Molecular characterization of β thalassaemia intermedia in Sri Lanka: Genotypic heterogeneity and phenotypic diversity

Abstract

Previous studies on thalassaemia in Sri Lanka revealed that one third of patients attending thalassaemia clinics have intermedia phenotype and has shown to be presented with a wide pheno$pic diversity. The majority of them have Hb E/J3 thalassaemia, but a significant minority has been shown to have co-inheritance of {3 thalassaemia minor with extra a globin genes. In this study, a group of (3 thalassaemia intermedia patients without HbE/p thalassaemia was systematically identified for further investigations. Fifty unrelated thalassaemia intermedia patients were identified from the five main thalassaemia centres in the island with mostly presented to the adult thalassaemia clinic at Ragama, Sri Lanka; Patients were assessed clinically and categorized into mild, moderate and severe phenotypic groups according to the clinical severity. Deoxyribonucleic acid (DNA) analysis was performed by Southern blot analysis, Gap polymerase chain reaction (PCR), multiplex ligation-dependant probe amplifications (MLPA) and DNA sequence analysis of PCR products. There were twenty six patients in mild and twelve patients each in moderate and severe phenotypic group in the p thalassaemia intermedia study. Ages were ranged from 5-65 years. Seventeen (34%) of the total thalassaemia intermedia population were homozygous or compound heterozygous for p mutations. Five of the homozygotes carried two mild p alleles including a rare promoter mutation - 90 C-»T, Hb variant alleles of Hb G-Szuhu and Hb G-Coushatta. Nine of the seventeen had inherited with two severe p alleles with a globin gene deletions either one or two. Of the other three individuals in this group one had Xmn I y +/+ polymorphism. In the other two individuals, phenotypes were unable to explain with the molecular analysis data obtained through this study. Individuals who had inherited with two mild |3 alleles or a severe allele with either Hb G-Szuhu or Hb G-Coushatta invariably had a mild phenotype while those individuals who had inherited two severe p alleles with a globin gene deletions invariably had a severe phenotype. Thirty three (66%) q£ the total thalassaemia intermedia population were heterozygous for P mutations, of which twenty eight carried excess a globin gene and had a mild to moderate phenotype. Five phenotypes in this group were unable to explain with the molecular analysis data obtained through this study. About forty three (86%) of the cases found with p thalassaemia intermedia had mutations on the a and p globin gene clusters that would account for their clinical presentation with co-inheritance of additional cc-globin genes and p thalassaemia minor the most common finding. However, seven (14%) of the study population remains to be explained that will result in an extraordinary diversity of lesions revealed within this island cohort.