Genomic and Phenotypic Characterization of Experimentally Selected Resistant Leishmania donovani Reveals a Role for Dynamin-1-Like Protein in the Mechanism of Resistance to a Novel Antileishmanial Compound

Hefnawy, Aya; Negreira, Gabriel H.; Jara, Marlene; Cotton, James A.; Maes, Ilse; D’haenens, Erika; Imamura, Hideo; Cuypers, Bart; Monsieurs, Pieter; Mouchtoglou, Christina; Winter, Hans De; Pintelon, Isabel; Timmermans, Jean‐Pierre; Berriman, Matt; Sanders, Mandy; Martín, Julio; Muylder, Géraldine De; Dujardin, Jean Claude; Sterckx, Yann G.J.; Domagalska, Malgorzata A.

doi:10.1128/mbio.03264-21

Cited by 8 publications

(7 citation statements)

References 42 publications

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“…Although the growth curves of both WT strain and LiR line exhibit logarithmic and stationary phases, the LiR line showed significant lower growth rate than the WT strain ( Figure 1 ). These results agree with the decrease in the growth rate observed in promastigotes of L. infantum strains selected for miltefosine resistance as well as in other Leishmania species selected for resistance to different drugs [ 31 , 32 , 50 ]. This phenotype has been associated with a decreased fitness of the resistant parasites.…”

Section: Resultssupporting

confidence: 87%

In-Depth Quantitative Proteomics Characterization of In Vitro Selected Miltefosine Resistance in Leishmania infantum

et al. 2022

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Visceral leishmaniasis (VL) is a neglected disease caused by Leishmania parasites. Although significant morbidity and mortality in tropical and subtropical regions of the world are associated with VL, the low investment for developing new treatment measures is chronic. Moreover, resistance and treatment failure are increasing for the main medications, but the emergence of resistance phenotypes is poorly understood at the protein level. Here, we analyzed the development of resistance to miltefosine upon experimental selection in a L. infantum strain. Time to miltefosine resistance emergence was ~six months and label-free quantitative mass-spectrometry-based proteomics analyses revealed that this process involves a remodeling of components of the membrane and mitochondrion, with significant increase in oxidative phosphorylation complexes, particularly on complex IV and ATP synthase, accompanied by increased energy metabolism mainly dependent on β-oxidation of fatty acids. Proteins canonically involved in ROS detoxification did not contribute to the resistant process whereas sterol biosynthesis enzymes could have a role in this development. Furthermore, changes in the abundance of proteins known to be involved in miltefosine resistance such as ABC transporters and phospholipid transport ATPase were detected. Together, our data show a more complete picture of the elements that make up the miltefosine resistance phenotype in L. infantum.

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Section: Resultssupporting

confidence: 87%

In-Depth Quantitative Proteomics Characterization of In Vitro Selected Miltefosine Resistance in Leishmania infantum

et al. 2022

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“…In each MePOP a different profile was seen, although 3 out of 4 shared the amplification of chromosome 31, and two MePOPs displayed a dosage increase in chromosome 23. The fact that an increase in copy number of chromosome 31 was observed under strong Sb III and miltefosine pressure, as well as under pressure of other drugs (Hefnawy et al , 2022 ) might indicate that the dosage increase in this chromosome has also a general role against multiple types of stresses. Noteworthy, there are several ABC transporters in that chromosome (ABCC4‐7 and ABCD3) which could play a role in drug efflux (Leprohon et al , 2006 ).…”

Section: Resultsmentioning

confidence: 99%

The adaptive roles of aneuploidy and polyclonality in Leishmania in response to environmental stress

et al. 2023

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Aneuploidy is generally considered harmful, but in some microorganisms, it can act as an adaptive mechanism against environmental stress. Here, we use Leishmania—a protozoan parasite with remarkable genome plasticity—to study the early steps of aneuploidy evolution under high drug pressure (using antimony or miltefosine as stressors). By combining single‐cell genomics, lineage tracing with cellular barcodes, and longitudinal genome characterization, we reveal that aneuploidy changes under antimony pressure result from polyclonal selection of pre‐existing karyotypes, complemented by further and rapid de novo alterations in chromosome copy number along evolution. In the case of miltefosine, early parasite adaptation is associated with independent point mutations in a miltefosine transporter gene, while aneuploidy changes only emerge later, upon exposure to increased drug levels. Therefore, polyclonality and genome plasticity are hallmarks of parasite adaptation, but the scenario of aneuploidy dynamics depends on the nature and strength of the environmental stress as well as on the existence of other pre‐adaptive mechanisms.

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“…In the present study, we have applied multiple technologies (i) to understand how environmental stresses, in particular high drug-pressure, promote changes in aneuploidy in vitro, and (ii) to assess the evolutionary dynamics of these aneuploidy changes. Our flash selection models were quite different from conventional drug resistance selection experiments in which parasites are progressively submitted to increasing concentrations of drug (22)(23)(24). In our Sb III flash selection model, we consistently observed drastic changes in aneuploidy emerging in a short period of ~3 weeks, affecting multiple chromosomes, and with different karyotypic outcomes between experimental replicates and repetitions.…”

Section: Discussionmentioning

confidence: 61%

“…At 100 µM, aneuploidy changes were specific to each of the 4 MePOP replicates, with the exception of chromosome 31 that consistently showed a higher somy than the control. The fact that an increase in copy number of chromosome 31 was observed under strong Sb III and miltefosine pressure, as well as under pressure of other drugs (23) might indicate that the dosage increase in this chromosome has a general role against multiple types of stresses.…”

Section: Discussionmentioning

confidence: 99%

The role of aneuploidy and polyclonality in the adaptation of the Protozoan parasiteLeishmaniato high drug pressure

Negreira

Groote

Giel

et al. 2022

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Aneuploidy is generally considered harmful, but in some microorganisms, it can act as an adaptive mechanism against environmental stresses. Here, we usedLeishmania-a protozoan parasite with a remarkable genome plasticity- to study the early evolution of aneuploidy under high drug pressure (antimony or miltefosine) as stressor model. By combining single-cell genomics, lineage tracing with cellular barcodes and longitudinal genome characterization, we revealed that antimony-induced aneuploidy changes result from the polyclonal selection of pre-existing karyotypes, complemented by further and rapid de novo alterations in chromosome copy number along evolution. In the case of miltefosine, early parasite adaptation was associated with independent pre-existing point mutations in a miltefosine transporter gene and aneuploidy changes only emerged later, upon exposure to increased concentration of the drug. Thus, polyclonality and genome plasticity are hallmarks of parasite adaptation, but the scenario of aneuploidy dynamics is dependent on the nature and strength of the environmental stress as well as on the existence of other pre-adaptive mechanisms.

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Genomic and Phenotypic Characterization of Experimentally Selected Resistant Leishmania donovani Reveals a Role for Dynamin-1-Like Protein in the Mechanism of Resistance to a Novel Antileishmanial Compound

Cited by 8 publications

References 42 publications

In-Depth Quantitative Proteomics Characterization of In Vitro Selected Miltefosine Resistance in Leishmania infantum

In-Depth Quantitative Proteomics Characterization of In Vitro Selected Miltefosine Resistance in Leishmania infantum

The adaptive roles of aneuploidy and polyclonality in Leishmania in response to environmental stress

The role of aneuploidy and polyclonality in the adaptation of the Protozoan parasiteLeishmaniato high drug pressure

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