The effect of declining exposure on T cell-mediated immunity to Plasmodium falciparum - an epidemiological "natural experiment".

BACKGROUND: Naturally acquired immunity to malaria may be lost with lack of exposure. Recent heterogeneous reductions in transmission in parts of Africa mean that large populations of previously protected people may lose their immunity while remaining at risk of infection. METHODS: Using two ethnically similar long-term cohorts of children with historically similar levels of exposure to Plasmodium falciparum who now experience very different levels of exposure, we assessed the effect of decreased parasite exposure on antimalarial immunity. Peripheral blood mononuclear cells (PBMCs) from children in each cohort were stimulated with P. falciparum and their P. falciparum-specific proliferative and cytokine responses were compared. RESULTS: We demonstrate that, while P. falciparum-specific CD4+ T cells are maintained in the absence of exposure, the proliferative capacity of these cells is altered considerably. P. falciparum-specific CD4+ T cells isolated from children previously exposed, but now living in an area of minimal exposure ("historically exposed") proliferate significantly more upon stimulation than cells isolated from children continually exposed to the parasite. Similarly, PBMCs from historically exposed children expressed higher levels of pro-inflammatory cytokines and lower levels of anti-inflammatory cytokines after stimulation with P. falciparum. Notably, we found a significant positive association between duration since last febrile episode and P. falciparum-specific CD4+ T cell proliferation, with more recent febrile episodes associated with lower proliferation. CONCLUSION: Considered in the context of existing knowledge, these data suggest a model explaining how immunity is lost in absence of continuing exposure to P. falciparum.

Plasmodium falciparum antigenic variation: relationships between widespread endothelial activation, parasite PfEMP1 expression and severe malaria.

BACKGROUND: Plasmodium falciparum erythrocyte membrane protein 1(PfEMP1) is a family of variant surface antigens (VSA) that mediate the adhesion of parasite infected erythrocytes to capillary endothelial cells within host tissues. Opinion is divided over the role of PfEMP1 in the widespread endothelial activation associated with severe malaria. In a previous study we found evidence for differential associations between defined VSA subsets and specific syndromes of severe malaria: group A-like PfEMP1 expression and the "rosetting" phenotype were associated with impaired consciousness and respiratory distress, respectively. This study explores the involvement of widespread endothelial activation in these associations. METHODS: We used plasma angiopoietin-2 as a marker of widespread endothelial activation. Using logistic regression analysis, we explored the relationships between plasma angiopoietin-2 levels, parasite VSA expression and the two syndromes of severe malaria, impaired consciousness and respiratory distress. RESULTS: Plasma angiopoietin-2 was associated with both syndromes. The rosetting phenotype did not show an independent association with respiratory distress when adjusted for angiopoietin-2, consistent with a single pathogenic mechanism involving widespread endothelial activation. In contrast, group A-like PfEMP1 expression and angiopoietin-2 maintained independent associations with impaired consciousness when adjusted for each other. CONCLUSION: The results are consistent with multiple pathogenic mechanisms leading to severe malaria and heterogeneity in the pathophysiology of impaired consciousness. The observed association between group A-like PfEMP1 and impaired consciousness does not appear to involve widespread endothelial activation.

Transfer of 4-hydroxynonenal from parasitized to non-parasitized erythrocytes in rosettes. Proposed role in severe malaria anemia.

Severe anaemia is a life-threatening complication of falciparum malaria associated with loss of predominantly non-parasitized red blood cells (npRBCs). This poorly elucidated process might be influenced by (i) rosettes, i.e. npRBCs cytoadherent to haemozoin-containing parasitized RBCs (pRBCs) and (ii) generation in pRBCs of 4-hydroxynonenal (4-HNE) through haemozoin-catalysed lipid peroxidation. We explored whether close proximity in rosettes may facilitate 4-HNE transfer to npRBCs, which is likely to enhance their phagocytosis and contribute to malaria anaemia. Fluorescence microscopy and flow cytometry data indicated 4-HNE transfer to npRBCs in rosettes. Rosettes were formed by 64·8 ± 1·8% varO-expressing pRBCs, and 8·7 ± 1·1% npRBCs were positive for 4-HNE-protein-conjugates, while low-rosetting parasites generated only 2·4 ± 1·1% 4-HNE-conjugate-positive npRBCs. 4-HNE transfer decreased after blocking rosetting by monoclonal antibodies. A positive linear relationship between rosette frequency and 4-HNE-conjugates in npRBCs was found in 40 malaria patients, a first indication for a role of rosetting in npRBCs modifications in vivo. Children with severe malaria anaemia had significantly higher percentages of 4-HNE-conjugate-positive npRBCs compared to children with uncomplicated malaria. In conclusion, 4-HNE transfer from pRBCs to npRBCs in rosettes is suggested to play a role in the phagocytic removal of large numbers of npRBCs, the hallmark of severe malaria anaemia.

Prognostic indicators of life-threatening malaria are associated with distinct parasite variant antigen profiles.

PfEMP1 is a family of cytoadhesive surface antigens expressed on erythrocytes infected with Plasmodium falciparum, the parasite that causes the most severe form of malaria. These surface antigens play a role in immune evasion and are thought to contribute to the pathogenesis of the malaria parasite. Previous studies have suggested a role for a specific subset of PfEMP1 called "group A" in severe malaria. To explore the role of group A PfEMP1 in disease, we measured the expression of the var genes that encode them in parasites from clinical isolates collected from children suffering from malaria. We also looked at the ability of these clinical isolates to induce rosetting of erythrocytes, which indicates a cytoadhesion phenotype that is thought to be important in pathogenesis. These two sets of data were correlated with the presence of two life-threatening manifestations of severe malaria in the children: impaired consciousness and respiratory distress. Using regression analysis, we show that marked rosetting was associated with respiratory distress, whereas elevated expression of group A-like var genes without elevated rosetting was associated with impaired consciousness. The results suggest that manifestations of malarial disease may reflect the distribution of cytoadhesion phenotypes expressed by the infecting parasite population.