Plasmodium vivax Cysteine-Rich Protective Antigen Polymorphism at Exon-1 Shows Recombination and Signatures of Balancing Selection

Abstract

Plasmodium vivax Cysteine-Rich Protective Antigen (CyRPA) is a merozoite protein participating in the parasite invasion of human reticulocytes. During natural P. vivax infection, antibody responses against PvCyRPA have been detected. In children, low anti-CyRPA antibody titers correlated with clinical protection, which suggests this protein as a potential vaccine candidate. This work analyzed the genetic and amino acid diversity of pvcyrpa in Mexican and global parasites. Consensus coding sequences of pvcyrpa were obtained from seven isolates. Other sequences were extracted from a repository. Maximum likelihood phylogenetic trees, genetic diversity parameters, linkage disequilibrium (LD), and neutrality tests were analyzed, and the potential amino acid polymorphism participation in B-cell epitopes was investigated. In 22 sequences from Southern Mexico, two synonymous and 21 nonsynonymous mutations defined nine private haplotypes. These parasites had the highest LD-R2 index and the lowest nucleotide diversity compared to isolates from South America or Asia. The nucleotide diversity and Tajima’s D values varied across the coding gene. The exon-1 sequence had greater diversity and Rm values than those of exon-2. Exon-1 had significant positive values for Tajima’s D, β-α values, and for the Z (HA: dN > dS) and MK tests. These patterns were similar for parasites of different origin. The polymorphic amino acid residues at PvCyRPA resembled the conformational B-cell peptides reported in PfCyRPA. Diversity at pvcyrpa exon-1 is caused by mutation and recombination. This seems to be maintained by balancing selection, likely due to selective immune pressure, all of which merit further study.

Genetic Diversity of Plasmodium vivax Cysteine-Rich Protective Antigen (PvCyRPA) in Field Isolates from Five Different Areas of the Brazilian Amazon

“…To analyze hypothetical PvCyRPA protein derivatives from P. vivax genome data available and Mexico isolates already available, multiple sequence alignments were conducted with Clustal Omega using the MegAlign Pro 15 (Lasergene DNASTAR) program. The following sequences were used: India VII-GenBank: gb|KMZ81773.1; North Korean-GenBank: gb|KNA00954.1; Mauritania I-Genbank: gb|KMZ94334.1; Brazil I-GenBank: gb|KMZ87926.1; Sanger Institute: SCO 66052.1; Sanger Institute: SCO71483.1; Sanger Institute: SGX76259.1; Mexico-Southern Mexican [23]. Additionally, our Brazilian Amazon isolates were compared to P01 strain (PVP01_0532400), a new reference genome for P. vivax from an Indonesian clinical isolate [32], and the circular map of protein alignment was generated using the software GenVision v15 (Lasergene DNASTAR).…”

“…As observed in the protein sequence alignments, the PvCyRPA coding gene had an excess of non-synonymous mutations, which were more frequent in exon-1 than in exon-2. In addition, we subsequently aligned the protein sequence of these mutant field isolates with other hypothetical CyRPA proteins derivative from P. vivax genome data available in the GenBank database and also aligned with the isolate Mexico-Southern Mexican [23]. Among a total of 31 in PvCyRPA protein observed in P. vivax sequences worldwide, 26 amino acid substitutions are also present in our isolates.…”

“…The genetic diversity and population structure of P. vivax for each candidate antigen is an important priority to the understanding of the malaria transmission dynamics [21]. In this scenario, many studies have been proposed to investigate the global diversity of leading vaccine antigens [22], and only one was recently addressed to the PvCyRPA [23], which does not include Brazilian malaria-endemic areas. Therefore, to understand the potential of PvCyRPA in vaccine development, we proposed to identify pvcyrpa gene in clinical isolates from different regions of the Brazilian Amazon and to study the potential impacts of the genetic diversity in predicted epitopes through bioinformatics tools.…”