Independent Origin and Global Distribution of Distinct Plasmodium vivax Duffy Binding Protein Gene Duplications

Abstract

BackgroundPlasmodium vivax causes the majority of malaria episodes outside Africa, but remains a relatively understudied pathogen. The pathology of P. vivax infection depends critically on the parasite’s ability to recognize and invade human erythrocytes. This invasion process involves an interaction between P. vivax Duffy Binding Protein (PvDBP) in merozoites and the Duffy antigen receptor for chemokines (DARC) on the erythrocyte surface. Whole-genome sequencing of clinical isolates recently established that some P. vivax genomes contain two copies of the PvDBP gene. The frequency of this duplication is particularly high in Madagascar, where there is also evidence for P. vivax infection in DARC-negative individuals. The functional significance and global prevalence of this duplication, and whether there are other copy number variations at the PvDBP locus, is unknown.Methodology/Principal FindingsUsing whole-genome sequencing and PCR to study the PvDBP locus in P. vivax clinical isolates, we found that PvDBP duplication is widespread in Cambodia. The boundaries of the Cambodian PvDBP duplication differ from those previously identified in Madagascar, meaning that current molecular assays were unable to detect it. The Cambodian PvDBP duplication did not associate with parasite density or DARC genotype, and ranged in prevalence from 20% to 38% over four annual transmission seasons in Cambodia. This duplication was also present in P. vivax isolates from Brazil and Ethiopia, but not India.Conclusions/SignificancePvDBP duplications are much more widespread and complex than previously thought, and at least two distinct duplications are circulating globally. The same duplication boundaries were identified in parasites from three continents, and were found at high prevalence in human populations where DARC-negativity is essentially absent. It is therefore unlikely that PvDBP duplication is associated with infection of DARC-negative individuals, but functional tests will be required to confirm this hypothesis.

Genetic diversity in twoPlasmodium vivaxprotein ligands for reticulocyte invasion

“…vivax isolates collected in various malaria endemic settings had previously revealed that the PvDBP gene is frequently duplicated [17,18]. Here, we confirm again that amplification of PvDBPII is a common genomic event, which can frequently occur in isolates from areas where Duffy-negative phenotype is virtually absent such as Cambodia.…”

“…Initially, this epidemiological pattern suggested that the duplication of this gene was likely associated with the capability of the parasite to overcome the barrier of Duffy negativity. Since then, this hypothesis has been challenged by Hostetler et al [18] who found that PvDBP gene duplications were widespread even in malaria endemic areas in Southeast Asia where Duffy-negativity is not present. Another recent study [19] reported evidence of PvDBP gene amplification (3 and 8 copies) in two Duffynegative Ethiopian isolates.…”

Molecular interactions governing host-specificity of blood stage malaria parasites

“…To the contrary, Hostetler et al . [65] recently demonstrated that P. vivax isolates with this Pv DBP duplication are globally widespread and are unlikely to be associated with the African DARC-negative phenotype. Rather, these authors proposed that upregulation of an alternative invasion pathway—perhaps mediated by the recently described Pv EBP ligand [66] or the greatly expanded Pv RBP family [67,68]— may facilitate invasion of DARC-negative erythrocytes.…”

Development of copy number assays for detection and surveillance of piperaquine resistance associated plasmepsin 2/3 copy number variation in Plasmodium falciparum

“…The PCR primers used to detect the Cambodian breakpoint observed to date may not amplify a product in samples that contain different breakpoints. As previous studies have suggested [13,14], copy number variations in the same gene can arise independently on different genetic backgrounds and result in distinct molecular breakpoints of amplification. Since qPCR methods do not rely on the location of primers in reference to an estimated breakpoint, they can be used broadly.…”