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1.
Mem Inst Oswaldo Cruz ; 102(3): 335-9, 2007 Jun.
Article in English | MEDLINE | ID: mdl-17568939

ABSTRACT

In previous immuno-epidemiological studies of the naturally acquired antibody responses to merozoite surface protein-1 (MSP-1) of Plasmodium vivax, we had evidence that the responses to distinct erythrocytic stage antigens could be differentially regulated. The present study was designed to compare the antibody response to three asexual erythrocytic stage antigens vaccine candidates of P. vivax. Recombinant proteins representing the 19 kDa C-terminal region of MSP-1(PvMSP19), apical membrane antigen n-1 ectodomain (PvAMA-1), and the region II of duffy binding protein (PvDBP-RII) were compared in their ability to bind to IgG antibodies of serum samples collected from 220 individuals from the state of Pará, in the North of Brazil. During patent infection with P. vivax, the frequency of individuals with IgG antibodies to PvMSP1(19), PvAMA-1, and PvDBP-RII were 95, 72.7, and 44.5% respectively. Although the frequency of responders to PvDBP-RII was lower, this frequency increased in individuals following multiple malarial infections. Individually, the specific antibody levels did not decline significantly nine months after treatment, except to PvMSP1(19). Our results further confirm a complex regulation of the immune response to distinct blood stage antigens. The reason for that is presently unknown but it may contribute to the high risk of re-infection in individuals living in the endemic areas.


Subject(s)
Antibodies, Protozoan/immunology , Immunoglobulin G/immunology , Malaria Vaccines/immunology , Malaria, Vivax/immunology , Merozoite Surface Protein 1/immunology , Plasmodium vivax/immunology , Animals , Enzyme-Linked Immunosorbent Assay , Humans , Merozoite Surface Protein 1/chemistry , Recombinant Proteins/immunology
2.
Mem. Inst. Oswaldo Cruz ; 102(3): 335-340, June 2007. tab, graf
Article in English | LILACS | ID: lil-452511

ABSTRACT

In previous immuno-epidemiological studies of the naturally acquired antibody responses to merozoite surface protein-1 (MSP-1) of Plasmodium vivax, we had evidence that the responses to distinct erythrocytic stage antigens could be differentially regulated. The present study was designed to compare the antibody response to three asexual erythrocytic stage antigens vaccine candidates of P. vivax. Recombinant proteins representing the 19 kDa C-terminal region of MSP-1(PvMSP19), apical membrane antigen n-1 ectodomain (PvAMA-1), and the region II of duffy binding protein (PvDBP-RII) were compared in their ability to bind to IgG antibodies of serum samples collected from 220 individuals from the state of Pará, in the North of Brazil. During patent infection with P. vivax, the frequency of individuals with IgG antibodies to PvMSP1(19), PvAMA-1, and PvDBP-RII were 95, 72.7, and 44.5 percent respectively. Although the frequency of responders to PvDBP-RII was lower, this frequency increased in individuals following multiple malarial infections. Individually, the specific antibody levels did not decline significantly nine months after treatment, except to PvMSP1(19). Our results further confirm a complex regulation of the immune response to distinct blood stage antigens. The reason for that is presently unknown but it may contribute to the high risk of re-infection in individuals living in the endemic areas.


Subject(s)
Humans , Animals , Antibodies, Protozoan/immunology , Immunoglobulin G/immunology , Malaria, Vivax/immunology , Merozoite Surface Protein 1/immunology , Plasmodium vivax/immunology , Enzyme-Linked Immunosorbent Assay , Merozoite Surface Protein 1/chemistry , Recombinant Proteins/immunology
3.
Am J Trop Med Hyg ; 73(5 Suppl): 25-31, 2005 Nov.
Article in English | MEDLINE | ID: mdl-16291763

ABSTRACT

Invasion of human erythrocytes by Plasmodium vivax requires interaction between Duffy binding protein (PvDBP) and the Duffy blood group antigen. The receptor-binding domain of PvDBP lies in a conserved N-terminal, cysteine-rich region, region II (PvRII). PvRII is a valuable malaria subunit vaccine candidate for asexual blood stages. We have evaluated in Aotus monkeys the immunogenicity and protective efficacy of recombinant PvRII formulated in Freund's and Montanide ISA720 adjuvants. Specific antibody titers were determined by an enzyme-linked immunosorbent assay after each of three doses of 50 microg of protein administered by the subcutaneous route. Immunization with PvRII formulated in Freund's adjuvant yielded higher antibody titers than immunization with the Montanide ISA720 formulation and offered partial protection. Although the Montanide ISA720 formulation was immunogenic, it did not provide any protection. Given the immunogenicity and partial protection observed, further studies are needed to optimize the PvRII vaccine formulation with adjuvants suitable for human use.


Subject(s)
Antigens, Protozoan/chemistry , Antigens, Protozoan/immunology , Cebidae , Disease Models, Animal , Duffy Blood-Group System/metabolism , Malaria Vaccines/immunology , Malaria, Vivax/prevention & control , Protozoan Proteins/chemistry , Protozoan Proteins/immunology , Receptors, Cell Surface/chemistry , Receptors, Cell Surface/immunology , Vaccines, Synthetic/immunology , Adjuvants, Immunologic/administration & dosage , Animals , Antibodies, Protozoan/blood , Antigens, Protozoan/genetics , Female , Freund's Adjuvant/administration & dosage , Humans , Immunization , Malaria Vaccines/administration & dosage , Malaria, Vivax/parasitology , Male , Mannitol/administration & dosage , Mannitol/analogs & derivatives , Oleic Acids/administration & dosage , Plasmodium vivax/immunology , Plasmodium vivax/pathogenicity , Protozoan Proteins/genetics , Receptors, Cell Surface/genetics , Vaccines, Synthetic/administration & dosage
4.
Am J Trop Med Hyg ; 73(5 Suppl): 44-9, 2005 Nov.
Article in English | MEDLINE | ID: mdl-16291766

ABSTRACT

The Duffy antigen (Fy) is necessary for Plasmodium vivax invasion of human erythrocytes. Some populations have a highly prevalent Fy-negative phenotype; such persons are naturally protected from P. vivax blood infection but are expected to completely support the P. vivax pre-erythrocytic cycle, representing a valuable model for studying the immune response during these parasitic stages. We typed 214 individuals, mostly Afro-Colombians, from a P. vivax-endemic area for Fy expression and determined the antibody response to P. vivax pre-erythrocytic (sporozoites and CS) and blood-stage antigens (blood forms, P. vivax merozoite surface protein 1, and P. vivax Duffy binding protein [PvDBP]). Antibody titers to P. vivax circumsporozoite protein, P11, and N-terminal peptides and the number of responders were similar in Fy-negative and Fy-positive individuals. The number of responders to sporozoites, blood forms, and PvDBP were different between these groups. Thus, Fy-negative individuals from malaria-endemic areas can be used to study the immune response to the P. vivax liver phase without interference of the erythrocytic cycle.


Subject(s)
Antibodies, Protozoan/blood , Antigens, Protozoan/immunology , Duffy Blood-Group System/metabolism , Malaria, Vivax/epidemiology , Plasmodium vivax/immunology , Receptors, Cell Surface/metabolism , Adolescent , Adult , Aged , Aged, 80 and over , Animals , Child , Colombia/epidemiology , Erythrocytes/parasitology , Female , Humans , Malaria, Vivax/immunology , Male , Middle Aged , Plasmodium vivax/growth & development , Prevalence , Rural Population , Sporozoites/immunology
5.
Am J Trop Med Hyg ; 73(2): 244-55, 2005 Aug.
Article in English | MEDLINE | ID: mdl-16103583

ABSTRACT

Naturally acquired antibody reactivity to two major Plasmodium vivax vaccine candidates was investigated in 294 donors from three malaria-endemic communities of Rondônia state, Brazil. Antibody recognition of recombinantly expressed antigens covering five different regions of P. vivax reticulocyte binding protein 1 (PvRBP1) and region II of P. vivax Duffy binding protein (PvDBP-RII) were compared. Positive IgG responses to these antigens were significantly related to the level of malaria exposure in terms of past infections and years of residence in the endemic area when corrected for age. The highest prevalence of anti-PvRBP1 total IgG antibodies corresponded to the amino acid regions denoted PvRBP1(431-748) (41%) and PvRBP1(733-1407) (47%). Approximately one-fifth of positively responding sera had titers of at least 1:1,600. Total IgG responses to PvDBP-RII were more prevalent (67%), of greater magnitude, and acquired more rapidly than those to individual PvRBP1 antigens. Responses to both PvRBP1 and PvDBP-RII were biased toward the cytophilic subclasses IgG1 and IgG3. These data provide the first insights on acquired antibody responses to PvRBP1 and a comparative view with PvDBP-RII that may prove valuable for understanding protective immune responses to these two vaccine candidates as they are evaluated as components of multitarget blood-stage vaccines.


Subject(s)
Antigens, Protozoan/immunology , Immunoglobulin G/blood , Malaria, Vivax/epidemiology , Membrane Proteins/immunology , Plasmodium vivax/immunology , Protozoan Proteins/immunology , Adolescent , Adult , Aged , Aged, 80 and over , Animals , Antibodies, Protozoan/blood , Antigens, Protozoan/genetics , Brazil/epidemiology , Child , Duffy Blood-Group System/metabolism , Female , Humans , Immunoglobulin G/classification , Malaria Vaccines , Malaria, Vivax/immunology , Male , Membrane Proteins/genetics , Middle Aged , Plasmodium vivax/growth & development , Protozoan Proteins/genetics , Recombinant Proteins/immunology
6.
Cytometry A ; 63(1): 59-66, 2005.
Article in English | MEDLINE | ID: mdl-15584018

ABSTRACT

BACKGROUND: The malaria parasite Plasmodium vivax preferentially invades reticulocytes. It is therefore relevant for vaccine development purposes to identify and characterize P. vivax proteins that bind specifically to the surface of reticulocytes. We have developed a two-color flow cytometric erythrocyte binding assay (F-EBA) that has several advantages over traditional erythrocyte binding assays (T-EBAs) used in malaria research. We demonstrate the use of F-EBA using the P. vivax Duffy binding protein region II (PvDBP-RII) recombinant protein as a model. This protein binds to all erythrocytes that express the Duffy receptor (Fy) and discriminates binding between normocytes and reticulocytes. METHODS: F-EBAs were performed by incubating freshly isolated Aotus nancymai, Macaca mulatta, Saimiri boliviensis, and human erythrocytes with PvDBP-RII, a fluorescent anti-His tag detection antibody, and thiazole orange before flow cytometric analysis. T-EBAs employing immunoblot detection with an anti-His antibody were performed concomitantly. RESULTS: PvDBP-RII bound to A. nancymai, M. mulatta, and human Fy+ erythrocytes, but not human Fy- erythrocytes, by F-EBAs and T-EBAs. However, F-EBAs exhibited higher sensitivity and better concordance between experiments compared with T-EBAs. CONCLUSIONS: F-EBA is a rapid, simple, and reliable method for quantifying the ability of malaria proteins to bind to the surface of erythrocytes. F-EBA can discriminate binding between erythrocyte subpopulations without enrichment protocols and may be more reliable and sensitive than T-EBAs in identifying novel erythrocyte binding proteins.


Subject(s)
Antigens, Protozoan/metabolism , Duffy Blood-Group System/metabolism , Erythrocytes/metabolism , Plasmodium vivax/metabolism , Protozoan Proteins/metabolism , Receptors, Cell Surface/metabolism , Animals , Aotidae , Erythrocytes/cytology , Flow Cytometry , Humans , In Vitro Techniques , Macaca mulatta , Recombinant Proteins/metabolism , Saimiri
7.
Biochem J ; 374(Pt 1): 193-8, 2003 Aug 15.
Article in English | MEDLINE | ID: mdl-12775212

ABSTRACT

Plasmodium vivax and P. knowlesi use the Duffy antigen as a receptor to invade human erythrocytes. Duffy-binding ligands belong to a family of erythrocyte-binding proteins that bind erythrocyte receptors to mediate invasion. Receptor-binding domains in erythrocyte-binding proteins lie in conserved cysteine-rich regions called Duffy-binding-like domains. In the present study, we report an analysis of the overall three-dimensional architecture of P. vivax and P. knowlesi Duffy-binding domains based on mild proteolysis and supportive-functional assays. Our proteolysis experiments indicate that these domains are built of two distinct subdomains. The N-terminal region from Cys-1-4 (C1-C4) forms a stable non-functional subdomain. The region spanning C5-C12 forms another subdomain, which is capable of binding Duffy antigen. These subdomains are joined by a protease-sensitive linker. Results from deletion constructs, designed for expression of truncated proteins on COS cell surface, show that regions containing C5-C8 of the Duffy-binding domains are sufficient for the binding receptor. Therefore the central region of Duffy-binding domains, which is flanked by two non-functional regions, is responsible for receptor recognition. Moreover, the minimal Duffy-binding region identified here is capable of folding into a functionally competent module. These studies pave the way for understanding the architecture of Duffy-binding domains and their interactions with host receptors.


Subject(s)
Antigens, Protozoan/physiology , Erythrocytes/parasitology , Plasmodium knowlesi/pathogenicity , Plasmodium vivax/pathogenicity , Protozoan Proteins/physiology , Receptors, Cell Surface/physiology , Animals , Antigens, Protozoan/chemistry , Antigens, Protozoan/genetics , Base Sequence , Binding Sites , Cell Adhesion Molecules/chemistry , Cell Adhesion Molecules/physiology , Cloning, Molecular , DNA Primers , Escherichia coli/genetics , Humans , Molecular Sequence Data , Mutagenesis , Peptide Fragments/chemistry , Polymerase Chain Reaction , Protozoan Proteins/chemistry , Protozoan Proteins/genetics , Receptors, Cell Surface/chemistry , Receptors, Cell Surface/genetics , Recombinant Proteins/chemistry , Recombinant Proteins/metabolism , Sequence Deletion , Trypsin
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