Publications
2020 | ||
Adriaensen, Wim; Cuypers, Bart; Cordero, Carlota F; Mengasha, Bewketu; Blesson, Séverine; Cnops, Lieselotte; Kaye, Paul M; Alves, Fabiana; Diro, Ermias; van Griensven, Johan Host transcriptomic signature as alternative test-of-cure in visceral leishmaniasis patients co-infected with HIV Journal Article EBioMedicine, 55 , 2020, ISSN: 2352-3964. Abstract | Links | BibTeX | Altmetric | Tags: immunology, leishmania, machine-learning, RNA-seq, transcriptomics @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 = {immunology, leishmania, machine-learning, RNA-seq, transcriptomics}, pubstate = {published}, tppubtype = {article} } 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. | ||
Uzureau, Sophie; Lecordier, Laurence; Uzureau, Pierrick; Hennig, Dorle; Graversen, Jonas H; Homblé, Fabrice; Mfutu, Pepe Ekulu; Arcolino], Fanny [Oliveira; Ramos, Ana Raquel; Rovere], Rita [La M; Luyten, Tomas; Vermeersch, Marjorie; Tebabi, Patricia; Dieu, Marc; Cuypers, Bart; Deborggraeve, Stijn; Rabant, Marion; Legendre, Christophe; Moestrup, Søren K; Levtchenko, Elena; Bultynck, Geert; Erneux, Christophe; Pérez-Morga, David; Pays, Etienne APOL1 C-Terminal Variants May Trigger Kidney Disease through Interference with APOL3 Control of Actomyosin Journal Article Cell Reports, 30 (11), pp. 3821 – 3836.e13, 2020, ISSN: 2211-1247. Abstract | Links | BibTeX | Altmetric | Tags: NGS @article{UZUREAU20203821, title = {APOL1 C-Terminal Variants May Trigger Kidney Disease through Interference with APOL3 Control of Actomyosin}, author = {Sophie Uzureau and Laurence Lecordier and Pierrick Uzureau and Dorle Hennig and Jonas H Graversen and Fabrice Hombl\'{e} and Pepe Ekulu Mfutu and Fanny [Oliveira Arcolino] and Ana Raquel Ramos and Rita [La M Rovere] and Tomas Luyten and Marjorie Vermeersch and Patricia Tebabi and Marc Dieu and Bart Cuypers and Stijn Deborggraeve and Marion Rabant and Christophe Legendre and S\oren K Moestrup and Elena Levtchenko and Geert Bultynck and Christophe Erneux and David P\'{e}rez-Morga and Etienne Pays}, url = {http://www.sciencedirect.com/science/article/pii/S2211124720302321}, doi = {https://doi.org/10.1016/j.celrep.2020.02.064}, issn = {2211-1247}, year = {2020}, date = {2020-01-01}, journal = {Cell Reports}, volume = {30}, number = {11}, pages = {3821 – 3836.e13}, abstract = {Summary The C-terminal variants G1 and G2 of apolipoprotein L1 (APOL1) confer human resistance to the sleeping sickness parasite Trypanosoma rhodesiense, but they also increase the risk of kidney disease. APOL1 and APOL3 are death-promoting proteins that are partially associated with the endoplasmic reticulum and Golgi membranes. We report that in podocytes, either APOL1 C-terminal helix truncation (APOL1Δ) or APOL3 deletion (APOL3KO) induces similar actomyosin reorganization linked to the inhibition of phosphatidylinositol-4-phosphate [PI(4)P] synthesis by the Golgi PI(4)-kinase IIIB (PI4KB). Both APOL1 and APOL3 can form K+ channels, but only APOL3 exhibits Ca2+-dependent binding of high affinity to neuronal calcium sensor-1 (NCS-1), promoting NCS-1-PI4KB interaction and stimulating PI4KB activity. Alteration of the APOL1 C-terminal helix triggers APOL1 unfolding and increased binding to APOL3, affecting APOL3-NCS-1 interaction. Since the podocytes of G1 and G2 patients exhibit an APOL1Δ or APOL3KO-like phenotype, APOL1 C-terminal variants may induce kidney disease by preventing APOL3 from activating PI4KB, with consecutive actomyosin reorganization of podocytes.}, keywords = {NGS}, pubstate = {published}, tppubtype = {article} } Summary The C-terminal variants G1 and G2 of apolipoprotein L1 (APOL1) confer human resistance to the sleeping sickness parasite Trypanosoma rhodesiense, but they also increase the risk of kidney disease. APOL1 and APOL3 are death-promoting proteins that are partially associated with the endoplasmic reticulum and Golgi membranes. We report that in podocytes, either APOL1 C-terminal helix truncation (APOL1Δ) or APOL3 deletion (APOL3KO) induces similar actomyosin reorganization linked to the inhibition of phosphatidylinositol-4-phosphate [PI(4)P] synthesis by the Golgi PI(4)-kinase IIIB (PI4KB). Both APOL1 and APOL3 can form K+ channels, but only APOL3 exhibits Ca2+-dependent binding of high affinity to neuronal calcium sensor-1 (NCS-1), promoting NCS-1-PI4KB interaction and stimulating PI4KB activity. Alteration of the APOL1 C-terminal helix triggers APOL1 unfolding and increased binding to APOL3, affecting APOL3-NCS-1 interaction. Since the podocytes of G1 and G2 patients exhibit an APOL1Δ or APOL3KO-like phenotype, APOL1 C-terminal variants may induce kidney disease by preventing APOL3 from activating PI4KB, with consecutive actomyosin reorganization of podocytes. | ||
Cuypers, Bart; Dumetz, Franck; Meysman, Pieter; Laukens, Kris; Muylder, Géraldine De; Dujardin, Jean-Claude; Domagalska, Malgorzata Anna The Absence of C-5 DNA Methylation in Leishmania donovani Allows DNA Enrichment from Complex Samples Journal Article Microorganisms, 8 (8), 2020, ISSN: 2076-2607. Abstract | Links | BibTeX | Altmetric | Tags: bisulfite-sequencing, genomics, leishmania, NGS @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 = {bisulfite-sequencing, genomics, leishmania, NGS}, pubstate = {published}, tppubtype = {article} } 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. | ||
2019 | ||
Shome, S; Parra, RG; Fatima, N; Monzon, AM; Cuypers, B; Moosa, Y; Coimbra, NDR; Assis, J; Giner-Delgado, C; Dönerta?, HM; Cuesta-Astroz, Y; Saarunya, G; Allali, I; Gupta, S; Srivastava, A; Kalsan, M; Valdivia, C; Olguin-Orellana, G J; Papadimitriou, S; Parisi, D; Kristensen, NP; Rib, L; Guebila, MB; Bauer, E; Zaffaroni, G; Bekkar, A; Ashano, E; Paladin, L; Necci, M; Moreyra, NN; Rydén, M; Villalobos-Solís, J; Papadopoulos, N; Rafael, C; Karakulak, T; Kaya, Y; Gladbach, Y; Dhanda, SK; ?o?tari?, N; Alex, A; DeBlasio, D; Rahman, F Global network of computational biology communities: ISCB’s Regional Student Groups breaking barriers Journal Article F1000Research, 8 (1574), 2019. Links | BibTeX | Altmetric | Tags: @article{10.12688/f1000research.20408.1, title = {Global network of computational biology communities: ISCB’s Regional Student Groups breaking barriers}, author = {S Shome and RG Parra and N Fatima and AM Monzon and B Cuypers and Y Moosa and NDR Coimbra and J Assis and C Giner-Delgado and HM D\”{o}nerta? and Y Cuesta-Astroz and G Saarunya and I Allali and S Gupta and A Srivastava and M Kalsan and C Valdivia and G J. Olguin-Orellana and S Papadimitriou and D Parisi and NP Kristensen and L Rib and MB Guebila and E Bauer and G Zaffaroni and A Bekkar and E Ashano and L Paladin and M Necci and NN Moreyra and M Ryd\'{e}n and J Villalobos-Sol\'{i}s and N Papadopoulos and C Rafael and T Karakulak and Y Kaya and Y Gladbach and SK Dhanda and N ?o?tari? and A Alex and D DeBlasio and F Rahman}, doi = {10.12688/f1000research.20408.1}, year = {2019}, date = {2019-01-01}, journal = {F1000Research}, volume = {8}, number = {1574}, keywords = {}, pubstate = {published}, tppubtype = {article} } | ||
Cuypers, B; Dumetz, F; Meysman, P; Laukens, K; Muylder, De G; Dujardin, J-C; Domagalska, MA The absence of C-5 DNA methylation in Leishmania donovani allows DNA enrichment from complex samples Journal Article bioRxiv, 2019. Abstract | Links | BibTeX | Altmetric | Tags: bisulfite-sequencing, epigenomics, genomics, leishmania, NGS @article{Cuypers747063, title = {The absence of C-5 DNA methylation in Leishmania donovani allows DNA enrichment from complex samples}, author = {B Cuypers and F Dumetz and P Meysman and K Laukens and G De Muylder and J-C Dujardin and MA Domagalska}, url = {https://www.biorxiv.org/content/early/2019/12/04/747063}, doi = {10.1101/747063}, year = {2019}, date = {2019-01-01}, journal = {bioRxiv}, publisher = {Cold Spring Harbor Laboratory}, abstract = {Cytosine C5 methylation is an important epigenetic control mechanism in a wide array of Eukaryotic organisms and generally carried out by proteins of 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 show that the Leishmania donovani genome contains a C-5 DNA methyltransferase (DNMT) from the DNMT6 subfamily, of which the function is still unclear, and verified its expression at RNA level. We created viable overexpressor and knock-out lines of this enzyme and characterised their genome-wide methylation patterns using whole-genome bisulfite sequencing, together with promastigote and amastigote control lines. Interestingly, despite 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 genome 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. A similar absence of DNA methylation was observed for the blood form of Trypanosoma brucei, another Trypanosomatid species. We demonstrate that this difference in DNA methylation between the parasite (no detectable DNA methylation) and the vertebrate host (DNA methylation), allows enrichment of parasite DNA using Methyl-CpG-binding domain columns, readily available in commercial kits. As such, we depleted methylated DNA from 1) mixes of Leishmania promastigote and amastigote DNA with human DNA and 2) THP-1 cells infected with Leishmania amastigotes. This resulted in 62x to 263x Leishmania:human enrichment, depending on the dilution and type of sample studied. These results open a promising avenue for including an unmethylated DNA enrichment step as a pre-enrichment before sequencing clinical samples.}, keywords = {bisulfite-sequencing, epigenomics, genomics, leishmania, NGS}, pubstate = {published}, tppubtype = {article} } Cytosine C5 methylation is an important epigenetic control mechanism in a wide array of Eukaryotic organisms and generally carried out by proteins of 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 show that the Leishmania donovani genome contains a C-5 DNA methyltransferase (DNMT) from the DNMT6 subfamily, of which the function is still unclear, and verified its expression at RNA level. We created viable overexpressor and knock-out lines of this enzyme and characterised their genome-wide methylation patterns using whole-genome bisulfite sequencing, together with promastigote and amastigote control lines. Interestingly, despite 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 genome 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. A similar absence of DNA methylation was observed for the blood form of Trypanosoma brucei, another Trypanosomatid species. We demonstrate that this difference in DNA methylation between the parasite (no detectable DNA methylation) and the vertebrate host (DNA methylation), allows enrichment of parasite DNA using Methyl-CpG-binding domain columns, readily available in commercial kits. As such, we depleted methylated DNA from 1) mixes of Leishmania promastigote and amastigote DNA with human DNA and 2) THP-1 cells infected with Leishmania amastigotes. This resulted in 62x to 263x Leishmania:human enrichment, depending on the dilution and type of sample studied. These results open a promising avenue for including an unmethylated DNA enrichment step as a pre-enrichment before sequencing clinical samples. | ||
2018 | ||
Hassan, Mehedi; Namasivayam, Aishwarya Alex; DeBlasio, Dan; Fatima, Nazeefa; Siranosian, Benjamin; Parra, Gonzalo R; Cuypers, Bart; Shome, Sayane; Monzon, Alexander Miguel; Fumey, Julien; Rahman, Farzana Reflections on a journey: a retrospective of the ISCB Student Council symposium series Journal Article BMC Bioinformatics, 19 (12), pp. 347, 2018, ISSN: 1471-2105. Abstract | Links | BibTeX | Altmetric | Tags: @article{Hassan2018, title = {Reflections on a journey: a retrospective of the ISCB Student Council symposium series}, author = {Mehedi Hassan and Aishwarya Alex Namasivayam and Dan DeBlasio and Nazeefa Fatima and Benjamin Siranosian and Gonzalo R Parra and Bart Cuypers and Sayane Shome and Alexander Miguel Monzon and Julien Fumey and Farzana Rahman}, url = {https://doi.org/10.1186/s12859-018-2369-x}, doi = {10.1186/s12859-018-2369-x}, issn = {1471-2105}, year = {2018}, date = {2018-10-09}, journal = {BMC Bioinformatics}, volume = {19}, number = {12}, pages = {347}, abstract = {This article describes the motivation, origin and evolution of the student symposia series organised by the ISCB Student Council. The meeting series started thirteen years ago in Madrid and has spread to four continents. The article concludes with the highlights of the most recent edition of annual Student Council Symposium held in conjunction with the 25th Conference on Intelligent Systems for Molecular Biology and the 16th European Conference on Computational Biology, in Prague, in July 2017.}, keywords = {}, pubstate = {published}, tppubtype = {article} } This article describes the motivation, origin and evolution of the student symposia series organised by the ISCB Student Council. The meeting series started thirteen years ago in Madrid and has spread to four continents. The article concludes with the highlights of the most recent edition of annual Student Council Symposium held in conjunction with the 25th Conference on Intelligent Systems for Molecular Biology and the 16th European Conference on Computational Biology, in Prague, in July 2017. | ||
Krivoshiev, Boris V; Beemster, Gerrit T S; Sprangers, Katrien; Cuypers, Bart; Laukens, Kris; Blust, Ronny; Husson, Steven J Transcriptome profiling of HepG2 cells exposed to the flame retardant 9,10-dihydro-9-oxa-10-phosphaphenanthrene 10-oxide (DOPO) Journal Article Toxicology research, 7 (3), pp. 492-502, 2018, ISSN: 2045-452X, (c8tx00006a[PII]). Abstract | Links | BibTeX | Altmetric | Tags: transcriptomics @article{Krivoshiev2018b, title = {Transcriptome profiling of HepG2 cells exposed to the flame retardant 9,10-dihydro-9-oxa-10-phosphaphenanthrene 10-oxide (DOPO)}, author = {Boris V Krivoshiev and Gerrit T S Beemster and Katrien Sprangers and Bart Cuypers and Kris Laukens and Ronny Blust and Steven J Husson}, url = {https://doi.org/10.1039/c8tx00006a}, doi = {10.1039/c8tx00006a}, issn = {2045-452X}, year = {2018}, date = {2018-03-12}, journal = {Toxicology research}, volume = {7}, number = {3}, pages = {492-502}, publisher = {Royal Society of Chemistry}, abstract = {The flame retardant, 9,10-dihydro-9-oxa-10-phosphaphenanthrene 10-oxide (DOPO), has been receiving great interest given its superior fire protection properties, and its predicted low level of persistence, bioaccumulation, and toxicity. However, empirical toxicological data that are essential for a complete hazard assessment are severely lacking. In this study, we attempted to identify the potential toxicological modes of action by transcriptome (RNA-seq) profiling of the human liver hepatocellular carcinoma cell line, HepG2. Such insight may help in identifying compounds of concern and potential toxicological phenotypes. DOPO was found to have little cytotoxic potential, with lower effective concentrations compared to other flame retardants studied in the same cell line. Differentially expressed genes revealed a wide range of molecular effects including changes in protein, energy, DNA, and lipid metabolism, along with changes in cellular stress response pathways. In response to 250 $mu$M DOPO, the most perturbed biological processes were fatty acid metabolism, androgen metabolism, glucose transport, and renal function and development, which is in agreement with other studies that observed similar effects of other flame retardants in other species. However, treatment with 2.5 $mu$M DOPO resulted in very few differentially expressed genes and failed to indicate any potential effects on biology, despite such concentrations likely being orders of magnitude greater than would be encountered in the environment. This, together with the low levels of cytotoxicity, supports the potential replacement of the current flame retardants by DOPO, although further studies are needed to establish the nephrotoxicity and endocrine disruption of DOPO.}, note = {c8tx00006a[PII]}, keywords = {transcriptomics}, pubstate = {published}, tppubtype = {article} } The flame retardant, 9,10-dihydro-9-oxa-10-phosphaphenanthrene 10-oxide (DOPO), has been receiving great interest given its superior fire protection properties, and its predicted low level of persistence, bioaccumulation, and toxicity. However, empirical toxicological data that are essential for a complete hazard assessment are severely lacking. In this study, we attempted to identify the potential toxicological modes of action by transcriptome (RNA-seq) profiling of the human liver hepatocellular carcinoma cell line, HepG2. Such insight may help in identifying compounds of concern and potential toxicological phenotypes. DOPO was found to have little cytotoxic potential, with lower effective concentrations compared to other flame retardants studied in the same cell line. Differentially expressed genes revealed a wide range of molecular effects including changes in protein, energy, DNA, and lipid metabolism, along with changes in cellular stress response pathways. In response to 250 $mu$M DOPO, the most perturbed biological processes were fatty acid metabolism, androgen metabolism, glucose transport, and renal function and development, which is in agreement with other studies that observed similar effects of other flame retardants in other species. However, treatment with 2.5 $mu$M DOPO resulted in very few differentially expressed genes and failed to indicate any potential effects on biology, despite such concentrations likely being orders of magnitude greater than would be encountered in the environment. This, together with the low levels of cytotoxicity, supports the potential replacement of the current flame retardants by DOPO, although further studies are needed to establish the nephrotoxicity and endocrine disruption of DOPO. | ||
Neuter, Nicolas De; Bittremieux, Wout; Beirnaert, Charlie; Cuypers, Bart; Mrzic, Aida; Moris, Pieter; Suls, Arvid; Tendeloo, Viggo Van; Ogunjimi, Benson; Laukens, Kris; Meysman, Pieter On the feasibility of mining CD8+ T cell receptor patterns underlying immunogenic peptide recognition Journal Article Immunogenetics, 70 (3), pp. 159-168, 2018, ISSN: 1432-1211. Abstract | Links | BibTeX | Altmetric | Tags: immunology, machine-learning, virology @article{DeNeuter2018, title = {On the feasibility of mining CD8+ T cell receptor patterns underlying immunogenic peptide recognition}, author = {Nicolas De Neuter and Wout Bittremieux and Charlie Beirnaert and Bart Cuypers and Aida Mrzic and Pieter Moris and Arvid Suls and Viggo Van Tendeloo and Benson Ogunjimi and Kris Laukens and Pieter Meysman}, url = {https://doi.org/10.1007/s00251-017-1023-5}, doi = {10.1007/s00251-017-1023-5}, issn = {1432-1211}, year = {2018}, date = {2018-03-01}, journal = {Immunogenetics}, volume = {70}, number = {3}, pages = {159-168}, abstract = {Current T cell epitope prediction tools are a valuable resource in designing targeted immunogenicity experiments. They typically focus on, and are able to, accurately predict peptide binding and presentation by major histocompatibility complex (MHC) molecules on the surface of antigen-presenting cells. However, recognition of the peptide-MHC complex by a T cell receptor (TCR) is often not included in these tools. We developed a classification approach based on random forest classifiers to predict recognition of a peptide by a T cell receptor and discover patterns that contribute to recognition. We considered two approaches to solve this problem: (1) distinguishing between two sets of TCRs that each bind to a known peptide and (2) retrieving TCRs that bind to a given peptide from a large pool of TCRs. Evaluation of the models on two HIV-1, B*08-restricted epitopes reveals good performance and hints towards structural CDR3 features that can determine peptide immunogenicity. These results are of particular importance as they show that prediction of T cell epitope and T cell epitope recognition based on sequence data is a feasible approach. In addition, the validity of our models not only serves as a proof of concept for the prediction of immunogenic T cell epitopes but also paves the way for more general and high-performing models.}, keywords = {immunology, machine-learning, virology}, pubstate = {published}, tppubtype = {article} } Current T cell epitope prediction tools are a valuable resource in designing targeted immunogenicity experiments. They typically focus on, and are able to, accurately predict peptide binding and presentation by major histocompatibility complex (MHC) molecules on the surface of antigen-presenting cells. However, recognition of the peptide-MHC complex by a T cell receptor (TCR) is often not included in these tools. We developed a classification approach based on random forest classifiers to predict recognition of a peptide by a T cell receptor and discover patterns that contribute to recognition. We considered two approaches to solve this problem: (1) distinguishing between two sets of TCRs that each bind to a known peptide and (2) retrieving TCRs that bind to a given peptide from a large pool of TCRs. Evaluation of the models on two HIV-1, B*08-restricted epitopes reveals good performance and hints towards structural CDR3 features that can determine peptide immunogenicity. These results are of particular importance as they show that prediction of T cell epitope and T cell epitope recognition based on sequence data is a feasible approach. In addition, the validity of our models not only serves as a proof of concept for the prediction of immunogenic T cell epitopes but also paves the way for more general and high-performing models. | ||
Cuypers, Bart A systems biology approach for a comprehensive understanding of molecular adaptation in** Leishmania donovani PhD Thesis University of Antwerp, 2018. BibTeX | Tags: genomics, LC-MS, leishmania, multi-omics, NGS, omics-integration, proteomics, RNA-seq, transcriptomics @phdthesis{cuypers2018systems, title = {A systems biology approach for a comprehensive understanding of molecular adaptation in** Leishmania donovani}, author = {Bart Cuypers}, year = {2018}, date = {2018-01-01}, school = {University of Antwerp}, keywords = {genomics, LC-MS, leishmania, multi-omics, NGS, omics-integration, proteomics, RNA-seq, transcriptomics}, pubstate = {published}, tppubtype = {phdthesis} } | ||
Dumetz, F; Cuypers, B; Imamura, H; Zander, D; textquoteright, E D; Maes, I; Domagalska, M A; Clos, J; Dujardin, J -C; Muylder, De G Molecular Preadaptation to Antimony Resistance in Leishmania donovani on the Indian Subcontinent Journal Article mSphere, 3 (2), 2018. Abstract | Links | BibTeX | Altmetric | Tags: LC-MS, leishmania, metabolomics, multi-omics, omics-integration, RNA-seq, transcriptomics @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 = {LC-MS, leishmania, metabolomics, multi-omics, omics-integration, RNA-seq, transcriptomics}, pubstate = {published}, tppubtype = {article} } 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. | ||
Krivoshiev, Boris V; Beemster, Gerrit T S; Sprangers, Katrien; Cuypers, Bart; Laukens, Kris; Blust, Ronny; Husson, Steven J Toxicogenomics of the flame retardant tris (2-butoxyethyl) phosphate in HepG2 cells using RNA-seq Journal Article Toxicology in Vitro, 46 , pp. 178 – 188, 2018, ISSN: 0887-2333. Abstract | Links | BibTeX | Altmetric | Tags: RNA-seq, transcriptomics @article{KRIVOSHIEV2018178, title = {Toxicogenomics of the flame retardant tris (2-butoxyethyl) phosphate in HepG2 cells using RNA-seq}, author = {Boris V Krivoshiev and Gerrit T S Beemster and Katrien Sprangers and Bart Cuypers and Kris Laukens and Ronny Blust and Steven J Husson}, url = {http://www.sciencedirect.com/science/article/pii/S0887233317303089}, doi = {https://doi.org/10.1016/j.tiv.2017.10.011}, issn = {0887-2333}, year = {2018}, date = {2018-01-01}, journal = {Toxicology in Vitro}, volume = {46}, pages = {178 – 188}, abstract = {Tris (2-butoxyethyl) phosphate (TBOEP) is a compound produced at high volume that is used as both a flame retardant and a plasticizer. It is persistent and bioaccumulative, yet little is known of its toxicological modes of action. Such insight may aid risk assessment in a weight-of-evidence approach supplementing current testing strategies. We used an RNA sequencing approach as an unbiased and sensitive tool to explore potential negative health effects of sub-cytotoxic concentrations of TBOEP on the transcriptome of the human liver hepatocellular carcinoma cell line, HepG2, with the lowest concentration used potentially holding relevance to human physiological levels. Over-representation and gene set enrichment analysis corresponded well and revealed that TBOEP treatments resulted in an upregulation of genes involved in protein and energy metabolism, along with DNA replication. Such increases in cell and macromolecule metabolism could explain the increase in mitochondrial activity at lower TBOEP concentrations. In addition, TBOEP affected a wide variety of biological processes, the most notable one being the general stress response, wound healing. Finally, TBOEP showed effects on steroid hormone biosynthesis and activation, regulation, and potentiation of immune responses, in agreement with other studies. As such, this study is the first study investigating genome-wide changes in gene transcription in response to TBOEP in human cells.}, keywords = {RNA-seq, transcriptomics}, pubstate = {published}, tppubtype = {article} } Tris (2-butoxyethyl) phosphate (TBOEP) is a compound produced at high volume that is used as both a flame retardant and a plasticizer. It is persistent and bioaccumulative, yet little is known of its toxicological modes of action. Such insight may aid risk assessment in a weight-of-evidence approach supplementing current testing strategies. We used an RNA sequencing approach as an unbiased and sensitive tool to explore potential negative health effects of sub-cytotoxic concentrations of TBOEP on the transcriptome of the human liver hepatocellular carcinoma cell line, HepG2, with the lowest concentration used potentially holding relevance to human physiological levels. Over-representation and gene set enrichment analysis corresponded well and revealed that TBOEP treatments resulted in an upregulation of genes involved in protein and energy metabolism, along with DNA replication. Such increases in cell and macromolecule metabolism could explain the increase in mitochondrial activity at lower TBOEP concentrations. In addition, TBOEP affected a wide variety of biological processes, the most notable one being the general stress response, wound healing. Finally, TBOEP showed effects on steroid hormone biosynthesis and activation, regulation, and potentiation of immune responses, in agreement with other studies. As such, this study is the first study investigating genome-wide changes in gene transcription in response to TBOEP in human cells. | ||
Cuypers, Bart; Berg, Maya; Imamura, Hideo; Dumetz, Franck; Muylder], Géraldine [De; Domagalska, Malgorzata A; Rijal, Suman; Bhattarai, Narayan Raj; Maes, Ilse; Sanders, Mandy; Cotton, James A; Meysman, Pieter; Laukens, Kris; Dujardin, Jean-Claude Integrated genomic and metabolomic profiling of ISC1, an emerging Leishmania donovani population in the Indian subcontinent Journal Article Infection, Genetics and Evolution, 62 , pp. 170 – 178, 2018, ISSN: 1567-1348. Abstract | Links | BibTeX | Altmetric | Tags: genomics, LC-MS, metabolomics, multi-omics, NGS @article{CUYPERS2018170, title = {Integrated genomic and metabolomic profiling of ISC1, an emerging Leishmania donovani population in the Indian subcontinent}, author = {Bart Cuypers and Maya Berg and Hideo Imamura and Franck Dumetz and G\'{e}raldine [De Muylder] and Malgorzata A Domagalska and Suman Rijal and Narayan Raj Bhattarai and Ilse Maes and Mandy Sanders and James A Cotton and Pieter Meysman and Kris Laukens and Jean-Claude Dujardin}, url = {http://www.sciencedirect.com/science/article/pii/S1567134818302004}, doi = {https://doi.org/10.1016/j.meegid.2018.04.021}, issn = {1567-1348}, year = {2018}, date = {2018-01-01}, journal = {Infection, Genetics and Evolution}, volume = {62}, pages = {170 – 178}, abstract = {Leishmania donovani is the responsible agent for visceral leishmaniasis (VL) in the Indian subcontinent (ISC). The disease is lethal without treatment and causes 0.2 to 0.4 million cases each year. Recently, reports of VL in Nepalese hilly districts have increased as well as VL cases caused by L. donovani from the ISC1 genetic group, a new and emerging genotype. In this study, we perform for the first time an integrated, untargeted genomics and metabolomics approach to characterize ISC1, in comparison with the Core Group (CG), main population that drove the most recent outbreak of VL in the ISC. We show that the ISC1 population is very different from the CG, both at genome and metabolome levels. The genomic differences include SNPs, CNV and small indels in genes coding for known virulence factors, immunogens and surface proteins. Both genomic and metabolic approaches highlighted dissimilarities related to membrane lipids, the nucleotide salvage pathway and the urea cycle in ISC1 versus CG. Many of these pathways and molecules are important for the interaction with the host/extracellular environment. Altogether, our data predict major functional differences in ISC1 versus CG parasites, including virulence. Therefore, particular attention is required to monitor the fate of this emerging ISC1 population in the ISC, especially in a post-VL elimination context.}, keywords = {genomics, LC-MS, metabolomics, multi-omics, NGS}, pubstate = {published}, tppubtype = {article} } Leishmania donovani is the responsible agent for visceral leishmaniasis (VL) in the Indian subcontinent (ISC). The disease is lethal without treatment and causes 0.2 to 0.4 million cases each year. Recently, reports of VL in Nepalese hilly districts have increased as well as VL cases caused by L. donovani from the ISC1 genetic group, a new and emerging genotype. In this study, we perform for the first time an integrated, untargeted genomics and metabolomics approach to characterize ISC1, in comparison with the Core Group (CG), main population that drove the most recent outbreak of VL in the ISC. We show that the ISC1 population is very different from the CG, both at genome and metabolome levels. The genomic differences include SNPs, CNV and small indels in genes coding for known virulence factors, immunogens and surface proteins. Both genomic and metabolic approaches highlighted dissimilarities related to membrane lipids, the nucleotide salvage pathway and the urea cycle in ISC1 versus CG. Many of these pathways and molecules are important for the interaction with the host/extracellular environment. Altogether, our data predict major functional differences in ISC1 versus CG parasites, including virulence. Therefore, particular attention is required to monitor the fate of this emerging ISC1 population in the ISC, especially in a post-VL elimination context. | ||
2017 | ||
Cuypers, Bart; Domagalska, Malgorzata A; Meysman, Pieter; de Muylder, Géraldine; Vanaerschot, Manu; Imamura, Hideo; Dumetz, Franck; Verdonckt, Thomas Wolf; Myler, Peter J; Ramasamy, Gowthaman; Laukens, Kris; Dujardin, Jean-Claude Multiplexed Spliced-Leader Sequencing: A high-throughput, selective method for RNA-seq in Trypanosomatids Journal Article Scientific Reports, 7 (1), pp. 3725, 2017, ISSN: 2045-2322. Abstract | Links | BibTeX | Altmetric | Tags: leishmania, NGS, RNA-seq, transcriptomics @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 = {leishmania, NGS, RNA-seq, transcriptomics}, pubstate = {published}, tppubtype = {article} } 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. | ||
Cuypers, Bart; den Broeck, Frederik Van; Reet, Nick Van; Meehan, Conor J; Cauchard, Julien; Wilkes, Jonathan M; Claes, Filip; Goddeeris, Bruno; Birhanu, Hadush; Dujardin, Jean-Claude; Laukens, Kris; Büscher, Philippe; Deborggraeve, Stijn Genome-Wide SNP Analysis Reveals Distinct Origins of Trypanosoma evansi and Trypanosoma equiperdum Journal Article Genome Biology and Evolution, 9 (8), pp. 1990-1997, 2017, ISSN: 1759-6653. Abstract | Links | BibTeX | Altmetric | Tags: evolution, genomics @article{Cuypers2017b, title = {Genome-Wide SNP Analysis Reveals Distinct Origins of Trypanosoma evansi and Trypanosoma equiperdum}, author = {Bart Cuypers and Frederik Van den Broeck and Nick Van Reet and Conor J Meehan and Julien Cauchard and Jonathan M Wilkes and Filip Claes and Bruno Goddeeris and Hadush Birhanu and Jean-Claude Dujardin and Kris Laukens and Philippe B\”{u}scher and Stijn Deborggraeve}, url = {https://doi.org/10.1093/gbe/evx102}, doi = {10.1093/gbe/evx102}, issn = {1759-6653}, year = {2017}, date = {2017-05-25}, journal = {Genome Biology and Evolution}, volume = {9}, number = {8}, pages = {1990-1997}, abstract = {Trypanosomes cause a variety of diseases in man and domestic animals in Africa, Latin America, and Asia. In the Trypanozoon subgenus, Trypanosoma brucei gambiense and Trypanosoma brucei rhodesiense cause human African trypanosomiasis, whereas Trypanosoma brucei brucei, Trypanosoma evansi, and Trypanosoma equiperdum are responsible for nagana, surra, and dourine in domestic animals, respectively. The genetic relationships between T. evansi and T. equiperdum and other Trypanozoon species remain unclear because the majority of phylogenetic analyses has been based on only a few genes. In this study, we have conducted a phylogenetic analysis based on genome-wide SNP analysis comprising 56 genomes from the Trypanozoon subgenus. Our data reveal that T. equiperdum has emerged at least once in Eastern Africa and T. evansi at two independent occasions in Western Africa. The genomes within the T. equiperdum and T. evansi monophyletic clusters show extremely little variation, probably due to the clonal spread linked to the independence from tsetse flies for their transmission.}, keywords = {evolution, genomics}, pubstate = {published}, tppubtype = {article} } Trypanosomes cause a variety of diseases in man and domestic animals in Africa, Latin America, and Asia. In the Trypanozoon subgenus, Trypanosoma brucei gambiense and Trypanosoma brucei rhodesiense cause human African trypanosomiasis, whereas Trypanosoma brucei brucei, Trypanosoma evansi, and Trypanosoma equiperdum are responsible for nagana, surra, and dourine in domestic animals, respectively. The genetic relationships between T. evansi and T. equiperdum and other Trypanozoon species remain unclear because the majority of phylogenetic analyses has been based on only a few genes. In this study, we have conducted a phylogenetic analysis based on genome-wide SNP analysis comprising 56 genomes from the Trypanozoon subgenus. Our data reveal that T. equiperdum has emerged at least once in Eastern Africa and T. evansi at two independent occasions in Western Africa. The genomes within the T. equiperdum and T. evansi monophyletic clusters show extremely little variation, probably due to the clonal spread linked to the independence from tsetse flies for their transmission. | ||
Jara, Marlene; Berg, Maya; Caljon, Guy; de Muylder, Geraldine; Cuypers, Bart; Castillo, Denis; Maes, Ilse; del Orozco, María Carmen; Vanaerschot, Manu; Dujardin, Jean-Claude; Arevalo, Jorge PLOS ONE, 12 (7), pp. 1-22, 2017. Abstract | Links | BibTeX | Altmetric | Tags: LC-MS, leishmania, metabolomics @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 = {LC-MS, leishmania, metabolomics}, pubstate = {published}, tppubtype = {article} } 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. | ||
Dumetz, F; Imamura, H; Sanders, M; Seblova, V; Myskova, J; Pescher, P; Vanaerschot, M; Meehan, C J; Cuypers, B; Muylder, De G; Späth, G F; Bussotti, G; Vermeesch, J R; Berriman, M; Cotton, J A; Volf, P; Dujardin, J C; Domagalska, M A mBio, 8 (3), 2017. Abstract | Links | BibTeX | Altmetric | Tags: genomics, leishmania, multi-omics, RNA-seq, transcriptomics @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 = {genomics, leishmania, multi-omics, RNA-seq, transcriptomics}, pubstate = {published}, tppubtype = {article} } 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—a protozoan parasite that kills more than 30,000 people each year—is 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. | ||
2016 | ||
Imamura, Hideo; Downing, Tim; den Broeck, Frederik Van; Sanders, Mandy J; Rijal, Suman; Sundar, Shyam; Mannaert, An; Vanaerschot, Manu; Berg, Maya; Muylder, Géraldine De; Dumetz, Franck; Cuypers, Bart; Maes, Ilse; Domagalska, Malgorzata; Decuypere, Saskia; Rai, Keshav; Uranw, Surendra; Bhattarai, Narayan Raj; Khanal, Basudha; Prajapati, Vijay Kumar; Sharma, Smriti; Stark, Olivia; Schönian, Gabriele; Koning, Harry De P; Settimo, Luca; Vanhollebeke, Benoit; Roy, Syamal; Ostyn, Bart; Boelaert, Marleen; Maes, Louis; Berriman, Matthew; Dujardin, Jean-Claude; Cotton, James A Evolutionary genomics of epidemic visceral leishmaniasis in the Indian subcontinent Journal Article eLife, 5 , pp. e12613, 2016, ISSN: 2050-084X. Abstract | Links | BibTeX | Altmetric | Tags: evolution, genomics, NGS @article{10.7554/eLife.12613, title = {Evolutionary genomics of epidemic visceral leishmaniasis in the Indian subcontinent}, author = {Hideo Imamura and Tim Downing and Frederik Van den Broeck and Mandy J Sanders and Suman Rijal and Shyam Sundar and An Mannaert and Manu Vanaerschot and Maya Berg and G\'{e}raldine De Muylder and Franck Dumetz and Bart Cuypers and Ilse Maes and Malgorzata Domagalska and Saskia Decuypere and Keshav Rai and Surendra Uranw and Narayan Raj Bhattarai and Basudha Khanal and Vijay Kumar Prajapati and Smriti Sharma and Olivia Stark and Gabriele Sch\”{o}nian and Harry P De Koning and Luca Settimo and Benoit Vanhollebeke and Syamal Roy and Bart Ostyn and Marleen Boelaert and Louis Maes and Matthew Berriman and Jean-Claude Dujardin and James A Cotton}, editor = {Dominique Soldati-Favre}, url = {https://doi.org/10.7554/eLife.12613}, doi = {10.7554/eLife.12613}, issn = {2050-084X}, year = {2016}, date = {2016-03-01}, journal = {eLife}, volume = {5}, pages = {e12613}, publisher = {eLife Sciences Publications, Ltd}, abstract = {textitLeishmania donovani causes visceral leishmaniasis (VL), the second most deadly vector-borne parasitic disease. A recent epidemic in the Indian subcontinent (ISC) caused up to 80% of global VL and over 30,000 deaths per year. Resistance against antimonial drugs has probably been a contributing factor in the persistence of this epidemic. Here we use whole genome sequences from 204 clinical isolates to track the evolution and epidemiology of textitL. donovani from the ISC. We identify independent radiations that have emerged since a bottleneck coincident with 1960s DDT spraying campaigns. A genetically distinct population frequently resistant to antimonials has a two base-pair insertion in the aquaglyceroporin gene LdAQP1 that prevents the transport of trivalent antimonials. We find evidence of genetic exchange between ISC populations, and show that the mutation in LdAQP1 has spread by recombination. Our results reveal the complexity of textitL. donovani evolution in the ISC in response to drug treatment.}, keywords = {evolution, genomics, NGS}, pubstate = {published}, tppubtype = {article} } textitLeishmania donovani causes visceral leishmaniasis (VL), the second most deadly vector-borne parasitic disease. A recent epidemic in the Indian subcontinent (ISC) caused up to 80% of global VL and over 30,000 deaths per year. Resistance against antimonial drugs has probably been a contributing factor in the persistence of this epidemic. Here we use whole genome sequences from 204 clinical isolates to track the evolution and epidemiology of textitL. donovani from the ISC. We identify independent radiations that have emerged since a bottleneck coincident with 1960s DDT spraying campaigns. A genetically distinct population frequently resistant to antimonials has a two base-pair insertion in the aquaglyceroporin gene LdAQP1 that prevents the transport of trivalent antimonials. We find evidence of genetic exchange between ISC populations, and show that the mutation in LdAQP1 has spread by recombination. Our results reveal the complexity of textitL. donovani evolution in the ISC in response to drug treatment. | ||
Wilkins, Katie; Hassan, Mehedi; Francescatto, Margherita; Jespersen, Jakob; Parra, Gonzalo R; Cuypers, Bart; DeBlasio, Dan; Junge, Alexander; Jigisha, Anupama; Rahman, Farzana; Laenen, Griet; Willems, Sander; Thorrez, Lieven; Moreau, Yves; Raju, Nagarajan; Chothani, Sonia Pankaj; Ramakrishnan, C; Sekijima, Masakazu; Gromiha, Michael M; Slator, Paddy J; Burroughs, Nigel J; ł, Przemys{ł}aw Sza; Tang, Zhonghui; Michalski, Paul; Luo, Oskar; Li, Xingwang; Ruan, Yijun; Plewczynski, Dariusz; Fiscon, Giulia; Weitschek, Emanuel; Ciccozzi, Massimo; Bertolazzi, Paola; Felici, Giovanni; Meysman, Pieter; Vanaerschot, Manu; Berg, Maya; Imamura, Hideo; Dujardin, Jean-Claude; Laukens, Kris; Domanova, Westa; Krycer, James R; Chaudhuri, Rima; Yang, Pengyi; Vafaee, Fatemeh; Fazakerley, Daniel J; Humphrey, Sean J; James, David E; Kuncic, Zdenka Highlights from the 11th ISCB Student Council Symposium 2015 Journal Article BMC Bioinformatics, 17 (3), pp. 95, 2016, ISSN: 1471-2105. Abstract | Links | BibTeX | Altmetric | Tags: @article{Wilkins2016, title = {Highlights from the 11th ISCB Student Council Symposium 2015}, author = {Katie Wilkins and Mehedi Hassan and Margherita Francescatto and Jakob Jespersen and Gonzalo R Parra and Bart Cuypers and Dan DeBlasio and Alexander Junge and Anupama Jigisha and Farzana Rahman and Griet Laenen and Sander Willems and Lieven Thorrez and Yves Moreau and Nagarajan Raju and Sonia Pankaj Chothani and C Ramakrishnan and Masakazu Sekijima and Michael M Gromiha and Paddy J Slator and Nigel J Burroughs and Przemys{\l}aw Sza{\l}aj and Zhonghui Tang and Paul Michalski and Oskar Luo and Xingwang Li and Yijun Ruan and Dariusz Plewczynski and Giulia Fiscon and Emanuel Weitschek and Massimo Ciccozzi and Paola Bertolazzi and Giovanni Felici and Pieter Meysman and Manu Vanaerschot and Maya Berg and Hideo Imamura and Jean-Claude Dujardin and Kris Laukens and Westa Domanova and James R Krycer and Rima Chaudhuri and Pengyi Yang and Fatemeh Vafaee and Daniel J Fazakerley and Sean J Humphrey and David E James and Zdenka Kuncic}, url = {https://doi.org/10.1186/s12859-016-0901-4}, doi = {10.1186/s12859-016-0901-4}, issn = {1471-2105}, year = {2016}, date = {2016-02-25}, journal = {BMC Bioinformatics}, volume = {17}, number = {3}, pages = {95}, abstract = {A1 Highlights from the eleventh ISCB Student Council Symposium 2015}, keywords = {}, pubstate = {published}, tppubtype = {article} } A1 Highlights from the eleventh ISCB Student Council Symposium 2015 | ||
Mondelaers, Annelies; Sanchez-Cañete, Maria P; Hendrickx, Sarah; Eberhardt, Eline; Garcia-Hernandez, Raquel; Lachaud, Laurence; Cotton, James; Sanders, Mandy; Cuypers, Bart; Imamura, Hideo; Dujardin, Jean-Claude; Delputte, Peter; Cos, Paul; Caljon, Guy; Gamarro, Francisco; Castanys, Santiago; Maes, Louis PLOS ONE, 11 (4), pp. 1-15, 2016. Abstract | Links | BibTeX | Altmetric | Tags: genomics, leishmania, NGS @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 = {genomics, leishmania, NGS}, pubstate = {published}, tppubtype = {article} } 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. | ||
Cuypers, Bart; Lecordier, Laurence; Meehan, Conor J; den Broeck, Frederik Van; Imamura, Hideo; Büscher, Philippe; Dujardin, Jean-Claude; Laukens, Kris; Schnaufer, Achim; Dewar, Caroline; Lewis, Michael; Balmer, Oliver; Azurago, Thomas; Kyei-Faried, Sardick; Ohene, Sally-Ann; Duah, Boateng; Homiah, Prince; Mensah, Ebenezer Kofi; Anleah, Francis; Franco, Jose Ramon; Pays, Etienne; Deborggraeve, Stijn Apolipoprotein L1 Variant Associated with Increased Susceptibility to Trypanosome Infection Journal Article mBio, 7 (2), 2016. Abstract | Links | BibTeX | Altmetric | Tags: evolution, NGS @article{Cuyperse02198-15, title = {Apolipoprotein L1 Variant Associated with Increased Susceptibility to Trypanosome Infection}, author = {Bart Cuypers and Laurence Lecordier and Conor J Meehan and Frederik Van den Broeck and Hideo Imamura and Philippe B\”{u}scher and Jean-Claude Dujardin and Kris Laukens and Achim Schnaufer and Caroline Dewar and Michael Lewis and Oliver Balmer and Thomas Azurago and Sardick Kyei-Faried and Sally-Ann Ohene and Boateng Duah and Prince Homiah and Ebenezer Kofi Mensah and Francis Anleah and Jose Ramon Franco and Etienne Pays and Stijn Deborggraeve}, editor = {Michael P Barrett and John C Boothroyd}, url = {https://mbio.asm.org/content/7/2/e02198-15}, doi = {10.1128/mBio.02198-15}, year = {2016}, date = {2016-01-01}, journal = {mBio}, volume = {7}, number = {2}, publisher = {American Society for Microbiology}, abstract = {African trypanosomes, except Trypanosoma brucei gambiense and Trypanosoma~brucei~rhodesiense, which cause human African trypanosomiasis, are lysed by the human serum protein apolipoprotein L1 (ApoL1). These two subspecies can resist human ApoL1 because they express the serum resistance proteins T. b. gambiense glycoprotein (TgsGP) and serum resistance-associated protein (SRA), respectively. Whereas in T.~b.~rhodesiense, SRA is necessary and sufficient to inhibit ApoL1, in T.~b.~gambiense, TgsGP cannot protect against high ApoL1 uptake, so different additional mechanisms contribute to limit this uptake. Here we report a complex interplay between trypanosomes and an ApoL1 variant, revealing important insights into innate human immunity against these parasites. Using whole-genome sequencing, we characterized an atypical T.~b.~gambiense infection in a patient in Ghana. We show that the infecting trypanosome has diverged from the classical T.~b.~gambiense strains and lacks the TgsGP defense mechanism against human serum. By sequencing the ApoL1 gene of the patient and subsequent in vitro mutagenesis experiments, we demonstrate that a homozygous missense substitution (N264K) in the membrane-addressing domain of this ApoL1 variant knocks down the trypanolytic activity, allowing the trypanosome to avoid ApoL1-mediated immunity.IMPORTANCE Most African trypanosomes are lysed by the ApoL1 protein in human serum. Only the subspecies Trypanosoma b. gambiense and T.~b.~rhodesiense can resist lysis by ApoL1 because they express specific serum resistance proteins. We here report a complex interplay between trypanosomes and an ApoL1 variant characterized by a homozygous missense substitution (N264K) in the domain that we hypothesize interacts with the endolysosomal membranes of trypanosomes. The N264K substitution knocks down the lytic activity of ApoL1 against T.~b.~gambiense strains lacking the TgsGP defense mechanism and against T.~b.~rhodesiense if N264K is accompanied by additional substitutions in the SRA-interacting domain. Our data suggest that populations with high frequencies of the homozygous N264K ApoL1 variant may be at increased risk of contracting human African trypanosomiasis.}, keywords = {evolution, NGS}, pubstate = {published}, tppubtype = {article} } African trypanosomes, except Trypanosoma brucei gambiense and Trypanosoma~brucei~rhodesiense, which cause human African trypanosomiasis, are lysed by the human serum protein apolipoprotein L1 (ApoL1). These two subspecies can resist human ApoL1 because they express the serum resistance proteins T. b. gambiense glycoprotein (TgsGP) and serum resistance-associated protein (SRA), respectively. Whereas in T.~b.~rhodesiense, SRA is necessary and sufficient to inhibit ApoL1, in T.~b.~gambiense, TgsGP cannot protect against high ApoL1 uptake, so different additional mechanisms contribute to limit this uptake. Here we report a complex interplay between trypanosomes and an ApoL1 variant, revealing important insights into innate human immunity against these parasites. Using whole-genome sequencing, we characterized an atypical T.~b.~gambiense infection in a patient in Ghana. We show that the infecting trypanosome has diverged from the classical T.~b.~gambiense strains and lacks the TgsGP defense mechanism against human serum. By sequencing the ApoL1 gene of the patient and subsequent in vitro mutagenesis experiments, we demonstrate that a homozygous missense substitution (N264K) in the membrane-addressing domain of this ApoL1 variant knocks down the trypanolytic activity, allowing the trypanosome to avoid ApoL1-mediated immunity.IMPORTANCE Most African trypanosomes are lysed by the ApoL1 protein in human serum. Only the subspecies Trypanosoma b. gambiense and T.~b.~rhodesiense can resist lysis by ApoL1 because they express specific serum resistance proteins. We here report a complex interplay between trypanosomes and an ApoL1 variant characterized by a homozygous missense substitution (N264K) in the domain that we hypothesize interacts with the endolysosomal membranes of trypanosomes. The N264K substitution knocks down the lytic activity of ApoL1 against T.~b.~gambiense strains lacking the TgsGP defense mechanism and against T.~b.~rhodesiense if N264K is accompanied by additional substitutions in the SRA-interacting domain. Our data suggest that populations with high frequencies of the homozygous N264K ApoL1 variant may be at increased risk of contracting human African trypanosomiasis. | ||
Cuypers, B; Jacobsen, A; Siranosian, B; Schwahn, K; Conard, AM; Aben, N; Hassan, M; Fatima, N; Hermans, SMA; Woghiren, M; Meysman, P; Rahman, F; Jigisha, A Highlights from the ISCB Student Council Symposia in 2016 Journal Article F1000Research, 5 (2852), 2016. Links | BibTeX | Altmetric | Tags: @article{10.12688/f1000research.10389.1, title = {Highlights from the ISCB Student Council Symposia in 2016}, author = {B Cuypers and A Jacobsen and B Siranosian and K Schwahn and AM Conard and N Aben and M Hassan and N Fatima and SMA Hermans and M Woghiren and P Meysman and F Rahman and A Jigisha}, doi = {10.12688/f1000research.10389.1}, year = {2016}, date = {2016-01-01}, journal = {F1000Research}, volume = {5}, number = {2852}, keywords = {}, pubstate = {published}, tppubtype = {article} } | ||
2015 | ||
Berg, Maya; ‘i, Raquel Garc; Cuypers, Bart; Vanaerschot, Manu; Manzano, José I; Poveda, José A; Ferragut, José A; Castanys, Santiago; Dujardin, Jean-Claude; Gamarro, Francisco Experimental Resistance to Drug Combinations in Leishmania donovani: Metabolic and Phenotypic Adaptations Journal Article Antimicrobial Agents and Chemotherapy, 59 (4), pp. 2242–2255, 2015, ISSN: 0066-4804. Abstract | Links | BibTeX | Altmetric | Tags: LC-MS, leishmania, leishmaniasis, metabolomics @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 = {LC-MS, leishmania, leishmaniasis, metabolomics}, pubstate = {published}, tppubtype = {article} } 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. | ||
Gryseels, Sophie; Rieger, Toni; Oestereich, Lisa; Cuypers, Bart; Borremans, Benny; Makundi, Rhodes; Leirs, Herwig; Günther, Stephan; de Bellocq], Joëlle [Goüy Virology, 476 , pp. 249 – 256, 2015, ISSN: 0042-6822. Abstract | Links | BibTeX | Altmetric | Tags: evolution, virology @article{GRYSEELS2015249, title = {Gairo virus, a novel arenavirus of the widespread Mastomys natalensis: Genetically divergent, but ecologically similar to Lassa and Morogoro viruses}, author = {Sophie Gryseels and Toni Rieger and Lisa Oestereich and Bart Cuypers and Benny Borremans and Rhodes Makundi and Herwig Leirs and Stephan G\”{u}nther and Jo\”{e}lle [Go\”{u}y de Bellocq]}, url = {http://www.sciencedirect.com/science/article/pii/S0042682214005480}, doi = {https://doi.org/10.1016/j.virol.2014.12.011}, issn = {0042-6822}, year = {2015}, date = {2015-01-01}, journal = {Virology}, volume = {476}, pages = {249 – 256}, abstract = {Despite its near pan-African range, the Natal multimammate mouse, Mastomys natalensis, carries the human pathogen Lassa virus only in West Africa, while the seemingly non-pathogenic arenaviruses Mopeia, Morogoro, and Luna have been detected in this semi-commensal rodent in Mozambique/Zimbabwe, Tanzania and Zambia, respectively. Here, we describe a novel arenavirus in M. natalensis from Gairo district of central Tanzania, for which we propose the name “Gairo virus”. Surprisingly, the virus is not closely related with Morogoro virus that infects M. natalensis only 90km south of Gairo, but clusters phylogenetically with Mobala-like viruses that infect non-M. natalensis host species in Central African Republic and Ethiopia. Despite the evolutionary distance, Gairo virus shares basic ecological features with the other M. natalensis-borne viruses Lassa and Morogoro. Our data show that M. natalensis, carrying distantly related viruses even in the same geographical area, is a potent reservoir host for a variety of arenaviruses.}, keywords = {evolution, virology}, pubstate = {published}, tppubtype = {article} } Despite its near pan-African range, the Natal multimammate mouse, Mastomys natalensis, carries the human pathogen Lassa virus only in West Africa, while the seemingly non-pathogenic arenaviruses Mopeia, Morogoro, and Luna have been detected in this semi-commensal rodent in Mozambique/Zimbabwe, Tanzania and Zambia, respectively. Here, we describe a novel arenavirus in M. natalensis from Gairo district of central Tanzania, for which we propose the name “Gairo virus”. Surprisingly, the virus is not closely related with Morogoro virus that infects M. natalensis only 90km south of Gairo, but clusters phylogenetically with Mobala-like viruses that infect non-M. natalensis host species in Central African Republic and Ethiopia. Despite the evolutionary distance, Gairo virus shares basic ecological features with the other M. natalensis-borne viruses Lassa and Morogoro. Our data show that M. natalensis, carrying distantly related viruses even in the same geographical area, is a potent reservoir host for a variety of arenaviruses. | ||
2013 | ||
Berg, Maya; Vanaerschot, Manu; Jankevics, Andris; Cuypers, Bart; Maes, Ilse; Mukherjee, Sandip; Khanal, Basudha; Rijal, Suman; Roy, Syamal; Opperdoes, Fred; Breitling, Rainer; Dujardin, Jean-Claude Metabolic adaptations of Leishmania donovani in relation to differentiation, drug resistance, and drug pressure Journal Article Molecular Microbiology, 90 (2), pp. 428-442, 2013. Abstract | Links | BibTeX | Altmetric | Tags: LC-MS, leishmania, metabolomics @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 = {LC-MS, leishmania, metabolomics}, pubstate = {published}, tppubtype = {article} } 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. | ||
Rai, Keshav; Cuypers, Bart; Bhattarai, Narayan Raj; Uranw, Surendra; Berg, Maya; Ostyn, Bart; Dujardin, Jean-Claude; Rijal, Suman; Vanaerschot, Manu mBio, 4 (5), 2013. Abstract | Links | BibTeX | Altmetric | Tags: leishmania @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 = {leishmania}, pubstate = {published}, tppubtype = {article} } 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. | ||
Berg, Maya; Vanaerschot, Manu; Jankevics, Andris; Cuypers, Bart; Breitling, Rainer; Dujardin, Jean-Claude LC-MS Metabolomics from study design to data-analysis – using a versatile pathogen as a test case Journal Article Computational and Structural Biotechnology Journal, 4 (5), pp. e201301002, 2013, ISSN: 2001-0370. Abstract | Links | BibTeX | Altmetric | Tags: LC-MS, leishmania, metabolomics @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 = {LC-MS, leishmania, metabolomics}, pubstate = {published}, tppubtype = {article} } 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. | ||