Transcriptome profiling of<i>Plasmodium vivax</i>in<i>Saimiri</i>monkeys identifies potential ligands for invasion

Gunalan, Karthigayan; Sá, Juliana M.; Barros, Roberto R. Moraes; Anzick, Sarah L.; Caleon, Ramoncito L.; Mershon, J. Patrick; Kanakabandi, Kishore; Paneru, Monica; Virtaneva, Kimmo; Martens, Craig; Barnwell, John W.; Ribeiro, José M. C.; Miller, Louis H.

doi:10.1073/pnas.1818485116

“…Prior studies have implicated that high levels of invasion related proteins like Duffy binding protein (DBP1) could enable parasites to invade duffy negative erythrocytes, and P. vivax may additionally have alternative invasion pathways independent of DBP1 [29][30][31] . Recent studies of SalI parasites infecting Aotus and Saimiri monkeys looked at potentially unique invasion pathways and implicated the differential expression of the tryptophan-rich protein family in mediating this process 18 . No clear pattern of differential expression was shown in the four patient isolates studied, however the expression heterogeneity of these erythrocyte-binding proteins in this study and several others warrants further investigation about the involvement of these in host recognition.…”

Section: Discussionmentioning

confidence: 99%

“…The first study to use RNA-seq in P. vivax was able to describe the features of the transcriptome more comprehensively, as previously unannotated untranslated regions (UTRs) and splice sites were outside the technical capabilities of microarrays 14 . Other previous studies of bulk RNA-seq from patient isolates have been performed on both a time course and with asynchronous samples [15][16][17] , and with primates in laboratory conditions 18 .…”

mentioning

confidence: 99%

Analysis of Plasmodium vivax schizont transcriptomes from field isolates reveals heterogeneity of expression of genes involved in host-parasite interactions

Siegel

¹

,

Chappell

²

,

Hostetler

³

et al. 2020

Sci Rep

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Plasmodium vivax gene regulation remains difficult to study due to the lack of a robust in vitro culture method, low parasite densities in peripheral circulation and asynchronous parasite development. We adapted an RNA-seq protocol “DAFT-seq” to sequence the transcriptome of four P. vivax field isolates that were cultured for a short period ex vivo before using a density gradient for schizont enrichment. Transcription was detected from 78% of the PvP01 reference genome, despite being schizont-enriched samples. This extensive data was used to define thousands of 5′ and 3′ untranslated regions, some of which overlapped with neighbouring transcripts, and to improve the gene models of 352 genes, including identifying 20 novel gene transcripts. This dataset has also significantly increased the known amount of heterogeneity between P. vivax schizont transcriptomes from individual patients. The majority of genes found to be differentially expressed between the isolates lack Plasmodium falciparum homologs and are predicted to be involved in host-parasite interactions, with an enrichment in reticulocyte binding proteins, merozoite surface proteins and exported proteins with unknown function. An improved understanding of the diversity within P. vivax transcriptomes will be essential for the prioritisation of novel vaccine targets.

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“…Prior studies have implicated that high levels of invasion related proteins like Duffy binding protein (DBP1) could enable parasites to invade duffy negative erythrocytes, and P. vivax may additionally have alternative invasion pathways independent of DBP1 [29][30][31] . Recent studies of SalI parasites infecting Aotus and Saimiri monkeys looked at potentially unique invasion pathways and implicated the differential expression of the tryptophanrich protein family in mediating this process 18 . No clear pattern of differential expression was shown in the four patient isolates studied, however the expression heterogeneity of these erythrocyte-binding proteins in this study and several others warrants further investigation about the involvement of these in host recognition.…”

Section: Discussionmentioning

confidence: 99%

Analysis ofPlasmodium vivaxschizont transcriptomes from field isolates reveals heterogeneity of expression of genes involved in host-parasite interactions

Siegel

¹

,

Chappell

²

,

Hostetler

³

et al. 2020

Preprint

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Plasmodium vivax gene regulation remains difficult to study due to the lack of a robust in vitro culture method, low parasite densities in peripheral circulation and asynchronous parasite development. We adapted an RNA-seq protocol "DAFT-seq" to sequence the transcriptome of four P. vivax field isolates that were cultured for a short period ex vivo before using a density gradient for schizont enrichment. Transcription was detected from 78% of the PvP01 reference genome, despite being schizont-enriched samples. This extensive data was used to define thousands of 5' and 3' untranslated regions (UTRs), some of which overlapped with neighbouring transcripts, and to improve the gene models of 352 genes, including identifying 20 novel gene transcripts. This dataset has also significantly increased the known amount of heterogeneity between P. vivax schizont transcriptomes from individual patients. The majority of genes found to be differentially expressed between the isolates lack Plasmodium falciparum homologs and are predicted to be involved in host-parasite interactions, with an enrichment in reticulocyte binding proteins, merozoite surface proteins and exported proteins with unknown function. An improved understanding of the diversity within P. vivax transcriptomes will be essential for the prioritisation of novel vaccine targets. Results Preparation of purified late-stage schizont transcriptomesFour blood samples from Cambodian patients were selected to undergo short-term ex-vivo culture. After the majority of parasites had matured, as judged by microscopy, late-stage schizonts were purified using Percoll gradients. Four RNA-seq libraries were generated using a modified version of the DAFT-seq protocol, which was optimised for highly AT-rich Plasmodium parasites 23 , and sequenced using the Illumina platform to generate 55-63 million reads per patient sample. In all cases >85% of reads mapped to the P. vivax PvP01 reference genome (Table S1). This data was used to improve the gene models of 352 genes in the PvP01 genome, including identifying 20 novel gene transcripts (Table S2). Comparison of the expression values of these RNA-seq libraries to blood stage microarray time course from a P. vivax dataset (containing three patient isolates) 12 (Fig. S1) and a more densely sampled P. falciparum dataset 24 (Fig. S2) confirmed that the samples were late-stage schizonts (as they correlated most strongly to these time points in the prior datasets) and that the transcriptomes were highly similar to each other (as the correlation of the normalised expression values of the RNA-seq libraries was close to 1) (Fig. S3). More apparent heterogeneity was found between the patient isolates using the PVP01 genome than using only the gene IDs present in the Sal1 genome (Fig. S4), consistent with most of the heterogeneity within the patient isolate transcriptomes being present in multigene families that are difficult to assemble and annotate, and are thus under-represented in the Sal1 genome.

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“…Similarly, by comparing the gene expression profiles of parasites infecting Duffy-negative or Duffy-positive people, the signature of genes specifically regulated to respond to the Duffy negativity barrier might be identified. Such an approach could also make use of the Saimiri and Aotus nonhuman primate model-as both species can be infected by P. vivax, while PvDBP can bind only to Aotus erythrocytes and not to Saimiri ones, indicating an alternative invasion pathway reminiscent of what is observed for Duffy-negative human erythrocytes [90]. Alternative models to decipher erythrocyte invasion mechanisms could also rely on the genetically tractable P. knowlesi that shares many biological similarities with P. vivax, including the requirement of the Duffy receptor for erythrocyte invasion [91] or even on P. cynomolgi, closely related to P. vivax and recently adapted to in vitro culture [92].…”

Section: Future Directions and Conclusive Remarksmentioning

confidence: 99%

The enigmatic mechanisms by which Plasmodium vivax infects Duffy-negative individuals

Popovici

¹

,

Roesch

²

,

Rougeron

³

2020

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The absence of the Duffy protein at the surface of erythrocytes was considered for decades to confer full protection against Plasmodium vivax as this blood group is the receptor for the key parasite ligand P. vivax Duffy binding protein (PvDBP). However, it is now clear that the parasite is able to break through this protection and induce clinical malaria in Duffy-negative people, although the underlying mechanisms are still not understood. Here, we briefly review the evidence of Duffy-negative infections by P. vivax and summarize the current hypothesis at the basis of this invasion process. We discuss those in the perspective of malaria-elimination challenges, notably in African countries.

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Transcriptome profiling ofPlasmodium vivaxinSaimirimonkeys identifies potential ligands for invasion

Cited by 27 publications

References 66 publications

Analysis of Plasmodium vivax schizont transcriptomes from field isolates reveals heterogeneity of expression of genes involved in host-parasite interactions

Analysis of Plasmodium vivax schizont transcriptomes from field isolates reveals heterogeneity of expression of genes involved in host-parasite interactions

Analysis ofPlasmodium vivaxschizont transcriptomes from field isolates reveals heterogeneity of expression of genes involved in host-parasite interactions

The enigmatic mechanisms by which Plasmodium vivax infects Duffy-negative individuals

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