Abstract Hemoglobin (Hb) Jax (HBA2:c.44G>C, Trp→Ser, TGG>TCG) is a rare α-globin variant. In this study, we reported the identification of Hb Jax in a 34-year-old Thai woman with unexplained microcytic anemia. She was referred for further hematologic evaluation as part of routine thalassemia screening. Hemoglobin analysis by high-performance liquid chromatography (HPLC) showed no identifiable abnormal peaks. However, hematological findings revealed a mild hypochromic microcytic profile. Molecular analysis found the absence of common α⁰- and α⁺-thalassemia deletions (--^SEA, --^THAI, --^{Chiang Rai}, -α^3.7 and -α^4.2) and direct DNA sequencing identified a heterozygous G>C substitution at codon 14 of the HBA2 gene, corresponding to the Hb Jax variant. Further family screening confirmed the same mutation in her father, who presented with similar hematologic features, while her mother had a normal α-globin genotype. Although clinically mild, the presence of this variant may complicate diagnosis, particularly in compound states with other α-globin mutations. This case highlights the importance of molecular diagnostics for the accurate identification of rare globin variants, particularly in regions with high prevalence of α-thalassemia and hemoglobinopathies.

Keyword: Hb Jax, α-globin chain, Hemoglobin variant, α-thalassemia, Hemoglobinopathies

Funding: MMP had been supported by the CMU Presidential Scholarship from the Graduate School, Chiang Mai University, Chiang Mai, Thailand, since the academic year 2024. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.  

Citation:  Paing, M.M., Boonthong, T., Ruengdit, C., Punyamung, M., and Pornprasert, S. 2026. Case report: Hemoglobin Jax (HBA2:c.44G>C, Trp→Ser, TGG>TCG) in a Thai family. Natural and Life Sciences Communications. 25(2): e2026038.

Graphical Abstract:

INTRODUCTION

Abnormal hemoglobins (Hb) arise from alterations in the molecular structure of the globin chains, typically produced by point mutations in globin genes. To date, over 1,000 naturally occurring human Hb variants have been discovered. In addition, there are more than 3,500 entries in ITHANET database and more than 2,300 entries in IthaGenes, an interactive database for hemoglobin variations and epidemiology (Kountouris et al., 2014), highlighting the genetic diversity present within the globin gene clusters. Among these variants, HbS (HBB:c.20A>T), HbC (HBB:c.19G>A), and HbE (HBB:c.79G>A) are the most common and clinically significant worldwide, especially in areas where they provide selective benefits against malaria. Although most Hb variants are pathologically silent and do not impact red cell function, a subset is associated with a wide range of hematologic diseases, from mild asymptomatic anemia to severe hemolytic anemia and thalassemia-like syndromes (Thom et al., 2013). Hb Jax (HBA2:c.44G>C) is an α-globin chain variant caused by a nucleotide substitution at codon 14 in the exon 1 of the HBA2 gene, resulting in an amino acid change from tryptophan to serine (Trp→Ser) (Panyasai et al., 2025). In heterozygous individuals, Hb Jax may affect mild hematological abnormalities resembling those seen in α⁺-thalassemia. However, when co-inherited with deletional α⁰-thalassemia, it can result in a more severe phenotype similar to HbH (β4) disease, characterized by chronic anemia and elevated reticulocyte counts (Singha et al., 2024; Panyasai et al., 2025).

In Thailand, Hb Jax variant was first reported in 2024 by Srichairatanakool et al. in a 59-year-old Thai female patient. A compound heterozygosity for Hb Jax and Hb Constant Spring (CS; HBA2:c.427T>C) resulted in chronic microcytic anemia (Srichairatanakool et al., 2024). In 2025, Panyasai et al. documented another case in a Thai family, involving a woman with unexplained anemia and her daughter, both found to carry the Hb Jax variant (Panyasai et al., 2025). Hb Jax is often underdiagnosed or misdiagnosed due to its broad spectrum of clinical manifestations. Accurate identification of Hb Jax is crucial to prevent diagnostic confusion with thalassemia and other hemoglobinopathies. Recognition of Hb Jax facilitates the detection of compound heterozygosity, where co-inheritance with α-thalassemia or α-globin gene variants can lead to severe clinical presentations (Srichairatanakool et al., 2024; Panyasai et al., 2025). Rare variants like Hb Jax expand hemoglobinopathy databases, improve understanding of genotype-phenotype correlations, and inform public health strategies, mainly in regions with a high prevalence of hemoglobin variants. In the present study, we reported our awareness of Hb Jax in a Thai family and offered a detailed clinical and hematological characterization of the affected members.

CASE PRESENTATION

The proband is a 34-year-old female from Hang Dong, a district of Chiang Mai Province in Northern Thailand. She was referred to the thalassemia and hemoglobinopathy screening program. The blood sample was collected using ethylenediaminetetraacetic acid (EDTA) as an anticoagulant, and the hematological data were analyzed by the ADVIA 2120i hematology analyzer (Siemens Healthcare Diagnostic, Deerfield, IL, USA). Her characteristics and hematological indices are shown in Table 1. The hemoglobin analysis was conducted by using HPLC (VARIANT β-thalassemia Short Program; Bio-Rad Laboratories, Hercules, CA, USA). The levels of HbA, HbA₂, and HbF were found within the normal adult values (Figure 1A). Additionally, the NucleoSpin^® kit (Macherey-Nagel, KG., Duren, Germany) was used to extract the genomic DNA from the blood sample according to the manufacturer's instructions. The α⁰-thalassemia deletions, including --^SEA, --^Thai, and --^{Chiang Rai} type, were identified by single-tube multiplex real-time polymerase chain reaction (PCR) with EvaGreen dye (Biotium, Fremont, CA, USA) and high-resolution melting (HRM) analysis (Ruengdit et al., 2023). The common α⁺-thalassemia -α^3.7 and -α^4.2 deletions were investigated through a multiplex Gap-PCR technique (Fucharoen et al., 2002). Moreover, the next-generation sequencing (NGS) panel, which targets the coding regions of the HBA1, HBA2, and HBB genes, was performed using the thalassemia Gene Detection Kit (BGI Group, Shenzhen, China) (He et al., 2017). The sequence readings were mapped to the hg19 version of the human genome, and the sequence variations were assessed using the THACARE HALOS analysis software (BGI Group). Molecular analysis revealed no detectable common α⁰-thalassemia or α⁺-thalassemia mutations in the proband. NGS result of the α-globin genes revealed a heterozygous single nucleotide substitution at codon 14 of the α2-globin gene (TGG>TCG) (Figure 2), resulting in the replacement of tryptophan with serine. This variant corresponds to the previously reported unstable hemoglobin known as Hb Jax (Srichairatanakool et al., 2024).

In order to assess the inheritance pattern and possible clinical implications, family screening was undertaken. The proband’s 65-year-old father presented with microcytic anemia, with hematological indices shown in Table 1. In contrast, her 57-year-old mother demonstrated normal hematologic parameters (Table 1). The Hb analysis of her parents by HPLC demonstrated Hb fractions for HbA, HbA₂, and HbF within or close to normal ranges (Figure 1B and C). NGS result revealed that the proband’s father carried the heterozygous HBA2:c.44G>C mutation, confirming the presence of Hb Jax while the mother was found to have normal HBA1, HBA2, and HBB genotypes (data not shown).

Table 1. The characteristics and hematological parameters of the proband and her parents.

Note: Normal range of adults: red blood cell counts (RBCs) 4.2-6.1 x 10¹²/L, total Hb 12.0-18.0 g/dL, packed cell volume (PCV) 0.37-0.52 L/L, mean corpuscular volume (MCV) 80-100 fL, mean corpuscular Hb (MCH) 27.0-31.0 pg, mean corpuscular Hb concentration (MCHC) 320-360 g/L, red cell distribution width (RDW) 11.0-16.0%, ferritin 30-300 ng/mL, serum iron 50-150 µg/dL, total iron-binding capacity (TIBC) 240-450 µg/dL, transferrin saturation 20-55%, HbA 95.0-98.0%, HbA₂ 1.5-3.5%, HbF 0.0-1.0%, *Hb analysis by HPLC

Figure 1. Representative of the HPLC chromatogram of (A) the proband, (B) her father, and (C) her mother.

Figure 2. Representative of NGS results exported from the integrative genomics viewer (IGV) of the heterozygosity for Hb Jax (HBA2:c.44G>C).

DISCUSSION

Hb Jax is a rare non-deletional α-globin variant caused by a single nucleotide substitution in exon 1 of the HBA2 gene (c.44G>C), resulting in the amino acid replacement of tryptophan by serine at codon 14. This amino acid substitution is particularly significant due to its impact on the structural integrity of the α-globin chain. The substitution of the hydrophobic tryptophan with the more hydrophilic serine at a critical internal position (A-helix) disrupts the normal hydrophobic interactions essential for protein folding and stability. Consequently, this molecular change compromises the structural stability of the Hb molecule. The functional instability of Hb Jax has been demonstrated through isopropanol precipitation testing, and computational analyses have predicted the variant to be deleterious (Panyasai et al., 2025). The current investigation conducted dichlorophenolindophenol (DCIP) and osmotic fragility (OF) tests, yielding a negative result for both tests in the proband and her father (Table 1). Indicating the heterozygosity for Hb Jax has minimal impact on the structural stability and precipitation of the Hb molecule. Moreover, Hb Jax was reported by James D. Hoyer on the HbVar: A database of Human Hemoglobin Variants and Thalassemias website in 2022. According to the HbVar database, the hematological findings in an adult male with a history of deep vein thrombosis who shows microcytosis (MCV 76.7 fL) and hypochromia (MCH 24.2 pg) with normal Hb levels (13.5 g/dL), suggesting a clinically silent or mildly affected phenotype (Giardine et al., 2014). Standard electrophoretic techniques, including capillary electrophoresis (CE) and isoelectric focusing, are unable to differentiate Hb Jax from normal HbA due to the minimal impact of the Trp→Ser substitution on the net charge of the hemoglobin molecule. As both tryptophan and serine are electrically neutral at physiological pH, Hb Jax co-migrates with HbA during CE method, thereby making it undetectable by this technique (Giardine et al., 2014; Panyasai et al., 2025). The HPLC has been shown to detect the variant as a subtle abnormality, typically presenting as a low, broad, elevated baseline, thereby providing a more sensitive detection method (Giardine et al., 2014; Panyasai et al., 2025). The accurate monitoring and identification of unstable hemoglobins hold significant clinical importance. However, a normal result from CE or HPLC does not exclude the presence of unstable hemoglobins, which undergo rapid denaturation and destruction within the red blood cells, leaving only normal hemoglobin available for being recognized (Risinger et al., 2019; Yates et al., 2010). The finding of Hb Jax becomes more feasible in the presence of co-inherited α⁰-thalassemia, although it remains undetectable by CE in both heterozygous and compound heterozygous. Moreover, the chromatographic profile of individuals heterozygous for Hb Jax did not reveal a distinct peak by HPLC (Panyasai et al., 2025). This observation aligns with our study, in which no identifiable Hb Jax peak was observed by HPLC in the proband (Figure 1A) or her father (Figure 1B), both are carriers of Hb Jax. In this study, we identified heterozygous Hb Jax in a Thai proband and her father. They presented with microcytic anemia in the absence of common α⁰- or α⁺-thalassemia deletions. Hematologic parameters showed reduced MCV, MCH, total Hb, and RBC counts, further supporting the hypothesis that Hb Jax alone might lead to mild instability or reduced globin synthesis. Their iron studies also revealed normal values of ferritin, serum iron and total iron-binding capacity (TIBC), and transferrin saturation (Table 1), so refusing iron deficiency anemia.

Additionally, the hematological profiles they demonstrate are in line with heterozygous states of non-deletional α-thalassemia variants like Hb CS (MCV 80.4 fL and MCH 26.0 pg) and Hb Quong Sze (QS; HBA2:c.377T>C) (MCV 75.2 fL and MCH 24.5 pg) (Yang et al., 2014; Jiang et al., 2020). Like Hb Jax, several non-deletional α-globin variants, including Hb CS, Hb QS, and Hb Westmead (WM; HBA2:c.369C>G), also involve the mutation in the HBA2 gene (Li et al., 2010). Notably, pathogenic variants in HBA2 are associated with more severe phenotypes than those in HBA1, as HBA2 contributes to approximately two to three times more α-globin production than HBA1 (Tamary and Dgany, 1993). Consequently, mutations affecting the dominantly expressed HBA2 gene are likely to result in more severe clinical manifestations than those involving HBA1 (Farashi and Harteveld, 2018). When these mutations are co-inherited with an α⁰-thalassemia allele, they can cause non-deletional HbH disease, which is usually more severe than deletional HbH disease. Future study in a broader population is therefore necessary to investigate the clinical and hematological features of individuals who co-inherited with α⁰-thalassemia and Hb Jax. Moreover, it is important to identify these mutations in carrier detection and prenatal diagnosis, to ensure the integrity in the diagnosis of α-thalassemia (Li et al., 2010). In this case, based on the genetic findings and inheritance pattern observed, it is likely that the Hb Jax variant identified in the proband was inherited from her father. Accurate detection and characterization of Hb Jax variant is essential for effective carrier screening, genetic counseling, and prenatal diagnosis, particularly in regions with high prevalence of thalassemia. Therefore, it is recommended that NGS panel which targets the coding regions of the HBA1, HBA2, and HBB genes be performed on all microcytic hypochromic anemic samples, but common causes of anemia remain undetected and regardless of the presence or absence of abnormal hemoglobin identified during Hb typing analysis.

CONCLUSION

This report describes a rare case of heterozygous Hb Jax with a focus on its possible clinical significance beyond its typically silent nature. Despite its rarity, our findings provide a compelling argument for its integration into routine genetic screening panels for anemia, particularly within the Thai population. Accurate identification and characterization of this hemoglobin variant is fundamental to enhancing diagnostic yield, recommending suitable therapeutic options, and being effective in pre-counseling to persons afflicted with this disease and their families.

ACKNOWLEDGEMENTS

The authors thank the technicians at the Department of Medical Technology, Hang Dong Hospital, Chiang Mai, Thailand, for their help and assistance.   

ETHICAL APPROVAL

This research received ethical approval from the Ethics Committee at the Faculty of Associated Medical Sciences, Chiang Mai University, Thailand (Approval No. AMSEC-68EM-014).

AUTHOR CONTRIBUTIONS  

Moe Moe Paing: Conceptualization (Equal), Validation (Equal), Writing – Original Draft (Lead), Visualization (Equal), Formal Analysis (Equal); Thun Boonthong: Methodology (Equal), Software (Lead), Formal analysis (Lead), Investigation (Equal), Data Curation (Lead), Writing – Review & Editing (Supporting); Chedtapak Ruengdit: Software (Equal), Validation (Equal), Formal Analysis (Equal), Investigation (Lead), Writing - Review & Editing (Supporting); Manoo Punyamung: Software (Equal), Investigation (Lead), Data Curation (Equal), Writing - Review & Editing (Supporting); Sakorn Pornprasert: Conceptualization (Lead), Methodology (Equal), Formal Analysis (Equal), Funding Acquisition (Lead), Investigation (Lead), Supervision (Lead), Validation (Equal), Visualization (Equal), Writing – Review & Editing (Lead), Resources (Lead), Project Administration (Lead).

CONFLICT OF INTEREST

The authors declare that they hold no competing interests.

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