A study of Plasmodium vivax Duffy binding protein (DBP) in West Papuan isolates

The invasion of immature red blood cells called reticulocytes is a critical event for the Plasmodium vivax merozoite to continue its life-cycle in humans. Plasmodium vivax employs the Duffy binding protein (DBP) ligand to engage its host receptor, the Duffy antigen receptor for chemokines (DARC), in initiating the essential formation of a tight junction between the merozoite and erythrocyte for reticulocyte invasion. Because of this critical step, PvDBP is a promising antigen to be developed as vaccine against P. vivax infection. However, genetic diversity of this putative vaccine antigen particularly in the central domain of region II of DBP that serves as the critical binding motif to the DARC might pose a serious problem in designing an effective vaccine which elicits neutralizing immunity toward this ligand. Information about PvDBPIl polymorphisms has so far been obtained from the Papua New Guinea (PNG), Colombia, Iran, Korea, Brazil, and Thailand, but none reported from Indonesia. This thesis seeks to develop a better understanding of the host-parasite interaction by characterizing the extent of polymorphisms of PvDBPII from clinical isolates obtained from Timika on the south central coast of Papua, Indonesia. Sequence analyses of nucleotide and amino acid of PvDBPII obtained from Timikan isolates, demonstrated that P. vivax populations from Indonesia were genetically highly diverse, as shown by the large number of substitutions found in this region of DBP compared with the reference strain, Sal-I. Mutations were dominated by non-synonymous substitutions resulting in amino acid changes and mainly occurred in the critical binding motif of PvDBPII. Statistical analyses and the presence of polymorphisms associated with neutralizing B-cell linear epitopes confirmed the action of positive selection by host immune pressure to region II of PvDBP among Timikan isolates, which may play an important role in generating and maintaining certain haplotypes in P. vivax populations circulating in Timika. The study presented in this thesis revealed that the most prevalent substitutions found in Indonesia are also observed in other distinct geographical regions. Furthermore, the phylogenetic tree study showed that Timikan PvDBPII isolates clustered together with isolates from certain other geographic regions, including Brazil, Sri Lanka, and Papua New Guinea. In this study, we also produced an allele of PvDBPII-INA21 type as full-length sequence recombinant protein, expressed in E. coli bacterial system, to be used for serological assay. Western blot analysis demonstrated that most sera tested from P. vivax-infected individuals living in P. vivax highly endemic area Timika, Papua, Indonesia, contained antibodies that responded to this recombinant of PvDBPII derived from INA21 haplotype. This suggested cross-reactivity of Timikan sera to conserved PvDBP antigens.