Approach
Folate deficiency classically presents as megaloblastic anaemia with macro-ovalocytes (oval macrocytosis with a mean corpuscular volume [MCV] >100 femtolitres), hypersegmented polymorphonuclear neutrophils (PMNs), and low reticulocyte count.
Severe folate deficiency presents as symptomatic macrocytic anaemia and pancytopenia; non-severe folate deficiency includes signs of macrocytosis without anaemia.
Once folate deficiency is diagnosed, recognition of the underlying precipitating cause is important to prevent ongoing folate deficiency states.
Note that neurological signs and symptoms are not usually seen in patients with folate deficiency. Co-existing vitamin B12 (cobalamin) deficiency, thiamine deficiency, or alcohol-use disorder should be considered in all cases of macrocytic anaemia with neurological signs and symptoms.
History
Symptomatic inquiry should cover symptoms associated with anaemia, including fatigue, palpitations, shortness of breath, dizziness, headaches, jaundice, loss of appetite, and weight loss. Patients may complain of painful swallowing, inflammation of the tongue (glossitis), and discomfort at the corners of the mouth (angular stomatitis) in severe folate deficiency.[Figure caption and citation for the preceding image starts]: Angular cheilitisFrom the collection of Dr Wanda C. Gonsalves; patient consent obtained [Citation ends].
Elicit relevant risk factors for folate deficiency. These include:[3][4][10][11][12][13][14][15]
Dietary deficiency, special diets (e.g., patients with phenylketonuria)
Malabsorption (tropical sprue, coeliac disease)
Use of certain drugs and toxins (e.g., sulfasalazine, trimethoprim, methotrexate, pyrimethamine, anticonvulsants); alcohol-use disorder
Increased physiological demand for folate (pregnancy, lactation, and prematurity)
States of increased cell turnover (e.g., exfoliative dermatitis; haemolysis of any aetiology, but particularly haemoglobinopathies, malaria)
Chronic dialysis
Symptoms of underlying disease should be elucidated, such as chronic diarrhoea and weight loss, or failure to thrive in intestinal disorders.
Physical examination
Look for signs of anaemia (pallor, tachycardia, tachypnoea, heart murmurs, signs of heart failure, jaundice) and underlying disease (e.g., signs of alcohol-use disorder, haemolytic anaemia, exfoliative dermatitis). Petechiae may be present in patients with thrombocytopenia.
Neurological dysfunction is not typically present in pure folate deficiency. Rare exceptions include children with inborn errors of folate absorption and metabolism, who often have severe myelopathy and neurological dysfunction.
Initial tests
All patients with suspected folate deficiency should have a full blood count with peripheral blood smear and reticulocyte count.
Haematological findings
Classically, folate deficiency presents as a megaloblastic anaemia with macro-ovalocytes (oval macrocytosis with MCV >100 femtolitres), hypersegmented PMNs, and a low reticulocyte count.
In early folate deficiency states, the haemoglobin and MCV are normal. As the deficiency progresses, MCV and mean corpuscular haemoglobin increase, followed by reduced haemoglobin. In advancing anaemia, poikilocytes and teardrop cells appear. Neutropenia and thrombocytopenia are often present in advanced folate deficiency.
In extremely severe cases of anaemia, receding macrocytosis has been reported, a probable effect of poikilocytosis and red blood cell (RBC) fragmentation.[30]
The presence of hypersegmented polymorphonuclear leukocytes is a characteristic feature of folate deficiency. It is defined as the presence of 5 lobes in >5% of neutrophils, or the presence of one or more neutrophils with 6 or more lobes. Hypersegmentation often precedes anaemia, but it is not found in sub-clinical vitamin deficiencies. It can be present in patients receiving drugs that inhibit DNA synthesis (e.g., 5-fluorouracil, hydroxyurea) and, rarely, in patients with myelofibrosis, chronic myelogenous leukaemia, or a benign hereditary condition.[Figure caption and citation for the preceding image starts]: Megaloblastic macrocytic anaemia: A. Peripheral blood smear of a patient with megaloblastic anaemia. B. Peripheral blood smear of healthy individualPhotomicrograph from Mark J. Koury, MD; used with permission [Citation ends].
Confirmatory tests
As the initial screen, measuring serum folate level is recommended in suspected folate deficiency.
Serum folate depends on intake, and falls rapidly into the deficient range during deprivation.[44][45] Approximately 5% of patients who have folate deficiency will have normal serum folate levels.[46][47] The RBC folate level decreases more slowly during the 3- to 4-month turnover period of RBCs. Although RBC folate may be a better indicator of tissue folate levels, the assay is complex and more expensive than serum folate assessment. RBC folate level can be low in vitamin B12 deficiency.
Serum folate <7 nanomol/L (<3 nanograms/mL) indicates folate deficiency and often leads to morphological features (megaloblastic changes in bone marrow and macrocytic anaemia).[46] A folate level between 7 nanomol/L and 11 nanomol/L (3 nanograms/mL and 5 nanograms/mL) can cause metabolic changes (elevated plasma homocysteine), and therefore is suspicious for folate deficiency.
When serum folate levels are normal or borderline, in the presence of a strong clinical suspicion, RBC folate (lower limit of normal: <317 nanomol/L [<140 nanograms/mL]) and plasma homocysteine levels (>15 micromol/L; may be indicative of folate deficiency) can be obtained to aid diagnosis.[46][48]
False-negative serum folate test results can be found after recent folate intake. False-positive low serum folate test results may be found in patients with anorexia, alcohol consumption, normal pregnancy, and in patients on anticonvulsants.[48]
Additional tests to consider
Further testing may be considered.
Biochemical tests
The laboratory signs of ineffective erythropoiesis and haemolysis co-exist with marked megaloblastic anaemia. These include elevated LDH, increased unconjugated bilirubin, and low haptoglobin. Serum iron, ferritin, and transferrin receptor levels are elevated, which can mask co-existing iron deficiency.[30]
Iron deficiency should be suspected where anaemia is not accompanied by classic findings of megaloblastosis, but hypersegmented PMNs are present on peripheral blood smear.
Bone marrow aspirate and biopsy
Examination of bone marrow typically shows megaloblastic erythropoiesis. The morphological hallmark of megaloblastic anaemia are erythroblasts that are larger than expected based on cytoplasmic appearance, with large, uncondensed nuclei. Late-stage erythroblasts may have lobulated nuclei. Giant band cells and metamyelocytes are seen.[Figure caption and citation for the preceding image starts]: Megaloblastic marrow cellsPhotomicrograph from Mark J. Koury, MD; used with permission [Citation ends].
Bone marrow examination is not necessary to confirm the diagnosis of folate deficiency, although it can be used to exclude important causes of macrocytic anaemia and pancytopenia, such as myelodysplasia or aplastic anaemia.
Exclude vitamin B12 (cobalamin) deficiency
Co-existing vitamin B12 deficiency should be ruled out before implementing therapy with folic acid because such therapy may resolve the haematological manifestations of vitamin B12 deficiency but allow the neurological complications of untreated vitamin B12 deficiency to progress.
Plasma or serum methylmalonic acid levels increase in vitamin B12 deficiency but are normal in folate deficiency. Homocysteine levels increase in both vitamin B12 deficiency and folate deficiency.[49][50] Levels of methylmalonic acid and homocysteine can be raised in the presence of impaired kidney function.
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