Malaria parasites rely on cycles of cellular invasion and intracellular growth to proliferate within the blood stream, a process underpining symptoms of the disease. Two major protein families have been studied extensively in P. falciparum (Pf): the erythrocyte binding-like proteins (DBPs/EBAs) and the reticulocyte binding-like proteins (RBLs/RHs). These proteins are hypothesized to have overlapping but critical roles during the invasion process. The zoonotic malaria parasite P. knowlesi much larger and polarised invasive stages known as merozoites, has larger merozoites and a smaller repertoire of these proteins – providing an ideal model to study their role with live microscopy.
Employing a conditional knockout approach, we've demonstrated distinct roles for the two families at different invasion stages, including a specific role for RBL proteins in the initial identification and deformation of target host erythrocytes. Furthermore, we've unearthed new features that prompt a significant reassessment of the invasion process. These proteins are also major candidates for blood stage vaccines against P. vivax, and we have been able to show that by swapping PvDBP orthologues into P. knowlesi we have developed a scalable model to determine which antibodies/sera are most effective at blocking invasion, enabling rationale antigen design. Finally, by employing a new Bar-Seq based approach we call P. knowlesi Multi Allelic Pools (PkMAPs) we can screen antibodies against pools of parasites expressing polymorphic variants of PvDBP – enabling us to study the effect of the majority of known PvDBP diversity combined in a single well of a 96-well plate.