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PA16.1-3 | Acquired Haemolytic Anaemias & Smear Morphology — Part 3

Infections, Hypersplenism & March Haemoglobinuria

A four-panel medical diagram compares malaria-related haemolysis, blackwater fever, hypersplenism, and march haemoglobinuria as causes or mechanisms of haemolytic anaemia. — **Panel A:** Malaria with infected RBCs, ring trophozoites, banana-shaped P. falciparum gametocyte, ruptured RBC, free haemoglobin, spleen sequestration, and high parasitaemia marker.. **Panel B:** Blackwater fever showing massive intravascular haemolysis, free haemoglobin passage to kidney, and dark haemoglobinuric urine.. **Panel C:** Hypersplenism showing enlarged spleen, splenic red pulp, sequestered RBCs, macrophage removal, mild normocytic anaemia, and normal or mildly raised reticulocytes.. **Panel D:** March haemoglobinuria showing repetitive foot-strike trauma, capillary RBC mechanical lysis, intravascular haemolysis, and exertional haemoglobinuria..

Malaria — the most common infectious cause globally. Plasmodium falciparum causes the most severe haemolysis:
• Parasites invade and rupture red cells directly → intravascular haemolysis.
• Parasitised cells are rigid and sequestered in the spleen → extravascular component.
• Smear: ring trophozoites, banana-shaped gametocytes (P. falciparum), infected cells of variable size depending on species. High parasitaemia (>5%) correlates with severity.
• Severe falciparum: blackwater fever — massive intravascular haemolysis → haemoglobinuria.

Hypersplenism — any cause of marked splenomegaly (portal hypertension, Gaucher disease, lymphoma) increases red cell sequestration time → mildly shortened RBC lifespan. Anaemia is typically mild, normocytic, with normal/mildly elevated reticulocytes. No specific smear morphology beyond mild anisopoikilocytosis.

March haemoglobinuria — rare; repetitive mechanical trauma (prolonged marching, hand drumming) ruptures red cells in small vessels of feet/palms → acute intravascular haemolysis, self-limited, haemoglobinuria. Smear may show mild polychromasia; usually normal.

Peripheral Smear Morphology: Pattern Recognition Guide

Systematic recognition of red cell morphology is the fastest triage tool in haemolytic anaemia.

Four-panel microscopic blood smear comparison showing different red blood cell abnormalities in hemolytic anemias under oil immersion microscopy. — **Panel A:** Spherocytes in warm AIHA - dense RBCs lacking central pallor with polychromasia. **Panel B:** Schistocytes/helmet cells in MAHA - fragmented triangular and crescent-shaped RBCs. **Panel C:** Bite cells/blister cells in G6PD deficiency - RBCs with semicircular membrane defects. **Panel D:** RBC agglutination in cold AIHA - grape-like clumps of red blood cells.

Morphology	What it tells you	Key diagnoses
Spherocytes	Loss of membrane → ↓surface:volume; no central pallor	Warm AIHA (IgG-mediated), hereditary spherocytosis, ABO-incompatible transfusion reaction
Schistocytes (helmet cells, fragments)	Mechanical shearing in vessels	MAHA (TTP, HUS, DIC, valve haemolysis)
Bite cells (blister cells)	Heinz body pitting by spleen	G6PD deficiency (oxidative stress), unstable haemoglobins
RBC agglutination (clumps)	IgM-mediated cold agglutination	Cold AIHA (Mycoplasma, EBV, lymphoma)
Target cells	↑surface:volume (↑membrane or ↓Hb)	Thalassaemia, liver disease, HbC; NOT specific for haemolysis
Polychromasia	Reticulocytosis	Any active haemolysis or blood loss

Practical rule: Schistocytes → think MAHA (non-immune, mechanical). Spherocytes → think immune (warm AIHA) or membrane disorder. Bite cells → think oxidative. Agglutination → think cold IgM.

SELF-CHECK

On peripheral smear of a patient with acute haemolytic anaemia following ingestion of fava beans, you observe red cells with irregular projections on one side and a clear zone ('bite') on the other, along with Heinz bodies on supravital stain. What is the mechanism of this morphology?

A. IgG-mediated partial phagocytosis by splenic macrophages

B. Mechanical shearing by fibrin strands in vessels

C. Splenic pitting of denatured haemoglobin (Heinz bodies) from oxidised cells

D. Complement-mediated membrane attack complex formation

Reveal Answer

Answer: C. Splenic pitting of denatured haemoglobin (Heinz bodies) from oxidised cells

In G6PD deficiency, oxidative stress (fava beans, primaquine, infections) overwhelms the reduced glutathione system. Haemoglobin is oxidised and denatured, forming Heinz bodies (precipitates attached to the inner membrane). Splenic macrophages 'pit' (remove) these Heinz body inclusions, creating the characteristic bite cell (blister cell). This is different from spherocyte formation (partial IgG-mediated phagocytosis) or schistocyte formation (mechanical shearing). The mechanism is oxidative — not immune and not mechanical.

SELF-CHECK

You are shown four peripheral smear images (Panel A–D). Match each finding to its haemolytic mechanism: - Panel A: Small dense RBCs with no central pallor, polychromasia - Panel B: Fragmented triangular RBCs, helmet cells - Panel C: Large clumps of agglutinated RBCs - Panel D: Cells with irregular 'bitten-out' area on one side Which panel represents MICROANGIOPATHIC haemolytic anaemia?

A. Panel A

B. Panel B

C. Panel C

D. Panel D

Reveal Answer

Answer: B. Panel B

Panel B (schistocytes/helmet cells/triangular fragments) is the hallmark of MAHA — red cells physically sheared by fibrin strands or turbulent flow. Panel A shows spherocytes (warm AIHA). Panel C shows RBC agglutination (cold AIHA). Panel D shows bite cells (G6PD/oxidative haemolysis). Recognising these four patterns instantly narrows the differential to a single mechanism.

Diagnostic Approach: Integrating DAT, Smear, and Lab Indices

A four-panel diagnostic algorithm integrates hemolysis markers, peripheral smear findings, DAT results, and confirmatory tests for hemolytic anemia. — **Panel A:** Confirm hemolysis using increased LDH, increased indirect bilirubin, decreased haptoglobin, increased reticulocyte count, plus intravascular markers hemoglobinemia, hemoglobinuria, and hemosiderinuria.. **Panel B:** Peripheral smear first in practice, showing spherocytes, schistocytes, bite cells, agglutination, and parasites with arrows to DAT, MAHA workup, G6PD assay, cold agglutinin testing, and malaria testing.. **Panel C:** DAT interpretation showing IgG positive warm AIHA, C3d-only positive cold AIHA, and DAT-negative non-immune hemolysis pathways.. **Panel D:** Specific tests and clinical pearl showing PNH flow cytometry CD55/CD59, ADAMTS13 activity less than 10 percent for TTP, G6PD quantitative assay timing, and the haptoglobin false-normal trap in infection or inflammation..

A systematic algorithm prevents over-reliance on any single test:

Step 1 — Confirm haemolysis:
• ↑LDH (released from lysed cells), ↑indirect bilirubin, ↓haptoglobin (binds free Hb, consumed in haemolysis), ↑reticulocyte count.
• Intravascular: also haemoglobinaemia, haemoglobinuria, haemosiderinuria.

Step 2 — Peripheral smear (performed FIRST in practice):
• Spherocytes → proceed to DAT.
• Schistocytes → MAHA workup (platelet count, PT/APTT, fibrinogen, D-dimer, ADAMTS13).
• Bite cells → G6PD assay.
• Agglutination → cold agglutinin titre, DAT (C3d pattern).
• Parasites → malaria thick/thin smear, rapid antigen test.

Step 3 — DAT:
• Positive IgG → warm AIHA → screen for secondary cause (ANA, SPEP, CT neck/chest/abdomen for lymphoma).
• Positive C3d only → cold AIHA → Mycoplasma serology, EBV, protein electrophoresis.
• Negative → non-immune → MAHA/PNH/mechanical/infectious workup.

Step 4 — Specific tests:
• PNH suspected: flow cytometry CD55/CD59.
• ADAMTS13 activity (TTP): <10% activity highly specific.
• G6PD: quantitative assay (avoid during acute crisis — falsely normal because reticulocytes have higher G6PD).

CLINICAL PEARL

Haptoglobin is the most sensitive single marker of haemolysis — but has a false-positive trap. Haptoglobin is an acute-phase reactant (rises in infection/inflammation). A patient with simultaneous haemolytic anaemia and active infection may have a 'normal' haptoglobin that is actually suppressed relative to their inflammatory baseline. In such cases, LDH + bilirubin + smear together confirm haemolysis even when haptoglobin looks reassuringly normal.