Reticulocyte Preference and Stage Development of Plasmodium vivax Isolates

Abstract

Plasmodium vivax, the most widely distributed human malaria parasite, is restricted to reticulocytes, limiting its asexual proliferation. In recent years, cases of severe and high-level P. vivax parasitemia have been reported, challenging the assumption that all isolates are equally restricted. In this article, we analyze the reticulocyte preference of a large number of Indian P. vivax isolates. Our results show that P. vivax isolates significantly vary in their level of reticulocyte preference. In addition, by carefully staging the parasites, we find that P. vivax schizonts are largely missing in peripheral blood, with the presence of schizonts in circulation correlating with a high reticulocyte preference.

Keywords: Plasmodium vivax; asexual stage; reticulocyte preference.

Figure 1.

Parasitemia range, reticulocytemia range, and stage distribution of Plasmodium vivax isolates. A, P. vivax–infected patients exhibit a range of parasitemia and reticulocytemia percentages at admission, but these values are not correlated (Spearman correlation; P = .8892). B, Schematic asexual blood stage development time line. The parasitized cells were categorized into 5 stages of development (stages I–IV and the gametocyte stage [G]). C, The ring proportion (stage I) of each sample was plotted against the reticulocytemia percentage. There was no correlation between the 2 values (Spearman correlation; P = .7726). D, The proportions of each stage were calculated for all samples. The number above each stage corresponds to the percentage of samples that had at least 1 of the each stage in circulation (n = 113).

Figure 2.

Reticulocyte preference measurement and correlation with stages. A, Blood samples were stained for both parasites and reticulocytes, allowing us to detect the red blood cell type the parasite is in. Black arrowheads point to parasitized cells, and the empty arrows point to reticulocytes. B, As the percentage of reticulocytes is much lower than that of normocytes, the parasitemia percentage among infected normocytes is generally higher than the percentage of infected reticulocytes (left panel). When compared as proportion of each cell type that is infected (that is, the proportion of all reticulocytes that contain a parasite), all samples have a higher proportion of reticulocytes (right panel). C, The ratio of each data point from the 2 graphs in panel (B) results in the relative frequency of infection in reticulocytes (RFR). RFRs have a wide range and correlate with their corresponding selectivity index (Spearman correlation; P = .0193). D, The samples were divided into 4 groups of increasing RFR (<50, 50 to <100, 100 to <200, and ≥200). The number of samples in each bin is indicated above each column. The proportions of the later stages (IV) and gametocyte (G) stage for each RFR bin were compared. There were significantly more schizonts in samples with high RFRs (≥200, indicating higher restriction and selectivity; P = .042, by the Kruskal-Wallis test), with comparable proportions of gametocytes (P = .1836).