Identification of double heterozygous -alpha(4.2I)/-alpha(4.2II) using third-generation sequencing

Objective: The 4.2 kb deletion (-alpha(4.2I)) is a common a(+)-thalassemia with a carrier rate, followed by the South-East Asian deletion (-(SEA)) and the 3.7 kb deletion (-alpha(3.7)/). There are few reports about 4.2 kb deletion sub-types. Herein, we present a patient with double heterozygous -alpha(4.2I)/-alpha(4.2II) who was identified using third-generation sequencing (TGS). Methods: Hematology and hemoglobin fraction analysis were carried out by complete blood count (CBC) and capillary electrophoresis (CE). Gap-PCR was used to detect the common deletional alpha-thalassemia, and multiple ligation-dependent probe amplification (MLPA) was performed to screen the large deletion. Sanger sequencing identified the variant. The different deletions were confirmed by TGS. Results: CBC showed the patient with microcytic hypochromic anemia, and CE indicated the presence of a Hb variant. Gap-PCR and MLPA detected 4.2 kb deletion homozygotes (-alpha(4.2)/alpha(4.2)). The Hb variant was confirmed as Hb Q-Thailand by Sanger sequencing. The patient was identified as compound heterozygous of 4.2 kb deletion and Hb Q-Thailand (-alpha(4.2)/-alpha(4.2-Q-Thailand), -alpha(4.2I)/-alpha(4.2II)) using TGS. Conclusions: Hb Q-Thailand (-alpha(4.2-Q-Thailand)/) complex 4.2 kb deletion heterozygote (-alpha(4.2)/) is easily misdiagnosed as 4.2 kb homozygous using Gap-PCR and MLPA. The TGS enables the identification of the two different 4.2 kb deletion sub-types.

Automated summary

Using third-generation sequencing (TGS), a patient with double heterozygous -alpha(4.2I)/-alpha(4.2II) was identified, indicating the ability of TGS to detect different sub-types of the 4.2 kb deletion.