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Immunoglobulin response to Plasmodium falciparum RESA proteins in uncomplicated and severe malaria.

Badaut C, Guyonnet L, Milet J, Renard E, Durand R, Viwami F, Sagbo G, Layla F, Deloron P, Bonnefoy S, Migot-Nabias F - Malar. J. (2015)

Bottom Line: Taking into account age, fever duration and parasitaemia, a multiple logistic regression performed on children clustered according to their antibody responses' profiles concluded to an increased risk of severe malaria for P2 (representative of RESA-1) responders (P = 0.007).Increased IL-10 plasma levels were found in children harbouring multiclonal P. falciparum infections on the basis of the T1526G resa2 gene polymorphism (P = 0.004).It suggested the measurement of plasma antibodies raised against specific peptides to serve as predictive immunologic markers for disease severity.

View Article: PubMed Central - PubMed

Affiliation: Equipe résidente de recherche en infectiologie tropicale, Institut de Recherche Biomédicale des Armées (IRBA), Brétigny sur Orge, France. cbadaut@gmail.com.

ABSTRACT

Background: The three members of the ring-infected erythrocyte surface antigen (RESA) proteins family share high sequence homologies, which impair the detection and assignment to one or another protein of some pathogenic processes inherent to Plasmodium falciparum malaria. The present study was intended to determine if the antibody and inflammatory responses of children living in a malaria-endemic area varied depending on the RESA-1, RESA-2 or RESA-3 proteins and the severity of the disease, two groups of severe and uncomplicated malaria cases being considered.

Methods: Two synthetic peptides representing predicted B cell epitopes were designed per RESA protein, all located outside of the 3' and 5' repetition blocks, in order to allow an antibody detection specific of each member of the family. Recombinant rRESA-1B and rRESA-3B proteins were also engineered. Two groups of Beninese children admitted to hospital in 2009 for either uncomplicated or severe malaria were compared for their plasma levels of IgG specifically recognizing each recombinant RESA protein or synthetic peptide, and for their plasma inflammatory cytokine levels (IFN-γ, TNF-α and IL-10), taking into account host and parasite genetic factors.

Results: The absence of IgG cross-reactivity between rRESA proteins and their protein carrier as well as between each RESA peptide and a non-epitopic RESA control peptide validated the use of the engineered recombinant proteins and peptides for the measurement of plasma IgG. Taking into account age, fever duration and parasitaemia, a multiple logistic regression performed on children clustered according to their antibody responses' profiles concluded to an increased risk of severe malaria for P2 (representative of RESA-1) responders (P = 0.007). Increased IL-10 plasma levels were found in children harbouring multiclonal P. falciparum infections on the basis of the T1526G resa2 gene polymorphism (P = 0.004).

Conclusions: This study provided novel tools to dissect the seroreactivity against the three members of the RESA protein family and to describe its relation to protection against malaria. It suggested the measurement of plasma antibodies raised against specific peptides to serve as predictive immunologic markers for disease severity. Lastly, it reinforced previous observations linking the T1526G resa2 gene mutation to severe malaria.

No MeSH data available.


Related in: MedlinePlus

Schematic map of the location of RESA peptides and recombinant proteins on RESA protein sequences. RESA-1 is encoded by the gene PFA0110w, RESA-2 by PF11_0512 and RESA-3 by PF11_0509. The schematic representation of the three RESA proteins (a) is detailed at the amino acid level for the regions containing peptides P1, P5 and P6 (b), P2 (c), P3 and P4 (d) as well as P Ctl (e). * internal stop codon
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Fig1: Schematic map of the location of RESA peptides and recombinant proteins on RESA protein sequences. RESA-1 is encoded by the gene PFA0110w, RESA-2 by PF11_0512 and RESA-3 by PF11_0509. The schematic representation of the three RESA proteins (a) is detailed at the amino acid level for the regions containing peptides P1, P5 and P6 (b), P2 (c), P3 and P4 (d) as well as P Ctl (e). * internal stop codon

Mentions: RESA family proteins share high homology sequences. Protein sequence alignments were performed with RESA-1 (accession number: PFA0110W/PF3D7_0102200), RESA-2 (accession number: PF11_0512/PF3D7_1149500) and RESA-3 (accession number: PF11_0509/PF3D7_1149200) using the Geneious software 6.1.4. from Biomatters Ltd, by means of the integrated multiple sequence comparison by log-expectation (Muscle) algorithm and the default settings. RESA-1 and -2 (see Additional file 1) shared 38% (333 amino-acids) homology, RESA-1 and -3, 49.1% (535 amino-acids) and RESA-2 and -3, 38.9% (361 amino-acids). Thus, cross-reactive antibodies should exist, preventing specific detection of the antigenic protein. To perform ELISA to evaluate the presence of RESA specific antibodies, six synthetic peptides that reproduced putative B cell epitopes of the RESA proteins were used. These peptides, whose sequences are shown in Table 1, were identified following B cell epitope predictions [43]. They were chosen to be located outside of the C-terminal and central repeats, which contain already known Pf155/RESA (RESA-1) epitopes [44], and to allow detection of protein specific antibodies. More precisely, they were chosen to be on a location where sequences are not conserved across the RESA proteins, leading to a given RESA protein-specific antibody detection. These peptides will be subsequently referred to as P1–P6, P1 and P2 being located on RESA-1, P3 and P4 on RESA-2, and P5 and P6 on RESA-3, as illustrated in Figure 1a. The peptides P1 (Figure 1b) and P2 (Figure 1c) shared 100% homology with RESA-1 and respectively, 11.1% (2 aa) and 26.7% (4 aa) homology with RESA-2, and 38.9% (7 aa) and 33.3% (5 aa) homology with RESA-3. The peptides P3 and P4 (Figure 1d), homologous to RESA-2, shared respectively 26.3% (5 aa) and 20% (3 aa) homology with both RESA-1 and -3. The peptides P5 (Figure 1b) homologous to RESA-3 shared no homology with RESA-1 or RESA-2 and P6 share 44.4% (4 aa) with RESA-1. A seventh peptide located on RESA-3 and not corresponding to a B cell epitope was assigned as control peptide (P Ctl) for antibody measurement. P Ctl (Figure 1e) was 60, 40 and 100% homologous to RESA-1 -2 and -3, respectively. The peptides were synthesized with an N-terminal biotin group. They were solubilized and used at a final concentration of 0.3 µM.Table 1


Immunoglobulin response to Plasmodium falciparum RESA proteins in uncomplicated and severe malaria.

Badaut C, Guyonnet L, Milet J, Renard E, Durand R, Viwami F, Sagbo G, Layla F, Deloron P, Bonnefoy S, Migot-Nabias F - Malar. J. (2015)

Schematic map of the location of RESA peptides and recombinant proteins on RESA protein sequences. RESA-1 is encoded by the gene PFA0110w, RESA-2 by PF11_0512 and RESA-3 by PF11_0509. The schematic representation of the three RESA proteins (a) is detailed at the amino acid level for the regions containing peptides P1, P5 and P6 (b), P2 (c), P3 and P4 (d) as well as P Ctl (e). * internal stop codon
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
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getmorefigures.php?uid=PMC4502540&req=5

Fig1: Schematic map of the location of RESA peptides and recombinant proteins on RESA protein sequences. RESA-1 is encoded by the gene PFA0110w, RESA-2 by PF11_0512 and RESA-3 by PF11_0509. The schematic representation of the three RESA proteins (a) is detailed at the amino acid level for the regions containing peptides P1, P5 and P6 (b), P2 (c), P3 and P4 (d) as well as P Ctl (e). * internal stop codon
Mentions: RESA family proteins share high homology sequences. Protein sequence alignments were performed with RESA-1 (accession number: PFA0110W/PF3D7_0102200), RESA-2 (accession number: PF11_0512/PF3D7_1149500) and RESA-3 (accession number: PF11_0509/PF3D7_1149200) using the Geneious software 6.1.4. from Biomatters Ltd, by means of the integrated multiple sequence comparison by log-expectation (Muscle) algorithm and the default settings. RESA-1 and -2 (see Additional file 1) shared 38% (333 amino-acids) homology, RESA-1 and -3, 49.1% (535 amino-acids) and RESA-2 and -3, 38.9% (361 amino-acids). Thus, cross-reactive antibodies should exist, preventing specific detection of the antigenic protein. To perform ELISA to evaluate the presence of RESA specific antibodies, six synthetic peptides that reproduced putative B cell epitopes of the RESA proteins were used. These peptides, whose sequences are shown in Table 1, were identified following B cell epitope predictions [43]. They were chosen to be located outside of the C-terminal and central repeats, which contain already known Pf155/RESA (RESA-1) epitopes [44], and to allow detection of protein specific antibodies. More precisely, they were chosen to be on a location where sequences are not conserved across the RESA proteins, leading to a given RESA protein-specific antibody detection. These peptides will be subsequently referred to as P1–P6, P1 and P2 being located on RESA-1, P3 and P4 on RESA-2, and P5 and P6 on RESA-3, as illustrated in Figure 1a. The peptides P1 (Figure 1b) and P2 (Figure 1c) shared 100% homology with RESA-1 and respectively, 11.1% (2 aa) and 26.7% (4 aa) homology with RESA-2, and 38.9% (7 aa) and 33.3% (5 aa) homology with RESA-3. The peptides P3 and P4 (Figure 1d), homologous to RESA-2, shared respectively 26.3% (5 aa) and 20% (3 aa) homology with both RESA-1 and -3. The peptides P5 (Figure 1b) homologous to RESA-3 shared no homology with RESA-1 or RESA-2 and P6 share 44.4% (4 aa) with RESA-1. A seventh peptide located on RESA-3 and not corresponding to a B cell epitope was assigned as control peptide (P Ctl) for antibody measurement. P Ctl (Figure 1e) was 60, 40 and 100% homologous to RESA-1 -2 and -3, respectively. The peptides were synthesized with an N-terminal biotin group. They were solubilized and used at a final concentration of 0.3 µM.Table 1

Bottom Line: Taking into account age, fever duration and parasitaemia, a multiple logistic regression performed on children clustered according to their antibody responses' profiles concluded to an increased risk of severe malaria for P2 (representative of RESA-1) responders (P = 0.007).Increased IL-10 plasma levels were found in children harbouring multiclonal P. falciparum infections on the basis of the T1526G resa2 gene polymorphism (P = 0.004).It suggested the measurement of plasma antibodies raised against specific peptides to serve as predictive immunologic markers for disease severity.

View Article: PubMed Central - PubMed

Affiliation: Equipe résidente de recherche en infectiologie tropicale, Institut de Recherche Biomédicale des Armées (IRBA), Brétigny sur Orge, France. cbadaut@gmail.com.

ABSTRACT

Background: The three members of the ring-infected erythrocyte surface antigen (RESA) proteins family share high sequence homologies, which impair the detection and assignment to one or another protein of some pathogenic processes inherent to Plasmodium falciparum malaria. The present study was intended to determine if the antibody and inflammatory responses of children living in a malaria-endemic area varied depending on the RESA-1, RESA-2 or RESA-3 proteins and the severity of the disease, two groups of severe and uncomplicated malaria cases being considered.

Methods: Two synthetic peptides representing predicted B cell epitopes were designed per RESA protein, all located outside of the 3' and 5' repetition blocks, in order to allow an antibody detection specific of each member of the family. Recombinant rRESA-1B and rRESA-3B proteins were also engineered. Two groups of Beninese children admitted to hospital in 2009 for either uncomplicated or severe malaria were compared for their plasma levels of IgG specifically recognizing each recombinant RESA protein or synthetic peptide, and for their plasma inflammatory cytokine levels (IFN-γ, TNF-α and IL-10), taking into account host and parasite genetic factors.

Results: The absence of IgG cross-reactivity between rRESA proteins and their protein carrier as well as between each RESA peptide and a non-epitopic RESA control peptide validated the use of the engineered recombinant proteins and peptides for the measurement of plasma IgG. Taking into account age, fever duration and parasitaemia, a multiple logistic regression performed on children clustered according to their antibody responses' profiles concluded to an increased risk of severe malaria for P2 (representative of RESA-1) responders (P = 0.007). Increased IL-10 plasma levels were found in children harbouring multiclonal P. falciparum infections on the basis of the T1526G resa2 gene polymorphism (P = 0.004).

Conclusions: This study provided novel tools to dissect the seroreactivity against the three members of the RESA protein family and to describe its relation to protection against malaria. It suggested the measurement of plasma antibodies raised against specific peptides to serve as predictive immunologic markers for disease severity. Lastly, it reinforced previous observations linking the T1526G resa2 gene mutation to severe malaria.

No MeSH data available.


Related in: MedlinePlus