Associations between Functional Variations in Fc'yriiia and TLR 9 and Susceptibility to Severe Malaria Anemia in Cidldren below 3 Years in Western Kenya

Abstract

Plasmodium Jalciparum malaria in holoendemic transmission areas is a major cause of childhood morbidity and mortality. The greatest burden of this morbidity and mortality is as a consequence of severe malaria anemia (SMA) in children aged below 5 years. Understanding the immunogenetic basis of naturally acquired immunity to P. Jalciparum infection and SMA would aid in designing a rationally based malaria vaccine. Variants withinthe FcyR mediate immunity through engagement of immunoglobulin (Ig) G and other immune mediators such as interferon gamma (IFN-y), resulting into erythrophagocytosis and production of inflammatory cytokines. Toll like receptors (TLRs) trigger transcription of pro-inflammatory cytokines and induction of adaptive immune responses. The two receptors collectively through both innate and adaptive immune responses may determine malaria disease pathogenesis. The role of these single nucleotide polymorphisms (SNPs) in conditioning susceptibility to SMA has however not been investigated in P. Jalciparum holoendemic transmission areas such as in western Kenya. In this cross-sectional study, the association between FcyRIIIA -176FN, TLR-9 (-1237T/C) genotypes/SNPs combinations, SMA.(Hb<6.0g/dL) and circulating levels of IFN-y were investigated in children (n=301) withJalciparum malaria from Siaya in western Kenya. Haematological and parasitological parameters were determined in all study participants. FcyRIIIA -176FN and TLR-9 (1237T/C) genotypes were determined using TaqMan 5' allelic discrimination assay. Circulating levels of IFN-y were determined using Cytokine 25-plex Ab Bead Kit. All data analysis was performed using SPSS (Version 19.0). Multivariate logistic regression analysis (controlling for confounding factors) revealed that children with FcyRIIIA -176V/ TLR-9 1237C SNP combinations had a 64% reduced odds of developing SMA (OR=0.36, 95%CI 0.20-0.64; p=O.OOI) while carriers of FcyRIIIA -176V/ TLR-9 -1237T SNP combination were more susceptible to SMA (OR=2.04, 95%CI, 1.19-3.50;p=O.009). Children with SMA had higher circulating IFN-y levels compared to non-SMA (p=0.008). Consistently, the FcyRIllA -176VITLR-9 -1237T (VT) carriers had higher levels of circulating IFN-y (p=0.01l) relative to non-carriers supporting the observation that higher IFN-y levels are associated with SMA. These results demonstrate that FcyRIIIA-176FN and TLR-9 (1237T/C) variants condition susceptibility to SMA and functional changes in circulating IFNy levels. This study therefore provides critical information that can be utilized in the design of an effective malaria vaccine since the intermediate step is to understand the natural immune response in a population before designing a long-lasting effective vaccine against the disease.