Overview of Psittacine Blood

Analysis and Comparative

R e t ro s p e c t i v e S t u d y o f C l i n i c a l
Diagnosis, Hematology and Blood
Chemistry in Selected Psittacine
S p ec ie s
a, b
Raffaella Capitelli, Dr Med Vet *, Lorenzo Crosta, Dr Med Vet, PhD

KEYWORDS

Psittacines
Hematology
Biochemistry
Psittacines diseases

Abnormal and normal blood values

KEY POINTS

Avian blood cells are fragile and samples should be processed as soon as possible after
collection.

It is important how the samples are taken, transported, and processed; therefore it is
essential that suitable structures are available before taking samples.

Mistakes when collecting, processing, and shipping the sample can cause many artifacts
and can make a normal panel abnormal, or mask out the abnormalities caused by various
pathologies.

Avian hematology and clinical chemistry are important in the diagnosis of many diseases
in psittacine species, but a proper interpretation of the results of a sample from a particular
species can only be achieved if its reference values are available and were obtained using
the same methods.

PART I
Introduction
Avian patients are not showing clear symptoms, therefore hematology and biochem-
istry are important aids in the diagnostic process of birds, including the species
belonging to the order Psittaciformes. Our need to widen diagnostic investigations
is made more difficult by the many differences among the various psittacine species
and the physiologic differences with the mammals.

a
CSV-Labvet, via Kennedy 10, 23873 Missaglia, Lecco, Italy; b Clinica Veterinaria Valcurone, via
Kennedy 10, 23873 Missaglia, Lecco, Italy
* Corresponding author.
E-mail address: capitelli@[Link]

Vet Clin Exot Anim 16 (2013) 71–120

[Link] [Link]
1094-9194/13/$ – see front matter Ó 2013 Elsevier Inc. All rights reserved.
72 Capitelli & Crosta

Avian laboratory medicine is relatively new to veterinary medicine. Before 1980,

avian hematology was primarily used as a research tool in the poultry industry.1 The
first description of avian hematology was by Lucas and Jamroz in 1961.2 Their atlas
is the basis of avian hematology and it describes the various hemic cells in poultry.2
In 1980, the Association of Avian Veterinarians reported a gradual increase in the
numbers of pet birds (primarily psittacines) being presented to their clinical practices.1

Blood Analysis
Avian blood cells are fragile and samples should be processed as soon as possible
after collection. It is important how the samples are taken, transported, and pro-
cessed, therefore it is essential that suitable structures are available before taking
samples.3 Mistakes when collecting, processing, and shipping the sample can cause
many artifacts and can make a normal panel abnormal or mask out abnormalities
caused by various pathologies.4–6
It is important to know whether the referring laboratory, either an in-house or
external laboratory

is equipped to process avian blood samples, using specific and validated tech-
niques for the different avian species, working with smaller blood collection size;

has knowledge of the normal reference values for many species, including pet
birds, and provides a chemistry panel suitable for each avian species/group;

is prepared to give information about the correct methods for blood collection,
sampling, and shipping and good professional advice for the interpretation of
the results.6

Blood Sampling
Animals differ in sex, age, environment, and possible stress factors, and sampling
conditions are also different making it difficult to interpret the data.4 Blood sampling
should be done with minimal stress. The practitioner should have the skill to obtain
a blood sample without anesthesia.4,5 A blood volume representing 1% or less of
the body weight can usually be withdrawn from healthy birds without risk. The sample
size taken from severely ill birds, however, must be reduced. For routine hematologic
evaluations in birds, a sample size of 0.2 mL is usually adequate. A variety of collection
methods has been used to obtain blood from birds, and the choice of method
depends on the size of the bird, peculiarities of the species, preference of the
collector, volume of blood needed, and the physical condition of the patient.1,3–5,7
Some examples are show in Table 1.
Routinely it would be safe to collect a blood volume that ranges between 0.5% and
0.7% of the patient’s body weight. The method of restraint to a safely collection of

Table 1
Maximum blood sample volume for selected Psittaciformes

Maximum Volume of Blood Sample

Species Body Weight (g) (1% Body Weight) (mL)
Amazon 250–600 4.5–2.5
Cockatoo 250–800 5.0
Cockatiel 40–80 0.4
African gray parrot 450 4
Macaw 200–1800 2.5–5
 Overview of Psittacine Blood Analysis 73

blood sample is dependent on the size of the bird, its capacity to inflict an injury, its
familiarity with humans, and the skills of the practitioner. Most birds require physical
restraint and it can be useful to use a towel or cloth to wrap them in order to prevent
self-inflicted injuries such as wing fractures. The three available sites for blood collec-
tion from psittacines birds, in order of preference, are the right jugular vein, the cuta-
neous ulnar (brachial) or wing vein, and the medial metatarsal vein. For tame birds, it is
possible to collect a blood sample from the jugular vein with a minimal amount of
restraint.
The right jugular vein is the most commonly used for blood collection in psittacine
birds. The right jugular vein is large and easy to locate and access in the featherless
area of the neck (apterium), providing a quick and easy method for collection of
adequate amounts of blood with minimum restraint even in small psittacines. The
disadvantage is that the jugular vein is mobile and must be well stabilized.7–10
An alternative site for blood collection in medium-large psittacine birds is the cuta-
neous ulnar (brachial) vein. Most parrots and cockatoos have short tarsometatarsi,
which makes access to the medial metatarsal vein difficult (Table 2).7
Because psittacine birds are small to medium in size and the total blood collected
cannot exceed a milliliter, not even a drop of the blood collected can be wasted. To
optimize the results, and using Psittaciformes as an example, the blood collected
should be used as follows:

2 drops of whole blood to make 2 blood smears at the time of collection using
nonanticoagulated blood and possibly using precleaned, bevel-edged micro-
scope slides

2 drops of whole blood to soak absorbent paper diskettes for Chlamydia
Immunocomb

2 to 3 drops of whole blood into a tube with a preserving solution suitable for
polymerase chain reaction analysis (PCR) (depending on the laboratory tech-
niques available)

2 to 3 drops of whole blood to fill a microhematocrit capillary tube for measuring
packed cell volume (PCV) and total plasma protein

The surplus into a lithium heparin container or other tests as needed
The best device to collect blood from a bird is a syringe with a 22- to 25-gauge
hypodermic needle. Thinner needles may cause hemolysis and it is advisable to avoid
a heparinized syringe.1,3–5,7,8,10,15 In smaller psittacine species, insulin syringes are
frequently used, better with detachable needles. Ejecting blood through a 25-gauge
or smaller needle may cause moderate to marked hemolysis, and this can invalidate
most biochemical analyses.16,17
Samples are collected in microhematocrit capillary tubes, EDTA tubes, lithium
heparin tubes, and serum separator tubes. EDTA tubes are not used for hematology

Table 2
Reported sites for the collection of blood from selected species of psittacines

African gray parrot Brachial Hawkey et al,11 1982

Amazon parrot Cutaneous ulnar, jugular Tell & Citino,12 1992
Budgerigar Right jugular Scope et al,13 2005
Red lory Right jugular Scope et al,14 2000

The terminology to describe the vein is as used in the original publication.

Data from Clark P, Boardman W, Raidal SR. Atlas of clinical avian hematology. Oxford (United
Kingdom): Wiley-Blackwell; 2009.
74 Capitelli & Crosta

Fig. 1. Normal avian erythrocytes and some nondifferentiable leukocytes due to adverse
effects on the morphology and staining by lithium heparin in a blood film of a healthy
Psittacus erithacus (Hemacolor, 400).

because they can cause adverse effects in some avian species, although many
authors advise them for hematology in psittacines because they allow for proper stain-
ing of cells and leukocytes tend not to clump.1,4,5,8,15,18,19 Heparin is useful for chem-
istry and is widely used in avian hematology.4,5,9,20 Many authors state that heparin
has adverse effects on the morphology and staining of avian hemic cells (Fig. 1)
and that, after 12 hours, it can cause leukocytes and thrombocytes to clump so inter-
fering with manual and automated cell counts.1,8,15,18 Blood samples collected
without anticoagulant require immediate processing because whole blood ships
poorly and cellular degeneration takes places quickly.1 Unseparated, clotted, or
hemolyzed samples frequently cause unreliable values. Lumeij21 noted that serum
potassium levels decrease in samples that are left unseparated for more than 2 hours.
Fudge says that samples shipped unseparated for 12 hours or longer results in
increased values of lactate dehydrogenase (LDH), potassium, and total proteins,
decreased values of glucose, and variable changes to calcium and phosphorus
levels.4 Lumeji22 suggests that lithium heparin is the best method for biochemistry
although in pigeons, serum bile acid levels are higher. Lipemia is a normal finding in
ovulating birds and can severely alter most photometric analyses. Lipemia is also
present in certain pathologies, such as some liver diseases. Fasting is not usually
necessary before blood sampling in a bird. In excitable or roughly handled pet birds,
or for intramuscular injections, creatine phosphokinase (CPK) levels can increase.
Unfortunately, bacterial contamination of avian samples received by laboratories can
easily occur, causing cloudiness in serum or plasma, which can affect photometric
assays of various analytes such as marked decrease in CPK and glucose levels.4,5
A blood film should be made immediately after collection. An air-dried thin smear
provides superior morphology compared with samples stored in EDTA or heparin.
The use of precleaned, bevel-edged microscope slides minimizes cell damage. The
wedge smear or slide and long coverglass techniques normally used by general prac-
titioners can produce damaged cells (smudged cells). The presence of numerous
smudged cells, which are probably granulocytes, causes inaccurate hematologic
results.4,5 Blood films can be prepared by using a slide and coverslip or by using 2
coverslips.4,15,23 Conventional Romanosvsky stain (Wright and Giemsa stains) usually
gives good results for avian blood smears. Quick stains can also produce good quality
blood films.1,9,20
 Overview of Psittacine Blood Analysis 75

Hematology
The blood of avian psittacines contains nucleated erythrocytes, nucleated thrombo-
cytes, heterophils, eosinophils, basophils, lymphocytes, and monocytes. The main
difference between avian blood and mammalian blood is that the avian blood contains
nucleated erythrocytes and nucleated thrombocytes, which can interfere with auto-
mated blood counting procedures.9 Moreover, avian white blood cell morphology
differs from that of mammals because the predominant avian granulocyte is the
heterophil versus the neutrophil in mammalian species. The other leukocytes are
morphologically similar to their mammalian counterparts. There also are some physi-
ologic factors that can influence the hematologic results such as age, gender, anes-
thesia, nutrition, environmental conditions, physiologic status, and genetic factors
(species).

Measurement techniques for avian hematology

The laboratory evaluation of psittacine blood involves the same routine procedures
used for mammalian hematology with some modifications.1

PCV or hematocrit PCV or hematocrit expresses the proportion of the volume of the
corpuscular part in whole blood.7 The standard method for determination of the
hematocrit (PCV) uses a microhematocrit tube that is centrifuged at 12,000g for
5 minute or at 3000g for 30 minutes.1 Normal values for PCV range from 35% to
55% in most psittacine species. A decreased PCV suggests anemia, whereas an
increased PCV indicates dehydration if total proteins are increased or erythrocytosis
if total proteins are normal or low.

Hemoglobin Hemoglobin concentration is determined using spectrophotometric

analysis of cyanmethemoglobin at 540 nm, after centrifugation to remove the free
nuclei from lysed erythrocytes to avoid overestimation. Recent reports have used
a colorimetric method assessed at both 570 and 880 nm to compensate for turbidity
of the sample.7,24,25
The normal hemoglobin concentration is 11 to 18 g/dL in most psittacine species.
The hemoglobin of mature birds contains myoinositol pentophosphate and not
2,3-diphosglycerate, as in mammals. In avian erythrocytes, the phosphate
compounds differ from those of mammals and therefore avian tissues can extract
oxygen more readily from hemoglobin than mammalian tissues.1

Hemic cell counts

A major limiting factor in avian hematology is the lack of a validated automated method
for the measuring hemic cell counts, and in particular for analyzing leukocytes. Thus,
manual counting methods have been well described and are the most common tech-
niques used in all birds. These methods are often time consuming, not accurate, and
suitable only for laboratories that handle many avian samples. Impedentiometric elec-
tronic cell counters are the gold standard in mammalian hematology but they cannot
be used in birds.20 Since 1990, studies performed in avian hematology (particularly on
ostriches and chickens), using laser and impedance technologies (Cell-Dyn 3500),
have given encouraging and reproducible results for erythrocyte and leukocyte
counts.26–30 The Cell-Dyn 3500 hematology system (Abbot laboratories, Abbot
Park, IL, USA) has veterinary software with a special application for avian species.
The most common techniques used in birds for erythrocyte and leukocyte counts
are manual by hemacytometer. The total red blood count (TRBC) is done using
Natt-Herrick solution or the erythrocyte Unopette system. For total white blood cell
counts (TWBC), direct methods with Natt-Herrick solution and indirect methods
76 Capitelli & Crosta

with the eosinophil Unopette system are used. In the direct method, blood is diluted
with Natt-Herrick solution. The advantage of this method is that using the same
charged hemocytometer, total erythrocyte and thrombocyte counts can also be ob-
tained.20 The disadvantage is that differentiating thrombocytes from small lympho-
cytes is often difficult, thus count errors are common. However, staining the blood
for 60 minutes in Natt-Herrick solution improves the ability to differentiate between
different cells.31
The Unopette eosinophil system uses a 1% phloxine B solution, a standard Neuba-
uer hemocytometer, and a rectangular coverglass. It is based on the principle that
phloxine B stains the eosinophils and heterophils red. With this system, only granulo-
cytes (eosinophils and eosin-stained heterophils), which appear as round, refractile,
and red-orange cells, can be counted in both chambers of the hemocytometer. A
differential cell count on a stained blood film is performed by obtaining the percentage
of eosinophils and heterophils. The TWBC count is calculated using the following
formula1,8,30:


TWBC cells=mL

total eosin  stained cells counted in 18 squares  1760 ðor 1:1  16  100Þ
5
% heterophils 1 % eosinophils ðfrom differential countÞ

Alternatively, when it is not possible to obtain a sufficient volume of blood to perform

a quantitative count of TWBC or when the count cannot be performed within a few
hours, some investigators have suggested estimating the WBC count from a blood
smear. This technique does not require special equipment and is available to most
practitioners. The estimated WBC count can be obtained from a stained blood smear,
and requires only a drop of blood. This method is based on the ratio of RBCs to WBCs,
and the total estimated WBC count is determined by counting all leukocytes in 10
microscope fields (40) of a monolayer blood smear and multiplying the mean number
by 2000.4,5,9,20 A comparison of the leukocyte concentrations, obtained by a quantita-
tive hemacytometer method, have found significant differences between these 2
methods, with greater values recorded for the hemacytometer counts.32 On the other
hand both the direct and indirect method can be inaccurate due to many reasons:
manual dilution, the use of a hemacytometer, the difficulty in identifying the hemic cells
(leukocytes vs erythrocytes and thrombocytes), and the possibility of an incorrect
differential count, which can change the final result of the Unopette eosinophil count.9
A reliable thrombocyte count is difficult to determine because they tend to clump.
For this reason, their concentration is often indicated as normal, increased, or
decreased depending on estimates from peripheral blood films. In a blood film of
a healthy psittacine, it is normal to see 1 to 5 thrombocytes in a monolayer 1000
(oil immersion) field unless thrombocytes clump excessively.1,10

Erythrocyte morphology
Evaluation of avian erythrocyte morphology is based on observation of the cells in
monolayer 1000 fields.1,33 Avian erythrocytes should be evaluated for their size,
shape, color, nucleus, and cellular inclusions.1,34,35 Mature psittacine erythrocytes
are elliptical with an elliptical, centrally positioned nucleus, and vary in size depending
on the species.
The hemoglobin within the cytoplasm stains a light orange-pink color. During matu-
ration, the erythrocyte assumes its characteristic oval shape and hemoglobinization
occurs. Nuclear chromatin is evenly clumped and becomes increasingly condensed
 Overview of Psittacine Blood Analysis 77

Fig. 2. Presence of increased polychromasia, anisocytosis, immature erythrocytes at different

stages of erythrocyte maturation and 1 mitotic figure in the blood smear from an anemic
Ara ararauna with 12% of PCV as part of a regenerative response (Hemacolor, 1000).

with cellular aging; small numbers of polychromatophils (less than 5%) may be
present. They are characterized by a slightly larger and more rounded appearance.
The nucleus is less condensed and the cytoplasm is bluish.1,8–10
Avian erythrocytes have a short lifespan compared with mammalian erythrocytes.
Erythropoiesis is controlled by circulating levels of erythropoietin, which is produced
in the kidney in response to blood oxygen and cortical, estrogen, and androgen
hormone levels.9
The degree of erythrocyte polychromasia and reticulocytosis, the presence of binu-
cleate immature erythrocytes, and an increase of immature erythrocytes in the periph-
eral blood are evidence of active erythropoiesis (Figs. 2 and 3). However, these cells
indicate abnormal erythropoiesis in nonanemic birds and may suggest the stimulation
of the hematopoietic tissue following an anoxic insult or toxicity.1 The presence of
a large number of hypochromatic erythrocytes indicates an erythrocyte disorder,
such as iron deficiency or iron sequestration caused by infectious diseases.

Fig. 3. Immature erythrocytes and thrombocytes at different stages of erythrocyte and

thrombocyte maturation, in the blood smear from the same birds as in Fig. 2. The cytoplasm
of immature erythrocytes (large cells) contains spots of pink material due to the develop-
ment of hemoglobin (Hemacolor, 1000).
78 Capitelli & Crosta

Fig. 4. The blood film reveals abnormal erythrocyte shape with increased membrane
fragility (ballooning). The presence of atypical erythrocytes should not be confused with
artifacts of slide preparation (Hemacolor, 1000).

A slight anisocytosis is considered normal for psittacine birds, but marked anisocy-
tosis is usually observed in birds with regenerative anemia associated with polychro-
masia. Similarly, a slight poikilocytosis (variation of the shape) (Fig. 4) is normal in the
peripheral blood of birds, whereas marked poikilocytosis may indicate erythrocytic
dysgenesis.
Anuclear erythrocytes (erythroplastids) or cytoplasmic fragments are occasionally
observed in the peripheral blood of healthy birds. Mitotic activity associated with
erythrocytes in blood films suggests a marked regenerative response (Fig. 5) or eryth-
rocytic dyscrasia.10 Binucleate erythrocytes rarely occur in blood smears of normal
birds; however, the presence of large numbers of binucleated erythrocytes plus other
features of RBC dyscrasia is suggestive of neoplastic, viral, or genetic diseases.10,36

RBC indexes
The RBC indexes are calculated from the PCV, hemoglobin level, and TRBC count.
Using the measured values, additional characteristics of erythrocytes can be

Fig. 5. Immature erythrocytes and thrombocytes at different stages of erythrocyte and

thrombocyte maturation in the blood smear from the same birds as in Fig. 2. On the right
side of the image, an activated thrombocyte may be observed (Hemacolor, 1000).
 Overview of Psittacine Blood Analysis 79

Table 3
Morphologic classification of anemia based on the RBC indexes

Normal MCH and MCHC Less Than MCH and MCHC

Normal MCV Normocytic normochromic Normocytic hypochromic
>MCV Macrocytic normochromic Macrocytic hypochromic
<MCV Microcytic hypochromic Microcytic hypochromic

calculated. The mean corpuscular volume (MCV) and mean corpuscular hemoglobin
(MCH) values in birds are larger than in mammals, due to a wider red cell size in birds,
whereas the MCH concentration (MCHC) is smaller than in mammals due to the pres-
ence of an erythrocyte nucleus. The evaluation of the RBC indexes gives additional
information on erythrocytes that may help in the morphologic classification of anemia
(as summarized in Table 3).

Response and abnormalities of the erythrogram

Absolute anemia (decreased PCV and TRBC) in psittacine birds can be caused by
blood loss (hemorrhagic anemia), increased destruction of erythrocytes (hemolytic
anemia), and decreased or no cell production (depression/aplastic anemia).9 The
most common causes of hemorrhagic anemia are trauma and hemorrhagic lesions
of internal organs, such as ulcerated neoplasms and gastric ulcerations. Less
common causes of blood loss anemia in psittacines include heavy infestation with
blood-sucking ectoparasites or gastrointestinal endoparasites, coagulopathies asso-
ciated with toxins, septicemic and viral infections, neoplasms, coagulation factor
deficiencies, or severe liver diseases. With increasing chronicity, hemorrhagic
anemia can be normocytic normochromic, macrocytic hypochromic, and microcytic
hypochromic. Hemolytic anemia can result from parasitemia, bacterial septicemia,
heavy metal toxicosis, acute aflatoxicosis, and extensive burns. Although rare,
immune-mediated anemia may result in hemolysis with red cell agglutination present
in the blood film. Hemolytic anemia is mostly regenerative characterized by marked
polychromasia, macrocytosis, anisocytosis, reticulocytosis, and hypocroma-
sia.1,8–10,19,36–39 Depression anemia indicates decreased erythropoiesis and is there-
fore nonregenerative, and can commonly develop in birds with chronic inflammatory
diseases, mainly those caused by infectious agents (Fig. 6). Psittacines can develop
anemia due to decreased erythropoiesis much faster than mammals do, possibly
because of the relatively short half-life of avian erythrocytes.10,24,40 Disorders
frequently associated with depression anemia in psittacine birds include chlamydio-
sis, aspergillosis, egg yolk peritonitis, mycobacteriosis, chronic air sacculites,
chronic hepatic or renal disease, hypothyroidism, hyperestrogenism, protein and
iron deficiency, starvation, neoplasia, and other chronic inflammatory diseases.10,41
Depression anemia is usually normocytic normochromic and can be associated
with the presence of hypochromasia, anisocytosis, and poikilocytosis.9
Hypochromasia is associated with nutritional deficiencies, especially iron defi-
ciency anemia, and can be found in birds with chronic inflammatory diseases,
presumably related to iron sequestration as part of the bird’s defense against infec-
tious agents.1
Erythrocytosis (polycythemia) is characterized by an increase in circulating red cells
and is rarely reported in psittacines.1,42 Absolute polycythemia can be primary (poly-
cytemia vera) or secondary, caused by hypoxia resulting in increased circulating
RBCs, and often occurs in chronic respiratory, circulatory, and cardiac diseases.9,10
80 Capitelli & Crosta

Fig. 6. The blood film reveals poor erythrocytic regenerative response in an anemic Psittacus
erithacus affected by PBFD (Hemacolor, 400).

Leukocyte morphology
The leukocytes in avian peripheral blood smears include granulocytes and mononu-
clear cells. Granulocytes are classified as heterophils, eosinophils, and basophils,
and mononuclear cells as monocytes and lymphocytes.1
Heterophils are the predominant leukocyte in the peripheral blood of most psitta-
cines, whereas lymphocytes are the most abundant in some species. This could be
due to the variation associated with the different interpretation of the leukocyte differ-
ential count performed in several laboratories.10 Heterophils are round cells with
a bilobed or trilobed nucleus that stains poorly. Its cytoplasm is clear or colorless
with numerous prominent eosinophilic granules (orange-pink, brick-red, or brown).
The cytoplasmic granules are typically spindle, rod, or spiculated shaped, but may
appear oval to round in some species (Figs. 7 and 8). Heterophil granules degranulate
with many stains showing clear cytoplasmic vacuoles containing a distinct round
orange granule (central body). Heterophils are the main phagocyte and are involved
in bacterial killing by chemotaxis, opsonization, phagocytosis, and lysis. These

Fig. 7. A normal heterophil and erythrocytes in a healthy Cacatua alba. The heterophils,
larger in size than erythrocytes, can vary among the species. The differences are observed
predominantly on the affinity of dyeing granules and on their form (Hemacolor, 1000).
 Overview of Psittacine Blood Analysis 81

Fig. 8. A normal heterophil and erythrocytes in an asymptomatic Psittacus erithacus. The

erythrocyte on the left side of the image shows a Haemoproteus gametocyte within the
cytoplasm (Hemacolor, 1000).

granulocytes increase due to inflammatory conditions. Abnormal heterophils include

immature and toxic heterophils. Immature heterophils are large round cells and
have increased cytoplasmic basophilia, nonsegmented nuclei, and round cytoplasmic
eosinophilic (secondary) and basophilic (primary) granules. Immature heterophils
possibly seen in the blood are myelocytes and metamyelocytes.1,2,7,9,10
Eosinophils are the least common granulocyte in psittacines. Eosinophils are
medium-sized round cells with a clear or light-blue cytoplasm in contrast to the color-
less cytoplasm of normal mature heterophils. They have a bilobed nucleus character-
ized by clumped chromatin that stains more intensely than heterophils. Most
psittacine eosinophils are the same size as heterophils but have round, strongly eosin-
ophilic, cytoplasmic granules. The cytoplasmic granules of eosinophils lack the central
refractile body seen in heterophils and stain more intensely than heterophil gran-
ules.1,7–9 The term eosinophil is incorrect for many psittacines species, such as the
cockatoo, which has eosinophils that lack the classically eosinophilic cytoplasmic
granules. The granules of eosinophils may stain differently in the various species
and may be eosinophilic, aqua, light blue or gray, or dark blue, and they can be round
or rod-shaped.7 The real function of these cells is unknown.1,7,24
Basophils of psittacines are smaller than heterophils, round with a round to oval
nonlobed nucleus and contain many round deeply metachromic granules that can
frequently degranulate; they fill the cytoplasm and obscure the nucleus. Basophils
seem to be involve in the initial phase of acute inflammation.1,9
Lymphocytes are the most common leukocyte in some psittacine species,10 and
they are similar to mammalian lymphocytes. Avian lymphocytes can be divided into
3 groups1,2,8–10,24 according to their size (small, medium, and large lymphocytes).
Most lymphocytes in peripheral blood smears are small and medium sized and are
smaller than the average heterophil; large lymphocytes are rare and are larger than
heterophils. The lymphocyte is typically a round cell, sometimes irregular when its
margins are in contact with surrounding cells or when it shows cytoplasmic blebs or
pseudopodes on the periphery. Normal small lymphocytes have a homogeneous
weakly basophilic cytoplasm, without a perinuclear area (Golgi apparatus) and lack
vacuoles and granules. In reactive lymphocytes, the cytoplasm tends to be strongly
basophilic and can have a perinuclear area (Figs. 9 and 10). Medium and large
lymphocytes have a moderate amount of blue-pale cytoplasm and may contain
82 Capitelli & Crosta

Fig. 9. Mature erythrocytes and a small mature lymphocyte with irregular cytoplasmic
protrusions in pancytopenic Psittacus erithacus affected by chronic virus disease (PBFD)
(Hemacolor 1000).

a few distinct azurophilic granules or magenta bodies. The nucleus is centrally placed
and shows an increased nucleus to cytoplasm ratio (except in large lymphocytes).
Cytoplasmic appearance is important to differentiate small lymphocytes from throm-
bocytes or medium and large lymphocytes from monocytes. Abnormal psittacine
lymphocytes are classified as reactive or blast-transformed lymphocytes. The function
of lymphocytes is the same as in mammals. B lymphocytes (depending on the bursa of
Fabricius) are involved in humoral immunity and T lymphocytes (depending on the
thymus) are involved in cell-mediated immunity.1,2,8,24,43
In peripheral blood films of psittacine birds, the monocyte is typically the largest
leukocyte and has an irregular to round shape, a gray–pale blue vacuolated cytoplasm
with fine eosinophilic dustlike granules (a pinkish tinged cytoplasm), and an irregular
and eccentric nucleus (sometimes with a flat or smoothly indented side). Monocytes
are involved in cell-mediated immunity.1,2,8,9,24,43

Differential leukocyte count

The differential WBC count is obtained on a monolayer blood smear by microscopic
observation at 100 and oil immersion of the central third of the blood film, counting

Fig. 10. A toxic heterophil, a small mature lymphocyte, and a reactive lymphocyte among
mature erythrocytes in a young Ara ararauna with chlamydiosis (Hemacolor, 1000).
 Overview of Psittacine Blood Analysis 83

at least 100 leukocytes and differentiating and calculating the relative (percentage)
and absolute number. The avian differential WBC count is technically more difficult
to interpret than the mammals one, because of some problematic cell pairs such as
heterophils versus eosinophils, monocytes versus large lymphocytes, small lympho-
cytes versus thrombocytes, reactive lymphocytes versus rubricytes, toxic heterophils
versus immature heterophils and basophils.
The distinction between eosinophils and heterophils is the first difficulty to be over-
come by the hematologist. The heterophils are the major avian granulocytes, whereas
eosinophils are uncommon in many psittacine species. Eosinophils do not degranulate
as quickly as heterophils and their granules appear brighter and rounder than hetero-
phils; their cytoplasm is blue and the nucleus is deeper blue, whereas, in heterophils,
the cytoplasm is clear or colorless and the nucleus is less condensed.1,8,9,18,19,24,30,43
The distinction between large lymphocytes and monocytes is based on fact that mono-
cytes have a homogeneous blue-gray cytoplasm, usually vacuolated, and have an
irregular shape and eccentric nucleus, whereas large lymphocytes have a pale blue
unvacuolated cytoplasm and a centrally located nucleus.1,8,9,18,19,24,30,43 Small
lymphocytes and reactive lymphocytes may be confused with thrombocytes and rubri-
cytes. The lymphocytes are typically round and with homogeneous pale blue cyto-
plasm and only rarely have magenta granules scattered throughout the cytoplasm,
whereas thrombocytes are oval with a colorless, vacuolated cytoplasm and often
have eosinophilic bipolar bodies (Fig. 11). The rubricytes are numerous when a marked
regenerative response is present; the cytoplasm is basophilic, gray or eosinophilic-
gray depending on the maturation stage and is darker than reactive lymphocytes
(Fig. 12). Toxic mature heterophils have a recognizable nuclear lobulation and some-
times a swollen appearance with nuclear karyolysis in severe cases, whereas immature
heterophils lack any lobulation. A high basophil count can indicate an inaccurate result,
probably caused by mistakenly counting toxic heterophils as basophils. The differential
count achieved on low-quality smears gives incorrect results.1,8,9,18,19,24,30,43 The
smudge cells are probably granulocytes and an increase in the relative count of
lymphocytes could also depend on smudging artifacts.4,5,30
Response and abnormalities of the leukogram
In the interpretation of a leukogram, the variations compared with normal reference
values can have physiologic causes (sex, environment, diet, and stress), pathologic

Fig. 11. Mature and polychromatic erythrocytes and a vacuolated thrombocyte in an Ara
ararauna. The blood film also reveals some ruptured cells that can cause inaccurate hema-
tologic results (Hemacolor, 1000).
84 Capitelli & Crosta

Fig. 12. Rubriblasts, prorubrocytes, basophilic rubricytes, polychromatophilic rubricytes, and

polychromatic erythrocytes in a peripheral blood smear of an anemic Ara ararauna (Hema-
color, 1000).

causes (leukocytosis, leucopenia, and myeloproliferative disorders) and causes unre-

lated to the patient such as differential and total WBC count performed by various
laboratories, inadequate techniques leading to inaccurate results, and artifacts due
to improper blood collection, sampling, and processing.
As already mentioned, avian leukograms can vary widely between normal psitta-
cines of the same species. Many young normal psittacines have mature hetero-
philia.9,18,19,39 Normal TWBC count reference intervals reported in psittacine birds
are generally broad, therefore only significant variations from normal ranges have
a diagnostic meaning. Most psittacine birds get frightened or excited when handled
for blood sampling, causing a physiologic leukocytosis resulting in an increase of
the concentration of heterophils and lymphocytes in the peripheral blood .This effect
(mature heterophilia) decreases within 24 hours if the stressor is removed.1,9,10
Leukocytosis is more frequently due to increased heterophils or lymphocytes rather
than monocytes, eosinophils, or basophils.9,18,19,39 The principal causes of leukocy-
tosis in psittacines include inflammation with infectious or noninfectious causes, expo-
sure to toxins, hemorrhages in the body cavity, rapidly growing neoplasms, and
leukemia. A differential leukocyte count helps to identify which component is most
responsible. Heterophils are responsible for the first defense against microbial infec-
tion and inflammatory reactions and heterophilia is generally seen when leukocytosis
is caused by inflammation.1,9,18,19,39 Heterophilia is found as a response to systemic or
local inflammation caused by infectious agents and noninfectious causes such as
trauma or toxicity. The severity of heterophilia is dependent on the degree of inflam-
mation and the cause of disease.10,44,45 Slight to moderate leukocytosis and lympho-
penia can occur with excess endogenous or exogenous glucocorticoids (stress
leukogram). Early or mild bacterial infection, metabolic disease, neoplasm, trauma,
and surgery can cause mature heterophilia. Marked leukocytosis and heterophilia
are often seen in psittacines with infections caused by common pathogens such as
Chlamydia, Mycobacterium, and Aspergillus.1
Heterophil morphology is more important than the absolute number to evaluate the
severity of disease and to make a prognosis. A left-shift leukocytosis is mild “regen-
erative” if only band heterophils (Fig. 13) are increased and is moderate to severe if
metamyelocytes and myeloblasts are also increased, whereas if the band heterophils
and immature cells are larger than mature heterophils, the left shift is referred to as
 Overview of Psittacine Blood Analysis 85

Fig. 13. Marked heterophilia in the blood film of an Amazona aestiva affected by acute
chlamydiosis (Hemacolor, 1000).

“degenerative”, indicating a severe inflammatory response. Regenerative left shifts

are correlated with severe or chronic bacterial, fungal, and chlamydial infection, and
less with toxins and neoplasms, but they can occur in severe blood loss anemia asso-
ciated with erythroid regeneration.9
Degenerative left shifts occur in septicemia, toxins, and some viral diseases.
Marked increases in immature heterophils may also result from granulocytic leukemia,
which is a rare condition in birds1,10 Toxic heterophils occur with severe systemic
illness such as septicemia, viremia, chlamydiosis, mycotic infections, and severe
tissue necrosis. The degree of heterophil toxicity characterizes the severity of disease
and a marked grade (41) indicates a poor prognosis.1,9,10
Eosinophilia is rare in birds. The true functions of avian eosinophils are unknown and
so eosinophilia is difficult to interpret in psittacine birds.1,10 Basophilia is also rare and
has been related to tissue necrosis occurring in chlamydiosis, respiratory diseases,
autoimmune hemolytic anemia, and traumas; it could also suggest early inflammation
or an immediate hypersensitivity reaction, because avian basophils produce, store,
and release histamine.1,9,18,19,39,46
Lymphocytosis is not common when compared with heterophilia and is related to
antigenic stimulation. A few reactive lymphocytes in the peripheral blood smear of
healthy psittacines may be normal, but the presence of many reactive lymphocytes
is suggestive of lymphocytosis with chronic antigenic stimulation associated with
infectious diseases, or during convalescence from infections. Severe lymphocytosis
can also occur with lymphocytic leukemia. In some cases of lymphocytic leukemia,
immature atypical lymphocytes may be present in the blood film.1,9,10,19,36,39 Anemia,
mitotic figures, and marked lymphocytosis, when most small mature lymphocytes
have scalloped cytoplasm, may also be present.1,8,10,19
Monocytosis is often associated with infectious diseases related to granulomatous
inflammation with necrosis (Mycobacterium, Chlamydia, and fungi) and massive tissue
loss. Monocytosis is often accompanied by heterophilia.1,9
Leukopenia can depend on pancytopenia, panleukopenia, granulocytopenia, and
lymphopenia. Heteropenia is the most frequent leukopenia and is associated with
severe septicemic and endotoxemic infections or with some viral diseases such as
Pacheco parrot disease.1,9,10,47 Other causes could be toxemia (drugs, plants, and
chemical substances), nutritional disease anaphylaxis, radiation, and autoimmune
disorders.9 Leukopenia, heteropenia, immature heterophils, and toxic heterophils
86 Capitelli & Crosta

suggest a degenerative response and poor prognosis. In these cases, it is useful to

monitor the decrease in leukocyte count and the variation in leukocyte morphology
with serial leukograms. Lymphopenia can occur with endogenous and exogenous
steroid excess and viremia.1,8–10
Eosinopenia is difficult to assess in psittacines because of their low normal values.

Thrombocyte morphology and abnormalities

Avian thrombocytes originate from mononuclear precursors in the bone marrow, and
occasionally immature thrombocytes are present in the peripheral blood.1,10 Mature
thrombocytes are round to oval cells with a round or oval centrally placed nucleus
with densely clumped chromatin. The nucleus is rounder, more dense, and bigger
than the erythrocyte nucleus, and thrombocytes have a high nucleus to cytoplasm
ratio. Their cytoplasm appears clear or colorless or pale gray-blue and often has
a reticulated aspect. In activated or phagocytic thrombocytes, numerous vacuolations
may be found depending on the degree of the reaction. Thrombocytes can frequently
have some distinct small eosinophilic granules located in the polar area of the cell.
Thrombocyte cells are usually aggregated in clumps of 2, 3, or more and show degran-
ulation of polar bodies, cellular degeneration, and nuclear pyknosis.1,2,9,19,24,43
Avian thrombocytes have an important function in blood clotting and in phagocy-
tosis, participating in the removal of foreign materials from the blood.10,48 They can
become reactive in chronic conditions.43 Thrombocytosis is frequently present in
a marked regenerative response after anemia. Thrombocytopenia is a result of
a depressed bone marrow production either due to excessive peripheral consumption
or destruction, and it is seen in psittacines with severe septicemia and/or diffuse intra-
vascular coagulation.9,10

Clinical Chemistry
Clinical chemistry is important in the diagnosis of many diseases in psittacine species,
but a proper interpretation of the results of a sample from a particular species can only
be achieved if its reference values are available and were obtained using the same
methods.49 Plasma is preferred for routine blood biochemical evaluations, and lithium
heparin is the preferred anticoagulant.4,5
Avian plasma samples are frequently yellow due to the presence of carotenoid
pigments and not bilirubin.16,50 Pink or red plasma is generally caused by hemolysis.50
Common causes of hemolysis are forcing whole blood through a small needle (25–26
gauge), mixing too vigorously or improperly storing the sample (freezing, exposing to
high temperatures or maintaining at room temperature for a long time).5,16 Furthermore,
lipemia can interfere with many biochemical analysis. It is not routinely possible to obtain
fasting samples from birds because of the particular nature of their digestive physiology
and anatomy, and because it is not advisable to subject a sick bird to fasting.4,5,51
Veterinary laboratories, clinical pathologists, and practitioners must develop the
techniques and services needed for avian biochemical analysis and interpretation.
Biochemical reference ranges for common species of psittacines have been obtained
by specialized laboratories and published.11–14,49,52–56 A computerized database can
be downloaded ([Link]). The physiologic and anatomic conditions in birds are
different from mammals and require the use of different tests. Correct knowledge of
avian physiology is important for the proper interpretation of routine chemistry values.16

Nitrogenous waste products

The role of the avian kidney is to eliminate nitrogenous waste products and regulate
homeostasis with water and electrolytic exchanges. Birds are uricotelic animals and
 Overview of Psittacine Blood Analysis 87

are able to eliminate or accumulate large amounts of uric acid with little water. The
avian kidney has two different types of nephrons: the mammalian-type nephron and
the “reptilian” nephron. The reptilian nephron (uricotelic) lacks the Henle’s loop and
receives blood from the renal portal system. The mammalian-type nephron instead
contains the Henle’s loop and is involved in the osmotic gradient process. A single
avian nephron has a lower glomerular filtration rate (GFR) than a mammalian nephron
but birds have a higher number of nephrons resulting in an overall similar GFR. Avian
kidneys bypass blood from reptilian to mammalian-type nephrons if the GFR is
reduced, improving the ability to concentrate urine. The ability to concentrate urine
is inverse with size, so that smaller birds have a greater ability to concentrate
urine.16,57–59

Uric acid
Uric acid is the major nitrogenous waste product and is an oxidized form of hypoxan-
thine.16 It is synthesized predominantly in the liver from purine metabolism and, in
a lesser amount in the renal tubules. In normal conditions, most of the uric acid
(80%–90%) is secreted actively in the proximal convoluted tubules, whereas a small
part (10%) is filtered by the glomerulus.59 Uric acid secretion is active through the renal
tubules and is not affected by dehydration until it is severe that is when the flow of
urine is insufficient to get the condensed uric acid into the convolute tubules. The
blood levels of uric acid can be influenced by species, age, and diet. In young birds,
the levels are lower than in adults. The concentration of uric acid can increase only
with serious kidney damage (<30% functionality) or severe dehydration. In general,
a concentration of uric acid in the blood greater than 15 mg/dL indicates renal impair-
ment from a variety of causes such as nephrotoxic aminoglycoside antibiotics or lead
toxicity, urinary tract obstruction, nephritis, nephrocalcinosis, and nephropathy asso-
ciated with hypovitaminosis A. It can also increase after a hyperproteic meal intake or
with severe tissue necrosis or starvation (increased nitrogenous catabolism). Uric acid
levels in the blood are the sum of production and secretion and can be normal even if
the kidney function is compromised when the bird is not eating or has a liver disease at
the same time. Furthermore, in polyuric and polydipsic nephropathic birds, the filtra-
tion of urine can increase compensating the decreased secretory capacity.16,51,59
Evaluation of uric acid levels is useful for monitoring treatment or progress of
disease when used with serial blood tests. When the plasma uric acid concentration
exceeds the solubility of sodium urate, uric acid (in the form of urate crystals) precip-
itate in the tissues (especially in the synovial joint and on the visceral surface) and
cause gout. In these cases, blood uric acid levels are greatly increased (eg, fivefold
greater than normal) and result from severe renal dysfunction.51

Creatinine and urea

Both urea and creatinine levels in birds are low compared with those of mammals and
may be less than the minimum assay level for standard laboratory techniques. Nor-
mally, creatinine plasma concentrations in African gray parrots and macaws are higher
than in Amazons and cockatoos. Increased levels are rare and occur in severe kidney
disease. Routine estimations of this constituent have poor diagnostic value in avian
medicine.49,60
Urea and uric acid, together, can be useful to evaluate the urea to uric acid ratio to
differentiate between dehydration, postprandial effects, and renal pathologies. This
ratio, as well as being an indicator of the state of dehydration, may also be useful
as an indicator of urinary fluid flow, being markedly increased in cases of renal failure
with reduced urinary flow.16,49
88 Capitelli & Crosta

Enzymes in liver and muscle tissue

Experimental studies on Melopsittacus ondulatus and Psittacus erithacus involving the
sensitivity and specificity of the enzymes in liver and muscle have been performed.
The specificity and sensitivity of these enzymes may vary with the species and the
type of disease because the organ distribution of these enzymes is different among
the species. Generally, the normal activity of a specific enzyme is a reflection of the
amount of tissue that contains it and increased enzyme values give a measure of
the damage of an organ and not of a decrease in its function.61
Aspartate aminotransferase High aspartate aminotransferase (AST) activity has been
found in a multitude of tissues but mainly in liver and muscle tissues, and its distribu-
tion among tissues can vary with the species. Plasma AST activity is sensitive but not
specific for hepatocellular damage and muscle injury, and is frequently evaluated with
CPK (a muscle-specific enzyme) to differentiate between liver and muscle
damage.16,61 Increases in plasma AST activity in birds are moderate when greater
than 350 IU/L (depending on different species and reference laboratory values) and
can result from hepatic or muscle damage. Increased plasma AST activity is marked
when greater than 800 IU/L. Increases of this magnitude may occur in severe hepatic
damage in the presence of biliverdinuria or biliverdinemia.16,51,61 In psittacines,
increases in AST activity occur with all liver diseases, including Pacheco disease, chla-
mydiosis, exposure to toxic chemicals, the use of some drugs (eg, doxycycline injec-
tion and antifungal drugs such as ketoconazole, fluconazole, and itraconazole).60
Lactate dehydrogenase Lactate dehydrogenase (LDH) isoenzymes are found in most
avian tissues and they are not specific for hepatocellular disease. Plasma LDH activity
is often markedly increased in association with hepatocellular disease or muscle
damage, but compared with plasma AST and alanine aminotransferase (ALT) activity,
it increases and declines more rapidly. Hemolysis causes a high increase in plasma
LDH activity because avian red cells have a high LDH activity.16,51,61
CPK Increased plasma CPK activity is found only in muscle tissue and is a muscle-
specific enzyme to detect muscle cell damage in birds. The normal plasma CPK
activity in most psittacine species is between 100 and 500 IU/L.51 Plasma CPK activity
is higher if any muscle cell damage is present, even with marked exertion or in birds
struggling during blood collection or suffering from seizure disorders. Muscle tissue
damage can happen with trauma, intramuscular injection of irritating fluids, or
systemic infections that affect the skeletal or cardiac muscles. It can be useful for
comparison with AST activity to differentiate muscle injury from hepatocellular injury,
which can be a cause of increased AST activity. When AST, CPK, and LDH are
increased, skeletal muscle injury should be considered. When the plasma CPK activity
is increased due to intramuscular injection, forced exercise, handling, and traumatic
injuries, AST and LDH activity can be lower. Under these conditions, if a bird has a pre-
existing hepatocellular injury, it could have an increase in both AST and the CPK
activity leading to misinterpretation. Plasma AST has a longer half-life than CPK,
and after a single muscle injury, as with an intramuscular injection of an irritating
drug, CPK activity may return to normal before the AST activity. In these cases, an
erroneous diagnosis of hepatobiliary disease may be made.16,61
Amylase Amylase is not a specific enzyme. Information and studies on birds are poor
and pancreatic diseases are rare. Amylase activity is greatly increased in pancreatic
disease but also in other pathologies. Zinc toxicosis can cause pancreatitis and the
higher the toxic levels, the more it is associated with the increases in plasma amylase
levels.53,62
 Overview of Psittacine Blood Analysis 89

Electrolytes
Potassium Potassium concentrations can increase or decrease, if blood samples not
separated immediately and the degree of artifactual changes depends on the
species.53 In macaws the potassium levels in blood samples increase by approxi-
mately 30% within 4 hours.16 Decreased values have been associated with chronic
diarrhea, starvation, and alkalosis. True hyperkalemia is possible in renal disease,
acidosis, and severe tissue necrosis.16,51
Sodium Sodium is the primary osmotically active electrolyte in avian plasma and
urine.51,63 Dietary sodium is absorbed in the intestinal tract, carried to the kidneys
and excreted by glomerular filtration. According to the bird’s need for sodium, it may
be reabsorbed into the plasma or secreted by the renal tubules, and then excreted.51
Hyponatremia (<130 mEq/L) can be associated with an excessive loss of sodium,
such as in renal disease and diarrhea, or with excessive hydration, such as polydipsia
or iatrogenic low-sodium intravenous fluids. Hypernatremia (>160 mEq/L) is seen rarely
with excessive dietary salt intake, dehydration due to decreased water intake, or
increased water loss such as in diarrhea, renal failure, or rarely, diabetes insipidus.51

Calcium The values of total calcium can be much higher than the normal physiologic
values tolerated by mammals. Marked increases in plasma total calcium levels are
seen in oviparous females in the reproductive phase, due to estrogen stimulation for
the production of calcium-binding proteins such as vitellogenin and albumin.16,63
The calcium metabolism is controlled by parathyroid hormone (PTH), calcitonin, and
vitamin D3 (ie, 1,25-dihydrocholecalciferol, calciferol). Estrogens, corticosteroids, total
thyroxine (T4), and glucagon also influence calcium metabolism.
PTH maintains normal calcium plasma levels affecting bone, kidney, and intestinal
mucosa; it mobilizes the calcium from bones, increases calcium absorption by the
intestinal mucosa and calcium reabsorbtion by the renal tubules, aiding in the mainte-
nance of the normal plasma concentration of ionized calcium. Calcitonin has the
opposite action to PTH, to prevent excessive calcium reabsorbtion from bones.51,63
The normal plasma calcium levels in nonlaying females is around 8 to 11 mg/dL and
one-third or one-half of this calcium is bound to albumin. Consequently, the total
plasma calcium concentration is influenced by plasma albumin levels and can
decrease with hypoalbuminemia and increase with hyperalbuminemia.51 A significant
relationship was only observed between albumin and calcium concentrations in
African gray parrots and correction formulas have been developed (adjusted Ca
(mmol/L) 5 Ca (mmol/L)  0.015 albumin (g/L) 1 0.4).52 The ionized calcium (active
fraction) is not affected by changes in the albumin plasma level and could differentiate
pathologic from physiologic increases in total calcium concentration. Some avian
veterinarians use ionized calcium considering the normal range values to be between
1.0 and 1.3 mmol/L (P. Gibbons, personal communication)16 or between 0.96 and
1.22 mml/L in African gray parrots.64 Blood samples for ionized calcium assay should
be tested as soon as possible because pH changes can affect this parameter.65 Hypo-
calcemia (<8.0 mg/dL) in most psittacine species may be caused by dietary calcium
and vitamin D3 deficiency, excessive dietary phosphorus, alkalosis, and hypoalbumi-
nemia.51 Marked hypocalcemia may be caused by malnutrition or reproductive abnor-
malities such as chronic egg laying.16 African gray parrots often are affected by
a hypocalcemic syndrome rarely seen in other psittacines64,65 with a plasma calcium
concentration of less than 6.0 mg/dL that results in seizure disorders.51,63
Hypercalcemia (>11 mg/dL) in most psittacine species has been correlated
with hypervitaminosis D3, osteolytic bone lesions secondary to neoplasm, and
hyperalbuminemia.51
90 Capitelli & Crosta

Phosphorus Plasma phosphorus is primarily maintained by renal excretion. Young

birds have a normal plasma phosphorus level higher than adults. Hypophosphatemia
is present with hypovitaminosis D3, malabsorption, or starvation, and even long-term
corticosteroid therapies. Hyperphosphatemia can occur with severe renal diseases
and hypervitaminosis D3. Artifactual hyperphosphatemia may be caused by hemolysis
and unseparated whole blood can produce a leakage of intracellular phosphorus.51

Metabolites
Biliverdin and bilirubin The major bile pigment in birds is biliverdin and not bilirubin
(due to a lack of biliverdin reductase); it is a green pigment and biliverdinuria is respon-
sible for the green discoloration of the urates often seen in avian liver disease.16,51,66
Biliverdin is not currently measured in clinical laboratories, only in research laborato-
ries.16,67 The presence of green serum or plasma occurs in severe hepatobiliary
diseases and is associated with a poor prognosis.51,66

Bile acids The determination of bile acids is a sensitive test for liver function in some
species of birds. Bile acids are synthesized in the liver from cholesterol and excreted in
the bile; more than 90% of these acids are reabsorbed by the jejunum and ileum into
the portal circulation, and carried from the blood by the hepatocytes.16,66 This process
is referred to as the enterohepatic circulation.51,66 Unlike mammals, a single sample
after fasting for 12 hours overnight is preferred to eliminate random postprandial
increases in bile acids, but in debilitated birds and smaller species fasting is not rec-
ommended.16,66 Fasting plasma bile acid concentrations do not differ with or without
a gallbladder.22,66 The bile acid concentration can vary depending on the analytical
method used (radioimmumoassay test or enzymatic colorimetric method).16,51,66
The enzymatic method for determination of bile acids is preferable but has been
only validated in a few species and lipemia and hemolysis can interfere with this spec-
tophotometric method.16,66 The reference range for bile acids has been obtained
using the enzymatic method.49 The reference range for African gray parrots, cocka-
toos and macaws is 18 to 71 mmol/L. In healthy Amazon parrots, normal fasting bile
acid levels are slightly higher than in other psittacines, between 19 and 144 mmol/
L.49,51 In other references, the normal plasma values of bile acids in Amazons and
African gray parrots differ slightly from these.66 The increases in fasting plasma bile
acid concentrations are suggestive of abnormal hepatic uptake, bile acid storage,
excretion, or hepatic perfusion.51 In one report, some psittacine species were tested
to evaluate bile acid levels, AST and CPK activity, and electrophoresis. The results
showed that enzymatic activities correlate poorly with high levels of bile acids,
whereas bile acids have the highest association with the positive results of hepatic
biopsies.68 This suggests that liver disease can be present without increases in
enzyme activity but it is probably present with high plasma levels of bile acids. Bile
acids levels cannot differentiate the kind of liver disease but they can be useful to
detect the presence of a hepatic disease and to assess the need for a liver biopsy
or to monitor birds undergoing therapy for liver disease.66

Cholesterol Plasma cholesterol is eliminated in the form of bile acids and its concen-
tration increases in extrahepatic biliary obstruction, hepatic fibrosis, and bile duct
hyperplasia. Hypercholesterolemia can also be caused by hypothyroidism, high-fat
diets, and lipemia. Hypocholesterolemia may be associated with end-stage liver
failure, maldigestion or malabsorption, and starvation.51

Glucose Glucose metabolism in birds is controlled by insulin and glucagon as

in mammals. Healthy psittacines have blood glucose concentrations greater than
 Overview of Psittacine Blood Analysis 91

150 to 200 mg/dL. In mammals, insulin has a primary role in glucose homeostasis; in
birds this role belongs to glucagon with some differences among the species.16,51,63
Diabetes mellitus in psittacines is attributed to increased glucagon secretion but
there are some reports describing a decreased blood insulin concentration and
a significant response to insulin therapy. Therefore, either hyperglucagonemia or
hypoinsulinemia can be involved in diabetes in psittacines.51 Hypoglycemia (<150–
200 mg/dL) occurs with prolonged starvation, severe liver disease (eg, Pacheco
disease), septicemia, enterotoxemia, endocrine disorders such as hypothyroidism,
and maldigestion or malasbsorption. The artifactual decrease in glucose levels caused
by nonseparation of plasma or serum from blood as in mammals, is not significant in
birds, because avian erythrocytes use fatty acids rather than glucose for energy. A
mild to moderate hyperglycemia (>500–600 mg/dL) may be induced by high levels
of endogenous or exogenous glucocorticoids as in exertion, excitement, extreme
temperature, stress, or glucocorticoid administration.16,51,64 Marked hyperglycemia
in psittacines (>700 mg/dL and in budgerigars also >900 mg/dL) raises the suspicion
of diabetes mellitus.51,63

Total proteins The normal total protein levels in birds are substantially lower than
those of mammals16,51,63 and generally range from 2.5 to 4.5 g/dL.51 The plasma
proteins consist of albumin, transport proteins, proteins of coagulation, fibrinogen,
enzymes, hormones, and immunoglobulins.63 An essential function of plasma protein
levels is preservation of the normal colloidal osmotic pressure, which maintains the
normal blood volume and pH.51
Some differences exist among psittacine species when different protein standards
and different methods are used in the various laboratories.52 The refractometer
method (easiest and common technique to test total proteins) in psittacines can
produce inaccurate results caused by interference from high concentrations of other
refractive substances such as blood glucose (higher in Psittacines) or lipemia and
hemolysis that can alter the accuracy of the refractometric method.16,51,63,69,70 The
Biuret method (BCG) is more accurate and gives reliable results when the total protein
levels are between 1 and 10 g/dL, but it has not been validated in birds.16,51 The
comparison between results obtained using the BCG method and gel electrophoresis
has shown many differences, probably caused by the use of human albumin stan-
dards and controls, whose binding affinity with the dye is different from avian albumin.
The difference between the total protein levels in psittacines is probably caused by the
use of bovine or human standards lacking species-specific protein standards.16,49,53
The total protein concentration obtained with a BCG method combined with gel elec-
trophoresis represents an absolute accurate concentration of the plasma protein51
and can be used to determine albumin concentration and globulin distribution.16 Elec-
trophoresis is useful for staging acute and chronic inflammatory conditions and for
monitoring therapeutic response in birds.16,63,68 The primary plasma protein fractions
include prealbumin, albumin, a-globulins (a1 and a2), b-globulins, and g-globulins. The
prealbumin fraction may be found in some psittacines. Albumin migration differs
among the species. These different migration patterns are related to variable confor-
mation and surface charge distribution of albumin molecules; this may also explain the
differential binding of BCG, and inaccurate results using human standards.16,71 These
difference were also present in globulin fractions with a lower normal concentration of
g-globulins and an increase primarily in the a- and b-globulin fractions (rather than the
g-globulin fraction) in acute inflammatory disease.72
A recent study has compared electrophoresis results using 3 different commercial
electrophoresis systems in African gray parrots, and has found many differences in
92 Capitelli & Crosta

protein fraction migration.73 Normal concentrations for albumin, a-globulin, b-globulin,

and g-globulin obtained with protein electrophoresis in psittacine birds ranged from
1.5 to 3.0 g/dL, 0.1 to 0.5 g/dL, 0.2 to 0.6 g/dL, and 0.2 to 0.7 g/dL, respectively.
The normal albumin to globulin (A/G) ratio for most psittacine birds is between 1.5
and 3.5.51 Reference intervals for each species of bird should be established in
each laboratory and because the results for protein fractions from psittacine species
are variable in terms of migrations pattern, they should be interpreted with
caution.73–75 Hypoalbuminemia has been found in birds with maldigestion, malab-
sorption, protein-losing enteropathy or nephropathy, and liver disease.16,51,75 Hypo-
proteinemia in birds is frequently associated with hypoalbuminemia (decreased A/G
ratio) and occurs with marked external hemorrhage, renal disease with chronic
proteinuria, or protein-losing enteropathies.16,51,63 A physiologic decrease in A/G ratio
is recurrent in egg-laying females with estrogen-induced hyperproteinemia to
complete egg formation because most of the yolk and chalazae proteins are globulins
and cause a marked increase in globulin fractions.16,51,53,63,75 The acute phase
proteins (a- and b-globulin fractions) are typically increased during inflammation.
Chronic inflammatory disorders such as active hepatitis may increase the a- and b-
globulin fractions.51,75 Immunoglobulins (IgM and IgG) may migrate in the latter frac-
tions. The g-globulin peak contains IgA, IgM, IgG, and IgE immunoglobulins. In
many chronic infections (eg, Chlamydia, Aspergillus, and Mycobacterium spp.), poly-
gammopathy can be seen indicating active chronic inflammatory diseases. Immuno-
deficiency may be present when g-globulin fractions are lowered.51 Hyperproteinemia
can be found in dehydration (normal A/G ratio), in acute or chronic inflammation
(decreased A/G ratio), or in a preovulatory condition. In chronic disturbances, g-glob-
ulins are the most increased fraction. Hyperproteinemia associated with hyperalbumi-
nemia and hypoglobulinemia with an increased A/G ratio is suggestive of dehydration,
chronic stress, or other immunosuppressive conditions (virus such as psittacine beak
and feather disease [PBFD]).16,51,75
Several organ-specific and species profiles used in our clinic and laboratory to eval-
uate the patient in a more complete way are given in Tables 4 and 5.

PART II

This part of the article considers whether there are significant changes in hematologic
and biochemical parameters and if these abnormalities have similarities with those
reported in literature. Possible analytical errors, diagnostic omissions, and potential
risks of overestimated or underestimated diagnoses in some clinical cases presented
to our practice are also considered.

Materials and Methods

A list of all tests evaluated are summarized in Table 6. Blood samples were collected
over 5 years from 150 different psittacines of 29 different species and grouped into 7
genera as summarized in Tables 7 and 8. The study is limited to those birds for which
we had a complete set of results for analysis that could lead to a proper diagnosis.
Most birds were adult (128 of 150 birds ranged between 1 and 25 years); a small number
of birds was young (8 birds were between 3 months and 12 months of age), or were aging
(14 birds were aged between 25 and 50 years). Gender was not determined in all
patients. Usually in our practice a venous blood sample is collected from the right jugular
or subcutaneous ulnar (wing) vein and the selected site depends on bird species, age,
and size; the veterinarian’s/technician’s experience is also an issue. Some samples
were collected from anesthetized birds (isoflurane), others from unanesthetized
 Overview of Psittacine Blood Analysis 93

Table 4
Examples of biochemical profiles of organs

Complete Renal Liver Pancreas Feather Picker

Uric acid Uric acid Uric acid Glucose Uric acid
LDH LDH LDH Lipase LDH
CPK CPK CPK Amylase CPK
AST AST ALT Cholesterol AST
Glucose Glucose AST Triglycerides Glucose
Creatinine Creatinine Glucose Zinc Creatinine
Electrophoresis Electrophoresis Bile acid Complete CBC Plasma proteins
Calcium Calcium Electrophoresis Calcium
Ionized calcium Sodium Complete CBC Phosphorus
Sodium Phosphorus Fecal examination
Phosphorus Potassium Gram stain
Potassium Complete CBC Total thyroxine
Bile acids Complete CBC
Amylase Zinc
Complete blood count

Abbreviations: ALT, alanine aminotransferase; AST, aspartate aminotransferase; CBC, complete

blood count.

patients, using 1- or 2-mL syringes and 23- to 25-gauge detachable needles. The
samples (from 0.8 to 2.0 mL of whole blood) were mixed immediately with lithium
heparin in pediatric collection tubes. Two monolayer blood smears were made with non-
heparinized fresh blood immediately after collection. Most of the blood tests were run

Table 5
Some biochemical profiles for selected species

Amazon Macaw Cockatiel Cockatoo African Gray Budgerigar

Uric acid Uric acid Uric acid Uric acid Uric acid Bile acids
Calcium Calcium AST AST Glucose Fecal stain
AST Phosphorus Glucose Plasma AST CBC
proteins
Glucose Chloride Plasma proteins Microbiology Ionized
calcium
Cholesterol LDH Microbiology CBC Sodium
Plasma AST Chlamydophila Potassium
proteins psittaci AC
Microbiology CPK CBC Plasma
proteins
Chlamydophila Glucose Fecal stain Microbiology
psittaci AC
CBC Plasma CBC
proteins
Cholesterol
CBC

Abbreviations: ALT, alanine aminotransferase; CBC, complete blood count.

94 Capitelli & Crosta

Table 6
Total number of tests performed

Test Number
Hematocrit 134
Total protein 125
Hemoglobin 115
TRBC 115
TWBC 150
Lymphocytes (%) 150
Monocytes (%) 150
Heterophils (%) 150
Eosinophils (%) 150
Basophils (%) 150
Anisocytosis/poikilocytosis 150
Polychromasia 150
Band heterophils 150
Toxic heterophils 20
Zinc 31
Calcium 68
Ionized calcium 37
Phosphorus 51
Sodium 51
Potassium 55
Bile acids 74
Glucose 128
Uric acid 130
Amylase 41
Creatinine 41
CPK 107
Cholesterol 39
LDH 103
AST 129
Prealbumin 75
Albumin 75
a1-Globulin 74
a2-Globulin 74
b-Globulin 74
g-Globulin 74
A/G ratio 75
Total data number 3465

within 4 to 12 hours after collection, with the exception of samples collected from birds
located in other premises. Our laboratory stains blood smears with Giemsa stain; once
the stained smears are dried, they are fixed and mounted with a coverslide glass. For
each sample, a microhematocrit tube is filled with blood with lithium heparin and centri-
fuged at 12,000  g for 5 minutes to determine the PCV. The erythrocyte total count, the
hemoglobin, and granulocyte total count were performed using an automated analyzer
 Overview of Psittacine Blood Analysis 95

Table 7
Total number of selected psittacine species

Species Number
Amazona aestiva 13
Amazona amazonica 2
Amazona auropalliata 1
Amazona autumnalis 1
Amazona brasiliensis 2
Amazona farinosa 1
Amazona festiva 1
Amazona finschii 1
Amazona ochrocephala 10
Amazona o. panamensis 6
Amazona vinacea 2
Amazona xantholora 2
Hybrid Amazon 2
Ara ararauna 18
Ara chloroptera 10
Ara macao 4
Ara militaris 2
Diopsittaca nobilis 4
Ara severa 5
Cacatua alba 8
Cacatua ducorpsii 2
Cacatua galerita 5
Cacatua sanguinea 1
Cacatua s. citrinocristata 2
Eclectus roratus 1
Aratinga solstitialis 1
Aratinga (Nandayus) nenday 1
Nymphicus hollandicus 3
Psittacus erithacus 39
Total 150

(Cell-Dyn 3500 hematology system; Abbot Laboratories, Abbott Park, IL, USA) updated
with specific veterinary software. The hemoglobin concentration needs to be corrected.
The Cell-Dyn analyzer uses the correction factors from the Vet 2.3 software, which are
adapted for some genus/groups or species of psittacines (Ara spp, Amazona spp,
Cacatua spp, Psittacus erithacus, Melopsittacus undulatus) using adequate correction
factors previously developed. For the granulocyte total count, only the optical leukocyte
count (WOC) is used, or (in the presence of interference by erythrocytes and thrombo-
cytes nuclei) the absolute count of neutrophils that corresponds to heterophils. The esti-
mated TWBC count is routinely performed for all samples to evaluate the number of
lymphocytes to compare it with the automated TWBC count and to double check any
abnormal results. The estimated TWBC count and WBC differential count are always
performed by the same operator. The biochemical results were obtained (see
Table 6) using standard spectrophotometric methods, according to the standard
96 Capitelli & Crosta

Table 8
Number of specimens for each group of selected psittacines

Group Number
Amazons 44
Macaws 43
Eclectus 1
Conures 2
Cockatoos 18
Cockatiels 3
African gray parrots 39
Total number of birds sampled 150

protocol provided with the system. Colorimetric methods were used to evaluate
calcium, phosphorus, glucose, cholesterol, creatinine, and uric acid concentrations.
Total bile acids were also measured using an enzymatic colorimetric method; total
proteins were assessed with the BCG method using a human standard. All enzymes
were measured with a kinetic optimized Scandinavian Committee on enzymes (SCE)
method. Sodium, potassium, and ionized calcium have all been tested with ion selective
electrode (ISE). Albumin, prealbumin, and globulin fractions were obtained by plasma
protein electrophoresis using a manual method (cellulose acetate membranes, barbitu-
rate buffer, for 30 minutes at 180 V and stained with Coomassie Blue). The biochemical
analysis were run using the Roche Cobas Mira Classic chemistry analyzer (Roche Diag-
nostics) and AVL 9180 electrolyte analyzer (Roche Diagnostics). The data collected over
a 5-year period were compared with the published reference values for psittacines, as
summarized in Tables 9–11. These results were correlated with the suspected diag-
nosis proposed by the different practitioners and the list of the diagnosed diseases as
summarized in Box 1. The diseases are grouped to facilitate the understanding of the
data.

Results
Table 12 summarizes the correlation between the species/group and the diagnosis.
Table 13 summarizes the rating for each parameter evaluated (total number and
percentage of the hematologic and biochemical normal, decreased, and increased
values, in comparison with the reference range).
Tables 14–16 present a score for each hematologic or blood chemistry parameter
(mild, moderate, marked, decreased, or increased). Data referring to conditions with
less than 3 specimens are excluded from the tables. The 2 cases of neurologic disor-
ders, regarding the same bird (African gray parrot), revealed mild heterophilia and, in
the first control, marked total hypocalcemia and decreased ionized calcium, mild
hyperphosphatemia, marked hypokalemia, moderate hypoglycemia, and an increase
in all muscle enzymes. In the 2 cases of osteomalacia, we observed a marked total
hypocalcemia and marked decreased ionized calcium with light hyperphosphatemia
and moderate increase in amylase, CPK, and LDH activity. In the 2 cases of trauma,
1 only had moderate heteropenia; the other showed marked hyperkalemia. In the
only neoplasm observed, the bird showed moderate hyperproteinemia, leukocytosis
with lymphocytosis and monocytosis, moderate hyperphosphatemia and high
amylase activity, mild increase of CPK activity, a marked decrease in A/G ratio with
mild albumin decrease and severe increase in a2-globulin and b-globulin fractions.
 Overview of Psittacine Blood Analysis 97

Discussion
This study refers only to psittacines living in Italy. According to our data, Amazons as
a group, seem to be more susceptible to bacterial infections and are the second group
for sensitivity to psittacosis. On the other hand, Amazons are less prone to be infected
by viruses compared with macaws. Macaws are apparently more susceptible to infec-
tious disease; specifically, they seem more prone to chlamydiosis and viral infections,
followed by bacterial, mycotic infections, and air sac infections (aerosacculitis).
African gray parrots are more susceptible to aerosacculitis and fungal infections
and are the second group for sensitivity to bacterial infections; conversely they
seem to be less sensitive to viruses and chlamydiosis as reported in the literature.65
Cockatoos are less susceptible to bacterial and viral infections; they also seem to
be more resistant to psittacosis compared with other large parrots.65
As suggested in the literature,65 organ-related diseases are more typically diag-
nosed in Amazons; liver diseases are the most represented followed by renal disease.
In macaws, there are some cases of enteritis, cloacal diseases, and reproductive
disorders but liver diseases are rare. African gray parrots are the second group of
parrots most susceptible to organ-related diseases and particularly to heart disease,
neurologic disorders, and osteomalacia. The latter two may be related to the typical
hypocalcemic syndrome of African gray parrots.51,52,63–65 The other psittacine species
show no evidence of these diseases. The last group is represented by some typical
psittacine issues and miscellaneous diseases. The most susceptible group is the
African gray parrots, which have the most dietary disorders followed by some cases
of feather and beak disorders, behavioral disorders, trauma, and gizzard foreign
bodies according to Rosskopf and Woerpel.65 Cockatoos are more susceptible to
feather and beak disorders and macaws seem to be more susceptible to behavioral
disease and zinc toxicosis. Even though there is only 1 Eclectus in our study, and
thus it is not useful from a statistical point of view, it is interesting that the bird was
referred for zinc toxicosis. It is reported in the literature that Eclectus parrots and cock-
atoos have higher normal levels of zinc in plasma or serum than other psittacines. The
authors suggest taking into consideration the species of parrots when evaluating zinc
toxicosis.76 The rest of the organ-related pathologies are rare or absent in the other
psittacine species. The group of diseases referred to as check are not considered
in this evaluation because they do not have any specific diagnostic value.
Considering the data summarized in Table 13, all the abnormal values obtained may
have an explanation compatible with what has been described earlier; that the hemo-
globin level is higher than the PCV and RBC count might depend on an artifactual
increase due to lysis of erythrocytes and thrombocytes. Leukocytosis, which is
commonly observed in psittacines, is normally seen with heterophilia.9,18,19,39 Leuko-
penia is rarely observed in birds, and is normally caused by a low number of hetero-
phils (heteropenia). Monocytosis is less common than heterophilia, but is more
common than lymphocytosis.1,9 As frequently reported, sick parrots are brought to
the veterinarian at an advanced, chronic stage of the disease; this could explain the
frequency of birds presenting with monocytosis. Eosinophilia and eosinopenia are
rarely observed in birds, as highlighted in Table 13. Also basophils are rare or
absent.1,9 Banded and toxic heterophils are observed less frequently and this happen
because it is not easy to recognize them and because they are not often present
unless in severe inflammatory diseases.1,9,10 Electrolytes (Na, K, phosphorus) show
a higher percentage of abnormal values but they are also the parameters most
affected by artifactual interference.4,5,16,21,53 Glucose and uric acid have lesser
percentages of abnormal values and are decreased most of the time; that can
 98
Capitelli & Crosta
Table 9
Reference values for hematology and clinical chemistry parameters in selected psittacine species
Cacatua Cacatua
Psittacus Amazona galerita Cacatua sulphurea Eclectus Nandayus
erithacus Group Nymphicus Cacatua (Sulfur- sanguinea (Yellow- roratus nenday Aratinga
(African (Amazon Ara Group hollandicus Group Crested (Little Crested (Eclectus (Nanday solstitialis
Units Gray Parrot) Parrots) (Macaws) (Cockatiel) (Cockatoos) Cockatoo) Corella) Cockatoo) Parrot) Conure) (Sun Conure)
Hematocrit % 43–51a 44–56a 45–55a 45–57a 40–54b 41–49a — — 35–47c 39–49c,d 39–49c,d
Red blood cells 10 /mL 3.0–3.6
6 a
2.6–3.5 a
2.5–4.5 a
2.5–4.7 a
2.5–2.95 b
2.4–3.0 a
— — 2.4–3.9 c
2.5–4.0 c,d
2,5–4.0c,d
White blood 103/mL 6.0–12b 5.0–17b 6.0–13.5a 5.0–10a 5.0–13b 12–16b 7.0–13b 9.0–16b 9.0–15b 5.0–11b 6.0–11b
cells
Hemoglobin g/dL 14.2–17.0a 13.8–17.9a 14.8–18.9a — 12.0–14.8b 13.8–17.1a — — 11.5–16.0c 12.0–16.0c,d 12.0–16.0c,d
Lymphocytes % 19–50b 20–67b 20–50a 25–55a 20–50b 22–56b 22–51b 21–56b 23–57b 24–47b 20–49b
Monocytes % 0–2b 0–2b 0–3a 0–2a 0–2b 0–2b 0–1b 0–2b 0–1b 0–1b 0–1b
Heterophils % 45–73b 31–71b 45–70a 40–70a 45–72b 44–75b 45–72b 44–79b 46–70b 48–71b 44–72b
Eosinophils % 0–1b 0–0b 0–2a 0–2a 0–2b 0–1b 0–2b 0–2b 0–1b 0–1b 0–1b
b b a a b b b b b b
Basophils % 0–1 0–2 0–5 0–6 0–1 0–1 0–1 0–1 0–1 0–2 0–2b
a a c b a c c,d
MCV fL 137–155 156–194 90–185 — 154–170 145–187 — — 95–220 90–190 90–190c,d
a a c b a c c,d
MCH pg 41.9–52.8 44.7–58.6 27–53 — 45.0–55.5 53.8–60.6 — — 27.0–55.0 28.0–55.0 28.0–55.0c,d
a a c b a c c,d
MCHC g/dL 28.9–34.0 28.9–35.8 23.0–32.0 — 24.1–32.9 33.3–37.6 — — 22.0–33.0 23.0–31.0 23.0–31.0c,d
Thrombocytes K/mL 11–42a 10–67a — — 11–34a 7–24a — — — — —
Amylase U/L 415–626b 184–478b 239–564b 113–870b 228–876b — 273–536b 136–750b 150–645c — —
AST U/L 110–340b 150–344b 65–168b 128–396b 140–360b 120–208b 140–350b 134–276b 144–339b 147–378b 138–355b
 Bile acids mmol/L 12–96b 33–154b 7–100b 44–108b 34–112b 24–98b 15–96b 24–95b 30–110b — 12–92b
b b b b b b b b c b
Calcium mg/dL 8–14 8–13.9 8.4–11.9 8.2–10.9 8.2–11.5 8–12 8–11.4 8.1–10.3 7–13.0 8–11 8–11.2b
b b b b b b b b
Cholesterol mg/dL 100–250 148–228 96–264 — 96–212 146–248 — 113–206 100–261 117–228 112–255b
CPK U/L 140–411b 117–425b 88–361b 160–420b 147–418b 164–396b 144–439b 157–408b 132–410b 135–400b 153–372b
Creatinine mg/dL 0.1–0.4c 0.1–0.4c 0.1–0.5c 0.1–0.4c 0.1–0.4c — — — 0.1–0.4c — —
Ionized meq/L 0.96–1.22e — — — — — — — — — —
calcium
Glucose mg/dL 256–360b 246–378b 210–360b 228–440b 206–418b 200–345b 220–412b 138–304b 216–396b 231–393b 167–380b
LDH U/L 154–378b 160–368b 70–220b 122–378b 208–414b 118–374b 198–452b 178–398b 198–386b 204–396b 155–346b
c c c c c c
Phosphorus mg/dL 3.2–5.4 3.1–5.5 2.0–12.0 3.2–4.8 2.5–5.5 — — — 2.9–6.5 — —
Potassium meq/L 2.9–4.6c 3.0–4.5c 2.0–5.0c 2.4–4.6c 2.5–4.5c — — — 3.5–4.3c — —
Sodium meq/L 157–165c 125–155c 140–165c 130–153c 130–155c — — — 130–145c — —
Total protein g/dL 2.7–4.4b 2.6–4.5b 2.4–4.4b 2.1–4.8b 2.6–4.8b 2.4–4.8b 2.4–4.3b 2.8–4.6b 3.2–4.3b 1.9–3.7b 2.4–4.5b
Uric acid mg/dL 2.2–11b 2.2–10b 1.8–12b 3.4–11b 3.8–11b 2.2–11b 3.5–11b 2.1–11b 2.0–11b 2.3–11b 2.2–12b
a
Data for adults of both sexes, as obtained from Hawkey CM, Samour JH. The value of clinical hematology in exotic birds. In: Jacobson ER, Kollias GV, Jr, editors.
Exotic animals: contemporary issues in small animal practice. London: Churchill Livingstone; 1988.
b
Data for adults of both sexes, as obtained from The Californian Avian Laboratory, Citrus Heights, CA 95621. Reference range 1998 from Fudge AM. Laboratory

Overview of Psittacine Blood Analysis

reference ranges for selected avian, mammalian, and reptilian species. In: Fudge AM, editor. Laboratory medicine: avian and exotic pets. Philadelphia: WB Saun-
ders; 2000. p. 376–400.
c
Data for adults of both sexes, as obtained from The Avian and wildlife Laboratory, University of Miami School Medicine, Miami, FL 33136.
d
The data are referred to the Conure group.
e
Data for adults of both sexes, as obtained from Stanford M. The significance of serum ionized calcium and 25-hydroxy-cholecalciferol (vitamin D3) assays in
African grey parrots. Exotic DVM 2003;5(3):1–6.

99
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Capitelli & Crosta
Table 10
Reference values for hematology and clinical chemistry in selected macaws and Amazon species
Amazona Amazona Amazona Amazona Ara
aestiva amazonica auropalliata Amazona Amazona ochrocephala Ara chloroptera Ara Ara Ara Diopsittaca
(Blue- (Orange- (Yellow- autumnalis farinose (Yellow- ararauna (Green- macao militaris severa nobilis (Red
Fronted Winged Naped (Red-Lored (Mealy Crowned (Blue and Winged (Scarlet (Military (Severe Shouldered
Units Amazon) Amazon) Amazon) Amazon) Amazon) Amazon) Gold Macaw) Macaw) Macaw) Macaw) Macaw) Macaw)
Hematocrit % 44–56a,b 44–56a,b 44–56a,b 44–56a,b 44–56a,b 44–56a,b 41–51a 41–56a 46–52a 44–50a 45–53a 41–51a,b
Red cells 10 /mL 2.6–3.5
6 a,b
2.6–3.5 a,b
2.6–3.5 a,b
2.6–3.5 a,b
2.6–3.5 a,b
2.6–3.5 a,b
2.7–3.5 a
2.8–3.3 a
2.7–3.2 a
2.7–3.1 a
3.1–3.6 a
2.7–3.5a,b
White cells 103/mL 6.0–13c 7.0–16c 6.0–17c 6.0–16c 9.0–16c 8.0–15c 8.0–16c 11.0–16.0c 10.0–14.0c 12.0–20.0c 9.0–14.0c 8.0–22.0c
Hemoglobin g/dL 13.8–17.9a,b 13.8–17.9a,b 13.8–17.9a,b 13.8–17.9a,b 13.8–17.9a,b 13.8–17.9a,b 14.8–18.9a 14.7–18.8a 15.8–18.4a 14.8–19.6a 16.0–17.1a 14.8–18.9a,b
Lymphocytes % 22–65c 20–64c 20–67c 22–66c 23–65c 18–63c 18–53c 22–60c 23–50c 25–40c 25–47c 32–66c
c c c c c c c c c c c
Monocytes % 0–1 0–1 0–1 0–1 0–1 0–1 0–2 0–2 0–2 0–2 0–2 0–1c
Heterophils % 33–72c 31–73c 31–73c 33–73c 38–76c 37–69c 49–71c 49–73c 48–73c 58–85c 45–80c 34–63c
Eosinophils % 0–1c 0–0c 0–0c 0–0c 0–0c 0–1c 0–1c 0–2c 0–1c 0–1c 0–1c 0–1c
c c c c c c c c c c c
Basophils % 0–1 0–2 0–2 0–2 0–1 0–1 0–1 0–1 0–1 0–1 0–2 0–1c
a,b a,b a,b a,b a,b a,b a a a a a
MCV fL 156–194 156–194 156–194 156–194 156–194 156–194 132–157 141–174 143–175 154–160 145–156 132–157a,b
MCH pg 44.7–58.6a,b 44.7–58.6a,b 44.7–58.6a,b 44.7–58.6a,b 44.7–58.6a,b 44.7–58.6a,b 49.4–56.4a 50.5–57.7a 51.1–64.2a 40.6–59.6a 47.9–55.2a 49.4–56.4a,b
MCHC g/dL 28.9–35.8a,b 28.9–35.8a,b 28.9–35.8a,b 28.9–35.8a,b 28.9–35.8a,b 28.9–35.8a,b 34.7–39.8a 29.1–38.3a 32.6–38.5a 34.7–37a 32.1–35.6a 34.7–39.8a,b
a,b a,b a,b a,b a,b a,b a a a a
Thrombocytes K/mL 10–67 10–67 10–67 10–67 10–67 10–67 11–34 8–15 17–30 19–30 10–13a 11–34a,b
c,b c,b c c c c,b c c,b c,b c,b c,b
Amylase U/L 184–478 184–478 187–546 156–472 36–574 184–478 239–516 239–564 239–564 239–564 239–564 239–564c,b
AST U/L 146–408c 114–389c 150–390c 150–408c 138–377c 150–380c 64–168c 62–168c 74–177c 76–166c 72–170c 126–215c
 Bile acids mmol/L 34–140c 29–157c 36–144c 24–120c 31–145c 59–188c 27–86c 15–78c 21–100c 24–130c 25–82c 7–100c,b
Calcium mg/dL 8.2–13.8c 8–14.5c 8.4–13.2c 8–13.2c 8.6–12.9c 8.6–11.7c 8.4–11.8c 8.4–11.8c 8.4–10.9c 8.4–10.9c 8.4–10.8c 8.4–11.9c,b
Cholesterol mg/dL 100–270c 180–254c 100–256c 150–228c 111–249c 125–246c 96–249c — — — 100–252c 96–264c
c c c c c c c c c c c
Creatinine U/L 130–417 120–455 132–402 136–420 208–348 260–490 92–380 96–368 98–366 126–321 123–356 88–361c,b
kinase
Creatinine mg/dL 0.1–0.4d,b 0.1–0.4d,b 0.1–0.4d,b 0.1–0.4d,b 0.1–0.4d,b 0.1–0.4d,b 0.1–0.5d,b 0.1–0.5d,b 0.1–0.5d,b 0.1–0.5d,b 0.1–0.5d,b 0.1–0.5d,b
Ionized meq/L — — — — — — — — — — — —
calcium
Glucose mg/dL 246–389c 249–482c 249–377c 250–388c 201–337c 170–316c 210–368c 210–360c 210–333c 214–340c 220–368c 231–300c
LDH U/L 158–366c 148–451c 160–360c 150–412c 159–381c 171–265c 69–220c 72–224c 60–210c 72–216c 66–210c 70–220c,b
d,b d,b d,b d,b d,b d,b d,b d,b d,b d,b d,b
Phosphorus mg/dL 3.1–5.5 3.1–5.5 3.1–5.5 3.1–5.5 3.1–5.5 3.1–5.5 2.0–12.0 2.0–12.0 2.0–12.0 2.0–12.0 2.0–12.0 2.0–12.0d,b
Potassium meq/L 3.0–4.5d,b 3.0–4.5d,b 3.0–4.5d,b 3.0–4.5d,b 3.0–4.5d,b 3.0–4.5d,b 2.0–5.0d,b 2.0–5.0d,b 2.0–5.0d,b 2.0–5.0d,b 2.0–5.0d,b 2.0–5.0d,b
Sodium meq/L 125–155d,b 125–155d,b 125–155d,b 125–155d,b 125–155d,b 125–155d,b 140–165d,b 140–165d,b 140–165d,b 140–165d,b 140–165d,b 140–165d,b
Total protein g/dL 3.5–6.5c 2.7–4.9c 2.8–4.7c 2.3–4.4c 2.4–4.2c 3.2–4.6c 2.5–4.2c 2.7–4.2c 2.4–3.8c 3.0–3.9c 2.3–3.9c 2.4–4.4c,b
c c c c c c c c c c c
Uric acid mg/dL 2.3–10 2.3–11 2.3–11 2.2–9.9 2.3–12 2.3–11 1.9–11 1.5–11 2.1–11 2.0–11 2.2–12 2.2–10c

a
Data for adults of both sexes, as obtained from Hawkey CM, Samour JH. The value of clinical hematology in exotic birds. In: Jacobson ER, Kollias GV, Jr, editors. Exotic animals: contem-
porary issues in small animal practice. London: Churchill Livingstone; 1988.
b
The data are referred to the Amazona and Ara groups.
c

Overview of Psittacine Blood Analysis

Data for adults of both sexes, as obtained from The Californian Avian Laboratory, Citrus Heights, CA 95621, Reference range 1998 from Fudge AM. Laboratory reference ranges for
selected avian, mammalian, and reptilian species. In: Fudge AM, editor. Laboratory medicine: avian and exotic pets. Philadelphia: WB Saunders; 2000. p. 376–400.
d
Data for adults of both sexes, as obtained from The Avian and Wildlife Laboratory, University of Miami School Medicine, Miami, FL 33136.

101
102 Capitelli & Crosta

Table 11
Reference ranges for protein electrophoresis in some psittacines species/groups

Fraction Reference Range (g/dL)

Amazona spp.
Prealbumin 0.35–1.05
Albumin 1.9–3.52
Alpha-1 0.05–0.32
Alpha-2 0.07–0.32
Beta 0.12–0.72
Gamma 0.17–0.76
Ratio A/G 1.9–5.9
Ara spp.
Prealbumin 0.05–0.7
Albumin 1.24–3.11
Alpha-1 0.04–0.25
Alpha-2 0.04–0.31
Beta 0.14–0.62
Gamma 0.1–0.62
Ratio A/G 1.6–4.3
Cacatua spp.
Prealbumin 0.24–1.18
Albumin 1.8–3.1
Alpha-1 0.05–0.18
Alpha-2 0.04–0.36
Beta 0.22–0.82
Gamma 0.21–0.65
Ratio A/G 2.0–4.5
Nymphicus hollandicus
Prealbumin 0.8–1.6
Albumin 0.7–1.8
Alpha-1 0.05–0.4
Alpha-2 0.05–0.44
Beta 0.21–0.58
Gamma 0.11–0.43
Ratio A/G 1.5–4.3
Psittacus erithacus
Prealbumin 0.03–1.35
Albumin 1.57–3.23
Alpha-1 0.02–0.27
Alpha-2 0.12–0.31
Beta 0.15–0.56
Gamma 0.11–0.71
Ratio A/G 1.6–4.3
Eclectus
Prealbumin 0.4–1.04
Albumin 2.3–2.6
(continued on next page)
 Overview of Psittacine Blood Analysis 103

Table 11
(continued)

Fraction Reference Range (g/dL)

Alpha-1 0.09–0.33
Alpha-2 0.11–0.27
Beta 0.17–0.43
Gamma 0.18–0.55
Ratio A/G 2.62–4.05
Conure
Prealbumin 0.18–0.98
Albumin 1.9–2.6
Alpha-1 0.04–0.23
Alpha-2 0.08–0.26
Beta 0.07–0.47
Gamma 0.12–0.61
Ratio A/G 2.2–4.3

Values from the Avian and Wildlife Laboratory, University of Miami, School Medicine, Miami, FL
33136. Beckman Paragon system protein electrophoresis.

mean an aspecific pathologic condition if the data are considered by themself. Bile
acids have a medium percentage of abnormal values and show increased values.
The muscle enzymes except AST/GPT have a higher percentage (highest LDH) of
abnormal values. This fact may also depend on intramuscular injection, forced exer-
cise, handling and traumatic injuries while travelling to the practice or during restraint.
The marked artifactual effects that can affect LDH plasma activity should not be
forgotten. Creatinine values are nearly always within the range. This confirms that
this test is rarely useful in routine profiles.60 Considering protein electrophoresis, there
is a higher percentage of abnormal decreased values of prealbumin, whereas the
percentage of abnormal decreased values of albumin are much lower. This could
also be an analytical error because of the different chemical and molecular features
of the different protein fractions (prealbumin) of different species of psittacines and
different methods.72–74 a- and b-globulin fractions have a higher percentage of
increased values that are compatible with leukocytosis and heterophilia (acute-
subacute inflammatory condition), and a high percentage of decreased values of
the A/G ratio. We have only seen 2 cases of increased A/G ratio in relation to
a decrease in globulin fractions and an increase in albumin. The g-globulins have
a higher percentage of marked decreased values. This could be explained by the
fact that the normal g-globulin fractions are usually lower72 or by a state of chronic
stress or immunodeficiency.16,51,75 The percentage of total proteins with increased
values (24%) is not increased and the percentage of decreased values is even less
(7%).
Considering the abnormal results of hematologic and biochemical tests found in
different diseases, in the first group of infectious diseases (as seen in Table 14), during
air sac infection, there are only abnormal values for the total leukocyte count with 1
case of heteropenia and 2 cases of moderate heterophilia, marked monocytosis,
and lymphopenia. Toxic heterophils are always present to a marked degree and this
suggests a degenerative response and poor prognosis.1,8–10 We observed hypogly-
cemia (compatible with severe anorexia and/or septicemia), an increase in plasma
levels of uric acid (dehydration or secondary renal disorders),16,51,59 and 1 case of
104 Capitelli & Crosta

Box 1
List of diseases diagnosed

Infectious diseases
Air sac infection (4)
Bacterial infection (13)
Fungal infection (3)
Chlamydiosis (22)
Viral infection (14)
Organic-related diseases
Enteritis (5)
Cloacal disease (3)
Liver disease (12)
Renal disease (3)
Heart disease (3)
Neurologic disorders (2)
Osteomalacia (2)
Reproductive disorders (3)
Miscellaneous diseases
Feather and beak disorders (8)
Behavioral disorders (8)
Dietary disorders (11)
Gizzard foreign bodies (6)
Zinc toxicosis (4)
Pediatric pathologies (4)
Trauma (2)
Neoplasia (1)
Check (17)a
Total (150)
a
The check group represents the data obtained for those psittacines (for the most part from
breeding centers or pet shops) with a high risk of disease but still without symptoms.

marked increase in all globulin fractions and marked decrease in the A/G ratio
(compatible with a heavy inflammatory and antibody stimulus).75 In the second group
(bacterial infections), (where there are more cases), we observed 5 cases of marked
anemia with only 2 of them showing a regenerative response and almost every time
moderate to marked leukocytosis with heterophylia, monocytosis, lymphopenia and
2 cases of heteropenia.
Biochemical parameters did not provide useful information about the abnormal
serum biochemistry values due to organ infection, as reported in the literature,77
except the plasma glucose levels and electrophoretic patterns. The plasma glucose
levels were severely decreased in 4 cases (similar to aerosacculitis) and there were
3 cases of hypoalbuminemia with a markedly decreased A/G ratio; in 1 of them, the
 Table 12
Comparison between different psittacine species and different diseases

Infection Disease Amazons (44) Macaws (43) Ecletto (1) Conures (2) Cockatoos (18) Cockatiels (3) African Grays (39) Total
Air sac infection (4) 0 2 0 0 0 0 2 4
Bacterial infection (13) 5 2 0 0 2 1 3 13
Chlamydiosis (22) 7 14 0 0 0 0 1 22
Fungal infection (3) 0 1 0 0 0 0 2 3
Viral infection (14) 2 8 0 0 2 0 2 14
Organ-related disease
Enteritis (5) 1 2 0 0 0 0 2 5
Cloacal disease (3) 1 1 0 0 0 0 1 3
Liver disease (12) 10 1 0 0 1 0 0 12
Renal disease (3) 2 0 0 0 0 0 1 3
Hearth disease (3) 1 0 0 0 0 0 2 3
Neurologic disorders (2) 0 0 0 0 0 0 2 2
Osteomalacia (2) 0 0 0 0 0 0 2 2

Overview of Psittacine Blood Analysis

Reproductive disorders (3) 1 2 0 0 0 0 0 3
Miscellaneous diseases
Feather and beak disorders (8) 1 1 0 0 4 0 2 8
Behavioral disorders (8) 0 3 0 0 1 2 2 8
Dietary disorders (11) 2 0 0 1 2 0 6 11
Gizzard foreign bodies (6) 1 0 0 1 2 0 2 6
Zinc toxicosis (4) 0 2 1 0 0 0 1 4
Pediatric pathologies (4) 0 2 0 0 2 0 0 4
Check (17)a 9 2 0 0 2 0 4 17
Total (150) 44 43 1 2 18 3 39 150
a
The group named check represents data that were obtained in psittacines (for the most part from breeding centers or pet shops) at high risk of disease but still
without symptoms.

105
106 Capitelli & Crosta

Table 13
Total absolute number and percentage for tests with normal, decreased, and increased values

Decreased Increased
Test Total Value Value n % Decreased % Increased % Abnormal
Hematocrit 134 29 28 77 22 21 43
Total protein 125 9 30 86 7 24 31
Hemoglobin 115 16 57 42 14 50 64
RBC 115 27 21 67 24 18 42
WBC 150 30 67 53 20 45 65
Lymphocytes (%) 150 38 27 85 25 18 43
Monocytes (%) 150 0 67 83 0 45 45
Heterophils (%) 150 31 56 63 21 37 58
Eosinophils (%) 150 0 3 147 0 2 2
Basophils (%) 150 0 0 150 0 0 0
Anisocytosis/ 150 0 13 137 0 9 9
poikilocytosis
Polychromasia 150 0 24 126 0 16 16
Band heterophils 150 0 23 127 0 15 15
Toxic heterophils 20 0 21 129 0 14 14
Zinc 31 0 21 10 0 67 67
Calcium 68 3 19 46 4 28 32
Ionized calcium 37 8 8 21 22 22 44
Phosphorus 51 10 15 26 20 29 49
Sodium 51 4 21 26 8 41 49
Potassium 55 20 15 20 36 28 64
Bile acids 74 12 26 36 16 36 52
Glucose 128 43 2 83 34 1 35
Uric acid 130 31 12 87 24 9 33
Amylase 41 6 20 15 15 49 64
Creatinine 41 3 7 31 7 17 24
CPK 107 9 50 48 8 47 55
Cholesterol 39 1 16 22 3 41 44
LDH 103 18 54 31 17 52 69
AST 129 18 32 79 14 25 39
Prealbumin 75 27 2 46 36 3 39
Albumin 75 17 3 55 23 4 27
a1-Globulin 74 9 14 51 12 19 31
a2-Globulin 74 9 27 38 12 36 48
b-Globulin 74 7 25 42 10 34 44
g-Globulin 74 29 12 33 39 16 55
A/G ratio 75 30 2 43 40 3 43

globulin fractions were all increased. In another case, we observed a decrease in

g-globulin. This can be explained by the fact that in bacterial infections, the changes
are most evident in the acute phases, whereas the changes tend to be moderate in the
chronic phases.72,75 Moreover, the continued stress of some chronic forms can lead
to a decrease in the g-globulin fractions. According to literature, there have been the
 Overview of Psittacine Blood Analysis 107

greatest number of abnormalities in chlamydiosis cases occurring in more parame-

ters.78 Even in this case the complete blood cell counts have given useful indications.
In 5 cases the hematocrit was higher and in 4 cases hyperproteinemia was present.
This may indicate dehydration and/or inflammation. No cases of regenerative or
depressive anemia have been observed although they are reported in literature.1,10,78
More severe leukocytosis than in bacterial diseases has been observed with lympho-
cytosis and monocytosis. It is reported that in Amazons the classic chlamydial leuko-
gram shows marked leukocytosis with monocytosis; lymphocytosis with reactive
lymphocytes may be seen. In macaws, leukocytosis can be important and in acute
case can exceeded 100,000/mm3.78 Leukopenia with marked heteropenia was also
observed. The high-grade toxic heterophils were not always present, but when
present, they confirm the severity of the disease, especially in ocular and upper respi-
ratory tract, chronic forms or in sepsis. Major changes in biochemical parameters were
also seen in this disease. Uric acid and glucose plasma levels seem to be markedly
decreased in half of the cases and AST and CPK plasma activity markedly increased.
Increased levels of bile acids in plasma were less frequent. Seeing such a large variety
of abnormalities should induce the clinician to widen the investigation to confirm or
exclude chlamydiosis. Even the electrophoretic pattern is often altered with peaks
in a- and b-globulins (acute phase proteins and acute inflammatory process), hyper-
proteinemia, and decreased A/G ratio. In some cases, the electrophoresis test helps
in the interpretation of serologic antibody tests for the diagnosis of this disease,
because the globulin fractions may increase before the antibody response does.75
In fungal diseases, (only 3 cases) we have found 2 cases of heterophilic leukocytosis
with toxic heterophils and 1 case of degenerative heteropenia. These diseases affect
the respiratory system and they are often nonresponsive to treatment and have a poor
prognosis. The only abnormality concerning biochemical parameters was the marked
decrease of ionized calcium, which could be due to alkalosis,64 artifactual errors, or to
the many causes of organ-related loss.
In cases of viral disease, we have seen some increases in PCV together with
increased proteins (severe dehydration). There were more cases of leukocytosis
with lymphocytosis and monocytosis or heterophilia (antibody stimulation and/or viral
infection complicated by other diseases), but there were also cases of leukopenia with
heteropenia and monocytosis and/or lymphopenia with monocytosis (as reported in
the literature on viral diseases).1,8–10,47 In this case, we have observed hypoglycemia
and marked increases in both muscular and hepatic enzyme activities. These
increases may be caused by neurologic disorders or vomiting (muscle enzymes) or
concomitant liver disease (AST). Alterations in potassium, phosphorus, and sodium
could be explained by the involvement of the kidneys or gastrointestinal tract. These
data are not specific, but might be useful to guide nutritional support and therapy.
Furthermore, we have observed 4 cases showing increased b; and g-globulin frac-
tions, but rare increases in a-globulin fractions, and no hypoalbuminemia and
decreases in the A/G ratio. These results can be related to acute-subacute forms in
the presence of an immune response. In enteritis, we found only rare outliers repre-
sented by the frequent increase in LDH activity (LDH activity is a nonspecific test
because it is present in all organs) and a decrease in glucose and uric acid levels (pro-
longed fasting). In cloacal diseases (even though there were few cases), we observed
alterations caused by chronic damage (mostly papillomatosis with chronic prolapses).
Leukocytosis with moderate heterophilia and marked monocytosis were registered
and severe leucopenia and heteropenia was seen only in 1 case (cloacal calculi). In
the electrophoretic pattern, the authors noted a decreased A/G ratio with increased
b- or a1-globulin fractions (acute phase inflammatory proteins). Occasional increases
 108
Capitelli & Crosta
Table 14
Hematologic and biochemical data grouped by infectious disease

Air Sac Infection (4) Bacterial Infection (13) Chlamydiosis (22) Funfal Infection (3) Viral Infection (14)
L1 L2 L3 D1 D2 D3 L1 L2 L3 D1 D2 D3 L1 L2 L3 D1 D2 D3 L1 L2 L3 D1 D2 D3 L1 L2 L3 D1 D2 D3
Hematocrit 1 1 4 1 2 5 1 1 1 4
TRBC — 1 4 1 2 2 1 3 1
TWBC 1 2 2 3 3 3 4 6 1 6 1 2 3 5 1 1
Hemoglobin — 1 3 1 1 3 9 1 1 6
Lymphocytes 1 5 3 1 1 2 2 1 1 3 2
Monocytes 3 3 2 1 4 6 1 3
Heterophils 1 1 6 6 3 5 1 2 3 3 2
Eosinophils 1 — — — —
Amylase — 3 1 2 1 2 1
AST 2 2 4 4 2 2 2 1 3 2 1
Bile acid 1 1 1 6 1 — 1 1
Calcium — 1 7 1 2 — —
Cholesterol — 1 1 1 — 1
CPK 1 1 1 4 2 2 3 2 1 1 1 6
 Creatinine — 1 1 — —
Ionized calcium — 1 1 2 —
Glucose 2 2 2 3 3 2 3 2 1
LDH 1 1 1 4 3 2 — 1 4 2
Phosphorus — 1 1 3 2 1 1 2
Potassium 1 4 1 1 2 1 1 2
Sodium — — 2 1 — 3
Total protein — 2 1 2 2 1 1 3 2
Uric acid 1 1 1 1 3 1 3 1 1 1 1 1
Zinc 1 — 2 1 — —
Prealbumin 1 1 1 1 1 1 1
Albumin 1 1 2 2 1 1
a1-Globulin 1 2 1 2 2 — 1 1
a2-Globulin 1 1 1 1 2 2 2 1 1
b-Globulin 1 1 1 3 4 1 1 2 1
g-Globulin 1 2 1 3 3 1 — 2 2

Overview of Psittacine Blood Analysis

A/G ratio 1 1 2 2 5 1 1 1 2

Score: 1, low decrease; 2, mild decrease; 3, severe decrease; 11, low increase; 12, mild increase; 13, severe increase.

109
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Capitelli & Crosta
Table 15
Hematologic and biochemical data grouped by organ-related disease
Reproductive
Enteritis (3) Cloacal Disease (3) Liver Disease (12) Heart Disease (3) Renal Disease (3) Disorders (3)
L1 L2 L3 D1 D2 D3 L1 L2 L3 D1 D2 D3 L1 L2 L3 D1 D2 D3 L1 L2 L3 D1 D2 D3 L1 L2 L3 D1 D2 D3 L1 L2 L3 D1 D2 D3
Hematocrit 1 1 — 3 1 3 1 1 — 1
TRBC 1 1 3 1 — 1 1 1
TWBC — 1 2 3 3 1 2 1 2
Hemoglobin 1 1 6 1 1 2 3
Lymphocytes 1 2 1 1 2 1 2
Monocytes 2 1 1 2 2 3 3 1
Heterophils 1 1 1 2 3 — 2 1 1
Eosinophils — — 1 — — —
Amylase — — 1 2 — 1 1
AST 2 1 3 2 — — 1
Bile acid — — 1 3 3 1 — —
Calcium 1 1 1 1 1 —
Cholesterol — — 1 — 1 —
CPK 1 1 1 2 2 1 1
 Creatinine — — 1 2 1 1 —
Ionized — — 2 1 1 1 —
calcium
Glucose 2 1 1 1 1 — 1 —
LDH 1 1 3 1 2 2 2 2 1 1 1 —
Phosphorus 1 — 1 2 1 1 1 1
Potassium — — 1 1 2 1 3 2
Sodium — — 1 — 3 2
Total protein 1 1 3 1 — 1 1 1
Uric acid 2 1 — 1 2 3 1 2 1 1 1
Zinc 1 1 2 1 — —
Prealbumin — — — — — —
Albumin — — 1 1 — 1 —
a1-Globulin — — 1 1 — —
a2-Globulin — 1 1 2 1 1 1 1
b-Globulin — 1 1 2 2 1 — — 2
g-Globulin — — 1 2 1 — —

Overview of Psittacine Blood Analysis

A/G ratio — 2 2 1 2 — — —

Score: 1, low decreased; 2, mild decreased; 3, severe decreased; 11, low increased; 12, mild increased; 13, severe increased.

111
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Capitelli & Crosta
Table 16
Hematologic and biochemical data grouped by miscellaneous disease
Feather and Beak Behavioral Dietary Gizzard Foreign Pediatric
Disorders (8) Disorders (8) Disorders (8) Bodies (6) Zinc Toxicosis (4) Pathologies (4)
L1 L2 L3 D1 D2 D3 L1 L2 L3 D1 D2 D3 L1 L2 L3 D1 D2 D3 L1 L2 L3 D1 D2 D3 L1 L2 L3 D1 D2 D3 L1 L2 L3 D1 D2 D3
Hematocrit 1 2 1 3 2 1 2 1 2
TRBC 1 3 3 1 3 1 — —
TWBC 3 1 5 3 1 5 1 1 3 1 4
Hemoglobin 1 4 3 2 2 3 1 1
Lymphocytes 2 2 2 1 3 3 3 1 2 1 1
Monocytes 3 1 3 1 3 2 2 2 2
Heterophils 1 1 1 1 1 1 4 2 1 5 1 3 2 1 1 1 1
Eosinophils — — 1 — — —
Amylase — 1 — — 1 2
AST 2 4 3 1 1 1 —
Bile acid 2 1 1 1 1 — — —
Calcium 1 — 1 — — —
Cholesterol 1 — 3 — — 1
CPK 3 1 3 3 3 1 1
Creatinine — — — — 1 —
 Ionized — — — — — —
calcium
Glucose 1 — — 1 2 4
LDH 1 1 1 1 1 1 5 1 2 1 — 2
Phosphorus — — — — 1 1
Potassium 2 1 1 — — 1 1
Sodium — — — — 3 4
Total protein 1 1 3 1 2 — 3
Uric acid 2 1 1 1 1 1 3 1 1 1 —
Zinc 2 1 3 2 1 3 —
Prealbumin 1 — 3 1 1 1 —
Albumin 1 — 1 1 — —
a1-Globulin — — 1 1 1 1 — 2
a2-Globulin — — 1 3 2 — 2 2
b-Globulin 1 — 1 1 1 1
g-Globulin 1 1 — — 2 1 1 1 1
A/G ratio 1 1 — 4 1 1 1 1

Overview of Psittacine Blood Analysis

Score: 1, low decreased; 2, mild decreased; 3, severe decreased; 11, low increased; 12, mild increased; 13, severe increased.

113
114 Capitelli & Crosta

in enzyme muscle activities and glucose could depend on chronic stress and/or on the
muscular effort for defecation.
Liver disease may not always have an inflammatory cause and therefore may not
induce hematologic changes. We observed both anemia and false polycythemia
(dehydration). Moderate heterophilia (25%) and marked leukopenia and heteropenia
(25%) were not constant findings, whereas we always found relative monocytosis.
Marked leukopenia and heteropenia may indicate a poor prognosis and may occur
in viral diseases (eg, Pacheco parrot disease).47 The most common alteration was
increased bile plasma levels (50%); this value cannot differentiate the kind of liver
disease but it always means hepatic damage.9 These increases are not necessarily
accompanied by increases in AST activities.75 They can increase in cases of multior-
gan lesions, such as in chlamydiosis or in bacteremia and septicemia, but sometimes
they do not increase in liver disease as shown here. We have reported 2 cases of
reduction of these enzyme activities as reported in the case of hepatic mass reduc-
tion.4 The sporadic increases of other enzymes were certainly nonspecific. Plasma
levels of uric acid (as expected) were found increased, decreased, and in most cases
normal; this metabolite in the blood is the sum of secretion by the liver and renal elim-
ination. Electrophoresis can help in the diagnosis of hepatitis and in monitoring this
disease. As reported in literature, not all increases in all globulin fractions have
been highlighted, and the highest incidence was in the increase of a2-globulin and
b-globulin, with a consequent decrease in the A/G ratio.75 Two cases of increase in
the A/G ratio due to a decrease in the b- and g-globulin fractions (possible virus
infection or analytical error for an altered migration of the bands) should also be
noted.16,51,75
In kidney disease, we observed an increase in those parameters traditionally linked
to kidney disorders (creatinine and phosphorus), although these could be possible
artifactual increases in both tests.4,5,49,79 The decreases in electrolytes, such as K,
Na and phosphorus, may depend on the marked polydipsia and polyuria. All these
parameters do not allow a diagnosis of kidney disease and certainly the most useful
test could be the electrophoresis.75 In our study, we noted a single case of increased
a1-globulin fractions without other changes.
In heart disease, we only observed increases in muscle enzyme activities (CPK and
LDH), always relative to monocytosis and marked reduction of plasma levels of uric
acid. These data show that aspecific muscle damage is possibly related or not related
to this disease.79
In neurologic disorders and osteomalacia (not shown in Table 15 because of the
low number of cases), the psittacine species most affected were African gray parrots
and there was a marked decrease both in total calcium and ionized calcium plasma
levels and a marked increase in muscle enzyme activities. This confirms what is
reported in the literature, that an assessment of ionized calcium is useful for
confirming hypocalcemia, which, in this case, was also responsible for neurologic
disorders.16,64
Regarding reproductive disorders, there are no significant alterations. We have re-
ported only 1 marked increase in plasma levels of potassium and uric acid in the
absence of any marked abnormalities. It is possible that in reproductive phases or
reproductive disorders, lipemic plasma arose4,5,53 creating an analytical error.
In the third group (miscellaneous diseases as in Box 1), we found some typical
parrot diseases such as feather and beak disorders. In these conditions, the TWBC
counts showed moderate decreases or increases and the most variable component
was lymphocytes followed by heterophils and monocytes. A possible explanation
for lymphocytosis may be a continuous immune stimulus or an immune-mediated
 Overview of Psittacine Blood Analysis 115

form, whereas lymphopenia may be caused by a chronic stressor (prolonged sexual

feather picking or severe feather picking) or viruses (PBFD).1,8–10,64 Apart from an
increase in CPK activity (caused by forced exercise or excitement), we did not observe
anything else.
In behavioral disorders, which can be a cause of feather picking or at least be linked
to this, no significant abnormalities were registered, besides 1 case where there was
leukocytosis with lymphocytosis and others presenting stress leukograms. Monocyto-
sis was always present. There were also increases in CPK plasma enzyme activities,
which can be due to excess exercise. So, being cautious and because the issue is
manifold and still misunderstood, we could observe some small differences in the leu-
kogram (lymphocytosis or lymphopenia vs stress leukogram) between feather
destructive behaviors and pure behavioral disorders.
In the hematologic and biochemical values of dietary disorders, there were a high
number of cases of moderated leukopenia with lymphopenia or heteropenia and
only 1 case of moderated leukocytosis with lymphocytosis and heterophilia. This
occurrence may not even have any diagnostic usefulness. We have also noted
moderate increases in bile acids and an increase in cholesterol in 3 cases, which so
often appears only in this disease. High cholesterol levels may be observed in meta-
bolic and liver disease and are frequently related to dietary disorders.51,60,79 We also
reported increases in a1- and a2-globulin fractions with decreases in the A/G ratio in
35% of cases. This occurrence could correlate with the increase in cholesterol,
possible liver disease, or with underlying hormonal conditions such as
hypothyroidism.
In gizzard foreign bodies, we often found leukocytosis with heterophilia and
monocytosis and lymphopenia. The absence of banded heterophils suggests that
they are stress leukograms and monocytosis can be caused by a penetrating injury
and irritation of the tissues. Other abnormal data found were decreased plasma
levels of uric acid and a case of hypoglycemia, both probably connected to pro-
longed starving and/or absorption disorders. There were marked increases in
muscle enzymes (with exclusion of AST) caused by possible perforations through
the muscle layer or excessive efforts related to dysphagia and vomiting. The finding
of increased of plasma levels of zinc, often feared in gizzard metallic foreign bodies,
does not show differences with increased values found in other diseases. As re-
ported by Fudge,79 the identification of a metal foreign body in the gastrointestinal
tract is not specific for zinc or heavy metal toxicosis. Evaluation of plasma levels of
zinc, if not markedly increased and accompanied by typical symptoms, can be
misleading and in this case must be assessed with care, also considering the
different species and their feeding habits; for example, cockatiels, typical seed
eaters, normally show lower levels of zinc than Amazons.80 In the 4 cases reported
as suspected zinc poisoning, abnormal hematologic and blood chemistry values
were absent or nonspecific, such as moderate heterophilia as reported in the liter-
ature.80 In these cases, the zinc value exceeded 400 mg/dL and the animals showed
symptoms of zinc poisoning. Some species such as cockatoos and the Eclectus
parrots already mentioned have higher plasma levels of zinc even in healthy birds.76
In 3 cases that do not belong to this group, we found nonspecific increases in
plasma levels of zinc, even though the birds did not have symptoms of zinc toxi-
cosis; in 2 of these cases, the diagnosis was feather picking in accordance with
what it is reported.80
With regard to pediatric diseases, all patients showed heterophilia and 1 bird had
marked lymphocytosis. These data should be critically evaluated and considered
pathologic when marked and in young healthy birds, there can also be a higher total
116 Capitelli & Crosta

white cell count with heterophilia. Lymphocytosis is usually rare in young animals but
can be caused by stimulation of the immune system. The total proteins were also
increased. This, however, can indicate inflammation or dehydration because normally
young birds have lower values. In the 2 cases, a high PCV can suggest dehydration,
whereas an increase in a1- and a2-globulin in the same patient could suggest
a problem of acute inflammation. The 2 cases of trauma did not give any indications,
but excluded the presence of severe bleeding and needed more controls. The case of
neoplasm, even though unique, showed a marked increase in a2- and b-globulin frac-
tions and a marked decrease in the A/G ratio. In this case, the animal had a neoplasm
but also a serious bacterial infection with a granulomatous process; furthermore, we
believe this had been ongoing for a long time. This bird also had marked leukocytosis
and lymphomonocytosis as a response to a chronic bacterial process and malignant
neoplasm. The other values were not specific to add useful information for the diag-
nosis and prognosis. Clearly in this case, as in others, the final diagnosis was histo-
logic for the neoplasm and microbiologic for the bacterial infection. In this article,
the results of histology, serology (antibody test), microbiology for bacterial and fungal
diseases, and PCR tests, are not fully described although these, along with other diag-
nostic tests, have often allowed a diagnosis to be confirmed that would not otherwise
be made.
In the cases in the check group, the abnormal values are not specific, for example,
moderated leukopenia with heteropenia (23%) and moderated leukocytosis with het-
erophilia and/or lymphocytosis (23%). The other abnormalities that can be highlighted
are nonspecific moderate decreases in many parameters and moderate increases in
plasma levels of LDH. These results could be artifactual decreases, because most of
these blood samples were collected from birds located in other premises and conse-
quentially were processed 12 hours after collection. For this reason, these data should
be critically evaluated and eventually rechecked to clarify the underlying pathologies.
The laboratory must remember that it is not the blood sample that is sick, but the
patient, and the veterinarian needs to understand that sure and universal analytical
information cannot be obtained from a single sample of blood, maybe even badly
taken.

ACKNOWLEDGMENTS

We are grateful to the staff of the CSV laboratory for their assistance in testing the
psittacine blood samples and particularly to Silvia Lubelli and Simona Eterno as well as
the clinical pathologist Gabriele Ghisleni for improving this article. We are extremely
grateful to Enrico Parravicini for his contribution in writing the text.

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