Studies on Immunological Responses to Parasite Surface Antigen as a Potential Vaccine Candidate against Human Leishmaniasis and Estimation of Asymptomatic Leishmania Infection in the Endemic Areas

At present, there is no licensed vaccine available against any form of human leishmaniasis, although, the development of an anti-Leishmania vaccine has been a longterm goal. Besides, for the sustainable elimination of VL, the presence of asymptomatic VL may prove an important impediment as they could play a vital role in maintaining transmission dynamics of Leishmania infection in the region. Here, we evaluated the recombinant native form of LaPSA protein for its ability to induce cellular and humoral immune responses in active cases of VL and PKDL, healed VL (HVL) and naïve individuals, to assess its potential use as a vaccine candidate against human VL, caused by L. donovani. Further, using serological and molecular tools, we assessed Leishmania parasites in the blood samples of recruited healthy individuals living in two VL-endemic regions (Malda and Murshidabad) of the state of West Bengal, to understand the magnitude of asymptomatic Leishmania infection present in the region. The study demonstrated Leishmania-specific cellular immune responses in terms of lymphoproliferation and cytokine profile upon PBMCs stimulation with Leishmania antigens i.e.; TSLA and LaPSA, in the active cases of VL and PKDL and HVL individuals. Lymphoproliferation in response to TSLA in HVL and PKDL group showed heightened response whereas LaPSA did not induce lymphoproliferation in either of Leishmania pre-exposed groups i.e., HVL or PKDL group. The cytokine profile upon LaPSA and TSLA stimulation corroborated our lymphoproliferation data; LaPSA did not induce significant pro-inflammatory cytokines response in either HVL or PKDL while TSLA elicited a strong protective cytokine response in the same group. Additionally, the significant activation of both CD4 + and CD8 + T cells along with high granzyme B production upon in vitro TSLA stimulation indicated for the first time the role of cytotoxic cells in resistance to L. donovani infection in HVL and PKDL groups. This finding indicated Leishmania-specific cell-mediated cytotoxicity as a part of the naturally acquired immunity developed in Leishmania pre-exposed groups (HVL and PKDL). In contrast and corroborating our cellular data, LaPSA did not induce significant activation in either CD8 + or CD4 + T cells. Generalized cellular immunity based on the phenotype and proportion of the different peripheral blood lymphoid subsets revealed impaired cellular immunity in active VL while PKDL showed raised proportion of CD3 + T cells. Our approach to investigate the ability of a given antigen, for its ability to elicit cellular protective responses in different groups of human populations would be applicable to select vaccine candidates and prioritize antigens for the clinical development of a vaccine against leishmaniasis. The other part of the study identified 22.35% asymptomatic individuals among the enrolled study population living in the two VL endemic districts of West Bengal. These could be the potential reservoir for Leishmania parasite and might play an important role in the disease transmission in the region. Besides, a qPCR method was more sensitive and specific than the serological method in assessment of asymptomatic VL individuals. Additionally, anti-TSLA or anti-LaPSA IgG based detection of asymptomatic infection in healthy individuals may not be a good choice. Thus, deployment of combined molecular and serological methods could effectively help estimate the asymptomatic Leishmania infection in healthy individuals living in endemic regions, and may contribute to early case detection and treatment. Furthermore, the estimation of asymptomatic VL individuals in the endemic area will be useful to take appropriate measures for the sustainable elimination of VL from India.