Search

Anemia is a common condition in HIV-infected children; however, its pathophysiology and the contribution of frequent causes of anemia such as iron deficiency (ID) and malaria are poorly understood. We carried out an ancillary study on the effect of HIV on anemia as part of a case-control study on risk factors of anemia among Mozambican children aged 1-59 months with documented HIV status. Of them, 390 children were admitted to the hospital with anemia (hemoglobin [Hb] < 11 g/dL), whereas 272 children without anemia (Hb ≥ 11 g/dL) were recruited in the community. We assessed differences by HIV status in the presentation of anemia etiological factors and the effect of HIV infection on the association of each factor with anemia. Among the 99 HIV-infected and 563 uninfected children included, HIV-infected anemic children had an increased risk of undernutrition (P < 0.0001), Epstein-Barr virus infection (P < 0.0001), bacteremia (P = 0.0060), a decreased risk of malaria (P < 0.0001), and a similar risk of ID (P = 0.7371) compared with anemic-uninfected children. HIV-infected children were significantly less likely to have anemia associated with Plasmodium falciparum hyperparasitemia (P = 0.0444) and had a lower prevalence of parasitemia in the bone marrow (BM) (P < 0.0001) than anemic-uninfected children. Levels of BM erythropoiesis and dyserythropoiesis were comparable between groups. These findings suggest that the pathophysiology of anemia among HIV-infected malaria-exposed children is not related to HIV-specific effects. For unclear reasons, HIV-infected children had reduced risk of malaria infection, whereas ID prevalence was comparable in HIV-infected and uninfected children, suggesting that iron supplementation recommendations should not be different in HIV-infected children.

Additional File 2. Supplementary methods and results for immunogenicity assessments of GAP vaccine. Text S1. Methodology for determining antibody mediated immune response to GAP vaccine. Text S2. Methodology for determining cell mediated immune response to GAP vaccine. Table S1. Antibodies used for flow cytometry. Figure S1. The gating strategy used to identify CD4+ T cells and subsets, as well as the expression of T cell activation markers. Figure S2. The anti-MSP2 IgM response over the course of the study in each subject. Figure S3. The IgG and IgM response to whole parasite lysate over the course of the study in each subject. Figure S4. The frequency of CD4+ T cells, Treg cells, Th1 cells and Tfh cells in peripheral blood mononuclear cells over the course of the study in each subject.

Additional file 1: Table S1. Flow chart of the feeding parameters that were tested progressively. The selected parameters are in bold and were used in the subsequent tests

Additional File 3. Supplementary methods and results for in vitro assessments of GAP vaccine. Text S1. Methodology for parasite genotyping. Text S2. Methodology for cytoadherence assays. Text S3. Methodology for scanning electron microscopy. Text S4. Methodology for var gene expression analysis. Table S1. Duration of in vitro culture of P. falciparum prior to harvest for RNA extraction for var gene transcription analysis. Text S5. Results of parasite genotyping. Text S6. Results of scanning electron microscopy analysis of parasite-infected erythrocytes. Text S7. Results of cytoadherence assays. Text S8. Results of var gene expression analysis. Table S2. Most abundantly expressed var genes.

Additional File 1. Supplementary methods and results for subject eligibility and safety assessments. Text S1: Subject eligibility criteria. Table S1: Incidence of adverse events by subject. Table S2: Red blood cell alloantibody test results for Cohort 4 and phenotype of donor red blood cells used for GAP master cell bank manufacture.

Human malaria parasites proliferate in different erythroid cell types during infection. Whilst Plasmodium vivax exhibits a strong preference for immature reticulocytes, the more pathogenic P. falciparum primarily infects mature erythrocytes. In order to assess if these two cell types offer different growth conditions and relate them to parasite preference, we compared the metabolomes of human and rodent reticulocytes with those of their mature erythrocyte counterparts. Reticulocytes were found to have a more complex, enriched metabolic profile than mature erythrocytes and a higher level of metabolic overlap between reticulocyte resident parasite stages and their host cell. This redundancy was assessed by generating a panel of mutants of the rodent malaria parasite P. berghei with defects in intermediary carbon metabolism (ICM) and pyrimidine biosynthesis known to be important for P. falciparum growth and survival in vitro in mature erythrocytes. P. berghei ICM mutants (pbpepc-, phosphoenolpyruvate carboxylase and pbmdh-, malate dehydrogenase) multiplied in reticulocytes and committed to sexual development like wild type parasites. However, P. berghei pyrimidine biosynthesis mutants (pboprt-, orotate phosphoribosyltransferase and pbompdc-, orotidine 5'-monophosphate decarboxylase) were restricted to growth in the youngest forms of reticulocytes and had a severe slow growth phenotype in part resulting from reduced merozoite production. The pbpepc-, pboprt- and pbompdc- mutants retained virulence in mice implying that malaria parasites can partially salvage pyrimidines but failed to complete differentiation to various stages in mosquitoes. These findings suggest that species-specific differences in Plasmodium host cell tropism result in marked differences in the necessity for parasite intrinsic metabolism. These data have implications for drug design when targeting mature erythrocyte or reticulocyte resident parasites.

Improving the efficiency of malaria diagnosis is one of the main goals of current malaria research. We have recently developed a magneto-optical (MO) method which allows high-sensitivity detection of malaria pigment (hemozoin) crystals via their magnetically induced rotation in blood. Here, we validate this technique on laboratory derived blood samples infected with \textit{Plasmodium falciparum}. Using two parasite cultures, the first containing mostly ring stages and the second corresponding to the end of the parasite life cycle, we demonstrate that our novel method can detect parasite densities as low as $\sim$40 and $\sim$10\,parasites per microliter of blood for ring and schizont stage parasites, respectively. This detection limit exceeds the performance of rapid diagnostic tests and competes with the threshold achievable by light microscopic observation of blood smears. Our method can be performed with as little as 50\,microliter of capillary blood and is sensitive to the presence of hemozoin micro-crystals down to ppm concentrations. The device, designed to a portable format for clinical and in-field tests, requires no special training of the operator or specific reagents, except for an inexpensive lysis solution to release intracellular hemozoin. Beyond diagnostics, this technique may offer an efficient tool to study hemozoin formation, trace hemozoin kinetics in the body and test susceptibility/resistance of parasites to new antimalarial drugs inhibiting hemozoin formation.

The infection of mice with Plasmodium berghei ANKA constitutes the best available mouse model for human Plasmodium falciparum-mediated cerebral malaria, a devastating neurological syndrome that kills nearly 2.5 million people every year. Experimental data suggest that cerebral disease results from the sequestration of parasitized erythrocytes within brain blood vessels, which is exacerbated by host proinflammatory responses mediated by cytokines and effector cells including T lymphocytes. Here, T cell responses to P. berghei ANKA were analyzed in cerebral malaria-resistant and -susceptible mouse strains. CD4+ T-cell proliferation and interleukin-2 (IL-2) production in response to parasite-specific and polyclonal stimuli were strongly inhibited in cerebral malaria-resistant mice. In vitro and in vivo depletion of CD4+ CD25+ regulatory T (T(reg)) cells significantly reversed the inhibition of CD4+ T-cell proliferation and IL-2 production, indicating that this cell population contributes to the suppression of T-cell function during malaria. Moreover, in vivo depletion of T(reg) cells prevented the development of parasite-specific TH1 cells involved in the induction of cerebral malaria during a secondary parasitic challenge, demonstrating a regulatory role for this cell population in the control of pathogenic responses leading to fatal disease.

Additional file 4. Influence of age and exposure on antibody levels to PfGPI in Papua New Guinean children.

Additional file 4. Influence of age and exposure on antibody levels to PfGPI in Papua New Guinean children.

Additional file 3. Association between IgG to PfGPI and protection against clinical malaria in Papua New Guinean children.

Additional file 3. Association between IgG to PfGPI and protection against clinical malaria in Papua New Guinean children.

Additional file 2. IgG to PfGPI by village of residency.

Additional file 2. IgG to PfGPI by village of residency.