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