Co-inheritance of α-and β-thalassemia in Khuzestan Province, Iran

Abstract

Background and aims: β-thalassemia is one of the most frequent hemoglobinopathies and single gene disorders in Iran. About 13 β globin mutations encompass 70–90% of mutations spectrum in Iran, the rest are rare or unknown. People who do not produce enough alpha globin protein chains have α-thalassemia. This is commonly found in Africa, the Middle East, India, Southeast Asia, southern China, and occasionally the Mediterranean region. There are normally four α globin genes, two on each chromosome 16. Individuals who have one or two abnormal alpha globin genes have α-thalassemia trait. The aim of this study was to detect α-thalassemia in β-thalassemia carriers during prenatal screening.Materials and methods: A total of 158 couples were diagnosed to be discordant α- and β-thalassemia carriers. We used the routine screening for thalassemia which includes full blood counts and indices, hemoglobin electrophoresis and measurement of Hb A2 level. The standard diagnostic marker for β-thalassemia is elevation of the Hb A2 level (>3·5%). Low mean corpuscular volume (MCV) and mean cell hemoglobin (MCH) with a normal Hb A2 indicate an α-thalassemia carrier. Staining for HbH inclusion bodies is also carried out as part of the screening for α thalassemia. The 59 and 39 ends of the breakpoint regions of the −α4·2 allele and the normal homologous segments were sequenced in selected individuals.Results: Of the 158 β-thalassemia partners, seven (4·4%) were found to have co-inheritance of α+-thalassemia, and three (1·9%) found to have co-inheritance of α0-thalassemia. Two pregnancies affected with Hb Bart's hydrops fetalis were terminated in the 158 couples. A sequence haplotype was found in all of the five Iranian −α4·2 thalassaemia alleles studied. Based on these findings, a novel polymerase chain reaction (PCR)/denaturing high performance liquid chromatography (DHPLC) assay was developed for rapid genotyping of the −α4·2 allele instead of traditional Southern blotting or Gap-PCR. This method involves amplification of the α globin target sequence encompassing these four polymorphic sites, followed by a partially denaturing HPLC analysis using the transgenomic WAVE DNA fragment analysis system. The major genotypes (−α4·2/αα, −α4·2/--MED, −α3·7/αα, −α3·7/−α3·7, αα/--MED and αα/αα) could be distinguished through the characteristic chromatograms generated by the WAVE system.Discussion: The results showed that molecular analysis must be used for accurate diagnosis of double heterozygotes in couples presumed to be discordant for α- and β-thalassemia on hematologic testing. The accuracy of this technique was evaluated blindly, and the results were 100% (40 of 40) concordant with the genotypes previously characterized by Southern blotting or Gap-PCR.