VetInfo Digest                                                                                                    April 2006

Table of Contents:

General Problems with Drawing Blood

Problems with Specific Test Procedures

Avian Flu

Lyme Disease

This Month's Note

Last month I didn't quite accomplish the goal that I set out to achieve. I wanted to provide some insight into the kinds of problems that cause errors in lab work and also to provide a list of common errors. Hopefully last month's issue had enough explanation that it is possible to see where lab errors occur. This month I just want to provide a list of the common tests and the major complicating factors in interpreting them, from lab error to common misconceptions. In order to get this all into the newsletter it will be necessary to use outline form. If you have problems reading this in the e-mail text version remember that you can always check out the newsletters online in the subscriber area, too.

Avian flu is a hot topic these days and it may affect pet owners in areas in which it occurs. This is more true for cat owners than for dog owners at the present time but since we know that both dogs and cats can be infected with some strains of flu virus it is best to keep up with information that is published on this topic.

General Problems with Drawing Blood

This is something that I forgot to include last month. Often when the report of blood tests is examined carefully there will be a note that the sample was hemolyzed, had high bilirubin levels or was lipemic. Any of these conditions can affect the lab results or in some cases completely invalidate them.

Hemolysis is breakdown of red blood cells. It occurs for a number of reasons, most of them related to problems drawing the blood. In some patients it is almost impossible to get a blood sample without hemolysis due to difficulties getting the patient to cooperate or in finding a vein to draw blood from in a very small patient. Hemolysis affects blood samples in two ways. There are some substances, such as potassium, that may be in higher concentrations in blood cells than in the plasma. These substances can be falsely elevated when red blood cells have broken down. Some of the tests run on serum or plasma are judged by color or wavelength measurements that are affected by the reddish color imparted to the serum when blood cells break down.

The best way to avoid hemolysis in blood samples is to have a calm gentle blood draw into a blood collection tube but this is often difficult to accomplish. In some cases it is necessary to sedate or anesthetize a patient to get a blood sample that is not hemolyzed. When patients are given Oxyglobin (tm), a synthetic source of hemoglobin used in lieu of transfusion, it mimics hemolysis in the blood stream. This isn't a common problem at the present time since Oxyglobin is currently unavailable, but this may change.

Increased bilirubin in a blood sample causes similar problems to hemolysis but is not as easily dealt with because it is generally due to liver disease or pre-existing hemolysis (blood break down happening inside the patient).

Lipemia is a vexing problem in blood samples. By far the most common cause of lipemia in blood samples is simply drawing the blood too close to the last meal. It can take up to 12 hours after a meal for lipid levels to fall to low enough levels to allow for good test results in some dogs and cats. Samples that are very lipemic may also have hemolysis since the lipids destabilize the blood cells. For routine blood testing it is reasonable just to redraw the blood sample after a 12 hour fast when it is apparent that lipemia is present. In emergencies it may be necessary to use lab results from a lipemic patient as a guide, remembering that it will cause false increases in sugar, protein, calcium and phosphorous levels and decreases in a number of blood tests, especially the liver enzyme test results.

If lipemia does not clear up after a 12 hour fast there may be naturally occurring lipemia. This can make it hard for a veterinarian to use in-house lab equipment but many commercial labs can clear most of the fat (lipid) from the blood using ultracentrifugation. There is small centrifuge (StatSpin tm) that can clear lipids in conjunction with a special test solution (Lipoclear tm) but many vets do not have this technology, either.

I think that most general practice vets tend to ignore lipemia and hemolysis unless they are very severe but this sometimes does lead to problems with interpretation of lab work. If there is reason to question lab results and one of these conditions is present make sure that additional blood samples are taken and analyzed if possible.

Red blood cell count (RBC)

• Puppies and kittens have normal RBCs that would be in the anemic range if they were adults.
• Many of the lab machines must be calibrated by species so if the lab report says Fluffy is a canine and you know that she is a cat, remember that the values may be a little off.
• When a dog or cat is anemic it is important to know if the pet's body is trying to respond to the anemia. A reticulocyte count helps in determining this.

White blood cell count (WBC)

• Cats can increase their white blood cell count substantially when they are frightened or stressed, both of which occur in conjunction with vet visits.
• Dogs can also increase their WBC when stressed but don't seem to do this as often as cats.
• Corticosteroid use increases the WBC in most pets and this must be accounted for when interpreting WBC results during corticosteroid use.

Platelets

• Platelet counts are notoriously unreliable. It is very important to examine a blood smear when platelet counts are low in order to help confirm that they are not low due to clumping in the blood or microscopic clotting.
• When a platelet disorder is suspected but hard to confirm it is important to draw two blood samples (using the same needle or Vacutainer collector without removing it from the vein) and to test the second one. This reduces the effect of clotting in the blood due to the needle trauma.

Potassium (low with kidney disease, rises with hypoadrenocorticism (Addison's disease)

• Akitas and English Springer Spaniels can have increased potassium levels in hemolyzed blood as their red blood cells are higher in potassium concentration than most breeds. This also occurs in very young puppies and kittens but is rarely of clinical importance.

Blood Urea Nitrogen (BUN) (rise with kidney disease)

• There is a very critical error in assessing BUN levels that is also very common. It is not possible to assess the meaning of a rise in BUN with certainty without assessing urine specific gravity at the same time. Most veterinarians make this error on a regular basis and our practice is no exception. It is hard to get a urine sample from many patients within the confines of an office visit that is reasonable in length. The BUN level is highly dependent on the hydration status of the patient and whether the kidneys are responding appropriately to that state. From a practical standpoint it is likely that this error doesn't cause too many problems but it is extremely important not to make the error of euthanizing a pet due to very high BUN levels without assessing urine specific gravity at the same time based on a single BUN measurement. I know of instances in which pets were unnecessarily euthanized due to high BUN levels that were actually appropriate responses to the pet's situation -- where correctable problems existed.
• Severe liver disease can cause the BUN levels to be artificially low.
• Protein restriction in the diet decreases BUN and excessive protein increases BUN.

Creatinine (rise with kidney disease)

• It is a good idea to consider the urine specific gravity when assessing creatinine levels but it is not quite as critical as it is for BUN.
• Pets who are rapidly losing muscle weight and pregnant pets may have artificially low creatinine levels. This can sometimes interfere with interpretation of lab work in dogs being treated for cancers that are causing rapid weight loss.

Calcium

• Calcium levels are very subject to laboratory error and are affected more by lipemia and increased bilirubin levels than most tests. It is always best to assume there may have been a laboratory error when calcium levels are out of the normal ranges.
• Some of the blood tubes used for drawing blood contain substances that complex with calcium. It is important that the right tubes be used for collecting samples for calcium measurement but this is widely known and so errors are infrequent.
• Ionized calcium (free calcium) levels are very subject to errors in handling the blood as the serum must be separated without exposure to air and the sample must remain frozen in transport to the lab. An iStat (tm) machine made for use in clinics gives an accurate ionized calcium level and helps to avoid the problems with sample handling.
• Corticosteroids and diuretics decrease calcium levels which can be important when monitoring levels over time.

Glucose

• The most common error affecting glucose levels is failure to separate the blood cells from the plasma or serum within 30 minutes of drawing the sample. RBCs and WBCs continue to use glucose in the blood until they are separated from it. The glucose level can fall as much as 20% per hour until the serum or plasma is separated from the cells.
• Failure to apply a sufficient blood sample causes some handheld glucose meters to give low glucose levels rather than an error message. If your pet is diabetic and you are using a glucometer to keep track of glucose levels at home don't panic if the sugar is low but your pet is normal. Redraw the sample first and if the results are the same, then administer oral glucose and contact your vet. If your pet is acting odd, give a sugar source first and worry about retesting later.
• Corticosteroid use increases blood sugar levels as does naturally occurring hyperadrenocorticism (Cushing's disease). It is important to remember this when monitoring patients likely to have high blood cortisol levels.

Cholesterol and Triglycerides

• By far the most common cause of high cholesterol and trigylceride levels is the rise in these levels from a recent meal. A true 12 hour fast is necessary when measuring cholesterol and triglyceride levels.
• Cholesterol levels rise with a number of diseases, especially hypothyroidism, diabetes mellitus, hyperadrenocorticism, pancreatitis and kidney disease. It is necessary to think about other primary causes before assuming that there is a naturally occurring high cholesterol level.
• Corticosteroid use (usually prednisone) can increase cholesterol levels in both dogs and cats. Megestrol acetate ( Ovaban tm, Megace tm) can cause cholesterol rises in cats.

  Total thyroxine ( TT4) and free thyroxine (fT4ed ) measured by equilibrium dialysis

  • Total thyroxine levels can be low if antithyroid antibodies are present, which can be a problem in dogs. The presence of antithyroid antibodies does not always indicate that hypothyroidism will occur. Free T4 levels are not affected by antithyroid antibodies.
  • Thyroxine levels vary during the course of the day, so if clinical signs of hyperthyroidism are present in a cat but thyroid levels are within the normal range it is best to recheck the thyroid levels another day at a different time. This variation is less of a problem with testing for hypothyroidism in dogs.
  • Almost any concurrent illness can cause thyroid hormone levels to drop. A diagnosis of hypothyroidism that is made when a dog is severely ill should always be suspect. Ruling out hyperthyroidism in a sick cat is also difficult due to this problem because a very ill cat will have lower thyroid levels, possibly dropping high values into the normal range. Taking the time to re-test when the dog or cat feels better can be very helpful and may prevent the use of a medication lifelong in a dog or allow accurate assessment of hyperthyroidism in a cat.
  • The use of glucocorticoids (prednisone, DepoMedrol Rx, etc) will lower T4 levels. This can have a very significant effect and is important to consider when interpreting this blood test.
• Greyhounds and other sighthounds have lower thyroxine levels than other dog breeds.

Lipase and Amylase -- Used to evaluate pancreatic disease

• High triglyceride levels can cause rises in lipase measurements in dogs.
• Amylase and lipase both rise in some dogs and cats early in kidney disease in some pets and will sometimes rise to levels that can confuse clinicians who are looking for rises in these enzymes due to pancreatitis.
• Neither amylase or lipase levels are very specific indicators of any disease state. There are better tests for pancreatitis now in both dogs and cats. Immunoreactive pancreatic lipase testing for dogs and cats is more accurate and is available through the GI Laboratory at Texas A&M veterinary school.

Alanine Transferase (ALT) -- Liver associated

• Alanine transferase is an enzyme that is found in liver cells. It is only found in the circulation when liver cells have been damaged sufficiently to leak it, or have died. Very high rises in ALT levels can occur with damage to a fairly small number of cells so this test is not a good way to assess the overall health of the liver, even though it is a very sensitive test for liver injury.
• Over 30 drugs are known to cause ALT rises in at least some patients.

Serum Alkaline Phosphatase (SAP, alk phos) -- Rises with liver damage or bone cancer

• Alkaline phosphatase levels are much higher in young dogs than in mature dogs so high levels in young dogs may be normal.
• Corticosteroid use very frequently causes rises in alkaline phosphatase levels without causing liver damage. Phenobarbital and primidone can also cause significant elevations in SAP.
• Many veterinarians seem to automatically assume that high alkaline phosphatase levels in an older dog when other liver enzyme levels are normal indicates that hyperadrenocorticism (Cushing's disease) is present. This is not true. Another common cause is benign nodular hypertrophy of the liver and there are several other possible causes, as well.
  
  

Serum Protein Levels

• Total protein is a measurement of albumin levels and antibody (immunoglobulin) levels in the blood stream. Abnormal levels of either albumin or immunoglobulins can cause increases on decreases in total protein so the value of this number alone is limited. It is necessary to measure albumin levels in the serum at the same time as total protein to truly evaluate serum protein levels.

It may seem to you that proper handling of blood and other laboratory specimens should just be a routine part of veterinary practice. For the most part this is true. However there are always going to be exceptions to this even in the best managed veterinary practices. It is simply outside the bounds of human behavior to perform perfectly in every situation. Just a general awareness that almost every test has pitfalls and keeping in mind that humans make mistakes when they are busy or distracted is helpful. If you have any reason not to trust lab results ask your vet to explain the interpretation or proper testing procedures so that you can air your worries.

Avian Flu

Several subscribers have written to me over the last month asking for information about avian flu and its possible effect on pets. Prior to the last year or so I was pretty confident that flu viruses were not very important in small animal practice with the exception of vets who treat ferrets. This situation has changed dramatically over the last few months, though.

Flu viruses have become much more important to small animal vets since the reported cases of avian flu (Influenza A, type H5N1 virus) infection in cats in Europe and infection of dogs with a flu virus (Influenza A, type H3N8) that originated in horses. Cats and dogs were thought to be resistant to influenza viruses in general prior to these recent developments. While there are still no reported cases of avian flu in dogs that I am presently aware of, it is hard to say they won't be affected at some point, now that it is clear that flu viruses can mutate to infect dogs in some cases.

The risk to pets in the United States and other areas that have not seen bird flu at the present time is very low. When avian flu reaches the United States, which seems likely, there are some precautions that can be taken to lessen the possibility of infection in individual cats and dogs. The major risk to pets is ingestion of infected birds. Therefore, the first step is to keep cats inside where they won't be tempted to hunt infected birds who may be easy targets since they don't feel well. Keeping dogs on a leash when they are out so that they won't find and ingest dead birds is a good idea as well. If the avian flu has affected poultry in your area it would be unwise to feed uncooked poultry meat to pets.

There is evidence that cats can pass the H5N1 influenza virus from one cat to another. This does make it necessary to consider precautions such as keeping cats from possible contact with other cats. This a another reason to keep cats indoors if the flu viruses occur in your region. This problem would make visits to the veterinarian's a little scary as there would be an increased risk of exposure to infected cats. It is always a good idea to use a cat carrier when taking your vet to the vet and this would increase the need for that precaution. A cat in a carrier is not nearly as likely to be exposed to the virus.

At the present time there is no evidence that the H5N1 virus can spread from an infected cat to a human. While this situation could change over time it is not necessary to worry much over this possibility right now but it is always a good idea to wash your hands after handling your cat and this would be especially true for cats who appear to be ill.

I am hopeful that the avian flu virus H5N1 will not turn out to be the one that causes a global pandemic. Unfortunately, I think that even if this virus isn't the one, there will be a future pandemic. If it does appear that it can infect either dogs or cats it will be necessary to consider measures to keep you pets safe. Limiting contact with sick animals would be a good first step and paying attention to the web sites from Cornell http://www.vet.cornell.edu/fhc/news/influenza.htm

and the AVMA http://www.avma.org/public_health/influenza/avian_faq.asp is probably a good idea. The AVMA site has links to a number of government sites with additional information.

Don't panic, but do pay attention!

Lyme Disease

The American College of Veterinary Internal Medicine released a " report" on Lyme disease this month. While there isn' total consensus among the reports authors, there is valuable information in the report and so I would like to share an overview with you.

Lyme disease is caused by a bacteria, Borrelia burgdorferi. This bacteria is carried primarily Ixodes species of ticks. While these ticks have a wide range in the United States, Lyme disease cases are concentrated in the Northeast, upper MidAtlantic states and the Northern Midwest. In order of the number of infections reported, the following 12 states had 95% of the infections in the U.S: PA, NY, NJ, MA, CT, RI, WI, MD, MN, DE, VA and NH. It is thought that this distribution is probably due to the preferred hosts for the Ixodes tick in these areas having habitats that overlap those of pets and people in the region. To best assess the risk in your area check the CDC incidence map at this URL: http://www.cdc.gov/ncidod/dvbid/lyme/ld_Incidence.htm

This may come as a surprise, but except for one study done in puppies less than 12 weeks of age, the gold standard for proof that a disease exists, meeting Koch' postulates, has not been met for Lyme disease in dogs, making it somewhat questionable whether this disease affects dogs by itself. Cats are not thought to be susceptible to Lyme disease at the present time. Over 90% of dogs in some endemic areas have antibodies against B. burgdorferi and most have not had a clinically detectable illness resulting from exposure to the Lyme disease organism. It is possible that immunosuppression or a second infectious organism from tick bites may be necessary to cause disease. If so, this might explain the difference between the large number of dogs exposed to the disease and the small percentage that develop clinical signs.

Due to the problem proving that B. burgdorferi can cause Lyme disease in dogs as a sole agent, there is a lot of controversy over whether to routinely test for the disease, whether to routinely vaccinate for the disease and what to do when a test is positive. I think that if you live in one of the 12 states that account for the majority of the disease cases you should probably work with your vet to decide the risk to your pets and make vaccination decisions accordingly. Try to limit exposure to ticks and use a good tick control product. Do this even in areas in which Lyme disease isn' a major problem as other tick diseases should also be controlled. In the rest of the states I think you should avoid vaccination as the risk of this disease is very low. Treatment is warranted if signs of Lyme disease are present, even in lower risk states. At least one month of daily doxycycline is current thought to be best. Dogs with severe kidney disease characterized by protein loss in the urine (proteinuria) may have Lyme nephropathy. Prompt and aggressive treatment is necessary for dogs to survive this form of the disease. Hopefully we will know more about this disease in the future so that there is a true consensus over how to identify and treat Lyme disease in dogs.

Thanks for your Support!

VetInfo Digest

P.O. Box 476

Cobbs Creek VA 23035

All opinions in this newsletter are those of the author, Michael Richards, DVM

Copyright 2006