What is the diagnostic approach for a patient of African or Mediterranean descent presenting with suspected hemolysis, considering potential underlying conditions such as sickle cell disease or G6PD (Glucose-6-Phosphate Dehydrogenase) deficiency?

Diagnostic Approach for Hemolysis in Patients of African or Mediterranean Descent

Initial Laboratory Confirmation of Hemolysis

Begin by confirming hemolysis with the following laboratory markers: elevated reticulocyte count, elevated lactate dehydrogenase (LDH), reduced or absent haptoglobin, elevated unconjugated bilirubin, and in cases of intravascular hemolysis, hemoglobinemia and hemoglobinuria. 1, 2, 3

Key Laboratory Tests

• Reticulocyte count: Elevated in hemolysis, though may be disproportionately low in certain conditions like pyruvate kinase deficiency 1
• LDH: Elevated due to red cell destruction 1
• Haptoglobin: Reduced or absent (haptoglobin binds free hemoglobin) 1, 3
• Unconjugated bilirubin: Typically elevated, usually <5 mg/dL in chronic hemolysis 1
• Hemoglobinuria/hemosiderinuria: Present only in severe intravascular hemolysis 3

Critical History Elements for African/Mediterranean Patients

Specifically query for medication exposures (especially antibiotics like cephalosporins and penicillins, antimalarials including primaquine, NSAIDs, and dapsone), recent infections (viral, bacterial, mycoplasma), and family history of anemia or neonatal jaundice. 4, 2

Essential Historical Points

• Medication exposure: Dapsone and methylene blue are absolutely contraindicated in G6PD deficiency and can cause severe hemolysis 4
• Oxidant stress triggers: Fava beans, infections, specific medications 2
• Recent infections: Viral, bacterial (especially mycoplasma), or parasitic infections can trigger hemolysis 2
• Family history: Chronic anemia, neonatal jaundice, or known hemolytic disorders 1, 2
• Ethnic background: Mediterranean, African, Indian, or Southeast Asian descent increases G6PD deficiency risk 4, 2

Peripheral Blood Smear Examination

Examine the peripheral smear immediately for diagnostic morphologic features: sickle cells (sickle cell disease), blister cells or bite cells (G6PD deficiency), spherocytes (hereditary spherocytosis or autoimmune hemolysis), and echinocytes (pyruvate kinase deficiency, especially post-splenectomy). 1, 5

Morphologic Clues by Condition

• Sickle cell disease: Sickle-shaped cells, target cells 1
• G6PD deficiency: Blister cells, bite cells, Heinz bodies (with supravital staining) 1, 5
• Pyruvate kinase deficiency: Usually unremarkable with anisocytosis and poikilocytosis; 3-30% echinocytes, especially after splenectomy 1
• Autoimmune hemolytic anemia: Spherocytes, agglutination 2

Specific Testing Algorithm for African/Mediterranean Patients

First-Line Screening Tests

Perform G6PD enzyme activity testing and hemoglobin electrophoresis/HPLC as first-line screening in all patients of African or Mediterranean descent presenting with hemolysis. 4, 2

• G6PD enzyme activity: Qualitative screening is sufficient for initial assessment; quantitative testing may be needed for borderline cases 4
• Hemoglobin electrophoresis/HPLC: Identifies sickle cell disease, sickle cell trait, and hemoglobin variants 1

Critical Timing Consideration for G6PD Testing

Test G6PD levels during steady state, not during acute hemolysis, as young reticulocytes have higher G6PD activity and can produce false-negative results. 6 If testing during crisis is unavoidable, repeat testing 2-3 months after resolution when the reticulocyte count normalizes.

Variant-Specific Risk Stratification

• Mediterranean variant (Gdmed): Causes more severe, potentially life-threatening hemolysis; found predominantly in Mediterranean, Indian, and Southeast Asian populations 4, 7
• African variant (GdA-): Causes milder, self-limited hemolysis; found in 10-15% of Black individuals 4, 7

Second-Line Testing When Initial Workup is Negative

If G6PD and hemoglobin electrophoresis are normal, proceed with direct antiglobulin test (DAT/Coombs), pyruvate kinase enzyme activity, and consideration of targeted next-generation sequencing panels for hereditary hemolytic anemias. 1, 2

Additional Diagnostic Tests

• Direct antiglobulin test (DAT): Identifies autoimmune hemolytic anemia or drug-induced immune hemolysis 1, 2
• Pyruvate kinase enzyme activity: Spectrophotometric assay using the ICSH standardized method 1
• Osmotic fragility: May be abnormal in membrane disorders but is not specific for pyruvate kinase deficiency 1
• Unstable hemoglobin testing: Isopropanol or heat stability testing for unstable hemoglobin variants 1
• Targeted NGS panels: Include globin genes (HBA1, HBA2, HBB), CYB5R3, PKLR, and membrane protein genes 1

Special Considerations for Sickle Cell Disease Patients

In patients with known sickle cell disease presenting with acute hemolysis beyond their baseline, consider delayed hemolytic transfusion reaction (DHTR) with hyperhemolysis, especially if recently transfused, as this can cause hemoglobin to drop below pre-transfusion levels and requires avoidance of further transfusion unless life-threatening. 1

• DHTR with hyperhemolysis: Can occur with negative DAT and no identifiable antibody 1
• Management: Avoid additional transfusions if possible; if transfusion is life-threatening necessary, use extended matched red cells (C/c, E/e, K, Jka/Jkb, Fya/Fyb, S/s) 1
• Immunosuppressive therapy: Consider IVIg (0.4-1 g/kg/day for 3-5 days), high-dose steroids (methylprednisolone 1-4 mg/kg/day), or eculizumab for severe cases 1

Critical Pitfalls to Avoid

• Do not test G6PD during acute hemolytic crisis: Young reticulocytes have higher enzyme activity and can mask deficiency 6
• Do not assume G6PD deficiency is clinically insignificant in sickle cell patients: While it may not worsen baseline disease severity, it mandates avoidance of oxidant medications 6, 8
• Do not perform splenectomy in hereditary stomatocytosis: This condition is contraindicated for splenectomy due to thrombophilic risk 2
• Do not give methylene blue to G6PD-deficient patients: This is absolutely contraindicated and can cause severe hemolytic anemia 4
• Do not overlook drug-induced hemolysis: Antibiotics (cephalosporins, penicillins, piperacillin) and antimalarials are common culprits 2

When to Consider Genetic Testing

Proceed with targeted NGS panels or specific gene sequencing when enzyme assays are equivocal, family studies are needed, or when pyruvate kinase deficiency or rare hemoglobinopathies are suspected after excluding common causes. 1

• Family testing: Study parents and first-degree relatives to confirm heterozygous states, particularly in transfusion-dependent cases 1, 7
• Molecular confirmation: DNA sequencing of PKLR for pyruvate kinase deficiency, globin genes for hemoglobin variants, CYB5R3 for methemoglobinemia 1