29 Oct 2024
Stacey Newton
Job Title
Image: © Duncan Noakes/ Adobe Stock
The use of haematology and biochemistry is vital in diagnosing many conditions in our equine patients. The timings of taking samples, the tests we request and the interpretation of results are skills we develop as practising clinicians. However, it is not uncommon to be reaching for the phone when our patient’s results come back to discuss the case with the pathologist.
We are extremely fortunate to have experienced pathologists working in our diagnostic labs in the UK. Here, the author shares her expertise, answering questions asked by and relevant to first opinion vets, shedding light on what we are seeing and how best to interpret results.
Q: Acute inflammatory markers are useful to evaluate the kinetics of inflammation, but are there any other reasons to use them?
A: The acute phase proteins include globulins, fibrinogen, high serum amyloid A (SAA), iron and albumin. As well as giving us useful information on inflammation, they can also provide insights to other conditions.
Fibrinogen can be used as a marker of disseminated intravascular coagulation (DIC) in horses, as it is low in many cases of DIC. In these cases, it should be considered alongside D-dimers, platelets, prothrombin time (PT) and activated partial thromboplastin time.
Iron (Fe) is reduced with inflammation, but can also be used as a marker of Fe deficiency anaemia alongside a reduced mean corpuscular volume (MCV; microcytic). But be aware that it is not specific on its own as it may also decrease with haemolysis and chronic anaemia.
Albumin is a negative acute phase protein and when significantly decreased, suggests other causes such as protein loss – for example, protein-losing enteropathy, protein-losing nephropathy and, although rare, hepatic insufficiency.
Q: What is the importance of D-dimers in inflammation in horses, both young and adults? Also, can I combine fibrinogen, SAA and D-dimers?
A: D-dimers are not specific for inflammation as they are part of the coagulation cascade and then fibrinolysis. They are a product of fibrinolysis, and increases are seen in any age of patient.
A D-dimer is a specific degradation fragment of cross-linked fibrin and a high plasma D-dimer is an indicator of intravascular fibrin formation and plasmin-mediated fibrinolysis.
The measurement of plasma D-dimer concentration is useful to aid in the diagnosis of systemic thrombosis, including pulmonary thromboembolism and DIC.
D-dimers can be combined with fibrinogen if suspicion of DIC exists. Fibrinogen and SAA are acute phase proteins that are increased with inflammation.
However, SAA has a much shorter half-life than fibrinogen, so combining the two can be used to determine the course of inflammation and whether it is ongoing or resolving.
Q: Is there any evidence that higher levels of SAA indicate an infectious process rather than just an inflammatory process?
A: We do not have a particular level of SAA that is more likely to indicate an infectious process, but bacterial disease is more likely to be present in many cases (Long and Nolen-Walston, 2020).
Elevations in SAA can increase with inflammation in the absence of infection and are not specific for a certain disease process.
Therefore, elevations in SAA should be evaluated in conjunction with physical examination findings and the results of other diagnostic tests.
Q: Why is polychromasia not seen in horses and why, then, can we use mean corpuscular volume and mean corpuscular haemoglobin concentration?
A: Horses don’t release their red blood cells from the bone marrow until they are more mature than other species, and as such we don’t see polychromasia. By the stage they do release their red blood cells they will have also lost their RNA, which gives the blue tinge to polychromatic cells with normal haematoxylin and eosin (H and E) stains.
However, the red blood cells are larger in size, meaning the MCV is greater. This can be of use in some cases to suggest regeneration, depending on the time of sampling in relation to the incidence of haemorrhage or haemolysis. If mean corpuscular haemoglobin concentration (MCHC) is increased, which in normal circumstances is not physically possible, MCHC suggests intravascular haemolysis, indicating that free haemoglobin is present, as well as that measured in the red blood cells.
Q: Is it worth taking a blood sample when a horse is dull and you are unsure as to why?
A: It is important that you take a full history, clinical examination and signalment along with noting the duration of any clinical signs, such as pyrexia or colic. But alongside this, taking a blood sample in these cases can be helpful and may highlight an abnormality.
When requesting a test, it pays to make sure you know why you want it and what the results will tell you, and bear in mind that if several tests are done, you are more likely to have one or more come back abnormal. The significance of an abnormal result depends on the test and the value – it’s important to remember that reference ranges only cover about 95% of the “normal” population.
Q: Why do different laboratories have differences in reference ranges of haematology parameters?
A: Different laboratories usually establish reference ranges from the population of patients that they receive samples from, which will vary between labs, regions and countries.
Q: How can we tell the difference between bacterial neutrophilia and neutrophilia due to bacterial complications of a viral infection?
A: In most cases, this cannot be differentiated on haemograms. Therefore, other tests run for underlying viral disease based on the clinical signs are used in correlation. For example, primary bacterial causes of respiratory disease are uncommon compared to viral causes. So, if a primary viral cause with a secondary bacterial component exists, we may find that lymphocytosis and neutrophilia is present.
If the bacterial infection becomes severe with a coexisting viral disease also present, we may then see a marked neutropenia with lymphocytosis with or without monocytosis. It may be more likely that thrombocytosis is seen with more severe bacterial infections.
Q: Which parameters of haematology could be helpful in the prognosis of colic?
A: Several parameters may be important in indicating prognosis in colic. However, the underlying cause of the colic should be considered, as some causes will obviously have a much poorer prognosis than others. Also, horses with surgical compared to medical colic will likely have a poorer prognosis.
A recent publication in Veterinary Clinical Pathology looked at some parameters and survival times for colic in a group of 17 horses (Nikvand et al, 2019). Although testing a relatively small group of animals, the study concluded that a combination of prolonged PT, increased D-dimer and low fibrinogen could indicate that an individual is less likely to survive. Also, very high levels of renal and liver parameters would also have a poorer prognosis. This does appear to suggest these horses likely had DIC.
Lactate is also a sign of inadequate tissue oxygenation and high levels may indicate a poor prognosis; however, this is a point-of-care or in-house test as it is too labile to send to an external lab. Cases with persistent high SAA, globulins (acute phase proteins) and low albumin may hold a poorer prognosis.
Q: How can laboratory findings help for diagnosis of laminitis or colitis?
A: Laboratory tests may help with determining the underlying cause in some cases of laminitis and colitis.
For example, laminitis may be due to insulin dysregulation caused by equine metabolic syndrome or equine pituitary pars intermedia dysfunction. Also, underlying systemic disease or neoplasia could be a cause of colitis in rare cases and an overall blood picture may be helpful.
Q: What is the difference between total proteins and total solids?
A: Total protein is run on an analyser that uses a colorimetric method and is measured on serum. Total solids (TS) are usually measured on plasma via a refractometer.
The TS value is usually lower than total protein as the reading can be affected by other serum component “solids” such as sodium, chloride, phosphate, glucose, cholesterol, urea and colloids, such as hetastarch and gelatin.
Q: What happens if there is a high protein level in the blood of an equine patient?
A: High blood protein is not a specific disease or condition, but it might indicate that an individual has a disease.
High blood protein rarely causes signs or symptoms on its own and elevated proteins more often indicate either dehydration (water loss) or inflammation (usually increased globulins).
If the proteins become very high due to either of these causes then the blood has increased viscosity.
Q: How does fasting increase gamma glutamyl transferase?
A: The mechanism for this is not clear, but it is likely similar to the process wherein increased bilirubin results from hepatic dysfunction. The longer the duration of fasting, the greater the increase in gamma glutamyl transferase (GGT).
A longer duration may be due to a metabolic effect on lipid metabolism (particularly in ponies and donkeys), with a resulting mobilisation of triglycerides and development of lipid deposition in the liver causing some hepatocellular swelling and cholestasis.
Q: Are small increases in alkaline phosphatase or GGT significant?
A: If levels of alkaline phosphatase (ALP) or GGT are just above the reference range then the result may not be significant. Correlation with other liver enzymes and other parameters is therefore advised. Repeat blood samples are useful to check for persistence, or presence of an upward trend that may then be significant.
Normal ALP and GGT can be seen with significant liver disease; this is likely due to the decrease in normal hepatic tissue and the number of hepatocytes/biliary epithelial cells. Be aware that bile acids and/or bilirubin are expected to increase in cases of severe or end-stage liver disease.
Q: What is the artefactual effect of hyperlipaemia on biochemical electrolyte analysis?
A: Hyperlipaemia causes a number of analytes to be affected. It causes increased red blood cell fragility and therefore may cause or accentuate current haemolysis, which is then also an added factor that may affect analytes. As an effect on direct measurement of analytes, hyperlipaemia can decrease acute pancreatitis (AP)/ALP and total calcium.
It may, however, increase aspartate aminotransferase (AST), GGT, glutamate dehydrogenase (GLDH), creatine kinase (CK), phosphorus, proteins, electrolytes, bilirubin and lactate dehydrogenase. Other effects may be due to clinical hyperlipaemia and its main effect on the liver with cholestasis, and therefore AP/ALP, GGT, GLDH, AST, bile acids and also triglycerides.
Q: What are the most common bloodwork anomalies in horses with a Fasciola hepatica (liver fluke) infestation?
A: Given the fact that Fasciola hepatica affects the liver then cases with an infestation are more likely to have elevated liver enzymes. However, studies suggest that the changes in haematology and biochemistry in horses with liver fluke are variable, with some cases having no significant changes (Howell et al, 2019).
An increased likelihood appears to exist of testing positive with recently validated F hepatica excretory-secretory antibody detection ELISA if clinical signs are consistent with liver disease.
