What is the differential diagnosis and appropriate work‑up for a patient with non‑immune hemolytic anemia (low haptoglobin, elevated reticulocyte count, negative direct antiglobulin (Coombs) test), macrocytosis, mild leukopenia, and a low absolute CD4⁺ T‑cell count?

Differential Diagnosis and Work-Up for Non-Immune Hemolytic Anemia with Macrocytosis and Low CD4 Count

This patient has confirmed non-immune hemolysis (low haptoglobin <0.10, elevated reticulocytes 177, negative Coombs) with macrocytosis (MCV 100) and mild leukopenia with low absolute CD4 count (335), requiring immediate evaluation for drug-induced hemolysis, nutritional deficiencies (B12/folate), HIV status, hereditary hemolytic disorders, and thrombotic microangiopathy.

Immediate Diagnostic Priorities

Confirm and Characterize the Hemolysis

The diagnostic triad is present: elevated reticulocytes, undetectable haptoglobin, and the negative Coombs test confirms non-immune etiology 1, 2. You must now obtain:

• Peripheral blood smear examination immediately to identify schistocytes (suggesting TMA), spherocytes (membrane defects), bite cells (G6PD deficiency), or other diagnostic morphologies—this test has 90% sensitivity and 95% specificity for differentiating hemolysis types 2, 3
• LDH and indirect (unconjugated) bilirubin to quantify hemolysis severity; the combination of elevated LDH and decreased haptoglobin is specific for hemolysis 1, 2
• Complete blood count with platelet count because thrombocytopenia alongside hemolysis mandates urgent TMA evaluation 2

Rule Out Life-Threatening Emergencies First

Before pursuing chronic causes, exclude acute emergencies 3:

• Thrombotic microangiopathy (TMA): If platelets are low, immediately order ADAMTS13 activity and inhibitor titer 2. Check creatinine and urinalysis for the TMA triad: microangiopathic hemolysis, thrombocytopenia, and organ involvement 2
• Medication-induced hemolysis: Review all current and recent medications for common culprits including daptomycin, cephalosporins, penicillins, NSAIDs, quinine/quinidine, rifampin, ribavirin, and immune checkpoint inhibitors 4, 1
• Severe infections: The negative CMV is helpful, but consider malaria, babesiosis, and other infectious causes of hemolysis 4, 3

Address the Macrocytosis (MCV 100)

The combination of macrocytosis with hemolysis and elevated reticulocytes creates two possibilities:

• Megaloblastic anemia superimposed on hemolysis: Markedly elevated LDH (>3000 IU/L) with macrocytosis and normal/low reticulocytes suggests B12/folate deficiency, but your patient has elevated reticulocytes 1. Still, check vitamin B12 and folate levels because deficiency can coexist with hemolysis and blunt the reticulocyte response 4, 5
• Reticulocytosis-induced macrocytosis: Reticulocytes are larger than mature RBCs, so the elevated reticulocyte count (177) may explain the MCV of 100 6. Calculate the corrected reticulocyte count and reticulocyte production index to assess marrow response adequacy 7

Investigate the Low CD4 Count and Leukopenia

The absolute CD4 count of 335 with leukocytes 3.95 and monocytes 0.23 raises concern for immunodeficiency:

• HIV testing is mandatory despite negative CMV, as HIV can cause both hemolysis and immunosuppression 4
• Consider autoimmune disorders: Check ANA, anti-Ro/La antibodies, and rheumatoid factor, as autoimmune conditions can cause both cytopenias and hemolysis 5
• Evaluate for lymphoproliferative disorders: The combination of cytopenias and hemolysis may indicate underlying hematologic malignancy requiring bone marrow examination 4, 5

Second-Line Testing for Hereditary Hemolytic Disorders

If the above workup is unrevealing and peripheral smear shows specific morphologies 3:

• G6PD enzyme assay if bite cells or blister cells are present, especially with history of oxidative stress exposure 4, 3
• Eosin-5'-maleimide (EMA) binding test for hereditary spherocytosis if spherocytes are seen 3
• Hemoglobin electrophoresis/phenotyping to screen for hemoglobinopathies (sickle cell, thalassemia, unstable hemoglobins) 3
• Paroxysmal nocturnal hemoglobinuria (PNH) flow cytometry if intravascular hemolysis markers are prominent (marked LDH elevation, hemoglobinuria) 4, 3

Additional Essential Tests

• Iron studies, ferritin, and copper levels to assess for deficiencies that may impair reticulocyte response and to monitor for iron overload in chronic hemolysis 4, 1
• Thyroid function (TSH) as thyroid disorders can cause both anemia and cytopenias 4
• Protein electrophoresis and cryoglobulin analysis to exclude paraprotein-related hemolysis 4
• Parvovirus B19 serology if reticulocyte response seems inadequate for the degree of anemia 4

Critical Pitfalls to Avoid

• Do not dismiss TMA based on "rare" schistocytes alone—low schistocyte counts can occur in early or evolving TMA with 80% sensitivity 2
• Do not delay drug discontinuation if medication-induced hemolysis is suspected; cessation is the most critical intervention 1
• Do not transfuse aggressively—transfuse RBCs only if hemoglobin <7-8 g/dL or the patient is symptomatic, using the minimum units necessary 4, 2
• Do not miss coexisting nutritional deficiencies—reticulocytopenia occurs in 20-40% of hemolytic anemia cases due to iron/vitamin deficiency and is a poor prognostic factor 7
• Do not overlook bone marrow involvement—if cytopenias persist across multiple cell lines or reticulocyte response is inadequate, bone marrow examination with cytogenetic analysis is required to exclude myelodysplastic syndromes 4, 5

Monitoring Strategy

• Serial hemoglobin, reticulocyte count, LDH, and haptoglobin every 2-3 days initially to assess disease progression and response 1, 2
• Weekly CBC monitoring once stabilized until hemolysis resolves completely 5, 2
• Reassess CD4 count and immune function after treating any identified underlying cause 4