Antibody Reactivity to Merozoite Antigens in Ghanaian Adults Correlates With Growth Inhibitory Activity Against Plasmodium falciparum in Culture.

BACKGROUND: Plasmodium falciparum uses a repertoire of merozoite-stage proteins for invasion of erythrocytes. Antibodies against some of these proteins halt the replication cycle of the parasite by preventing erythrocyte invasion and are implicated as contributors to protective immunity against malaria. METHODS: We assayed antibody reactivity against a panel of 9 recombinant antigens based on erythrocyte-binding antigen (EBA) and reticulocyte-like homolog (Rh) proteins in plasma from children with malaria and healthy adults residing in 3 endemic areas in Ghana using enzyme-linked immunosorbent assay. Purified immunoglobulin (Ig)G from adult plasma samples was also tested for invasion inhibition against 7 different P falciparum culture lines, including clinical isolates. RESULTS: Antibodies against the antigens increased in an age-dependent manner in children. Breadth of reactivity to the different antigens was strongly associated with in vitro parasite growth inhibitory activity of IgG purified from the adults. The strongest predictors of breadth of antibody reactivity were age and transmission intensity, and a combination of reactivities to Rh2, Rh4, and Rh5 correlated strongly with invasion inhibition. CONCLUSIONS: Growth inhibitory activity was significantly associated with breadth of antibody reactivity to merozoite antigens, encouraging the prospect of a multicomponent blood-stage vaccine.

Assessing the impact of differences in malaria transmission intensity on clinical and haematological indices in children with malaria.

BACKGROUND: Malaria control interventions have led to a decline in transmission intensity in many endemic areas, and resulted in elimination in some areas. This decline, however, will lead to delayed acquisition of protective immunity and thus impact disease manifestation and outcomes. Therefore, the variation in clinical and haematological parameters in children with malaria was assessed across three areas in Ghana with varying transmission intensities. METHODS: A total of 568 children between the ages of 2 and 14 years with confirmed malaria were recruited in hospitals in three areas with varying transmission intensities (Kintampo > Navrongo > Accra) and a comprehensive analysis of parasitological, clinical, haematological and socio-economic parameters was performed. RESULTS: Areas of lower malaria transmission tended to have lower disease severity in children with malaria, characterized by lower parasitaemias and higher haemoglobin levels. In addition, total white cell counts and percent lymphocytes decreased with decreasing transmission intensity. The heterozygous sickle haemoglobin genotype was protective against disease severity in Kintampo (P = 0.016), although this was not significant in Accra and Navrongo. Parasitaemia levels were not a significant predictor of haemoglobin level after controlling for age and gender. However, higher haemoglobin levels in children were associated with certain socioeconomic factors, such as having fathers who had any type of employment (P < 0.05) and mothers who were teachers (P < 0.05). CONCLUSIONS: The findings demonstrate significant differences in the haematological presentation and severity of malaria among areas with different transmission intensity in Ghana, indicating that these factors need to be considered in planning the management of the disease as the endemicity is expected to decline after control interventions.

Variation in Plasmodium falciparum erythrocyte invasion phenotypes and merozoite ligand gene expression across different populations in areas of malaria endemicity.

Plasmodium falciparum merozoites use diverse alternative erythrocyte receptors for invasion and variably express cognate ligands encoded by the erythrocyte binding antigen (eba) and reticulocyte binding-like homologue (Rh) gene families. Previous analyses conducted on parasites from single populations in areas of endemicity revealed a wide spectrum of invasion phenotypes and expression profiles, although comparisons across studies have been limited by the use of different protocols. For direct comparisons within and among populations, clinical isolates from three different West African sites of endemicity (in Ghana, Guinea, and Senegal) were cryopreserved and cultured ex vivo after thawing in a single laboratory to assay invasion of target erythrocytes pretreated with enzymes affecting receptor subsets. Complete invasion assay data from 67 isolates showed no differences among the populations in the broad range of phenotypes measured by neuraminidase treatment (overall mean, 40.6% inhibition) or trypsin treatment (overall mean, 83.3% inhibition). The effects of chymotrypsin treatment (overall mean, 79.2% inhibition) showed heterogeneity across populations (Kruskall-Wallis P = 0.023), although the full phenotypic range was seen in each. Schizont-stage transcript data for a panel of 8 invasion ligand genes (eba175, eba140, eba181, Rh1, Rh2a, Rh2b, Rh4, and Rh5) were obtained for 37 isolates, showing similar ranges of variation in each population except that eba175 levels tended to be higher in parasites from Ghana than in those from Senegal (whereas levels of eba181 and Rh2b were lower in parasites from Ghana). The broad diversity in invasion phenotypes and gene expression seen within each local population, with minimal differences among them, is consistent with a hypothesis of immune selection maintaining parasite variation.

Analysis of Erythrocyte Invasion Mechanisms of Plasmodium falciparum Clinical Isolates Across 3 Malaria-Endemic Areas in Ghana.

BACKGROUND: Plasmodium falciparum invades human erythrocytes by using an array of ligands that interact with several receptors, including sialic acid (SA), complement receptor 1 (CR1), and basigin. We hypothesized that in malaria-endemic areas, parasites vary invasion pathways under immune pressure. Therefore, invasion mechanisms of clinical isolates collected from 3 zones of Ghana with different levels of endemicity (from lowest to highest, Accra, Navrongo, and Kintampo) were compared using standardized methods. METHODS: Blood samples were collected from children aged 2-14 years in whom malaria was diagnosed, and erythrocyte invasion phenotypes were determined using the enzymes neuraminidase, chymotrypsin, and trypsin, which differentially cleave receptors from the erythrocyte surface. In addition, antibodies against CR1 and basigin were used to determine the contributions of these receptors to invasion. Gene expression levels of P. falciparum invasion ligands were also examined. RESULTS: The parasites generally expressed SA-independent invasion phenotypes across the malaria-endemic areas, with parasites from Kintampo showing the highest invasion rates in neuraminidase-treated erythrocytes. CR1 was a major mediator of SA-independent invasion, while basigin was essential for both SA-dependent and SA-independent invasion mechanisms. Furthermore, expression of the basigin ligand PfRh5 was the best predictor of donor parasitemia. CONCLUSIONS: Erythrocyte invasion phenotypes expressed by P. falciparum are influenced by endemicity levels, and the PfRh5-basigin pathway is a potential vaccine target.