Abstract
Leishmaniasis is a widespread neglected tropical disease transmitted by infected sand flies resulting in either benign cutaneous infection or fatal visceral disease. Leishmania donovani is the principal species responsible for visceral leishmaniasis, yet an atypical L. donovani has become attenuated in several countries including Sri Lanka and causes cutaneous leishmaniasis. Previous studies have identified 91 genes altered in the atypical cutaneous L. donovani compared to typical visceral disease associated L. donovani including mutations in the RagC and Raptor genes that are part of the eukaryotic conserved TOR pathway and its upstream sensing pathway. In the present study, we investigate whether the RagC R231C mutation present in atypical cutaneous L. donovani introduced into the virulent L. donovani 1S2D chromosome by CRISPR gene editing could affect virulence for survival in visceral organs. Through bioinformatic analysis, we further investigated the presence of sensing pathway components upstream of TOR in L. donovani including RagC complexing proteins, RagA and Raptor. L. donovani 1S2D edited to express mutant RagC R231C were viable in promastigote but had reduced visceral parasitemia in infected BALB/c mice. The RagC R231C mutant retained the ability to interact with RagA and gene knockout experiments revealed that although the RagA gene was essential, the RagC gene was not essential under promastigote culture conditions but was essential for survival in the liver of experimentally infected mice. These results provide evidence that the TOR associated sensing pathway plays a prominent role in L. donovani visceral disease and the RagC R231C mutation contributed to the atypical pathology of cutaneous L. donovani in Sri Lanka.
Highlights
Leishmaniasis is a neglected tropical disease present throughout developing countries and is caused by Leishmania protozoa parasites that are transmitted by infected sandflies [1]
The Leishmania donovani parasite is transmitted by infected sandflies and is the principal causative agent of visceral leishmaniasis, a neglected tropical disease with no available vaccine
This study investigated whether members of the TOR and associated sensing pathways could be identified by bioinformatic analysis and the potential role a RagC mutation plays in the attenuation of L. donovani in Sri Lanka for causing visceral disease
Summary
Leishmaniasis is a neglected tropical disease present throughout developing countries and is caused by Leishmania protozoa parasites that are transmitted by infected sandflies [1]. There are over 20 species of Leishmania that infect humans of which most remain in the dermal layer of the skin at the site of the sand fly bite resulting in cutaneous leishmaniasis that usually selfcures [2]. The Leishmania donovani species typically disseminates from the dermis to the liver, spleen and bone marrow resulting in visceral leishmaniasis with persistent fever, hepatosplenomegaly, internal bleeding, anemia and is fatal if not treated. Humans are the only known reservoir for L. donovani while virtually all cutaneous leishmaniasis causing species have animal reservoirs [3]. What controls disease pathology and tropism during infection remains poorly understood this is largely dependent on the species of Leishmania [4]
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