A 2-year-old Chinese boy was referred to Hospital UKM for investigation of recurrent episodes of dark-coloured urine and pallor since birth. He was born prematurely at 34 weeks gestation and developed severe early-onset neonatal jaundice requiring exchange blood transfusion. Screening at birth showed Glucose-6-phosphate dehydrogenase (G6PD) deficiency. On admission, physical examination revealed pallor, jaundice and mild hepatomegaly. Results of laboratory investigations showed a hemoglobin level of 11.0 g/dl with a hemolytic blood picture, reticulocytosis of 20% and red cell G6PD activity reported as undetectable. The patient's DNA was analysed for G6PD mutations by PCR-based techniques and DNA sequencing and results showed a 24 bp deletion of nucleotide 953-976 in the exon 9 of the G6PD gene. DNA analysis was also performed on blood samples of the patient's mother and female sibling confirming their heterozygous status, although both showed normal red cell G6PD activity levels. The patient was discharged well and his parents were appropriately advised on the condition and the importance of taking folic acid regularly. This is a first case report in Malaysia of G6PD deficiency causing chronic-hemolytic anemia. The rare 24 bp deletion causes the G6PD Nara variant, previously reported only in two other unrelated males, a Japanese and a Portuguese both with chronic hemolytic anemia.
We performed DNA analysis on cord blood samples of 128 Chinese male neonates diagnosed as G6PD deficiency in Hospital Universiti Kebangsaan Malaysia by a combination PCR-restriction enzyme digest technique, Single Stranded Conformation Polymorphism analysis and DNA sequencing. We found 10 different G6PD-deficient mutations exist. The two commonest alleles were G6PD Canton 1376 G>T (42.3%) and Kaiping 1388 G>A (39.4%) followed by G6PD Gaohe 592 G>A (7.0%), Chinese-5 1024 C>T, Nankang 517 T>C (1.5%), Mahidol 487 G>A (1.6%), Chatham 1003 G>T (0.8%), Union 1360 C>T (0.8%), Viangchan 871 G>A (0.8%) and Quing Yang 392 G>T (0.8%). Sixty eight percent (88/125) neonates in this study had neonatal jaundice and 29.7% developed hyperbilirubinemia >250 micromol/l. The incidence of hyperbilirubinemia >250 micromol/l was higher in G6PD Kaiping (43.8%) than G6PD Canton (22%) (p< 0.05). There was no significant difference in the incidence of neonatal jaundice, mean serum bilirubin, mean age for peak serum bilirubin, percentage of babies requiring phototherapy and mean duration of phototherapy between the two major variants. None of the 88 neonates required exchange transfusion. In conclusion we have completely characterized the molecular defects of a group of Chinese G6PD deficiency in Malaysia. The mutation distribution reflects the original genetic pool and limited ethnic admixture with indigenous Malays.
We performed DNA analysis using cord blood samples on 86 male Malay neonates diagnosed as G6PD deficiency in the National University of Malaysia Hospital by a combination of rapid PCR-based techniques, single-stranded conformation polymorphism analysis (SSCP) and DNA sequencing. We found 37.2% were 871G>A (G6PD Viangchan), 26.7% were nt 563 C>T (G6PD Mediterranean) and 15.1% were 487G>A (G6PD Mahidol) followed by 4.7% 1376G>T (G6PD Canton), 3.5% 383T>C (G6PD Vanua Lava), 3.5% 592C>T (G6PD Coimbra), 2.3% 1388G>A (G6PD Kaiping), 2.3% 1360C>T (G6PD Union), 2.3% 1003G>A (G6PD Chatham), 1.2% 131C>G (G6PD Orissa) and 1.2% 1361G>A (G6PD Andalus). Seventy-one (82.6%) of the 86 G6PD-deficient neonates had neonatal jaundice. Fifty seven (80%) of the 71 neonates with jaundice required phototherapy with only one neonate progressing to severe hyperbilirubinemia (serum bilirubin >340 micromol/l) requiring exchange transfusion. There was no significant difference in the incidence of neonatal jaundice, mean serum bilirubin level, mean age for peak serum bilirubin, percentage of babies requiring phototherapy and mean number of days of phototherapy between the three common variants. In conclusion, the molecular defects of Malay G6PD deficiency is heterogeneous and G6PD Viangchan, Mahidol and Mediterranean account for at least 80% of the cases. Our findings support the observation that G6PD Viangchan and Mahidol are common Southeast Asian variants. Their presence in the Malays suggests a common ancestral origin with the Cambodians, Laotians and Thais. Our findings together with other preliminary data on the presence of the Mediterranean variant in this region provide evidence of strong Arab influence in the Malay Archipelago.
Neonatal screening for G6PD deficiency has long been established in many countries. The aim of the study was to determine whether the routine semiquantitative fluorescent spot test could detect all cases of G6PD deficiency, including those cases with partial deficiency (residual red cell G6PD activity between 20-60% of normal). We compared the results of G6PD screening by the semiquantitative fluorescent spot test and quantitative G6PD activity assay on a group of 976 neonates and 67 known female heterozygotes. The values for mean G6PD activity of G6PD-normal neonates and 293 healthy adult females were determined. There was no significant difference in the mean normal G6PD activity between the two racial groups in the neonates (669 Malays, 307 Chinese) and in the 293 healthy adult females (150 Malays, 143 Chinese) group. The values for the upper limits of total deficiency (20% of normal residual activity) for neonates and adult females were 2.92 U/gHb and 1.54 U/gHb, respectively. The upper limits of partial deficiency (60% of normal residual activity) were 8.7 U/gHb and 4.6 U/gHb respectively. The prevalence of G6PD deficiency among the male neonates was 5.1% (26) by both the fluorescent spot test and the enzyme assay method. The G6PD activity levels of all 26 cases of G6PD-deficient male neonates were < 20% normal (severe enzyme deficiency). In the female neonate group, the frequency of G6PD deficiency was 1.3% (6 of 472) by the fluorescent spot test and 9.35% (44 of 472) by enzyme assay. The 6 cases diagnosed as deficient by the fluorescent spot test showed severe enzyme deficiency (< 2.92 U/gHb). The remaining 38 female neonates had partial enzyme deficiency and all were misdiagnosed as normal by the fluorescent spot test. In the female heterozygote group, G6PD deficiency was diagnosed in 53% (35 of 67) by enzyme assay and in 7.5% (4 of 67) of cases by the fluorescent spot test. The 4 cases detected by fluorescent spot test had severe enzyme deficiency (<1.6 U/gHb). The remaining 31 (46.3%) cases, diagnosed as normal by fluorescent spot test, showed partial G6PD deficiency. In conclusion, we found that the semiquantitative fluorescent spot test could only diagnose cases of total G6PD deficiency and misclassified the partially-deficient cases as normal. In this study, the overall prevalence of G6PD deficiency was 3.28% by the semiquantitative fluorescent spot test and 7.17% by enzyme assay. This means that 3.9% of G6PD-deficient neonates were missed by the routine fluorescent spot test and they were found to be exclusively females. This study demonstrates a need to use a method that can correctly classify female heterozygotes with partial G6PD deficiency. The clinical implication is that these individuals may be at risk of the hemolytic complication of G6PD deficiency.