The diversity of Plasmodium falciparum antigens is a major obstacle in designing a highly efficacious vaccine against malaria. PfRH5 is necessary for erythrocyte invasion of the parasite through interaction with host erythrocyte surface protein, basigin (CD147) making PfRH5 a promising blood-stage subunit vaccine candidate. PfRH5 is previously shown to be highly conserved with only five common non-synonymous single nucleotide polymorphisms (nsSNPs). Using longitudinal cohorts of children from the endemic areas of the Southwest Pacific (Papua New Guinea, PNG), we aimed to confirm whether genetic mutations lead to antigenic escape in natural infections, and which alleles to be included in PfRH5 vaccine formulation. A total of PCR amplified 534 antigen sequences from both clinical and non-clinical infections of 204 PNG children under 3 years of age were assembled and analyzed. Thirteen common nsSNPs were identified, which resulted in 44 unique PfRH5 haplotypes. However, the top 10 most frequent haplotypes accounted for 87% of the diversity and the 3D7 (reference) haplotype was only present amongst 2.7% of infections. These nsSNPs were found near the Basigin binding site as well as on the surface of PfRH5 protein, and four amino acid changes were found only in clinical infections. Significant balancing selection surrounded most polymorphisms however directional selection was also present. Two polymorphisms (S381L, S203Y) are strongly associated with clinical infections, both of which fall within regions of directional selection. Cryo-EM structure of the PfRh5/CyRPA/Ripr complex shows that S203Y is in close proximity to the Basigin binding site, and S381L is close to the contact point of its interaction partner Cysteine Rich Protective Antibody (CyRPA). These polymorphisms may constitute adaptations to antibody-mediated inhibition of Basigin binding and complex formation respectively. These results suggest that a single allele RH5 vaccine may have limited efficacy in PNG, and will allow the identification of critical PfRH5 haplotypes for inclusion in the vaccine.
Keywords: Plasmodium falciparum, PfRH5, Vaccine, Polymorphisms, Haplotypes