Hb SKMC and an unprecedented γδß-thalassemia: first report from Iraq.

BACKGROUND: Thalassemias are genetic disorders of globin chain synthesis. In Iraq, ß-thalassemia is more prevalent than α-thalassemia. This study identifies two unpredicted globin gene mutations, a rare α-globin gene mutation (Hb SKMC) and a novel γδß-thalassemia deletion. METHODS: Over 2 years, the Genetics unit at PAR hospital in Erbil, northern Iraq processed 137 ß-thalassemia and 97 α-thalassemia genetic testing requests. Three symptomatic thalassemia cases with unreported genotypes were identified. Proband-1α and proband-2α had Hb H disease, while proband-1ß had severe transfusion-dependent ß-thalassemia (TDT). Molecular studies included multiplex PCR, reverse hybridization, multiplex ligation-dependent probe amplification (MLPA), and globin gene sequencing. RESULTS: The α-thalassemia probands exhibited moderate microcytic hypochromic anemia with irregular transfusions and splenomegaly. Hb H disease was confirmed by positive Hb H tests and high-performance liquid chromatography (HPLC). Molecular analysis revealed heterozygous -MED deletion in proband-1α and α2Poly-A2 mutation in proband-2α. Sequencing identified the Hb SKMC (HBA1:c.283_300+3dup) mutation in both probands. The ß-thalassemia proband showed anemia and regular transfusions. Molecular studies detected the IVS1.110 G>A mutation and a novel γδß-thalassemia deletion in compound heterozygous form. The maternal sample showed the IVS1.110 G>A mutation, and MLPA confirmed the γδß-thalassemia deletion in the paternal sample. CONCLUSION: These findings highlight the genetic diversity of thalassemias in the region and emphasize the importance of advanced molecular diagnostics in detecting rare mutations.

Genotype and phenotype analysis of α-thalassemia fusion gene in southern China.

OBJECTIVE: The α-globin fusion gene between the HBA2 and HBAP1 genes, is clinically important in thalassemia screening because this fusion gene can cause severe hemoglobin (Hb) H disease when combined with α0 -thalassemia (α0 -thal). In this study, we evaluate the red blood cell parameters of α-thalassemia fusion gene in southern China. METHOD: Study samples suspected of α-thalassemia fusion gene were collected and confirmed by PCR-sequencing from one medical lab center in southern China. Their genotypes and phenotypes were analyzed. RESULTS: A total of 266 cases of α-thalassemia fusion gene were confirmed in our lab from 2017 to 2023, most of them were from Hainan province (169 cases) and Huadu district of Guangzhou (21 cases), the nationality of 143 cases from Hainan was identified, with 71.3% (102/143) being from the Li minority. The Hb, MCV, MCH for αα/(αα)fusion in adult males were 143.5±11.83g/L, 81.51±4.39 fl, and 26.26±1.29 pg, respectively; and in females, they were 126.69±12.89 g/L, 80.10±4.05 fl, 25.8±2.04 pg, respectively. All 12 cases (αα) Fusion/ --SEA showed anemia with decreased Hb, MCV and MCH. CONCLUSION: The carriers of α-globin fusion gene heterozygotes are clinically silent and exhibit an α+ phenotype. Individuals with (αα)Fusion/--SEA show apparent anemia. This α-globin fusion gene is relatively common in southern China, specifically among the Li minority of Hainan province. Therefore, it should be taken into account for genetic counseling purposes.

Novel Insights into Hb Shaare Zedek Associated with ß0-Thalassemia: Molecular Characteristics, Genetic Origin and Diagnostic Approaches.

Hemoglobin Shaare Zedek (Hb SZ) is a rare structural α-Hb variant. Characterizing its genotype-phenotype relationship and genetic origin enhances diagnostic and clinical management insights. We studied a proband and six family members using high-performance liquid chromatography (HPLC), capillary electrophoresis (CE), PCR, and sequencing to analyze α- and ß-globin genes and α-globin haplotypes. Pathogenicity predictions and a rapid diagnostic method were developed. The proband, his father, grandfather, and aunt had Hb migrating to the HbH-zone on CE and elevated fetal hemoglobin (HbF) on HPLC. Direct sequencing identified an A to G mutation at codon 56 of the α2-globin gene, characteristic of Hb SZ. Additionally, the proband carried a ß-globin gene mutation [HBB.52A>T]. Mild thalassemia-like changes were observed in the proband, whereas individuals with only the Hb SZ variant did not exhibit these changes. Pathogenicity predictions indicated that Hb SZ is benign. The variant can be identified using restriction fragment length polymorphism (RFLP) and allele-specific PCR. The Thai variant of Hb SZ is associated with the haplotype [- - M - - - -]. Hb SZ is a non-pathological variant that minimally affects red blood cell parameters, even when it coexists with ß0-thalassemia. HPLC and CE systems cannot distinguish it from other Hbs, necessitating DNA analysis for accurate diagnosis.

A rare case of compound heterozygous Southeast Asian double α-globin gene deletion and Haemoglobin Quong Sze in a Malay proband.

INTRODUCTION: Haemoglobin (Hb) Quong Sze is a non-deletional α-thalassaemia subtype that occurs due to missense mutation at codon 125 of the HBA2 gene. Interaction between Hb QS with Southeast Asian double α-globin gene deletion results in non-deletional HbH disease, which is more severe than deletional HbH. CASE REPORT: A 3-month-old baby boy was presented with neonatal anaemia and mild hepatomegaly. Full blood count revealed severe hypochromic microcytic anaemia. There was an abundance of HbH inclusion bodies in his red blood cells. High-performance liquid chromatography showed a reduced HbA2 level with the presence of pre-run peak. Capillary electrophoresis showed the presence of HbH and Hb Barts. Molecular analysis found a common α0-thalassaemia (--SEA) in one allele and mutation in codon 125 in the other allele. DISCUSSION: Non-deletional HbH disease due to a combination of deletional and non-deletional mutations may present with severe clinical manifestations than those with deletion mutations, which warrants accurate diagnosis using molecular techniques.

Hb Guigang [α90 (FG2)Lys→Asn; HBA1:c.273G˃T]: a Novel α-Globin Chain Variant.

BACKGROUND: New hemoglobin (Hb) variants are constantly being updated as assays are developed and the testing population expands. Here, we report a novel Hb variant, named Hb Guigang. METHODS: Hemoglobin (Hb) analysis was analyzed by capillary electrophoresis (CE) and high-performance liquid chromatography (HPLC). Glycated hemoglobin was performed by CE and HPLC. Routine genetic analysis was done with Gap-PCR and PCR-reverse dot-blot hybridization. The hemoglobin variant was identified by Sanger sequencing. RESULTS: CE of three cases showed the presence of Hb variants in Zone 5 and Zone 12, respectively. HPLC indicated an elevated P3 peak, suggesting the possible presence of the Hb variant. Hb A1c was measured by CE and HPLC, and the results were 6.7% and 4.76%, respectively. Sanger sequencing confirmed an AAG˃AAT mutation at codon 90 of the HBA1 gene. This mutation was reported for the first time, and we named it Hb Guigang based on the proband's place of residence. CONCLUSIONS: Hb Guigang with normal hematological parameters was separated and quantified by CE, whereas HPLC suggested that Hb Guigang co-eluted with the P3 peaks and could not be quantified.

De-novo ATR-16 syndrome associated with inherited hemoglobin Evanston causing HbH phenotype: a rare occurrence.

Abnormality of three α-globin genes, either deletion or point mutation results in symptomatic Hemoglobin H (HbH) phenotype. Most of such cases of α-globin defects are inherited from the parents, de-novo cases are exceedingly rare. Herein, a case of HbH is reported where the proband inherited one α-globin gene with a point mutation (αEvanston) from the mother. This was associated with large de-novo deletion of chromosome 16p13.3 resulting in α-thalassemia and mental retardation (ATR-16) syndrome. This deletion also encompassed two α-globin genes from chromosome 16, eventually leading to --/ααEvanston genotype, explaining the clinical presentation of the proband. The challenges in screening of such cases and confirming the molecular diagnosis along with the mode of inheritance has been discussed.

α-Globin mutations and Genetic Variants in γ-globin Promoters are Associated with Unelevated Hemoglobin F Expression of Atypical ß0-thalassemia/HbE.

BACKGROUND: Excessive expression of hemoglobin F (HbF) is a characteristic feature and important diagnostic marker of ß0-thalassemia/HbE disease. However, some patients may exhibit low-HbF levels, leading to misdiagnosis and precluding genetic counseling. The genetic factors influencing these differences in HbF expression in this atypical disease are not completely understood. AIMS: To investigate determinants contributing to the non-elevation of HbF expression in ß0-thalassemia/HbE disease. METHODS: We studied 231 patients with ß0-thalassemia/HbE confirmed by DNA analysis; classified them into the low-HbF (n = 62) and high-HbF (n = 169) groups; analyzed hematological parameters and hemoglobin levels in both groups; and characterized mutations in ß- and α-globin genes and genetic variants in γ-globin promoters. RESULTS: Both groups showed similar rates of type ß0-thalassemia mutations but significantly different proportions of α-globin mutations: approximately 88.7% (95% confidence interval [CI] = 66.8-115.5) and 39.1% (95% CI = 30.2-49.7) in the low- and high-HbF groups, respectively. The results revealed single-nucleotide polymorphisms (SNPs) at -158 (C>T) in the Gγ-globin promoters and novel SNPs at the 5' untranslated region position 25 (G>A) in Aγ-globin promoters. The distribution of CC genotypes of the Gγ-globin promoter in the low-HbF group was significantly higher than that in the high-HbF group. CONCLUSIONS: Cases with HbE predominance with low-HbF levels and undetectable HbA may not be as conclusive as those with homozygous HbE until DNA analysis is performed. Concomitant inheritance of α-thalassemia is an important inherent factor modifying HbF expression in a typical ß0-thalassemia/HbE, and SNPs with the CC genotype in the Gγ-globin promoter may indicate unelevated HbF expression in patients with this disease.

[Molecular Diagnosis and Pedigree Analysis of Rare Mutations in Non-coding Region of HBA2 Gene].

OBJECTIVE: To perform molecular diagnosis and pedigree analysis for one case with α-thalassemia who does not conform to the genetic laws, and explore the effects of a newly discovered rare mutation (HBA2:c.*12G>A) on clinical phenotypes. METHODS: Blood samples of the proband and her family members were collected for blood routine analysis, and the hemoglobin components were analyzed by capillary electrophoresis. The common α- and ß-globin gene loci in Chinese population were detected by conventional techniques (Gap-PCR, RDB-PCR). The α-globin gene sequences (HBA1, HBA2) were analyzed by Sanger sequencing. RESULTS: By analyzing the test results of proband and her family members, the genotype of the proband was -α3.7/HBA2:c.*12G>A, her father was HBA2:c.*12G>A heterozygous mutation carrier. CONCLUSION: This study identifies a rare α-globin gene mutation (HBA2:c.*12G>A) that has not been reported before. It is found that heterozygous mutation carriers present with static α-thalassemia.

[Analysis of the Gene Mutation Type and Frequency of Thalassemia Patients in Jingzhou Area].

OBJECTIVE: To analyze the gene mutation types and frequence of thalassemia patients in Jingzhou area. METHODS: A total of 721 suspected thalassemia patients who were visited in Jingzhou Central Hospital from June 2019 to June 2022 were selected as the research objects. There were 204 males and 517 females. PCR-reverse dot hybridization method was used to analyze the types and frequencies of 23 common α or ß thalassemia gene mutations. RESULTS: Among the 721 patients with suspected thalassemia, 228 cases were positive for α or ß thalassemia gene, with a total positive rate of 31.62%, including 87 cases of α-thalassemia, accounting for 38.16%, and 140 cases of ß-thalassemia, accounting for 61.40%. There was 1 case of α ß complex thalassemia, accounting for 0.44%. A total of 4 types of α-thalassemia gene mutations were detected, all of which were deletion types, including αα/--SEA (64/87, 73.56%), αα/-α3.7 (14/87, 16.09%), --SEA /-α3.7 (7/87, 8.05%), αα/-α4.2 (2/87, 2.30%). Among 140 patients with ß-thalassemia, 138 were pure heterozygotes, and the genotypes of IVS-II-654M (63/140, 45.00%), CD41-42M (34/140, 24.29%), CD17M (18/140, 12.86%) and CD27-28M (10/140, 7.14%) accounted for 89.29% of all mutations (125/140), 2 cases of double heterozygosity (2/140, 1.43%) were found, no homozygous ß-thalassemia were detected; 1 case of αß complex thalassemia with genotype -α3.7/IVS-II-654M was found. The incidence of difference types of thalassemia was statistically significant (χ2=194.250, P < 0.001). The percentage of positive thalassemia genes was not significantly difference between male and female suspected patients (χ2=0.199, P =0.655). CONCLUSION: The α-thalassemia gene mutation in Jingzhou area is dominated by αα/--SEA, and the IVS-II-654M mutation is more common in ß-thalassemia, and α ß complex thalassemia is relatively rare, which can provide a reference for the formulation of prevention and treatment measures for thalassemia in Jingzhou area.

A moonlighting job for α-globin in blood vessels.

ABSTRACT: Red blood cells express high levels of hemoglobin A tetramer (α2ß2) to facilitate oxygen transport. Hemoglobin subunits and related proteins are also expressed at lower levels in other tissues across the animal kingdom. Physiological functions for most nonerythroid globins likely derive from their ability to catalyze reduction-oxidation (redox) reactions via electron transfer through heme-associated iron. An interesting example is illustrated by the recent discovery that α-globin without ß-globin is expressed in some arteriolar endothelial cells (ECs). α-globin binds EC nitric oxide (NO) synthase (eNOS) and degrades its enzymatic product NO, a potent vasodilator. Thus, depletion of α-globin in ECs or inhibition of its association with eNOS causes arteriolar relaxation and lowering of blood pressure in mice. Some of these findings have been replicated in isolated human blood vessels, and genetic studies are tractable in populations in which α-thalassemia alleles are prevalent. Two small studies identified associations between loss of α-globin genes in humans and NO-regulated vascular responses elicited by local hypoxia-induced blood flow or thermal stimulation. In a few larger population-based studies, no associations were detected between loss of α-globin genes and blood pressure, ischemic stroke, or pulmonary hypertension. In contrast, a significant positive association between α-globin gene copy number and kidney disease was detected in an African American cohort. Further studies are required to define comprehensively the expression of α-globin in different vascular beds and ascertain their overall impact on normal and pathological vascular physiology.

Mitoxantrone ameliorates ineffective erythropoiesis in a ß-thalassemia intermedia mouse model.

ABSTRACT: ß-thalassemia is a condition characterized by reduced or absent synthesis of ß-globin resulting from genetic mutations, leading to expanded and ineffective erythropoiesis. Mitoxantrone has been widely used clinically as an antitumor agent considering its ability to inhibit cell proliferation. However, its therapeutic effect on expanded and ineffective erythropoiesis in ß-thalassemia is untested. We found that mitoxantrone decreased α-globin precipitates and ameliorated anemia, splenomegaly, and ineffective erythropoiesis in the HbbTh3/+ mouse model of ß-thalassemia intermedia. The partially reversed ineffective erythropoiesis is a consequence of effects on autophagy as mitochondrial retention and protein levels of mTOR, P62, and LC3 in reticulocytes decreased in mitoxantrone-treated HbbTh3/+ mice. These data provide significant preclinical evidence for targeting autophagy as a novel therapeutic approach for ß-thalassemia.

[Expert consensus on clinical genetic counseling of α-thalassemia gene analysis].

α-thalassemia is a type of microcytic hypochromic anemia caused by variants of alpha-globin gene, and is one of the most common monogenic disorders in southern China. The population screening model based on hematologic phenotype has achieved great results in areas with high incidence of thalassemia. However, with the continuous decline of the cost of genetic testing and implementation of screening programs for thalassemia gene carriers, more variants in the alpha-globin gene have been discovered, which also brings great challenges to clinical genetic counseling. From the perspective of alpha-globin genetic analysis, this consensus has discussed the contents of pre- and post-test genetic counseling, with an aim to provide standardized guidance for clinicians.

Ancestry of the major long-range regulatory site of the α-globin genes in the Portuguese population with the common 3.7 kb α-thalassemia deletion.

BACKGROUND: The α-Major Regulatory Element (α-MRE), also known as HS-40, is located upstream of the α-globin gene cluster and has a crucial role in the long-range regulation of the α-globin gene expression. This enhancer is polymorphic and several haplotypes were identified in different populations, with haplotype D almost exclusively found in African populations. The purpose of this research was to identify the HS-40 haplotype associated with the 3.7 kb α-thalassemia deletion (-α3.7del) in the Portuguese population, and determine its ancestry and influence on patients' hematological phenotype. METHODS AND RESULTS: We selected 111 Portuguese individuals previously analyzed by Gap-PCR to detect the presence of the -α3.7del: 50 without the -α3.7del, 34 heterozygous and 27 homozygous for the -α3.7del. The HS-40 region was amplified by PCR followed by Sanger sequencing. Four HS-40 haplotypes were found (A to D). The distribution of HS-40 haplotypes and genotypes are significantly different between individuals with and without the -α3.7del, being haplotype D and genotype AD the most prevalent in patients with this deletion in homozygosity. Furthermore, multiple correspondence analysis revealed that individuals without the -α3.7del are grouped with other European populations, while samples with the -α3.7del are separated from these and found more closely related to the African population. CONCLUSION: This study revealed for the first time an association of the HS-40 haplotype D with the -α3.7del in the Portuguese population, and its likely African ancestry. These results may have clinical importance as in vitro analysis of haplotype D showed a decrease in its enhancer activity on α-globin gene.

Comprehensive analysis of thalassemia alleles (CATSA) based on third-generation sequencing is a comprehensive and accurate approach for neonatal thalassemia screening.

Thalassemia is one of the most common and damaging monogenic diseases in the world. It is caused by pathogenic variants of α- and/or ß-globin genes, which disrupt the balance of these two protein chains and leads to α-thalassemia or ß-thalassemia, respectively. Patients with α-thalassemia or ß-thalassemia could exhibit a severe phenotype, with no simple and effective treatment. A three-tiered strategy of carrier screening, prenatal diagnosis and newborn screening has been established in China for the prevention and control of thalassemia, of which the first two parts have been studied thoroughly. The implementation of neonatal thalassemia screening is lagging, and the effectiveness of various screening programs has not yet been demonstrated. In this study, hemoglobin capillary electrophoresis (CE), hotspot testing method, and third-generation sequencing (TGS) were used in the variant detection of 2000 newborn samples, to assess the efficacy of these methods in neonatal thalassemia screening. Compared with CE (249, 12.45 %) and hotspot analysis (424, 21.2 %), CATSA detected the largest number of thalassemia variants (535, 26.75 %), which included 24 hotspot variants, increased copy number of α-globin gene, rare pathogenic variants, and three unreported potentially disease-causing variants. More importantly, CATSA directly determined the cis-trans relationship of variants in three newborns, which greatly shortens the clinical diagnosis time of thalassemia. CATSA showed a great advantage over other genetic tests and could become the most powerful technical support for the three-tiered prevention and control strategy of thalassemia.

Uncommon Combination of Hemoglobin Jax and Hemoglobin Constant Spring Leading to Microcytic Anemia.

BACKGROUND Thalassemia and hemoglobin (Hb) variants are the most common hereditary red blood cell disorders worldwide. Alpha-thalassemia and alpha-globin variants are caused by mutations of the alpha-globin genes (HBA2 and HBA1), resulting in impaired alpha-globin production and structurally abnormal globin, respectively. Clinical severity of alpha-thalassemia correlates with the number of affected alpha-globin genes, yielding a spectrum of clinical manifestations from mild to severe anemia. Routine diagnosis involves Hb analysis and PCR-based methods, yet identifying rare variants necessitates comprehensive clinical and hematologic laboratory data. The knowledge of phenotype and genotype correlation is useful for genetic counseling and treatment planning. CASE REPORT A 59-year-old Thai woman presented with chronic anemia. Her baseline Hb level ranged between 8.0 and 9.0 g/dL, with no history of transfusion. Physical examination showed mild pallor, without enlarged liver and spleen. Laboratory investigations showed microcytic, hypochromic anemia and abnormal Hb peak by Hb analysis (retention time 4.58 min by HPLC method). Common alpha-globin gene deletions, including the Southeast-Asian/Thai 3.7 kb and 4.2 kb deletions were tested using gap-PCR, with none of these deletions detected. Direct DNA sequencing revealed a compound heterozygosity of Hb Jax (HBA2: c.44G>C) and Hb Constant Spring (HBA2: c.427T>C). CONCLUSIONS Compound heterozygosity of Hb Jax and Hb Constant Spring results in microcytic anemia. Hb Jax can be identified by Hb analysis, and diagnosis can be confirmed by direct DNA sequencing method. Coinheritance of Hb Jax and alpha-globin variants should be considered in cases with microcytic anemia and a specific Hb peak seen in Hb chromatogram.

[Study of the types of mutations of Thalassemia in Shanghai area].

OBJECTIVE: To analyze the mutations of globin genes among patients suspected for thalassemia from the Shanghai area. METHODS: A total of 4 644 patients diagnosed at Ruijin Hospital, Shanghai Jiao Tong University School of Medicine between June 2016 and December 2019 were selected as the study subjects. The patients were tested for common mutations associated with thalassemia gene by Gap-PCR and reverse dot blotting (RDB). Patients were suspected to harbor rare mutations based on the inconsistency between hematological phenotypes and results of common mutation detection, and were further analyzed by Gap-PCR and Sanger sequencing. RESULTS: Among the 4 644 patients, 2 194 (47.24%) were found to carry common thalassemia mutations, among which 701 (15.09%) were α-thalassemia, 1 448 (31.18%) were ß-thalassemia, and 45 (0.97%) were both α- and ß-thalassemia. Forty six samples were found to harbor rare mutations, which included 17 α-globin gene and 29 ß-globin gene mutations. CD77(CCC>ACC) (HBA2: c.232C>A) of the α-globin gene, NG_000007.3: g.70567_71015del449, codon 102(-A) (HBB: c.308_308delA) and IVS-Ⅱ-636 (A>G) (HBB: c.316-215A>G) of the ß-globin gene were previously unreported new types of globin gene mutations. CONCLUSION: Among the 4 644 patients, the detection rate for common thalassemia mutations was 47.24%, whilst 46 samples were detected with rare gene mutations. The type of gene mutation types were diverse in the Shanghai area. The study has provided more accurate results for genetic diagnosis and counseling.

Hb H disease associated with compound heterozygosity for --SEA deletion and a novel alpha globin chain variant (HBA2:c.175C>A) on the distal histidine in a Chinese family.

OBJECTIVES: In clinical practice, the majority of α-thalassaemia cases arise from deletions of the α-globin genes. However, a subset of cases is attributed to rare haemoglobin variants, which can manifest with borderline or normal screening results, potentially leading to missed diagnoses in clinical practice. METHODS: Blood samples were collected from family members and underwent haematological, DNA and RNA analysis. RESULTS: The five-month-old proband presented a haematological phenotype consistent with Hb H disease. The mother's haematology profile was consistent with an α-thalassaemia carrier, while the father exhibited a borderline reduction in MCV and MCH. MALDI-TOF identified an abnormal α-chain in the proband. DNA analysis revealed a novel α-globin variant (HBA2:c.175C>A, α58His>Asn, Hb DG-Nancheng) affecting the distal histidine in the family. The father and the mother had α-genotype of --SEA/αα and αDG-Nanchengα/αα, respectively; while the proband inherited both mutant alleles (--SEA/αDG-Nanchengα). Sequencing of cDNA from HBA2 gene identified an equal ratio of normal and mutant alleles. CONCLUSION: This rare case highlighted the importance of identifying rare haemoglobin variant during prenatal screening. The clinical and genetic data provides useful information on the pathogenicity of this variant and further insight into the role of distal histidine residue of α-globin.

Severe Transfusion-Dependent Thalassemia in Compound Heterozygote Palestinian Siblings with Two α-Globin Gene Defects, Hb Taybe D HBA1: C.119_121delCCA Mutation and HBA2: C.*94A > G Mutation.

Alpha and Beta Thalassemia are autosomal recessive anemias that cause significant morbidity and mortality worldwide, especially in the Middle East and North Africa (MENA) region where carrier rates reach up to 50%. We report the case of two siblings of Palestinian origin born who presented to our tertiary healthcare center for the management of severe transfusion dependent hemolytic anemia. Before presentation to our center, the siblings were screened for a-thalassemia using the Alpha-globin StripAssay. They were found to carry the α2 polyA-1 [AATAAA > AATAAG] mutation in the heterozygous form, which was insufficient to make a diagnosis. No pathogenic variants were detected on Sanger sequencing of the HBB gene. Full sequencing of the a-gene revealed compound heterozygous variants (HBA1:c.119_121delCCA and the previously detected HBA2:c.*+94A > G Poly A [A->G]) with trans inheritance. This report highlights the impact of non-deletional mutations on α-globin chain stability. The compound heterozygosity of a rare α-globin chain pathogenic variant with a polyadenylation mutation in the probands leads to clinically severe a-thalassemia. Due to the high carrier status, the identification of rare mutations through routine screening techniques in our populations may be insufficient. Ongoing collaboration among hematologists, medical geneticists, and counselors is crucial for phenotypic-genotypic correlation and assessment of adequate genetic testing schemes.