Interpreting blood profiles in avians

Sample artefacts

Both haemolysis and lipaemia can have a significant effect on the results obtained (^{Table 1}).

Haemolysis is usually secondary to technical difficulties with sample collection, such as the use of a narrow gauge needle, or excessive agitation of the tube when mixing the sample with the anticoagulant (^{Figure 1}).

Lipaemia is usually associated with a high fat diet, liver disease or reproductive activity (^{Figure 2}).

Reference ranges

Species-specific reference ranges are available for birds commonly kept in captivity and specialist laboratories often have their own data, but for rarer species published material may not be available or may be based on a very small population size. In this situation, results have to be evaluated with reference to a similar species and care should be taken with interpretation taking into account the scientific validity of the range available.

Bearing this in mind, ^{Tables 2} and ³ present biochemical and haematological reference ranges for some more commonly encountered avian species. Where published ranges are not available, the boxes have been left blank.

Haematology

Even if only a small sample can be obtained from an avian patient, a packed cell volume (PCV) and blood smear is informative. Unlike mammals, the erythrocytes and thrombocytes of avian species are nucleated, so cannot be differentiated from white cells on an automated analyser; therefore, a manual white cell count must be performed on all samples.

Avian erythrocytes have a shorter lifespan than those of mammals and so a degree of polychromasia is commonly observed on a blood film. An increased PCV is indicative of dehydration or, less commonly, polycythaemia, which may be primary (very rare) or secondary. Concurrent assessment of total protein, urea and possibly uric acid levels will help to differentiate dehydration from true polycythaemia, which may occur with iron storage disease (most commonly seen in rhamphastids, such as toucans) or chronic respiratory disease, such as macaw respiratory hypersensitivity syndrome.

The aetiologies of anaemia in birds are similar to those seen in mammals. A regenerative anaemia may occur in response to haemorrhage or toxicosis (commonly heavy metal toxicity). Haemoparasites may also cause regenerative anaemia and can be identified on blood smear examination. A non-regenerative anaemia may occur with chronic inflammatory or infectious disease, or neoplasia. Depression anaemias associated with chronic disease are seen more commonly in birds than mammals, due to the shorter lifespan of birds’ red blood cells.

An overall leucocytosis is seen with infectious disease, inflammation or stress, while a leucopaenia may occur due to chronic inflammatory disease or overwhelming bacterial or viral infection. The differential white cell count will further characterise the changes. Psittacine beak and feather disease (PBFD) is an important differential diagnosis for severe leucopaenia in parrots and is often accompanied by a nonregenerative anaemia.

Birds have heterophils in place of the mammalian neutrophil, with these cells fulfilling a similar role, increasing in response to inflammatory disease and stress. Toxic change to heterophils may occur in association with severe inflammation or infection and may be associated with a poorer prognosis.

Monocytes are found in very low numbers in peripheral blood and a zero count is not uncommon, nor considered significant. However, a raised count can be very significant, usually occurring in association with chronic granulomatous infections, such as aspergillosis, chlamydophila and avian tuberculosis. Aspergillosis is an extremely common diagnosis in avian medicine (^{Figure 3}) and, alongside a characteristic monocytosis, affected birds usually present with a heterophilic leucocytosis, lymphopaenia, non-regenerative anaemia and hyperproteinaemia.

Elevated lymphocyte counts are associated with chronic infectious or inflammatory disease and rarely, lymphoid leukaemia. Their numbers will usually decrease in the face of viral infection.

Basophils are present in small numbers in avian species. They may increase in number in response to tissue damage or infection. Reduced numbers are not considered significant.

Biochemistry

• Proteins

Albumin is not accurately measured by most standard wet chemistry mammalian tests. Serum protein electrophoresis is recommended for accuracy and also to separate globulins into alpha, beta and gamma fractions to give more specific information on the nature of inflammatory conditions.

As albumin is synthesised in the liver it is often reduced in the presence of hepatic disease. Reductions in albumin may also occur with chronic disease processes, anorexia or starvation and protein loss through the kidneys or intestine. Increases to albumin are seen with dehydration and female reproductive activity.

Elevations to globulins are consistent with infectious and/ or inflammatory disease. The alpha globulin fraction tends to increase in acute disease, although elevations may also be seen in reproductively active females. The beta globulin fraction includes fibrinogen and increases reflect acute inflammation and infection; an associated increase in the white blood cell count may be seen. The gamma fraction contains the immunoglobulins and, therefore, indicates an immune system response, seen in a more chronic infectious or inflammatory disease state.

• Calcium and phosphorus

When evaluating calcium status it is important to assess ionised calcium alongside total calcium, because approximately one third of total blood calcium is protein bound and, therefore, directly influenced by plasma protein concentration.

Changes in total calcium levels usually have no pathophysiological significance in birds, with increased concentration reflecting dehydration (secondary to increased proteins) or reproductive activity in female birds. Ionised calcium indicates the physiologically active calcium concentration and changes are always significant as normally levels are kept within a tightly controlled range.

Low ionised calcium is very common in pet psittacine birds, especially grey parrots (Psittacus erithacus erithacus) and is almost inevitably secondary to a poor, seed-based diet containing inadequate calcium and vitamin D, together with excess phosphorus and a lack of exposure to natural sunlight (UVB). Nutritional secondary hyperparathyroidism is a common sequel and may lead to juvenile osteodystrophy in chicks (^{Figure 4}).

Hyperphosphataemia is usually related to metabolic bone disease or haemolysis of the sample. The ratio of calcium to phosphorus is also important, with the excess phosphorus relative to calcium found in most seed-based diets potentially leading to nutritional secondary hyperparathyroidism.

• Liver parameters

The diagnosis of avian liver disease can be a challenge because many of the typical mammalian parameters lack either sensitivity or specificity in birds.

Aspartate aminotransferase (AST) can be a sensitive indicator of hepatocellular damage, but it is relatively non-specific, also being found in skeletal muscle, heart, brain and kidney. The result should, therefore, be interpreted alongside creatine kinase (CK) levels as this enzyme is not found in liver cells. Elevation of AST with a normal CK is likely to indicate hepatocellular damage, whereas elevation of both AST and CK is consistent with tissue damage, with or without concurrent liver damage.

Gamma glutamyl transferase (GGT) increases in response to cholestasis and is more specific to liver disease than AST, but the sensitivity of the assay is low. High concentrations have been associated with bile duct carcinoma and elevations may also be identified in obese birds with fatty liver disease.

Glutamate dehydrogenase (GDH) is probably the most specific enzyme for the detection of hepatocellular damage. However, sensitivity is also low – limiting its diagnostic use.

Bile acid assays are the most useful test validated for the evaluation of hepatic function in birds, with increases reflecting impairment. Normal or low bile acids do not rule out advanced liver disease or cirrhosis, where a lack of production may confound the result.

Hepatic lipidosis is commonly encountered in sedentary captive parrots fed on a seed-based, high-fat diet (^{Figure 5}). The condition is particularly prevalent in Amazona species and cockatiels.

In addition to changes to the aforementioned liver enzymes, blood cholesterol and triglyceride levels are often raised in these cases. Cholesterol profiling has been researched in birds to provide a more accurate method of identifying liver disease specifically by measuring high and low density cholesterol. Elevated low-density cholesterol has been shown to correlate well with histopathologic evidence of hepatic lipidosis in psittacine birds and may, therefore, be a helpful diagnostic marker.

• Renal parameters

In comparison with mammals, the major end product of protein metabolism in birds is uric acid. This is produced by the liver and eliminated from the blood via the kidneys, mainly by active secretion by the proximal convoluted tubules of the kidney nephrons. Birds do produce a small amount of urea as a by-product of detoxification, but, importantly, this does not increase significantly with renal dysfunction and is useful only as a marker of hydration status.

Uric acid level can be used to assess renal function, but significant kidney damage must be present before the concentration rises. A rise in uric acid will occur with dehydration, but only when it is severe. To distinguish renal disease from dehydration plasma urea concentration, total protein and PCV may be evaluated concurrently. It is often useful to reassess these parameters after a period of fluid therapy – a persistently elevated uric acid level is likely to indicate renal disease, in which case renal biopsy may be necessary for further investigation. Elevated phosphorus is not usually a feature of avian renal disease.

A high risk exists of visceral or articular gout with prolonged, severe elevations of blood uric acid concentration (^{Figure 6}). It is important to note that significant elevations to uric acid are a normal postprandial finding in carnivorous birds, therefore a 24-hour fast is recommended prior to sampling these birds or care taken interpreting the results.

• Glucose

Hyperglycaemia may be observed and is usually associated with stress. Diabetes mellitus is reported in birds, most frequently budgerigars and cockatiels. This is characterised by repeatedly elevated blood glucose concentrations and clinical signs of polyuria and polydipsia and may be accompanied by hepatic lipidosis.

Hypoglycaemia is generally seen with starvation, sepsis or end-stage chronic disease.

Conclusion

Both the haematological and biochemical evaluation of blood samples are important to assist in rapid diagnosis of the diseases of birds, although care should be taken with the interpretation of results based on the validity of reference ranges available.

Acknowledgements

Thanks to Michael Stanford for his advice and comments and for providing photos. Thanks also to Michael Orsi for photos.

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