Abstract
BackgroundLeishmaniasis is a serious health problem in some parts of the world. In spite of the many known leishmaniasis control measures, the disease has continued to increase in endemic areas, and no effective vaccine has been discovered.MethodsIn this study, Leishmania tarentulae was used as a living factory for the production of two LACK and KMP11 immunogenic antigens in the mice body, and safety profiles were investigated. The sequences of the KMP11 and LACK L. major antigens were synthesized in the pLEXSY-neo 2.1 plasmid and cloned into E. coli strain Top10, and after being linearized with the SwaI enzyme, they were transfected into the genome of L. tarentolae. The L. tarentolae-LACK/KMP11/EGFP in the stationary phase with CpG ODN as an adjuvant was used for vaccination in BALB/c mice. Vaccination was performed into the left footpad. Three weeks later, the booster was injected in the same manner. To examine the effectiveness of the injected vaccine, pathogenic L. major (MRHO/IR/75/ER) was injected into the right footpad of all mice three weeks following the booster vaccination. In order to assess humoral immunity, the levels of IgG1, and IgG2a antibodies before and 6 weeks after the challenge were studied in the groups. In addition, in order to investigate cellular immunity in the groups, the study measured IFN-γ, IL-5, TNF-α, IL-6 and IL-17 cytokines before, 3 weeks and 8 weeks after the challenge, and also the parasite load in the lymph node with real-time PCR.ResultsThe lowest level of the parasitic load was observed in the G1 group (mice vaccinated with L. tarentolae-LACK/KMP11/EGFP with CpG) in comparison with other groups (L. tarentolae-LACK/KMP11/EGFP +non-CpG (G2); L. tarentolae-EGFP + CpG (G3, control); L. tarentolae-EGFP + non-CpG (G4, control); and mice injected with PBS (G5, control). Moreover, the evaluation of immune response showed a delayed-type hypersensitivity towards Th1.ConclusionsAccording to the results of this study, the live recombinant vaccine of L. tarentolae-LACK/KMP11/EGFP with the CpG adjuvant reduced the parasitic load and footpad induration in infected mice. The long-term effects of this vaccine can be evaluated in volunteers as a clinical trial in future planning.
Highlights
Leishmaniasis is a serious health problem in some parts of the world
Production of recombinant parasites Recombinant parasites L. tarentolae-LACK/Kinetoplastida membrane protein-11 (KMP11)/ EGFP and L. tarentolae-EGFP expressing antigens LACK/KMP11/EGFP and EGFP were constructed by transfecting their linear structure inside the locus of 18S rRNA, L. tarentolae
The sequences of KMP11 and LACK L. major antigens in the form of molecular constructs were fabricated inside the pLEXSY-neo 2.1 vector (Jena Bioscience, Germany) and cloned in E. coli strain Top10, and after linearization with SwaI enzyme, they were stably transfected inside the L. tarentolae genome
Summary
Leishmaniasis is a serious health problem in some parts of the world. In spite of the many known leishmaniasis control measures, the disease has continued to increase in endemic areas, and no effective vaccine has been discovered. According to the WHO report [1], leishmaniasis is a serious health problem in some parts of the world, especially in the Eastern Mediterranean region. Each type can exist in multiple foci within endemic regions. The cases of infection have been increasing over the past 10 years in the aforementioned countries [6]. In 2018, 100,000 new cases of this disease were reported to the WHO from these countries. The cases reported from Iran consisted of 8649 cases of ACL and 18175 cases of ZCL [7,8,9]
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