Low Antibody Levels to Pregnancy-specific Malaria Antigens and Heightened Cytokine Responses Associated With Severe Malaria in Pregnancy

BACKGROUND:  Pregnant women living in unstable malaria transmission settings may develop severe malaria (SM). The pathogenesis of SM in pregnancy is poorly understood. METHODS:  To determine whether SM in pregnancy is associated with lower malarial antibody responses and higher cytokine responses, plasma samples were collected from 121 Sudanese pregnant women of whom 39 were diagnosed with SM. Antibodies to pregnancy-specific and non-pregnancy-specific Plasmodium falciparum variant surface antigens (VSA) and concentrations of cytokines TNF, IFNγ, IL-1β, IL-6, IL-8 and IL-10 were measured. RESULTS:  Pregnant women with SM demonstrated significantly lower antibody levels to pregnancy-specific VSA (P = .020) and higher plasma IFNγ (P = .020), IL-10 (P = .0002) and IL-6 levels (P < .0001) than uninfected pregnant women. Concentrations of inflammatory cytokines IL-1β (P = .001), IL-6 (P = .004) and IL-8 (P = .020) were inversely correlated with antibodies to VAR2CSA-DBL5 in pregnant women with SM. Lower haemoglobin levels and higher parasite densities were associated with lack of pregnancy-specific antibodies (P = .028) and higher levels of inflammatory cytokines, in particular IL-6 and IL-8. CONCLUSIONS:  Pregnant women with SM lack pregnancy-specific malaria immunity, and this correlates with heightened inflammatory cytokine concentrations, low haemoglobin levels and high parasite density, suggesting that failure of antibody to control parasitaemia may contribute to SM pathogenesis

Low Antibody Levels to Pregnancy-specific Malaria Antigens and Heightened Cytokine Responses Associated With Severe Malaria in Pregnancy

BACKGROUND:  Pregnant women living in unstable malaria transmission settings may develop severe malaria (SM). The pathogenesis of SM in pregnancy is poorly understood. METHODS:  To determine whether SM in pregnancy is associated with lower malarial antibody responses and higher cytokine responses, plasma samples were collected from 121 Sudanese pregnant women of whom 39 were diagnosed with SM. Antibodies to pregnancy-specific and non-pregnancy-specific Plasmodium falciparum variant surface antigens (VSA) and concentrations of cytokines TNF, IFNγ, IL-1β, IL-6, IL-8 and IL-10 were measured. RESULTS:  Pregnant women with SM demonstrated significantly lower antibody levels to pregnancy-specific VSA (P = .020) and higher plasma IFNγ (P = .020), IL-10 (P = .0002) and IL-6 levels (P < .0001) than uninfected pregnant women. Concentrations of inflammatory cytokines IL-1β (P = .001), IL-6 (P = .004) and IL-8 (P = .020) were inversely correlated with antibodies to VAR2CSA-DBL5 in pregnant women with SM. Lower haemoglobin levels and higher parasite densities were associated with lack of pregnancy-specific antibodies (P = .028) and higher levels of inflammatory cytokines, in particular IL-6 and IL-8. CONCLUSIONS:  Pregnant women with SM lack pregnancy-specific malaria immunity, and this correlates with heightened inflammatory cytokine concentrations, low haemoglobin levels and high parasite density, suggesting that failure of antibody to control parasitaemia may contribute to SM pathogenesis

Effect of nutrient supplementation on the acquisition of humoral immunity to Plasmodium falciparum in young Malawian children

BACKGROUND: There is evidence that suggests that undernutrition has a detrimental effect on malarial immunity in children. The aim of the study was to discover whether nutrient supplementation improved development of malarial antibody immunity in children up to 18 months of age. METHODS: The study was conducted with a subset of 432 Malawian children from a randomized controlled trial of nutritional supplements. The arms included pre- and postnatal small-quantity lipid-based nutrient supplements for both mother and child; prenatal supplementation with iron and folic acid; and pre- and postnatal supplementation with multiple micronutrients. Paired plasma samples were collected at 6 and 18 months of age. The levels of antibodies against merozoite surface protein 1 (MSP1 19kD) and MSP2, erythrocyte binding antigen 175 (EBA175), reticulocyte binding protein homologue 2A (Rh2A9), schizont extract and variant antigens expressed on the surface of infected erythrocytes were measured. RESULTS: At 18 months of age, 5.4% of children were parasitaemic by microscopy and 49.1% were anaemic. Antibodies to the tested merozoite antigens and schizont extract increased between 6 and 18 months and this increase was statistically significant for MSP1, MSP2 and EBA175 (p < 0.0001) whereas IgG to variant surface antigens decreased with increasing age (p < 0.0001). However, the supplementation type did not have any impact on the prevalence or levels of antibodies at either 6 or 18 months of age to any of the tested malaria antigens in either univariate analysis or multivariate analysis after adjusting for covariates. CONCLUSIONS: Pre- and postnatal lipid-based nutrient supplementation did not alter malaria antibody acquisition during infancy, compared to prenatal supplementation with iron and folic acid or pre- and postnatal supplementation with multiple micronutrients. Trail registeration Clinicaltrials.gov registration number NCT01239693

Association between malaria immunity and pregnancy outcomes among Malawian pregnant women receiving nutrient supplementation

BACKGROUND: Malaria antibody responses measured at delivery have been associated with protection from maternal anaemia and low birth weight deliveries. Whether malarial antibodies present in the first half of pregnancy may protect from these or other poor birth outcomes is unclear. To determine whether malaria antibodies in the first half of pregnancy predict pregnancy outcomes, antibodies were measured to a range of merozoite antigens and to antigens expressed on the surface of parasitized red blood cells (pRBCs) in plasma samples collected at 14-20 weeks of gestation from Malawian women. The latter antibodies were measured as total IgG to pRBCs, and antibodies promoting opsonic phagocytosis of pRBCs. Associations between antibodies and maternal haemoglobin in late pregnancy or newborn size were investigated, after adjusting for potential covariates. RESULTS: Antibodies to pRBC surface antigens were associated with higher haemoglobin concentration at 36 weeks. Total IgG to pRBCs was associated with 0.4 g/l [(95% confidence interval (0.04, 0.8)] increase in haemoglobin, and opsonizing antibody with 0.5 (0.05, 0.9) increase in haemoglobin for each 10% increase in antibody. These antibodies were not associated with birthweight, placental malaria, or newborn anthropometrics. Antibodies to merozoite antigens and non-placental-binding IEs were not associated with decreased risk of any of these outcomes. In some instances, they were negatively associated with outcomes of interest. CONCLUSION: Antibodies to placental-binding infected erythrocytes may be associated with higher haemoglobin levels in pregnancy, whereas antibodies to other malaria antigens may instead be markers of malaria exposure. Trial registration clinicaltrials.gov NCT01239693. Registered Nov 10, 2010

The impact of lipid-based nutrient supplementation on anti-malarial antibodies in pregnant women in a randomized controlled trial

BACKGROUND: Malaria and undernutrition frequently coexist, especially in pregnant women and young children. Nutrient supplementation of these vulnerable groups might reduce their susceptibility to malaria by improving immunity. METHODS: Antibody immunity to antigens expressed by a placental-binding parasite isolate, a non-placental binding parasite isolate, merozoites and schizonts at enrolment (before 20 gestation weeks) and at 36 gestation weeks were measured in 1,009 Malawian pregnant women receiving a daily lipid-based nutrient supplement, multiple micronutrients or iron and folic acid, who were participants in a randomized clinical trial assessing the effects of nutrient supplementation on pregnancy outcomes and child development (registration ID: NCT01239693). RESULTS: Antibodies to placental-binding isolates significantly increased while antibodies to most merozoite antigens declined over pregnancy. Overall, after adjustment for covariates, the type of supplementation did not influence antibody levels at 36 gestation weeks or the rate of change in antibody levels from enrolment to 36 weeks. A negative association between maternal body mass index and opsonizing antibodies to placental-binding antigens (coefficient (95% CI) -1.04 (-1.84, -0.24), was observed. Similarly, women with higher socioeconomic status had significantly lower IgG and opsonizing antibodies to placental-binding antigens. Neither of these associations was significantly influenced by the supplementation type. CONCLUSIONS: In the current cohort nutrient supplementation did not affect anti-malarial antibody responses, but poor and undernourished mothers should be a priority group in future trials

Effect of nutrient supplementation on the acquisition of humoral immunity to Plasmodium falciparum in young Malawian children

Abstract Background There is evidence that suggests that undernutrition has a detrimental effect on malarial immunity in children. The aim of the study was to discover whether nutrient supplementation improved development of malarial antibody immunity in children up to 18 months of age. Methods The study was conducted with a subset of 432 Malawian children from a randomized controlled trial of nutritional supplements. The arms included pre- and postnatal small-quantity lipid-based nutrient supplements for both mother and child; prenatal supplementation with iron and folic acid; and pre- and postnatal supplementation with multiple micronutrients. Paired plasma samples were collected at 6 and 18 months of age. The levels of antibodies against merozoite surface protein 1 (MSP1 19kD) and MSP2, erythrocyte binding antigen 175 (EBA175), reticulocyte binding protein homologue 2A (Rh2A9), schizont extract and variant antigens expressed on the surface of infected erythrocytes were measured. Results At 18 months of age, 5.4% of children were parasitaemic by microscopy and 49.1% were anaemic. Antibodies to the tested merozoite antigens and schizont extract increased between 6 and 18 months and this increase was statistically significant for MSP1, MSP2 and EBA175 (p < 0.0001) whereas IgG to variant surface antigens decreased with increasing age (p < 0.0001). However, the supplementation type did not have any impact on the prevalence or levels of antibodies at either 6 or 18 months of age to any of the tested malaria antigens in either univariate analysis or multivariate analysis after adjusting for covariates. Conclusions Pre- and postnatal lipid-based nutrient supplementation did not alter malaria antibody acquisition during infancy, compared to prenatal supplementation with iron and folic acid or pre- and postnatal supplementation with multiple micronutrients. Trail registeration Clinicaltrials.gov registration number NCT0123969

CD14(hi)CD16+ monocytes phagocytose antibody-opsonised Plasmodium falciparum infected erythrocytes more efficiently than other monocyte subsets, and require CD16 and complement to do so

Background With more than 600,000 deaths from malaria, mainly of children under five years old and caused by infection with Plasmodium falciparum, comes an urgent need for an effective anti-malaria vaccine. Limited details on the mechanisms of protective immunity are a barrier to vaccine development. Antibodies play an important role in immunity to malaria and monocytes are key effectors in antibody-mediated protection by phagocytosing antibody-opsonised infected erythrocytes (IE). Eliciting antibodies that enhance phagocytosis of IE is therefore an important potential component of an effective vaccine, requiring robust assays to determine the ability of elicited antibodies to stimulate this in vivo. The mechanisms by which monocytes ingest IE and the nature of the monocytes which do so are unknown. Methods Purified trophozoite-stage P. falciparum IE were stained with ethidium bromide, opsonised with anti-erythrocyte antibodies and incubated with fresh whole blood. Phagocytosis of IE and TNF production by individual monocyte subsets was measured by flow cytometry. Ingestion of IE was confirmed by imaging flow cytometry. Results CD14hiCD16+ monocytes phagocytosed antibody-opsonised IE and produced TNF more efficiently than CD14hiCD16- and CD14loCD16+ monocytes. Blocking experiments showed that Fcγ receptor IIIa (CD16) but not Fcγ receptor IIa (CD32a) or Fcγ receptor I (CD64) was necessary for phagocytosis. CD14hiCD16+ monocytes ingested antibody-opsonised IE when peripheral blood mononuclear cells were reconstituted with autologous serum but not heat-inactivated autologous serum. Antibody-opsonised IE were rapidly opsonised with complement component C3 in serum (t1/2 = 2-3 minutes) and phagocytosis of antibody-opsonised IE was inhibited in a dose-dependent manner by an inhibitor of C3 activation, compstatin. Compared to other monocyte subsets, CD14hiCD16+ monocytes expressed the highest levels of complement receptor 4 (CD11c) and activated complement receptor 3 (CD11b) subunits. Conclusions We show a special role for CD14hiCD16+ monocytes in phagocytosing opsonised P. falciparum IE and production of TNF. While ingestion was mediated by Fcγ receptor IIIa, this receptor was not sufficient to allow phagocytosis; despite opsonisation with antibody, phagocytosis of IE also required complement opsonisation. Assays which measure the ability of vaccines to elicit a protective antibody response to P. falciparum should consider their ability to promote phagocytosis and fix complement