HbAA Children Research Articles

Relationship between red blood cell polymorphisms and effectiveness of seasonal malaria chemoprevention in 2020 in Dangassa, Mali.

Treatment failure with amodiaquine was reported in Dangassa, where red blood cell (RBC) polymorphisms are found and seasonal malaria chemoprevention (SMC) is underway. Here, we aimed at assessing the influence of RBC polymorphisms on SMC effectiveness. This was a secondary analysis of data from a study conducted in Dangassa. Children aged 5 to 14years enrolled in an open randomized study were assigned either to receive SMC (intervention arm) or not (control arm). SMC was implemented from July to November. For all children, hemoglobin type and blood group were determined at enrolment in July, and parasitemia and hemoglobin level were monthly monitored by finger-prick. Overall, 166 children were enrolled among which 82 (49.40%) in the control arm and 84 (50.60%) in the SMC arm. The prevalence of HbAS was 10.24% (17/166) with 12.20% in the control and 8.33% in the SMC arm. O group was the most common overall (45%) and in the SMC arm (54%), but the control arm had more B (39.02%) than O (36.59%). In the SMC arm, no case of Plasmodium infection and malaria disease was observed in the 7 HbAS children while in Non-HbAS children, peaks of infection and disease prevalence were respectively observed in October (24.66%) and November (7.14%). For the SMC arm, in group O and Non-group O, Plasmodium infection cases were observed from August to December. Plasmodium infection and malaria disease were more frequently observed in HbAS children in the control arm than in the SMC arm. Further studies are needed to assess factors associated with the asymptomatic carriage of parasites during SMC in Dangassa. NCT04149106.

Children with hemoglobin C or S trait have low serologic responses to a subset of malaria variant surface antigens

Children with hemoglobin AC or AS have decreased susceptibility to clinical malaria. Parasite variant surface antigen (VSA) presentation on the surface of infected erythrocytes is altered in erythrocytes with hemoglobin C (Hb AC) or sickle trait (Hb AS) mutations in vitro. The protective role of incomplete or altered VSA presentation against clinical malaria in individuals with Hb AC or AS is unclear. Using a high-throughput protein microarray, we sought to use serological responses to VSAs as a measure of host exposure to VSAs among Malian children with Hb AC, Hb AS, or wildtype hemoglobin (Hb AA). In uncomplicated malaria, when compared to Hb AA children, Hb AC children had significantly lower serological responses to extracellular Plasmodium falciparum erythrocyte membrane protein-1 (PfEMP1) domains but did not differ in responses to intracellular PfEMP1 domains and other VSAs, including members of the repetitive interspersed family (RIFIN) and subtelomeric variable open reading frame (STEVOR) family. Healthy children with Hb AC and Hb AS genotypes recognized fewer extracellular PfEMP1s compared to children with Hb AA, especially CD36-binding PfEMP1s. These reduced serologic responses may reflect reduced VSA presentation or lower parasite exposure in children with Hb AC or AS and provide insights into mechanisms of protection.

Impact of haemoglobinopathies on asymptomatic Plasmodium falciparum infection and naturally acquired immunity among children in Northern Ghana

BackgroundThe protective effect of certain haemoglobinopathies, such as HbS, HbC, and α-thalassaemia, against severe malaria has long been established; however, there is only limited and equivocal evidence regarding their impact on asymptomatic parasitaemia. Here, we investigated the effect of HbS, HbC, and α-thalassaemia on asymptomatic P. falciparum parasitaemia and acquired immunity among children in Northern Ghana.Materials and methodsA cross-sectional study was conducted among 1,017 healthy children (1-17 years) in 13 malaria-endemic communities in Northern Ghana. The children were screened for structural Hb phenotypes using SickleSCAN, for P. falciparum infection using anti-HRP2 malaria RDT and subsequently confirmed by capillary electrophoresis and PCR, respectively. α-thalassaemia genotyping was done using PCR. Levels of IgG specific for six recombinant malaria antigens (PfCSP, GLURP, MSP3, Pfs230, HB3VAR06, and IT4VAR60) and crude asexual blood-stage antigens were evaluated by ELISA.Results266 out of the 1,017 participants had either HbAC (18%) or HbAS (8.4%), whereas 35% had α‐thalassaemia. Twenty-five percent and 6% HbAC individuals co-inherited heterozygous and homozygous α-thalassaemia respectively. Similarly, 25% and 10.5% of HbAS co-inherited heterozygous and homozygous α-thalassaemia. Asymptomatic parasitaemia rates were 23%, 24%, and 19% in those with HbAA, HbAC and HbAS, respectively. The overall parasite carriage rates in heterozygous (21%) and homozygous α-thalassaemia (25%) individuals were similar to that of individuals without α-thalassaemia (23%). P. falciparum parasite carriage risk was about three times higher among homozygous α-thalassaemia individuals with HbAC (OR = 2.97; 95% CI 0.83-10.62) and heterozygous carriers with HbAS variants (OR = 2.86; 95% CI 0.85-9.60) compared to the wildtype. In HbAS individuals, IgG levels to IT4VAR60 and HB3VAR06 were significantly lower, whereas anti-CSP levels were higher than in HbAA and HbAC.ConclusionsCo-inheritance of HbAS and HbAC with α-thalassaemia increased the risk of asymptomatic parasitaemia, an indication of a negative epistatic effect between these Hb variants. Antibody levels against non-PfEMP1 antigens were slightly higher among HbAS children, but quite similar in all study groups, indicating differences in parasite exposure.

Sickle cell anaemia and severe Plasmodium falciparum malaria: a secondary analysis of the Transfusion and Treatment of African Children Trial (TRACT)

SummaryBackgroundSickle cell anaemia (SCA) has historically been associated with high levels of childhood mortality in Africa. Although malaria has a major contribution to this mortality, to date, the clinical pathology of malaria among children with SCA has been poorly described. We aimed to explore the relationship between SCA and Plasmodium falciparum malaria in further detail by investigating the burden and severity of malaria infections among children recruited with severe anaemia to the TRACT trial of blood transfusion in Africa.MethodsThis study is a post-hoc secondary analysis of the TRACT trial data, conducted after trial completion. TRACT was an open-label, multicentre, factorial, randomised controlled trial enrolling children aged 2 months to 12 years who presented with severe anaemia (haemoglobin <6·0 g/dL) to four hospitals in Africa. This secondary analysis is restricted to Uganda, where the birth prevalence of SCA is approximately 1% and malaria transmission is high. Children were classified as normal (HbAA), heterozygous (HbAS), or homozygous (HbSS; SCA) for the rs334 A→T sickle mutation in HBB following batch-genotyping by PCR at the end of the trial. To avoid confounding from SCA-specific medical interventions, we considered children with an existing diagnosis of SCA (known SCA) separately from those diagnosed at the end of the trial (unknown SCA). The outcomes considered in this secondary analysis were measures of P falciparum parasite burden, features of severe malaria, and mortality at day 28 in malaria-positive children.FindingsBetween Sept 17, 2014, and May 15, 2017, 3944 children with severe anaemia were enrolled into the TRACT trial. 3483 children from Uganda were considered in this secondary analysis. Overall, 1038 (30%) of 3483 Ugandan children had SCA. 1815 (78%) of 2321 children without SCA (HbAA) tested positive for P falciparum malaria, whereas the prevalence was significantly lower in children with SCA (347 [33%] of 1038; p<0·0001). Concentrations of plasma P falciparum histidine-rich protein 2 (PfHRP2), a marker of the total burden of malaria parasites within an individual, were significantly lower in children with either known SCA (median 8 ng/mL; IQR 0–57) or unknown SCA (7 ng/mL; 0–50) than in HbAA children (346 ng/mL; 21–2121; p<0·0001). In contrast to HbAA children, few HbSS children presented with classic features of severe and complicated malaria, but both the frequency and severity of anaemia were higher in HbSS children. We found no evidence for increased mortality at day 28 in those with SCA compared with those without SCA overall (hazard ratios 1·07 [95% CI 0·31–3·76] for known SCA and 0·67 [0·15–2·90] for unknown SCA).InterpretationThe current study suggests that children with SCA are innately protected against classic severe malaria. However, it also shows that even low-level infections can precipitate severe anaemic crises that would likely prove fatal without rapid access to blood transfusion services.FundingUK Medical Research Council, Wellcome, and UK National Institute for Health and Care Research.

Memory CD8+ T cell compartment associated with delayed onset of Plasmodium falciparum infection and better parasite control in sickle-cell trait children.

ObjectivesStudy of individuals with protection from Plasmodium falciparum (Pf) infection and clinical malaria, including individuals affected by the sickle‐cell trait (HbAS), offers the potential to identify cellular targets that could be translated for therapeutic development. We previously reported the first involvement of cellular immunity in HbAS‐associated relative protection and identified a novel subset of memory‐activated NK cells that was enriched in HbAS children and associated with parasite control. We hypothesised that other memory cell subsets might distinguish the baseline profile of HbAS children and children with normal haemoglobin (HbAA).MethodsSubsets of memory T cells and NK cells were analysed by flow cytometry in paired samples collected from HbAS and HbAA children, at baseline and during the first malaria episode of the ensuing transmission season. Correlations between cell frequencies and features of HbAS‐mediated protection from malaria were determined.ResultsHbAS children displayed significantly higher frequency of memory CD8+ T cells at baseline than HbAA children. Baseline frequency of memory CD8+ T cells correlated with features of HbAS‐mediated protection from malaria. Exploration of memory CD8+ T cell subsets revealed that central memory CD8+ T cell frequency was higher in HbAS children than in HbAA children.ConclusionThis study shows that HbAS children develop a larger memory CD8+ T cell compartment than HbAA children, and associates this compartment with better control of subsequent onset of infection and parasite density. Our data suggest that central memory CD8+ T cells may play an important role in the relative protection against malaria experienced by HbAS individuals, and further work to investigate this is warranted.

A novel population of memory-activated natural killer cells associated with low parasitaemia in Plasmodium falciparum-exposed sickle-cell trait children.

ObjectivesThe sickle‐cell trait phenotype is associated with protection from malaria. Multiple molecular mechanisms have been proposed to explain this protection, but the role of the host immune system has been poorly investigated. We hypothesised that cellular immunity to malaria may develop differently in sickle‐cell trait children (HbAS) and children with normal haemoglobin (HbAA) repeatedly exposed to Plasmodium falciparum (Pf).MethodsPaired samples collected prior to the Pf transmission season and during the first malaria episode of the ensuing transmission season from HbAS and HbAA children were analysed by multiplex bead‐based assay and comprehensive multi‐dimensional flow cytometry profiling.ResultsCellular immune profiles were enriched in HbAS relative to HbAA children before the start of the Pf transmission season, with a distinct NK subset. These cells were identified as a novel subset of memory‐activated NK cells characterised by reduced expression of the ecto‐enzyme CD38 as well as co‐expression of high levels of HLA‐DR and CD45RO. The frequency of this NK subset before the transmission season was negatively correlated with parasite density quantified during the first malaria episode of the ensuing transmission season. Functional assessment revealed that these CD38dim CD45RO+ HLA‐DR+ NK cells represent a important source of IFN‐γ.ConclusionOur data suggest that this novel memory‐activated NK cell subset may contribute to an accelerated and enhanced IFN‐γ‐mediated immune response and to control of parasite density in individuals with the sickle‐cell trait. This distinct cellular immune profile may contribute to predispose HbAS children to a relative protection from malaria.

Albumin, copper, manganese and cobalt levels in children suffering from sickle cell anemia at Kasumbalesa, in Democratic Republic of Congo

BackgroundSickle Cell Anemia (SCA) is characterized by high levels of oxidative stress markers and low levels of antioxidant capacity. Antioxidant defence mechanisms against the harmful effects of ROS requires cellular and extracellular enzymes. These enzymes requires micronutrient for complete activity. Information on micronutrients such as manganese, cobalt and copper in SCA population was poorly documented in the literature.MethodsPlasma copper, manganese, cobalt and albumin concentrations determined by atomic absorption spectrophotometry were compared between two groups of children: 76 with SCA (Hb-SS) and 76 without SCA (controls). This study was conducted in the Muhona Hospital of Kasumbalesa, which is situated in a rural and low in resources.ResultsThe mean age was 10.0 years (SD = 5.4) in SCA children and 9.2 years (SD = 4.7) in the control group. The levels of cobalt, manganese, copper and albumin were not different between the two groups (p > 0.05).ConclusionIn our study, albumin, manganese, cobalt and copper values did not differ between SCA children in steady state and Hb-AA children. The lack of differences in plasma elemental concentrations between the two groups in context of increased demands in the SCA group, may represent adequate compensatory intake or elemental dyshomeostasis in the SCA group.

Prevalence of Inherited Hemoglobin Disorders and Relationships with Anemia and Micronutrient Status among Children in Yaoundé and Douala, Cameroon.

Information on the etiology of anemia is necessary to design effective anemia control programs. Our objective was to measure the prevalence of inherited hemoglobin disorders (IHD) in a representative sample of children in urban Cameroon, and examine the relationships between IHD and anemia. In a cluster survey of children 12–59 months of age (n = 291) in Yaoundé and Douala, we assessed hemoglobin (Hb), malaria infection, and plasma indicators of inflammation and micronutrient status. Hb S was detected by HPLC, and α+thalassemia (3.7 kb deletions) by PCR. Anemia (Hb < 110 g/L), inflammation, and malaria were present in 45%, 46%, and 8% of children. A total of 13.7% of children had HbAS, 1.6% had HbSS, and 30.6% and 3.1% had heterozygous and homozygous α+thalassemia. The prevalence of anemia was greater among HbAS compared to HbAA children (60.3 vs. 42.0%, p = 0.038), although mean Hb concentrations did not differ, p = 0.38). Hb and anemia prevalence did not differ among children with or without single gene deletion α+thalassemia. In multi-variable models, anemia was independently predicted by HbAS, HbSS, malaria, iron deficiency (ID; inflammation-adjusted ferritin <12 µg/L), higher C-reactive protein, lower plasma folate, and younger age. Elevated soluble transferrin receptor concentration (>8.3 mg/L) was associated with younger age, malaria, greater mean reticulocyte counts, inflammation, HbSS genotype, and ID. IHD are prevalent but contribute modestly to anemia among children in urban Cameroon.

Sickle cell trait is associated with controlled levels of haem and mild proinflammatory response during acute malaria infection.

SummaryThe controlled induction of haemoxygenase‐1 (HO‐1), an enzyme that catabolizes haem, has been shown to reduce haem, preventing pathologies associated with haem toxicity. The hemoglobin genotype HbAS confers reduced susceptibility to severe complications of malaria by a mechanism that is not well understood. Using a longitudinal approach, we investigated the effect of baseline concentrations of HO‐1 on the accumulation of haem during acute Plasmodium falciparum malaria in HbAS and HbAA genotypes. Plasma concentrations of haem, HO‐1 and cytokines were quantified in venous blood obtained from children (9 months–5 years of age) during malaria infection, and at convalescence (baseline levels). Parasitaemia was determined during malaria infection. In patients with the HbAA genotype, there was a significant elevation in the plasma concentration of haem (P = 0.002), and a consequent increased induction of HO‐1 (P < 0.001) during falciparum malaria compared with levels at convalescence. Contrary to HbAA, plasma concentration of haem did not change in the HbAS genotypical group (P = 0·110), and the induction of HO‐1 was reduced during malaria compared with levels at convalescence (P = 0·006). Higher plasma levels of haem were observed in HbAS compared with HbAA at convalescence (P = 0·010), but this difference did not affect the levels of HO‐1 within each genotype (P = 0·450). Relatively milder proinflammatory responses were observed in HbAS children during malaria infection compared to HbAA children. Our findings suggest that a mechanism of reduced susceptibility to severe malaria pathologies by the HbAS genotype may involve the control of haem, leading to controlled levels of HO‐1 and milder proinflammatory responses during acute malaria.

Effect of red blood cell variants on childhood malaria in Mali: a prospective cohort study

Red blood cell variants protect African children from severe falciparum malaria. However, their individual and interactive effects on mild disease and parasite density, and their modification by age-dependent immunity, are poorly understood. In this study, we address these knowledge gaps in a prospective cohort study of malaria risk and Plasmodium falciparum densities in Malian children. The Kenieroba Innate Defense Study for Malaria (KIDS-Malaria) was a 4-year prospective cohort study of children aged 6 months to 17 years undertaken in Mali between 2008 and 2011. Red blood cell variants were haemoglobin S (HbS), haemoglobin C (HbC), α thalassaemia, ABO blood groups, and glucose-6-phosphate dehydrogenase (G6PD) deficiency encoded by the X-linked A- allele. The primary outcome was malaria incidence, measured as the number of uncomplicated or severe malaria episodes over time. The secondary outcome was parasite density at the time of a malaria episode. We modelled incidence rate ratios with quasi-Poisson regression and we analysed parasite densities using generalised estimating equations. This study is registered with ClinicalTrials.gov, number NCT00669084. Between May 1, 2008, and Dec 29, 2011, we enrolled 1586 children into the study. We successfully typed all five red blood cell variants for 1543 of these children, who therefore constituted the evaluable population and in whom we diagnosed 4091 malaria episodes over 2656 child-years of follow-up. In these 1543 children, red blood cell variants were common, and occurred at the following frequencies: sickle cell trait (HbAS) 220 (14%), HbC heterozygosity (HbAC) 103 (7%), α thalassaemia 438 (28%), type O blood group 621 (40%), and G6PD deficiency 72 (9%) in 767 boys and 158 (20%) in 776 girls. The overall incidence of malaria was 1.54 episodes per child-year of follow-up, ranging from 2.78 episodes per child-year at age 3 years to 0.40 episodes per child-year at age 17 years. The malaria incidence was lower in HbAS children than in HbAA children with normal haemoglobin (adjusted incidence rate ratio [aIRR] 0.66 [95% CI 0.59-0.75], p<0.0001) and lower in G6PD A-/A- homozygous girls than in G6PD A+/A+ girls (0.51 [0.29-0.90], p=0.020), but was higher in HbAC children than in HbAA children (1.15 [1.01-1.32], p=0.039). Parasite density was lower in HbAS children (median 10,550 parasites per μL [IQR 1350-26,250]) than in HbAA children (15,150 parasites per μL [4250-31,050]; p=0.0004). The HbAS-associated reductions in malaria risk and parasite density were greatest in early childhood. The individual and interactive effects of HbAS, HbAC, and G6PD A-/A- genotypes on malaria risk and parasite density define clinical and cellular correlates of protection. Further identification of the molecular mechanisms of these protective effects might uncover new targets for intervention. Intramural Research Program, National Institute of Allergy and Infectious Diseases, National Institutes of Health.

Acute-phase reactants, essential trace elements and some hematological parameters in Nigerian children with steady state sickle cell disorder

Background: Sickle cell disease (SCD) is an inherited chronic hematological disorder, with inflammatory responses arising from different pathways. Information is scarce about the levels of acute-phase reactants and essentials trace elements in HbSS Nigerian children. Understanding of these will further elucidate the pathophysiology of SCD, which may assist in the proper management of this condition in the pediatric population. Aim: To measure the levels of acute-phase reactants (C-reactive protein [CRP], C1q, C4, ferritin and transferrin), trace elements (Fe, Zn, Cu) and some hematological parameters in HbSS children Materials and Methods: A total number of 26 consecutive steady state HbSS children below the age of 5 years was recruited for the study. The same number of HbAA children was recruited as a control. Trace elements were determined with atomic absorption spectrophotometer. C4, C1q and CRP were quantified using immunoplates, and full blood count analysis was done according to standard hematological procedures. Results: There were no significant differences between serum mean levels of Zn, Cu, Ferritin, C4, C1q, albumin and CRP in steady state HbSS children compared to their HbAA counterparts. There was a significant increase in the level of serum iron in steady state HbSS children compared to HbAA children. There was also a significant reduction in the serum level of transferrin in steady state HbSS children compared to HbAA children. There were no significant differences between the white blood cell, red blood cell, mean corpuscular hemoglobin (Hb), mean corpuscular Hb concentration, platelets, lymphocytes, monocytes and neutrophils in steady state HbSS children compared to their HbAA counterparts. However, there was a significant reduction in the Hb concentration and hematocrit value in steady state HbSS children compared to HbAA children. Conclusion: The study observes reduced inflammation in steady state HbSS children below the age of 5 years. It is recommended that consumption of diets or use of iron containing drug by HbSS children be monitored to prevent iron overload.

α+Thalassemia Antagonizes the Malaria-protective Effects of Sickle-Cell Trait

α+Thalassemia Antagonizes the Malaria-protective Effects of Sickle-Cell Trait

Ex-Vivo Cytoadherence Phenotypes of Plasmodium falciparum Strains from Malian Children with Hemoglobins A, S, and C

Sickle hemoglobin (Hb) S and HbC may protect against malaria by reducing the expression of Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1) on the surface of parasitized red blood cells (RBCs), thereby weakening their cytoadherence to microvascular endothelial cells (MVECs) and impairing their activation of MVECs to produce pathological responses. Therefore, we hypothesized that parasites causing malaria in HbAS or HbAC heterozygotes have overcome this protective mechanism by expressing PfEMP1 variants which mediate relatively strong binding to MVECs. To test this hypothesis, we performed 31 cytoadherence comparisons between parasites from HbAA and HbAS (or HbAC) Malian children with malaria. Ring-stage parasites from HbAA and HbAS (or HbAC) children were cultivated to trophozoites, purified, and then inoculated in parallel into the same wildtype uninfected RBCs. After one cycle of invasion and maturation to the trophozoite stage expressing PfEMP1, parasite strains were compared for binding to MVECs. In this assay, there were no significant differences in the binding of parasites from HbAS and HbAC children to MVECs compared to those from HbAA children (HbAS, fold-change = 1.46, 95% CI 0.97–2.19, p = 0.07; HbAC, fold-change = 1.19, 95% CI 0.77–1.84, p = 0.43). These data suggest that in-vitro reductions in cytoadherence by HbS and HbC may not be selecting for expression of high-avidity PfEMP1 variants in vivo. Future studies that identify PfEMP1 domains or amino-acid motifs which are selectively expressed in parasites from HbAS children may provide further insights into the mechanism of malaria protection by the sickle-cell trait.

Comparative haematological parameters of HbAA and HbAS genotype children infected with Plasmodium falciparum malaria in Yemen

BackgroundSickle haemoglobin (HbS) is known to offer considerable protection against falciparum malaria. However, the mechanism of protection is not yet completely understood. In this study, we investigate how the presence of the sickle cell trait affects the haematological profile of AS persons with malaria, in comparison with similarly infected persons with HbAA. This study is based on the hypothesis that the sickle cell trait plays a protective role against malaria.MethodsChildren from an endemic malaria transmission area in Yemen were enrolled in this study. Hematological parameters were estimated using manual methods, the percentage of parasite density on stained thin smear was calculated, haemoglobin genotypes were determined on paper electrophoresis, ferritin was measured using enzyme-linked immunosorbent assay, serum iron and TIBC were assayed using spectrophotometer, transferrin saturation index was calculated by dividing serum iron by TIBC and expressing the result as a percentage. Haematological parameters were compared in HbAA- and HbAS-infected children.ResultsFalciparum malaria parasitaemia was confirmed in the blood smears of 62 children, 44 (55.7%) of AA and 18 (37.5%) AS, so there was higher prevalence in HbAA children (P = 0.047). Parasite density was lower in HbAS- than HbAA-infected children (P = 0.003). Anaemia was prominent in malaria-infected children, with high proportions of moderate and severe forms in HbAA (P = 0.001). The mean levels of haemoglobin, packed cell volume, reticulocyte count, platelets count, lymphocytes, eosinophils, and serum iron were significantly lower while total leukocytes, immature granulocytes, monocytes, erythrocyte sedimentation rate, transferrin saturation, and serum ferritin were significantly higher in HbAA-infected children than HbAS-infected children.ConclusionInfection with Plasmodium falciparum malaria caused more significant haematological alterations of HbAA children than HbAS. This study supports the observation that sickle cell trait seems to be a beneficial genetic factor that resists malaria, since inheriting it protects against significant haematological consequences of malaria.

Epistatic Interactions between Apolipoprotein E and Hemoglobin S Genes in Regulation of Malaria Parasitemia

Apolipoprotein E is a monomeric protein secreted by the liver and responsible for the transport of plasma cholesterol and triglycerides. The APOE gene encodes 3 isoforms Ɛ4, Ɛ3 and Ɛ2 with APOE Ɛ4 associated with higher plasma cholesterol levels and increased pathogenesis in several infectious diseases (HIV, HSV). Given that cholesterol is an important nutrient for malaria parasites, we examined whether APOE Ɛ4 was a risk factor for Plasmodium infection, in terms of prevalence or parasite density. A cross sectional survey was performed in 508 children aged 1 to 12 years in Gabon during the wet season. Children were screened for Plasmodium spp. infection, APOE and hemoglobin S (HbS) polymorphisms. Median parasite densities were significantly higher in APOE Ɛ4 children for Plasmodium spp. densities compared to non-APOE Ɛ4 children. When stratified for HbS polymorphisms, median Plasmodium spp. densities were significantly higher in HbAA children if they had an APOE Ɛ4 allele compared to those without an APOE Ɛ4 allele. When considering non-APOE Ɛ4 children, there was no quantitative reduction of Plasmodium spp. parasite densities for HbAS compared to HbAA phenotypes. No influence of APOE Ɛ4 on successful Plasmodium liver cell invasion was detected by multiplicity of infection. These results show that the APOE Ɛ4 allele is associated with higher median malaria parasite densities in children likely due to the importance of cholesterol availability to parasite growth and replication. Results suggest an epistatic interaction between APOE and HbS genes such that sickle cell trait only had an effect on parasite density in APOE Ɛ4 children. This suggests a linked pathway of regulation of parasite density involving expression of these genes. These findings have significance for understanding host determinants of regulation of malaria parasite density, the design of clinical trials as well as studies of co-infection with Plasmodium and other pathogens.

Effects of Age, Hemoglobin Type and Parasite Strain on IgG Recognition of Plasmodium falciparum–Infected Erythrocytes in Malian Children

BackgroundNaturally-acquired antibody responses to antigens on the surface of Plasmodium falciparum-infected red blood cells (iRBCs) have been implicated in antimalarial immunity. To profile the development of this immunity, we have been studying a cohort of Malian children living in an area with intense seasonal malaria transmission.Methodology/Principal FindingsWe collected plasma from a sub-cohort of 176 Malian children aged 3-11 years, before (May) and after (December) the 2009 transmission season. To measure the effect of hemoglobin (Hb) type on antibody responses, we enrolled age-matched HbAA, HbAS and HbAC children. To quantify antibody recognition of iRBCs, we designed a high-throughput flow cytometry assay to rapidly test numerous plasma samples against multiple parasite strains. We evaluated antibody reactivity of each plasma sample to 3 laboratory-adapted parasite lines (FCR3, D10, PC26) and 4 short-term-cultured parasite isolates (2 Malian and 2 Cambodian). 97% of children recognized ≥1 parasite strain and the proportion of IgG responders increased significantly during the transmission season for most parasite strains. Both strain-specific and strain-transcending IgG responses were detected, and varied by age, Hb type and parasite strain. In addition, the breadth of IgG responses to parasite strains increased with age in HbAA, but not in HbAS or HbAC, children.Conclusions/SignificanceOur assay detects both strain-specific and strain-transcending IgG responses to iRBCs. The magnitude and breadth of these responses varied not only by age, but also by Hb type and parasite strain used. These findings indicate that studies of acquired humoral immunity should account for Hb type and test large numbers of diverse parasite strains.

Relationship between Malaria Incidence and IgG Levels to Plasmodium falciparum Merozoite Antigens in Malian Children: Impact of Hemoglobins S and C

Heterozygous hemoglobin (Hb) AS (sickle-cell trait) and HbAC are hypothesized to protect against Plasmodium falciparum malaria in part by enhancing naturally-acquired immunity to this disease. To investigate this hypothesis, we compared antibody levels to four merozoite antigens from the P. falciparum 3D7 clone (apical membrane antigen 1, AMA1-3D7; merozoite surface protein 1, MSP1-3D7; 175 kDa erythrocyte-binding antigen, EBA175-3D7; and merozoite surface protein 2, MSP2-3D7) in a cohort of 103 HbAA, 73 HbAS and 30 HbAC children aged 3 to 11 years in a malaria-endemic area of Mali. In the 2009 transmission season we found that HbAS, but not HbAC, significantly reduced the risk of malaria compared to HbAA. IgG levels to MSP1 and MSP2 at the start of this transmission season inversely correlated with malaria incidence after adjusting for age and Hb type. However, HbAS children had significantly lower IgG levels to EBA175 and MSP2 compared to HbAA children. On the other hand, HbAC children had similar IgG levels to all four antigens. The parasite growth-inhibitory activity of purified IgG samples did not differ significantly by Hb type. Changes in antigen-specific IgG levels during the 2009 transmission and 2010 dry seasons also did not differ by Hb type, and none of these IgG levels dropped significantly during the dry season. These data suggest that sickle-cell trait does not reduce the risk of malaria by enhancing the acquisition of IgG responses to merozoite antigens.

Increased IgE level in Nigerian sickle cell disease children: Implication for severity of allergic reactions

Background: Susceptibility of sickle cell disease subjects to various infectious agents is on the increase, but information relating SCD to allergic condition is scarce. Hence, assessment of immunoglobulin status in SCD children may provide useful information to improve management of SCD children. Objectives: To determine the levels of immunoglobulin classes (IgG, IgA, IgM, and IgE) in Nigerian HbSS children below 5 years of age compared with sex- and age-matched HbAA children. Materials and Methods: Blood samples were collected from a total of 45 children less than 5 years of age who were recruited into the study as follows: 26 HbSS and 19 HbAA subjects for the estimation of serum immunoglobulin levels using enzyme-linked immunosorbent assay (ELISA) techniques and determination of genotype using electrophoresis technique. Results: IgG concentration was nonsignificantly higher (P = 0.997) in HbSS children (1022.56 ± 148.97 ng/ml) compared to HbAA children (933.68 ± 106.10 ng/ml). IgA (P = 0.906) and IgM (P = 0.986) concentrations were nonsignificantly lower in HbSS children (255.07 ± 133.71 ng/ml) compared to HbAA children. IgE was significantly higher (P = 0.000***) in HbSS (108.67 ± 69.22 IU/ml) compared to HbAA (24.51 ± 17.58 IU/ml) children.Conclusion: SCD children in steady state have adequate levels of Ig classes. Non-specific elevation of IgE levels may be a factor of inflammatory response in SCD children, and this may be proposed for reduced allergic reaction among SCD children.

Co‐inheritance of α+‐thalassaemia and sickle trait results in specific effects on haematological parameters

Both the sickle cell trait (HbAS) and alpha(+)-thalassaemia are common in many tropical areas. While their individual haematological effects are well described, few studies describe their effects when inherited together. We present data from the Kenyan coast, which suggest that HbAS and alpha(+)-thalassaemia may interact to produce specific effects on haematological parameters. Overall, the difference in Hb concentrations between non-thalassaemics (alphaalpha/alphaalpha) and alpha(+)-thalassaemia homozygotes (-alpha/-alpha) was greater in non-HbAS (HbAA) (0.63 g/dl) than in HbAS children (0.25 g/dl). HbAS also ameliorated both the reduced mean cell volume and mean cell haemoglobin normally associated with the -alpha/-alpha genotype. Potential mechanisms and implications are discussed.

Distribution of IgG subclass antibodies specific for Plasmodium falciparum glutamate-rich-protein molecule in sickle cell trait children with asymptomatic infections

Polymorphism in the β-globin gene (hemoglobin S) has been associated with protection against severe forms of malaria. In a cross-sectional study, 180 young Gabonese children with and without sickle cell trait and harboring asymptomatic Plasmodium falciparum infections, were assessed for the responses to recombinant protein containing the conserved region of glutamate-rich protein (GLURP). We reported increased age-dependence of antibody prevalence and levels of total IgG ( p < 0.0001), IgG1 ( p = 0.009), and IgG3 ( p < 0.03) antibodies to GLURP with a cut-off at 5 years of age. Whatever the hemoglobin type, cytophilic antibodies (IgG1 and IgG3) were prevalent, but GLURP-specific IgG4 antibodies were detected at significantly ( p < 0.05) lower levels in HbAS children. We showed that the distribution of non-cytophilic IgG antibodies differs according to the hemoglobin type and to the malaria antigens tested. This may have possible implication for the clearance of malaria parasites and for protection against severe malaria.