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

Tarikul Islam ,Syeda Kashfi Qadri,Mohammad Amir Hossain,Nafisa Nawal Islam,Mohammad Abdul Mannan,Shahida Akhter,Manzoor Hussain,Syed Saleheen Qadri,Shezote Talukder,Jakia Rahman,Hasan Mahmud,Firoza Begum,Mohammod Shahidullah,Suprovath Kumar Sarker,Narayan Chandra Saha ,Sarabon Tahura,Asifuzzaman Rahat,A K M Muraduzzaman,Nazmun Nahar,Firdausi Qadri,Tahmina Shirin,Sharif Akhteruzzaman,Golam Sarower Bhuyan,Kaiissar Mannoor

doi:10.1186/s12863-018-0664-1

Abstract

BackgroundLike glucose-6-phosphate dehydrogenase (G6PD) deficient hemizygous males and homozygous females, heterozygous females could also manifest hemolytic crisis, neonatal hyperbilirubinemia or kernicterus upon exposure to oxidative stress induced by certain foods such as fava beans, drugs or infections. Although hemizygous males and homozygous females are easily detected by conventional G6PD enzyme assay method, the heterozygous state could be missed by the conventional methods as the mosaic population of both normal and deficient RBCs circulates in the blood. Thus the present study aimed to apply high resolution melting (HRM) curve analysis approach to see whether HRM could be used as a supplemental approach to increase the chance of detection of G6PD heterozygosity.ResultsSixty-three clinically suspected females were evaluated for G6PD status using both enzyme assay and HRM analysis. Four out of sixty-three participants came out as G6PD deficient by the enzyme assay method, whereas HRM approach could identify nine participants with G6PD variants, one homozygous and eight heterozygous. Although only three out of eight heterozygous samples had G6PD enzyme deficiency, the HRM-based heterozygous G6PD variants detection for the rest of the samples with normal G6PD enzyme activities could have significance because their newborns might fall victim to serious consequences under certain oxidative stress.ConclusionsIn addition to the G6PD enzyme assay, HRM curve analysis could be useful as a supplemental approach for detection of G6PD heterozygosity.

Highlights

Like glucose-6-phosphate dehydrogenase (G6PD) deficient hemizygous males and homozygous females, heterozygous females could manifest hemolytic crisis, neonatal hyperbilirubinemia or kernicterus upon exposure to oxidative stress induced by certain foods such as fava beans, drugs or infections
Studies have shown that random X-chromosome inactivation can result in mosaic populations of normal and deficient erythrocytes [9] and heterozygous females can be affected in some circumstances and especially such a situation can occur when the population of deficient Red blood cells (RBCs) develops hemolysis under conditions of oxidative stress [10,11,12]
It has been illustrated that the proportion of defective to normal RBC population might be subjected to change over time, e.g., the age-related bias of X-chromosome inactivation leading to G6PD deficiency in octogenarian, nanogenarian, and centenarian females in a population with prevalent G6PD variants have been reported [16]

Summary

Introduction

Like glucose-6-phosphate dehydrogenase (G6PD) deficient hemizygous males and homozygous females, heterozygous females could manifest hemolytic crisis, neonatal hyperbilirubinemia or kernicterus upon exposure to oxidative stress induced by certain foods such as fava beans, drugs or infections. Studies have shown that random X-chromosome inactivation can result in mosaic populations of normal and deficient erythrocytes [9] and heterozygous females can be affected in some circumstances and especially such a situation can occur when the population of deficient RBCs develops hemolysis under conditions of oxidative stress [10,11,12]. Heterozygous females could give birth to hemizygous male newborns who could suffer from hemolytic crisis upon accidental exposure to oxidative stress which could lead to hyperbilirubinemia, kernicterus and even death [17, 18]

Methods

Results

Discussion

Conclusion