What is the approach to diagnosing thalassemia?

Diagnosis of Thalassemia

The diagnosis of thalassemia requires a stepwise approach beginning with complete blood count (CBC), hemoglobin analysis, and molecular genetic testing to identify specific mutations in the globin genes. 1, 2

Initial Laboratory Evaluation

• Complete Blood Count (CBC):

  • Look for microcytic (low MCV) and hypochromic (low MCH) anemia
  • Normal or elevated red blood cell count despite anemia
  • Normal or elevated ferritin levels (to distinguish from iron deficiency anemia)

• Hemoglobin Analysis:

  • Hemoglobin electrophoresis
  • High-performance liquid chromatography (HPLC)
  • Capillary electrophoresis
  • Isoelectric focusing

These tests reveal characteristic patterns depending on the type of thalassemia:

• Alpha thalassemia: Normal HbA2, normal or slightly increased HbF
• Beta thalassemia trait: Elevated HbA2 (>3.5%), normal or slightly increased HbF
• Beta thalassemia major: Predominantly HbF, absent or reduced HbA, elevated HbA2
• HbE/beta thalassemia: HbE, HbF, and reduced or absent HbA

Confirmatory Testing

• Molecular Genetic Testing:
  • PCR-based techniques to identify specific mutations
  • Gap-PCR for common deletions
  • DNA sequencing for point mutations
  • Reverse dot blot hybridization for common mutations 3, 1

Diagnostic Algorithm

1. Initial Screening:

   • CBC showing microcytic, hypochromic anemia with normal/elevated RBC count
   • Normal or elevated ferritin levels (ruling out iron deficiency)

2. Hemoglobin Analysis:

   • Abnormal hemoglobin pattern on electrophoresis or HPLC
   • Quantification of HbA, HbA2, HbF, and other hemoglobin variants

3. Molecular Confirmation:

   • Genetic testing to identify specific alpha or beta globin gene mutations
   • Essential for definitive diagnosis and classification 1, 2

Classification Based on Clinical Severity

After diagnosis, thalassemia is classified into:

• Transfusion-Dependent Thalassemia (TDT):

  • Requires regular blood transfusions to survive
  • Includes beta thalassemia major and severe HbE/beta thalassemia

• Non-Transfusion-Dependent Thalassemia (NTDT):

  • Does not require regular transfusions for survival
  • Includes beta thalassemia intermedia, HbH disease, and mild HbE/beta thalassemia 2

Special Considerations

• Family Screening: When thalassemia is diagnosed, family members should be screened to identify carriers 1

• Prenatal Diagnosis:

  • Offered to at-risk couples (both carriers)
  • Methods include chorionic villus sampling or amniocentesis with DNA analysis 1

• Newborn Screening:

  • Recommended in high-risk populations
  • Allows early intervention and management 1

Potential Diagnostic Pitfalls

• Co-inheritance of iron deficiency may mask elevated HbA2 in beta thalassemia carriers
• Concurrent alpha and beta thalassemia can normalize MCV and MCH
• HbE can be misidentified as HbA2 on some electrophoresis systems
• Apparent homozygosity for mutations may actually represent compound heterozygosity with a rare variant or deletion 3

Targeted Screening Recommendations

Targeted screening is recommended for individuals with ancestry from:

• Mediterranean regions
• Middle East
• Africa
• Southeast Asia 1

By following this diagnostic approach, clinicians can accurately diagnose thalassemia, determine its severity, and implement appropriate management strategies to improve patient outcomes and quality of life.