Haemoglobinopathies: Integrated Biochemical and Molecular Diagnosis in 5243 Patients
Abstract
1. Introduction
2. Materials and Methods
2.1. Patients
2.2. First-Level Screening
2.3. Second-Level Molecular Analysis
2.3.1. Genomic DNA Extraction
2.3.2. Molecular Analysis of Globin Genes
3. Results
4. Discussion
- NM_000518.5(HBB):c.118C>T (β039), detected in 44% of cases, typical of the Mediterranean region;
- NM_000518.5(HBB):c.93-21G>A (IVS-I-110), in 17.7%, common in Eastern Europe;
- NM_000518.5(HBB):c.92+6T>C (IVS-I-6), in 12.7%, prevalent in Eastern Europe and China;
- NM_000518.5(HBB):c.92+1G>A (IVS-I-1), in 12.3%, also typical of the Mediterranean basin.
- Among α-globin mutations, the most frequent were as follows:
- -α3·7 (NG_000006.1:g.34164_37967del3804), found in 48% of carriers, widespread in the Mediterranean and Middle East;
- α2 IVS1 -5nt (HbA2: c.95+2_95+6delTGAGG), in 15.4%, common in Morocco and the Middle East;
- -20.5 Kb (NG_000006.1:g.15164_37864del22701), in 14.2%;
- Triplicated α (NG_000001:g.34247_38050dup), in 11% of cases.
- (1)
- Serum iron and ferritin analysis;
- (2)
- NGS-based sequencing of the α-, β-, γ- and δ-globin genes in cases of isolated HbA2 elevation, suspected silent β-thalassaemia, or persistent diagnostic uncertainty.
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
Abbreviations
CNV | Copy number variation |
EDTA | ethylenediaminetetraacetic acid |
gDNA | Genomic DNA |
HbA | Haemoglobin A1, two α-chains and two β-chains |
HbA2 | Haemoglobin A2 |
HbF | Foetalhaemoglobin |
HBB | Haemoglobin Subunit Beta |
HbH | Haemoglobin H |
Hb Bart’s | Haemoglobin Bart’s |
HCT | Haematocrit |
HGVS | Human Genome Variation Society |
HPLC | High-performance liquid chromatography |
HPFH | Persistence of foetal haemoglobin |
MCV | Mean corpuscular volume |
MCH | Mean corpuscular haemoglobin |
MCHC | Mean corpuscular haemoglobin concentration |
NGS | Next-generation sequencing |
PLT | Platelet count |
RBC | Red blood cell count |
RDW | Red cell distribution width |
WHO | World Health Organization |
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Mutation HGVS Nomenclature Carriers of the α-thalassaemia Trait | Nomenclature Ithanet | RBC 106/μL | Hb g/dL | HCT % | MCV fL | MCH pg | MCHC g/dL | HBA2% | HBF% | |
---|---|---|---|---|---|---|---|---|---|---|
1 | NG_000006.1: g.34164_37967del3804 (n = 129) | -3.7 (tipo I) heterozygotes | 5.10 ± 0.58 | 13.1 ± 1.53 | 39.6 ± 4.66 | 77.8 ± 5.27 | 25.7 ± 2.03 | 33.0 ± 1.09 | 2.5 ± 0.84 | 1.3 ± 0.53 |
2 | HbA2: c.95+2_95+6delTGAGG (n = 41) | α2 IVS1 -5nt | 5.2 ± 0.71 | 12.4 ± 1.96 | 38.1 ± 5.06 | 74.0 ± 4.31 | 23.8 ± 2.01 | 32.3 ± 1.62 | 2.5 ± 1.11 | 0.8 ± 0.22 |
3 | NG_000006.1: g.15164_37864del22701 (n = 38) | -20.5 Kb double gene del | 5.4 ± 0.70 | 11.6 ± 1.20 | 36.0 ± 3.90 | 65.9 ± 6.60 | 21.7 ± 3.10 | 32.3 ± 0.91 | 2.3 ± 0.40 | 0.7 ± 0.30 |
4 | NG_000001: g.34247_38050dup (n = 29) | anti-3.7 (ααα−3.7) | 4.12 ± 0.52 | 12.7 ± 1.78 | 37.5 ± 4.03 | 81.7 ± 5.24 | 28.3 ± 3.32 | 33.7 ± 1.68 | 2.7 ± 0.63 | 1.7 ± 2.31 |
5 | HBA2: c.*+94A>G (n = 10) | α 2poly-A1 | 5.30 ± 0.45 | 12.4 ± 0.11 | 38.1 ± 0.23 | 72.6 ± 5.28 | 23.6 ± 1.87 | 32.4 ± 0.90 | 2.5 ± 0.23 | 0.6 ± 0.20 |
6 | NC_000016.10: g.169818_174075del (n = 10) | -4.2 | 5.02 ± 0.20 | 12.0 ± 0.75 | 40.3 ± 6.40 | 78.6 ± 6.37 | 24.1 ± 1.95 | 30.5 ± 5.22 | 2.3 ± 0.43 | 0.3 ± 0.25 |
7 | HbA2: c.427T>C (n = 5) | α 2 cd 142 Hb Constant Spring | 5.62 ± 0.43 | 13.0 ± 1.07 | 40.7 ± 3.93 | 71.8 ± 5.60 | 23.2 ± 1.38 | 32.3 ± 0.78 | 2.2 ± 0.21 | 0.4 ± 0.28 |
8 | HbA2: c.2T>C (n = 4) | α 2 init cd | 5.60 ± 0.77 | 13.3 ± 2.73 | 41.5 ± 4.82 | 78.6 ± 2.96 | 26.6 ± 0.50 | 32.5 ± 1.76 | 2.3 ± 0.50 | 0.7 ± 0.42 |
9 | NG_000006.1: g.24664_41064del 16401 (n = 1) | --MED double gene del | 5.50 | 12.1 | 37.0 | 68.9 | 22.4 | 32.0 | 3.5 | 0.1 |
Mutation HGVS Nomenclature Compound Heterozygotes and Homozygotes for the α-globin Genes | Nomenclature Ithanet | RBC 106/μL | Hb g/dL | HCT % | MCV fL | MCH pg | MCHC g/dL | HBA2 % | HBF % | HB variants | |
---|---|---|---|---|---|---|---|---|---|---|---|
1 | NG_000001:g.34247_38050 dup/HbA2:c.427T>C (n = 3) | Anti-3.7 (ααα-3.7)/α2 cd 142 Hb Constant Spring | 5.60 ± 1.32 | 13.2 ± 1.67 | 45.0 ± 0.82 | 75.2 ± 1.56 | 24.5 ± 1.06 | 32.6 ± 0.78 | 2.5 ± 0.14 | 0.4 ± 0.14 | ---- |
2 | NG_000006.1:g.34164_37967del3804/HBA2: c.*+94A>G (n = 1) | -3.7 (tipo I)/α 2 Poly A-1 (HBA2:c.*+94A>G) | 5.70 | 11.4 | 37.5 | 60.0 | 23.1 | 33.3 | 2.4 | 0.3 | ---- |
3 | HBA1:c.91G>C/NG_000006.1:g.34164_37967del3804 (n = 1) | α2 cd68 (C>G)/-3.7 (tipo I) HbG | 4.7 | 7.4 | 24.9 | 53.1 | 15.8 | 29.7 | ---- | ---- | 32.6 |
4 | NG_000006.1:g.34164_37967del3804 (n = 3) | -3.7 (tipo I) Omozigote | 5.42 ± 0.36 | 10.9 ± 0.81 | 34.5 ± 1.63 | 64.2 ± 3.64 | 20.3 ± 0.52 | 31.6 ± 1.56 | 2.7 ± 0.15 | 1.0 ± 0.10 | ---- |
Mutation HGVS Nomenclature Carriers of the β-thalassaemia Trait | Nomenclature Ithanet | RBC 106/μL | Hb g/dL | HCT % | MCV fL | MCH pg | MCHC g/dL | HBA2% | HBF% | |
---|---|---|---|---|---|---|---|---|---|---|
1 | NM_000518.5(HBB):c.118C>T (n = 210) | CODON 39 [C>T] | 5.76 ± 0.74 | 11.2 ± 1.39 | 35.0 ± 4.80 | 61.1 ± 5.10 | 19.6 ± 1.74 | 32.2 ± 1.72 | 5.4 ± 1.07 | 1.8 ± 2.63 |
2 | NM_000518.5(HBB):c.93-21G>A (n = 84) | IVS 1.110 [G>A] | 5.70 ± 0.63 | 11.8 ± 1.26 | 36.4 ± 3.83 | 63.8 ± 3.01 | 20.7 ± 1.42 | 32.2 ± 1.12 | 5.0 ± 0.67 | 1.2 ± 1.23 |
3 | NM_000518.5(HBB):c.92+6T>C (n = 60) | IVS 1.6 [T>C] | 5.76 ± 1.12 | 13.1 ± 3.91 | 39.0 ± 5.79 | 67.2 ± 7.7 | 22.6 ± 2.07 | 32.8 ± 0.97 | 3.8 ± 0.54 | 1.0 ± 0.38 |
4 | NM_000518.5(HBB):c.92+1G>A (n = 58) | IVS 1.1 [G>A] | 5.83 ± 0.70 | 11.1 ± 1.20 | 35.6 ± 3.60 | 64.2 ± 6.40 | 19.5 ± 0.8 | 30.7 ± 2.60 | 5.5 ± 0.4 | 1.3 ± 0.5 |
5 | NM_000518.5(HBB):c.316-106C>G (n = 22) | IVS 2.745 [C>G] | 5.60 ± 0.71 | 10.9 ± 1.60 | 33.8 ± 4.50 | 61.0 ± 4.23 | 19.5 ± 1.90 | 31.9 ± 1.53 | 5.1 ± 0.50 | 1.7 ± 1.5 |
6 | NM_000518.5(HBB):c.315+1G>A (n = 15) | IVS 2.1 [G>A] | 6.21 ± 0.61 | 12.1 ± 0.91 | 37.6 ± 3.56 | 61.2 ± 5.36 | 19.8 ± 1.63 | 32.4 ± 2.28 | 5.5 ± 0.88 | 1.6 ± 0.82 |
7 | NM_000518.5(HBB):c.-151C>T (n = 14) (silente) | -101 C>T (c.151C>T) | 4.63 ± 0.47 | 12.9 ± 1.32 | 38.3 ± 3.85 | 83.5 ± 2.62 | 28.8 ± 0.96 | 33.6 ± 0.89 | 3.7 ± 0.50 | 1.4 ± 0.98 |
8 | NM_000518.5(HBB):c.-137C>G (n= 3) (silente) | -87 [C>G] | 5.04 ± 0.86 | 11.8 ± 1.50 | 36, ± 4.53 | 72.2 ± 5.37 | 23.6 ± 1.30 | 32.7 ± 0.83 | 5.1 ± 1.06 | 2.55 ± 0.64 |
9 | NG_000007.3:g63632_7104 del (n = 3) | Hb Lepore BW | 5.40 ± 0.2 | 11.6 ± 0.60 | 32.6 ± 1.00 | 64.4 ± 0.90 | 22.1 ± 1.00 | 33.3 ± 0.4 | 2.7 ± 0.10 | 5.0 ± 0.60 |
10 | NM_000518.5(HBB):c.92+2T>A (n = 2) | IVS 1.2 [T>A] | 5.39 ± 0.10 | 10.5 ± 0.49 | 32.0 ± 0.26 | 59.8 ± 2.40 | 19.6 ± 0.77 | 33.0 ± 0.85 | 5.6 ± 0.14 | 1.55 ± 0.35 |
11 | NM_000518.5(HBB):c.47G>A (n = 2) | CODON 15 [TGG>TGA] | 5.31 ± 0.33 | 10.4 ± 0.64 | 32.9 ± 2.26 | 61.9 ± 0.49 | 19.7 ± 0.00 | 31.7 ± 0.21 | 6.20 ± 0.28 | 1.05 ± 0.35 |
Haemoglobin Variants HGVS Nomenclature | Nomenclature Ithanet | RBC 106/μL | Hb g/dL | HCT% | MCVfL | MCHpg | MCHCg/dL | HBA2% | HBF% | |
---|---|---|---|---|---|---|---|---|---|---|
1 | HBB:c.20A>T (n = 41) | CODON 6 [A>T] HbS | 4.49 ± 0.70 | 12.2 ± 2.40 | 35.0 ± 3.93 | 79.0 ± 6.70 | 27.2 ± 3.69 | 34.5 ± 1.19 | 3.3 ± 1.10 | 1.2 ± 1.03 |
2 | HBB:c.19G>A (n = 3) | CODON 6 [G>A] HbC | 4.22 ± 1.21 | 9.7 ± 1.57 | 34.5 ± 2.35 | 69.7 ± 3.04 | 24.0 ± 3.35 | 35.0 ± 2.25 | 2.8 ± 0.36 | 0.4 ± 0.51 |
3 | HBB:c.364G>C (n = 1) | HbD Punjab HbD | 4.70 | 12.2 | 37.4 | 79.9 | 26.3 | 37.7 | 2.36 | 0.37 |
4 | HBB:c.79G>A | HbE | 5.20 | 13.1 | 38.2 | 73.5 | 25.2 | 34.3 | 1.6 | |
5 | HBB:c.364G>A | Hb O-Arab | 5.21 | 13.4 | 39.4 | 74.0 | 25.4 | 34.4 | 3.7 | 0.8 |
HGVS Nomenclature Presence of both β- and α-thalassaemia traits | Nomenclature Ithanet | RBC 106/μL | HB g/L | HCT % | MCV fL | MCH pg | MCHC/dL | HBA2% | HBF % | |
---|---|---|---|---|---|---|---|---|---|---|
1 | NG_000006.1:g.34164_37967del3804/NM_000518.5(HBB):c.118C>T (n = 8) | α−3.7/CODON 39 | 5.33 ± 0.66 | 10.9 ± 1.37 | 33.5 ± 4.20 | 62.8 ± 3.03 | 20.5 ± 1.26 | 32.6 ± 1.14 | 5.3 ± 0.59 | 1.95 ± 1.16 |
2 | NG_000006.1:g.34164_37967del3804/HBB:c.20A>T (n = 3) | α−3.7/CODON 6 [A>T] | 4.41 ± 0.45 | 10.9 ± 0.46 | 33.4 ± 0.35 | 74.3 ± 5.01 | 26.9 ± 1.85 | 32.66 | 3 ± 0.85 | 0.7 ± 0.62 |
3 | HbA2: c.95+2_95+6delTGAGG/ NM_000518.5(HBB):c.118C>T (n = 3) | α2 IVS1 -5nt/CODON 39 | 5.77 ± 0.23 | 11.5 ± 0.61 | 35.4 ± 1.70 | 61.4 ± 0.45 | 19.4 ± 0.91 | 31.5 ± 1.23 | 5.7 ± 0.61 | 0.9 ± 0.83 |
4 | NG_000006.1:g.34164_37967del3804/NM_000518.5(HBB):c.93-21G>A (n = 2) | α−3.7/IVS 1.110 | 4.84 ± 0.70 | 13.1 ± 3.4 | 40.3 ± 8.2 | 85.5 ± 2.98 | 28.3 ± 4.32 | 32.6 ± 1.20 | 4.9 ± 1.70 | 1.6 ± 0.63 |
5 | NG_000001:g.34247_38050dup/NM_000518.5(HBB):c.93-21G>A (n = 2) | ααα−3.7/IVS 1.110 | 5.6 ± 0.46 | 11.0 ± 1.19 | 32.5 ± 3.17 | 57.3 ± 1.63 | 19.4 ± 0.50 | 33.9 ± 0.69 | 4.7 ± 0.68 | 2.1 ± 0.17 |
6 | NG_000006.1:g.34164_37967del3804/NM_000518.5(HBB):c.92+6T>C/ (n = 1) | α−3.7/IVS 1.6 | 5.06 | 11.4 | 36.1 | 71.3 | 22.5 | 31.6 | 3.9 | 0.5 |
7 | HbA2: c.95+2_95+6delTGAGG/NM_000518.5(HBB):c.93-21G>A (n = 1) | α2 IVS1 -5nt/IVS 1.110 | 5.24 | 11.8 | 36.5 | 69.7 | 22.5 | 32.3 | 5.5 | 5.4 |
8 | HbA2: c.95+2_95+6delTGAGG/NM_000518.5(HBB):c.92+1G>A/ (n = 1) | α2 IVS1 -5nt/IVS 1.1 | 6.39 | 13.7 | 40.9 | 64 | 21.4 | 33.5 | 6.3 | 0.8 |
9 | α−4.2/NG_000007.3:g63632_7104 del (n = 1) | α−4.2/HB LEPORE-BW | 5.81 | 14.9 | 45 | 77.4 | 25.6 | 33.1 | 3.5 | 2.5 |
10 | NG_000006.1: g.15164_37864del22701 /NM_000518.5(HBB):c.118C>T (n = 1) | -20.5 KB/CODON 39 | 5.03 | 11.8 | 36 | 71.6 | 23.5 | 32.8 | 4.7 | 0.3 |
11 | NG_000006.1:g.34164_37967del3804/HBB:c.20A>T (n = 1) | α−3.7/CODON 6 [A>T] | 4.8 | 13.1 | 38.5 | 80.5 | 28.6 | 33.5 | 2.9 | 0.1 |
12 | NG_000001:g.34247_38050dup/NM_000518.5(HBB):c.118C>T (n = 1) | ααα−3.7/CODON 39 | 4.79 | 10.9 | 30.1 | 60.2 | 22.1 | 34.5 | 5.4 | 1.1 |
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Dell’Edera, D.; Persia, B.; La Rocca, F.; Centoducati, C. Haemoglobinopathies: Integrated Biochemical and Molecular Diagnosis in 5243 Patients. Hemato 2025, 6, 36. https://doi.org/10.3390/hemato6040036
Dell’Edera D, Persia B, La Rocca F, Centoducati C. Haemoglobinopathies: Integrated Biochemical and Molecular Diagnosis in 5243 Patients. Hemato. 2025; 6(4):36. https://doi.org/10.3390/hemato6040036
Chicago/Turabian StyleDell’Edera, Domenico, Brunilde Persia, Francesco La Rocca, and Carmela Centoducati. 2025. "Haemoglobinopathies: Integrated Biochemical and Molecular Diagnosis in 5243 Patients" Hemato 6, no. 4: 36. https://doi.org/10.3390/hemato6040036
APA StyleDell’Edera, D., Persia, B., La Rocca, F., & Centoducati, C. (2025). Haemoglobinopathies: Integrated Biochemical and Molecular Diagnosis in 5243 Patients. Hemato, 6(4), 36. https://doi.org/10.3390/hemato6040036