Clinical utility of targeted gene enrichment and sequencing technique in the diagnosis of adult hereditary spherocytosis

Abstract

Background: The present study aimed to use the targeted capture and sequencing technique to diagnose adult hereditary spherocytosis (HS). These results were compared with clinical features and laboratory examinations to explore the diagnosis of HS.

Methods: Whole blood and clinical data from ten patients with HS were collected. Genomic DNA was extracted, and a library was prepared. Exomes of patients with ten HS-related genes encoding red cell membrane skeleton protein were captured and sequenced. Bioinformatics analyses were carried out throughout the 1000 Genomes Project, ExAC, dbSNP147, and 1000 Normal Han Population databases.

Results: Gene mutations were found in 9 out of 10 cases of HS. Our data validation showed 90% specificity. Three types of gene mutations were found, including 6 cases of SPTB, 3 cases of ANK1, and 2 cases of SLC4A1. There were 4 mutation forms, including nonsense mutation, missense mutation, shear mutation, and code shift mutation, all of which were new, heterozygous mutations. These variations were predicted to be pathogenic in four databases.

Conclusions: Our data demonstrate that targeted gene enrichment and sequencing methods were an efficient tool for determining genetic etiologies of red blood cell (RBC) membrane disorders and can facilitate accurate diagnosis and genetic counseling. They are also in good agreement with the clinical results.

Keywords: ANK1; Hereditary spherocytosis (HS); SLC4A1; SPTB; targeted capture.

Figure 1

Family tree, ANK1 mutation, light microscope (Wright-Giemsa stain, ×1,000) and scanning electron images (4,000×; scale bar =10 µm) of RBC morphology in No.1 (father, I-2), 2 (daughter, II-3), 3 (son, II-1).

Figure 2

Family tree, SLC4A1 mutation, light microscope (Wright-Giemsa stain, ×1,000), and scanning electron images (4,000×; scale bar =10 µm) of RBC morphology in No.4 (mother, I-2), and 5 (son, II-4).

Figure 3

Family tree, SPTB mutation, light microscope (Wright-Giemsa Stain,×1,000) and scanning electron images (4,000×; scale bar =10 µm) of RBC morphology in No.6 (second brother, II-5) and 7(first brother, II-2).

Figure 4

Prediction map of the three-dimensional structure of protein (protein structure prediction Website-SWISS-MODEL). (A) (ANK1): there is an additional hydrogen bond between 275 P → R and 278 amino acids, and the side chain of amino acids has also changed. (B) (SPTB): the side chain of amino acid changed when the amino acid V → M was in the 71st position. (C) (SLC4A1): the side chain of amino acid changed when the amino acid V → M was in the 71st position.