Microcytic Anemias: Thalassemia Lab Test Interpretation
If you haven’t done so already, read my blog post on understanding lab reference ranges to provide context for variation of lab reference ranges we see in practice.
Questions to always ask yourself when ordering a lab test:
Will this change my management? (i.e. have I already reached the treatment threshold? Will this test change my plan?)
Will this lab test help me rule in, or rule out, a specific diagnosis?
Refresh yourself on pre-test probability, sensitivity and specificity, and negative and positive predictive values if you haven’t in a while. These are important diagnostic test principles that help inform diagnostic testing in practice. Now - let’s get into navigating a diagnosis of thalassemia.
Case: you’re seeing a 30-year-old female with mild chronic fatigue. She tells you she is from Egypt, has a past medical history of low iron, and does not take any medications. She used to take iron supplements but stopped because they didn’t improve her fatigue. As part of your initial workup, you order a CBC to assess for anemia.
What is a CBC?
The CBC (complete blood count) evaluates different properties of the blood, including:
Hemoglobin (hgb): the iron-containing oxygen transport metalloprotein in red blood cells
Hematocrit (hct): the proportion of blood volume that is occupied by red blood cells; tells us about blood viscosity
Red blood cell count (RBC): the number of RBCs contained in a sample of blood
Red blood cell distribution width (RDW): measures variation of RBC size and volume
Mean corpuscular volume (MCV): tells us about the size of the RBCs (low = microcytic, high = macrocytic, normal = normocytic)
Mean corpuscular hemoglobin (MCH): tells us the average amount of hemoglobin in the RBCs
Mean corpuscular hemoglobin concentration (MCHC): tells us about the average amount of hemoglobin relative to the size of the RBC
Platelets: play a fundamental role in hemostasis (too low = bleeding risk, too high = clotting risk)
Normal reference ranges vary depending on age, sex, and the lab reporting the results.
Back to the Case:
Results of the CBC (bolded measurements are abnormal)
Interpretation pearls:
To diagnose anemia, either the hemoglobin, hematocrit, or RBCs are low (or can be all 3)
To diagnose a microcytic anemia, the MCV is low (typically <80)
In this case, a hgb of 105 (low) and MCV of 70 (low) provides you with a diagnosis of microcytic anemia. This could be contributing to her presenting complaint of fatigue - but you don’t have the full diagnosis yet.
Differential diagnoses for microcytic anemia (3):
Iron deficiency anemia
Anemia of chronic disease
Hemoglobinopathy
Sideroblastic anemia (rare)
Lead poisoning
Copper deficiency
Zinc toxicity
In this case, the main differentials to work up are iron deficiency and hemoglobinopathy based on her history.
What is your next step? Order ferritin to assess for iron deficiency anemia.
Results:
Ferritin: 50 (normal)
She does not have an inflammatory condition to suggest a falsely elevated ferritin, so you have ruled out iron deficiency anemia. The next differential diagnosis to assess for is hemoglobinopathy.
Tip: if you suspect an inflammatory disorder is causing elevated ferritin, the transferrin saturation (TSAT) is a helpful test to assess for iron deficiency anemia. If the TSAT is <20% = iron deficiency likely, if >20% = iron deficiency ruled out (1).
PEARL # 1: If the hemoglobin +/- RBC count +/- hematocrit is low, with a low MCV, this indicates a microcytic anemia. If the ferritin is normal, or TSAT is >20%, you have ruled out iron deficiency anemia.
What is Thalassemia?
Thalassemia is a heterogenous grouping of genetic disorders that affect the synthesis of alpha or beta chains of hemoglobin (2). It is autosomal recessive, meaning both parents must have the disease or be carriers of the disease to pass it along.
Pathophysiology recap: If the body cannot make enough of alpha or beta chains, the RBCs don’t form correctly and cannot carry sufficient oxygen to body tissues (2). This results in anemia from early childhood lasting throughout the lifetime. To inherit thalassemia, at least one parent must be a carrier. It results from a genetic mutation or a deletion of key gene fragments (2).
Thalassemia is classified as alpha thalassemia or beta thalassemia.
Alpha thalassemia: is caused by alpha globin gene deletion, resulting in reduced or absent production of alpha-globin chains (2). The alpha globin gene has 4 alleles, and disease severity ranges from mild to severe depending on how many alleles are deleted (2). Alpha thalassemia is prevalent in Asian and African populations (2). Up to 40% of African Americans carry the gene for alpha thalassemia, believed to be due to the fact it is protective against malaria. In this case, it is common to have microcytosis without anemia.
1 of 4 gene deletions: silent carrier (alpha thalassemia minima); asymptomatic, normal CBC (3)
2 of 4 gene deletions: trait (alpha thalassemia minor); microcytosis, mild anemia (3)
3 of 4 gene deletions: HbH disease (alpha thalassemia intermedia); microcytosis, mild-moderate anemia
4 of 4 gene deletions: alpha thalassemia major; usually fatal without treatment
Beta thalassemia: results from point mutations in the beta-globin gene (2). There are 3 categories of beta thalassemia.
Beta thalassemia trait/minor: single gene defect; mild or asymptomatic (3)
Beta thalassemia intermedia: 2 gene defects, mild-moderate impairment in beta-globin production; results in mild-moderate disease (3)
Beta thalassemia major: 2 gene defects, severe impairment in beta-globin production; signs and symptoms will be moderate to severe (e.g. jaundice, hepatosplenomegaly, failure to thrive) (3)
Beta-thalassemia is more prevalent in the Mediterranean population, and is relatively common in Southeast Asia and Africa (2).
Patients can have coinheritance of both alpha and beta thalassemia, and usually have a milder clinical course due to a less severe alpha-beta chain imbalance (2).
Patients can also have iron deficiency anemia and a hemoglobinopathy.
Pearl # 2:
Thalassemia trait/carrier states: asymptomatic or mild microcytic anemia with mild symptoms
Intermedia/major states of alpha or beta thalassemia: results in moderate to severe microcytic anemia with moderate-severe symptoms (e.g. fatigue, light-headedness, syncope)
How do we Diagnose Thalassemia?
Back to the case: your patient has a microcytic anemia with normal ferritin. You have ruled out iron deficiency anemia, and are now going to work her up for hemoglobinopathy.
Your next steps:
Peripheral blood smear: can be helpful to assess additional red cell properties like abnormal differences in RBC size, shape, and other physical appearances that are indicative of hemoglobinopathies like sickle cell disease and thalassemia. Where I practice, if there is an abnormal CBC, the peripheral blood smear is often reflex tested.
Clues on the peripheral blood smear to indicate a thalassemia: microcytic cells, hypochromic cells, increased % of reticulocytes (immature RBCs), target cells, Heinz bodies, anisocytosis (variation in size), and poikilocytosis (variation in shape) (2).
Hemoglobin electrophoresis: this test separates normal and abnormal types of hemoglobin in the blood. It screens for hemoglobinopathy and identifies abnormal hemoglobin (e.g. S, H).
In normal adults, hemoglobin A (HbA), composed of both alpha and beta-globin chains, makes up 95% to 98% of hemoglobin (2,3)
Hemoglobin A2 (HbA2) is normally 2% to 3% of hemoglobin (2,3)
Hemoglobin F (HbF) makes up less than 2% of hemoglobin(2,3)
Clues to indicate alpha thalassemia trait: normal HbA2
Clues to indicate alpha thalassemia intermedia: presence of HbH (a less common form of hemoglobin) (2,3)
Clues to indicate beta thalassemia trait: mild elevation HbA2 (3)
Clues to indicate beta thalassemia minor: mild elevation of HbA2 and mild decrease of HbA (2,3)
Clues to indicate beta thalassemia major: elevation of HbF and HbA2, absent or very low HbA (2,3).
Iron studies: are routinely done to rule out iron deficiency anemia as highlighted previously (serum iron, ferritin, total iron-binding capacity (TIBC), transferrin saturation (TSAT) (3)
PEARL # 3: Hemoglobinelectrophoresis
HbA makes up majority of hemoglobin in healthy adults
Assesses for any abnormal elevation in HbA2, HbF or abnormal decrease in HbA
Detects abnormal hemoglobin (e.g. HbS, HbH)
Back to the case:
Iron studies are normal. Peripheral blood smear reveals microcytosis and hypochromia. Results of hemoglobin electrophoresis:
Hemoglobin C: not detected
Hemoglobin E: not detected
Hemoglobin S: not detected
Hemoglobin H: not detected
Abnormal hemoglobin: not detected
Hemoglobin A2: 0.035 (mildly elevated)
Hemoglobin A: 0.93 (mildly decreased)
What are your next steps? You’ve made your diagnosis with clinical clues and lab results:
Anemia with microcytosis (low hemoglobin and low MCV)
Normal ferritin level
Microcytosis and hypochromia on peripheral blood smear
Hemoglobin electrophoresis reveals mildly elevated HbA2, no abnormal hemoglobin and mild decrease Hemoglobin A
Diagnosis: Beta thalassemia minor
Beta thalassemia can be diagnosed based on hemoglobin electrophoresis. Genetic testing is not necessary, and can be reserved for patients who are thinking about pregnancy and need genetic counselling. Alpha thalassemia requires genetic testing to confirm the diagnosis, and should be offered to patients planning pregnancy.
Because we all love a good algorithm…but it’s not fool proof!
Key Take Home Points:
The CBC is the initial test of choice to screen for anemia or hemoglobinopathy.
If hemoglobin +/- RBCs +/- hematocrit are low, and the MCV is low, order iron studies to rule out iron deficiency anemia.
If ferritin is normal or TSAT is >20%, order a peripheral blood smear to assess red cell properties and hemoglobin electrophoresis to assess for hemoglobinopathy (e.g. thalassemia, sickle cell).
If the hemoglobinopathy screen is positive and a patient is planning for pregnancy, send for genetic testing to confirm the diagnosis and arrange for genetic counselling.
Want to test your knowledge with more cases? Let’s go!
Because reference ranges vary based on the reporting lab, I’ve indicated abnormal results in bold.
Case # 1: Results for a male patient with fatigue and inflammatory bowel disease. What is the most likely diagnosis?
Case # 2: Results for a female patient with fatigue and heavy menses (no inflammatory conditions). What is the most likely diagnosis?
Case # 3: Results for a patient that is planning for pregnancy. What is the most likely diagnosis?
Case # 4: Results for a patient with severe fatigue and pre-syncopal episodes. What is the most likely diagnosis?
Answer Key
Case # 1:
Diagnosis: iron deficiency anemia. Low hemoglobin, hematocrit, RBC count, and microcytosis with a low TSAT. Ferritin is falsely elevated due to inflammation.
Case # 2:
Diagnosis: iron deficiency anemia. Low RBC count, microcytosis, and low ferritin.
Case # 3:
Diagnosis: alpha thalassemia minor/trait. Low hemoglobin with microcytosis, normal iron indices, and normal hemoglobin electrophoresis.
Case # 4:
Diagnosis: beta-thalassemia major. Microcytic anemia with increased Hemoglobin A2, Hemoglobin F, and severely decreased Hemoglobin A.
ReferencesElsayed ME, Sharif MU, Stack AG. Transferrin Saturation: A Body Iron Biomarker. Adv Clin Chem. 2016;75:71-97. doi: 10.1016/bs.acc.2016.03.002. Epub 2016 May 6. PMID: 27346617.
Bajwa H, Basit H. Thalassemia. [Updated 2023 Aug 8]. In: StatPearls [Internet]. Treasure Island (FL): StatPearls Publishing; 2024 Jan-. Available from: https://www.ncbi.nlm.nih.gov/books/NBK545151/
Baird DC, Batten SH, Sparks SK. Alpha- and Beta-thalassemia: Rapid Evidence Review. Am Fam Physician. 2022 Mar 1;105(3):272-280. PMID: 35289581.