Dataset Information

Infectivity and Drug Susceptibility Profiling of Different Leishmania-Host Cell Combinations.

ABSTRACT: Protozoan parasites of the genus Leishmania are the causative agents of leishmaniasis, a spectrum of a disease that threatens public health worldwide. Although next-generation therapeutics are urgently needed, the early stage of the drug discovery process is hampered by very low hit rates from intracellular Leishmania phenotypic high-throughput screenings. Designing and applying a physiologically relevant in vitro assay is therefore in high demand. In this study, we characterized the infectivity, morphology, and drug susceptibility of different Leishmania and host cell infection combinations. Primary bone marrow-derived macrophage (BMDM) and differentiated human acute monocytic leukemia (THP-1) cells were infected with amastigote or promastigote forms of Leishmania amazonensis and Leishmania donovani. Regardless of host cell types, amastigotes were generally well phagocytosed and showed high infectivity, whereas promastigotes, especially those of L. donovani, had predominantly remained in the extracellular space. In the drug susceptibility test, miltefosine and sodium stibogluconate (SSG) showed varying ranges of activity with 14 and >10-fold differences in susceptibility, depending on the host-parasite pairs, indicating the importance of assay conditions for evaluating antileishmanial activity. Overall, our results suggest that combinations of Leishmania species, infection forms, and host cells must be carefully optimized to evaluate the activity of potential therapeutic compounds against Leishmania.

SUBMITTER: Baek KH

PROVIDER: S-EPMC7281264 | biostudies-literature | 2020 May

REPOSITORIES: biostudies-literature

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Infectivity and Drug Susceptibility Profiling of Different Leishmania-Host Cell Combinations.

Baek Kyung-Hwa KH Piel Laura L Rosazza Thibault T Prina Eric E Späth Gerald F GF No Joo Hwan JH

Pathogens (Basel, Switzerland) 20200520 5

Protozoan parasites of the genus Leishmania are the causative agents of leishmaniasis, a spectrum of a disease that threatens public health worldwide. Although next-generation therapeutics are urgently needed, the early stage of the drug discovery process is hampered by very low hit rates from intracellular Leishmania phenotypic high-throughput screenings. Designing and applying a physiologically relevant in vitro assay is therefore in high demand. In this study, we characterized t ...[more]

PMID: 32443883

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Project description:BackgroundGenome sequencing of Leishmania species that give rise to a range of disease phenotypes in the host has revealed highly conserved gene content and synteny across the genus. Only a small number of genes are differentially distributed between the three species sequenced to date, L. major, L. infantum and L. braziliensis. It is not yet known how many of these genes are expressed in the disease-promoting intracellular amastigotes of these species or whether genes conserved between the species are differentially expressed in the host.Methods/principal findingsWe have used customised oligonucleotide microarrays to confirm that all of the differentially distributed genes identified by genome comparisons are expressed in intracellular amastigotes, with only a few of these subject to regulation at the RNA level. In the first large-scale study of gene expression in L. braziliensis, we show that only approximately 9% of the genes analysed are regulated in their RNA expression during the L. braziliensis life cycle, a figure consistent with that observed in other Leishmania species. Comparing amastigote gene expression profiles between species confirms the proposal that Leishmania transcriptomes undergo little regulation but also identifies conserved genes that are regulated differently between species in the host. We have also investigated whether host immune competence influences parasite gene expression, by comparing RNA expression profiles in L. major amastigotes derived from either wild-type (BALB/c) or immunologically compromised (Rag2(-/-) gamma(c) (-/-)) mice. While parasite dissemination from the site of infection is enhanced in the Rag2(-/-) gamma(c) (-/-) genetic background, parasite RNA expression profiles are unperturbed.Conclusion/significanceThese findings support the hypothesis that Leishmania amastigotes are pre-adapted for intracellular survival and undergo little dynamic modulation of gene expression at the RNA level. Species-specific parasite factors contributing to virulence and pathogenicity in the host may be limited to the products of a small number of differentially distributed genes or the differential regulation of conserved genes, either of which are subject to translational and/or post-translational controls.

Project description:BackgroundParasites of the genus Leishmania are the causative agents of leishmaniasis, a group of diseases that range in manifestations from skin lesions to fatal visceral disease. The life cycle of Leishmania parasites is split between its insect vector and its mammalian host, where it resides primarily inside of macrophages. Once intracellular, Leishmania parasites must evade or deactivate the host's innate and adaptive immune responses in order to survive and replicate.ResultsWe performed transcriptome profiling using RNA-seq to simultaneously identify global changes in murine macrophage and L. major gene expression as the parasite entered and persisted within murine macrophages during the first 72 h of an infection. Differential gene expression, pathway, and gene ontology analyses enabled us to identify modulations in host and parasite responses during an infection. The most substantial and dynamic gene expression responses by both macrophage and parasite were observed during early infection. Murine genes related to both pro- and anti-inflammatory immune responses and glycolysis were substantially upregulated and genes related to lipid metabolism, biogenesis, and Fc gamma receptor-mediated phagocytosis were downregulated. Upregulated parasite genes included those aimed at mitigating the effects of an oxidative response by the host immune system while downregulated genes were related to translation, cell signaling, fatty acid biosynthesis, and flagellum structure.ConclusionsThe gene expression patterns identified in this work yield signatures that characterize multiple developmental stages of L. major parasites and the coordinated response of Leishmania-infected macrophages in the real-time setting of a dual biological system. This comprehensive dataset offers a clearer and more sensitive picture of the interplay between host and parasite during intracellular infection, providing additional insights into how pathogens are able to evade host defenses and modulate the biological functions of the cell in order to survive in the mammalian environment.

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Infectivity and Drug Susceptibility Profiling of Different Leishmania-Host Cell Combinations.

Publications

Infectivity and Drug Susceptibility Profiling of Different <i>Leishmania</i>-Host Cell Combinations.

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