Antibody responses to plasmodium falciparum 3d7 rh5 and eba-175 antigens in an acute and convalescent cohort of children from Junju sub-location in Kilifi, Kenya.

Abstract

Repeated exposure to malaria parasites broadens the repertoire of antigens recognized by the host immune system leading to acquisition of immunity to the disease. This immunity is inefficient in the absence of exposure as it wanes with time. Antibodies mediate humoral immunity against malaria and are majorly directed towards the extracellular merozoite. The reticulocyte binding-like protein homologue 5 (Rh5) and erythrocyte binding antigen (EBA-175) are merozoite antigens that are immunogenic and they are potential malaria vaccine candidates. The sequence conservation of Rh5 across Plasmodium falciparum (P. falciparum) strains makes it a desirable vaccine candidate that elicits strain-transcending immune responses. Rh5 also maintains the malaria parasite’s viability as shown by futile attempts to delete the gene encoding the antigen. A recent study has shown that EBA-175 shed post merozoite invasion mediates clustering of uninfected red blood cells (RBCs) around infected RBCs promoting parasite growth as progeny merozoites can easily invade other uninfected RBCs. In naturally malaria-exposed populations, Rh5 and EBA-175 may be targets of protective immunity. However, there is limited knowledge on how the magnitude of the antigen-specific antibody titres changes following an acute malaria episode. Thus, the objective of the study was to determine whether the magnitude of antibody responses to Rh5 and EBA-175 from the P. falciparum 3D7 strain changes over time following an acute malaria episode. The two recombinant proteins were expressed by mammalian Expi293F cells. Antibody responses to the expressed antigens were measured by enzyme-linked immunosorbent assay (ELISA) using plasma samples collected during a cross-sectional bleed (baseline), acute malaria infection, 4, 6 and 24 weeks following treatment. The differences in antibody titres between all the time-points were determined using the non-parametric Kruskal-Wallis test while the differences between any two time-points were assessed using the non-parametric Wilcoxon signed-rank test. A non-linear mixed effects model was used to model antibody responses as a function of time. The two antigens were immunogenic (OD above 0.05) as they elicited antibody responses in the plasma samples tested. There was also a marked heterogeneity in antibody responses to both antigens in the children aged between 0-15 years from Junju, Kilifi. The data also showed that infection with P. falciparum boosted antibody responses to Rh5 (p = 0.044). For EBA-175, acute malaria infection did not significantly boost antibody responses to the antigen. However, children older than 5 years of age elicited greater antibody responses to EBA-175 (Estimate = 0.14, SE = 0.05, p = 0.01), suggesting an acquisition of malarial immunity as the children grew older. Generally, antibody responses to both antigens declined during the follow-up period of 24 weeks (Estimate = -0.04, SE= 0.01, p = <0.01) for Rh5 and (Estimate = -0.07, SE = 0.02, p = <0.001) for EBA-175, showing the short-lived nature of immune responses to the two antigens. In conclusion, the two antigens were immunogenic. Antibody responses to the two antigens were heterogeneous and waned after the acute malaria infection. While infection with P. falciparum boosted antibody responses to Rh5, there was an effect of age on antibody responses to EBA-175. These results have the implications for the development of a more efficacious malaria vaccine based on RH5 and EBA-175, and also inform the timing of serological studies. These two antigens are important blood stage antigens that could be included in a multi-component vaccine for a better vaccine efficacy.