Studies of <i>P. falciparum</i> invasion of and development in RBCs containing hemoglobin variants.

<p>Unless otherwise stated, assessments of RBC invasion and growth are relative to HbAA or non-thalassemic RBCs.</p><p>NR, not reported; HPFH, syndrome of hereditary persistence of fetal hemoglobin.</p

The major hemoglobinopathies: epidemiology, molecular pathology, and clinical phenotype.

<p>The human genome normally contains four copies of α-globin genes (in paired copies on chromosome 16: genotype αα/αα) and two copies of β-globin genes (on chromosome 11). Normal adult hemoglobin (HbAA) is a tetramer of two α-globin and two β-globin proteins.</p>a<p>Not technically a hemoglobinopathy but rather a normal hemoglobin variant of all newborns and infants.</p

General mechanisms by which hemoglobinopathies may attenuate the pathogenesis of falciparum malaria.

<p>(A) Restriction of red blood cell (RBC) invasion or intraerythrocytic growth, thereby suppressing parasite densities <i>in vivo</i>; (B) interference with parasite-derived mediators of pathogenesis, including those involved in the binding of parasite-infected RBCs (iRBCs) to extracellular host receptors; (C) modulation of innate host defenses to favor protective, anti-inflammatory responses over those that drive pathogenic, pro-inflammatory responses; (D) enhancement of adaptive cell-mediated and humoral immune responses that clear iRBCs from the blood.</p

The top <i>Rsb</i> hits for Cambodia using Thailand as the reference population.

<p>Dashed lines indicate the top 1% of <i>Rsb</i> values for imputed (above zero) and complete-case (below zero) data.</p

The top <i>Rsb</i> hits for Thailand, Cambodia, and Gambia, using Malawi as the reference population.

<p>Dashed lines indicate the top 1% of <i>Rsb</i> values (median threshold across 3 populations) for Beagle-imputed (above zero) and complete-case (below zero) data.</p

Linkage disequilibrium (LD) decay as a function of genomic distance in each population, using <i>r</i>^<i>2</i> between pairs of markers with MAF≥5% and within 1000 bp of each other.

<p>The plotted <i>r</i>^<i>2</i> is the mean calculated for pairwise distances binned in 5-bp increments. LD decays to <i>r</i>^<i>2</i> = 0.2 within approximately 187 bp for Cambodia, 162 bp for Thailand, 102 bp for Gambia, and 67 bp for Malawi.</p

Genes (n = 62) with ≥10 SNPs having <i>Tajima’s D</i> (TD) values >1 in populations from Thailand (total 793 genes), Cambodia (1208 genes), Gambia (723 genes), and Malawi (812 genes).

<p>Genes (n = 62) with ≥10 SNPs having <i>Tajima’s D</i> (TD) values >1 in populations from Thailand (total 793 genes), Cambodia (1208 genes), Gambia (723 genes), and Malawi (812 genes).</p

A Library of <i>Plasmodium vivax</i> Recombinant Merozoite Proteins Reveals New Vaccine Candidates and Protein-Protein Interactions

<div><p>Background</p><p>A vaccine targeting <i>Plasmodium vivax</i> will be an essential component of any comprehensive malaria elimination program, but major gaps in our understanding of <i>P</i>. <i>vivax</i> biology, including the protein-protein interactions that mediate merozoite invasion of reticulocytes, hinder the search for candidate antigens. Only one ligand-receptor interaction has been identified, that between <i>P</i>. <i>vivax</i> Duffy Binding Protein (PvDBP) and the erythrocyte Duffy Antigen Receptor for Chemokines (DARC), and strain-specific immune responses to PvDBP make it a complex vaccine target. To broaden the repertoire of potential <i>P</i>. <i>vivax</i> merozoite-stage vaccine targets, we exploited a recent breakthrough in expressing full-length ectodomains of <i>Plasmodium</i> proteins in a functionally-active form in mammalian cells and initiated a large-scale study of <i>P</i>. <i>vivax</i> merozoite proteins that are potentially involved in reticulocyte binding and invasion.</p><p>Methodology/Principal Findings</p><p>We selected 39 <i>P</i>. <i>vivax</i> proteins that are predicted to localize to the merozoite surface or invasive secretory organelles, some of which show homology to <i>P</i>. <i>falciparum</i> vaccine candidates. Of these, we were able to express 37 full-length protein ectodomains in a mammalian expression system, which has been previously used to express <i>P</i>. <i>falciparum</i> invasion ligands such as PfRH5. To establish whether the expressed proteins were correctly folded, we assessed whether they were recognized by antibodies from Cambodian patients with acute vivax malaria. IgG from these samples showed at least a two-fold change in reactivity over naïve controls in 27 of 34 antigens tested, and the majority showed heat-labile IgG immunoreactivity, suggesting the presence of conformation-sensitive epitopes and native tertiary protein structures. Using a method specifically designed to detect low-affinity, extracellular protein-protein interactions, we confirmed a predicted interaction between <i>P</i>. <i>vivax</i> 6-cysteine proteins P12 and P41, further suggesting that the proteins are natively folded and functional. This screen also identified two novel protein-protein interactions, between P12 and PVX_110945, and between MSP3.10 and MSP7.1, the latter of which was confirmed by surface plasmon resonance.</p><p>Conclusions/Significance</p><p>We produced a new library of recombinant full-length <i>P</i>. <i>vivax</i> ectodomains, established that the majority of them contain tertiary structure, and used them to identify predicted and novel protein-protein interactions. As well as identifying new interactions for further biological studies, this library will be useful in identifying <i>P</i>. <i>vivax</i> proteins with vaccine potential, and studying <i>P</i>. <i>vivax</i> malaria pathogenesis and immunity.</p><p>Trial Registration</p><p>ClinicalTrials.gov <a href="https://clinicaltrials.gov/ct2/show/NCT00663546" target="_blank">NCT00663546</a></p></div

The top |<i>iHS</i>| hits in 4 global <i>P</i>. <i>falciparum</i> populations.

<p>Dashed lines indicate the top 1% of |<i>iHS</i>| values (median threshold across 4 populations) for Beagle-imputed (above zero) and complete-case (below zero) data. The positions of genes known to confer drug resistance or encode vaccine candidates are annotated.</p

Relationship between imputation accuracy and minor allele frequency (MAF), tested on chromosome 13 SNPs.

<p>MAF-wise means are used to infer the trend in <i>r</i>^<i>2</i>. Only designs using LDhat-inferred recombination rates are considered for IMPUTE. Use of a cosmopolitan reference panel improves accuracy across both common and low-frequency SNPs in IMPUTE; however, accuracy decreases substantially in Beagle. SNPs in Malawi have a maximum MAF = 0.23.</p