Gene Mutation Spectrum among Alpha-Thalassaemia Patients in Northeast Peninsular Malaysia

Abstract

(1) Background: Alpha (α)-thalassaemia is a genetic disorder that affects 5% of the world population. Deletional or nondeletional mutations of one or both HBA1 and HBA2 on chromosome 16 will result in reduced production of α-globin chains, a component of haemoglobin (Hb) that is required for the formation of red blood cells (RBCs). This study aimed to determine the prevalence, haematological and molecular characterisations of α-thalassaemia. (2) Method: The parameters were based on full blood count, high-performance liquid chromatography and capillary electrophoresis. The molecular analysis involved gap-polymerase chain reaction (PCR), multiplex amplification refractory mutation system-PCR, multiplex ligation-dependent probe amplification and Sanger sequencing. (3) Results: With a total cohort of 131 patients, the prevalence of α-thalassaemia was 48.9%, leaving the remaining 51.1% with potentially undetected α gene mutations. The following genotypes were detected: -α^3.7/αα (15.4%), -α^4.2/αα (3.7%), --^SEA/αα (7.4%), α^CSα/αα (10.3%), α^Adanaα/αα (0.7%), α^{Quong Szeα}/αα (1.5%), -α^3.7/-α^3.7 (0.7%), α^CSα/α^CSα (0.7%), -α^4.2/α^CSα (0.7%), -^SEA/α^CSα (1.5%), -^SEA/α^{Quong Sze}α (0.7%), -α^3.7/α^Adanaα (0.7%), --^SEA/-α^3.7 (2.2%) and α^CSα/α^Adanaα (0.7%). Indicators such as Hb (p = 0.022), mean corpuscular volume (p = 0.009), mean corpuscular haemoglobin (p = 0.017), RBC (p = 0.038) and haematocrit (p = 0.058) showed significant changes among patients with deletional mutations, but not between patients with nondeletional mutations. (4) Conclusions: A wide range of haematological parameters was observed among patients, including those with the same genotype. Thus, a combination of molecular technologies and haematological parameters is necessary for the accurate detection of α-globin chain mutations.

Keywords: genotype; globin; molecular analysis; mutation; prevalence; α-thalassaemia.

Figure 1

Representative gel image for α deletional mutations: Lane M: 100 bp plus ladder; Lane 1: heterozygous 4.2 mutation (-α^4.2/αα); Lane 2: heterozygous 20.5 mutation (-α^20.5/αα); Lane 3: heterozygous SEA mutation (--^SEA/αα); Lane 4: heterozygous 4.2 mutation (-α^4.2/αα); Lane 5: heterozygous Fil mutation (--^FIL/αα); Lane 6: heterozygous 3.7 mutation (-α^3.7/αα); Lane 7: compound heterozygous (--^SEA/-α^3.7); Lane 8: negative control; Lane 9: heterozygous 4.2 mutation (-α^4.2/αα).

Figure 2

Representative gel image for α nondeletional mutations: Lane M: 100 bp ladder; Lane 1: initiation codon mutation; Lane 2: codon 30 mutation; Lane 3: codon 59 mutation (Hb Adana); Lane 4: codon 142 mutation (Hb Constant Spring); Lane 5–6: normal α; Lane 7: codon 142 mutation (Hb Constant Spring); Lane 8: negative control.

Figure 3

(a,b) A representative result of Hb Constant Spring by HPLC and CE analyses. The red arrow represents the Hb Constant Spring peak and zone in HPLC and CE respectively.