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Evidence that a naturally occurring single nucleotide polymorphism in the RagC gene of Leishmania donovani contributes to reduced virulence.

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.

SUBMITTER: Lypaczewski P

PROVIDER: S-EPMC7901767 | biostudies-literature | 2021 Feb

REPOSITORIES: biostudies-literature

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Evidence that a naturally occurring single nucleotide polymorphism in the RagC gene of Leishmania donovani contributes to reduced virulence.

Lypaczewski Patrick P Zhang Wen-Wei WW Matlashewski Greg G

PLoS neglected tropical diseases 20210223 2

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 disea ...[more]

PMID: 33621241

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Project description:BackgroundLeishmaniasis is a neglected tropical disease with diverse clinical phenotypes, determined by parasite, host and vector interactions. Despite the advances in molecular biology and the availability of more Leishmania genome references in recent years, the association between parasite species and distinct clinical phenotypes remains poorly understood. We present a genomic comparison of an atypical variant of Leishmania donovani from a South Asian focus, where it mostly causes cutaneous form of leishmaniasis.ResultsClinical isolates from six cutaneous leishmaniasis patients (CL-SL); 2 of whom were poor responders to antimony (CL-PR), and two visceral leishmaniasis patients (VL-SL) were sequenced on an Illumina MiSeq platform. Chromosome aneuploidy was observed in both groups but was more frequent in CL-SL. 248 genes differed by 2 fold or more in copy number among the two groups. Genes involved in amino acid use (LdBPK_271940) and energy metabolism (LdBPK_271950), predominated the VL-SL group with the same distribution pattern reflected in gene tandem arrays. Genes encoding amastins were present in higher copy numbers in VL-SL and CL-PR as well as being among predicted pseudogenes in CL-SL. Both chromosome and SNP profiles showed CL-SL and VL-SL to form two distinct groups. While expected heterozygosity was much higher in VL-SL, SNP allele frequency patterns did not suggest potential recent recombination breakpoints. The SNP/indel profile obtained using the more recently generated PacBio sequence did not vary markedly from that based on the standard LdBPK282A1 reference. Several genes previously associated with resistance to antimonials were observed in higher copy numbers in the analysis of CL-PR. H-locus amplification was seen in one cutaneous isolate which however did not belong to the CL-PR group.ConclusionsThe data presented suggests that intra species variations at chromosome and gene level are more likely to influence differences in tropism as well as response to treatment, and contributes to greater understanding of parasite molecular mechanisms underpinning these differences. These findings should be substantiated with a larger sample number and expression/functional studies.

Project description:Virulence, the degree to which a pathogen harms its host, is an important but poorly understood aspect of host-pathogen interactions. Virulence is not static, instead depending on ecological context and potentially evolving rapidly. For instance, at the start of an epidemic, when susceptible hosts are plentiful, pathogens may evolve increased virulence if this maximizes their intrinsic growth rate. However, if host density declines during an epidemic, theory predicts evolution of reduced virulence. Although well-studied theoretically, there is still little empirical evidence for virulence evolution in epidemics, especially in natural settings with native host and pathogen species. Here, we used a combination of field observations and lab assays in the Daphnia-Pasteuria model system to look for evidence of virulence evolution in nature. We monitored a large, naturally occurring outbreak of Pasteuria ramosa in Daphnia dentifera, where infection prevalence peaked at ~ 40% of the population infected and host density declined precipitously during the outbreak. In controlled infections in the lab, lifespan and reproduction of infected hosts was lower than that of unexposed control hosts and of hosts that were exposed but not infected. We did not detect any significant changes in host resistance or parasite infectivity, nor did we find evidence for shifts in parasite virulence (quantified by host lifespan and number of clutches produced by hosts). However, over the epidemic, the parasite evolved to produce significantly fewer spores in infected hosts. While this finding was unexpected, it might reflect previously quantified tradeoffs: parasites in high mortality (e.g., high predation) environments shift from vegetative growth to spore production sooner in infections, reducing spore yield. Future studies that track evolution of parasite spore yield in more populations, and that link those changes with genetic changes and with predation rates, will yield better insight into the drivers of parasite evolution in the wild.Supplementary informationThe online version contains supplementary material available at 10.1007/s10682-022-10169-6.

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Evidence that a naturally occurring single nucleotide polymorphism in the RagC gene of Leishmania donovani contributes to reduced virulence.

Publications

Evidence that a naturally occurring single nucleotide polymorphism in the RagC gene of Leishmania donovani contributes to reduced virulence.

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