Sequence polymorphism in two novel Plasmodium vivax ookinete surface proteins, Pvs25 and Pvs28, that are malaria transmission-blocking vaccine candidates

Background: For many malarious regions outside of Africa, development of ef fective transmission-blocking vaccines will require coverage against both P lasmodium falciparum and P. vivax. Work on P. vivax transmission-blocking v accines has been hampered by the inability to clone the vaccine candidate g enes from this parasite. Materials and Methods: To search for genes encoding the ookinete surface pr oteins from P. vivax, the DNA sequences of the eight known proteins in the P25 subfamily (Pfs25, Pgs25, Pys25, Pbs25) and in the P21/28 subfamily (Pfs 28, Pgs28, Pys21, Pbs21) of zygote/ookinete surface proteins were aligned. Regions of highest identity were used to design degenerate PCR oligonucleot ides. Genomic DNA from the Sal I strain of P. vivax and genomic and splinke rette DNA libraries were used as PCR templates. To characterize the polymor phisms of Pvs25 and Pvs28, these two genes were PCR amplified and the DNA s equences were determined from genomic DNA extracted from patients infected with P. vivax. Results: Analysis of the deduced amino add sequence of Pvs28 revealed a sec retory signal sequence, lour epidermal growth factor (EGF)-like domains, si x copies of the heptad amino acid repeat (GSGCE/D), and a short hydrophobic region. Because the fourth EGF-like domain has four rather than six cystei nes, the gene designated Pvs28 is the presumed homologue of P21/28 subfamil y members. Analysis of the deduced amino acid sequence of Pvs25 revealed a similar structure to that of Pvs28. The presence of six rather than four cy steines in the fourth EGF-like domain suggested that Pvs25 is the homologue of P25 subfamily members. Several regions of genetic polymorphisms in Pvs2 5 and Pvs28 were identified in field isolates of P. vivax. Conclusions: The genes encoding two ookinete surface proteins, Pvs28 and Pv s25, from P. vivax have been isolated and sequenced. Comparison of the prim ary structures of Pvs25, Pvs28, Pfs25, and Pfs28 suggest that there are reg ions of genetic polymorphism in the P25 and P21/28 subfamilies.