Molecular Analysis of Glucose-6-Phosphate Dehydrogenase Gene Mutations in Bangladeshi Individuals

Abstract

Glucose-6-phosphate dehydrogenase (G6PD) deficiency is a common X-linked human enzyme defect of red blood cells (RBCs). Individuals with this gene defect appear normal until exposed to oxidative stress which induces hemolysis. Consumption of certain foods such as fava beans, legumes; infection with bacteria or virus; and use of certain drugs such as primaquine, sulfa drugs etc. may result in lysis of RBCs in G6PD deficient individuals. The genetic defect that causes G6PD deficiency has been identified mostly as single base missense mutations. One hundred and sixty G6PD gene mutations, which lead to amino acid substitutions, have been described worldwide. The purpose of this study was to detect G6PD gene mutations in hospital-based settings in the local population of Dhaka city, Bangladesh. Qualitative fluorescent spot test and quantitative enzyme activity measurement using RANDOX G6PDH kit were performed for analysis of blood specimens and detection of G6PD-deficient participants. For G6PD-deficient samples, PCR was done with six sets of primers specific for G6PD gene. Automated Sanger sequencing of the PCR products was performed to identify the mutations in the gene. Based on fluorescence spot test and quantitative enzyme assay followed by G6PD gene sequencing, 12 specimens (11 males and one female) among 121 clinically suspected patient-specimens were found to be deficient, suggesting a frequency of 9.9% G6PD deficiency. Sequencing of the G6PD-deficient samples revealed c.C131G substitution (exon-3: Ala44Gly) in six samples, c.G487A substitution (exon-6:Gly163Ser) in five samples and c.G949A substitution (exon-9: Glu317Lys) of coding sequence in one sample. These mutations either affect NADP binding or disrupt protein structure. From the study it appears that Ala44Gly and Gly163Ser are the most common G6PD mutations in Dhaka, Bangladesh. This is the first study of G6PD mutations in Bangladesh.

Prevalence and Genetic Characterization of Glucose-6-Phosphate Dehydrogenase Deficiency in Anemic Subjects from Uttar Pradesh, India

“…10 This variant is due to a substitution of (C!T) nucleotide at 563, changing serine to phenylalanine at amino acid 188. 8 Similar to other reports, in our study the Mediterranean mutation was the most common.…”

“…7 As per available reports, the three most commonly deficient variants are the G6PD Mediterranean mutation (563C!T), Kerala-Kalyan variant (949G!A), and Orissa variant (131C!G), which are common in the Indian population. 8,9 The diagnosis of G6PD deficiency can be done by a qualitative or a quantitative method. The qualitative methods such as the fluorescent spot test (FST) and dichlorophenol-indolphenol (DCIP) decolorization method are the most preferred ones and are cost-effective methods used for mass screening of suspected G6PD-deficient patients.…”

High resolution melting curve analysis enables rapid and reliable detection of G6PD variants in heterozygous females

“…To diagnose how many of the suspected participants had an actual G6PD deficiency, quantitative enzyme assay was performed using RBC specimens of each participant. Among 63 participants, only 4 (6.34%) came out as G6PD enzyme deficient, provided that the cut-off for G6PD enzyme deficiency was set to 7.37 U/g Hb, as demonstrated in our previous study [ 26 ].…”

“…All 63 specimens were subjected to HRM analysis targeting three previously reported G6PD common variants in Bangladesh [ 26 ]. HRM curve analysis could identify 7 out of 63 samples that were found to vary from the wild-type alleles (Fig.…”

“…Thus, targeting a set of G6PD variants specific to the population of certain geographical location and ethnic background, HRM melting curve approach could be implemented for the screening of heterozygous G6PD variants. Hence, we planned to use HRM approach for screening of G6PD heterozygous females in Bangladesh because only three G6PD variants in the G6PD gene had been reported as the most common in the country [ 26 ]. In addition to three commonly occurring mutations, we had identified another G6PD variant, namely G6PD Mediterranean (c.C563T).…”