Investigation of Anaemia

Anaemia is a reduction in red cell mass and oxygen carrying capacity. It has many different causes, which can be divided into three basic groups:

• Blood loss – haemorrhagic anaemia
• Increased RBC destruction – haemolytic anaemia
• Reduced RBC production – hypoproliferative (if complete, aplastic) anaemia due to:
  • Primary bone marrow disease
  • Secondary to other systemic diseases

All three types can give rise to chronic anaemia but only haemorrhagic or haemolytic anaemia can be acute. This is because of the long life span of erythrocytes (110 days in the dog, 70 days in

the cat). Even if there were a sudden complete arrest in RBC production, a discernible fall in RBC mass would take several days to develop; hypoproliferative anaemias are always chronic.

Clinical signs of anaemia

Nonspecific clinical signs:

• Pale mucous membranes
• Weakness and poor exercise tolerance

Signs associated with physiological compensation:

• Rapid heartbeat and respiratory rate – if severe anaemia, even at rest
• Haemic murmur – a soft mid-systolic murmur in the aortic region

These signs are not seen with mild anaemia but become more pronounced as the severity increases.

All the signs except a haemic murmur could occur with severe shock. (Haemorrhagic shock occurs when 1⁄3 total blood volume is lost over a short period of time or 1⁄2 of the total blood volume is lost over 24 hours). In shock, capillary refill time is prolonged (>2s).

Mucous membranes can appear pale in many non-anaemic cats. With anaemia of comparable severity the signs are more pronounced:

• In the dog than the cat
• If rapid onset - clinical signs are more pronounced with acute than chronic anaemia
• In older animals due to reduced ability to compensate compared with younger animals n In haemolytic compared to haemorrhagic anaemia

Table showing the PCV at which marked clinical signs of anaemia appear

	Canine	Feline
Acute anaemia	PCV <20%	PCV<12-15%
Chronic anaemia	PCV<12-15%	PCV<8-10%

Specific clinical signs:

• Jaundice - associated with acute haemolytic anaemia or following severe haemorrhage into a body cavity
• Petechial haemorrhages - thrombocytopaenia, von Willebrand disease (see section on coagulopathies
• Pyrexia - often accompanies immune-mediated disease
• Hepatic and/or splenic enlargement - may occur with immune-mediated destruction of erythrocytes or platelets, extramedullary haematopoiesis (rarely causes hepatomegaly), lymphoma/leukaemia
• Abdominal pain/enlargement - may occur with intra-abdominal haemorrhage, for example canine splenic haemangiosarcoma or nodular hyperplasia, and hepatic rupture due to feline amyloidosis
• Lymphadenopathy - Lymphoma or possibly leukaemia (myeloid neoplasms can often involve lymph nodes)

An approach to diagnosis and investigation of anaemia

This approach assumes all anaemias are either regenerative or nonregenerative. In reality things are not as clear cut with anaemias often appearing to be (sometimes transiently) poorly regenerative.

How severe is the anaemia?

Very severe anaemia is life threatening and may warrant a blood transfusion.

• Is this a mild, moderate or severe anaemia? This is usually determined by the PCV.

Correlation of PCV with grade of anaemia

	Canine	Feline
Mild	30-37%	20-26%
Moderate	20-29%	14-19%
Severe	13-19%	10-13%
Very severe	<13%	<10%

Is there a statistical chance this is normal?

In any healthy population, for any analyte, there will be occasional individuals whose values for that analyte fall just outside the reference interval. In young puppies the lowest PCV is documented between 4-6 weeks of age (24-34%), gradually increasing to the adult range by 3-6 months.

• Mild anaemia may be secondary to coexisting disease and may not be the most relevant clinical sign to investigate. Proceed to section diagnosis of nonregenerative anaemia
• Moderate-severe anaemia. Proceed to section determine bone marrow response

Is the anaemia regenerative? Determine the bone marrow response.

This is an important question for all anaemias, but particularly for anaemias of moderate or greater severity

• No erythroid response is expected in the first 1-3 days (preregenerative phase)
• Polychromasia and reticulocytosis peak at 4-5 days. In the dog, if there is polychromasia, a reticulocyte count is usually unnecessary because the degree of polychromasia reflects the degree of regeneration (reticulocytosis). In the cat,the degree of polychromasia reflects the aggregate reticulocyte count

A reticulocyte count is useful to quantify the erythroid response in non- and poorly regenerative anaemias.

Erythroid response	Canine Reticulocytes x109/l.	Feline Aggregate reticulocytes x109/l.
None	60 (1%)	<15 (0.5%)
Mild	150 (4%)	50 (2%)
Moderate	300 (10%)	100 (4%)
Marked	>500 (25%)	>200 (>5%)

Late-stage response at 7-14 days. In cats, marked punctate reticulocytosis. In dogs, macrocytic hypochromic erythrocytes.

Red cell indices can be used to classify the anaemia. This is the traditional morphological classification of anaemia using the MCV and MCHC. Note these are mean values of all RBCs and are not as sensitive as some automated analysers which can identify small populations of abnormal cells.

Normocytic normochromic anaemia. Nonregenerative with too few reticulocytes or other immature cells to increase the MCV. This category includes preregenerative anaemia following acute haemolysis or haemorrhage.

Macrocytic hypochromic. This is a regenerative anaemia with increased reticulocytes, which are hypochromic because haemoglobin synthesis is incomplete.

Microcytic hypochromic. Usually indicative of iron deficiency anaemia preventing adequate haemoglobin synthesis. Microcytosis and altered iron metabolism is also seen in dogs with portosystemic shunts and certain other hepatopathies.

Macrocytic normochromic. This is an uncommon category, usually seen in cats as a nonregenerative anaemia associated with FeLV infection or myelodysplasia. Rarely macrocytic normochromic anaemia can indicate vitamin B12 deficiency, for example malabsorption. Macrocytic anaemia has also been associated with folate antagonists such as phenytoin.

Diagnosis of regenerative anaemia

Examine the blood smear. This is very important for diagnosis of haemolytic anaemia. Look for autoagglutination, spherocytes, Heinz bodies, polychromasia, schistocytes etc (see above).

Urinalysis. Haemoglobinuria is consistent with intravascular haemolysis.

Internal haemorrhage. Is or usually becomes strongly regenerative and may resemble haemolytic anaemia (spherocytes are not always seen with IMHA). Check for any evidence of internal haemorrhage with centesis and cytology.

External blood loss. History including possible access to rodenticide. Look for biochemical evidence of hypoproteinaemia and/or hypoalbuminaemia? Check for thrombocytopaenia or other bleeding tendencies (buccal mucosal bleeding time, OSPT, APTT). If the anaemia is microcytic, or faeces are dark, consider GI blood loss. Testing faeces for occult blood may clarify.

Diagnosis of nonregenerative anaemia

This is more complex, there are many potential causes so the diagnostic approach must be tailored to the available evidence.

Complete blood count

• Inflammatory leucogram – anaemia of inflammatory disease, the most common cause of mild anaemia
• Leukaemia, myelodysplasia
• Thrombocytopaenia
• Pancytopaenia or bicytopaenia. Proceed to bone marrow examination

Clinical chemistry profile

• Anaemia secondary to renal disease (usually chronic renal disease) or hepatic disease
• Chronic systemic disease may cause anaemia

Microcytic hypochromic RBCs

• This pattern suggests iron deficiency; the anaemia is often regenerative
• Evaluate total body iron (serum iron alone may be misleading). Reticulocyte haemoglobin concentration may be helpful
• Most common causes include chronic haemorrhage associated with the urinary tract or gastrointestinal tract with anaemia. Urinalysis and faecal occult blood may be useful. In the gastrointestinal tract consider ulceration and/or neoplasia
• Microcytic hypochromic cells may occur with metabolic disease and hepatic disease affecting iron metabolism, with or without concurrent anaemia

Macrocytic normochromic feline RBCs

• With no reticulocytosis, suggests FeLV induced myelodysplasia (see previous page)

Bone marrow examination

• Many indications including persistent nonregenerative severe anaemia, bi or pancytopaenia. Useful to diagnose myelofibrosis, myelophthisic disease, aplastic pancytopaenia, dyserythropoiesis, leukaemia, maturation arrest
• If investigating bone marrow hypoplasia take aspirates and a core biopsy to differentiate between a poor sample and bone marrow hypocellularity

Endocrinology

• Hypothyroidism or other endocrine dysfunction. Choice of test affected by the presence of other clinical signs

Toxicity

• Check for access to oestrogens or testicular neoplasia with feminisation
• Possibly withhold drug therapy, for example occasional blood dyscrasias are associated with certain licensed feline anti-thyroid therapies and anti-epileptic drugs
• Check for chemicals in the environment such as lead from stripping/sanding lead paint when renovating old houses
• Not all toxicities lead to nonregenerative anaemia, onion poisoning and paracetamol toxicity in cats can result in an acute haemolytic crisis

Virology, serology and PCR

• May be useful if suspect underlying chronic infection. Choice of test determined by clinical signs