Detection of the Prevalence of Sickle cell disease in Tharu Population of For-Western Nepal by RFLP Method

Abstract

Sickle cell disease (SCD) is an autosomal recessive disorder of hemoglobin (Hb) affecting individuals of common malarial region in the world. The genetic cause of SCD is a transversion mutation at 6th codon (GAG→GTG) of β-subunits of hemoglobin molecule known as sickle hemoglobin (HbS). The pathological complications in SCD arise due to HbS polymerization causing vaso-occlusion and hemolytic anemia which secondarily lead to other presentations. Because of poor health education and not well defined symptoms, the management procedure is costly, tedious and sensitive for developing country like Nepal. In this study, 116 samples were randomly collected from two Tharu sub-groups (Rana and Dangaura) of Kailali and Kanchanpur districts of Nepal. Initial screening of HbS was performed by hemoglobin solubility test, out of which only 26 showed positive results. To confirm the preliminary results genomic DNA was extracted and the targeted region of βglobin (539 bp) was PCR amplified. Validation of the point mutation that causes sickle cell disease was performed by restriction digestion of the amplified products. Twenty five samples were confirmed to be sickle cell carrier (heterozygous with only six exhibiting sickle cell trait) and none of the sample was found to be homozygous for the disease. The frequency of HbS carrier cases in male i.e. 13/60 (21.67%) was almost similar to that of female i.e.12/60 (21.43%). The highest frequency of HbS carrier cases were from >40 years groups that is 6/23 (26.08%) and least frequency from <20 years groups that is 2 (11.11%). Though the studied Tharu subgroups were originated from different geographical area and marriage practice, these groups were restricted and the distribution of HbS gene was found on both groups of people. The distribution of HbS was found to be higher in Rana compared to Dangaura. It demands a rapid, cheap and sensitive technique to perform the screening of the individuals living in the areas of high risks. RFLP technique has been widely used for detection of sickle cell disease. It can be performed in such disease risk population before the marriage and for the prenatal diagnosis of risks couple to predict, prevent and to reduce the frequency of diseased child born in near future. Keywords: Sickle cell disease, Tharu population, β-globin, hemoglobin solubility test, PCR- RFLP, Prenatal diagnosis.