Antibody response to a new member of the DBL family (EBP2) after a brief Plasmodium vivax exposure

Abstract

Plasmodium vivax blood-stage invasion into reticulocyte is critical for parasite development. Thus, validation of novel parasite invasion ligands is essential for malaria vaccine development. Recently, we demonstrated that EBP2, a Duffy binding protein (DBP) paralog, is antigenically distinct from DBP and could not be functionally inhibited by anti-DBP antibodies. Here, we took advantage of a small outbreak of P.vivax malaria, located in a non-malarious area of Brazil, to investigate for the first time IgM/IgG antibodies against EBP2 and DEKnull-2 (an engineering DBPII vaccine) among individuals who had their first and brief exposure to P.vivax (16 cases and 22 non-cases). Our experimental approach included 4 cross sectional surveys at 3-month interval (12-month follow-up). The results demonstrated that while a brief initial P.vivax infection was not efficient to induce IgM/ IgG antibodies to either EBP2 or DEKnull-2, IgG antibodies against DEKnull-2 (but not EBP2) were boosted by recurrent blood-stage infections following treatment. Of interest, in most recurrent P. vivax infections (4 out of 6 patients) DEKnull-2 IgG antibodies were sustained for 6 to 12 months. Polymorphisms in the ebp2 gene does not seem to explain EBP2 low immunogenicity as the ebp2 allele associated with the P.vivax outbreak presented high identity to the original EBP2 isolate used as recombinant protein. Although EBP2 antibodies were barely detectable after a primary episode of P.vivax infection, EBP2 was highly recognized by serum IgG from long-term malaria-exposed Amazonians (range from 35 to 92% according to previous malaria episodes). Taken together, the results showed that individuals with a single and brief exposure to P.vivax infection develop very low anti-EBP2 antibodies, which tend to increase after long-term malaria exposure. Finally, the findings highlighted the potential of DEKnull-2 as a vaccine candidate, as in non-immune individuals anti-DEKnull-2 IgG antibodies were boosted even after a brief exposure to P.vivax blood stages. Author summary Vaccines might be a crucial component of the current efforts to malaria control and elimination, and much of the vaccine-related research on P. vivax has been focused on region II of the Duffy binding protein (DBPII), a ligand for human blood-stage infection. Recently, the newly described Erythrocyte binding protein 2 (EBP2), a P.vivax DBP paralog that it is antigenically distinct from DBP, was identified as potential vaccine targets. To date, scarce data are available about the naturally acquired immunity to EBP2. In a small outbreak of P.vivax malaria, located in a non-malarious area, we investigated whether a first P.vivax exposure induces antibodies against EBP2 that could be boosted by P.vivax recurrent infections. In parallel, we included an engineered DBPII vaccine (named DEKnull-2) whose antibody response were previously associated with broadly neutralizing P.vivax antibodies. This study shows EBP2, compared with DEKnull-2, was poorly immunogenic among individuals who experienced their first blood-stage P. vivax malaria infection. However, EBP2 was highly immunogenic in long-term malaria exposed individuals, reinforcing its potential as a P. vivax blood-stage vaccine candidate. Finally, our results reinforce that multiple blood-stage antigens should be targeted for the development of efficient vaccines against P. vivax.