Reduction of missed diagnosis of G6PD deficiency in heterozygous females by G6PD/6PGD ratio assay combined with ARMS-PCR

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 4161 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/NGSresults as a reference, the missed diagnosis rate of the G6PD/6PGD ratio assay was 33.88%. Using ARMS-PCRto retest the samples with a G6PD/6PGD ratio between 1.00(similar to)1.10 or 1.00(similar to)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 methodfor discovering most carriers missed by the G6PD/6PGD ratio assay.

Automated summary

This study aims to improve the missed diagnosis issue in heterozygous females during G6PD deficiency screening. By supplementing with methods like ARMS-PCR and NGS, the missed diagnosis rate of G6PD deficiency screening can be significantly reduced.