Malaria vaccine development of Plasmodium vivax, by using ookinete surface proteins as vaccine antigens

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In many malarious regions outside of Africa, development of transmission-blocking vaccine will require activity against both Plasimodiun falciparum and P.vivax. Work on P.vivax transmission-blocking vaccines have been hampered by the inability to clone the vaccine candidate genes from this parasite. To scarch for genes encoding the ookinete surface proteins from P.vivax, the gene sequences of tile eight known proteins in P25 subfamily (Pfs25, Pgs25, Pys25, Pbs25) and in P21/28 subfamily (Pfs28, Pgs28, Pys2l, Pbs2l) were aligned. Regions of highest identity were used to design degenerate PCR oligonucleotides. P.vivax Sail genomic DNA and genomic and splinkerette DNA libraries were used as PCR templates. Analysis of tile deduced amino acid sequence of Pvs28 revealed a secretory signal sequence, four EGF-like domains, six copies of the heptad amino acid repeat (GSGGE/D) and then a short hydrophobic region. Pvs28 is the presumed homologue of P21/28 subfamily member because the fourth EGF-like domain has four rather than six cysteines. Analysis of the deduced amino acid sequence of Pvs25 revealed a similar structure to Pvs28. Tile presence of six rather than four cysteines in tile fourth EGF-like domain suggested that Pvs25 is the homologue of P25 subfamily member. Accordingly, we have successfully cloned novel ookinete surface protein genes, Pvs28 and Pvs25, as the transmission-blocking vaccille candidate antigens, from P.vivax.