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The repertoire of G6PD deficiency molecular markers is extensive and displays considerable spatial heterogeneity. G6PD is essential for the protection of cells against oxidative damage and G6PD deficiency is common in malaria endemic areas due to its association with protection of some manifestations of severe malaria. Examples of such mutations include polymorphisms in the X-linked glucose-6-phosphate dehydrogenase (G6PD) gene and in the HBB gene. Plasmodium parasites have co-evolved with human hosts and exert a considerable evolutionary pressure on mutations that confer a degree of protection against malaria. The screening is high throughput, adaptable to inclusion of multiple targets of interest and easily standardized. The bead based assay compares well to alternative measures of genotyping and phenotyping for G6PD. Flow cytometry data available for n = 61 samples further supported the concordance between % G6PD normal/deficient cells and genotype. Seventy-five samples from Burkina Faso (n = 75/78, 96.2%) and 58 samples from The Gambia (n = 58/61, 95.1%) had a G6PD and a HBB genotype successfully assigned by the bead-based assay. Genotyping data was compared to phenotypical data generated by flow cytometry and to established genotyping methods. The products were hybridized to magnetic beads and the median fluorescence intensity (MFI) was read on MAGPIX ® (Luminex corp.).
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Human DNA from whole blood samples was amplified in a novel, multiplex PCR reaction and extended with SNP-specific probes in an allele specific primer extension (ASPE) to simultaneously detect four epidemiologically important human markers including G6PD SNPs (G202A and A376G) and common haemoglobin mutations (HbS and HbC). Current methodologies in epidemiological studies commonly rely on individual target amplification and visualization here a method is presented to simultaneously detect the polymorphisms and that can be expanded to include other single nucleotide polymorphisms (SNPs) of interest. Measuring the prevalence of these inherited blood disorders in affected populations can improve understanding of disease epidemiology. All are associated to some degree with protection against clinical malaria whilst additionally G6PDd is associated with haemolysis following treatment with 8-aminoquinolines. Glucose-6-phosphate dehydrogenase deficiency (G6PDd), haemoglobin C (HbC) and S (HbS) are inherited blood disorders (IBD) common in populations in malaria endemic areas.