@article{vaccines9030270,

title = {HLA-DRB1 Alleles Associated with Lower Leishmaniasis Susceptibility Share Common Amino Acid Polymorphisms and Epitope Binding Repertoires},

author = {Nicky de Vrij and Pieter Meysman and Sofie Gielis and Wim Adriaensen and Kris Laukens and Bart Cuypers},

url = {https://www.mdpi.com/2076-393X/9/3/270},

doi = {10.3390/vaccines9030270},

issn = {2076-393X},

year  = {2021},

date = {2021-01-01},

journal = {Vaccines},

volume = {9},

number = {3},

abstract = {Susceptibility for leishmaniasis is largely dependent on host genetic and immune factors. Despite the previously described association of human leukocyte antigen (HLA) gene cluster variants as genetic susceptibility factors for leishmaniasis, little is known regarding the mechanisms that underpin these associations. To better understand this underlying functionality, we first collected all known leishmaniasis-associated HLA variants in a thorough literature review. Next, we aligned and compared the protection- and risk-associated HLA-DRB1 allele sequences. This identified several amino acid polymorphisms that distinguish protection- from risk-associated HLA-DRB1 alleles. Subsequently, T cell epitope binding predictions were carried out across these alleles to map the impact of these polymorphisms on the epitope binding repertoires. For these predictions, we used epitopes derived from entire proteomes of multiple Leishmania species. Epitopes binding to protection-associated HLA-DRB1 alleles shared common binding core motifs, mapping to the identified HLA-DRB1 amino acid polymorphisms. These results strongly suggest that HLA polymorphism, resulting in differential antigen presentation, affects the association between HLA and leishmaniasis disease development. Finally, we established a valuable open-access resource of putative epitopes. A set of 14 HLA-unrestricted strong-binding epitopes, conserved across species, was prioritized for further epitope discovery in the search for novel subunit-based vaccines.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{Hefnawy2021.01.05.425522,

title = {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 anti-leishmanial compound},

author = {Aya Hefnawy and Gabriel Negreira and Marlene Jara and James A Cotton and Ilse Maes and Erika D{textquoteright} Haenens and Hideo Imamura and Bart Cuypers and Pieter Monsieurs and Christina Mouchtoglou and Hans De Winter and Matt Berriman and Mandy Sanders and Julio Martin and Geraldine de Muylder and Jean-Claude Dujardin and Yann G -J Sterckx and Malgorzata Anna Domagalska},

url = {https://www.biorxiv.org/content/early/2021/01/06/2021.01.05.425522},

doi = {10.1101/2021.01.05.425522},

year  = {2021},

date = {2021-01-01},

journal = {bioRxiv},

publisher = {Cold Spring Harbor Laboratory},

abstract = {The implementation of prospective drug resistance (DR) studies in the R\&D pipelines is a common practice for many infectious diseases, but not for Neglected Tropical Diseases. Here, we explored and demonstrated the importance of this approach, using as paradigms Leishmania donovani, the etiological agent of Visceral Leishmaniasis (VL), and TCMDC-143345, a promising compound of the GSK textquoteleftLeishboxtextquoteright to treat VL. We experimentally selected resistance to TCMDC-143345 in vitro and characterized resistant parasites at genomic and phenotypic levels. We found that it took more time to develop resistance to TCMDC-143345 than to other drugs in clinical use and that there was no cross resistance to these drugs, suggesting a new and unique mechanism. By whole genome sequencing, we found two mutations in the gene encoding the L. donovani dynamin-1-like protein (LdoDLP1) that were fixed at highest drug pressure. Through phylogenetic analysis, we identified LdoDLP1 as a family member of the dynamin-related proteins, a group of proteins that impacts the shapes of biological membranes by mediating fusion and fission events, with a putative role in mitochondrial fission. We found that L. donovani lines genetically engineered to harbor the two identified LdoDLP1 mutations were resistant to TCMDC-143345 and displayed altered mitochondrial properties. By homology modeling, we showed how the two LdoDLP1 mutations may influence protein structure and function. Taken together, our data reveal a clear involvement of LdoDLP1 in the adaptation/resistance of L. donovani to TCMDC-143345.Importance Humans and their pathogens are continuously locked in a molecular arms race during which the eventual emergence of pathogen drug resistance (DR) seems inevitable. For neglected tropical diseases (NTDs), DR is generally studied retrospectively, once it has already been established in clinical settings. We previously recommended to keep one step ahead in the host-pathogen arms race and implement prospective DR studies in the R\&D pipeline, a common practice for many infectious diseases, but not for NTDs. Here, using Leishmania donovani, the etiological agent of Visceral Leishmaniasis (VL), and TCMDC-143345, a promising compound of the GSK textquoteleftLeishboxtextquoteright to treat VL, as paradigms, we experimentally selected resistance to the compound and proceeded to genomic and phenotypic characterization of DR parasites. The results gathered in the present study suggest a new DR mechanism involving the L. donovani dynamin-1 like protein (LdoDLP1) and demonstrate the practical relevance of prospective DR studies.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{Adriaensen2020,

title = {Host transcriptomic signature as alternative test-of-cure in visceral leishmaniasis patients co-infected with HIV},

author = {Wim Adriaensen and Bart Cuypers and Carlota F Cordero and Bewketu Mengasha and S\'{e}verine Blesson and Lieselotte Cnops and Paul M Kaye and Fabiana Alves and Ermias Diro and Johan van Griensven},

url = {https://doi.org/10.1016/j.ebiom.2020.102748},

doi = {10.1016/j.ebiom.2020.102748},

issn = {2352-3964},

year  = {2020},

date = {2020-05-01},

journal = {EBioMedicine},

volume = {55},

publisher = {Elsevier},

abstract = {BackgroundVisceral leishmaniasis (VL) treatment in HIV patients very often fails and is followed by high relapse and case-fatality rates. Hence, treatment efficacy assessment is imperative but based on invasive organ aspiration for parasite detection. In the search of a less-invasive alternative and because the host immune response is pivotal for treatment outcome in immunocompromised VL patients, we studied changes in the whole blood transcriptional profile of VL-HIV patients during treatment.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{microorganisms8081252,

title = {The Absence of C-5 DNA Methylation in Leishmania donovani Allows DNA Enrichment from Complex Samples},

author = {Bart Cuypers and Franck Dumetz and Pieter Meysman and Kris Laukens and G\'{e}raldine De Muylder and Jean-Claude Dujardin and Malgorzata Anna Domagalska},

url = {https://www.mdpi.com/2076-2607/8/8/1252},

doi = {10.3390/microorganisms8081252},

issn = {2076-2607},

year  = {2020},

date = {2020-01-01},

journal = {Microorganisms},

volume = {8},

number = {8},

abstract = {Cytosine C5 methylation is an important epigenetic control mechanism in a wide array of eukaryotic organisms and generally carried out by proteins of the C-5 DNA methyltransferase family (DNMTs). In several protozoans, the status of this mechanism remains elusive, such as in Leishmania, the causative agent of the disease leishmaniasis in humans and a wide array of vertebrate animals. In this work, we showed that the Leishmania donovani genome contains a C-5 DNA methyltransferase (DNMT) from the DNMT6 subfamily, whose function is still unclear, and verified its expression at the RNA level. We created viable overexpressor and knock-out lines of this enzyme and characterized their genome-wide methylation patterns using whole-genome bisulfite sequencing, together with promastigote and amastigote control lines. Interestingly, despite the DNMT6 presence, we found that methylation levels were equal to or lower than 0.0003% at CpG sites, 0.0005% at CHG sites, and 0.0126% at CHH sites at the genomic scale. As none of the methylated sites were retained after manual verification, we conclude that there is no evidence for DNA methylation in this species. We demonstrated that this difference in DNA methylation between the parasite (no detectable DNA methylation) and the vertebrate host (DNA methylation) allowed enrichment of parasite vs. host DNA using methyl-CpG-binding domain columns, readily available in commercial kits. As such, we depleted methylated DNA from mixes of Leishmania promastigote and amastigote DNA with human DNA, resulting in average Leishmania:human enrichments from 62\× up to 263\×. These results open a promising avenue for unmethylated DNA enrichment as a pre-enrichment step before sequencing Leishmania clinical samples.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{Dumetze00548-17,

title = {Molecular Preadaptation to Antimony Resistance in Leishmania donovani on the Indian Subcontinent},

author = {F Dumetz and B Cuypers and H Imamura and D Zander and E D{textquoteright}Haenens and I Maes and M A Domagalska and J Clos and J -C Dujardin and G De Muylder},

editor = {Margaret Phillips},

url = {https://msphere.asm.org/content/3/2/e00548-17},

doi = {10.1128/mSphere.00548-17},

year  = {2018},

date = {2018-01-01},

journal = {mSphere},

volume = {3},

number = {2},

publisher = {American Society for Microbiology Journals},

abstract = {Antimonials (Sb) were used for decades for chemotherapy of visceral leishmaniasis (VL). Now abandoned in the Indian subcontinent (ISC) because of Leishmania donovani resistance, this drug offers a unique model for understanding drug resistance dynamics. In a previous phylogenomic study, we found two distinct populations of L. donovani: the core group (CG) in the Gangetic plains and ISC1 in the Nepalese highlands. Sb resistance was only encountered within the CG, and a series of potential markers were identified. Here, we analyzed the development of resistance to trivalent antimonials (SbIII) upon experimental selection in ISC1 and CG strains. We observed that (i) baseline SbIII susceptibility of parasites was higher in ISC1 than in the CG, (ii) time to SbIII resistance was higher for ISC1 parasites than for CG strains, and (iii) untargeted genomic and metabolomic analyses revealed molecular changes along the selection process: these were more numerous in ISC1 than in the CG. Altogether these observations led to the hypothesis that CG parasites are preadapted to SbIII resistance. This hypothesis was experimentally confirmed by showing that only wild-type CG strains could survive a direct exposure to the maximal concentration of SbIII. The main driver of this preadaptation was shown to be MRPA, a gene involved in SbIII sequestration and amplified in an intrachromosomal amplicon in all CG strains characterized so far. This amplicon emerged around 1850 in the CG, well before the implementation of antimonials for VL chemotherapy, and we discuss here several hypotheses of selective pressure that could have accompanied its emergence.IMPORTANCE The textquotedblleftantibiotic resistance crisistextquotedblright is a major challenge for scientists and medical professionals. This steady rise in drug-resistant pathogens also extends to parasitic diseases, with antimony being the first anti-Leishmania drug that fell in the Indian subcontinent (ISC). Leishmaniasis is a major but neglected infectious disease with limited therapeutic options. Therefore, understanding how parasites became resistant to antimonials is of commanding importance. In this study, we experimentally characterized the dynamics of this resistance acquisition and show for the first time that some Leishmania populations of the ISC were preadapted to antimony resistance, likely driven by environmental factors or by drugs used in the 19th century.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{Cuypers2017,

title = {Multiplexed Spliced-Leader Sequencing: A high-throughput, selective method for RNA-seq in Trypanosomatids},

author = {Bart Cuypers and Malgorzata A Domagalska and Pieter Meysman and G\'{e}raldine de Muylder and Manu Vanaerschot and Hideo Imamura and Franck Dumetz and Thomas Wolf Verdonckt and Peter J Myler and Gowthaman Ramasamy and Kris Laukens and Jean-Claude Dujardin},

url = {https://doi.org/10.1038/s41598-017-03987-0},

doi = {10.1038/s41598-017-03987-0},

issn = {2045-2322},

year  = {2017},

date = {2017-06-16},

journal = {Scientific Reports},

volume = {7},

number = {1},

pages = {3725},

abstract = {High throughput sequencing techniques are poorly adapted for in vivo studies of parasites, which require prior in vitro culturing and purification. Trypanosomatids, a group of kinetoplastid protozoans, possess a distinctive feature in their transcriptional mechanism whereby a specific Spliced Leader (SL) sequence is added to the 5textasciiacutexend of each mRNA by trans-splicing. This allows to discriminate Trypansomatid RNA from mammalian RNA and forms the basis of our new multiplexed protocol for high-throughput, selective RNA-sequencing called SL-seq. We provided a proof-of-concept of SL-seq in Leishmania donovani, the main causative agent of visceral leishmaniasis in humans, and successfully applied the method to sequence Leishmania mRNA directly from infected macrophages and from highly diluted mixes with human RNA. mRNA profiles obtained with SL-seq corresponded largely to those obtained from conventional poly-A tail purification methods, indicating both enumerate the same mRNA pool. However, SL-seq offers additional advantages, including lower sequencing depth requirements, fast and simple library prep and high resolution splice site detection. SL-seq is therefore ideal for fast and massive parallel sequencing of parasite transcriptomes directly from host tissues. Since SLs are also present in Nematodes, Cnidaria and primitive chordates, this method could also have high potential for transcriptomics studies in other organisms.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{Dumetze00599-17,

title = {Modulation of Aneuploidy in Leishmania donovani during Adaptation to Different In Vitro and In Vivo Environments and Its Impact on Gene Expression},

author = {F Dumetz and H Imamura and M Sanders and V Seblova and J Myskova and P Pescher and M Vanaerschot and C J Meehan and B Cuypers and G De Muylder and G F Sp\"{a}th and G Bussotti and J R Vermeesch and M Berriman and J A Cotton and P Volf and J C Dujardin and M A Domagalska},

editor = {Keith Gull},

url = {https://mbio.asm.org/content/8/3/e00599-17},

doi = {10.1128/mBio.00599-17},

year  = {2017},

date = {2017-01-01},

journal = {mBio},

volume = {8},

number = {3},

publisher = {American Society for Microbiology},

abstract = {Aneuploidy is usually deleterious in multicellular organisms but appears to be tolerated and potentially beneficial in unicellular organisms, including pathogens. Leishmania, a major protozoan parasite, is emerging as a new model for aneuploidy, since in vitro-cultivated strains are highly aneuploid, with interstrain diversity and intrastrain mosaicism. The alternation of two life stages in different environments (extracellular promastigotes and intracellular amastigotes) offers a unique opportunity to study the impact of environment on aneuploidy and gene expression. We sequenced the whole genomes and transcriptomes of Leishmania donovani strains throughout their adaptation to in vivo conditions mimicking natural vertebrate and invertebrate host environments. The nucleotide sequences were almost unchanged within a strain, in contrast to highly variable aneuploidy. Although high in promastigotes in vitro, aneuploidy dropped significantly in hamster amastigotes, in a progressive and strain-specific manner, accompanied by the emergence of new polysomies. After a passage through a sand fly, smaller yet consistent karyotype changes were detected. Changes in chromosome copy numbers were correlated with the corresponding transcript levels, but additional aneuploidy-independent regulation of gene expression was observed. This affected stage-specific gene expression, downregulation of the entire chromosome 31, and upregulation of gene arrays on chromosomes 5 and 8. Aneuploidy changes in Leishmania are probably adaptive and exploited to modulate the dosage and expression of specific genes; they are well tolerated, but additional mechanisms may exist to regulate the transcript levels of other genes located on aneuploid chromosomes. Our model should allow studies of the impact of aneuploidy on molecular adaptations and cellular fitness.IMPORTANCE Aneuploidy is usually detrimental in multicellular organisms, but in several microorganisms, it can be tolerated and even beneficial. Leishmania\textemdasha protozoan parasite that kills more than 30,000 people each year\textemdashis emerging as a new model for aneuploidy studies, as unexpectedly high levels of aneuploidy are found in clinical isolates. Leishmania lacks classical regulation of transcription at initiation through promoters, so aneuploidy could represent a major adaptive strategy of this parasite to modulate gene dosage in response to stressful environments. For the first time, we document the dynamics of aneuploidy throughout the life cycle of the parasite, in vitro and in vivo. We show its adaptive impact on transcription and its interaction with regulation. Besides offering a new model for aneuploidy studies, we show that further genomic studies should be done directly in clinical samples without parasite isolation and that adequate methods should be developed for this.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{10.1371/journal.pone.0180532,

title = {Macromolecular biosynthetic parameters and metabolic profile in different life stages of Leishmania braziliensis: Amastigotes as a functionally less active stage},

author = {Marlene Jara and Maya Berg and Guy Caljon and Geraldine de Muylder and Bart Cuypers and Denis Castillo and Ilse Maes and Mar\'{i}a Carmen del Orozco and Manu Vanaerschot and Jean-Claude Dujardin and Jorge Arevalo},

url = {https://doi.org/10.1371/journal.pone.0180532},

doi = {10.1371/journal.pone.0180532},

year  = {2017},

date = {2017-01-01},

journal = {PLOS ONE},

volume = {12},

number = {7},

pages = {1-22},

publisher = {Public Library of Science},

abstract = {It was recently hypothesized that Leishmania amastigotes could constitute a semi-quiescent stage characterized by low replication and reduced metabolic activity. This concept developed with Leishmania (Leishmania) mexicana and Leishmania (Leishmania) major models might explain numerous clinical and sub-clinical features of Leishmania (Viannia) braziliensis infections, like reactivation of the disease, non-response to chemotherapy or asymptomatic infections. We compared here in vitro the proliferative capability of L. (V.) braziliensis amastigotes and promastigotes, assessed the expression of key molecular parameters and performed metabolomic analysis. We found that contrary to the highly proliferative promastigotes, amastigotes (axenic and intracellular) do not show evidence of extensive proliferation. In parallel, amastigotes showed a significant decrease of (i) the kDNA mini-circle abundance, (ii) the intracellular ATP level, (iii) the ribosomal components: rRNA subunits 18S and 28S α and ribosomal proteins RPS15 and RPL19, (iv) total RNA and protein levels. An untargeted metabolomic study identified clear differences between the different life stages: in comparison to logarithmic promastigotes, axenic amastigotes showed (a) a strong decrease of 14 essential and non-essential amino acids and eight metabolites involved in polyamine synthesis, (b) extensive changes in the phospholipids composition and (c) increased levels of several endogenous and exogenous sterols. Altogether, our results show that L. (V.) braziliensis amastigotes can show a phenotype with negligible rate of proliferation, a lower capacity of biosynthesis, a reduced bio-energetic level and a strongly altered metabolism. Our results pave the way for further exploration of quiescence among amastigotes of this species.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{10.1371/journal.pone.0154101,

title = {Genomic and Molecular Characterization of Miltefosine Resistance in Leishmania infantum Strains with Either Natural or Acquired Resistance through Experimental Selection of Intracellular Amastigotes},

author = {Annelies Mondelaers and Maria P Sanchez-Ca\~{n}ete and Sarah Hendrickx and Eline Eberhardt and Raquel Garcia-Hernandez and Laurence Lachaud and James Cotton and Mandy Sanders and Bart Cuypers and Hideo Imamura and Jean-Claude Dujardin and Peter Delputte and Paul Cos and Guy Caljon and Francisco Gamarro and Santiago Castanys and Louis Maes},

url = {https://doi.org/10.1371/journal.pone.0154101},

doi = {10.1371/journal.pone.0154101},

year  = {2016},

date = {2016-01-01},

journal = {PLOS ONE},

volume = {11},

number = {4},

pages = {1-15},

publisher = {Public Library of Science},

abstract = {During the last decade miltefosine (MIL) has been used as first-line treatment for visceral leishmaniasis in endemic areas with antimonial resistance, but a decline in clinical effectiveness is now being reported. While only two MIL-resistant Leishmania infantum strains from HIV co-infected patients have been documented, phenotypic MIL-resistance for L. donovani has not yet been identified in the laboratory. Hence, a better understanding of the factors contributing to increased MIL-treatment failure is necessary. Given the paucity of defined MIL-resistant L. donovani clinical isolates, this study used an experimental amastigote-selected MIL-resistant L. infantum isolate (LEM3323). In-depth exploration of the MIL-resistant phenotype was performed by coupling genomic with phenotypic data to gain insight into gene function and the mutant phenotype. A naturally MIL-resistant L. infantum clinical isolate (LEM5159) was included to compare both datasets. Phenotypically, resistance was evaluated by determining intracellular amastigote susceptibility in vitro and actual MIL-uptake. Genomic analysis provided supportive evidence that the resistance selection model on intracellular amastigotes can be a good proxy for the in vivo field situation since both resistant strains showed mutations in the same inward transporter system responsible for the acquired MIL-resistant phenotype. In line with previous literature findings in promastigotes, our data confirm a defective import machinery through inactivation of the LiMT/LiRos3 protein complex as the main mechanism for MIL-resistance also in intracellular amastigotes. Whole genome sequencing analysis of LEM3323 revealed a 2 base pair deletion in the LiMT gene that led to the formation an early stop codon and a truncation of the LiMT protein. Interestingly, LEM5159 revealed mutations in both the LiMT and LiRos3 genes, resulting in an aberrant expression of the LiMT protein. To verify that these mutations were indeed accountable for the acquired resistance, transfection experiments were performed to re-establish MIL-susceptibility. In LEM3323, susceptibility was restored upon expression of a LiMT wild-type gene, whereas the MIL-susceptibility of LEM5159 could be reversed after expression of the LiRos3 wild-type gene. The aberrant expression profile of the LiMT protein could be restored upon rescue of the LiRos3 gene both in the LEM5159 clinical isolate and a ΔLiRos3 strain, showing that expression of LdMT is dependent on LdRos3 expression. The present findings clearly corroborate the pivotal role of the LiMT/LiRos3 complex in resistance towards MIL.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{Berg2242,

title = {Experimental Resistance to Drug Combinations in Leishmania donovani: Metabolic and Phenotypic Adaptations},

author = {Maya Berg and Raquel Garc{'i}a-Hern\'{a}ndez and Bart Cuypers and Manu Vanaerschot and Jos\'{e} I Manzano and Jos\'{e} A Poveda and Jos\'{e} A Ferragut and Santiago Castanys and Jean-Claude Dujardin and Francisco Gamarro},

url = {https://aac.asm.org/content/59/4/2242},

doi = {10.1128/AAC.04231-14},

issn = {0066-4804},

year  = {2015},

date = {2015-01-01},

journal = {Antimicrobial Agents and Chemotherapy},

volume = {59},

number = {4},

pages = {2242--2255},

publisher = {American Society for Microbiology Journals},

abstract = {Together with vector control, chemotherapy is an essential tool for the control of visceral leishmaniasis (VL), but its efficacy is jeopardized by growing resistance and treatment failure against first-line drugs. To delay the emergence of resistance, the use of drug combinations of existing antileishmanial agents has been tested systematically in clinical trials for the treatment of visceral leishmaniasis (VL). In vitro, Leishmania donovani promastigotes are able to develop experimental resistance to several combinations of different antileishmanial drugs after 10 weeks of drug pressure. Using an untargeted liquid chromatography-mass spectrometry (LC-MS) metabolomics approach, we identified metabolic changes in lines that were experimentally resistant to drug combinations and their respective single-resistant lines. This highlighted both collective metabolic changes (found in all combination therapy-resistant [CTR] lines) and specific ones (found in certain CTR lines). We demonstrated that single-resistant and CTR parasite cell lines show distinct metabolic adaptations, which all converge on the same defensive mechanisms that were experimentally validated: protection against drug-induced and external oxidative stress and changes in membrane fluidity. The membrane fluidity changes were accompanied by changes in drug uptake only in the lines that were resistant against drug combinations with antimonials, and surprisingly, drug accumulation was higher in these lines. Together, these results highlight the importance and the central role of protection against oxidative stress in the different resistant lines. Ultimately, these phenotypic changes might interfere with the mode of action of all drugs that are currently used for the treatment of VL and should be taken into account in drug development.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{Raie00611-13b,

title = {Relapse after Treatment with Miltefosine for Visceral Leishmaniasis Is Associated with Increased Infectivity of the Infecting Leishmania donovani Strain},

author = {Keshav Rai and Bart Cuypers and Narayan Raj Bhattarai and Surendra Uranw and Maya Berg and Bart Ostyn and Jean-Claude Dujardin and Suman Rijal and Manu Vanaerschot},

editor = {Louis M Weiss},

url = {https://mbio.asm.org/content/4/5/e00611-13},

doi = {10.1128/mBio.00611-13},

year  = {2013},

date = {2013-01-01},

journal = {mBio},

volume = {4},

number = {5},

publisher = {American Society for Microbiology},

abstract = {Leishmania donovani is an intracellular protozoan parasite that causes leishmaniasis, which can range from a self-healing cutaneous disease to a fatal visceral disease depending on the infecting species. Miltefosine is currently the latest and only oral antileishmanial that came out of drug discovery pipelines in the past few decades, but recent reports indicate a significant decline in its efficacy against visceral leishmaniasis (also known as kala-azar) in the Indian subcontinent. This relapse rate of up to 20% within 12~months after treatment was shown not to be related to reinfection, drug quality, drug exposure, or drug-resistant parasites. We therefore aimed to assess other phenotypes of the parasite that may affect treatment outcome and found a significant association between the number of metacyclic parasites, parasite infectivity, and patient treatment outcome in the Indian subcontinent. Together with previous studies on resistance of L. donovani against pentavalent antimonials, these data suggest that the infectivity of the parasite, or related phenotypes, might be a more determinant factor for treatment failure in visceral leishmaniasis than drug susceptibility, warranting a reassessment of our current view on treatment failure and drug resistance in leishmaniasis and beyond.IMPORTANCE The high miltefosine relapse rate poses a major challenge for the current Kala-Azar Elimination Program in the Indian subcontinent and other leishmaniasis control programs worldwide. This relapse rate could not be related to reinfection, drug-resistant parasites, or reduced treatment quality. Here we report that an increased infectivity of the parasite is associated with miltefosine relapse of visceral leishmaniasis (VL) patients. These results supplement those obtained with antimonial-resistant L. donovani where an increased infectivity was also observed. This challenges the current view of Leishmania drug susceptibility being the biggest parasitic factor that contributes to treatment failure in leishmaniasis. These selected more infectious parasites may pose an additional burden to leishmaniasis control programs, highlighting the importance of multifaceted control measures to achieve leishmaniasis elimination in the Indian subcontinent and other regions where leishmaniasis is endemic.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{doi:10.1111/mmi.12374,

title = {Metabolic adaptations of Leishmania donovani in relation to differentiation, drug resistance, and drug pressure},

author = {Maya Berg and Manu Vanaerschot and Andris Jankevics and Bart Cuypers and Ilse Maes and Sandip Mukherjee and Basudha Khanal and Suman Rijal and Syamal Roy and Fred Opperdoes and Rainer Breitling and Jean-Claude Dujardin},

url = {https://onlinelibrary.wiley.com/doi/abs/10.1111/mmi.12374},

doi = {10.1111/mmi.12374},

year  = {2013},

date = {2013-01-01},

journal = {Molecular Microbiology},

volume = {90},

number = {2},

pages = {428-442},

abstract = {Summary Antimonial (sodium stibogluconate, SSG) resistance and differentiation have been shown to be closely linked in Leishmania donovani, with SSG-resistant strains showing an increased capacity to generate infectious (metacyclic) forms. This is the first untargeted LC-MS metabolomics study which integrated both phenomena in one experimental design and provided insights into metabolic differences between three clinical L. donovani strains with a similar genetic background but different SSG-susceptibilities. We performed this analysis at different stages during promastigote growth and in the absence or presence of drug pressure. When comparing SSG-resistant and SSG-sensitive strains, a number of metabolic changes appeared to be constitutively present in all growth stages, pointing towards a clear link with SSG-resistance, whereas most metabolic changes were only detected in the stationary stage. These changes reflect the close intertwinement between SSG-resistance and an increased metacyclogenesis in resistant parasites. The metabolic changes suggest that SSG-resistant parasites have (i) an increased capacity for protection against oxidative stress; (ii) a higher fluidity of the plasma membrane; and (iii) a metabolic survival kit to better endure infection. These changes were even more pronounced in a resistant strain kept under SbIII drug pressure.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{BERG2013e201301002,

title = {LC-MS Metabolomics from study design to data-analysis \textendash using a versatile pathogen as a test case},

author = {Maya Berg and Manu Vanaerschot and Andris Jankevics and Bart Cuypers and Rainer Breitling and Jean-Claude Dujardin},

url = {http://www.sciencedirect.com/science/article/pii/S2001037014600453},

doi = {https://doi.org/10.5936/csbj.201301002},

issn = {2001-0370},

year  = {2013},

date = {2013-01-01},

journal = {Computational and Structural Biotechnology Journal},

volume = {4},

number = {5},

pages = {e201301002},

abstract = {Thanks to significant improvements in LC-MS technology, metabolomics is increasingly used as a tool to discriminate the responses of organisms to various stimuli or drugs. In this minireview we discuss all aspects of the LC-MS metabolomics pipeline, using a complex and versatile model organism, Leishmania donovani, as an illustrative example. The benefits of a hyphenated mass spectrometry platform and a detailed overview of the entire experimental pipeline from sampling, sample storage and sample list set-up to LC-MS measurements and the generation of meaningful results with state-of-the-art data-analysis software will be thoroughly discussed. Finally, we also highlight important pitfalls in the processing of LC-MS data and comment on the benefits of implementing metabolomics in a systems biology approach.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}