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11 - Biochemistries
11 - Biochemistries
11 - Biochemistries
11
tries are necessary to evaluate cellular changes.
Properly evaluating a biochemical profile requires
knowledge of the diagnostic sensitivities and specifi-
cities of tests, correct intervals for a specific test in a
given species and a list of diseases that can induce
the observed changes.
precision of 9.0%, which is much better than its disease. Until reference intervals are established for
day-to-day variation, at least in humans, of 13.0%. birds free from subclinical infections (particularly
viral diseases) and maintained on adequate diets for
In birds, some blood chemistry variables may show a long periods, it will be impossible to define precise
circadian rhythm (ie, plasma corticosterone) or a reference intervals on a population basis. Many nor-
circannual rhythm (ie, plasma thyroxine).38a,49a Be- mal values published in the literature have been
cause of these inherent problems in determining spe- collected by diagnostic laboratories, which generally
cific serum chemistry values it is important to have receive samples from abnormal birds.
a basic knowledge of the technical and statistical
methods used in establishing the value of these tests. In addition to varying among populations, reference
Developing this working knowledge is further com- intervals may also vary among laboratories because
plicated in avian medicine due to a lack of knowledge of variation in test methods. A laboratory must be
concerning the day-to-day variations that occur in able to provide a reference interval established in
different biochemical parameters in different avian that laboratory for the species and variables con-
species. cerned, or the results from the laboratory will be of
little value.
To be of optimum use, clinical chemistry data must
be evaluated based on the values in healthy individu- In interpreting clinicopathologic data, it should be
als, the precision of quantitative measurements and noted that:
the clinical chemistry changes characteristically ex-
pected in various pathologic states. 1. There are subtle changes that exist between
health and disease. The concept of normality
rarely exists.
2. Not all values from healthy individuals will fall
within a normal reference interval (usually en-
Reference Values – compasses 95% of healthy individuals).
3. Some values from abnormal individuals will fall
Reference Intervals within the reference interval.
An International System of Units, the so-called SI An enzyme that is released into the serum/plasma
Unit System (Systeme International d’Unites) was must be easy to assay in order to be of diagnostic
adopted in the 1970’s to standardize measurements. value. In addition, the assay must be economically
feasible and indicate pathologic changes in a specific
Standardization created a change in the numerical organ, or a defined small group of organs. The en-
values of many frequently used tests. The mol, for zyme must also be stable in the serum/plasma for a
example, indicates the amount of a substance in sufficient time to permit its detection.
terms of molecules. The concentration of all sub-
stances is now expressed in terms of liters. For bio- It is important to realize that cells must be damaged
chemical substances, the molar concentration per before they release enzymes into the serum/plasma.
liter, which is expressed in sub-multiples (millimoles Therefore, enzymatic-based tests are a measure of
per liter - mmol/l or micromoles per liter - µmol/l) is cell damage, and not necessarily a measure of organ
the preferred standard of measurement. function. Anoxia causes the cell membrane to lose its
integrity so that soluble enzymes from the cytosol can
The advantages of an accepted, worldwide stand- leak into the serum/plasma.
ardized system are obvious. Unfortunately, the
standardized system is not always used to report With liver disease, it is common to have normal
data. Many refereed journals continue to use the histology with marked biochemical changes.This loss
conventional rather than SI units. All publications of integrity may be observed histologically as a swel-
before 1975 used the conventional units, and even in ling of the cell. Anoxic red blood cells, for example,
countries that are committed to SI units there are leak cytosolic LDH into serum/plasma, causing an
laboratory instruments still in use that produce re- increase of LDH activity in a sample. Combining the
sults in conventional units. Most enzyme activities values obtained for several enzymatic assays will
are still expressed in terms of international units per increase the diagnostic value of the biochemical
liter (IU/l or U/l) because the SI unit, the catal, has evaluation of a patient.
not been widely adopted. It is often necessary to Enzyme activities in tissue or serum/plasma are usu-
convert values expressed in conventional units to SI ally in such low concentrations that it is not practical
units. Conversion tables are provided in the Appendix. to quantitate the enzyme directly. Therefore, enzymes
are measured indirectly based on their in vitro activ-
ity under controlled or specific conditions at which
their activity is proportional to enzyme concentra-
tion. There are a multitude of methods used by differ-
ent laboratories for detecting enzyme activities, and
the reference intervals will vary among these labora-
tories despite all results being expressed in U/l. Test
227
CHAPTER 11 BIOCHEMISTRIES
values will vary depending on the substrate, buffer µl/parameter) or automatically (Autoanalyzer Two
and incubation temperature used by the laboratory. only requires 20 µl/parameter).
Historically, wet chemistry systems have been used Diagnostic Value: The validity of a parameter in
for evaluation of blood parameters. Wet chemistry suggesting or confirming the presence of disease.
means that liquid reagents act with a certain volume
of sample under strictly defined constant conditions Physiologic Influence: The influence of physiologic
(eg, temperature, pH, time) and produce a change of conditions on a test assay.
color that is proportional to the concentration of sub- Pathologic Changes: The effect that pathologic
stances or the activity of enzymes. As the indicator changes have on test values with reference to special
dye changes color, the reaction is read spectropho- literature.
tometrically. Because the reagents can be prepared in
the laboratory, the cost for frequently used tests is Reference intervals for different avian species using
inexpensive on a per test basis. The minimum sam- various testing methods are provided in the Appendix.
ple size often depends on whether reagents are added
by hand (older systems that may require 100 to 200
228
SECTION TWO PATIENT EVALUATION
TABLE 11.1 Recommended Samples for Biochemical Tests41 TABLE 11.2 Causes of Tissue Enzyme Increased Activities41
Calcium Heparinized plasma or serum (see text) GGT Biliary and renal tubular Hepatocellular damage
epithelium Some renal diseases
Cholesterol Heparinized plasma or serum
GLDH Mitochondrial enzyme Hepatocellular necrosis
Creatinine Heparinized plasma or serum found in most tissues
Glucose Heparinized plasma or serum (see text) Liver, kidney, brain
Iron Heparinized plasma or serum LDH Skeletal muscle, cardiac Hemolysis
muscle, liver, bone, Hepatic necrosis
TIBC Heparinized plasma or serum
kidney, RBCs Muscle damage
Lipase Heparinized plasma or serum
TP Heparinized plasma or serum involved in the conversion cascade, AST and ALT are
Triglycerides Heparinized plasma or EDTA plasma the two enzymes of greatest clinical importance.
Urea Heparinized plasma or serum
Uric Acid Heparinized plasma or serum Diagnostic Value: Alanine aminotransferase activ-
Electrolytes Sample ity occurs in many different tissues. Specific diagnos-
Chloride Heparinized plasma or serum tic value of these enzymes in birds is poor. In many
Potassium Heparinized plasma or serum cases, patients with severe liver damage have had
Sodium Heparinized plasma or serum normal ALT activities, reflecting a low level of en-
zyme activity in liver cells from certain species. Alan-
ine aminotransferase activities often increase due to
Enzymes damage in many different tissues. In some avian
species, normal ALT activities are below the sensitiv-
Alanine Aminotransferase ALT (GPT) ity of many analyzers.35,36
Method: It is not possible to monitor transaminase
(ALT and aspartate aminotransferase AST) reactions Pathologic Changes: Elevated activities are difficult
directly; however, continuous monitoring assays can to interpret, and this enzyme has limited usefulness
be performed by coupling the transaminase reactions in birds because it can be increased by pathologic
to specific dehydrogenase reactions. Because of the changes in almost all tissues. Activity in erythrocytes
value of AST and ALT activities in diagnosing dis- is 1.6 times higher than in plasma, and hemolysis will
ease, standardization of reference methods for these cause elevated activities.50
two enzymes have been given priority by national and
international groups. These groups have chosen a Physiologic Influence: Age-dependent elevation (in-
coupled-reaction with malate or lactate dehydro- creased activity with aging) of enzyme activity has
genase as the indicator enzymes. These methods been described in birds.25 In raptors, seasonal vari-
differ with respect to substrate concentration, nature ation in ALT activities has been reported. These
of buffer and assay temperature.68 changes were independent of reproductive activity.22
use make it difficult to compare AP activities between Physiologic Influence: Aspartate aminotransferase
laboratories or reference literature. values are age-dependent to varying degrees among
different species.25,15,16,17 The cause of this age-de-
Physiology: Alkaline phosphatase operates at an al- pendent increase in activity has not been defined.
kaline pH and is possibly involved in energy transfer Gender differences have not been described.4,32
for exchange of ions across the cell membrane. Alka-
line phosphatase activity has been found to occur Pathologic Changes: In general, AST activities in
predominantly in the duodenum and kidney. Low AP birds greater than 230 U/l are considered abnormal.
activities were reported in the liver with no activity Abnormal activities have been linked to vitamin E,
in other organs of pigeons.50,51 Similar findings have selenium or methionine deficiencies,25 liver damage
been described in chickens8 and turkeys.9 Most en- (particularly psittacosis or Pacheco’s disease vi-
zyme assays are used to document damage to cells rus),23,61,63 pesticide and carbon tetrachloride intoxi-
resulting in enzyme release. In contrast, plasma AP cation43 and muscle damage. Intramuscular injec-
activity is induced by increased cellular activity (in- tions of irritating substances may cause elevation of
creased synthesis) rather than cell damage. CK with no increases in AST activity. In other pa-
tients, both the CK and AST activities will increase
Diagnostic Value: Alkaline phosphatase activities post-injection.
may be elevated due to irritation of the cells in differ-
ent tissues. Increased activities have no specific im- Creatinine Kinase - CK (CPK)
portance. Method: Numerous colorimetric, fluorimetric and
coupled enzyme assays have been developed to detect
Physiologic Influence: Juvenile birds have signifi-
CK activity.68
cantly higher AP activities from bone growth and
development than adults.15,16,17,32 In hens, activities Physiology: Creatinine kinase functions in skeletal
are elevated prior to egg laying.28 Seasonal variations muscle, heart muscle and brain tissue. In muscle, this
in activities have been described.4 enzyme makes ATP available for contraction by the
phosphorylation of ADP from creatinine phosphate.
Pathologic Changes: Elevations are most common
There are three isoenzymatic forms of CK that can be
with liver disease even though the level of activity in separated by electrophoresis. In mammals, quantiza-
this organ is low. Hyperparathyroidism-induced tion of isoenzymes can be used to determine the tissue
stimulation of osteoplastic activity may also cause source of the enzyme.39,68 There have been no reported
increased AP activity. Enteritis has been described
attempts to separate tissue-specific creatinine ki-
as a cause of higher AP activities but activity of this
nases in birds.
isoenzyme is labile and difficult to measure.36 Afla-
toxin B1 intoxication with massive liver destruction Diagnostic Value: Elevations in activities are
in pigeons, cockatiels, Red-tailed Hawks and Great mostly seen because of muscle cell damage. This
Horned Owls was not found to significantly increase enzyme has value in distinguishing muscle from liver
AP activity.13 Low AP activities have been linked to cell damage. However, the clinician should consider
dietary zinc deficiencies. that muscle and liver cell damage can occur simulta-
neously from the same or different pathologic proc-
Aspartate Aminotransferase - AST (GOT) esses.
Method: See ALT.
Physiologic Influence: In mammals, CK activity is
Physiology: See ALT. subject to a number of physiologic variations (eg,
muscle mass of an individual, gender, age, physical
Diagnostic Value: High AST activity has been de-
activity).39,68 Physiologic changes of CK are well
scribed in liver, skeletal muscle, heart, brain and known and are also described in avian species. CK activ-
kidney cells. The distribution of AST in avian tissues
ity in healthy turkeys is extremely sensitive to physi-
varies among the species.43,25,50 Elevated activities are cal stress and exercise.50 Neither gender32 nor age15,16,17
usually indicative of liver or muscle damage. Aspar-
has been shown to significantly affect CK activity.
tate aminotransferase activity provides the best in-
formation when combined with other more specific Pathologic Changes: Increase in CK activity has
tests.49,36 Creatinine kinase activity can be used to been linked to muscle cell necrosis, convulsions, in-
exclude muscle damage as a cause of increased AST tramuscular injections (depending on the volume and
activity. degree of irritation), vitamin E and selenium defi-
230
SECTION TWO PATIENT EVALUATION
ciencies, neuropathies, lead toxicity and occasionally been observed in birds with liver disease, but few
chlamydiosis.36,43 reference intervals are available for avian species.36
Physiology: Glutamate dehydrogenase is a mito- Diagnostic Value: Although this enzyme is not spe-
chondrial enzyme found in numerous tissues. cific for any organ, elevations are most common with
hepatic disease in psittacines. Lactate dehydro-
Diagnostic Value: Significant amounts of this en- genase activities are thought to rise and fall more
zyme have been found in the liver, kidney and brain quickly than AST activities in birds with liver dis-
of chickens, ducks, turkeys and racing pigeons.50,51 In ease.61 These differences may provide information on
budgerigars, the highest enzyme activity has been the chronicity of liver disease.
reported in the kidney.50 Significant elevations have
231
CHAPTER 11 BIOCHEMISTRIES
activity of approximately 30% of avian samples will the blood and recycled. Only a small percentage of the
exceed the upper range limit of the test. Samples that total pool of bile acids is lost in the feces each day. A
exceed the test limit must be diluted and reanalyzed. small quantity of the total bile acids reabsorbed from
the gastrointestinal tract is not removed from the
Physiology: Amylase occurs in plasma as a number blood by the liver and reaches the general circulation.
of isoenzymes that are principally derived from the It is this fraction of unextracted bile acids that is
pancreas, liver and small intestine. In birds, the measured. The quantity of bile acids in the plasma
isoenzymes have not been separated, making it im- normally increases following the ingestion of food.
possible to determine which specific tissues are re-
sponsible for increased plasma amylase activity. Diagnostic Value: If liver function is impaired, bile
acids are not properly reabsorbed from the blood, and
Diagnostic Value: Little information is available on consequently the proportion of excreted bile acids
amylase activity in birds. In some cases it has been reaching the peripheral circulation increases. Circu-
found to be useful in the diagnosis of neuropathic lating bile acids can therefore be used as a sensitive
gastric dilatation.36 indicator of liver function, and of the integrity of the
Pathologic Changes: Increased enzymatic activity circulation through the liver, biliary tract and intes-
can be seen with acute pancreatitis. In these cases tines. It has been suggested that chronic liver disease
enzyme activity may exceed three times the upper that results in cirrhosis may decrease the production
limit of the reference interval. Activities less than of bile acids with a subsequent decrease in the
twice the upper limit of the reference interval are plasma. This may be particularly true in a post-
sometimes seen in macaws with severe enteritis in prandial sample. Further investigations are needed
the absence of pancreatic lesions. In most cases of to determine if decreased bile acid concentrations are
neuropathic gastric dilatation, amylase activity is a reasonable indicator of a loss of functional liver
normal or only slightly elevated.36 mass. Low bile acid concentrations are common in
birds with microhepatia (as detected radiographi-
Reference Values: Budgerigars (187-582 U/1); Afri- cally), poor feather formation and an overgrown, mal-
can Grey Parrots (211-519 U/1); Amazon parrots formed beak.
(106-524 U/1); macaws (276-594 U/1) (Kodak Ek-
tachem, Amylopectin, 25°C).33 Physiologic Influence: A significant postprandial in-
crease of bile acids has been documented in racing
Bile Acids pigeons and the Mallard Duck. Healthy birds with a
Method: Several assays have been used to quantitate gall bladder may not have significantly different post-
either total or individual bile acids. The most fre- prandial bile acid concentrations when compared to
quently used assays are gas liquid chromatography, species that do not have a gall bladder.55
high performance liquid chromatography, enzymatic
Pathologic Changes: Elevations in bile acids have
assays and immunoassays (RIA, ELISA). Among
been shown to correlate with liver disease in pi-
these, RIA and enzymatic methods are mainly used
geons,55 chickens10 and African Grey Parrots.50 With
by commercial laboratories. RIA-derived values are
further research, bile acid assays may prove to be one
not comparable to those detected using other meth-
of the best tests for liver function in birds.38 Bile acids
ods. Nonspecific cross reactions occur when human
are stable in plasma for prolonged periods, allowing
anti-bile acid antibodies are used to detect bile acids
shipment of specimens to distant laboratories for
in bird plasma; therefore, enzymatic methods seem
analysis.
to be the assay of choice for use in birds.
Reference Intervals: African Grey Parrots (18-71);
Physiology: The liver synthesizes the primary bile
Amazon parrots (19-144); cockatoos (23-70); macaws
acids (cholic acid and chenodeoxycholic acid). It then
(25-71).
excretes these acids as sodium salts into the bile.
With the ingestion of food, bile is carried via the bile Bilirubin
duct into the small intestine where the bile acids act Method: Most methods for measuring bilirubin are
principally as emulsifying agents in fat digestion and based on the diazo reaction, in which diazotized sul-
absorption. Most bile acids that enter the gastroin- fanilic acid reacts with bilirubin to produce two
testinal tract are reabsorbed in the distal small and azodipyrroles. These products are reddish purple at
large intestines where they return, via the portal neutral pH, and blue at low or high pH.
circulation, to the liver. They are then extracted from
233
CHAPTER 11 BIOCHEMISTRIES
Physiology: In birds, the major bile pigment is Diagnostic Value: Total calcium should always be
biliverdin. The enzyme biliverdin reductase is ab- interpreted along with albumin concentrations. Hy-
sent, and biliverdin is not converted into bilirubin.44,45 poalbuminemia will reduce the quantity of bound
calcium and result in a decreased total calcium con-
Diagnostic Value: Low concentrations of bilirubin centration without reducing biologically active cal-
were detected in the sera of healthy ducks. Concen- cium (ionized fraction).31,36,53 The hyperproteinemia
trations increased following infection with duck that occurs with dehydration may result in an in-
hepatitis virus.1 The diagnostic value of bilirubin creased total calcium concentration.
appears to vary among species. It has no value in
chickens that cannot form bilirubin, but may be of Physiologic Influence: Ovulating hens have signifi-
value in other species. cantly higher calcium levels than non-reproductively
active females. Female budgerigars were found to
Pathologic Changes: Bilirubin cannot normally be have significantly higher calcium concentrations
detected in plasma of normal psittacines. With severe than males. Young birds generally have lower cal-
hepatic disease (eg, chlamydiosis or Pacheco’s disease cium concentrations than adults.32,27
virus) bilirubin concentrations up to 44.5 µmol/l have
been reported. A slight yellow coloration (icterus) Pathologic Changes: Decreased calcium concentra-
could be seen in the facial skin of two macaws with tions are common in seizuring African Grey Parrots.
bilirubin concentrations exceeding 40 µmol/l. This hypocalcemia syndrome has been described as a
unique form of hypoparathyroidism in which calcium
Calcium is not properly released from bone.31,33,36 Glucocorti-
Sample: Heparinized plasma or serum can be used. coid therapy will decrease total calcium concentra-
Some calcium-binding anticoagulants, like EDTA, tions. Increased calcium concentrations have been
citrate and oxalate (fluoride oxalate is used for deter- reported with dietary excesses of Vitamin D,
mining glucose levels in mammals) will cause falsely osteolytic bone tumors and dehydration. Even in
low values. For the determination of ionized calcium cases of severe dietary calcium deficiency, parahor-
levels, whole blood, heparinized plasma or serum can mone will normally mobilize bone to maintain cal-
be used, but the pH of the specimen must be the same cium blood concentrations within physiologic limits.
as that of the patient’s blood at the time of sampling.
This is most readily achieved by collecting and proc- Cholesterol
essing the specimen quickly and anaerobically. Method: Cholesterol consists of both free cholesterol
and cholesterol esters, which are measured together
Method: Total calcium concentrations include the as total cholesterol. Either enzymatic or chemical
sum of biologically active ionized calcium, protein methods can be used for quantification. Enzymatic
bound calcium (which is bound mainly to albumin) procedures have virtually replaced chemical methods
and calcium chelated with anions, like phosphate or in the clinical laboratory. The initial reaction steps
citrate. Bound calcium is biologically inactive and can are common to all enzymatic procedures. These in-
be decreased (thus decreasing the measurement of clude the hydrolysis of cholesterol esters to form free
total calcium) without causing any clinical effects. Of cholesterol, which is measured after a subsequent
the many methods described to measure total cal- oxidation step utilizing O2 to produce H2O2.
cium, atomic adsorption spectrophotometry and spec-
trophotometry of calcium-dye complexes are most Physiology: Cholesterol is a major lipid that is a
often used. Ionized calcium levels have been shown precursor of all the steroid hormones and bile acids
to be clinically valuable; however, this is not a com- as well as a component of the plasma membrane of
monly available assay. cells. It is obtained from the animal protein sources
in the diet as well as being synthesized by the liver.
Physiology: As a major constituent of bone, calcium
plays a vital role in the structure of the body. It also has Diagnostic Value: Elevated and decreased choles-
important physiologic functions involving the trans- terol concentrations may occur from a number of
mission of nerve impulses, the permeability and excit- physiologic influences and different diseases; how-
ability of all membranes, the activation of enzyme ever, the diagnostic value of this test in birds appears
systems (eg, blood clotting), calcification of egg shells to be poor. Very high cholesterol concentrations usu-
and contraction of the uterus during oviposition. ally accompany lipemia, especially in Amazon par-
rots, macaws and Rose-breasted Cockatoos with fatty
liver degeneration.
234
SECTION TWO PATIENT EVALUATION
Physiologic Influence: Cholesterol concentrations septicemia (eg, chlamydiosis), renal trauma and
will vary with a bird’s diet. Carnivorous birds have nephrotoxic drugs.43
higher concentrations, whereas fruit- or grain-eating
birds have lower concentrations.43 Male budgerigars Glucose
were found to have significantly higher cholesterol Sample: Heparinized plasma or serum can be used.
concentrations than females.32 For reliable glucose determination in avian blood, it
is not necessary to prevent glycolysis as long as the
Pathologic Changes: Elevations can occur because blood is not stored for more than two hours.57 This is
of hypothyroidism, liver disease, bile duct obstruc- contrary to the situation in mammals in which so-
tion, starvation or high fat diets.2,25,73,36 High choles- dium fluoride is often used to ensure accurate glucose
terol concentrations have been reported in budgeri- determinations. This is because avian erythrocytes
gars with xanthomatosis.36 Decreased cholesterol consume very little, if any, glucose, and depend pri-
levels have been associated with some cases of liver marily on fatty acid metabolism for energy.
disease, aflatoxicosis,73 reduced fat in the diet, Es-
cherichia coli endotoxemia and spirochetosis.25 Method: Glucose levels may be determined using en-
zymatic (eg, hexokinase) or colorimetric (eg, toluid-
Creatinine ine) techniques. There is a reasonable agreement in
Method: Most currently used assays are based on the the values among the most commonly used meth-
Jaffe reaction.68 This reaction occurs between creat- ods.68 Simple colorimetric tests in the form of a dip
inine and the picrate ion formed in an alkaline me- stick have been used with some success in birds.
dium. Lipemia or hemolysis of the sample can interfere with
photometric methods of measurement, giving falsely
Physiology: Blood creatinine is derived mainly from
elevated values.11 This is less likely to occur with
the catabolism of creatine found in muscle tissue. kinetic assays that evaluate a change in optical den-
Phosphocreatine is used to store energy in muscle, sity over time and are therefore self-blanking.
and its catabolism to creatinine occurs at a steady
rate. Excretion of creatinine is solely via the kidneys. Physiology: Glucose is continuously required as an
It is freely filtered and reabsorbed in the tubules.25 In energy source by all the body cells and must be
birds, creatine is excreted in urine before it has been maintained at adequate levels in plasma. Glucose
converted to creatinine.6 The urinary excretion of levels are maintained principally through the conver-
creatine may be one reason that creatinine levels do sion of liver glycogen, with some being derived from
not provide an accurate assessment of avian renal non-carbohydrate sources (hepatic gluconeogenesis).
function. In periods of starvation, glucose is increasingly de-
rived from the breakdown of fats and proteins, pri-
Diagnostic Value: There is a slim margin between
marily from muscle tissue, through gluconeogenesis
the physiologic and pathologic levels of creatinine. in the liver and the kidneys. All plasma glucose is
For many analyzers, physiologic values are below the filtered from the blood through the renal glomeruli
detectable range. This test parameter is very insen- and then totally reabsorbed in the tubules.
sitive and is a relatively poor diagnostic test in birds.
Interestingly, 73 hours of starvation in pigeons in-
Physiologic Influence: Normally, creatinine produc-
duces hyperglycemia rather than starvation hypo-
tion is relatively constant and is minimally affected
glycemia.57 This finding has important consequences
by catabolism of dietary or tissue proteins. Theoreti-
for avian anesthesia and gastrointestinal surgery, as
cally, the pool of creatine from which creatinine is
presurgical fasting varying from four hours (empty-
liberated depends on the total muscle mass. How-
ing of the crop) to 24 hours (emptying of the entire
ever, in all avian species that have been investigated,
gastrointestinal tract) can be advantageous. Pro-
the reference interval for creatinine has been be-
longed fasting is not recommended in birds that
tween 0.1-0.4 mg/dl, with no significant differences
weigh less than 100 grams.
between species.
Diagnostic Value: Glucose is often a part of a labora-
Pathologic Changes: Severe kidney damage can
tory panel25,43,73 even though pathologic changes in
lead to increased creatinine levels, especially if the
birds are seldom detected.36 Glucose should be evalu-
filtration rate is decreased. Elevations have also been
ated in convulsing birds or those with glucosuria.
described in connection with egg-related peritonitis,
235
CHAPTER 11 BIOCHEMISTRIES
Physiologic Influence: Plasma glucose levels are Diagnostic Value: Changes in inorganic phospho-
higher in juvenile than adult budgerigars.32 Vari- rus concentration can occur with several diseases, but
ations also occur due to time of day and amount of not on a consistent basis. The diagnostic value is poor.
environmental stress.43 Plasma glucose concentra-
tions in fasted birds are subject to a circadian rhythm. Physiologic Influence: Diets that consist mostly of
A rise in plasma glucose concentration starts during seeds may lead to increased phosphorus levels. Juve-
the scotophase, reaching peak values early during the nile budgerigars were found to have higher concen-
photophase. Subsequently, a gradual increase can be trations than adults.32 No changes in inorganic phos-
observed with the lowest values at the end of the phorus levels were noted in laying hens.43
photophase. Afternoon plasma glucose concentra- Pathologic Changes: Increased plasma inorganic
tions in birds that are fed early during the photophase phosphate levels can be seen in some cases of severe
are significantly higher when compared to fasted kidney damage 2,36,73 due to vitamin D hypervitami-
birds.57 nosis,2 nutritional secondary hyperparathyroid-
Pathologic Changes: Increases in plasma glucose ism43,73 and hypoparathyroidism.31,33 False elevations
levels are due to increased glucose production or will occur if samples are hemolyzed. Occasionally,
release. For example, increases occur after meals, decreased plasma inorganic phosphate levels may
with excitement or stress or because of decreased occur from hypovitaminosis D (calcium level also
glucose usage (diabetes mellitus).2,25,50,73 Diabetes decreased), malabsorption because of phosphate
mellitus has been confirmed in budgerigars, cocka- binding agents in the diet (calcium normal) and long-
tiels, Amazon parrots, Scarlet Macaws, Umbrella term glucocorticoid therapy.
Cockatoos and a Toco Toucan.43 Transient elevations
Iron
in glucose have been reported in cockatiels with egg-
Sample: Heparinized plasma or serum can be used.
related peritonitis.63 Decreases in plasma glucose
Plasma specimens collected with EDTA, oxalate or
levels can be due to hepatic dysfunction (eg, Pacheco’s
citrate are unsatisfactory, because they bind iron.
disease virus), impaired glucose production or its
Markedly hemolyzed specimens are nondiagnostic
excessive utilization (eg, septicemia, neoplasia, as-
because free hemoglobin will increase the total serum
pergillosis).61,63 In young birds of prey, hypoglycemia
iron levels.
can cause convulsions.36 Starvation of up to four days’
duration will not cause hypoglycemia in all birds, but Method: For iron level assays, reduced Fe (II) is
in some (particularly raptor neonates), hypoglycemia complexed with a chromogen. This complex has a
can occur after a few days of anorexia.50 Glucose high light absorbance that is proportional to the iron
concentrations can be artificially decreased during concentration.68 Most assays require a large sample
storage if the blood sample is contaminated with size (200 µl).
bacteria.34
Physiology: Iron is an essential constituent of the
Phosphorus heme portion of hemoglobin. As the hemoglobin in
Sample: Heparinized plasma or serum is suitable. aged erythrocytes is broken down, iron is recycled and
Anticoagulants such as citrate, oxalate or EDTA fresh hemoglobin is synthesized. Iron is transported
should not be used because they interfere with the in the plasma attached to a β-1-globulin known as
formation of the phosphomolybdate complex. Hemo- transferrin.
lysis must be avoided, because the phosphate concen-
tration of erythrocytes is higher than that of plasma, Diagnostic Value: The value of determining iron in
and hemoglobin interferes with the colorimetric de- different avian species has not been thoroughly inves-
tection reaction used to determine phosphorus levels. tigated. A recent report shows a failure to correlate
serum iron levels with liver biopsy and subsequent
Method: Most assays for inorganic phosphate rely on toxicologic analysis for iron.75
the formation of a complex of phosphate ion with a
molybdate compound.20 Physiologic Influence: Prior to egg laying, iron lev-
els will increase two to three times normal in some
Physiology: Inorganic phosphorus is derived from species.25 Raptors maintained in captivity have sig-
the diet. It is a major constituent of bone and a vital nificantly lower values than their free-ranging coun-
cellular component, playing important roles in the terparts.36 Captive toucans have approximately three
storage, release and transfer of energy and in acid-
base metabolism.
236
SECTION TWO PATIENT EVALUATION
times higher reference values of iron than psittacine Total Protein (TP)
birds (see Chapter 47).74 Sample: When only small amounts of blood can be
collected, it may be advantageous to use plasma
Pathologic Changes: Severe and chronic loss of
instead of serum to determine the TP concentration.
blood will increase iron values. Iron deficiency ane- In pigeons, the concentration of TP in plasma is about
mia has been described in raptors.40 Changes in 1.5 g/l higher than in serum, because the former
plasma iron levels in mynah birds and toucans with contains fibrinogen.58a
iron storage disease are described in Chapter 47.
Method: Total protein levels may be determined us-
Total Iron-Binding Capacity (TIBC) ing a chemical method or a refractometer. The chemi-
Method: An excess amount of ferric ammonium cit- cal method of choice is the biuret method, which
rate is added to serum to saturate the transferrin measures the TP in fluids colorimetrically using the
iron-binding sites. The unbound Fe (III) is removed formation of a blue peptide (copper complex in alka-
and the iron content of the supernatant is assayed as line solution). This method is extremely accurate for
described for iron.68 the protein levels typically found in plasma or serum
(1 to 10 g/dl), but is not precise enough to determine
Physiology: Normally, only one-third of the iron-
the low concentrations of proteins that are found
binding sites of transferrin are occupied by Fe (III),
normally in other body fluids. Both wet and dry
creating a reserve of iron-binding sites. The total
chemistry methods use this technique, but the re-
iron-binding capacity (TIBC) is a measurement of the
sults vary with the instrument used.
maximum concentration of iron that serum proteins,
principally transferrin, can bind. A urine iron-bind- Most commercial laboratories use a human standard
ing capacity (UIBC) test is also available.75 when determining TP and albumin concentrations,
Diagnostic Value: Abnormalities in TIBC occur with and various studies have shown that there are sig-
some disorders of iron metabolism. Very little data nificant differences between TP concentrations when
from birds is available. This parameter appears to different standards are used (eg, human, bovine,
have little importance in diagnosing hemochroma- pigeon, chicken). Because it is impossible to have a
tosis, but insufficient research has been performed.74,75 species-specific standard for all species presented to
the avian practitioner and because there is a high
Pathologic Changes: TIBC may be increased with correlation between the results obtained with the
iron deficiency and decreased in chronic inflamma- various standards, it seems wise to establish refer-
tory disorders. ence values for the various species using the human
standard.53
Lipase
Method: There are various methods for determina- The refractometer is widely used by veterinarians to
tion of lipase activity, and the reference ranges de- measure change in the refractive index of a solution,
pend on the method used. which is caused mainly by the proteins in solution
and is proportional to the concentration of total solids
Physiology: Lipase measured in plasma or serum is or protein. Most refractometers are temperature-
produced in the pancreas. This enzyme functions in compensated and already calibrated in scales ex-
the digestion of fat in the diet. pressing TP concentration (g/dl) and specific gravity
of urine.
Diagnostic Value: Lipase and amylase activities
were high in a caique with clinical signs of pancreatic Information on the reliability of the refractometric
exocrine insufficiency when compared to the activi- method to determine TP concentrations in avian
ties of these enzymes in the mate (Ritchie BW, un- blood is conflicting. One study indicated that tem-
published). Hemolysis inhibits enzyme activity. perature compensated refractometers provide reli-
Pathologic Changes: Although no reference values able results when compared to non-temperature-
are currently available, birds do exhibit high lipase compensated devices.3 In another study, tempera-
activity in severe cases of acute pancreatitis. For ture-compensated and non-compensated refractome-
diagnostic purposes, a blood sample from a repre- ters yielded higher values than the biuret method,
sentative of the same species should be included for with the temperature-compensated instrument be-
comparison. ing consistently higher in readings than the non-
temperature-compensated refractometer. In juvenile
237
CHAPTER 11 BIOCHEMISTRIES
phoretic tracing. The length and height of each peak dietary protein, temperature stress, state of hydra-
ithin the pattern indicates the relative amount of a tion, hemorrhage and inflammation.66
particular protein or group of proteins. This can be
translated into percentage readings and, by combin- Pathologic Changes: Decreases in albumin concen-
ing this information with the TP concentration, abso- tration can occur from decreased synthesis due to
lute values for the concentration of each protein, or chronic liver disease or chronic inflammation, in-
protein group, can be calculated. creased albumin loss due to renal disease, parasitism
or over-hydration.72 A decrease in albumin causes
Physiology: Most frequently used electrophoresis edema because of a decrease in oncotic pressure.
methods identify five main protein fractions in birds: Increases are seen because of dehydration.
albumin, α1-, α2-, β- and γ-globulins.66 A pre-albumin
fraction has been described in pigeons and some Increases in α- and β-globulins may be caused by
parrot species.15,16,17,50,54 The α-globulins are acute acute nephritis, severe active hepatitis, systemic my-
phase proteins that typically increase with acute cotic diseases (γ-) and the nephrotic syndrome.72 In-
inflammation; β-globulins are composed of comple- creases in γ-globulins occur with acute or chronic
ment, hemopexin, ferritin, fibrinogen and lipoprote- inflammation, infection, chronic hepatitis and im-
ins.39 Some immunoglobulins, including IgM and mune mediated disorders.72
IgA, also migrate in the β-globulin range. The β-
Triglycerides
globulins are also acute phase proteins. The γ-globu-
Sample: Serum and lipemic specimens should be
lin fraction is mainly composed of immunoglobulins
warmed to 37°C and vigorously mixed prior to analy-
(IgA, IgM, IgE and IgG).66
sis.
Diagnostic Value: In healthy birds the albumin frac-
Method: Usually, triglycerides are enzymatically de-
tion is the largest protein fraction. An inflammatory
tected by breaking down the triglycerides and meas-
process will cause a rise in TP because of increased
uring the glycerol that is liberated.
concentrations of α, β or γ globulin fractions. Often
albumin concentrations are decreased in these situ- Physiology: Triglycerides are the major storage form
ations. The combined effect of these changes is a of lipids, and are a major energy source. Each mole-
decrease in the albumin/globulin (A/G) ratio. Often cule of triglyceride consists of three fatty acid mole-
the TP concentration is within the reference range, cules attached to a molecule of glycerol. They are
while the A/G ratio is decreased. Therefore, the A/G synthesized in the intestinal mucosa and liver from
ratio is of greater clinical importance than the TP the components of fat digestion and absorption.
concentration. Examples of diseases with a decrease
in the A/G ratio are egg-related peritonitis, and Diagnostic Value: Triglyceride values have been in-
chronic infectious diseases such as aspergillosis, psit- sufficiently evaluated in birds. Several factors can
tacosis and tuberculosis. influence the blood concentration and increases may
not be of clinical importance.
Serum or plasma protein electrophoresis can be used
to monitor response to treatment. When the bird Physiologic Influence: Triglyceride levels may vary
responds favorably, an increase in the albumin con- based on climate, hormone influence, diet and gen-
centration and a decrease in the globulin concentra- der. Increases may occur during starvation, particu-
tion can be observed, which leads to normalization of larly in obese birds. Estrogen injections have been
the A/G ratio. In birds with liver failure, extremely shown to elevate triglyceride concentrations in some
low plasma protein concentrations can occur in com- species.25
bination with a decreased A/G ratio. Gastrointestinal
Pathologic Changes: Egg-related peritonitis has
and renal diseases can also lead to severe hypoprote-
been associated with high concentrations of trigly-
inemia. These changes are caused by a loss of albu-
cerides.73 High concentrations (2000-5000 mg/dl) were
min. Elevated TP concentrations with a normal A/G
reported in Amazon parrots showing signs of hy-
ratio can be expected in dehydrated birds.
peradrenocorticism. Because triglyceride values are
Physiologic Influence: Physiologic factors that may determined based on enzymatically released glycerol,
change the protein concentration and therefore affect these values may be falsely elevated after exercise or
protein electrophoresis results include gender, age, following any event that causes increased levels of
blood glycerol (eg, catching birds in an aviary).
239
CHAPTER 11 BIOCHEMISTRIES
Diagnostic Value: Urea is present in very small Pathologic Changes: Hyperuricemia can be ex-
amounts in avian plasma, and determining urea lev- pected if the glomerular filtration is decreased more
els has generally been considered of little value. How- than 70 to 80%. Decreased filtration may occur from
ever, recent investigations have shown good correla- hypovitaminosis A-induced damage to renal epi-
tion between increased plasma urea concentrations thelial cells, dehydration, intoxications or from some
and renal disease in pigeons.50 In other avian species, bacterial and viral (Newcastle disease) infec-
urea may have little value in detecting renal disease tions.2,5,21,25,36,73 Uric acid levels may also be increased
but can be used as a sensitive indicator of dehydration. from the release of nucleic acids caused by severe
tissue damage or starvation. If a toenail clip is used
Physiologic Influence: Physiologic conditions are for blood collection and urates from the droppings
known to change urea concentrations in mammals, contaminate the sample, the uric acid levels may be
but similar effects have not been documented in falsely elevated.21,43
birds.
If the blood uric acid concentration exceeds its solu-
Pathologic Changes: High urea plasma levels can bility it will be deposited in different locations in the
occur in all conditions that cause low urine flow, such body. High plasma or serum concentrations of uric
as dehydration or bilateral ureteral obstruction.50 acid are a prognostic indicator that gout may occur.
Use of nephrotoxic drugs may also lead to hyper-
Uric Acid
uricemia. Hypervitaminosis D3-induced renal dam-
Method: Both wet and dry chemistry systems use
age is frequently associated with gout and extremely
oxidation of urates by uricase as a detection method.
high uric acid levels. This problem is particularly
Most uricase methods are extremely specific and only
common in macaws. This has been described for
a few structural analogues to uric acid will interfere
aminoglycosides (gentamicin),2,25,43 and allopurinol in
with the test. In general, the concentrations of these
Red-tailed Hawks.56 Interestingly, in most species,
analogues are low in biological fluids.68
allopurinol is effective in treating, not inducing gout.
Physiology: In birds, uric acid is the major product of
Hypouricemia is much less common in birds than
the catabolism of nitrogen. Synthesis occurs mainly
hyperuricemia. Severe hepatocellular disease with
in the liver50 and in the renal tubules.14 Approxi-
reduced synthesis of uric acid has been suggested as
mately 90% of blood uric acid is eliminated by secre-
one etiology.
tion into the lumen of the tubules. Only 50% of the
healthy avian kidney is actually used for excreting
protein waste, providing a large functional reserve.25 Electrolytes
Physiology: Chloride is the major extracellular an- Pathologic Changes: Hyperkalemia can be caused
ion. Sodium and chloride together represent the ma- by severe tissue damage, reduced potassium excre-
jority of the osmotically active constituents of tion by diseased kidneys,25,73 adrenal disease73 or be-
plasma. cause of redistribution of potassium from the intra-
cellular to the extracellular fluid (acidosis).25
Diagnostic Value: Elevations in chloride concentra- Dehydration25,73 and hemolytic anemia25 can also
tions rarely are detected. cause hyperkalemia.
Physiologic Influence: In budgerigars, no gender or
Hypokalemia may be caused by decreased potassium
other physiologic variables have been observed.32 intake, increased potassium loss due to chronic diar-
Pathologic Changes: Hyperchloridemia can occur rhea or diuretic therapy (seldom used in birds)73 and
with dehydration.25,36 The role of chloride in main- the shift of potassium from the extracellular to the
taining acid-base balance has not been sufficiently intracellular fluid (alkalosis).25
evaluated in birds.
Sodium
Potassium Sample: Either heparinized plasma or serum is ap-
Sample: Either heparinized plasma or serum is ap- propriate for sodium assays. With ion-selective elec-
propriate for detecting potassium. If ion-selective trodes, whole blood may be used. Electrolyte concen-
electrode methods are used, whole blood is also an trations are different between serum and plasma.
effective sample. Differences in the electrolyte con- Hyperlipemia and hyperproteinemia will cause
centrations in serum and plasma must be considered falsely low potassium levels by a mechanism similar
when interpreting results. Potassium levels are usu- to that described for potassium.
ally higher in serum due to the release of potassium
Method: Sodium may be determined by atomic ad-
from thrombocytes damaged in the coagulation proc-
sorption spectrophotometry, flame emission spectro-
ess. Hemolysis will elevate the plasma concentration
photometry or electrochemically with a sodium ion-
of potassium (500 to 700%).39 Potassium concentra-
selective electrode. The last two systems are most
tions were found to rapidly decline in pigeon and commonly used.30,68
chicken plasma allowed to sit for two hours.46 For
accurate results, plasma should be separated within Physiology: Sodium is present mainly in the ex-
minutes of collection. Hyperproteinemia and hyper- tracellular fluid and is primarily responsible for de-
lipemia will result in falsely low potassium levels termining the volume of the extracellular fluid and
caused by a decreased aqueous fraction of the total its osmotic pressure. Intracellular sodium levels are
plasma volume. kept low by a relatively impermeable cell membrane
and a sodium pump which removes sodium from the
Method: Potassium may be determined by atomic
cell. The amount of sodium in the body is regulated
adsorption spectrophotometry, flame emission spec- by the kidney. In addition, many avian species have
trophotometry or electrochemically with a sodium a specialized nasal gland (salt or supraorbital gland)
ion-selective electrode. The last two systems are most that is able to secrete large quantities of sodium in
commonly used.30,68 response to osmotic changes, thus allowing these
Physiology: Only two percent of the body’s potassium birds to drink salt water. When sea birds are kept in
is in the extracellular fluid. The other 98% is kept fresh water for a period of time the gland shrinks so
within the cells by “potassium pumps” in the cell that when returned to salt water the birds can no
membranes. longer tolerate high sodium levels.This mechanism of
decreasing sodium concentration in the serum and
Diagnostic Value: Alternatives in potassium ho- urine of birds is mediated by a pituitary-adrenal
meostasis have serious consequences. Decreased ex- response.66
tracellular potassium is characterized by muscle weak-
ness, paralysis and cardiac effects. Many potassium Diagnostic Value: Abnormal sodium levels that are
abnormalities are the result of hemolytic samples. not caused by technical failures are rarely seen in
birds. If they do occur, they are good indicators of a
Physiologic Influence: High amounts of potassium pathologic situation. Salt poisoning, mainly from
in the diet can elevate plasma concentrations. high salt foods, may occur more frequently in com-
panion birds than is documented.
241
CHAPTER 11 BIOCHEMISTRIES
Physiologic Influence: Sodium plasma levels are sis, which occurs at 37°C, the values are being influ-
maintained within narrow limits, despite wide fluc- enced by temperature.
tuations in dietary intake.
Method: An expensive blood gas instrument is neces-
Pathologic Changes: Hypernatremia can occur from sary.
increased sodium intake (peanuts, crackers), exces-
sive water loss or decreased water intake. Diagnostic Value: Clinical significance in compan-
ion birds has not been thoroughly investigated.
Hyponatremia may be due to increased sodium loss
as in kidney disease73 or severe diarrhea.25.73 It may Pathologic Changes: Acidemia (decrease in blood or
also be caused by over-hydration as in psychogenic plasma pH) has been reported in some birds with
polydipsia or after intravenous fluid therapy with renal disease.
sodium-free or low sodium solutions. The relative Reference Values for Budgerigars:32 pH (7.334 to
over-hydration, which follows a reduction in renal 7.489); pCO2 (30.6 to 43.2 mm Hg) (see Chapter 39).
perfusion possibly because of decreased colloid os-
motic pressure, may also cause hyponatremia.
Other Tests
Total Carbon Dioxide Content (Bicarbonate)
Method: Heparinized plasma or serum can be used.
Delta-Aminolevulinic Acid Dehydratase
Bicarbonate levels are determined by mixing the Method: Plasma or serum can be used to measure
sample with a strong acid and measuring the carbon delta-aminolevulinic acid dehydratase colorimetrically.
dioxide (CO2) release. Most of the carbon dioxide Diagnostic Value: Delta-aminolevulinic acid dehy-
produced is derived from bicarbonate, but a small dratase can be used to detect lead intoxication, and
amount is generated from dissolved carbonic and decreased plasma activity is pathologic.
carbamino acids.
Pathologic Changes: The activity can decrease de-
Physiology: Alterations of bicarbonate and CO2 dis- pending on the dosage of lead and the species up to
solved in plasma are characteristic of acid-base bal- 50% of the normal value.18,19,25 Central nervous sys-
ance. For clinical purposes, the total CO2 content is tem changes have been reported if plasma activity is
the same as the bicarbonate content.11 below 86 U/l (see Chapter 37).
Diagnostic Value: Bicarbonate levels are useful for
Acid Phosphatase
establishing whether or not acidosis or alkalosis is
This enzyme consists of a number of isoenzymes in a
present and, if so, how severe it is.
variety of organs. The activity is much lower than
Pathologic Changes: Increases are mainly due to that of alkaline phosphatase. Ovulation has been
metabolic alkalosis and decreases due to metabolic shown to increase acid phosphatase activities.25
acidosis. Reference intervals for most avian species
are not available. Copper
Method: Atomic adsorption spectrophotometry after
Reference Values for Adult Budgerigars: 21 to 26 direct dilution is the method of choice for determining
mmol/l.32 serum copper.68
Blood Gases - pCO2, pO2 and pH Physiology: Copper is a component of several major
Sample: Venous heparinized blood is the most likely enzymes and plays a vital role in hemapoiesis. It is
specimen that will be collected for blood gas analysis. involved in the absorption and the transfer of iron
Determination should be performed as quickly as and hemoglobin synthesis. In the plasma it is mainly
possible (in house).68 When measuring blood gases bound to ceruloplasmin.
and acid-base status in birds, it is necessary to collect
Diagnostic Value: Elevation occurs with copper in-
blood samples in pre-cooled syringes and store the
samples on ice to stop the metabolism of the erythro- toxication. In postmortem specimens, copper concen-
cytes. The nucleated avian erythrocytes possess vir- tration in the liver provides the best diagnostic sam-
tually all the enzymes typical of metabolically active ple.25
cells and consume oxygen seven to ten times faster Physiologic Influence: Copper levels are generally
than mammalian erythrocytes. Even during analy- higher in female mammals under the influence of
242
SECTION TWO PATIENT EVALUATION
estrogens. In birds, the effect of estrogens on copper Many factors can influence the color of avian urine.
levels has not been investigated. It can change with the ingestion of water-soluble
vitamins (especially Vitamin B), the amount of uric
Pathologic Changes: Copper intoxications will in- acid and feces mixed with the urine, the specific
crease the serum level. gravity and certain diseases (see Color 8). Macaws
often have very dark yellow urine, which is not normal.
Plasma Dye Clearance Test
In many animal species, the hepatic uptake and The white crystalline portion of the urine in birds is
excretion of different organic dyes injected intrave- seldom evaluated except for color. Birds that are in a
nously has been used for diagnosis of liver disease. negative nitrogen balance (severe cachexia, catabolic
Indocyanine green has been successfully used to de- disease) usually have an increased quantity of
tect liver disease in three raptor species.70 The dye urates.
was non-irritating if accidentally injected perivascu-
larly and clearance occurred. Pathologic Changes: Lead intoxication in some
species may result in chocolate milk-colored urine
In contrast, Bromsulphalein must be injected with and urates. This hemoglobinuria is common and nor-
care, because perivascular injection causes severe mal for some nervous birds. Severe liver disease, like
pain. In chickens, the clearance is markedly influ- that induced by chlamydia or Pacheco’s disease virus,
enced by age and gender.6,58 The clinical value of can increase the secretion of biliverdin, which results
these two tests has been insufficiently studied in in yellow-green or mustard-colored urine and urates.
birds. Because many other severe clinical diseases cause
this color to be present, it is not pathognomonic.
Urinalysis
Specific Gravity
Urinalysis is indicated if renal disease is suspected. Normal: The specific gravity varies with the state of
Polyuria is a common clinical presentation in com- hydration and with the individual bird. In the
panion birds.71 It may be caused by excitement, in polyuric bird, values from 1.005 to 1.020 are consid-
which the content of the cloaca is shed before the ered normal. A refractometer can be used for this
water is reabsorbed; by the intake of large amounts determination. Water deprivation should be used to
of fluids (fruits, vegetables); by renal disease, neo- evaluate the kidney’s ability to concentrate low lev-
plasia, diabetes, sepsis, toxins, adrenal disorders or els, often due to psychogenic polydipsia.
gout; after administration of some medications; and
Pathologic Changes: Increased loss of water without
with impending egg laying. In all of these cases, it is
an increased loss of solute will create a low specific
relatively easy to separate the urine from the feces
gravity. This situation can be caused by intravenous
via aspirating the liquid deposited on a water-resis-
fluid therapy, hyperthyroidism, liver disease, pitui-
tant surface. Transient polyuria can be induced by
tary neoplasia, progesterone or glucocorticoid ther-
administering water by crop tube. This will usually
apy. Any disease that causes polyuria and polydipsia
result in urine production within 30 minutes after
can cause a low specific gravity. A reduced ability to
administration.73 In pigeons, urine for analysis has
concentrate or dilute the glomerular filtrate will lead
been collected directly from the cloaca using a can-
to an increased specific gravity and severe renal
nula.26 Urine samples can be collected from individ-
pathology.
ual ureters of anesthetized parrots using a speculum.
Specific Evaluation
Volume, Color and Consistency73,25
Substances filtered by the normal kidney generally
Urine evaluation should include a measurement of
have a molecular weight of less than 68,000 (eg,
volume, a record of appearance (color, consistency)
water, uric acid, urea, glucose, electrolytes). Two sub-
and determination of specific gravity. Normal com-
stances that are on the border of this molecular
panion birds produce a small quantity of urine, and
weight cutoff are hemoglobin and albumin. Most
if it can easily be collected it is generally abnormal
other physiologic proteins have higher molecular
(stress or disease). The urine is usually clear in most
weights. Most substances that are filtered by the
companion bird species, but in other birds, such as
kidneys are critical to normal bodily functions and
ratites and Anseriformes, it is normally opaque,
are completely reabsorbed (eg, amino acids, glucose,
cloudy or slightly flocculent.
vitamins). The excretion or retention of other sub-
stances are regulated according to the body’s needs.
243
CHAPTER 11 BIOCHEMISTRIES
Urinary pH and the concentration of some chemical in most birds when the blood glucose level exceeds
constituents in the urine can be measured using 600 mg/dl. In diabetes mellitus, birds may have blood
commercial test strips designed for use with human glucose concentrations above 800 mg/dl.
urine. It should be noted that the sensitivity of these
tests has been adjusted to detect what would be Ketones
regarded as abnormal levels of certain substances in Ketones should be absent from the urine of birds. Any
human urine. These sensitivities are not necessarily significant shift in energy production from carbohy-
applicable to birds and the fact that a “higher” read- drates to fats results in the increased oxidation of
ing is obtained on an area of the test strip does not fatty acids and the production of intermediate meta-
necessarily imply an abnormality. For example, alka- bolites that accumulate faster than they can be oxi-
line urine can produce falsely elevated protein levels. dized by the tissues. Catabolic processes such as
The color of the urine sample may also affect the severe hepatitis in combination with low blood glu-
results of some test parameters. cose concentrations and diabetes mellitus can cause
ketonuria.
Normal pH: Most pet birds have a pH between 6.0
to 8.0, which is largely related to the diet. Birds fed Bilirubin
large amounts of protein (carnivores) have an acidic Bilirubin is not normally present in birds. Biliverdin
urine, while grain-eating birds have more alkaline is the major bile pigment, but will not react with the
urine. bilirubin portion of a mammalian urine dip stick.
Pathologic Changes: Companion birds with urine Urinary Urobilinogen
pH lower than 5.0 are considered acidotic.73 Increased Normal readings are 0.0 to 0.1 in healthy birds.
protein catabolism will cause a lower pH. Bacterial Pathologic changes would be expected in cases of
metabolism tends to cause an alkaline pH. Compan- intravascular hemolysis and severe liver disease, but
ion birds with papillomatosis and other disorders are seldom reported. Falsely high levels of urobilino-
that typically cause tenesmus may have acidic urine. gen in urine can be due to drugs which appear red in
Presumably this is caused by excretion of fluids from acid urine (eg, Vitamin B12) or if sulphonamides are
the upper intestinal tract.8 It has been suggested that present.
the cloacal mucosa of a normal companion bird is
neutral to slightly alkaline when measured with lit- Blood
mus paper (Harrison GJ, unpublished). Commercial strip tests are available that can distin-
guish hematuria (ie, an abnormally large number of
Urinary Protein
intact RBCs in the urine) and hemoglobinuria (ie,
Normal: Trace amounts of protein can be detected in
hemoglobin that is free within the urine and not
the urine of 90% of birds tested.73
contained within cells). With hematuria, individual
Pathologic Changes: Many renal disorders will re- erythrocytes lyse on the test area, giving individual
sult in a mild to moderate proteinuria. Non-renal spots of color. If there is free pigment, the color
sources of proteinuria include hematuria, hemoglo- change is uniform throughout.
binuria and hyperproteinemia, which are usually
Normal readings are negative or trace. Blood in the
caused by an increase in the production of immuno-
urine may originate from the cloaca or from the
globulins. Inaccurate protein levels will be detected
urinary, reproductive or gastrointestinal tracts.
if the urine is alkaline or if the strip is soaked in urine
Hemoglobinuria can be due to intravascular lysis of
(instead of briefly dipped), which leaches out the
RBCs (rare) or lysis of RBCs present in the urine.
citrate buffer.
Urinary Nitrite
Glucose
Normal: Avian urine normally contains no glucose. This test is included on many commercial test strips
In healthy pigeons, reference values between 0 and and is used to screen for bacteriuria. It is an unreli-
3.2 mmol/l were established by the hexokinase reac- able test for avian urine.
tion.50 Trace glucose readings may be detected in
Urinary Sediment
normal avian urine by using dip sticks.73
Examination of the urinary sediment is a valuable
Pathologic Changes: The threshold for glucosuria to part of urinalysis but one that is often omitted. A
occur varies with the species.73 Glucosuria will occur fresh or refrigerated sample is required. With time,
244
SECTION TWO PATIENT EVALUATION
there is increasing alkalinity causing progressive cases of renal disease. Granular casts are most com-
lysis of blood cells and casts. Usually centrifugation mon. Cellular casts (which incorporate cells like
is used to concentrate the sediment to approximately RBCs, WBCs or tubular epithelial cells) and hyaline
ten percent of its original urinary volume. casts (consisting of mucoprotein gel) may also be
seen.
White and Red Blood Cells
The number of RBCs and WBCs in the sediment is Bacteria
reported as the number per high power field (HPF). In mammals, it is believed that bacteria in excess of
Normal urine contains 0 to 3 RBCs/HPF and 0 to 3 3 x 104/ml of urine must be present before they are
WBCs/HPF.73 More than 6 white or red blood cells per detectable in urinary sediment.68 Gram-positive cocci
HPF is a cause for concern. All cells noted within the and rods may be noted in the avian urinary sediment
urine sediment may have origins within the cloaca or if the sample has been contaminated with fecal ma-
the urinary, reproductive or gastrointestinal tracts. terial.
39. Kaneko JJ: Clinical Biochemistry of 49. Lumeij JT, Westerhof I: Blood chemis- 57. Lumeij JT: The influence of blood Textbook of clinical chemistry, Phila-
Domestic Animals. San Diego, Aca- try for the diagnosis of hepatobiliary sample treatment, feeding and star- delphia, WB Saunders Co, pp 356-
demic Press, 1989. disease in birds. Vet Quarterly 9:255- vation on plasma glucose concentra- 386, 1986.
40. Kösters J, Meisters B: Hematokrit 261, 1987. tions in racing pigeons. In Lumeij JT: 66. Sturkie PD: Avian Physiology 2nd
und Hemoglobinwerte von verschie- 49a. Lumeij JT, Westerhof I: Clinical evalu- A Contribution to Clinical Investi- ed. Auflage, Ithaca, New York. S 624-
denen Falken und Eulen. (PCV and ation of thyroid function in racing pi- gate Methods for Birds with Special 625, 1982.
hemoglobin of different birds of prey). geons (Columba livia domestica). Reference to the Racing Pigeon. PhD 67. Tell LA, Citino SB: Hematologic and
Prakt Tierarzt 63(5):444-448, 1982. Avian Pathol 17:63-70, 1988. thesis, Utrecht University, 1987, pp serum chemistry reference intervals
41. Kraft W, Dürr UM: Kompendium der 50. Lumeij TJ: A Contribution to Clinical 26-30. for Cuban Amazon Parrots (Ama-
klinischen Laboratoriumsdiagnostik Investigative Methods for Birds, with 58. JO, Smith AH: BSP clearance and zona leucocephala leucocephala). J
bei Hund, Katze, Pferd. (Clinical labo- Special Reference to the Racing Pi- fatty infiltration of the liver in domes- Zoo & Wildlife Med 23(1):65-71,
ratory diagnosis in dogs, cats and geon (Columba livia domestica). tic fowl. Physiologist 10:264, 1967. 1992.
horses). Stuttgart, M & H Schaper, Utrecht, Proefschrift, 1987. 58a. Mclean B, Lumeij JT: Evaluation of 68. Tietz NW: Textbook of Clinical
1981. 51. Lumeij JT: Avian clinical pathology: different refractometric methods for Chemistry. Philadelphia, WB Saun-
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