way ANOVA; n = 3 to 8). (I to K) PBMCs stimulated with Leishmania antigen (Leish) 2 weeks after challenge in 8 to 10 NHP. Selection was based on cell number and viability. (I) IFN-g SFC by ELISPOT (P = 0.0075, one-way ANOVA; n = 3 to 10). (J) Percent of CD4+IFN-g+ lymphocytes by flow cytometry (P = 0.0002, one-way ANOVA; n = 3 to 10). (K) Frequency of CD4+ lymphocytes producing cytokines (P = 0.0418, one-way ANOVA; n = 4 to 6). (L) LST induration size 48 hours after the injection of Leishmania antigen at 12 weeks after challenge (P = 0.0269, oneway ANOVA; n = 3 to 10). Cumulative data for 11 CTL and 10 PdSP15 NHP from two independent experiments are shown. Lines and bars indicate the mean, and error bars indicate SEM.
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