VAR2CSA Signatures of High Plasmodium falciparum Parasitemia in the Placenta

Plasmodium falciparum infected erythrocytes (IE) accumulate in the placenta through the interaction between Duffy-binding like (DBL) domains of parasite-encoded ligand VAR2CSA and chondroitin sulphate-A (CSA) receptor. Polymorphisms in these domains, including DBL2X and DBL3X, may affect their antigenicity or CSA-binding affinity, eventually increasing parasitemia and its adverse effects on pregnancy outcomes. A total of 373 DBL2X and 328 DBL3X sequences were obtained from transcripts of 20 placental isolates infecting Mozambican women, resulting in 176 DBL2X and 191 DBL3X unique sequences at the protein level. Sequence alignments were divided in segments containing combinations of correlated polymorphisms and the association of segment sequences with placental parasite density was tested using Bonferroni corrected regression models, taking into consideration the weight of each sequence in the infection. Three DBL2X and three DBL3X segments contained signatures of high parasite density (P<0.003) that were highly prevalent in the parasite population (49–91%). Identified regions included a flexible loop that contributes to DBL3X-CSA interaction and two DBL3X motifs with evidence of positive natural selection. Limited antibody responses against signatures of high parasite density among malaria-exposed pregnant women could not explain the increased placental parasitemia. These results suggest that a higher binding efficiency to CSA rather than reduced antigenicity might provide a biological advantage to parasites with high parasite density signatures in VAR2CSA. Sequences contributing to high parasitemia may be critical for the functional characterization of VAR2CSA and the development of tools against placental malaria

VAR2CSA signatures of high Plasmodium falciparum parasitemia in the placenta.

Plasmodium falciparum infected erythrocytes (IE) accumulate in the placenta through the interaction between Duffy-binding like (DBL) domains of parasite-encoded ligand VAR2CSA and chondroitin sulphate-A (CSA) receptor. Polymorphisms in these domains, including DBL2X and DBL3X, may affect their antigenicity or CSA-binding affinity, eventually increasing parasitemia and its adverse effects on pregnancy outcomes. A total of 373 DBL2X and 328 DBL3X sequences were obtained from transcripts of 20 placental isolates infecting Mozambican women, resulting in 176 DBL2X and 191 DBL3X unique sequences at the protein level. Sequence alignments were divided in segments containing combinations of correlated polymorphisms and the association of segment sequences with placental parasite density was tested using Bonferroni corrected regression models, taking into consideration the weight of each sequence in the infection. Three DBL2X and three DBL3X segments contained signatures of high parasite density (P<0.003) that were highly prevalent in the parasite population (49-91%). Identified regions included a flexible loop that contributes to DBL3X-CSA interaction and two DBL3X motifs with evidence of positive natural selection. Limited antibody responses against signatures of high parasite density among malaria-exposed pregnant women could not explain the increased placental parasitemia. These results suggest that a higher binding efficiency to CSA rather than reduced antigenicity might provide a biological advantage to parasites with high parasite density signatures in VAR2CSA. Sequences contributing to high parasitemia may be critical for the functional characterization of VAR2CSA and the development of tools against placental malaria

Association of severe malaria outcomes with platelet-mediated clumping and adhesion to a novel host receptor.

INTRODUCTION: Severe malaria has been attributed partly to the sequestration of Plasmodium falciparum-infected erythrocytes (IEs) in the microvasculature of vital host organs. Identification of P. falciparum cytoadherence phenotypes that are associated with severe malaria may lead to the development of novel strategies against life-threatening malaria. METHODS AND FINDINGS: Forty-six P. falciparum isolates from Mozambican children under 5 years of age with severe malaria (cases) were examined and compared to 46 isolates from sex and age matched Mozambican children with uncomplicated malaria (controls). Cytoadherence properties such as platelet-mediated clumping, rosetting and adhesion to purified receptors (CD36, ICAM1 and gC1qR), were compared between these matched pairs by non-parametric tests. The most common clinical presentation associated with severe malaria was prostration. Compared to matched controls, prevalence of platelet-mediated clumping was higher in cases (P = .019), in children presenting with prostration (P = .049) and in children with severe anaemia (P = .025). Prevalence of rosetting and gC1qR adhesion were also higher in isolates from cases with severe anemia and multiple seizures, respectively (P = .045 in both cases), than in controls. CONCLUSIONS: These data indicate a role for platelet-mediated clumping, rosetting and adhesion to gC1qR in the pathogenesis of severe malaria. Inhibition of these cytoadherence phenotypes may reduce the occurrence or improve the prognosis of severe malaria outcomes

Functional and Immunological Characterization of a Duffy Binding-Like Alpha Domain from Plasmodium falciparum Erythrocyte Membrane Protein 1 That Mediates Rosetting ▿

The Duffy binding-like (DBL) domains are common adhesion modules present in Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1) variants, which are responsible for immune evasion and cytoadherence. Knowledge about how immune responses are acquired against polymorphic DBL domains of PfEMP1 can aid in the development of vaccines for malaria. A recombinant DBLα domain, encoded by R29 var1, which binds complement receptor 1 to mediate rosetting by the P. falciparum laboratory strain R29, was expressed in Escherichia coli, renatured by oxidative refolding to its native form, and purified to homogeneity. Antibody levels in 704 plasmas obtained from residents of areas of different levels of malaria endemicity in Orissa (India) and Manhiça (Mozambique) were assessed by enzyme-linked immunosorbent assay. The refolded DBLα domain was pure, homogeneous, and functional in that it bound human erythrocytes with specificity and was capable of inhibiting rosetting. The proportion of individuals who had measurable anti-DBLα immunoglobulin G responses was low in areas of low malaria endemicity in Orissa (6.7%) but high in areas of high endemicity in Orissa (87.5%) and Manhiça (74.5%). Seroprevalence and antibody levels against the recombinant protein increased with the age of inhabitants from areas with high transmission rates (P < 0.001). Half of the children in these areas had seroconverted by the age of 5 years. These findings suggest that in spite of the extreme polymorphism of PfEMP1 DBLα domains, the acquisition of specific antibodies is rapid and age related and reflects the reduced risk of malaria in areas with high transmission rates. Further studies are required to elucidate the role of these antibodies in protection from malaria

VAR2CSA signatures of high Plasmodium falciparum parasitemia in the placenta.

Plasmodium falciparum infected erythrocytes (IE) accumulate in the placenta through the interaction between Duffy-binding like (DBL) domains of parasite-encoded ligand VAR2CSA and chondroitin sulphate-A (CSA) receptor. Polymorphisms in these domains, including DBL2X and DBL3X, may affect their antigenicity or CSA-binding affinity, eventually increasing parasitemia and its adverse effects on pregnancy outcomes. A total of 373 DBL2X and 328 DBL3X sequences were obtained from transcripts of 20 placental isolates infecting Mozambican women, resulting in 176 DBL2X and 191 DBL3X unique sequences at the protein level. Sequence alignments were divided in segments containing combinations of correlated polymorphisms and the association of segment sequences with placental parasite density was tested using Bonferroni corrected regression models, taking into consideration the weight of each sequence in the infection. Three DBL2X and three DBL3X segments contained signatures of high parasite density (P<0.003) that were highly prevalent in the parasite population (49-91%). Identified regions included a flexible loop that contributes to DBL3X-CSA interaction and two DBL3X motifs with evidence of positive natural selection. Limited antibody responses against signatures of high parasite density among malaria-exposed pregnant women could not explain the increased placental parasitemia. These results suggest that a higher binding efficiency to CSA rather than reduced antigenicity might provide a biological advantage to parasites with high parasite density signatures in VAR2CSA. Sequences contributing to high parasitemia may be critical for the functional characterization of VAR2CSA and the development of tools against placental malaria

Parity and Placental Infection Affect Antibody Responses against Plasmodium falciparum during Pregnancy▿ †

Women are at higher risk of Plasmodium falciparum infection when pregnant. The decreasing risk of malaria with subsequent pregnancies is attributed to parity-dependent acquisition of antibodies against placental parasites expressing variant surface antigens, VAR2CSA, that mediate placental sequestration through adhesion to chondroitin sulfate A (CSA). However, modulation of immunity during pregnancy may also contribute to increase the risk of malaria. We compared antibody responses among 30 Mozambican primigravidae and 60 multigravidae at delivery, 40 men, and 40 children. IgG levels were measured against the surface antigens of erythrocytes infected with P. falciparum isolated from 12 pregnant women (4 placental and 8 peripheral blood isolates) and 26 nonpregnant hosts. We also measured IgG levels against merozoite recombinant antigens and total IgG. Placental P. falciparum infection was associated with increased levels of total IgG as well as IgG levels against merozoite antigens and parasite isolates from pregnant and nonpregnant hosts. We therefore stratified comparisons of antibody levels by placental infection. Compared to multigravidae, uninfected primigravidae had lower total IgG as well as lower levels of IgGs against peripheral blood isolates from both pregnant and nonpregnant hosts. These differences were not explained by use of bed nets, season at delivery, neighborhood of residence, or age. Compared to men, infected primigravidae had higher levels of IgGs against isolates from pregnant women and CSA-binding lines but not against other isolates, supporting the concept of a pregnancy-specific development of immunity to these parasite variants. Results of this study show that parity and placental infection can modulate immune responses during pregnancy against malaria parasites

Complement Receptor 1 availability on red blood cell surface modulates Plasmodium vivax invasion of human reticulocytes

Plasmodium vivax parasites preferentially invade reticulocyte cells in a multistep process that is still poorly understood. In this study, we used ex vivo invasion assays and population genetic analyses to investigate the involvement of complement receptor 1 (CR1) in P. vivax invasion. First, we observed that P. vivax invasion of reticulocytes was consistently reduced when CR1 surface expression was reduced through enzymatic cleavage, in the presence of naturally low-CR1-expressing cells compared with high-CR1-expressing cells, and with the addition of soluble CR1, a known inhibitor of P. falciparum invasion. Immuno-precipitation experiments with P. vivax Reticulocyte Binding Proteins showed no evidence of complex formation. In addition, analysis of CR1 genetic data for worldwide human populations with different exposure to malaria parasites show significantly higher frequency of CR1 alleles associated with low receptor expression on the surface of RBCs and higher linkage disequilibrium in human populations exposed to P. vivax malaria compared with unexposed populations. These results are consistent with a positive selection of low-CR1-expressing alleles in vivax-endemic areas. Collectively, our findings demonstrate that CR1 availability on the surface of RBCs modulates P. vivax invasion. The identification of new molecular interactions is crucial to guiding the rational development of new therapeutic interventions against vivax malaria