Glucose-6-phosphate Dehydrogenase Gene Mutations Research Articles

Identification of mutation in G6PD gene in Nung ethnic patients with glucose-6-phosphate dehydrogenase deficiency

Glucose-6-phosphate dehydrogenase (G6PD) is the key enzyme that initiates the pentose phosphate cycle in glucose metabolism. Physiologically, this pathway is the main source of nicotinamide adenine dinucleotide phosphate (NADPH) for red blood cells. This study aims to identify glucose-6-phosphate dehydrogenase (G6PD) mutations in Nung ethnic patients with G6PD deficiency. 18 pediatric patients of the Nung ethnic group were diagnosed with G6PD enzyme deficiency at the Vietnam National Children’s Hospital and applied PCR and gene sequencing to detect mutations in the G6PD gene. 8 types of mutation were detected in the G6PD gene in which the most common mutation wasKaiping (c.1388G>A) with 44.4%, followed by Canton (c.1376G>T), Viangchan (c.871G>A), Union (c.1360C>T), Gaohe (c.95A>G), Orissa (c.131C>G), and Chinese-5 (c.1024C>T). Silent mutations at the 1311C>T location were found in 3 cases.

Newborn screening for G6PD deficiency in HeFei, FuYang and AnQing, China: Prevalence, cut-off value, variant spectrum.

Glucose-6-phosphate dehydrogenase (G6PD) deficiency is an X-linked recessive Mendelian genetic disorder characterized by neonatal jaundice and hemolytic anemia, affecting more than 400 million people worldwide. The purpose of this research was to investigate prevalence rates of G6PD deficiency and to evaluate and establish specific cut-off values in early prediction of G6PD deficiency by regions (HeFei, FuYang, AnQing) on different seasons, as well as to investigate the frequencies of G6PD gene mutations among three regions mentioned above. A total of 31,482 neonates (21,402, 7680, and 2340 for HeFei, FuYang, and AnQing cities, respectively) were recruited. Positive subjects were recalled to attend genetic tests for diagnosis. G6PD activity on the Genetic screening processor (GSP analyzer, 2021-0010) was measured following the manufactureržs protocol. The cut-off value was first set to 35 U/dL. The receiver operating characteristics (ROC) curve was employed to assess and compare the efficiency in predicting G6PD deficiency among HeFei, FuYang, and AnQing cities in different seasons.

Severe G6PD deficiency leads to recurrent infections and defects in ROS production: Case report and literature review.

Purpose: Severe glucose-6-phosphate dehydrogenase (G6PD) deficiency can lead to reduced nicotinamide adenine dinucleotide phosphate oxidase activity in phagocytes, resulting in immunodeficiency, with a limited number of reported cases. Here, we aimed to report a child with severe G6PD deficiency in China and investigate the mechanism of his recurrent infections. Methods: The clinical manifestations and immunological phenotypes of this patient were retrospectively collected. Gene mutation was detected by whole-exome sequencing and confirmed by Sanger sequencing. Dihydrorhodamine (DHR) analysis was performed to measure the respiratory burst of neutrophils. Messenger ribonucleic acid and protein levels were detected in the patient under lipopolysaccharide stimulation by real-time quantitative reverse transcription polymerase chain reaction and Western blot. A review of the literature was performed. Results: A male child with G6PD deficiency presented with recurrent respiratory infections, Epstein‒Barr virus infection and tonsillitis from 8 months of age. Gene testing revealed that the proband had one hemizygous mutation in the G6PD gene (c.496C>T, p. R166C), inherited from his mother. This mutation might affect hydrophobic binding, and the G6PD enzyme activity of the patient was 0. The stimulation indexes of the neutrophils in the patient and mother were 22 and 37, respectively. Compared with healthy controls, decreased reactive oxygen species (ROS) production was observed in the patient. Activation of nuclear factor kappa-B (NF-κB) signaling was found to be influenced, and the synthesis of tumor necrosis factor alpha (TNF-α) was downregulated in the patient-derived cells. In neutrophils of his mother, 74.71% of the X chromosome carrying the mutated gene was inactivated. By performing a systematic literature review, an additional 15 patients with severe G6PD deficiency and recurrent infections were identified. Four other G6PD gene mutations have been reported, including c.1157T>A, c.180_182del, c.514C>T, and c.953_976del. Conclusion: Severe G6PD deficiency, not only class I but also class II, can contribute to a chronic granulomatous disease-like phenotype. Decreased reactive oxygen species synthesis led to decreased activation of the NF-κB pathway in G6PD-deficient patients. Children with severe G6PD deficiency should be aware of immunodeficiency disease, and the DHR assay is recommended to evaluate neutrophil function for early identification.

Cut-off values for diagnosis of G6PD deficiency by flow cytometry in Thai population

In heterozygous females, X-inactivation causes a change in glucose-6-phosphate dehydrogenase (G6PD) activity from normal to deficient. Most G6PD screening tests are used to accurately diagnose hemizygous males, but they are less reliable for diagnosing heterozygous females. This study established flow cytometric cut-off values for screening of G6PD deficiency in hemizygous males and heterozygous or homozygous females. We studied 205 (125 females, 80 males) leftover blood samples from quantitative methemoglobin reduction (MR) screening. G6PD gene mutations determined by multiplex amplification refractory mutation system-polymerase chain reaction and direct DNA sequencing were used as the gold standard reference. Accuracy of the test, including the sensitivity, specificity, and positive and negative predictive values, was analyzed using MedCalc software. The optimal cut-off values for classification of %red blood cells with normal G6PD activity or %bright cells into homozygous normal, heterozygous, and homozygous deficiency in females were 85.4–100%, 6.3–85.3%, and 0–6.2%, respectively (sensitivity 93.2%, specificity 100%). The cut-offs for classification into hemizygous normal and hemizygous deficiency in males were 76.5–100% and 0–76.4%, respectively (sensitivity 100%, specificity 96.5%). Flow cytometry can be used to differentiate heterozygous females with intermediate phenotype from homozygous females, but cannot distinguish between heterozygous females with extreme phenotype and homozygous females. By flow cytometry, heterozygous and homozygous deficiency was detected in 29.6% and 3.2% of females, respectively. Among males, hemizygous deficiency was found in 31.3%. Flow cytometry can be used to screen patients with G6PD deficiency, and reliably and efficiently identify heterozygous and homozygous females, and hemizygous males based on cellular G6PD activity.

Molecular Characterization and Genotype-Phenotype Correlation of G6PD Mutations in Five Ethnicities of Northern Vietnam.

Glucose-6-phosphate dehydrogenase (G6PD) deficiency is the most common enzyme disorder and is caused by G6PD gene mutations. To date, more than 400 variants in the G6PD gene have been discovered, and about 160 identified variants are associated with a significant decrease in the G6PD enzyme activity. However, the molecular characterization and epidemiological study of G6PD deficiency are still limited in Vietnam. Therefore, we conducted this study to determine the G6PD variants among the Vietnamese populations and evaluate their correlation to G6PD enzyme activity. A total of 339 patients (302 males and 37 females) were enrolled in this study. The G6PD variants were identified by Sanger sequencing. Our results indicate that males are more severely deficient in G6PD than females. This enzyme activity in males (1.27 ± 1.06 IU/g·Hb) is significantly lower than in females (2.98 ± 1.57 IU/g·Hb) (p < 0.0001). The enzyme activity of the heterozygous-homozygous females and heterozygous females-hemizygous males was found to be significantly different (p < 0.05), which is interpreted due to random X-inactivation. For G6PD molecular characteristics, Viangchan (c.871G>A), Canton (c.1376G>T) and Kaiping (c.1388G>A) variants were the most dominant, accounting for 24.48%, 17.70%, and 22.42%, respectively, whereas the highest frequency of complex variants was observed in Viangchan/Silent with 20.35%. In terms of G6PD activity, the Union variant presented the lowest mean value (1.03 IU/g·Hb) compared to the other variants (p < 0.05). Computational analysis using Polyphen-2 tool investigated that all variants were relative to G6PD deficiency and separated the levels as benign and damaged. The result will establish effective methods to screen G6PD variants in Vietnam.

Molecular genotyping of G6PD mutations for neonates in Ningbo area.

The aim of this study is to determine the cut‐off value of glucose‐6‐phosphate dehydrogenase (G6PD) activity and the mutation spectrum of G6PD gene in neonates with G6PD deficiency at Ningbo. Around 82233 neonatal blood samples were measured to determine G6PD activity. The positive samples were further detected with gene analysis. A total of 445 neonates were confirmed as G6PD deficiency, and the incidence in Ningbo was 1/185. 17 types of G6PD gene mutations were found, including 11 single‐site mutations and 6 double‐site mutations. Considering the significant differences in G6PD activity, the cut‐off value was detected to be 2.35 and 3.65 U/gHb for males and females, respectively. Significant differences in G6PD activities were noted and found to be varied from 4.61 to 6.02 U/gHb in different seasons (p < 0.0001). G6PD deficiency screening is a significant detection test for neonatal G6PD deficiency prevention. Our study highlights that the screening should be done using different cut‐off values according to the sexes in different seasons.

Level of apoptosis in Saudi patients with a defect in Glucose-6-phosphate dehydrogenase (G6PD)

IntroductionGlucose-6-phosphate dehydrogenase (G6PD) is an enzyme responsible for the production of NADPH pathway. Mutation in G6PD gene results in abnormal functional and structural changes in red blood cells (RBCs). The aim of the current study is to compare the levels of apoptosis in Saudi people suffering from G6PD deficiency. Materials and methodsTwenty samples from unrelated Saudi children and neonates between the ages of 1 month to 10 years were collected. Ten ml Blood samples were collected by venipuncture in EDTA tubes, and hematological parameters assessed using Coulter Counter Analyzer. MNC cells were isolated. Apoptotic leukocytes were determined and concentaration of IL-107 release was measured using the FACS machine. ResultThere was a significant increase in white blood cells (WBCs) count in G6PD deficiency group compared to healthy controls. Also, there was a sgnificant increase in the level of apoptosis observed in the freshly isolated RBCs obtained from G6PD deficiency patients. ConclusionThe findings may indicate that apoptosis of RBCs represents an important factor in the pathogenesis of G6PD.

Glucose-6-phosphate dehydrogenase deficiency in the Han Chinese population: molecular characterization and genotype\u2013phenotype association throughout an activity distribution

Glucose-6-phosphate dehydrogenase (G6PD) deficiency is a common hereditary disorder in China. The existing prevalence and molecular epidemiology of G6PD deficiency in China were geographically limited. In this study, the spectrum of G6PD gene mutations was well characterized in a large and diverse population all over the country; and the correlation of genotype and enzyme activity phenotype was explored for the first time. The results showed that the overall prevalence of G6PD deficiency in China was 2.10% at the national level. The top six common mutations were c.1388 G>A, c.1376 G>T, c.95 A>G, c.392 G>T, c.871 G>A and c.1024 C>T, accounting for more than 90% of G6PD deficient alleles. Compound mutation patterns were frequently observed in females with severe deficiency. The distribution of G6PD activities depended on the type of mutation patterns and genders. Hemizygote, homozygote, and compound heterozygote were predominantly associated with severe G6PD deficiency, whereas heterozygotes with single mutation mainly presented moderate enzyme deficiency. A significant gap between G6PD activities in hemizygous and normal males was observed, and yet, the overall distribution of that in females carrying missense mutations was a continuum from G6PD severely deficient to normal. This is the first report of discussing the association between G6PD genetic variants in the Chinese and enzyme activity phenotypes.

PB1968 TWO NEW MUTATIONS AND MOLECULAR CHARACTERIZATION OF G6PD GENE AMONG THAI CHILDREN WITH G6PD DEFICIENCY

Background:Glucose‐6‐phosphate dehydrogenase (G6PD) deficiency is the most common X‐linked inherited erythroenzymopathy in Thailand. The clinical and hematological manifestations of G6PD deficiency is variable.Aims:This study aimed to characterize the spectrum of G6PD gene mutations in Thai pediatric patients who were followed‐up in Phramongkutklao Hospital, a tertiary center in central Thailand.Methods:A total of 102 children including 73 males (71.6%) and 29 females (28.4%) were included in our study. Mutation analysis was performed by direct DNA sequencing of the G6PD gene. Neonatal hyperbilirubinemia was the most common (87.3%) clinical manifestation in this study.Results:Molecular analysis of G6PD gene in 102 G6PD‐deficient Thai children identified a total of 12 mutational types. G6PD Viangchan (871G&gt;A) was the most common (46.2%) identified mutation. G6PD Canton (1376G &gt; T) was the second most common and was identified in 15.4%, followed by G6PD Kaiping (1388G &gt; A) 14.4% and G6PD Mahidol (487G &gt; A) 8.6%, respectively. Eight uncommon G6PD mutations including G6PD Quing Yan (392G &gt; T) (3.8%), G6PD Aures (143C &gt; T) (1.9%), G6PD Coimbra (592C &gt; T) (1.9%), G6PD Songklanagarind (196T &gt; A) (1.9%), G6PD Union (1360C &gt; T) (1.9%), G6PD Valladolid (406C &gt; T) (1.9%), G6PD Chinese‐5 (1024C &gt; T) (1%) and G6PD Mediterranean (563C &gt; T) (1%) were identified in this study. G6PD Valladolid and G6PD Aures were firstly identified in Thai populations.Summary/Conclusion:This study characterizes the molecular heterogeneity of G6PD variants causing G6PD deficiency in Thai children. Our study demonstrated the efficiency of direct DNA sequencing which can identify mutations in all 102 patients.

Impact of glucose-6-phosphate dehydrogenase deficiency on dengue infection in Myanmar children.

Glucose-6-phosphate dehydrogenase (G6PD) deficiency may affect the clinical presentation of dengue due to the altered redox state in immune cells. We aimed to determine the association between G6PD deficiency and severity of dengue infection in paediatric patients in Myanmar. A cross-sectional study was conducted among paediatric patients aged 2–13 years with dengue in Yankin Children Hospital, Myanmar. One hundred and ninety-six patients positive for dengue infection, as determined via PCR or ELISA, were enrolled. Dengue severity was determined according to the 2009 WHO classification guidelines. Spectrophotometric assays determined G6PD levels. The adjusted median G6PD value of males in the study population was used to define various cut-off points according to the WHO classification guidelines. G6PD genotyping for Mahidol, Kaiping and Mediterranean mutations was performed for 128 out of 196 samples by real-time multiplex PCR. 51 of 196 (26.0%) patients had severe dengue. The prevalence of G6PD phenotype deficiency (< 60% activity) in paediatric patients was 14.8% (29/196), specifically, 13.6% (14/103) in males and 16.2% (15/93) in females. Severe deficiency (< 10% activity) accounted for 7.1% (14/196) of our cohort, occurring 11.7% (12/103) in males and 2.2% (2/93) in females. Among 128 samples genotyped, the G6PD gene mutations were detected in 19.5% (25/128) of patients, with 20.3% (13/ 64) in males and 18.8% (12/64) in females. The G6PD Mahidol mutation was 96.0% (24/25) while the G6PD Kaiping mutation was 4.0% (1/25). Severe dengue was not associated with G6PD enzyme deficiency or presence of the G6PD gene mutation. Thus, no association between G6PD deficiency and dengue severity could be detected.Trial registration: The study was registered following the WHO International Clinical Trials Registry Platform (WHO-ICTRP) on Thai Clinical Trials Registry (TCTR) website, registration number # TCTR20180720001

Chinese Newborn Screening for G6PD Deficiency, with Activity and Gene

Background: Glucose-6-phosphate dehydrogenase (G6PD) deficiency is one of the most common X-linked enzymopathies caused by G6PD gene mutation. We aimed to provide the characteristics of G6PD deficiency and G6PD gene mutation distribution in a large Chinese newborn screening (NBS) population. Methods: We collected screening information of all the NBS centers around China, from January 2013 to December 2017. G6PD activity fluorescence screenings and G6PD/6PGD tests were performed in 1,764,299 birth population from 29 centers in 12 representative provinces in 2016. We then performed multicolour melting curve analysis (MMCA) to classify G6PD gene mutations in 10,357 neonates with activity-confirmed G6PD deficiency, and DNA Sanger sequencing for G6PD coding exons if hot site mutations were not found. Finally, we investigated G6PD gene mutation with MMCA in 6,443 female and 7,358 male neonates with normal G6PD activity respectively. Findings: The screened population, organizations and provinces of G6PD deficiency were increased from 2013 to 2017 in China, high in South and low in North. The top 5 frequency of G6PD gene mutations were c.1376G>T, c.1388G>A, c.95A>G, c.1024C>T, and c.871G>A and varied in different provinces, with regional and ethnic features, and 4 pathogenic mutation sites (c.152C>T, c.290A>T, c.697G>C, and c.1285A>G ) were firstly reported. Three hundred and seventy-two (372/6,443, 5·77%) female neonates with normal G6PD activity were detected as G6PD gene heterozygotes and 50 (17·42%, 50/287) female G6PD heterozygotes showed G6PD deficiency symptom in two years, suggesting that a supplementary NBS strategy of G6PD gene first screening for female G6PD deficiency in high-incidence regions of South China. Interpretation: G6PD deficiency mainly occurs in South China, and the frequency of G6PD gene mutation varies in different regions and ethnicities. Our suggested NBS strategy for G6PD deficiency could enhance the detection rate of female G6PD heterozygotes. Funding Statement: This work is partially financial supported by Natural Scientific Foundation of China (81570142), and the Ministry of Science and Technology (2016YFA0101300, 2017YFC1001700). Declaration of Interests: All the authors have nothing to disclose. Ethics Approval Statement: This study was approved by the Research Ethics Committees of Children’s Hospital of Chongqing Medical University and registered on Chinese Clinical Trail Registry (ChiCTR-SOC-17014057).

Molecular diagnosis and mutation identification in newborn with glucose-6-phosphate dehydrogenase deficiency by DNA sequencing

Objective To evaluate the clinical significance of molecular diagnosis for detecting neonatal glucose-6-phosphate dehydrogenase (G6PD) deficiency and to identify the novel mutations in G6PD gene. Methods Totally 231 newborn blood samples with decreased G6PD deficiency assessed by first-line newborn screening were recruited from Children's Hospital, Chongqing Medical University. G6PD gene mutation analysis was performed by PCR following by Sanger DNA sequencing, and then hotspot mutation sites were confirmed. The relationship between the enzymatic phenotype and genotype was evaluated by comparing the results between the G6PD activity detection and the molecular diagnosis. Novel variations were mutation predicted to identity the pathogenic mutations. Results Totally 125 G6PD patients were identified in present study group, among which 102 patients were classified as positive by G6PD/6PGD quantitative tests, giving out a detecting rate of 81.6%. The 123 patients were confirmed by molecular diagnosis with a detecting of 98.4%. Within the 16 probably positive cases, 11 definite mutations were confirmed by molecular diagnosis. DNA sequencing also confirmed another group of 12 patients from the 113 negative samples, giving a detecting rate of 10.6%. Two novel mutations of G6PD gene, c. 29C>A and c. 361C>T were found in the study, of which, c. 29C>A was observed in 7.3% (9/123) of the samples and might be a new hotspot site in Chinese population. Conclusions Quantitative ratio testing may be not efficient enough for G6PD deficiency diagnosis in patients carrying heterozygous mutations. Novel mutations can be found in G6PD gene and obviously there are regional diversities in the frequency of hotspot mutations for G6PD deficiency. Therefore, DNA sequencing based molecular testing may serve as the most valuable method for the diagnosis of neonatal G6PD deficiency.(Chin J Lab Med, 2016, 39: 843-847) Key words: Glucosephosphate dehydrogenase deficiency; Glucosephosphate dehydrogenase; Sequence analysis, DNA; Molecular diagnostic techniques; Mutation; Neonatal screening

Application of Mass-array gene chip to detect glucose-6-phosphate dehydrogenase gene mutations

Objective To develop the Mass-array gene chip to detect glucose-6-phosphate dehydrogenase (G6PD) gene mutations, and to evaluate its quality. Methods Randomly choosing the children who perform neonatal screening in Neonatal Screening Center of Fujian Maternity and Children Health Hospital from 2006 to 2013. Children were divided into control group and G6PD patient group. Using Genotyping Tools from Sequenom company and the software of Mass-array Assay Design to design the PCR amplification primer of 33 G6PD gene mutations which were well-known in Chinese. Then depending on Mass-array gene chip technology to detect glucose-6-phosphate dehydrogenase gene mutations. DNA Sanger sequencing was used to verify the accuracy of the testing results of gene chip. Results In the 300 children with G6PD deficiency, we detected 9 kinds of simplex single point mutations: 1376G>T, 1388G>A, 95A>G, 1024C>T, 392G>T, 1360C>T, 487G>A, 517T>C, 1365-13T>C and 7 kinds of compound mutations: 871G>A/1365-13T>C/1311C>T, 1004C>A/1311C>T/1365-13T>C, 1376G>T/ 1365-13T>C/1311C>T, 1365-13T>C/1311C>T, 1376G>T/1365-13T>C, 95A>G/1365-13T >C/1311C>T, 1388G>A/1365-13T >C. No G6PD gene mutation was detected in 300 normal children. The results of DNA Sanger sequencing were identical to that of gene chip. Conclusion Mass-array of G6PD gene is an effective, accurate and efficient way for G6PD gene mutation screening.(Chin J Lab Med, 2015, 38: 822-826) Key words: Glucosephosphate dehydrogenase; Mutation; Oligonucleotide array sequence analysis; Sequence analysis, DNA; Polymerase chain reaction

Glucose-6-phosphate dehydrogenase deficiency in Japan

In the past 10 years, we have diagnosed congenital hemolytic anemia in 294 patients, approximately 33% of whom were found to have glucose-6-phosphate dehydrogenase (G6PD) deficiency. It is becoming more common for Japanese to marry people of other ethnic origins, such that G6PD deficiency is becoming more prevalent in Japan. Japanese G6PD deficiency tends to be diagnosed in the neonatal period due to severe jaundice, while G6PD-deficient patients with foreign ancestors tend to be diagnosed at the onset of an acute hemolytic crisis before the age of six. It is difficult to predict the clinical course of each patient by G6PD activity, reduced glutathione content, or the presence/absence of severe neonatal jaundice. We propose that both neonatal G6PD screening and systematic analyses of G6PD gene mutations may be useful for personalized management of patients with G6PD-deficient hemolytic anemia.

A multiplex method for detection of glucose-6-phosphate dehydrogenase (G6PD) gene mutations.

Glucose-6-phosphate dehydrogenase (G6PD) deficiency is the most common human enzyme defect caused by G6PD gene mutations. This study aimed to develop a cost-effective, multiplex, genotyping method for detecting common mutations in the G6PD gene. We used a SNaPshot approach to genotype multiple G6PD mutations that are common to human populations in South-East Asia. This assay is based on multiplex PCR coupled with primer extension reactions. Different G6PD gene mutations were determined by peak retention time and colors of the primer extension products. We designed PCR primers for multiplex amplification of the G6PD gene fragments and for primer extension reactions to genotype 11 G6PD mutations. DNA samples from a total of 120 unrelated G6PD-deficient individuals from the China-Myanmar border area were used to establish and validate this method. Direct sequencing of the PCR products demonstrated 100% concordance between the SNaPshot and the sequencing results. The SNaPshot method offers a specific and sensitive alternative for simultaneously interrogating multiple G6PD mutations.

Molecular epidemiology analysis of glucose-6-phosphate dehydrogenase gene mutations among infertile patients in Shenzhen

To explore the incidence and genotypes of glucose-6-phosphate dehydrogenase (G6PD) gene mutations among infertile patients in Shenzhen. DNA samples from 851 infertile patients were tested for 25 G6PD gene mutation sites using a multiplex SNaPshot assay. The incidence of G6PD gene mutations among infertile patients in Shenzhen was 17.63%. Male and female abnormal rates were 15.13% and 20.09% respectively. Most of the female abnormal cases were heterozygotes. Mutations involved 11 haplotypes in 10 sites. 1311C> T/IVS-11 93T> C was the most common mutation, accounting for 72.00% (108/150) abnormal cases. Forty three cases of missense mutations were detected, including 19 cases of 1376G> T, 9 cases of 1388G> A, 5 cases of 95A> G and 871G> A/1311C> T/IVS-11 93T> C, 1 case of 202G> A, 835A> T, 1360C> T, 1376G> T and 392G> T/1311C> T/IVS-11 93T> C. The incidence of G6PD gene mutations among infertile patients in Shenzhen was high and the mutation types were various. Therefore, the G6PD deficiency genetic screening should be performed prior to assisted reproduction. This investigated results provided valuable basic data for genetic counseling, preimplantation genetic diagnosis and prenatal diagnosis.

DNA hypermethylation and X chromosome inactivation are major determinants of phenotypic variation in women heterozygous for G6PD mutations.

Glucose-6-phosphate dehydrogenase (G6PD) deficiency is an X-linked incompletely dominant enzyme deficiency that results from G6PD gene mutations. Women heterozygous for G6PD mutations exhibit variation in the loss of enzyme activity but the cause of this phenotypic variation is unclear. We determined DNA methylation and X-inactivation patterns in 71 G6PD-deficient female heterozygotes and 68 G6PD non-deficient controls with the same missense mutations (G6PD Canton c.1376G>T or Kaiping c.1388G>A) to correlate determinants with variable phenotypes. Specific CpG methylations within the G6PD promoter were significantly higher in G6PD-deficient heterozygotes than in controls. Preferential X-inactivation of the G6PD wild-type allele was determined in heterozygotes. The incidence of preferential X-inactivation was 86.2% in the deficient heterozygote group and 31.7% in the non-deficient heterozygote group. A significant negative correlation was observed between X-inactivation ratios of the wild-type allele and G6PD/6-phosphogluconate dehydrogenase (6PGD) ratios in heterozygous G6PD Canton (r=-0.657, p<0.001) or Kaiping (r=-0.668, p<0.001). Multivariate logistic regression indicated that heterozygotes with hypermethylation of specific CpG sites in the G6PD promoter and preferential X-inactivation of the wild-type allele were at risk of enzyme deficiency.

Prevalence of Three Common Glucose-6-Phosphate Dehydrogenase Gene Mutations in Neonates in Province of Mazandaran, North of Iran, 2012

In northern provinces of Iran high rates of incidence of glucose-6-phosphate dehydrogenase (G6PD) enzyme deficiency have been reported and most of these patients carry one of the three common G6PD gene mutations: Mediterranean, Chatham or Cosenza. The aim of this study was to investigate prevalence of each of these mutations among neonates in Mazandaran, a northern province of Iran. Four hundred and twelve blood samples were collected and using standard protocols DNA was extracted. In order to detect the above mutations PCR-RFLP method was applied. Fifty- three of neonates had G6PD gene mutation (12.9%, CI 95%: 9.66-16.14). About 17% of female and 9% of male newborns were carriers for one of the three common G6PD gene mutations. The Mediterranean type had the highest gene frequency (0.0607) among the three examined mutations. The present study shows around 17% (CI 95%: 11.97-22.03) of Mazandarani female population is carrier for one of the three mutations and since the likelihood of having an affected child in a carrier woman is 1 in every 4 child births, the researchers recommended all women to be screened for the presence of three common G6PD gene mutations prior to pregnancy.

Glucose-6-phosphate dehydrogenase (G6PD) gene mutations detection by improved high-resolution DNA melting assay

Glucose-6-phosphate dehydrogenase (G6PD) deficiency is a common inherited disorder worldwide including southern China. G6PD gene mutations cause deficiency of the enzyme and a large spectrum of diseases. High-resolution DNA melting (HRM) assay was recently introduced as a rapid, inexpensive and effective method for genotyping. But there was a shortcoming of this method that hemizygous and homozygous genotypes were not easily distinguished from wild-types. Here we used improved HRM method for a small-scale screening of G6PD-deficient variants among people of Meizhou region. Then all amplicons were ascertained by direct DNA sequencing. These results indicated that HRM method was a major technical advance for G6PD mutations screening.

A novel point mutation in a class IV glucose-6-phosphate dehydrogenase variant (G6PD São Paulo) and polymorphic G6PD variants in São Paulo State, Brazil

In this study, we used red cell glucose-6-phosphate dehydrogenase (G6PD) activity to screen for G6PD-deficient individuals in 373 unrelated asymptomatic adult men who were working with insecticides (organophosphorus and carbamate) in dengue prevention programs in 27 cities in São Paulo State, Brazil. Twenty-one unrelated male children suspected of having erythroenzymopathy who were attended at hospitals in São Paulo city were also studied. Fifteen of the 373 adults and 12 of the 21 children were G6PD deficient. G6PD gene mutations were investigated in these G6PD-deficient individuals by using PCR-RFLP, PCR-SSCP analysis and DNA sequencing. Twelve G6PD A-202A/376G and two G6PD Seattle844C, as well as a new variant identified as G6PD São Paulo, were detected among adults, and 11 G6PD A-202A/376G and one G6PD Seattle844C were found among children. The novel mutation c.660C > G caused the replacement of isoleucine by methionine (I220M) in a region near the dimer interface of the molecule. The conservative nature of this mutation (substitution of a nonpolar aliphatic amino acid for another one) could explain why there was no corresponding change in the loss of G6PD activity (64.5% of normal activity in both cases).