Complexity of beta thalassemia diagnosis: experience of a case study

Background: Beta thalassemia is an autosomal recessive disorder that is caused by reduced synthesis of beta-globin chains. Mutations in the HBB gene are responsible for the disease. Most of the common genetic defects in β-thalassemia occur due to point mutations and small deletions in the β-globin gene.Method: Proband and his family were referred to Kawsar Human Genetics Research Center (KHGRC, Tehran, Iran). The proband was a 10 year old boy who underwent blood transfusion for three times during his life. Blood samples were collected, and genomic DNA was extracted using salting out procedure. DNA analyses were performed using ARMS PCR (Amplification Refractory Mutation System), MLPA (Multiplex Ligation-dependent Probe Amplification) and Sanger sequencing. Results: According to the CBC electrophoresis result, the proband’s mother was normal (MCV: 96.4, MCH: 31.6, Hb: 13.1, HbA2: 3, HbF: 0.4); the proband’s father was carrier of beta thalassemia (MCV: 63.6, MCH: 19.7, Hb: 14, HbA2: 4.9, HbF: 0). Using ARMS-PCR showed homozygote mutation in the proband and heterozygote mutation in probands’s father. Both mutations were the same (frameshift (–AA)) but no mutation were found in the mother. This could be four possibilities for it: 1. Simultaneous occurrence of Alpha triplication with heterozygote frameshift (–AA). 2. Denovo deletion or duplication in HBB 3. Non-Parental Event 4. Denovo frameshift (–AA) mutation. 5. Maternal gonadal mosaicism. Possibility of 1-3 was rule out using MLPA and Identifiler kit. The only explanation could be denovo frameshift mutation in mother or gonadal mosaicism which needs more investigation.Conclusion: The result of this case study suggested that complete molecular testing must be performed in complex cases like what we have in this study which can help the family in prenatal diagnosis of their future embryos.

Keywords:

Beta thalassemia , Beta globin gene , Iran , ARMS-PCR , MLPA