Reduction of Missed Diagnosis of G6PD Deficiency in Heterozygous Females by G6PD/6PGD Ratio Assay Combined with Amplification Refractory Mutation System PCR

Abstract

Objective: Glucose-6-phosphate dehydrogenase (G6PD) deficiency is an X-linked genetic disorder that results in impaired enzyme activity. The G6PD/6PGD ratio assay was routinely used for G6PD deficiency screening in China, but there is an apparent defect of missed diagnosis in heterozygous females. The study aims to explore the means to improve its accuracy. Methods: A total of 4,161 Chinese females of childbearing age were collected in this retrospective study. All samples were first subjected to G6PD/6PGD ratio assay and then screened by amplification refractory mutation system PCR (ARMS-PCR) for six hotspot mutants in Chinese population (c.1376G>T, c.1388G>A, c.95A>G, c.1024C>T, c.392G>T, and c.871G>A). For the samples with G6PD/6PGD ratio<1.0 and no mutations were found by ARMS-PCR, next-generation sequencing (NGS) was performed. Sanger sequencing was finally used to verify all the variants. Results: The prevalence of G6PD deficiency in Shenzhen females of childbearing age was 7.31%. The proportion of the six hotspot mutations accounted for 98.03% of all 304 G6PD variants carriers. Taking the ARMS-PCR/NGS results as a reference, the missed diagnosis rate of the G6PD/6PGD ratio assay was 33.88%. Using ARMS-PCR to retest the samples with a G6PD/6PGD ratio between 1.00 and ∼1.10 or 1.00 and ∼1.15 could reduce the missed diagnosis rate from the original 33.88% to 18.09% or 12.05% separately. Conclusion: ARMS-PCR is an appropriate supplementary method for discovering most carriers missed by the G6PD/6PGD ratio assay.

© 2022 The Author(s). Published by S. Karger AG, Basel

References Luzzatto L, Nannelli C, Notaro R. Glucose-6-phosphate dehydrogenase deficiency. Hematol Oncol Clin North Am. 2016;30(2):373–93. Frank JE. Diagnosis and management of G6PD deficiency. Am Fam Physician. 2005;72(7):1277–82. Belfield KD, Tichy EM. Review and drug therapy implications of glucose-6-phosphate dehydrogenase deficiency. Am J Health Syst Pharm. 2018;75(3):97–104. Gómez-Manzo S, Marcial-Quino J, Vanoye-Carlo A, Serrano-Posada H, Ortega-Cuellar D, González-Valdez A, et al. Glucose-6-phosphate dehydrogenase: update and analysis of new mutations around the world. Int J Mol Sci. 2016;17(12):2069. Minucci A, Moradkhani K, Hwang MJ, Zuppi C, Giardina B, Capoluongo E. Glucose-6-phosphate dehydrogenase (G6PD) mutations database: review of the “old” and update of the new mutations. Blood Cells Mol Dis. 2012;48(3):154–65. Beutler E, Yeh M, Fairbanks VF. The normal human female as a mosaic of X-chromosome activity: studies using the gene for G-6-PD-deficiency as a marker. Proc Natl Acad Sci U S A. 1962;48(1):9–16. Cappellini MD, Fiorelli G. Glucose-6-phosphate dehydrogenase deficiency. Lancet. 2008;371(9606):64–74. Luzzatto L. Glucose 6-phosphate dehydrogenase deficiency: from genotype to phenotype. Haematologica. 2006;91(10):1303–6. He M, Lin K, Huang Y, Zhou L, Yang Q, Li S, et al. Prevalence and molecular study of G6PD deficiency in the Dai and Jingpo ethnic groups in the Dehong Prefecture of the Yunnan Province. Hum Hered. 2018;83(2):55–64. Jiang W, Yu G, Liu P, Geng Q, Chen L, Lin Q, et al. Structure and function of glucose-6-phosphate dehydrogenase-deficient variants in Chinese population. Hum Genet. 2006;119(5):463–78. Bhutani VK, Kaplan M, Glader B, Cotten M, Kleinert J, Pamula V. Point-of-care quantitative measure of glucose-6-phosphate dehydrogenase enzyme deficiency. Pediatrics. 2015;136(5):e1268–75. Roper DR, De la Salle B, Soni V, Fletcher K, Green JA. Abrogation of red blood cell G6PD enzyme activity through Heat treatment: development of survey material for the UK NEQAS G6PD scheme. Int J Lab Hematol. 2017;39(3):308–16. Adu P, Simpong DL, Takyi G, Ephraim RKD. Glucose-6-phosphate dehydrogenase deficiency and sickle cell trait among prospective blood donors: a cross-sectional study in Berekum, Ghana. Adv Hematol. 2016;2016:7302912. Henriques G, Phommasone K, Tripura R, Peto TJ, Raut S, Snethlage C, et al. Comparison of glucose-6 phosphate dehydrogenase status by fluorescent spot test and rapid diagnostic test in Lao PDR and Cambodia. Malar J. 2018;17(1):243. Tseng CP, Huang CL, Chong KY, Hung IJ, Chiu DTY. Rapid detection of glucose-6-phosphate dehydrogenase gene mutations by denaturing high-performance liquid chromatography. Clin Biochem. 2005;38(11):973–80. Fan Z, Weng X, Huang G, Pan Z, Long Z, Fan Q, et al. STARD-rapid screening for the 6 most common G6PD gene mutations in the Chinese population using the amplification refractory mutation system combined with melting curve analysis. Medicine. 2018;97(17):e0426. Maffi D, Pasquino MT, Caprari P, Caforio MP, Cianciulli P, Sorrentino F, et al. Identification of G6PD Mediterranean mutation by amplification refractory mutation system. Clin Chim Acta. 2002;321(1–2):43–7. Islam MT, Sarker SK, Talukder S, Bhuyan GS, Rahat A, Islam NN, et al. High resolution melting curve analysis enables rapid and reliable detection of G6PD variants in heterozygous females. BMC Genet. 2018;19(1):58. Bogari NM. Next generation sequencing (NGS) in glucose-6-phosphate dehydrogenase (G6PD) deficiency studies. Bioinformation. 2016;12(2):41–3. Howes RE, Battle KE, Satyagraha AW, Baird JK, Hay SI. G6PD deficiency: global distribution, genetic variants and primaquine therapy. Adv Parasitol. 2013;81:133–201. Mehta A, Mason PJ, Vulliamy TJ. Glucose-6-phosphate dehydrogenase deficiency. Baillieres Best Pract Res Clin Haematol. 2000;13(1):21–38. Du CS, Xu YK, Hua XY, Wu QL, Liu LB. Glucose-6-phosphate dehydrogenase variants and their frequency in Guangdong, China. Hum Genet. 1988;80(4):385–8. Lin F, Lou ZY, Xing SY, Zhang L, Yang LY. The gene spectrum of glucose-6-phosphate dehydrogenase (G6PD) deficiency in Guangdong province, China. Gene. 2018;678:312–7. Nkhoma ET, Poole C, Vannappagari V, Hall SA, Beutler E. The global prevalence of glucose-6-phosphate dehydrogenase deficiency: a systematic review and meta-analysis. Blood Cells Mol Dis. 2009;42(3):267–78. Lyu RY, Chen XW, Zhang M, Chen YS, Yu J, Wen FQ. Detection of gene mutation in glucose-6-phosphate dehydrogenase deficiency by RT-PCR sequencing. Zhongguo Dang Dai Er Ke Za Zhi. 2016;18(7):630–4. Nanfack AJ, Agyingi L, Noubiap JJN, Ngai JN, Colizzi V, Nyambi PN. Use of amplification refractory mutation system PCR assay as a simple and effective tool to detect HIV-1 drug resistance mutations. J Clin Microbiol. 2015;53(5):1662–71. Du CS, Ren X, Chen L, Jiang W, He Y, Yang M. Detection of the most common G6PD gene mutations in Chinese using amplification refractory mutation system. Hum Hered. 1999;49(3):133–8. Liu Z, Yu C, Li Q, Cai R, Qu Y, Wang W, et al. Chinese newborn screening for the incidence of G6PD deficiency and variant of G6PD gene from 2013 to 2017. Hum Mutat. 2020;41(1):212–21. Li L, Zhou YQ, Xiao QZ, Yan TZ, Xu XM. Development and evaluation of a reverse dot blot assay for the simultaneous detection of six common Chinese G6PD mutations and one polymorphism. Blood Cells Mol Dis. 2008;41(1):17–21. Gao J, Lin S, Chen S, Wu Q, Zheng K, Su J, et al. Molecular characterization of glucose-6-phosphate dehydrogenase deficiency in the Shenzhen population. Hum Hered. 2020;85(3–6):110–6. He Y, Zhang Y, Chen X, Wang Q, Ling L, Xu Y. Glucose-6-phosphate dehydrogenase deficiency in the Han Chinese population: molecular characterization and genotype-phenotype association throughout an activity distribution. Sci Rep. 2020;10(1):17106. Article / Publication Details

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Abstract of Research Article

Received: March 03, 2022
Accepted: September 27, 2022
Published online: October 31, 2022

Number of Print Pages: 7
Number of Figures: 2
Number of Tables: 2

ISSN: 0001-5652 (Print)
eISSN: 1423-0062 (Online)

For additional information: https://www.karger.com/HHE

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