FLUORESCENT SPOT TEST SCREENING FOR GLUCOSE-6-PHOSPHATE DEHYDROGENASE DEFICIENCY IN DALMATIA

V. Krzelj L. Balarin, S. Zlodre, J. Terzic, J. Jaksic, E. Marusic, B. Petric.

Department of Pediatrics, Clinical Hospital Split, Croatia and Medical School of the University, Split, Croatia and Department of Pediatrics, General Hospital, Sibenik, Croatia

OBJECTIVE: Glucose-6-phosphate dehydrogenase (G-6-PD) deficiency is one of the commonest enzymopathy in humans. Acute acquired hemolytic anemia, congenital nonspherocytic hemolytic anemia, favism and neonatal hyperbilirubinaemia are the most frequently clinical expressions of G-6-PD deficiency. High incidence of G-6-PD deficiency in the Mediterranean area is a result of selective advantage in an environment ofholoendemic malaria. Dalmatia is a region of Croatia which lies on the East Coast of the Adriatic Sea, i.e. part of Mediterranean basin and the prevalence of G-6-PD deficiency has not been determined yet. The International Committee of Standardization in Hematology has recommended the fluorescent spot test as the most acceptable method for screening control. The test is reliable for the diagnosis of hemizygotes, homozygotes and heterozygotes with a higher degree of the G-6-PD deficiency . The objective of this study was to determine the incidence of glucose-6-phosphate dehydrogenase G-6-PD deficiency in Dalmatia and the reliability of the fluorescent spot test.

METHODS: The principle of the fluorescent spot test depends upon the fluorescence of NADPH under long-wave UV light. The reduction of NADP⁺ to NADPH occurs in the presence of G-6-PD and the rate of NADPH formation is proportional to G-6-PD activity. The fluorescent spot test was used to analyze 2726 randomly selected blood samples of high school boys and girls in Dalmatia. They represent 10% of the whole high school population. Screening for G-6-PD deficiency was done in 1340 (49%) male and 1386 (51%) female pupils. Fluorescence reading was performed at the beginning and in the 3^rd, 6^th, 10^th and 25^th minutes of incubation. Results were classified in three groups: "bright fluorescence" (BF), "weak fluorescence" (WF), and "no fluorescence" (NF). All NF and WF samples in the 3^rd minute were quantitatively measured using spectrophotometric method.

RESULTS: G-6-PD deficiency was found in 12 (0.44%) of the samples. The incidence was 0.75% in males, and 0.14% in females. All NF samples at fluorescent spot test were G-6-PD deficiency. WF in the 3^rd minute of the incubation was present in 33 (1.2%) examinees, and only 2 (6%) were true positive. The fluorescence reading in the 10^th minute of incubation omits 5 (41%) of the G-6-PD deficiency samples. These individuals were moderate G-6-PD deficient.

CONCLUSIONS: The study results show that frequency of G-6-PD deficiency in Dalmatia is 0.44%. Moderate enzyme deficiency can be found with an early fluorescence reading.