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Identification and characterization of the Plasmodium vivax thrombospondin-related apical merozoite protein.

Mongui A, Angel DI, Moreno-Perez DA, Villarreal-Gonzalez S, Almonacid H, Vanegas M, Patarroyo MA - Malar. J. (2010)

Bottom Line: Despite its epidemiological importance, few antigens from this parasite species have been characterized to date compared to Plasmodium falciparum, due in part to the difficulties of maintaining an in vitro culture of P. vivax.Recombinant PvTRAMP is recognized by sera from infected patients, therefore, indicating that it is targeted by the immune system during a natural infection with P. vivax.The results of this work support conducting further studies with PvTRAMP to evaluate its immunogenicity and protection-inducing ability in the Aotus animal model.

View Article: PubMed Central - HTML - PubMed

Affiliation: Fundación Instituto de Inmunología de Colombia (FIDIC), Bogotá, Colombia.

ABSTRACT

Background: Malaria caused by Plasmodium vivax is a major public health problem worldwide that affects 70-80 million people in the Middle East, Asia, Western Pacific, South America and the Caribbean. Despite its epidemiological importance, few antigens from this parasite species have been characterized to date compared to Plasmodium falciparum, due in part to the difficulties of maintaining an in vitro culture of P. vivax. This study describes the identification of the P. falciparum thrombospondin-related apical merozoite protein homologue in P. vivax (PvTRAMP) and examines its potential to be further evaluated as vaccine candidate.

Methods: The gene encoding PvTRAMP was identified through an extensive search of the databases hosting the genome sequence of P. vivax. Genes adjacent to pvtramp were identified in silico to determine the degree of similarity between the protein sequences encoded by equivalent chromosomic fragments in P. falciparum and Plasmodium knowlesi. The pvtramp gene was amplified from cDNA of P. vivax schizont stages, cloned and expressed in Escherichia coli. Anti-PvTRAMP antisera was obtained by inoculating rabbits with PvTRAMP B cell epitopes produced as synthetic peptides in order to assess its recognition in parasite lysates by Western blot and in intact parasites by indirect immunofluorescence. The recognition of recombinant PvTRAMP by sera from P. vivax-infected individuals living in endemic areas was also assessed by ELISA.

Results: The PfTRAMP homologue in P. vivax, here denoted as PvTRAMP, is a 340-amino-acid long antigen encoded by a single exon that could have a potential role in cytoadherence, as indicated by the presence of a thrombospondin structural homology repeat (TSR) domain. According to its transcription and expression profile, PvTRAMP is initially located at the parasite's apical end and later on the parasite surface. Recombinant PvTRAMP is recognized by sera from infected patients, therefore, indicating that it is targeted by the immune system during a natural infection with P. vivax.

Conclusions: The results of this work support conducting further studies with PvTRAMP to evaluate its immunogenicity and protection-inducing ability in the Aotus animal model.

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Cellular localization of PvTRAMP as assessed by IFA using hyper-immune anti-PvTRAMP rabbit sera as primary antibody. (A-C) Detection of P. vivax in early schizont stages. (D-I) Parasites in late schizont stage (segmented). The figure shows fluorescence with DAPI and FITC staining, and the merging of both.
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Figure 3: Cellular localization of PvTRAMP as assessed by IFA using hyper-immune anti-PvTRAMP rabbit sera as primary antibody. (A-C) Detection of P. vivax in early schizont stages. (D-I) Parasites in late schizont stage (segmented). The figure shows fluorescence with DAPI and FITC staining, and the merging of both.

Mentions: Immunofluorescence assays using blood smears from P. vivax-infected Aotus monkeys and pooled rabbit hyperimmune sera as primary antibody detected PvTRAMP in schizont stages. As shown in Figure 3, a fluorescence pattern typical of merozoite surface proteins with some apparent concentration towards the apical pole was observed, which is similar to the fluorescence pattern reported for PfTRAMP. Localization assays performed by Thompson et al [26] using a combination of antisera against the rhoptry-associated protein RAP1 and the apical membrane antigen AMA1 showed that PfTRAMP co-localizes with AMA-1 in apical organelles from early schizonts, but displays a distinct localization pattern compared to RAP1 [26,42]. This has lead the authors to suggest that PfTRAMP originates at the micronemes and relocalizes to merozoite surface before initiation of RBC invasion. Further confocal microscopy studies with more specific antibodies would help defining whether P. vivax TRAMP exhibits this localization pattern.


Identification and characterization of the Plasmodium vivax thrombospondin-related apical merozoite protein.

Mongui A, Angel DI, Moreno-Perez DA, Villarreal-Gonzalez S, Almonacid H, Vanegas M, Patarroyo MA - Malar. J. (2010)

Cellular localization of PvTRAMP as assessed by IFA using hyper-immune anti-PvTRAMP rabbit sera as primary antibody. (A-C) Detection of P. vivax in early schizont stages. (D-I) Parasites in late schizont stage (segmented). The figure shows fluorescence with DAPI and FITC staining, and the merging of both.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3020679&req=5

Figure 3: Cellular localization of PvTRAMP as assessed by IFA using hyper-immune anti-PvTRAMP rabbit sera as primary antibody. (A-C) Detection of P. vivax in early schizont stages. (D-I) Parasites in late schizont stage (segmented). The figure shows fluorescence with DAPI and FITC staining, and the merging of both.
Mentions: Immunofluorescence assays using blood smears from P. vivax-infected Aotus monkeys and pooled rabbit hyperimmune sera as primary antibody detected PvTRAMP in schizont stages. As shown in Figure 3, a fluorescence pattern typical of merozoite surface proteins with some apparent concentration towards the apical pole was observed, which is similar to the fluorescence pattern reported for PfTRAMP. Localization assays performed by Thompson et al [26] using a combination of antisera against the rhoptry-associated protein RAP1 and the apical membrane antigen AMA1 showed that PfTRAMP co-localizes with AMA-1 in apical organelles from early schizonts, but displays a distinct localization pattern compared to RAP1 [26,42]. This has lead the authors to suggest that PfTRAMP originates at the micronemes and relocalizes to merozoite surface before initiation of RBC invasion. Further confocal microscopy studies with more specific antibodies would help defining whether P. vivax TRAMP exhibits this localization pattern.

Bottom Line: Despite its epidemiological importance, few antigens from this parasite species have been characterized to date compared to Plasmodium falciparum, due in part to the difficulties of maintaining an in vitro culture of P. vivax.Recombinant PvTRAMP is recognized by sera from infected patients, therefore, indicating that it is targeted by the immune system during a natural infection with P. vivax.The results of this work support conducting further studies with PvTRAMP to evaluate its immunogenicity and protection-inducing ability in the Aotus animal model.

View Article: PubMed Central - HTML - PubMed

Affiliation: Fundación Instituto de Inmunología de Colombia (FIDIC), Bogotá, Colombia.

ABSTRACT

Background: Malaria caused by Plasmodium vivax is a major public health problem worldwide that affects 70-80 million people in the Middle East, Asia, Western Pacific, South America and the Caribbean. Despite its epidemiological importance, few antigens from this parasite species have been characterized to date compared to Plasmodium falciparum, due in part to the difficulties of maintaining an in vitro culture of P. vivax. This study describes the identification of the P. falciparum thrombospondin-related apical merozoite protein homologue in P. vivax (PvTRAMP) and examines its potential to be further evaluated as vaccine candidate.

Methods: The gene encoding PvTRAMP was identified through an extensive search of the databases hosting the genome sequence of P. vivax. Genes adjacent to pvtramp were identified in silico to determine the degree of similarity between the protein sequences encoded by equivalent chromosomic fragments in P. falciparum and Plasmodium knowlesi. The pvtramp gene was amplified from cDNA of P. vivax schizont stages, cloned and expressed in Escherichia coli. Anti-PvTRAMP antisera was obtained by inoculating rabbits with PvTRAMP B cell epitopes produced as synthetic peptides in order to assess its recognition in parasite lysates by Western blot and in intact parasites by indirect immunofluorescence. The recognition of recombinant PvTRAMP by sera from P. vivax-infected individuals living in endemic areas was also assessed by ELISA.

Results: The PfTRAMP homologue in P. vivax, here denoted as PvTRAMP, is a 340-amino-acid long antigen encoded by a single exon that could have a potential role in cytoadherence, as indicated by the presence of a thrombospondin structural homology repeat (TSR) domain. According to its transcription and expression profile, PvTRAMP is initially located at the parasite's apical end and later on the parasite surface. Recombinant PvTRAMP is recognized by sera from infected patients, therefore, indicating that it is targeted by the immune system during a natural infection with P. vivax.

Conclusions: The results of this work support conducting further studies with PvTRAMP to evaluate its immunogenicity and protection-inducing ability in the Aotus animal model.

Show MeSH
Related in: MedlinePlus