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  • JANUARY 2014
    JANUARY 2014
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January 2014 - Test your knowledge in the quiz!

1.QUESTION Juvenile myelo-monocytic leukaemia
Acute lymphatic leukemia under chemotherapy
Sickle cell crisis anaemia
Megaloblastic anaemia under unsuccessful vitamin B12 therapy

Online version of this month`s case:

The correct answer is:

Megaloblastic anaemia under unsuccessful vitamin B12 therapy.

Scattergrams and Microscopy

Patient history: a two and a half year old child, under treatment for extreme anaemia.

The presence of immature granulocytes (IG) and atypical lymphocytes in the WDF scattergram resulted in the appearance of the flags ‘IG Present’ and ‘Atypical Lympho?’.
The flag ‘NRBC Present’ indicated that nucleated red blood cells (NRBC) were detected in the WNR scattergram.
The RET scattergram indicated reticulocytosis, while the low RET-He value resulted in a negative Delta-He.
PLT-F analysis triggered the appearance of the ‘Thrombocytopenia’ flag and also showed a slightly increased immature platelet fraction (IPF).
Peripheral blood smear
Bone marrow smear


Interpretation and differential diagnosis

Our interpretation of this case

The correct answer to January’s quiz is:

Megaloblastic anaemia under unsuccessful vitamin B12 therapy
The answer can be inferred from…

Case history

A two and a half year old child was brought to the hospital in a comatose state. The initial complete blood count showed a pancytopenia as well as a macrocytic, hyperchromic anaemia with ineffective erythropoiesis and thrombopoiesis. Cobalamin (vitamin B12) concentrations were extremely low due to insufficient nutrition (breast milk from a vegan mother). Intravenous vitamin B12 therapy was immediately started.

Case results

The full blood count results on the third day after the start of vitamin B12 therapy is shown. The increased reticulocyte count is an effect of ongoing vitamin B12 therapy and suggests successful DNA synthesis. However, the newly built red blood cells (reticulocytes) contain merely low amounts of haemoglobin, indicated by the low RET-He and Delta-He values. This suggests an absolute or functional iron deficiency in addition to the vitamin B12 deficiency, which was confirmed by a low ferritin concentration (15 ng/mL).

The presence of immature granulocytes (IG) and a slightly increased immature platelet fraction (IPF) may indicate the improvement of DNA synthesis in myelopoiesis and thrombopoiesis, respectively, while the 'Atypical Lympho?' flag may indicate the presence of reactive lymphocytes, which is often seen in young children. The presence of NRBC indicates bone marrow stress or extramedullary erythropoiesis due to the extreme anaemia.

Differential diagnosis

The following answers are incorrect for the described reasons.

Juvenile myelomonocytic leukaemia (JMML)

JMML is a myelodysplastic, myeloproliferative leukaemia that mostly affects children aged 4 and younger. In addition, the observed anaemia, thrombocytopenia, increased IG and NRBC counts and the abnormal intracellular structure of the neutrophils (increased NEUT-SSC) might indicate a myelodysplastic condition and could fit the JMML diagnosis. However, the absence of a leukocytosis and a monocytosis and the observed reticulocytosis and hyperchromia are not associated with myeloproliferative disorders and this makes the JMML diagnosis very unlikely. In addition, the extremely negative Delta-He value indicates an acute change in haemoglobinisation within the last couple of days, which is very unusual for a chronic bone marrow disease such as JMML.

Acute lymphatic leukaemia under chemotherapy

Acute lymphoblastic leukaemia very often occurs in paediatric patients and a pancytopenia with a macrocytic anaemia can occur during cytotoxic chemotherapy. However, the combination of a reticulocytosis, low RET-He value and extremely negative Delta-He value is highly unlikely in a cytotoxic chemotherapy. A bleeding or haemolysis may cause an increased reticulocyte count but never in combination with a low RET-He and negative Delta-He.

Sickle cell anaemia crisis

Sickle cell anaemia (SCA) or drepanocytosis is a hereditary blood disorder. The sickling of red blood cells occurs because of a mutation in the haemoglobin gene (HbS). The terms ‘sickle cell crisis’ or ‘sickling crisis’ can be used to describe several independent acute conditions occurring in SCA patients, such as the vaso-occlusive, aplastic, sequestration or haemolytic crises. This case shows a macrocytic anaemia with a reticulocytosis (erythropoietic response), which could be the result of a haemolytic crisis. However, the increase in hyperchromic cells (HYPER-He), the increased MCH and extremely negative Delta-He show that the reticulocytosis cannot be a response to haemolysis. In addition, the abnormal intracellular structure of the neutrophils (increased NEUT-SSC) is not a sign of SCA.



Underlying disease

Megaloblastic anaemia

Megaloblastic anaemias are a heterogeneous group of disorders sharing common morphological characteristics. Megaloblasts are larger than their normal counterparts and have more cytoplasm relative to the size of their nuclei. Nuclear chromatin condenses more slowly than usual and with increasing maturity of the cytoplasm leads to so-called nuclear-cytoplasmic asynchrony. Granulocyte precursors also display nuclear-cytoplasmic asynchrony and enlargement, the giant metamyelocyte often being conspicuous in the bone marrow. Hypersegmented neutrophils are prominent in the peripheral blood. Ineffective thrombopoiesis is present with a reduced platelet count with a functional abnormality. The aetiology of megaloblastic anaemias is diverse, but common factors are impaired DNA synthesis and assembly. The most common causes of megaloblastosis are vitamin B12 (cobalamin) and folate deficiencies. Cobalamin and folate metabolism are intricately related. Dietary intake is the source of cobalamin and folate because humans cannot synthesize these substances. The body stores large amounts of cobalamin, sufficient for 2–6 years.

The differential diagnosis of macrocytosis can be divided into two broad categories based simply on RBC morphology.


  1. Round macrocytosis is due to abnormal lipid composition of the erythrocyte membrane and common causes are: alcoholism, liver disease, renal disease, hypothyroidism.
  2. Oval macrocytosis indicates a problem with cell DNA replication. Common causes are:

  • Drug effects including cytotoxic chemotherapy
  • Megaloblastic anaemias – folate or vitamin B12 deficiency and hypersegmented neutrophils
  • Myelodysplasia – hyposegmented neutrophils and abnormal platelet morphology


Patients with RBC cold agglutinins can show a spurious MCV increase due to red cell clumping at room temperature. Patients with increased reticulocyte counts can also have an increased MCV due to the larger size of the reticulocyte.


Cobalamin deficiency

The causes of cobalamin (vitamin B12) deficiency are multiple and differ with age. Impaired gastric or intestinal absorption, inadequate dietary intake, drugs, or congenital errors in metabolism can cause cobalamin deficiency.

(A) Nutritional disorders: This is rare but can occur in individuals who are on strict vegetarian diets without milk, cheese, and eggs (vegans) over a number of years. Cobalamin deficiency has also been described in infants born to severely cobalamin-deficient mothers, mainly Indian vegans.

(B) Malabsorption of cobalamin: Causes may be gastric or intestinal in origin.

  • Gastric: Addisonian pernicious anaemia, juvenile pernicious anaemia, congenital intrinsic factor deficiency or abnormality. Total or partial gastrectomy.
  • Intestinal: Stagnant loop syndrome (jejunal diverticulosis, ileocolic fistula, anatomical blind loop, intestinal stricture, etc.), ileal resection, Crohn’s disease, selective malabsorption with proteinuria (Imerslund’s syndrome), tropical sprue and fish tapeworm infestation.

(C) Malabsorption of cobalamin but without severe cobalamin deficiency: Simple atrophic gastritis, gluten-induced enteropathy, severe chronic pancreatitis, HIV infection, Zollinger-Ellison syndrome, radiotherapy, graft versus host disease, drugs (neomycin, colchicine, phenytoin, phenformin, slow release potassium chloride), alcohol.

(D) Abnormalities of cobalamin metabolism: Congenital transcobalamin II (TCII) deficiency or abnormality, congenital methylmalonic aciduria, nitrous oxide inhalation.

(E) Drugs that affect cobalamin metabolism: p-Aminosalicylic acid, metformin, phenformin, colchicine, neomycin.




Megaloblastic anaemia

  1. Hoffbrand A.V. (1999): Megaloblastic anaemia. In: Hoffbrand A.V., Lewis S.M., Tuddenham E.G.D. (Editors):  Postgraduate Haematology. 4th Edition. Oxford, UK. Butterworth Heinemann; 47–67.
  2. Rasmussen K., Moller J. (2001): Methodologies of testing. In: Carmel R., Jacobsen D.W. (Editors): Homocysteine in health and disease. Cambridge, UK. Cambridge University Press; p. 199–211.
  3. Clarke R., Grimley Evans J., Schneede J., et al (2004):  Vitamin B12 and folate deficiency in later life. Age und Ageing, 33, p. 34–41.

Advanced clinical parameters

Reference ranges

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