Abstract
Visceral leishmaniasis (VL) is a vector-borne disease caused by the digenetic protozoan parasite Leishmania donovani complex. So far there is no effective vaccine available against VL. The DDX3 DEAD-box RNA Helicase (Hel67) is 67 kDa protein which is quite essential for RNA metabolism, amastigote differentiation, and infectivity in L. major and L. infantum. To investigate the role of Hel67 in the L. donovani, we created L. donovani deficient in the Hel67. Helicase67 null mutants (LdHel67−/−) were not able to differentiate as axenic amastigotes and were unable to infect the hamster. So, we have analyzed the prophylactic efficacy of the LdHel67−/− null mutant in hamsters. The LdHel67−/− null mutant based candidate vaccine exhibited immunogenic response and a higher degree of protection against L. donovani in comparison to the infected control group. Further, the candidate vaccine displayed antigen-specific delayed-type hypersensitivity (DTH) as well as strong antibody response and NO production which strongly correlates to long term protection of candidate vaccine against the infection. This study confirms the potential of LdHel67−/− null mutant as a safe and protective live attenuated vaccine candidate against visceral leishmaniasis.
Highlights
Visceral Leishmaniasis (VL) is the most deadly disease among the neglected tropical diseases that targets tissue macrophages
Metacyclic LdHel67−/− parasites when used to infect hamster peritoneal macrophages (Fig. 2A) or when injected in susceptible golden Syrian hamsters (Fig. 2B,C), manifested attenuation in growth. This result can be directly correlated with the deficiency of Hel[67], because LdHel67−/− AB expressing Helicase of 67 kDa (HEL67) protein episomally can revert the infectivity in hamster peritoneal macrophages as well as in animal model
We have evaluated the infectivity of LdHel67−/− null mutant in hamsters by suppressing the immune system using cyclophosphamide (CPA) as immunosuppressant and found that LdHel67−/− null mutant parasites never regained any infectivity on days 45, 60 and 90 p.i
Summary
Visceral Leishmaniasis (VL) is the most deadly disease among the neglected tropical diseases that targets tissue macrophages. Up to now several studies using targeted gene deletion strategy have been reported for developing Leishmania-attenuated vaccine strains. Among the vaccine candidates studied for VL, deletion of biopterin transporter (BT1) gene in L. donovani reduced its infectivity in mice and this BT1deleted mutant elicited protective immunity against L. donovani. L. donovani p27 null mutant parasite (Ldp27) exhibited reduced virulence in different in vivo models and reported as nonpathogenic and renders long-term protective immunity in BALB/c mice[12]. To evaluate the role of DEAD Box RNA Helicase 67 in L. donovani, in this study we have constructed DEAD Box RNA Helicase 67 null mutant in L. donovani (LdHel67−/−) and confirmed its prophylactic efficacy as a live attenuated vaccine candidate in hamster model
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