Targets and Patterns of Erythrocyte Invasion Inhibitory Antibodies in Malaria

Abstract

Background As malaria transmission continues to decline, a better understanding of how transmission intensity and patterns influence clinical presentation of disease and the acquisition of immunity to malaria is important for our understanding of the pathophysiology of malaria and the changes in immune responses. Erythrocyte invasion is a crucial step in the life cycle of Plasmodium falciparum. However, the targets of naturally acquired invasion inhibitory antibodies remain unclear. In this study, the differences in clinical and haematological presentation of disease, the predictors of anaemia severity in children with malaria, living in three ecologically distinct areas of Ghana, with different transmission intensities was determined. Patterns of invasion inhibitory antibodies were also assessed to explore the impact of transmission intensity on these parameters. Additionally, the relationship between antibody levels and functionality of antibodies to key invasion ligands were evaluated in adults living in a holoendemic area of Ghana. Methods Blood samples were taken from children between the ages of 2 and 14 years with confirmed malaria in hospitals in three areas with different transmission intensities (Kintampo>Navrongo>Accra). Whole blood was used to perform comprehensive analysis of parasitological, clinical and haematological variables, whilst socio-economic details were collected using a questionnaire. Plasma samples separated from the whole blood collected were tested by enzyme linked immunosorbent assays for antibodies to P. falciparum invasion antigens, including erythrocyte binding antigens (EBA) 175, EBA140, EBA181, and reticulocyte binding-like homologue (Rh) 2, Rh4 and Rh5. To identify the targets of invasion inhibitory antibodies, plasma samples were collected from 50 male adults living in a high transmission area with no recent history of clinical malaria. Antibodies against EBA and Rh proteins were detected using enzyme linked immunosorbent assays (ELISA). Immunoglobulin (IgG) fractions were purified from the plasma and used in erythrocyte invasion assays. Results Severity of malarial anaemia was more pronounced in children living in areas of high malaria transmission compared to children living in low endemic areas. Sickle cell trait was protective against anaemia severity in Kintampo (P=0.016), although this association was not statistically significant in Accra (P=0.379) and) and Navrongo (P=0.529). Parasitaemia was not a significant predictor of haemoglobin level, after controlling for age and gender. In addition, antibodies against invasion antigens were negatively correlated with parasitaemia, and increased in an age-dependent manner. Regression analysis showed that breadth of antibody reactivity was exposure and age-dependent. Levels of antibodies to EBA and RH antigens generally correlated with invasion inhibitory activity and breadth of antibody reactivity was predictive of inhibitory activity of purified IgG (ρ= 0.437, P= 0.009). Conclusions Our findings demonstrate significant differences in the haematological presentation and severity of malaria among children residing in areas with different transmission intensities. Patterns of antibody responses against invasion proteins are both antigen and exposure dependent. Growth inhibitory activity was significantly associated with antibody levels and the number of different antigens recognized.