Immunological and Genetic Correlates of Immunity to Plasmodium Falciparum Malaria

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Malaria is a major cause of morbidity and mortality in children under five years old and pregnant women in sub-Saharan Africa. Previous studies have shown that immunity to malaria is mediated by host immunological and genetic factors and parasite genetics. In this study which sought to identify immunological and genetics correlates of immunity against Plasmodium falciparum malaria, data from two consecutive malaria transmission community-based cohort studies and a hospital-based malaria case-control were analyzed. The Afro-immuno assay (AIA) enzyme linked immuno-sorbent assay (ELISA) protocol was used to measure baseline isotype IgG and IgM and IgG1-4 subclasses levels to eight (8) malaria antigens GLURP-R0, GLURP-R2, MSP1 hybrid, EBA-175, AMA1-FVO, MSP3-FVO, LR146 and AS202.11. FcγIIA-131H/R, FcγIIB-232I/T, FcγIIIA-158V/F and FcγIIIB-NA1/NA2/SH together with IgG3 hinge region length polymorphisms were genotyped using various polymerase chain reaction (PCR) and sequencing techniques. There was a strong correlation between IgG1 and 3 for MSP3-FVO; IgG2 and 3 for GLURP-R2; IgG1 and 2 for AMA1-FVO and IgG1 and 2 for EBA-175 with age (0.413 ≤ r ≤ 0.202, p < 0.00001). In an age adjusted univariate analysis to investigate the association between the antibodies and clinical malaria, IgG1 to MSP3 FVO (IRR=0.83 [95% CI, 0.64, 1.06, p= 0.13]), IgG3 to GLURP R0 (IRR= 0.89 [95% CI, 0.77, 1.03, p=0.12], IgG3 to EBA-175 (IRR=0.88 [95% CI, 0.73, 1.05, p= 0.14]) and IgG4 to MSP3 FVO (IRR=0.81 [95% CI, 0.65, 1.02, p= 0.08]) showed some association with reduced risk to clinical malaria. In a final analysis using a model that comprises, age and the immunological variables with trends 23 suggestive of protection, only IgG4 to MSP3 FVO (IRR=0.85 [95% CI, 0.66, 1.08, p= 0.19]) and IgG3 to GLURP R0 (IRR= 0.92 [95% CI, 0.79, 1.07, p=0.29] showed persistent trends towards association with protection though statistically insignificant. The FcγRIIIB-NA1/NA1 genotype was significantly associated with reduced risk to uncomplicated malaria (p=0.007) and severe malaria (p=0.0002) while the FcγRIIIB-NA2/NA2 genotype was associated with susceptibility in both cases. There was an over-representation of the heterozygous long-medium (LM) (46.0%) genotype among individuals apparently protected against clinical malaria while the homozygous medium-medium (MM) (13.9%) was under-represented. For all the antigens, the MM and the homozygous long-long (LL) genotypes were associated with the highest and the lowest IgG3 levels respectively. Trends towards protection approaching significance observed for IgG3 to GLURP-R0 might be due to the cytophilic nature of IgG3 enabling effective parasite clearance through ADCI while IgG4 to MSP3-FVO might probably be a surrogate marker to some specific cytokine response critical for protection against malaria. The association of the FcγRIIIB-NA1/NA1 genotype with protection against and the FcγRIIIB-NA2/NA2 with susceptibility to severe and uncomplicated malaria in this study is consistent with the observation that NA1 has a higher affinity for both IgG1 and IgG3 than the NA2. The over-representation of LM among individuals with reduced risk to clinical malaria might be explained by the fact that such individuals benefit from both an efficient L-allele and a perhaps not so efficient but high IgG3 titre inducing M-allele. Findings from this study support the development of a multivalent vaccine from GLURP-R0 and MSP3-FVO. FcγRIIIB-NA1/NA1 and IgG3 hinge region 24 length polymorphisms have been implicated in this study as potential confounders to be adequately corrected for in malaria vaccine trials and pre-clinical studies to enable accurate interpretation of data.
A Thesis Submitted to the Department of Biochemistry and Biotechnology, Kwame Nkrumah University of Science and Technology, Kumasi in Partial Fulfilment of the Requirements for the Degree of Master of Philosophy (M.Phil.) in Biochemistry.