Abstract
BackgroundThe distribution of the enzymopathy glucose-6-phosphate dehydrogenase (G6PD) deficiency is linked to areas of high malaria endemicity due to its association with protection from disease. G6PD deficiency is also identified as the cause of severe haemolysis following administration of the anti-malarial drug primaquine and further use of this drug will likely require identification of G6PD deficiency on a population level. Current conventional methods for G6PD screening have various disadvantages for field use.MethodsThe WST8/1-methoxy PMS method, recently adapted for field use, was validated using a gold standard enzymatic assay (R&D Diagnostics Ltd ®) in a study involving 235 children under five years of age, who were recruited by random selection from a cohort study in Tororo, Uganda. Blood spots were collected by finger-prick onto filter paper at routine visits, and G6PD activity was determined by both tests. Performance of the WST8/1-methoxy PMS test under various temperature, light, and storage conditions was evaluated.ResultsThe WST8/1-methoxy PMS assay was found to have 72% sensitivity and 98% specificity when compared to the commercial enzymatic assay and the AUC was 0.904, suggesting good agreement. Misclassifications were at borderline values of G6PD activity between mild and normal levels, or related to outlier haemoglobin values (<8.0 gHb/dl or >14 gHb/dl) associated with ongoing anaemia or recent haemolytic crises. Although severe G6PD deficiency was not found in the area, the test enabled identification of low G6PD activity. The assay was found to be highly robust for field use; showing less light sensitivity, good performance over a wide temperature range, and good capacity for medium-to-long term storage.ConclusionsThe WST8/1-methoxy PMS assay was comparable to the currently used standard enzymatic test, and offers advantages in terms of cost, storage, portability and use in resource-limited settings. Such features make this test a potential key tool for deployment in the field for point of care assessment prior to primaquine administration in malaria-endemic areas. As with other G6PD tests, outlier haemoglobin levels may confound G6PD level estimation.
Highlights
The distribution of the enzymopathy glucose-6-phosphate dehydrogenase (G6PD) deficiency is linked to areas of high malaria endemicity due to its association with protection from disease
Erythrocytes with insufficient G6PD are unprotected against oxidative injury, and individuals with G6PD deficiency may develop haemolytic anaemia in response to a number of stresses, including infection and exposure to medications such as the 8 amino-quinoline, primaquine [17]
WST8/1-methoxy PMS test use in a field setting Timeframe and temperature storage conditions affect assay performance Bloodspot storage Following collection, a random selection of 150 Filter paper blood spots (FPBS) were stored at two different temperatures (4°C and 24°C), and assayed at various days (1, 2, 4, 5, 6, 9, 10) to determine optimal storage times before degradation of G6PD occurs and risk of misclassification increases
Summary
The distribution of the enzymopathy glucose-6-phosphate dehydrogenase (G6PD) deficiency is linked to areas of high malaria endemicity due to its association with protection from disease. Glucose-6-phosphate dehydrogenase (G6PD) is an X-linked recessive hereditary disorder that currently affects 200–400 million people worldwide, with over 160 mutations identified [3,4,5,6] and there is pronounced geographical overlap between areas of G6PD deficiency prevalence and malaria endemicity [2,7,8,9,10,11,12,13,14,15]. Given the risk of haemolysis in G6PD deficient individuals, and the genetic and phenotypic variability of G6PD deficiency across geographic areas where primaquine treatment is considered, estimation of G6PD enzyme function prior to drug administration is recommended [23]. Primaquine therapy without prior determination of G6PD enzyme function, perhaps due to a lack of reliable tests, is thought to be common [24]
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