Association between Interferon-gamma Promoter -183 (G/T), -1616 (A/G) and Intronic +2200 (A/G) Variants and Malarial Disease Outcomes in Children Exposed to Plasmodium falciparum

Evans Ouma Raballah Okoth


Infection with Plasmodium falciparum, the most common human plasmodial parasite causes significant global morbidity and mortality. Interferon-gamma (IFN-) is a type-1 cytokine with T helper (Th) 1-biased immune responses for intracellular pathogen control. Elevated IFN- levels contribute to protective immunity against clinical and cerebral malaria. High IFN- responses are associated with reduced asexual parasite multiplication rates following a primary infection with P. falciparum. Previous studies have associated IFN- single nucleotide polymorphisms (SNPs) with susceptibility to asthma, tuberculosis and functional changes in circulating IFN- levels. However, the role of IFN- polymorphisms in conditioning malarial disease outcomes in children residing in P. falciparum holoendemic transmission areas remains unexplored. The current study, using a hospital-based cross-sectional design, investigated the functional associations between IFN- genotypic and haplotypic promoter [-183G/T and -1616A/G] and intronic [+2200A/G] variation in conditioning malarial disease outcomes [(severe malarial anaemia (SMA; Haemoglobin (Hb)<6.0 g/dL), malarial anaemia (MA; Hb<8.0 g/dL and high-density parasitaemia (HDP; ≥10,000 parasites/μL)] in infants and young children (age 3-36 months, n=574) enrolled at Siaya District Hospital, western Kenya. Blood samples (3mL) for malaria diagnosis and complete haematological measurements were obtained upon consent from parent/guardian. In addition, plasma was separated and frozen at -80°C until determination of circulating IFN- levels. DNA was extracted from blood spotted on filter paper using Chelex method. Genotyping of the IFN- SNPs was performed using a Taqman® 5′ allelic discrimination Assay-By-Design method according to manufacturer’s instructions and by PCR followed by restriction fragment length polymorphism (RFLP), IFN- levels were measured as part of human325-cytokine plex assay. Multivariate logistic regression models controlling for the confounders revealed that GT heterozygosity at the IFN- -183G/T variants showed a trend of reduced risk of developing SMA (OR, 0.603; 95% CI, 0.286-1.271; P=0.183) relative to the wild type (GG homozygotes). Additionally, the IFN- -1616A/G variants demonstrated that individuals with GG had a two-fold increased risk of developing MA (OR, 2.306; 95% CI, 1.141-4.660; P=0.020) relative to the homozygous A (wild type). The IFN- +2200A/G variants were not associated with SMA, though heterozygous individuals (AG) demonstrated a tendency towards reduced risk of developing MA (OR, 0.597; 95% CI, 0.331-1.076; P=0.086). In addition, the GAA haplotype tended to reduce risk of developing MA (OR, 0.635; 95% CI, 0.391-1.030; P=0.066), while the GAG haplotype tended towards increased risk of developing MA (OR, 1.794; 95% CI, 0.957-3.364; P=0.068). However, carriage of these genotypes and haplotypes did not influence the circulating levels of IFN-. The results demonstrate that genetic variation in the three IFN- genotypes investigated in this study may be important in regulating MA outcomes in paediatric P. falciparum infection. These findings provide an enhanced understanding of how variation in IFN- conditions malaria pathogenesis and the important role that IFN- plays in children with falciparum malaria. From these results it is recommended that future work should focus on comprehensive longitudinal studies to delineate the role of IFN- variants in modulating malarial outcomes in this population.


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