A "One-Stop" Screening Protocol for Haemoglobinopathy Traits and Iron Deficiency in Sri Lanka.

Introduction: The high frequencies of carriers of severe haemoglobinopathies and of iron deficiency in Southeast Asia require reliable and affordable tests to improve on current screening procedures. Objectives: We evaluate a "one stop" approach using the THALCON dichlorophenolindophenol (DCIP) and one-tube osmotic fragility (OF) tests and measurement of Zinc Protoporphyrin (ZPP) to detect and distinguish HbE and ß-thalassaemia traits from iron deficiency. We compare findings with current screening practice in Sri Lanka that relies on the identification of low mean red cell volume and/or mean red cell hemoglobin for this purpose. Methods: Between November 2017 and May 2018, we undertook a cross-sectional survey of secondary school students in Gampaha district, Sri Lanka. The THALCON-DCIP and OF tests were compared to capillary electrophoresis (CE), used as a gold standard to detect haemoglobinopathies. ZPP was measured in whole blood. Plasma ferritin and C-reactive protein (CRP) were measured in students with a raised ZPP concentration. Results: We collected venous blood samples from 1,324/1,332 (99.4%) students. The median age of the students was 17 (IQR 16-18) years, all were Sinhalese and 814/1,324 (61.5%) were female. CE identified 3 students with HbE trait and 26 students with ß-thalassaemia trait. The THALCON-DCIP test was positive only in the 3 students with HbE (sensitivity 100%, 95% CI 29.2-100.0; specificity 100%, 95% CI 99.7-100.0). The THALCON-OF test identified all 26 students with ß-thalassaemia trait (sensitivity = 100%, 95% CI 86.8-100.0) and 287 students with a normal CE result (specificity = 77.9%; 95% CI 75.5-80.1). It was also positive in 2/3 (66.7%) students with HbE trait. Iron deficiency (ZPP > 70 µmol/mol heme) was present in 118/1,240 (9.5%) students with a normal hemoglobin genotype, all of whom had plasma ferritin <15 ng/ml and CRP <5 mg/L. Conclusion: This one-stop approach offers reliable and affordable population screening for both haemoglobinopathy traits and iron deficiency in resource-limited settings where these conditions are common and ensures that iron supplements are targeted only to those who require them. Further work is warranted to refine the OF test to reduce the number of false positive results.

Haemoglobin variants, iron status and anaemia in Sri Lankan adolescents with low red cell indices: A cross sectional survey.

Iron deficiency complicates the use of red cell indices to screen for carriers of haemoglobin variants in many populations. In a cross sectional survey of 7526 secondary school students from 25 districts of Sri Lanka, 1963 (26.0%) students had low red cell indices. Iron deficiency, identified by low serum ferritin, was the major identifiable cause occurring in 550/1806 (30.5%) students. Low red cell indices occurred in iron-replete students with alpha-thalassaemia including those with single alpha-globin gene deletions. Anaemia and low red cell indices were also common in beta-thalassaemia trait. An unexpected finding was that low red cell indices occurred in 713 iron-replete students with a normal haemoglobin genotype. It is common practice to prescribe iron supplements to individuals with low red cell indices. Since low red cell indices were a feature of all forms of α thalassaemia and also of iron deficiency, in areas where both conditions are common, such as Sri Lanka, it is imperative to differentiate between the two, to allow targeted administration of iron supplements and avoid the possible deleterious effects of increased iron availability in iron replete individuals with low red cell indices due to other causes such as α thalassaemia.

Mutations in the chromatin-associated protein ATRX.

ATRX belongs to the SNF2 family of proteins, many of which have been demonstrated to have chromatin remodeling activity. Constitution mutations in the X-encoded gene give rise to alpha thalassemia mental retardation (ATR-X) syndrome and a variety of related conditions that are often associated with profound developmental delay, facial dysmorphism, genital abnormalities, and alpha thalassemia. Acquired mutations in ATRX are observed in the preleukemic condition alpha thalassemia myelodysplastic syndrome (ATMDS). Mutations in ATRX have been shown to perturb gene expression and DNA methylation. This is a comprehensive report of 127 mutations including 32 reported here for the first time. Missense mutations are shown to cluster in the two main functional domains. The truncating mutations appear to be "rescued" to some degree and so it appears likely that most if not all constitutional ATRX mutations are hypomorphs.

A novel mutation in the last exon of ATRX in a patient with alpha-thalassemia myelodysplastic syndrome.

We describe a patient with acquired alpha-thalassemia myelodysplastic syndrome (ATMDS). A previously healthy 66-year-old man presented with hemoglobin of 9.3 g/dL, mean corpuscular volume 59 fL, and a bone marrow aspirate with increased erythroid precursors and hypolobulated megakaryocytes. Hemoglobin H inclusions were seen in most red cells after 1% brilliant cresyl blue supravital stain of the peripheral blood. At the molecular level, we identified of a novel mutation in the most 3' exon of the ATRX gene (CGA-->TGA substitution in codon 2407) resulting in a premature termination codon (p.R2407X). This case provides further evidence for a link between ATRX mutations and ATMDS, and suggests a possible role for the conserved Q-box element in ATRX function.