Use of an attenuated leishmanial parasite as an immunoprophylactic and immunotherapeutic agent against murine visceral leishmaniasis

Abstract

The ability of the leishmanial parasite UR6 to act as an immunoprophylactic and immunotherapeutic agent against Leishmania donovani infection in BALB/c mice was investigated. Unlike the virulent L. donovani AG83 (MOHOM/IN/1983/AG83), UR6 given through intracardiac route failed to induce visceral infection, but when it was injected subcutaneously, UR6 induced a short-lived and localized self-healing skin lesion. Priming of peritoneal macrophages with UR6 in vitro induced superoxide (O(2)(-)) generation, whereas similar experiments with virulent AG83 inhibited O(2)(-) generation. It was observed that priming of mice with either live or sonicated UR6 in the absence of any adjuvant provided strong protection against subsequent virulent challenge. Further, UR6-primed infected mice not only displayed a strong antileishmanial delayed-type hypersensitivity (DTH) response but also showed an elevated level of the serum antileishmanial immunoglobulin G2a (IgG2a) isotype, whereas infected mice failed to mount any antileishmanial DTH response and showed an elevated level of IgG1. This indicates that UR6 priming and subsequent L. donovani infection allowed the expansion of Th1 cells. Our studies indicate that UR6 has potential to be used as an immunoprophylactic and immunotherapeutic agent against experimental visceral leishmaniasis.

FIG. 1

Parasite burdens in spleens and livers of BALB/c mice in a progressive infection after inoculation with AG83 amastigotes and UR6 promastigotes through the tail vein.

FIG. 2

Skin lesions in BALB/c mice in response to s.c. challenge with live UR6. I, lesion at site of injection; II, healing response.

FIG. 3

Superoxide (O₂⁻) generation (in terms of NBT reduction) in peritoneal Mφs of BALB/c mice. Mφs were stimulated or not with LPS or incubated in presence of UR6 or AG83. + ve, positive. Results are means and SDs.

FIG. 4

Antileishmanial DTH response in the BALB/c mice. I, normal mice; II saline-treated infected mice; III, normal mice primed with 10⁸ UR6 organisms; IV, infected mice primed with 10⁸ UR6 organisms. Sonicated UR6 was given to the primed animals as described in the text. Animals were injected with UR6 on days 0, 15, and 30, and 15 days after the last injection, animals were injected with L. donovani amstigotes.

FIG. 5

Antileishmanial isotypes present in the sera of BALB/c mice.

FIG. 6

TNF-α (A) and nitric oxide (B) induction from BALB/c Mφs stimulated with UR6 or AG83. Released TNF-α and nitric oxide were measured as indicated in the text. (A) TNF-α released by BALB/c Mφs alone (I) or stimulated with UR6 (III) or AG83 (IV) was compared to that released by Mφs stimulated with S. aureus Cowan 1 (1,850 pg/ml) (II) (B) Nitric oxide released by BALB/c Mφs stimulated with AG83 or UR6 was corrected for the background stimulation with Mφs alone. For comparison, the nitric oxide released by LPS-stimulated Mφs was 23.8, 22.0, and 23.7 μM at 12, 18, and 24 h, of incubation, respectively. The data shown represent one experiment of five.