New PCR assays to measure expression of plasmodium vivax reticulocyte binding proteins

Abstract

Invasion of red blood cells is a crucial step in malaria infection. Different malaria parasites have different preference for host cells. Plasmodium falciparum infects both reticulocytes and fully mature erythrocytes; Plasmodium vivax only infects reticulocytes. This difference is explained by the different ligand-receptor interactions used by these parasites. Two invasion ligand families have been implicated in host cell sensing during invasion: I. the Erythrocyte Binding Antigens (EBA) and II. the Reticulocyte Binding Proteins (RBP). The latter are believed to mediate specific binding of P. vivax merozoites to reticulocytes. Protective immunity for malaria has been linked to the presence of antibodies against parasite invasion ligands. These proteins thus represent prime targets for vaccine development. While extensive studies have been performed on P. falciparum proteins and P. vivax Duffy Binding protein, a member of the EBA family, little has been done on P. vivax RBPs which comprise the vast majority of invasion ligands of this parasite. In this study, we aim to use quantitative polymerase chain reaction (qPCR) to measure the expression of the 9 different RBPs identified in the reference genome of P. vivax. The repertoire of RBP expression across different parasite isolates will provide valuable insights into P. vivax invasion and vaccine development. Here, we report our qPCR assays which have been optimized for robust quantification of RBP transcripts as well as the transcripts of house-keeping and schizont-specific genes. Preliminary measurements of a few clinical isolates are presented.