Hb E/β-thalassemia shows phenotypic heterogeneity with presentations ranging from mild to severe . The reasons for such variability in presentations are not completely understood and are an active area of research . To that end, we studied the clinical and genetic profile and explored their association in Hb E/β-thalassemia patients in Bangladesh from a specialized referral hematology unit of a tertiary care hospital.
We found that nearly two-thirds of patients had moderate to severe disease according to the Mahidol Scoring System. Our finding nearly corresponds to the original work that produced the scale at Mahidol University, Thailand, which found that more than two-thirds of patients had moderate to severe disease in a large sample of 950 patients . However, an Indonesian study found that nearly four-fifth of patients had moderate to severe disease using the same scale, which is higher than that found in our study. Although no such classification was done previously among similar patients in Bangladesh, the Mahidol score was used to classify β-thalassemia intermedia patients in the country. The study by Mannan and his colleagues found that 35.3% had moderate disease, and 6% had severe disease. Various genetic interactions underlie intermediate forms of β-thalassemia and are less severe phenotypically than the forms compounded by hemoglobin E variants .
We noted a wide age range at which the diagnosis was made, and transfusions were started among the patients. Earlier age at presentation was usually associated with a higher severity score requiring a higher number of transfusions and a higher proportion of growth retardation and hepatosplenomegaly among Hb E/β-thalassemia patients. Olivieri and colleagues  extensively studied the phenotypic variability of such a group of thalassemic patients, which endorses our findings. Male patients were more frequent in our study, congruent with previous estimates .
The IVS-1-5 (G > C) mutation was the most common mutation found in more than 50% of the participants of this study. Mutational analysis of β-thalassemic individuals by Ayub et al.  reported that the splice site mutation IVS-1-5 (G > C) was the most common mutation in Bangladesh. Moreover, this mutation happens to be the most prevalent among thalassemic patients of South Asia . However, unlike previous studies [6, 18] that argued against any association of the mutational specificity of the HBB gene with the phenotypic variability of Hb E/β-thalassemia, we noted that the IVS-1-5 (G > C) mutation was predominantly associated with moderate to severe disease. In particular, all severe phenotypes in our study were associated with this mutation. This relationship could be explained by the fact that IVS-1-5 (G > C) mutations produce β+-thalassemia alleles leading to defective splicing of mRNA and impaired globin chain production . Which, when compounded by Hb E mutation (codon 26 G > A), leads to impaired Hb A production, increased Hb A2 and Hb E synthesis, and an overall decrease in hemoglobin level . However, one might argue that the presence of IVS-1-5 (G > C) mutations across all severities of presentation in Hb E β-thalassemic patients, as described by Olivieri et al. , points against such an association. However, their findings could be explained by other secondary and tertiary modifiers, including other genetic and environmental attenuating factors which might be responsible for such phenotypic heterogeneity of the same mutation . On the other hand, classification schemes used for severity grading might have affected the distribution seen in Olivieri et al. . As we could not characterize the α-globin gene mutation among the participants, an exploration of the concomitant taming effect of alpha-mutation on the severity  was not possible in our study. Hence, further large sample studies are suggested to conclude any association.
As mild diseases could often be managed without transfusion preventing iron overload in these patients, a mutational analysis of the Hb E/β-thalassemia patients could guide individually tailored management for them.
Finally, one case among our participants needs particular attention. Interestingly, we noted that this case had a Mahidol score classified as severe but needed occasional blood transfusion only. This patient had a low steady-state hemoglobin of 5 g/dl, early age at first presentation (6 years), large spleen size (18 cm), severe growth retardation (< 3rd percentile), and an IVS-1-5 (G > C) mutation. Interestingly, the patient started blood transfusion at the age of 17 years, which indicates a lack of awareness and follow-up in the patient’s part and might explain why the patient developed overt clinical presentation. However, a general lack of awareness about thalassemia and inadequate access to dedicated treatment facilities due to location and/or poor socioeconomic condition often led to delayed presentation or follow-up at hospitals. Hossain et al. , in a review of thalassemia in South Asian countries, noted that health awareness of the general population is very poor among the general population, and many thalassemia patients may die without knowing about their disease. However, this case also shows the variety of presentation Hb E/β-thalassemia patients can have and emphasize the importance of early detection and regular follow-up.
The major limitation of our study is the small sample size and characterization of patients from a single center. Another limitation is the inability to characterize α-globin gene mutations among the patients. However, our study is one of the earliest attempts to explore any association between phenotypic severity and mutations of the HBB gene among Hb E/β-thalassemia patients, and the results could aid in determining management strategies of patients carrying both Hb E and β-thalassemia mutations.