The evolution and diversity of a low complexity vaccine candidate, merozoite surface protein 9 (MSP-9), in Plasmodium vivax and closely related species

Abstract

The merozoite surface protein-9 (MSP-9) has been considered a target for an anti-malarial vaccine since it is one of many proteins involved in the erythrocyte invasion, a critical step in the parasite life cycle. Orthologs encoding this antigen have been found in all known species of Plasmodium parasitic to primates. In order to characterize and investigate the extent and maintenance of MSP-9 genetic diversity, we analyzed DNA sequences of the following malaria parasite species: Plasmodium falciparum, Plasmodium reichenowi, Plasmodium chabaudi, Plasmodium yoelii, Plasmodium berghei, Plasmodium coatneyi, Plasmodium gonderi, Plasmodium knowlesi, Plasmodium inui, Plasmodium simiovale, Plasmodium fieldi, Plasmodium cynomolgi and Plasmodium vivax and evaluated the signature of natural selection in all MSP-9 orthologs. Our findings suggest that the gene encoding MSP-9 is under purifying selection in P. vivax and closely related species. We further explored how selection affected different regions of MSP-9 by comparing the polymorphisms in P. vivax and P. falciparum, and found contrasting patterns between these two species that suggest differences in functional constraints. This observation implies that the MSP-9 orthologs in human parasites may interact differently with the host immune response. Thus, studies carried out in one species cannot be directly translated into the other.

Keywords: ABRA; Binding sites; Genetic diversity; MSP-9; Merozoite surface proteins; Plasmodium.

Fig. 1

Diagram of the MSP-9 Plasmodium spp protein alignment (partial sequence of P. inui was not included). The length of the protein is indicated for each species. The putative signal peptide, cysteines, RBC binding regions (Kushwaha et al., 2002, Li et al., 2004) and repetitive regions (R, R1, R2, R3 and R4) are shown. Blocks with different patterns represent different repetitive regions. Gaps in the protein alignment are represented by blank areas.

Fig. 2

Repetitive regions (R1, R2, R3 and R4) of PvMSP-9. Letters A-H represent repetitive motifs and numbers next to the letters represent motifs with non-synonymous substitutions (see Supplementary Table S1).

Fig. 3

Alignment of putative T-cell epitopes of PvMSP-9 (Lima-Junior et al., 2008), including different Plasmodium species. The positions underlined correspond to the carboxy-terminal region of peptide pL.

Fig. 4

Bayesian phylogenetic tree of the gene encoding MSP-9 using MrBayes. The phylogeny was estimated using solely alignable sites on the N-terminal (1,294 bp). The numbers on nodes of the tree are posterior probabilities represented as percentage based on 3×10⁶ Markov Chain Monte Carlo (MCMC) steps; the first 1,000,000 (50%) were discarded as a burn-in. Sampling was performed every 100 generations.