The Experts below are selected from a list of 360 Experts worldwide ranked by ideXlab platform
Andrew F. Read - One of the best experts on this subject based on the ideXlab platform.
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a nutrient mediates intraspecific competition between Rodent Malaria parasites in vivo
Proceedings of The Royal Society B: Biological Sciences, 2017Co-Authors: Nina Wale, Andrew F. ReadAbstract:Hosts are often infected with multiple strains of a single parasite species. Within-host competition between parasite strains can be intense and has implications for the evolution of traits that im...
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a nutrient mediates intraspecific competition between Rodent Malaria parasites in vivo
Proceedings of The Royal Society B: Biological Sciences, 2017Co-Authors: Nina Wale, Derek G Sim, Andrew F. ReadAbstract:Hosts are often infected with multiple strains of a single parasite species. Within-host competition between parasite strains can be intense and has implications for the evolution of traits that impact patient health, such as drug resistance and virulence. Yet the mechanistic basis of within-host competition is poorly understood. Here, we demonstrate that a parasite nutrient, para-aminobenzoic acid (pABA), mediates competition between a drug resistant and drug susceptible strain of the Malaria parasite, Plasmodium chabaudi We further show that increasing pABA supply to hosts infected with the resistant strain worsens disease and changes the relationship between parasite burden and pathology. Our experiments demonstrate that, even when there is profound top-down regulation (immunity), bottom-up regulation of pathogen populations can occur and that its importance may vary during an infection. The identification of resources that can be experimentally controlled opens up the opportunity to manipulate competitive interactions between parasites and hence their evolution.
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immune mediated competition in Rodent Malaria is most likely caused by induced changes in innate immune clearance of merozoites
Malaria Journal, 2014Co-Authors: Jayanthi Santhanam, Andrew F. Read, Lars Raberg, Nicholas J SavillAbstract:Malaria infections often consist of more than one strain of the same parasitic species. Understanding the within-host competition between these various strains is essential to understanding the evolution and epidemiology of drug resistance in Malarial infections. The infection process and the competition between strains involve complicated biological processes that are explained by various hypotheses. Mathematical models tested against experimental data provide quantitative measures to compare these hypotheses and enable us to discern the actual biological processes that contribute to the observed dynamics. We use a group of models against experimental data on Rodent Malaria to test various hypotheses. Such quantitative measures, in understanding Rodent Malaria, can be considered as a step towards understanding within-host parasite dynamics. Our work presented here demonstrates how confronting mathematical models with data allows the discovery of subtle and novel interactions between hosts and parasites that would be impractical to do in an experiment and allows the rejection of hypotheses that are incorrect. It is our contention that understanding the forces controlling within-host parasite dynamics in well-defined experimental model is a necessary step towards understanding these features in natural infections.
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quantifying variation in the potential for antibody mediated apparent competition among nine genotypes of the Rodent Malaria parasite plasmodium chabaudi
Infection Genetics and Evolution, 2013Co-Authors: Karen Fairlieclarke, Andrew F. Read, Judith E Allen, Andrea L GrahamAbstract:Within-host competition among parasite genotypes affects epidemiology as well as the evolution of vir- ulence. In the Rodent Malaria Plasmodium chabaudi, competition among genotypes, as well as clone-spe- cific and clone-transcending immunity are well documented. However, variation among genotypes in the induction of antibodies is not well understood, despite the important role of antibodies in the clearance of Malaria infection. Here, we quantify the potential for antibodies induced by one clone to bind another (i.e., to cause antibody-mediated apparent competition) for nine genetically distinct P. chabaudi clones. We hypothesised that clones would vary in the strength of antibody induction, and that the propensity for clone-transcending immunity between a pair of clones would increase with increasing genetic relat- edness at key antigenic loci. Using serum collected from mice 35 days post-infection, we measured titres of antibody to an unrelated antigen, Keyhole Limpet Haemocyanin (KLH), and two Malaria antigens: recombinant Apical Membrane Antigen-1 (AMA-1) and Merozoite Surface Protein-119 (MSP-119). Amino acid sequence homology within each antigenic locus was used as a measure of relatedness. We found sig- nificant parasite genetic variation for the strength of antibody induction. We also found that relatedness at MSP-119 but not AMA-1 predicted clone-transcending binding. Our results help explain the outcome of chronic-phase mixed infections and generate testable predictions about the pairwise competitive ability of P. chabaudi clones.
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virulence drug sensitivity and transmission success in the Rodent Malaria plasmodium chabaudi
Proceedings of The Royal Society B: Biological Sciences, 2012Co-Authors: Petra Schneider, Simon Blanford, Andrew F. Read, Andrew S Bell, Derek G Sim, Aidan J Odonnell, Krijn P Paaijmans, Sarah E ReeceAbstract:Here, we test the hypothesis that virulent Malaria parasites are less susceptible to drug treatment than less virulent parasites. If true, drug treatment might promote the evolution of more virulent parasites (defined here as those doing more harm to hosts). Drug-resistance mechanisms that protect parasites through interactions with drug molecules at the sub-cellular level are well known. However, parasite phenotypes associated with virulence might also help parasites survive in the presence of drugs. For example, rapidly replicating parasites might be better able to recover in the host if drug treatment fails to eliminate parasites. We quantified the effects of drug treatment on the in-host survival and between-host transmission of Rodent Malaria (Plasmodium chabaudi) parasites which differed in virulence and had never been previously exposed to drugs. In all our treatment regimens and in single- and mixed-genotype infections, virulent parasites were less sensitive to pyrimethamine and artemisinin, the two antiMalarial drugs we tested. Virulent parasites also achieved disproportionately greater transmission when exposed to pyrimethamine. Overall, our data suggest that drug treatment can select for more virulent parasites. Drugs targeting transmission stages (such as artemisinin) may minimize the evolutionary advantage of virulence in drug-treated infections.
Richard Culleton - One of the best experts on this subject based on the ideXlab platform.
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plasmodium vinckei genomes provide insights into the pan genome and evolution of Rodent Malaria parasites
BMC Biology, 2021Co-Authors: Richard Culleton, Abhinay Ramaprasad, Arnab Pain, Severina Klaus, Olga DouvropoulouAbstract:Rodent Malaria parasites (RMPs) serve as tractable tools to study Malaria parasite biology and host-parasite-vector interactions. Among the four RMPs originally collected from wild thicket rats in sub-Saharan Central Africa and adapted to laboratory mice, Plasmodium vinckei is the most geographically widespread with isolates collected from five separate locations. However, there is a lack of extensive phenotype and genotype data associated with this species, thus hindering its use in experimental studies. We have generated a comprehensive genetic resource for P. vinckei comprising of five reference-quality genomes, one for each of its subspecies, blood-stage RNA sequencing data for five P. vinckei isolates, and genotypes and growth phenotypes for ten isolates. Additionally, we sequenced seven isolates of the RMP species Plasmodium chabaudi and Plasmodium yoelii, thus extending genotypic information for four additional subspecies enabling a re-evaluation of the genotypic diversity and evolutionary history of RMPs. The five subspecies of P. vinckei have diverged widely from their common ancestor and have undergone large-scale genome rearrangements. Comparing P. vinckei genotypes reveals region-specific selection pressures particularly on genes involved in mosquito transmission. Using phylogenetic analyses, we show that RMP multigene families have evolved differently across the vinckei and berghei groups of RMPs and that family-specific expansions in P. chabaudi and P. vinckei occurred in the common vinckei group ancestor prior to speciation. The erythrocyte membrane antigen 1 and fam-c families in particular show considerable expansions among the lowland forest-dwelling P. vinckei parasites. The subspecies from the highland forests of Katanga, P. v. vinckei, has a uniquely smaller genome, a reduced multigene family repertoire and is also amenable to transfection making it an ideal parasite for reverse genetics. We also show that P. vinckei parasites are amenable to genetic crosses. Plasmodium vinckei isolates display a large degree of phenotypic and genotypic diversity and could serve as a resource to study parasite virulence and immunogenicity. Inclusion of P. vinckei genomes provide new insights into the evolution of RMPs and their multigene families. Amenability to genetic crossing and transfection make them also suitable for classical and functional genetics to study Plasmodium biology.
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plasmodium vinckei genomes provide insights into the pan genome and evolution of Rodent Malaria parasites
bioRxiv, 2020Co-Authors: Richard Culleton, Abhinay Ramaprasad, Arnab Pain, Severina KlausAbstract:Background Rodent Malaria parasites (RMPs) serve as tractable tools to study Malaria parasite biology and host-parasite-vector interactions. Plasmodium vinckei is the most geographically widespread of the four RMP species with isolates collected in five countries in sub-Saharan Central Africa between 1940s and 1970s. Several P. vinckei isolates are available but are relatively less characterized compared to other RMPs thus hampering its exploitation as Rodent Malaria models. We have generated a comprehensive resource for P. vinckei comprising of high-quality reference genomes, genotypes, gene expression profiles and growth phenotypes for ten P. vinckei isolates. This also allows for a comprehensive pan-genome analysis of the reference-quality genomes of RMPs. Results Plasmodium vinckei isolates display a large degree of phenotypic and genotypic diversity and potentially constitute a valuable resource to study parasite virulence and immunogenicity. The P. vinckei subspecies have diverged widely from their common ancestor and have undergone genomic structural variations. The subspecies from Katanga, P. v. vinckei, is unique among the P. vinckei isolates with a smaller genome size and a reduced multigene family repertoire. P. v. vinckei infections provide good schizont yields and is amenable to genetic manipulation, making it an ideal vinckei group parasite for reverse genetics. Comparing P. vinckei genotypes reveal region-specific selection pressures particularly on genes involved in mosquito transmission. RMP multigene family expansions observed in P. chabaudi and P. vinckei have occurred in their common ancestor prior to speciation. The erythrocyte membrane antigen 1 (ema1) and fam-c families have considerably expanded among the lowland forests-dwelling P. vinckei parasites with, however, most of the ema1 genes pseudogenised. Genetic crosses can be established in P. vinckei but are limited at present by low transmission success under the experimental conditions tested in this study. Conclusions We observe significant diversity among P. vinckei isolates making them particularly useful for the identification of genotype-phenotype relationships. Inclusion of P. vinckei genomes provide new insights into the evolution of RMPs and their multigene families. Plasmodium vinckei parasites are amenable to experimental genetic crosses and genetic manipulation, making them suitable for classical and functional genetics to study Plasmodium biology.
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a fast and cost effective microsampling protocol incorporating reduced animal usage for time series transcriptomics in Rodent Malaria parasites
Malaria Journal, 2019Co-Authors: Richard Culleton, Abhinay Ramaprasad, Amit Kumar Subudhi, Arnab PainAbstract:The transcriptional regulation that occurs in Malaria parasites during the erythrocytic stages of infection can be studied in vivo with Rodent Malaria parasites propagated in mice. Time-series transcriptome profiling commonly involves the euthanasia of groups of mice at specific time points followed by the extraction of parasite RNA from whole blood samples. Current methodologies for parasite RNA extraction involve several steps and when multiple time points are profiled, these protocols are laborious, time-consuming, and require the euthanization of large cohorts of mice. A simplified protocol has been designed for parasite RNA extraction from blood volumes as low as 20 μL (microsamples), serially bled from mice via tail snips and directly lysed with TRIzol reagent. Gene expression data derived from microsampling using RNA-seq were closely matched to those derived from larger volumes of leucocyte-depleted and saponin-treated blood obtained from euthanized mice with high reproducibility between biological replicates. Transcriptome profiling of microsamples taken at different time points during the intra-erythrocytic developmental cycle of the Rodent Malaria parasite Plasmodium vinckei revealed the transcriptional cascade commonly observed in Malaria parasites. Microsampling is a quick, robust and cost-efficient approach to sample collection for in vivo time-series transcriptomic studies in Rodent Malaria parasites.
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A fast and cost-effective microsampling protocol incorporating reduced animal usage for time-series transcriptomics in Rodent Malaria parasites
BMC, 2019Co-Authors: Abhinay Ramaprasad, Richard Culleton, Amit Kumar Subudhi, Arnab PainAbstract:Abstract Background The transcriptional regulation that occurs in Malaria parasites during the erythrocytic stages of infection can be studied in vivo with Rodent Malaria parasites propagated in mice. Time-series transcriptome profiling commonly involves the euthanasia of groups of mice at specific time points followed by the extraction of parasite RNA from whole blood samples. Current methodologies for parasite RNA extraction involve several steps and when multiple time points are profiled, these protocols are laborious, time-consuming, and require the euthanization of large cohorts of mice. Results A simplified protocol has been designed for parasite RNA extraction from blood volumes as low as 20 μL (microsamples), serially bled from mice via tail snips and directly lysed with TRIzol reagent. Gene expression data derived from microsampling using RNA-seq were closely matched to those derived from larger volumes of leucocyte-depleted and saponin-treated blood obtained from euthanized mice with high reproducibility between biological replicates. Transcriptome profiling of microsamples taken at different time points during the intra-erythrocytic developmental cycle of the Rodent Malaria parasite Plasmodium vinckei revealed the transcriptional cascade commonly observed in Malaria parasites. Conclusions Microsampling is a quick, robust and cost-efficient approach to sample collection for in vivo time-series transcriptomic studies in Rodent Malaria parasites
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a fast and cost effective microsampling protocol incorporating reduced animal usage for time series transcriptomics in Rodent Malaria parasites
bioRxiv, 2018Co-Authors: Richard Culleton, Abhinay Ramaprasad, Amit Kumar Subudhi, Arnab PainAbstract:The transcriptional regulation occurring in Malaria parasites during the clinically important life stages within host erythrocytes can be studied in vivo with Rodent Malaria parasites propagated in mice. Time-series transcriptome profiling commonly involves the euthanasia of groups of mice at specific time points followed by the extraction of parasite RNA from whole blood samples. Current methodologies for parasite RNA extraction involve several steps and when multiple time points are profiled, these protocols are laborious, time consuming, and require the euthanisation of large cohorts of mice. We designed a simplified protocol for parasite RNA extraction from blood volumes as low as 20 microliters (microsamples), serially bled from mice via tail snips and directly lysed with TRIzol reagent. Gene expression data derived from microsampling using RNA-seq were closely matched to those derived from larger volumes of leucocyte-depleted and saponin-treated blood obtained from euthanized mice and also tightly correlated between biological replicates. Transcriptome profiling of microsamples taken at different time points during the intra-erythrocytic developmental cycle of the Rodent Malaria parasite Plasmodium vinckei revealed the transcriptional cascade commonly observed in Malaria parasites. Microsampling is a quick, robust and cost-efficient approach to sample collection for in vivo time-series transcriptomic studies in Rodent Malaria parasites.
Abhinay Ramaprasad - One of the best experts on this subject based on the ideXlab platform.
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plasmodium vinckei genomes provide insights into the pan genome and evolution of Rodent Malaria parasites
BMC Biology, 2021Co-Authors: Richard Culleton, Abhinay Ramaprasad, Arnab Pain, Severina Klaus, Olga DouvropoulouAbstract:Rodent Malaria parasites (RMPs) serve as tractable tools to study Malaria parasite biology and host-parasite-vector interactions. Among the four RMPs originally collected from wild thicket rats in sub-Saharan Central Africa and adapted to laboratory mice, Plasmodium vinckei is the most geographically widespread with isolates collected from five separate locations. However, there is a lack of extensive phenotype and genotype data associated with this species, thus hindering its use in experimental studies. We have generated a comprehensive genetic resource for P. vinckei comprising of five reference-quality genomes, one for each of its subspecies, blood-stage RNA sequencing data for five P. vinckei isolates, and genotypes and growth phenotypes for ten isolates. Additionally, we sequenced seven isolates of the RMP species Plasmodium chabaudi and Plasmodium yoelii, thus extending genotypic information for four additional subspecies enabling a re-evaluation of the genotypic diversity and evolutionary history of RMPs. The five subspecies of P. vinckei have diverged widely from their common ancestor and have undergone large-scale genome rearrangements. Comparing P. vinckei genotypes reveals region-specific selection pressures particularly on genes involved in mosquito transmission. Using phylogenetic analyses, we show that RMP multigene families have evolved differently across the vinckei and berghei groups of RMPs and that family-specific expansions in P. chabaudi and P. vinckei occurred in the common vinckei group ancestor prior to speciation. The erythrocyte membrane antigen 1 and fam-c families in particular show considerable expansions among the lowland forest-dwelling P. vinckei parasites. The subspecies from the highland forests of Katanga, P. v. vinckei, has a uniquely smaller genome, a reduced multigene family repertoire and is also amenable to transfection making it an ideal parasite for reverse genetics. We also show that P. vinckei parasites are amenable to genetic crosses. Plasmodium vinckei isolates display a large degree of phenotypic and genotypic diversity and could serve as a resource to study parasite virulence and immunogenicity. Inclusion of P. vinckei genomes provide new insights into the evolution of RMPs and their multigene families. Amenability to genetic crossing and transfection make them also suitable for classical and functional genetics to study Plasmodium biology.
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plasmodium vinckei genomes provide insights into the pan genome and evolution of Rodent Malaria parasites
bioRxiv, 2020Co-Authors: Richard Culleton, Abhinay Ramaprasad, Arnab Pain, Severina KlausAbstract:Background Rodent Malaria parasites (RMPs) serve as tractable tools to study Malaria parasite biology and host-parasite-vector interactions. Plasmodium vinckei is the most geographically widespread of the four RMP species with isolates collected in five countries in sub-Saharan Central Africa between 1940s and 1970s. Several P. vinckei isolates are available but are relatively less characterized compared to other RMPs thus hampering its exploitation as Rodent Malaria models. We have generated a comprehensive resource for P. vinckei comprising of high-quality reference genomes, genotypes, gene expression profiles and growth phenotypes for ten P. vinckei isolates. This also allows for a comprehensive pan-genome analysis of the reference-quality genomes of RMPs. Results Plasmodium vinckei isolates display a large degree of phenotypic and genotypic diversity and potentially constitute a valuable resource to study parasite virulence and immunogenicity. The P. vinckei subspecies have diverged widely from their common ancestor and have undergone genomic structural variations. The subspecies from Katanga, P. v. vinckei, is unique among the P. vinckei isolates with a smaller genome size and a reduced multigene family repertoire. P. v. vinckei infections provide good schizont yields and is amenable to genetic manipulation, making it an ideal vinckei group parasite for reverse genetics. Comparing P. vinckei genotypes reveal region-specific selection pressures particularly on genes involved in mosquito transmission. RMP multigene family expansions observed in P. chabaudi and P. vinckei have occurred in their common ancestor prior to speciation. The erythrocyte membrane antigen 1 (ema1) and fam-c families have considerably expanded among the lowland forests-dwelling P. vinckei parasites with, however, most of the ema1 genes pseudogenised. Genetic crosses can be established in P. vinckei but are limited at present by low transmission success under the experimental conditions tested in this study. Conclusions We observe significant diversity among P. vinckei isolates making them particularly useful for the identification of genotype-phenotype relationships. Inclusion of P. vinckei genomes provide new insights into the evolution of RMPs and their multigene families. Plasmodium vinckei parasites are amenable to experimental genetic crosses and genetic manipulation, making them suitable for classical and functional genetics to study Plasmodium biology.
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a fast and cost effective microsampling protocol incorporating reduced animal usage for time series transcriptomics in Rodent Malaria parasites
Malaria Journal, 2019Co-Authors: Richard Culleton, Abhinay Ramaprasad, Amit Kumar Subudhi, Arnab PainAbstract:The transcriptional regulation that occurs in Malaria parasites during the erythrocytic stages of infection can be studied in vivo with Rodent Malaria parasites propagated in mice. Time-series transcriptome profiling commonly involves the euthanasia of groups of mice at specific time points followed by the extraction of parasite RNA from whole blood samples. Current methodologies for parasite RNA extraction involve several steps and when multiple time points are profiled, these protocols are laborious, time-consuming, and require the euthanization of large cohorts of mice. A simplified protocol has been designed for parasite RNA extraction from blood volumes as low as 20 μL (microsamples), serially bled from mice via tail snips and directly lysed with TRIzol reagent. Gene expression data derived from microsampling using RNA-seq were closely matched to those derived from larger volumes of leucocyte-depleted and saponin-treated blood obtained from euthanized mice with high reproducibility between biological replicates. Transcriptome profiling of microsamples taken at different time points during the intra-erythrocytic developmental cycle of the Rodent Malaria parasite Plasmodium vinckei revealed the transcriptional cascade commonly observed in Malaria parasites. Microsampling is a quick, robust and cost-efficient approach to sample collection for in vivo time-series transcriptomic studies in Rodent Malaria parasites.
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A fast and cost-effective microsampling protocol incorporating reduced animal usage for time-series transcriptomics in Rodent Malaria parasites
BMC, 2019Co-Authors: Abhinay Ramaprasad, Richard Culleton, Amit Kumar Subudhi, Arnab PainAbstract:Abstract Background The transcriptional regulation that occurs in Malaria parasites during the erythrocytic stages of infection can be studied in vivo with Rodent Malaria parasites propagated in mice. Time-series transcriptome profiling commonly involves the euthanasia of groups of mice at specific time points followed by the extraction of parasite RNA from whole blood samples. Current methodologies for parasite RNA extraction involve several steps and when multiple time points are profiled, these protocols are laborious, time-consuming, and require the euthanization of large cohorts of mice. Results A simplified protocol has been designed for parasite RNA extraction from blood volumes as low as 20 μL (microsamples), serially bled from mice via tail snips and directly lysed with TRIzol reagent. Gene expression data derived from microsampling using RNA-seq were closely matched to those derived from larger volumes of leucocyte-depleted and saponin-treated blood obtained from euthanized mice with high reproducibility between biological replicates. Transcriptome profiling of microsamples taken at different time points during the intra-erythrocytic developmental cycle of the Rodent Malaria parasite Plasmodium vinckei revealed the transcriptional cascade commonly observed in Malaria parasites. Conclusions Microsampling is a quick, robust and cost-efficient approach to sample collection for in vivo time-series transcriptomic studies in Rodent Malaria parasites
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a fast and cost effective microsampling protocol incorporating reduced animal usage for time series transcriptomics in Rodent Malaria parasites
bioRxiv, 2018Co-Authors: Richard Culleton, Abhinay Ramaprasad, Amit Kumar Subudhi, Arnab PainAbstract:The transcriptional regulation occurring in Malaria parasites during the clinically important life stages within host erythrocytes can be studied in vivo with Rodent Malaria parasites propagated in mice. Time-series transcriptome profiling commonly involves the euthanasia of groups of mice at specific time points followed by the extraction of parasite RNA from whole blood samples. Current methodologies for parasite RNA extraction involve several steps and when multiple time points are profiled, these protocols are laborious, time consuming, and require the euthanisation of large cohorts of mice. We designed a simplified protocol for parasite RNA extraction from blood volumes as low as 20 microliters (microsamples), serially bled from mice via tail snips and directly lysed with TRIzol reagent. Gene expression data derived from microsampling using RNA-seq were closely matched to those derived from larger volumes of leucocyte-depleted and saponin-treated blood obtained from euthanized mice and also tightly correlated between biological replicates. Transcriptome profiling of microsamples taken at different time points during the intra-erythrocytic developmental cycle of the Rodent Malaria parasite Plasmodium vinckei revealed the transcriptional cascade commonly observed in Malaria parasites. Microsampling is a quick, robust and cost-efficient approach to sample collection for in vivo time-series transcriptomic studies in Rodent Malaria parasites.
Arnab Pain - One of the best experts on this subject based on the ideXlab platform.
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plasmodium vinckei genomes provide insights into the pan genome and evolution of Rodent Malaria parasites
BMC Biology, 2021Co-Authors: Richard Culleton, Abhinay Ramaprasad, Arnab Pain, Severina Klaus, Olga DouvropoulouAbstract:Rodent Malaria parasites (RMPs) serve as tractable tools to study Malaria parasite biology and host-parasite-vector interactions. Among the four RMPs originally collected from wild thicket rats in sub-Saharan Central Africa and adapted to laboratory mice, Plasmodium vinckei is the most geographically widespread with isolates collected from five separate locations. However, there is a lack of extensive phenotype and genotype data associated with this species, thus hindering its use in experimental studies. We have generated a comprehensive genetic resource for P. vinckei comprising of five reference-quality genomes, one for each of its subspecies, blood-stage RNA sequencing data for five P. vinckei isolates, and genotypes and growth phenotypes for ten isolates. Additionally, we sequenced seven isolates of the RMP species Plasmodium chabaudi and Plasmodium yoelii, thus extending genotypic information for four additional subspecies enabling a re-evaluation of the genotypic diversity and evolutionary history of RMPs. The five subspecies of P. vinckei have diverged widely from their common ancestor and have undergone large-scale genome rearrangements. Comparing P. vinckei genotypes reveals region-specific selection pressures particularly on genes involved in mosquito transmission. Using phylogenetic analyses, we show that RMP multigene families have evolved differently across the vinckei and berghei groups of RMPs and that family-specific expansions in P. chabaudi and P. vinckei occurred in the common vinckei group ancestor prior to speciation. The erythrocyte membrane antigen 1 and fam-c families in particular show considerable expansions among the lowland forest-dwelling P. vinckei parasites. The subspecies from the highland forests of Katanga, P. v. vinckei, has a uniquely smaller genome, a reduced multigene family repertoire and is also amenable to transfection making it an ideal parasite for reverse genetics. We also show that P. vinckei parasites are amenable to genetic crosses. Plasmodium vinckei isolates display a large degree of phenotypic and genotypic diversity and could serve as a resource to study parasite virulence and immunogenicity. Inclusion of P. vinckei genomes provide new insights into the evolution of RMPs and their multigene families. Amenability to genetic crossing and transfection make them also suitable for classical and functional genetics to study Plasmodium biology.
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plasmodium vinckei genomes provide insights into the pan genome and evolution of Rodent Malaria parasites
bioRxiv, 2020Co-Authors: Richard Culleton, Abhinay Ramaprasad, Arnab Pain, Severina KlausAbstract:Background Rodent Malaria parasites (RMPs) serve as tractable tools to study Malaria parasite biology and host-parasite-vector interactions. Plasmodium vinckei is the most geographically widespread of the four RMP species with isolates collected in five countries in sub-Saharan Central Africa between 1940s and 1970s. Several P. vinckei isolates are available but are relatively less characterized compared to other RMPs thus hampering its exploitation as Rodent Malaria models. We have generated a comprehensive resource for P. vinckei comprising of high-quality reference genomes, genotypes, gene expression profiles and growth phenotypes for ten P. vinckei isolates. This also allows for a comprehensive pan-genome analysis of the reference-quality genomes of RMPs. Results Plasmodium vinckei isolates display a large degree of phenotypic and genotypic diversity and potentially constitute a valuable resource to study parasite virulence and immunogenicity. The P. vinckei subspecies have diverged widely from their common ancestor and have undergone genomic structural variations. The subspecies from Katanga, P. v. vinckei, is unique among the P. vinckei isolates with a smaller genome size and a reduced multigene family repertoire. P. v. vinckei infections provide good schizont yields and is amenable to genetic manipulation, making it an ideal vinckei group parasite for reverse genetics. Comparing P. vinckei genotypes reveal region-specific selection pressures particularly on genes involved in mosquito transmission. RMP multigene family expansions observed in P. chabaudi and P. vinckei have occurred in their common ancestor prior to speciation. The erythrocyte membrane antigen 1 (ema1) and fam-c families have considerably expanded among the lowland forests-dwelling P. vinckei parasites with, however, most of the ema1 genes pseudogenised. Genetic crosses can be established in P. vinckei but are limited at present by low transmission success under the experimental conditions tested in this study. Conclusions We observe significant diversity among P. vinckei isolates making them particularly useful for the identification of genotype-phenotype relationships. Inclusion of P. vinckei genomes provide new insights into the evolution of RMPs and their multigene families. Plasmodium vinckei parasites are amenable to experimental genetic crosses and genetic manipulation, making them suitable for classical and functional genetics to study Plasmodium biology.
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a fast and cost effective microsampling protocol incorporating reduced animal usage for time series transcriptomics in Rodent Malaria parasites
Malaria Journal, 2019Co-Authors: Richard Culleton, Abhinay Ramaprasad, Amit Kumar Subudhi, Arnab PainAbstract:The transcriptional regulation that occurs in Malaria parasites during the erythrocytic stages of infection can be studied in vivo with Rodent Malaria parasites propagated in mice. Time-series transcriptome profiling commonly involves the euthanasia of groups of mice at specific time points followed by the extraction of parasite RNA from whole blood samples. Current methodologies for parasite RNA extraction involve several steps and when multiple time points are profiled, these protocols are laborious, time-consuming, and require the euthanization of large cohorts of mice. A simplified protocol has been designed for parasite RNA extraction from blood volumes as low as 20 μL (microsamples), serially bled from mice via tail snips and directly lysed with TRIzol reagent. Gene expression data derived from microsampling using RNA-seq were closely matched to those derived from larger volumes of leucocyte-depleted and saponin-treated blood obtained from euthanized mice with high reproducibility between biological replicates. Transcriptome profiling of microsamples taken at different time points during the intra-erythrocytic developmental cycle of the Rodent Malaria parasite Plasmodium vinckei revealed the transcriptional cascade commonly observed in Malaria parasites. Microsampling is a quick, robust and cost-efficient approach to sample collection for in vivo time-series transcriptomic studies in Rodent Malaria parasites.
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A fast and cost-effective microsampling protocol incorporating reduced animal usage for time-series transcriptomics in Rodent Malaria parasites
BMC, 2019Co-Authors: Abhinay Ramaprasad, Richard Culleton, Amit Kumar Subudhi, Arnab PainAbstract:Abstract Background The transcriptional regulation that occurs in Malaria parasites during the erythrocytic stages of infection can be studied in vivo with Rodent Malaria parasites propagated in mice. Time-series transcriptome profiling commonly involves the euthanasia of groups of mice at specific time points followed by the extraction of parasite RNA from whole blood samples. Current methodologies for parasite RNA extraction involve several steps and when multiple time points are profiled, these protocols are laborious, time-consuming, and require the euthanization of large cohorts of mice. Results A simplified protocol has been designed for parasite RNA extraction from blood volumes as low as 20 μL (microsamples), serially bled from mice via tail snips and directly lysed with TRIzol reagent. Gene expression data derived from microsampling using RNA-seq were closely matched to those derived from larger volumes of leucocyte-depleted and saponin-treated blood obtained from euthanized mice with high reproducibility between biological replicates. Transcriptome profiling of microsamples taken at different time points during the intra-erythrocytic developmental cycle of the Rodent Malaria parasite Plasmodium vinckei revealed the transcriptional cascade commonly observed in Malaria parasites. Conclusions Microsampling is a quick, robust and cost-efficient approach to sample collection for in vivo time-series transcriptomic studies in Rodent Malaria parasites
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a fast and cost effective microsampling protocol incorporating reduced animal usage for time series transcriptomics in Rodent Malaria parasites
bioRxiv, 2018Co-Authors: Richard Culleton, Abhinay Ramaprasad, Amit Kumar Subudhi, Arnab PainAbstract:The transcriptional regulation occurring in Malaria parasites during the clinically important life stages within host erythrocytes can be studied in vivo with Rodent Malaria parasites propagated in mice. Time-series transcriptome profiling commonly involves the euthanasia of groups of mice at specific time points followed by the extraction of parasite RNA from whole blood samples. Current methodologies for parasite RNA extraction involve several steps and when multiple time points are profiled, these protocols are laborious, time consuming, and require the euthanisation of large cohorts of mice. We designed a simplified protocol for parasite RNA extraction from blood volumes as low as 20 microliters (microsamples), serially bled from mice via tail snips and directly lysed with TRIzol reagent. Gene expression data derived from microsampling using RNA-seq were closely matched to those derived from larger volumes of leucocyte-depleted and saponin-treated blood obtained from euthanized mice and also tightly correlated between biological replicates. Transcriptome profiling of microsamples taken at different time points during the intra-erythrocytic developmental cycle of the Rodent Malaria parasite Plasmodium vinckei revealed the transcriptional cascade commonly observed in Malaria parasites. Microsampling is a quick, robust and cost-efficient approach to sample collection for in vivo time-series transcriptomic studies in Rodent Malaria parasites.
Chris J. Janse - One of the best experts on this subject based on the ideXlab platform.
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the plasmodium phist and resa like protein families of human and Rodent Malaria parasites
PLOS ONE, 2016Co-Authors: Cristina K Moreira, Photini Sinnis, Blandine Frankefayard, Chris J. Janse, Bernina Naissant, Alida Coppi, Brandy L Bennett, Elena Aime, Isabelle CoppensAbstract:The phist gene family has members identified across the Plasmodium genus, defined by the presence of a domain of roughly 150 amino acids having conserved aromatic residues and an all alpha-helical structure. The family is highly amplified in P. falciparum, with 65 predicted genes in the genome of the 3D7 isolate. In contrast, in the Rodent Malaria parasite P. berghei 3 genes are identified, one of which is an apparent pseudogene. Transcripts of the P. berghei phist genes are predominant in schizonts, whereas in P. falciparum transcript profiles span different asexual blood stages and gametocytes. We pursued targeted disruption of P. berghei phist genes in order to characterize a simplistic model for the expanded phist gene repertoire in P. falciparum. Unsuccessful attempts to disrupt P. berghei PBANKA_114540 suggest that this phist gene is essential, while knockout of phist PBANKA_122900 shows an apparent normal progression and non-essential function throughout the life cycle. Epitope-tagging of P. falciparum and P. berghei phist genes confirmed protein export to the erythrocyte cytoplasm and localization with a punctate pattern. Three P. berghei PEXEL/HT-positive exported proteins exhibit at least partial co-localization, in support of a common vesicular compartment in the cytoplasm of erythrocytes infected with Rodent Malaria parasites.
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generation of transgenic Rodent Malaria parasites expressing human Malaria parasite proteins
Methods of Molecular Biology, 2015Co-Authors: Chris J. Janse, Ahmed M Salman, Catherin Marin Mogollon, Jingwen Lin, Fiona J A Van Pul, Shahid M. KhanAbstract:We describe methods for the rapid generation of transgenic Rodent Plasmodium berghei (Pb) parasites that express human Malaria parasite (HMP) proteins, using the recently developed GIMO-based transfection methodology. Three different genetic modifications are described resulting in three types of transgenic parasites. (1) Additional Gene (AG) mutants. In these mutants the HMP gene is introduced as an "additional gene" into a silent/neutral locus of the Pb genome under the control of either a constitutive or stage-specific Pb promoter. This method uses the GIMO-transfection protocol and AG mutants are generated by replacing the positive-negative selection marker (SM) hdhfr::yfcu cassette in a neutral locus of a standard GIMO mother line with the HMP gene expression cassette, resulting in SM free transgenic parasites. (2) Double-step Replacement (DsR) mutants. In these mutants the coding sequence (CDS) of the Pb gene is replaced with the CDS of the HMP ortholog in a two-step GIMO-transfection procedure. This process involves first the replacement of the Pb CDS with the hdhfr::yfcu SM, followed by insertion of the HMP ortholog at the same locus thereby replacing hdhfr::yfcu with the HMP CDS. These steps use the GIMO-transfection protocol, which exploits both positive selection for Pb orthologous gene-deletion and negative selection for HMP gene-insertion, resulting in SM free transgenic parasites. (3) Double-step Insertion (DsI) mutants. When a Pb gene is essential for blood stage development the DsR strategy is not possible. In these mutants the HMP expression cassette is first introduced into the neutral locus in a standard GIMO mother line as described for AG mutants but under the control elements of the Pb orthologous gene; subsequently, the Pb ortholog CDS is targeted for deletion through replacement of the Pb CDS with the hdhfr::yfcu SM, resulting in transgenic parasites with a new GIMO locus permissive for additional gene-insertion modifications.The different types of transgenic parasites can be exploited to examine interactions of drugs/inhibitors or immune factors with HMP molecules in vivo. Mice either immunized with HMP-vaccines or treated with specific drugs can be infected/challenged with these transgenic mutants to evaluate drug or vaccine efficacy in vivo.
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standardization in generating and reporting genetically modified Rodent Malaria parasites the rmgmdb database
Methods of Molecular Biology, 2012Co-Authors: Shahid M. Khan, Blandine Frankefayard, Hans Kroeze, Chris J. JanseAbstract:Genetically modified Plasmodium parasites are central gene function reagents in Malaria research. The Rodent Malaria genetically modified DataBase (RMgmDB) ( www.pberghei.eu ) is a manually curated Web - based repository that contains information on genetically modified Rodent Malaria parasites. It provides easy and rapid access to information on the genotype and phenotype of genetically modified mutant and reporter parasites. Here, we provide guidelines for generating and describing Rodent Malaria parasite mutants. Standardization in describing mutant genotypes and phenotypes is important not only to enhance publication quality but also to facilitate cross-linking and mining data from multiple sources, and should permit information derived from mutant parasites to be used in integrative system biology approaches. We also provide guidelines on how to submit information to RMgmDB on non-published mutants, mutants that do not exhibit a clear phenotype, as well as negative attempts to disrupt/mutate genes. Such information helps to prevent unnecessary duplication of experiments in different laboratories, and can provide indirect evidence that these genes are essential for blood-stage development.
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a novel gene insertion marker out gimo method for transgene expression and gene complementation in Rodent Malaria parasites
PLOS ONE, 2011Co-Authors: Takeshi Annoura, Onny Klop, Severine Chevalleymaurel, Mohammed Sajid, Blandine Frankefayard, Jai Ramesar, Chris J. Janse, Shahid M. KhanAbstract:Research on the biology of Malaria parasites has greatly benefited from the application of reverse genetic technologies, in particular through the analysis of gene deletion mutants and studies on transgenic parasites that express heterologous or mutated proteins. However, transfection in Plasmodium is limited by the paucity of drug-selectable markers that hampers subsequent genetic modification of the same mutant. We report the development of a novel ‘gene insertion/marker out’ (GIMO) method for two Rodent Malaria parasites, which uses negative selection to rapidly generate transgenic mutants ready for subsequent modifications. We have created reference mother lines for both P. berghei ANKA and P. yoelii 17XNL that serve as recipient parasites for GIMO-transfection. Compared to existing protocols GIMO-transfection greatly simplifies and speeds up the generation of mutants expressing heterologous proteins, free of drug-resistance genes, and requires far fewer laboratory animals. In addition we demonstrate that GIMO-transfection is also a simple and fast method for genetic complementation of mutants with a gene deletion or mutation. The implementation of GIMO-transfection procedures should greatly enhance Plasmodium reverse-genetic research.
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Selection by flow-sorting of genetically transformed, GFP-expressing blood stages of the Rodent Malaria parasite, Plasmodium berghei
Nature Protocols, 2006Co-Authors: Chris J. Janse, Blandine Franke-fayard, Andrew P WatersAbstract:This protocol describes a methodology for the genetic transformation of the Rodent Malaria parasite Plasmodium berghei and the subsequent selection of transformed parasites expressing green fluorescent protein (GFP) by flow-sorting. It provides methods for: transfection of the schizont stage with DNA constructs that contain gfp as the selectable marker; selection of fluorescent mutants by flow-sorting; and injection of flow-sorted, GFP-expressing parasites into mice and the subsequent collection of transformed parasites. The use of two different promoters for the expression of GFP is described; these two promoters require slightly different procedures for the selection of mutants. The protocol enables the collection of transformed parasites within 10–12 days after transfection. The genetic modification of P. berghei is widely used to investigate gene function in Plasmodium sp. The application of flow-sorting to the selection of transformed parasites increases the possibilities of parasite mutagenesis, by effectively expanding the range of selectable markers.
