Atlas of Canine and Feline Peripheral Blood Smears

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ATLAS OF
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CANINE AND FELINE

PERIPHERAL BLOOD
SMEARS
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ATLAS OF
CANINE AND FELINE
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PERIPHERAL BLOOD
SMEARS
First Edition
Amy C. Valenciano, DVM, MS, DACVP

Veterinary Clinical Pathologist
IDEXX Reference Laboratories, Inc.
Dallas, Texas
Rick L. Cowell, DVM, MS, MRCVS, DACVP
Veterinary Clinical Pathologist
IDEXX Laboratories, Inc.
Stillwater, Oklahoma
Theresa E. Rizzi, DVM, DACVP
Clinical Associate Professor
Department of Veterinary Pathobiology
Center for Veterinary Health Sciences
Oklahoma State University
Ronald D. Tyler, DVM, PhD, DACVP (Clinical and Anatomic
Pathology), DABT
Adjunct Professor
Department of Veterinary Pathobiology
Center for Veterinary Health Sciences
Oklahoma State University
&
Tamil Nadu Veterinary and Animal Sciences University
Madras Veterinary College
Vepery, Chennai, India
Consulting Editor:
Dennis B. DeNicola, DVM, PhD, DACVP
Clinical Pathologist
Chief Veterinary Educator
IDEXX Laboratories, Inc.
Westbrook, Maine
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3251 Riverport Lane

St. Louis, MO 63043
ATLAS OF CANINE AND FELINE PERIPHERAL BLOOD SMEARS

Edition 1
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Copyright © 2014 by Mosby, an imprint of Elsevier Inc.
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The Publisher
Library of Congress Cataloging-in-Publication Data
Valenciano, Amy C., author.

Atlas of canine and feline peripheral blood smears / Amy C. Valenciano, Rick L. Cowell,
Theresa E. Rizzi, Ronald D. Tyler.
p. ; cm.
ISBN 978-0-323-04468-4 (alk. paper)
I. Cowell, Rick L., author. II. Rizzi, Theresa E., author. III. Tyler, Ronald D., author. IV. Title.
[DNLM: 1. Clinical Laboratory Techniques—Atlases. 2. Blood Cells—pathology—Atlases.
3. Blood Cells—ultrastructure—Atlases. 4. Cat Diseases—blood—Atlases. 5. Dog Diseases—
blood—Atlases. SF 772.67]
SF769.5
636.089’607561—dc23
2013012786
Vice President and Publisher: Linda Duncan

Content Strategy Director: Penny Rudolph
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Printed in China
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PREFACE
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The Hematology Atlas of Canine and Feline evaluation, and serve as valued reference for
Peripheral Blood Smears is created to be a “scope- anyone interested in hematopathology.
side” visual atlas of normal and abnormal findings
on dog and cat peripheral blood smears. Our aim
ACKNOWLEDGMENTS
is to provide veterinary students, veterinary tech-
nicians, practicing veterinarians, and veterinary We thank our families for their support and under-
residents with a large number of logically orga- standing. Many other people deserve acknowl-
nized, high quality photomicrographs that are edgement and sincere thanks also. These include
useful in learning to identify normal and abnormal Elsevier’s excellent editors and staff, and the many
peripheral blood findings and enjoyable to view. veterinary pathologists at IDEXX laboratories and
Each entity is fully characterized by multiple pho- other colleagues who sent slides or pictures for use
tomicrographs, taken at varying magnifications in the text, especially: Drs. Debbie Bernreuter,
and encompassing variations in morphological Dean Cornwell, Jim Mathews, Desire Lipscomb,
features. Additionally, there are several special Liz Little, Jennifer Neel, Natalie Courtman, Carrie
mosaic photomicrographs constructed of similar- Flint, Dennis DeNicola, Shanon Zablotsky, and
appearing and often confused elements that Kari Velguth and Ms. Joan Shewmaker. We espe-
enables careful comparison for quick microscopic cially appreciate and recognize the editing work of
differentiation. The concise text provides a quick Drs. Dave Fisher and Pat McManus who served as
and detailed description of each microscopic additional reviewers for this atlas.
finding being emphasized to compliment the cor- Finally, we would like to thank IDEXX Labora-
responding photomicrographs. tories for generously supporting veterinary educa-
Emphasized are normal cellular findings and tion and this book in particular.
common abnormal findings, in particular, RBC
Amy C. Valenciano
and WBC morphology changes, hemoparasites,
Rick L. Cowell
and infectious agents. A section on leukemia is pro-
Theresa E. Rizzi
vided to offer an overview of the different types
Ronald D. Tyler
of leukemias in dogs and cats, with an emphasis
on the challenges of differentiating between the Supported in part by IDEXX Laboratories as part
different cellular origins of leukemia based solely of their commitment to Veterinarians, Veterinary
on light microscopy and thus the need for special- Technicians, and Veterinary Medical Education.
ist review and further diagnostics. The authors
sincerely hope this atlas will provide a means
to develop a strong foundation in blood smear
v
To Dr. Rick Cowell, an inspiration and excellent pathologist and mentor. Thank you
for sharing your projects, insights, and laughter. It was an honor to work alongside
you in creating this wonderful atlas. I dedicate my efforts to God and my family:
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Daniel (husband), Avery (daughter), Ty (son), Bonny (twin) and my dear parents.
I thank my wonderful mentors especially Drs. Dave Fisher, Sonjia Shelly,
Carol Grindem, Jan Andrews, Mary Jo Burkhard, Gregg Dean, Christine Stanton,
and Lon Rich. I also thank IDEXX laboratories for supporting academic growth
and for promoting excellence in veterinary pathology.
Amy Valenciano
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To my parents who taught me the value of honesty

and instilled in me a work ethic that has served me well through the years.
To my wife (Annette) and daughter (Anne) who have continually given support,
meaning and inspiration to my life.
To my daughter (Rebecca) who showed me the face of true courage, and taught me to
laugh and love even in the worst of times. While she lost her battle with cancer at the
age of 11 her memories and life lessons will forever be remembered.
To the many outstanding veterinary clinical pathologists I have had the
opportunity to learn from especially, Drs. Ronald D. Tyler,
James Meinkoth, and Dennis DeNicola.
To the many veterinary practitioners, residents, and students who taught
me much more than I could ever have hoped to teach them, and have
become colleagues and friends.
To IDEXX Laboratories for their continued support of veterinary education and
especially to Dr. Dean Cornwell for his support and encouragement.
Rick Cowell
To Deb, for your enduring love and support.

To my son, Aiden, who reminds me to laugh often and who always makes me proud.
Theresa E. Rizzi
To my mother who gave me my core tenants of faith, honesty and respect; my

wonderful wife, Reba, for her patience and support; my children who have made my
life truly wonderful, exciting, and worthwhile; and the many exceptional colleagues
who have taught, inspired and guided me—especially Roger Panciera for his
mentorship and Rick Cowell for his many contributions to my development as a
person and pathologist and, most of all, his friendship through the years.
Ronald D. Tyler
INTRODUCTION:
BLOOD SMEAR PREPARATION
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AND EXAMINATION
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Overview
Evaluation of blood smears is a fundamental step in overall health assessment and is included in stan-
dard hematologic profiles and as part of the diagnostic evaluation of virtually every ill patient. In addi-
tion to being a means of obtaining the differential white blood cell (WBC) count (i.e., determination of
the distribution of the different types of leukocytes, both as percentages and counts per microliter [µL]),
blood smear examination may yield a broad range of diagnostic information. For example, altered red
blood cell (RBC) morphology may suggest chronic blood loss, exposure to endogenous and exogenous
toxins, disorders of vasculature, or immune-mediated hemolysis. Changes in WBC morphology may be
the earliest laboratory finding of inflammation and may be diagnostic for certain inherited conditions
and leukemias. Some changes occur in leukocyte maturity, morphology, and type that may only be
detected through microscopic evaluation of peripheral blood smears (i.e., neutrophilic left-shift, neutro-
philic toxic change, and the presence of circulating mast cells). In some cases, infectious agents, pathog-
nomonic cellular inclusions, and neoplastic cells are observed on blood films, yielding an immediate,
definitive diagnosis. In addition, monitoring changes found in peripheral blood may help determine a
patient’s treatment, response to therapy, and short-term and long-term prognoses.
Blood smear evaluation should be performed as part of every complete blood count (CBC), either
in-house or in an outside laboratory. Evaluation provides morphologic confirmation of hematologic
parameters, assurance of the quality of values obtained from automated analyzers, and additional impor-
tant information not given by automated methods. The value of blood smear review is maximized when
the information is correlated with the patient’s medical history, current and previous laboratory findings,
and physical examination findings.
Blood smear preparation is easy and inexpensive, and experience in evaluation is readily acquired
with adequate background information and regular practice. This introduction describes applicable
techniques and interpretations of canine and feline blood smears. A brief introduction to the integration
of findings from blood smear evaluations with other values in the CBC is included.
Equipment and Supplies

An essential piece of equipment for blood smear evaluation is a well-maintained, binocular microscope
with high-quality 10, 20, 40 or 50 (ideally, oil-immersion 50×), and 100× (oil-immersion) objectives. Addi-
tionally, clean glass slides and good-quality, fresh stains and Coplin jars are needed for in-house
staining.
Standard plain or frosted glass slides may be used directly from the package without special treatment
or cleaning. Slides with frosted ends facilitate labeling with patient information. Slide surfaces should
be free of dust, fingerprints, and residue from detergent, alcohol, or tap water. The use of special cytologic
adhesives may result in background staining and are not recommended. An ample supply of slides
facilitates preparation of multiple smears per sample, avoiding the frustration of interpreting any poorly
made smears. These additional smears may also be reserved for alternative types of staining or, if indi-
cated, a specialist’s review.
Sample Collection
Ideally, blood samples should be collected on the first attempt from a medium to large vein of a calm
patient. Ethylenediaminetetraacetic acid (EDTA) is the preferred anticoagulant for blood used in cyto-
logic preparations. The liquid form of EDTA disperses more rapidly in samples and may be preferable
to the powder form, particularly for feline blood, but both forms of EDTA preserve general cellular
morphology in refrigerated samples for up to 4 hours. In either case, adequate mixing is essential and
x
INTRODUCTION xi
is accomplished by gently inverting the tube 6 to 10 times. Alternatively, blood without an anticoagulant
may be placed directly from the collection needle onto the slide. This is preferred by some because of
possible morphologic alterations caused by the EDTA and is particularly useful if sample volume is
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limited. Samples collected from superficial skin-puncture wounds, clipped toenails (due to excessive
contamination with tissue procoagulants), and blood anticoagulated with heparin (a relatively poor
preservative of cellular morphology and staining characteristics) are less acceptable. Blood anticoagu-
lated with citrate may be used to evaluate cell morphology on blood smears; however, the required 10%
sample dilution interferes with cell count estimates.
Collection of blood in proper proportion to anticoagulant is facilitated by the use of commercial vacu-
tainers. This is essential to avoid certain artifacts of cell morphology and also to achieve accurate cell
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counts, which may be affected by overfilling and underfilling the tube.
Smear Preparation
The goal of blood smear preparation is to spread the cells (RBC, WBC, and platelets), so a large,
uniform, single layer (monolayer) of cells is created. In the monolayer, RBC should be side by side, yet
infrequently touching, and should be devoid of large spaces without cells. In the monolayer, WBCs
flatten and stain well, allowing good evaluation of cytoplasmic and nuclear detail. WBCs will be
evenly distributed, allowing accurate differential counts to be performed and promoting examination
of WBC, RBC, and platelet morphologies. Inclusions such as rickettsial and viral inclusions may be
readily identified. In thick areas of the smear, cells often are too contracted, distorted, and poorly
stained for reliable evaluation. In excessively thin areas of the smear and in the feathered edge, cells are
often distorted, damaged, and ruptured, prohibiting reliable classification and morphologic evaluation.
Also, a differential WBC count performed near or in the feathered edge results in skewing toward the
larger cell types.
Well-made smears are essential for reliable identification, quantification, and evaluation of periph-
eral blood cells. Smears may be prepared on glass slides or coverslips. The glass slide technique is
generally easier and more reliable than preparing smears on coverslips. Glass slides may also be pro-
cessed through automatic stainers and are most suitable for laboratories using such equipment.
However, the coverslip method generally results in more uniform WBC distribution and less trauma to
fragile blood components such as large or neoplastic cells. For both methods, blood samples should be
fresh and adequately mixed just prior to smear preparation, and smears should be completely air-dried
before staining.
The blood smear preparation technique should be learned well, as it is necessary for reliable in-house
blood smear examination and hemogram results from samples submitted to referral laboratories. At least
one freshly prepared (in-house) blood smear should accompany all EDTA-anticoagulated blood samples
submitted to referral laboratories for CBC. Failure to submit a premade blood smear may result in several
artifacts; for example, blood parasites (e.g., Mycoplasma hemofelis) may fall off the RBC surfaces, resulting
in false-negative results; platelet numbers will decrease and may cause platelet estimates to be falsely
decreased; and WBCs become pyknotic (secondary to aging), and when 10% or more of the WBCs are
pyknotic, the differential count is invalid.
Smears are prepared on glass slides by placing a drop of blood (2 to 3 mm in diameter) on the broad
face of the slide about 1 to 1.5 cm from the frosted border (or edge of a nonfrosted slide). Another clean,
dry slide (spreader slide) is held loosely against the surface of the first slide at a 30-degree angle
and drawn smoothly toward the blood drop. The spreader slide should be brought to a position
where it just meets, but is not drawn into, the blood drop. When the spreader slide makes contact with
the blood, capillary action immediately distributes the blood between the two slides. Then, with no
downward pressure, the spreader slide is quickly and smoothly swept across the remaining length of
the underlying slide.
Ideally, blood smears have a smooth transition from the thick region to the feathered edge and cover
an area one half the length and slightly less than the width of the slide. If the edge of the blood film is
blunt instead of feathered, the second slide was probably raised off the first before the blood was spread
completely. Unequal smear thickness usually results from the spreader slide being held at too obtuse an
angle or placing too much pressure on the first slide while spreading the blood. Too much pressure on
the second slide may also result in WBCs clumping along the smear’s feathered edge. Too little pressure
may result in short, thick smears. Smear thickness may also be affected by the viscosity of the blood
sample. Adjusting the angle at which the second slide is held against the first may help compensate for
very viscous (hemoconcentrated) or thin (anemic) blood samples. A more obtuse (40- to 45-degree) angle
between the two slides makes thicker smears for very anemic blood, and an angle less than 30 degrees
xii INTRODUCTION
may be necessary for preparing smears of severely hemoconcentrated blood. Smears need to be thor-
oughly air-dried before staining; those that are thick may require additional drying time (and additional
time in fixative and stain).
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Common problems in blood smear technique include unclean slides, placing too large a drop of blood
on the blood smear slide, pulling the prep slide too far back, excessive pressure, slow movement of the
prep slide forward, and jerky or uneven forward movement of the prep slide.
Unclean microscope slides may cause spaces devoid of cells to be scattered throughout the smear,
prohibit smooth movement of the prep slide, and impair the staining of portions or the entire smear.
Using a drop of blood that is too large causes the smear to be too thick and to extend too far down
the blood smear slide. Often, the blood smear extends to the end of the slide without formation of a
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monolayer, precluding reliable microscopic examination.

Pulling the prep slide too far back causes blood to flow underneath the prep slide. When the prep
slide is moved forward, plasma and smaller cells (i.e., platelets, erythrocytes, and small lymphocytes)
pass back under the prep slide more easily than do larger cells (i.e., neutrophils and monocytes). As a
result, WBCs are unevenly distributed with small lymphocytes being in excess in the earlier formed
portion of the smear and neutrophils and monocytes being in excess in the later formed portion of the
smear (i.e., feathered edge). Obviously, these smears may give misleading differential WBC counts.
Excessive pressure on the prep slide causes uneven WBC spreading, where most of the WBCs become
concentrated at the feathered edge. Also, with extreme pressure it may be difficult to rapidly and
smoothly move the prep slide forward. Thus, excessive pressure may lead to irregular smears and
uneven distribution of WBC types, resulting in inaccurate differential WBC counts.
Stains
As with cytologic preparations, Romanowsky-type stains (Wright; Wright-Giemsa) are good general
stains for microscopic blood smear evaluation. Quick Romanowsky-type stains (Diff-Quik) are advan-
tageous because they are less sensitive to solution pH and staining time and less susceptible to precip-
itate formation compared with Wright stains. However, most quick stains are also less effective at
demonstrating polychromasia of immature erythrocytes, granules in some cells (i.e., granular lympho-
cytes and mast cells), and toxic change in neutrophils. Thus, whenever possible, Wright stains are
preferred.
Different brands of Wright and Wright-Giemsa stains are available, and following the manufacturer’s
staining protocol for each type is advised. In general, staining is achieved by first dipping the air-dried
slides into a methanol-containing Coplin jar for fixation (approximately 30 seconds). After fixation, the
slides are dipped into Coplin jars containing the stain reagent. Some manufacturers have one type
of stain reagent, others have two. After staining, the slides are dipped into Coplin jars containing
buffer and then rinsed with water (tap, deionized, or a combination of tap and deionized water). The
slides may be dried by blotting with bibulous paper or placed upright on an absorptive surface
to hasten drying. A blow dryer set on low power and held 8 to 10 inches from the slide also shortens
drying time.
Quick stain methods vary somewhat and should be used according to manufacturer’s recommenda-
tions. Diff-Quik requires three separate solutions: (1) an alcohol fixative, (2) a methylene blue dye
mixture, and (3) an eosin-containing solution. The smear is slowly dipped and slightly swished five or
more times into each solution in sequence, with one edge of the slide briefly blotted between solutions.
For each step, the fluid should drain smoothly off the slide between dips, with no bubbling or uneven-
ness, before proceeding to the next solution. Tap or distilled water is used to rinse the slide after the last
solution and before air-drying.
Hematologic Reference Intervals

Reference intervals, which are found in publications and texts, provided by a reference laboratory, or
provided by manufacturers of bench-top analyzers, are generally reliable. Certain physiologic factors
occasionally cause a healthy patient’s hematologic values to deviate from reference intervals. For example,
very young animals are rapidly expanding their vascular space and tend to have relatively low hemato-
crits. Young animals are also actively replacing fetal RBCs with adult RBCs and also have greater eryth-
rocytic anisocytosis and polychromasia and a higher frequency of nucleated RBCs compared with mature
animals. Relatively high lymphocyte counts are also common in young animals, and lymphopenia
is suggested in puppies and kittens less than 6 months of age if lymphocyte counts drop below
2000 cells/µL. Transient elevations above the reference interval for lymphocyte counts are common in
INTRODUCTION 1
excited or vigorously exercised patients, especially if they are immature. This epinephrine-induced
response may also result in temporarily increased counts of other WBC types in the peripheral blood of
healthy patients. At least one canine breed, the Greyhound (and potentially other sight hounds as well),
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has hematologic reference intervals reported to fall slightly outside of reference intervals commonly used
for the species. Although these normal physiologic conditions should be considered, they generally
explain less than 5% of the patient values that fall outside reference intervals for any single hematologic
result.
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SECTION 1:
GENERAL ASSESSMENT
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Blood Smears, 3
Monolayer, 4
Feathered Edge, 6
Body, 8
Background, 10
Increased Basophilia, 10
Cytoplasmic Fragments, 12
Infectious Agents, 14
Stain Precipitation, 16
Fibrin, 18
Skin Contaminants, 20
2
BLOOD SMEARS 3
Blood Smears
DISTINCTIVE FEATURES: Blood smears have three major areas: (1) the thick inner area (body); (2) the
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monolayer; and (3) the feathered edge (the most external area). The inner area is the thickest area of
the smear, and cells are usually too contracted, distorted, or poorly stained for reliable evaluation. The
monolayer is the best area for cell morphology evaluation and differential cell counts, whereas the feath-
ered edge is the best area to search for organisms (i.e., microfilaria), clumped platelets, and large atypical
cells, neoplastic cells, or both. Blood smears should have a smooth transition in thickness from the
proximal end to the distal end of the smear and have an adequate monolayer for evaluation of cell dis-
tribution and morphology. The monolayer is found within the distal half of the smear adjacent to the
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feathered edge and is luminescent when the unstained slide is held under indirect light.
DIAGNOSTIC SIGNIFICANCE: Initially, the entire smear should be quickly scanned using the 10 or 20×
objective from the thickest region to the feathered edge. At this low magnification, blood films may be
checked for staining, overall thickness, smooth transitions in thickness, cell distribution, adequacy of the
monolayer area, and general appearance of the background. The feathered edge may be scanned at low
power for platelet clumps, large parasites and large, atypical nucleated cells.
A patient’s hematocrit may be crudely estimated by examining a blood smear at low magnification.
This estimate assumes that smear thickness was not increased or decreased to compensate for anemia
or hemoconcentration by increasing or decreasing the angle of the spreader slide. Blood films from
nonanemic animals generally have red blood cells (RBCs) that are closely apposed in the monolayer as
well as several RBC layers at the thick end of the smear that obstruct penetrance of most of the condenser
light. In contrast, smears from animals that are moderately to markedly anemic usually have RBCs that
are widely separated from one another in the monolayer and only one to two RBC layers in the thick
end of the smear that allow considerable condenser light to penetrate. In smears prepared from hemo-
concentrated specimens, the monolayer will occupy a relatively smaller zone and cells may be crowded
and difficult to evaluate. Estimates should ultimately be checked against the patient’s measured hema-
tocrit or packed cell volume (PCV).
4 SECTION 1 GENERAL ASSESSMENT
Monolayer
DISTINCTIVE FEATURES: In the monolayer, blood cells are present in a single layer. Cells should be side
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by side, but infrequently touching, and should not be distorted. Also, the monolayer should be devoid
of large spaces without cells.
DIAGNOSTIC SIGNIFICANCE: The monolayer represents the limited region where cell morphology is
most reliably evaluated. White blood cells (WBCs) should be fairly uniformly distributed within this
region. WBC estimates, differential platelet estimates, and examination of WBC, RBC, and platelet mor-
phologies should be done in the monolayer. In the monolayer, WBCs flatten and stain well, allowing
good evaluation of cytoplasmic and nuclear detail. Also, inclusions such as rickettsial and viral inclusions
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may be readily identified.

On a well-made blood smear, the total WBC and differential cell counts may be estimated or simply
classified as low, normal, or high by examining the monolayer of the smear using the 40× or 50× objec-
tive, whereas platelet frequency is best estimated using the 100× objective.
Plate 1-1 Normal Monolayer

BLOOD SMEARS MONOLAYER 5
Plate 1-1 Normal Monolayer (con’t)

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Plate 1-1a
Plate 1-1b
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Plate 1-1d
Plate 1-1e
Plate 1-1f
Plate 1-1g
Plate 1-1h
Plate 1-1i
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Plate 1-1j
Plate 1-1a
Return to text. Return to image plate.
Plate 1-1b Plate 1-1c

Return to text. Return to image plate. Return to text. Return to image plate.
Plate 1-1d Plate 1-1e

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Plate 1-1f Plate 1-1g

Plate 1-1h Plate 1-1i

Plate 1-1j
Feathered Edge
DISTINCTIVE FEATURES: The feathered edge is the gently tapered end of the smear, located opposite the
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point of application of the blood drop.

DIAGNOSTIC SIGNIFICANCE: Large structures such as platelet clumps, microfilaria, cells containing
infectious agents, and large, normal or neoplastic nucleated cells may be concentrated in this area. Thus,
microscopic examination of the feathered edge is essential.
If large platelet clumps are found at the feathered edge, this may account for a low or low-normal
platelet estimate in the monolayer when evaluating platelet numbers and may also indicate that an
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instrument-generated platelet count may not be valid. Large neoplastic nucleated cells may be found in
low or high numbers in this area, suggesting leukemia or circulating neoplastic cells from nonhemato-
poietic neoplasia (i.e., mast cell tumor and lymphoma). Large parasites, in particular, microfilaria, may
be located in this region as well. Cells with intracellular organisms such as leukocytes containing Histo-
plasma organisms and RBCs containing Babesia spp. hemoparasites may concentrate in the feathered edge.
A differential WBC count may be inaccurately skewed toward larger cell types if the differential is per-
formed mostly at the feathered edge. Also, in the feathered edge, cells are often distorted, damaged, or
ruptured, prohibiting reliable classification and morphologic evaluation.
If excessive pressure is applied to the spreader smear during smear preparation, most of the nucleated
cells will end up in the feathered edge of the smear. Estimates of the total WBC count and determination
of percentages of the different types of leukocytes based only on examination of the monolayer of the
smear will be inaccurate. A total WBC count may be attempted by evaluating the number of nucleated
cells in the feathered edge and monolayer of the smear at 10× magnification.
BLOOD SMEARS FEATHERED EDGE 7
Plate 1-2 Normal Feathered Edge

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Plate 1-2g
Plate 1-2h
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Plate 1-2h
Body
DISTINCTIVE FEATURES: The body is the thick inner part of the smear. The cells are touching, and no
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space exists between cells.

DIAGNOSTIC SIGNIFICANCE: The cells in the body of the smear are distorted and compressed, prohibit-
ing evaluation of cell morphology and performing a differential WBC count. Sometimes, large parasites
such as microfilaria may be found in this region on a low-power scan.
Plate 1-3 Normal Body

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BLOOD SMEARS BODY 9
Plate 1-3 Normal Body (con’t)

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Plate 1-3g
Plate 1-3h
Plate 1-3i
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Plate 1-3a


9.e2
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Plate 1-3j
Background
DISTINCTIVE FEATURES: The background of the smear should be colorless and free of stain debris or
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precipitant.
Increased Basophilia
DISTINCTIVE FEATURES: The background of the smear stains a basophilic hue.
DIAGNOSTIC SIGNIFICANCE: If the background of the smear stains more intensely than normal, increased
plasma protein concentration should be considered, warranting evaluation for hyperglobulinemia.
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Plate 1-4 Increased Background Basophilia

BACKGROUND INCREASED BASOPHILIA 11
Plate 1-4 Increased Background Basophilia (con’t)

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Plate 1-4d
Plate 1-4e
Plate 1-4f
Plate 1-4g
Plate 1-4h
Plate 1-4i
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Cytoplasmic Fragments
DISTINCTIVE FEATURES: With some acute leukemias, the background may contain cytoplasmic fragments
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(sometimes called lymphoglandular bodies) formed from rapidly dividing or ruptured neoplastic cells.
DIAGNOSTIC SIGNIFICANCE: These are characterized by small, basophilic, anucleate blebs that could be
mistaken for platelets. If cytoplasmic fragments are found, assess the slide carefully for circulating neo-
plastic cells.
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BACKGROUND CYTOPLASMIC FRAGMENTS 13
Plate 1-5 Cytoplasmic Fragments

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Platelet clump
2 cytoplasmic blebs
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Plate 1-5e
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Plate 1-5a

Platelet clump
2 cytoplasmic blebs

Infectious Agents
DISTINCTIVE FEATURES: Free hemoparasites may be found in the background, warranting careful micro-
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scopic assessment of the extracellular space. Section VI, Extracellular Organisms, contains detailed
information on the different types of infectious agents that may be found free in the background of blood
films.
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BACKGROUND INFECTIOUS AGENTS 15
Plate 1-6 Extracellular Infectious Agents

A, Microfilaria B, Trypanosomes C, Spirochetes D, Bacteria
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A B
C D
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A
Plate 1-6a
B C
D
Plate 1-6d
Stain Precipitation
DISTINCTIVE FEATURES: Stain precipitant is usually dark blue, fine, and found in small clusters.
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DIAGNOSTIC SIGNIFICANCE: Stain precipitant may be confused with external parasites (especially Myco-
plasma hemofelis organisms that have dislodged from the RBC surface), and when overlapping or adjacent
to RBCs, the stain may be confused with hemoparasites.
Plate 1-7 Stain Precipitant

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BACKGROUND STAIN PRECIPITATION 17
Plate 1-7 Stain Precipitant (con’t)

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Make sure not to confuse stain precipitate with A, Mycoplasma hemocanis;

B, Mycoplasma hemofelis; C, Extracellular bacteria.
A B C
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Plate 1-7a Plate 1-7g
Plate 1-7b Plate 1-7h
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Plate 1-7d Plate 1-7j
Plate 1-7e
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Plate 1-7c Plate 1-7d

Plate 1-7e Plate 1-7f

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Plate 1-7i Plate 1-7j

A B C
Fibrin
DISTINCTIVE FEATURES: Fibrin may be found on the feathered edge of the smear, sometimes with entan-
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gled platelets. Fibrin forms large, slender, homogeneous, pale blue, intertwining strands.
DIAGNOSTIC SIGNIFICANCE: Fibrin should not be confused with microfilaria.
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BACKGROUND FIBRIN 19
Plate 1-8 Fibrin

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Skin Contaminants
DISTINCTIVE FEATURES: Skin contaminants are individualized, polygonal to flattened, basophilic, anucle-
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ate superficial squamous epithelial cells.

DIAGNOSTIC SIGNIFICANCE: Skin contaminants are from either the patient’s skin obtained during veni-
puncture or, more frequently, from fingerprints made on the slide during slide preparation or staining.
These are external contaminants and should not be mistaken for neoplastic cells or parasites.
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BACKGROUND SKIN CONTAMINANTS 21
Plate 1-9 Skin Contaminants

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Plate 1-9a

SECTION 2:
RED BLOOD CELLS
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Normal Morphology (Discocytes and Echinocytes (Crenated Cells, Burr Cells,

Normocytes), 24 Berry Cells), 60
Anemia, 26 Acanthocytes (Spur Cells), 62
Stomatocytes, 64
Polycythemia, 28
Crystalized Hemoglobin, 66
Morphologic Changes Associated with
Erythrocytic Ghosts (Lysed Red Blood Cells,
Disease, 30
Fading Erythrocytes), 68
Rouleaux, 30
Nucleated Red Blood Cells (Metarubricytes and
Agglutination, 32 Rubricytes), 70
Polychromasia, 34 Howell-Jolly Bodies, 74
Reticulocytes, 36 Basophilic Stippling, 78
Hypochromia, 38 Siderotic Inclusions (Pappenheimer Bodies), 82
Anisocytosis, 40 Heinz Bodies, 84
Macrocytosis, 42 Distemper Inclusions, 88
Microcytosis, 44 Dysplastic Changes, 90
Poikilocytosis, 46 Erythrocytic Parasites, 92
Leptocytes: Target Cells (Codocytes) and Mycoplasma hemocanis (previously
Folded Red Blood Cells, 48 Hemobartonella canis), 92
Spherocytes, 50 Mycoplasma hemofelis (previously
Schistocytes (Fragmented Hemobartonella felis), 94
Erythrocytes), 52 Cytauxzoon felis, 98
Keratocytes (Blister Cells and Helmet Babesia spp., 102
Cells), 54 Red Blood Cell Artifacts, 106
Eccentrocytes (Hemighosts), 56 Refractile Artifact, 106
Pyknocytes, 58 Platelet over Red Blood Cell, 108
23
24 SECTION 2 RED BLOOD CELLS
Normal Morphology (Discocytes and Normocytes)

DISTINCTIVE FEATURES: Mature canine and feline erythrocytes are anucleate, red, disc-like cells (disco-
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cytes). Canine red blood cells (RBCs) are approximately 7 micrometers (µm) in diameter and shaped like
biconcave discs with prominent central pallor. Feline erythrocytes are smaller, approximately 6 µm in
diameter, and more “cup” shaped, so they lack the prominent central pallor seen in canine RBCs.
DIAGNOSTIC SIGNIFICANCE: Discocytes are the expected finding in healthy nonanemic dogs and cats
but can be the predominant RBC morphology observed in many disease states as well.
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Plate 2-1
A, Canine red blood cell (RBC) B, Normal feline RBC
A A
A A
A
NORMAL MORPHOLOGY (DISCOCYTES AND NORMOCYTES) 25
Plate 2-1
A, Canine red blood cell (RBC) B, Normal feline RBC (con’t)
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B B
B B
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Plate 2-1b2
Plate 2-1b3
Plate 2-1b4
Plate 2-1b5
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A A
Plate 2-1a2 Plate 2-1a3
A A
Plate 2-1a4 Plate 2-1a5
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B B
Plate 2-1b2 Plate 2-1b3
B B
Plate 2-1b4 Plate 2-1b5
Anemia
DISTINCTIVE FEATURES: Anemia is a decrease in numbers of RBCs compared with established reference
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intervals and can be estimated by blood smear evaluation or quantitated by RBC count, hemoglobin
concentration, and measurement of the hematocrit.
DIAGNOSTIC SIGNIFICANCE: Two general mechanisms of anemia, each with several etiologies, exist.
The first mechanism is decreased RBC production from bone marrow failure or disease. Differentials
could include immune-mediated mechanisms, toxins, drug effects, infectious agents (feline leukemia
virus [FeLV], Ehrlichia spp.), endocrine disease, chronic liver and renal disease, and neoplasia (primary
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bone marrow neoplasia or metastatic neoplasia). The second major mechanism of anemia is increased
RBC loss, which could be secondary to immune-mediated RBC destruction, hemolysis (viral, bacterial,
or ehrlichial infections; RBC fragmentation; oxidative RBC damage), and acute and chronic blood loss
(coagulopathies, trauma, blood sucking endoparasites and ectoparasites).
NEXT STEPS: When anemia is diagnosed, careful blood smear assessment of RBC morphology and
investigation for hemoparasites are important for classifying the anemia and identifying the cause
and to assess for evidence of RBC regeneration. Additionally, evaluation of platelet numbers and
morphology and the white blood cell (WBC) line may help identify important etiologies of anemia.
ANEMIA 27
Plate 2-2 Anemic Blood Smears

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Plate 2-2h
Polycythemia
DISTINCTIVE FEATURES: Polycythemia is an increase in the number of RBCs compared with established
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reference intervals and may be estimated by blood smear evaluation or quantitated by RBC count,
hemoglobin concentration, and measurement of the hematocrit.
DIAGNOSTIC SIGNIFICANCE: The most common cause of polycythemia in dogs and cats is hemocon-
centration secondary to decreased plasma volume (dehydration and endotoxic shock). A mild or moder-
ate transient polycythemia may be secondary to epinephrine release (from excitement or fright), resulting
in splenic contraction, especially in the dog. The last major category is from increased RBC production,
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which may be physiologically appropriate and inappropriate. Appropriate erythroid production is in

response to hypoxia primarily secondary to cardiac disease, pulmonary disease, or both. Animals living
at high altitudes and some canine breeds (Greyhound and other sight breeds) also have greater hema-
tocrit values. Inappropriate polycythemia may result from inappropriate production of erythropoietin
(responsible for stimulating RBC production in the marrow), as from renal cysts or tumors and some
other types of neoplasia. Lastly, polycythemia vera (primary bone marrow neoplasia or leukemia) results
in increased RBC production independent of erythropoietin.
NEXT STEPS: When polycythemia is identified, dehydration must first be excluded by physical exami-
nation and by checking blood chemistry parameters (total protein concentration, blood urea nitrogen
[BUN]) and a urinalysis (urine specific gravity). If polycythemia is not secondary to dehydration, is
persistent, and the patient does not live at high altitudes and is not a sight hound breed, then assess-
ment for appropriate or inappropriate causes of increased RBC production should be considered and
may include blood smear assessment, measurement of erythropoietin levels, bone marrow cytology,
and exclusion of cardiac, pulmonary, and renal diseases.
POLYCYTHEMIA 29
Plate 2-3 Polycythemia

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Plate 2-3h
Morphologic Changes Associated with Disease

Rouleaux
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DISTINCTIVE FEATURES: Groups of RBCs forming peculiar linear stacks in a conformation similar to a
roll of coins, is called rouleaux and is secondary to the electrostatic association of RBCs. Rouleaux may
be seen in the thick area of the smear in healthy cats but should not be present in the monolayer of the
smear.
DIAGNOSTIC SIGNIFICANCE: Rouleaux may be seen occasionally in healthy dogs and especially cats.
Increased rouleaux formation is occasionally observed in sick or debilitated dogs and cats with increased
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fibrinogen or globulin concentration. Rouleaux is generally associated with inflammatory conditions but
also in some noninflammatory conditions such as lymphoproliferative disorders resulting in increased
globulin production. Lipemic blood samples may cause increased rouleaux as well.
NEXT STEPS: Careful attention should be made to not confuse rouleaux with RBC agglutination. Rou-
leaux are orderly linear stacks of RBCs, whereas RBC agglutination is formed by grapelike RBC
aggregates. To aid in differentiating between rouleaux and agglutination, a saline dilution test is
useful. Rouleaux may be easily dissociated by dilution of RBCs in saline, whereas true agglutination
persists despite saline dilution (for method, see Appendix). If rouleaux is confirmed, determine
plasma or serum total protein, albumin, globulin, fibrinogen values and lipemic index, and investigate
potential causes of any abnormalities found.
Plate 2-4 Rouleaux

MORPHOLOGIC CHANGES ASSOCIATED WITH DISEASE ROULEAUX 31
Plate 2-4 Rouleaux (con’t)

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Make sure not to confuse rouleaux and agglutination.

A, Rouleaux B, Agglutination
10 um
A B
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10 um
A B
Agglutination
DISTINCTIVE FEATURES: Agglutination is a random, disorganized clumping of RBCs as opposed to the
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organized stack-of-coins formation seen with rouleaux formation. Agglutination is most prominent in
the body of the blood film (thick area) and may occur in this area as an artifact. Agglutination is signifi-
cant if found in the monolayer.
DIAGNOSTIC SIGNIFICANCE: Agglutination occurs when antibodies on one RBC bind to antigen on
other RBCs, forming globular to amorphous, grapelike aggregates of RBCs. When present, RBC agglu-
tination is supportive of immune-mediated hemolytic anemia (IMHA). Agglutination is not observed in
most cases of IMHA, but when present, it occurs most commonly with immunoglobulin M (IgM) because
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of its pentavalent nature. However, extremely heavy IgG antibody coating of RBC membranes may cause
agglutination. Agglutination is generally considered diagnostic of IMHA.
NEXT STEPS: If necessary, agglutination may be differentiated from rouleaux formation by performing
a saline dilution (dispersion) test. The presence of RBC agglutination warrants careful assessment of
the blood film for other supportive evidence of IMHA such as spherocytes, and also for underlying
causes of IMHA such as neoplastic cells and hemoparasites.
Plate 2-5 RBC Agglutination
20 um 20 um
10 um
MORPHOLOGIC CHANGES ASSOCIATED WITH DISEASE AGGLUTINATION 33
Plate 2-5 RBC Agglutination (con’t)

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Make sure not to confuse agglutination with rouleaux.

A, Rouleaux B, Agglutination
10 um
A B
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10 um


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A B
Polychromasia
DISTINCTIVE FEATURES: Polychromatophils are young, anucleate erythrocytes that stain both blue and
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pink (hence polychromatophilic), making the cell appear gray or bluish-red with Romanowsky-type
stains. The blue color is caused by the presence of basophilic staining cellular organelles (ribosomes) that
are broken down within 12 to 24 hours after release from bone marrow. Typically, polychromatophilic
cells are larger than normochromic erythrocytes. Polychromatophil cells are counted as reticulocytes
when stained with new methylene blue (see next section).
DIAGNOSTIC SIGNIFICANCE: Significant polychromasia indicates a regenerative bone marrow response
to anemia. Hemorrhage and hemolysis are the two main causes of regenerative anemia and increased
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numbers of polychromatophilic RBCs. Remember that polychromasia does not occur instantly after
hemorrhage or hemolysis but takes 2 to 4 days to increase the number of polychromatophilic erythrocytes
in peripheral blood and may not exceed the reference interval or achieve maximum values for 5 to 7
days. Therefore, early or acute hemorrhagic or hemolytic anemias may have minimal polychromasia.
Also, polychromatophilic RBCs are often difficult to differentiate from normochromic RBCs when smears
are stained with rapid stains (i.e., Diff-Quik).
In the cat, mild anemias that are regenerative may have minimal or no polychromasia. The lack of
obvious regeneration (reticulocytosis) does not rule out either hemorrhage or hemolysis. Potential causes
include early hemorrhage or hemolysis, immune-mediated disease directed at RBC precursors, and
recent transfusion, as well as anemia caused by multifactorial causes. If polychromasia is not evident in
a feline blood smear, a reticulocyte count is recommended, which is a more accurate way to determine
regeneration in cats.
NEXT STEPS: Significant polychromasia may be used to call an anemia regenerative, but the absence
of polychromasia should be interpreted cautiously. The anemia may appear nonregenerative in early
acute hemorrhagic or hemolytic anemias, and polychromasia may be difficult to recognize with some
stains. Thus, if the onset of anemia is unknown, serial complete blood count (CBC) examinations may
be needed to evaluate for regeneration. Reticulocyte counts should also be performed in cases where
regeneration is questioned.
Plate 2-6 Polychromasia

MORPHOLOGIC CHANGES ASSOCIATED WITH DISEASE POLYCHROMASIA 35
Plate 2-6 Polychromasia (con’t)

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H-J body
Plate 2-6j
Reticulocytes
DISTINCTIVE FEATURES: When immature, anucleate erythrocytes are stained with a supravital stain such
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as new methylene blue (NMB), the stain penetrates the RBC membrane and binds to the ribosomes,
staining them dark blue and causing them to clump, which identifies these cells as reticulocytes.
The dog has only one identifiable form of reticulocytes in peripheral blood, called aggregate reticulo-
cytes. Cats, in contrast, have two: (1) punctate reticulocytes and (2) aggregate reticulocytes. Punctate
reticulocytes contain few, small, dotlike (punctate), blue-black structures (small polyribosome aggre-
gates) but no large aggregates of ribosomes. Aggregate reticulocytes contain one or more medium to
large, blue-black structures that may appear as a cluster or network of aggregated structures.
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The clinical usefulness of punctate reticulocyte counts in cats remains unclear. Many laboratories and
all hematology instruments that report reticulocyte counts in cats, count and report only aggregate
reticulocytes. Manual reticulocyte counts are done by counting the number of reticulocytes present in
1000 RBCs. The percentage is determined by dividing the number of reticulocytes counted by 10. The
absolute reticulocyte count is determined by multiplying the reticulocyte % by the RBC count. The degree
of reticulocytosis is best indicated by the absolute aggregate reticulocyte count (see the Appendix for
more information).
Canine reticulocytes released from the bone marrow mature rapidly from aggregate reticulocytes to
mature erythrocytes in approximately 24 hours. Punctate reticulocytes are present in low numbers in
dogs. Feline aggregate reticulocytes released from bone marrow mature within 12 to 24 hours to the
punctate reticulocyte stage. Feline punctate reticulocytes may take up to 2 weeks to develop into mature
erythrocytes.
Usually, good correlation exists between the degree of polychromasia observed on a blood film and
the aggregate reticulocyte numbers (with the exception of cats, see above). Punctate reticulocytes appear
the same as mature erythrocytes on a Wright-stained blood film.
DIAGNOSTIC SIGNIFICANCE: The effectiveness of bone marrow response to anemia is determined by
the number of aggregate reticulocytes in dogs and cats. Regenerative anemias occur secondary to blood
loss and IMHA if the bone marrow is healthy. Some heavy metal toxicities, in particular, lead poisoning,
result in excessive numbers (relative to the degree of anemia) of reticulocytes and nucleated RBCs
(nRBCs). A mild to moderate anemia may be seen typically only in chronic lead poisoning. Also, with
lead poisoning, basophilic stippling is noted within mature RBCs stained with Wright stain.
NEXT STEPS: If the anemia is regenerative (above reference interval), top considerations include blood
loss (internal or external hemorrhage) or hemolysis. Note that chronic blood loss anemia may be
nonregenerative because of iron deficiency. If the anemia is not adequately regenerative, it should be
investigated if adequate time has elapsed for a regenerative response to have occurred. If deemed
truly nonregenerative, further testing such as biochemical profile, urinalysis, and bone marrow cytol-
ogy are indicated.
Plate 2-7 Reticulocytes A, Canine B, Feline
Canine Reticulocytes Feline Aggregate Reticulocyte
Feline Punctate Reticulocyte
A B
MORPHOLOGIC CHANGES ASSOCIATED WITH DISEASE RETICULOCYTES 37
Plate 2-7 Reticulocytes (con’t)

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A A
A A
Aggregate
Punctates
B B
Aggregate Reticulocyte Feline Punctate Reticulocytes
Artifact
Heinz Bodies
Punctate Reticulocyte
B B
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Canine Reticulocytes
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A
Plate 2-7a
Feline Aggregate Reticulocyte
Feline Punctate Reticulocyte
B A
A A
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A B
Aggregate Reticulocyte
Aggregate
Artifact
Punctates
Heinz Bodies
Punctate Reticulocyte
B B
Feline Punctate Reticulocytes
B
Plate 2-7j
Hypochromia
DISTINCTIVE FEATURES: Hypochromic RBCs are excessively pale mature erythrocytes with increased
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central pallor, showing a gradual change from a pale central area to a light red periphery.
DIAGNOSTIC SIGNIFICANCE: Hypochromic RBCs are hemoglobin deficient mature erythrocytes.
Increased hypochromia indicates iron deficiency anemia, which occurs secondary to low-grade, chronic
blood loss. Iron deficiency anemia may be secondary to blood loss from the gastrointestinal tract (ulcers,
neoplasia, parasites); blood loss in urine (urinary tract infection, calculi, or neoplasia); or external blood
loss from blood sucking ectoparasites (fleas, ticks). Milk is low in iron; therefore, young, nursing animals
are very susceptible to developing iron deficiency.
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NEXT STEPS: Hypochromic RBCs must be differentiated from artifacts. Hypochromic RBCs show a
gradual change from red to pale. Artifacts appearing as punched-out centers with a sharp demarca-
tion from dark (red) to light (clear area) are called torocytes. Torocytes are artifacts and do not indicate
iron deficiency anemia. Dogs often develop hypochromic RBCs with iron deficiency anemia but feline
erythrocytes are less likely to become hypochromic. Significant hypochromia warrants ruling out
sources of chronic blood loss. An iron profile may also be considered.
Plate 2-8 Hypochromic Red Blood cell

MORPHOLOGIC CHANGES ASSOCIATED WITH DISEASE HYPOCHROMIA 39
Plate 2-8 Hypochromic Red Blood cell (con’t)

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Plate 2-8j
Anisocytosis
DISTINCTIVE FEATURES: Variation in cell size is called anisocytosis. Normal erythrocytes may vary (larger
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or smaller) up to one third of their size in normal animals.

DIAGNOSTIC SIGNIFICANCE: Increased anisocytosis occurs when significant numbers of small-diameter
RBCs, large-diameter RBCs, or a combination of both are present together with normal-diameter RBCs.
Anisocytosis is most often recognized with significant numbers of spherocytes in IMHA and large
numbers of macrocytic polychromatophils in regenerative anemias.
NEXT STEPS: The significance of anisocytosis is dependent on the numbers of variably sized RBCs and
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the specific morphology of the RBCs constituting the anisocytosis (i.e., spherocytes, polychromato-
phils, etc.).
MORPHOLOGIC CHANGES ASSOCIATED WITH DISEASE ANISOCYTOSIS 41
Plate 2-9 Anisocytosis

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Plate 2-9a
10 um


Macrocytosis
DISTINCTIVE FEATURES: Macrocytosis is defined as larger than normal RBCs.
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DIAGNOSTIC SIGNIFICANCE: Macrocytosis secondary to reticulocytosis is expected in regenerative

anemias. Macrocytosis is abnormal when secondary to feline leukemia virus (FeLV) infection in cats
(myelodysplastic syndrome, erythroleukemia) or inherited disorders such as macrocytosis of poodles
(poodle marrow dyscrasia).
NEXT STEPS: When significant numbers of macrocytic RBCs are evident, investigate for a regenerative
anemia (expect concurrent polychromasia). In anemic cats with asynchronous regeneration (i.e., large
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number of nRBCs and absent or minimal polychromasia or reticulocytosis), consider FeLV testing to
aid in identification FeLV associated myelodysplastic syndrome. Check for breed-associated disorders
as well. Note that agglutination may cause a falsely increased mean corpuscular volume (MCV), sug-
gesting macrocytosis; as aggregated RBCs are sometimes measured as a single RBC, careful examina-
tion of the blood film is important.
Plate 2-10 Macrocytes

MORPHOLOGIC CHANGES ASSOCIATED WITH DISEASE MACROCYTOSIS 43
Plate 2-10 Macrocytes (con’t)

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Microcytosis
DISTINCTIVE FEATURES: Microcytes are RBCs with reduced cell volume compared with normal RBCs.
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DIAGNOSTIC SIGNIFICANCE: Small RBCs may occur with acquired disorders (i.e., iron deficiency
anemia, schistocytosis, bone marrow disorders); congenital disorders (i.e., familial dyserythropoiesis of
English Springer Spaniels), and hereditary disorders (i.e., microcytosis of Akita and Shiba Inu dogs).
Microcytes may be seen in both congenital and acquired liver disease, in particular liver shunts. MCV
may be artifactually low if a short blood sample in an ethylenediaminetetraacetic acid (EDTA) tube is
evaluated (excess EDTA). Spherocytes are small-diameter cells, which are included in this section,
although these cells are not truly microcytic. Spherocytes may appear as small-diameter cells on a blood
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smear; however, this is caused by their spheric shape, which conserves volume after removal of mem-
brane. Thus, spherocytes are not considered by many to be true microcytes.
NEXT STEPS: When significant numbers of microcytes are evident on blood smear review, rule out
iron deficiency anemia, IMHA, microangiopathy, liver disease, and check for breed-associated
disorders.
Plate 2-11 Microcytes

MORPHOLOGIC CHANGES ASSOCIATED WITH DISEASE MICROCYTOSIS 45
Plate 2-11 Microcytes (con’t)

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Poikilocytes
DISTINCTIVE FEATURES: Abnormally shaped RBCs are generically termed poikilocytes.
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DIAGNOSTIC SIGNIFICANCE: Many different forms of poikilocytes exist, as well as many different causes
of poikilocytosis; a few important causes include the following:
• In vitro artifactual changes (echinocytes and some acanthocytes)
• RBC fragmentation (schistocytes and keratocytes)
• Membrane loss (spherocytes)
• RBC membrane abnormalities (acanthocytes and stomatocytes)
• Excess RBC membrane compared with cytoplasm (target cells)
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The diagnostic significance of poikilocytes depends on which of the many different types are present
and, in some cases, is dependent on the numbers of affected RBCs (i.e., low numbers of Heinz bodies
may be normally seen in feline blood, but Heinz bodies are not normal in canine blood). Anemia is often
associated with poikilocytes and may range from mild to moderate or severe. Some of the more impor-
tant poikilocytes are discussed individually below.
Plate 2-12 Poikilocytosis

MORPHOLOGIC CHANGES ASSOCIATED WITH DISEASE POIKILOCYTES 47
Plate 2-12 Poikilocytosis (con’t)

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Plate 2-12b
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Plate 2-12h
Plate 2-12i
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Plate 2-12j
Leptocytes: Target Cells (Codocytes) and Folded Red Blood Cells

DISTINCTIVE FEATURES: Leptocytes are thin erythrocytes and may appear as either target cells or folded
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cells. Target cells have a dark central area surrounded by a clear area, which is then encircled by a dark
peripheral region giving the appearance of a target. Folded cells have a fold in the membrane or have a
central bar of hemoglobin.
DIAGNOSTIC SIGNIFICANCE: Occasional target cells are normal in the peripheral blood of dogs. Increased
numbers of target cells may be observed in blood from dogs with regenerative anemia, and sometimes,
polychromatophilic cells appear as target cells, folded cells, or both. Leptocytes may be seen in pathologic
states such as iron deficiency anemia, congenital dyserythropoiesis, liver disease, or nephrotic
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syndrome.
NEXT STEPS: The presence of leptocytes should prompt investigation into disorders such as iron
deficiency anemia, IMHA, regenerative anemia, liver disease, and renal disease.
Plate 2-13 Leptocytes

POIKILOCYTES LEPTOCYTES: TARGET CELLS (CODOCYTES) AND FOLDED RED BLOOD CELLS 49
Plate 2-13 Leptocytes (con’t)

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Plate 2-13b
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Plate 2-13d
Plate 2-13e
Plate 2-13f
Plate 2-13g
Plate 2-13h
Plate 2-13i
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Plate 2-13a


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Spherocytes
DISTINCTIVE FEATURES: Spherocytes are small RBCs that are less than two thirds the diameter of normal
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erythrocytes. As they are spherical and not disc shaped, they lack central pallor and appear dense, often
darker (more red in color) than discocytes. Only spherocytes in the monolayer region should be identi-
fied, as in other areas of the smear artifactual spherocytes may be present.
DIAGNOSTIC SIGNIFICANCE: Spherocytes are fairly easy to recognize in dogs but are often impossible
to recognize in cats, as normal feline RBCs lack central pallor. When spherocytes are observed in moder-
ate to high numbers, IMHA is the most likely cause. IMHA may be a primary (autoimmune mediated)
disease or a secondary disease (initiated by primary infectious, inflammatory or neoplastic conditions).
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Low numbers of spherocytes may be seen in many disorders, including Heinz body hemolytic anemia,
hemoparasitism, envenomation (i.e., snake bites and bee stings), zinc toxicity, microangiopathy and
neoplasia (i.e., lymphoma and hemangiosarcoma).
NEXT STEPS: If significant numbers of spherocytes are present, look for RBC agglutination to support
the diagnosis of IMHA; if agglutination is not evident, consider Coombs test to lend additional
support to the IMHA diagnosis. Even if autoagglutination is not evident and the Coombs test is nega-
tive, significant numbers of spherocytes are still most likely secondary to IMHA. Carefully check for
hemoparasites, circulating neoplastic cells, evidence of neutrophil toxic changes, and other RBC mor-
phologic abnormalities, especially if secondary IMHA is suspected.
Plate 2-14 Spherocytes
nRBC
Polychromatophilic
RBC
nRBC Neutrophils
H-J
Body
Spherocytes
POIKILOCYTES SPHEROCYTES 51
Plate 2-14 Spherocytes (con’t)

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Plate 2-14a nRBC
Plate 2-14b
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Polychromatophilic
Plate 2-14c RBC
Plate 2-14d
Plate 2-14e
Plate 2-14f
Plate 2-14g
Plate 2-14h nRBC Neutrophils
Plate 2-14i
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H-J
Body
Spherocytes
Plate 2-14a


2
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Plate 2-14j
Schistocytes (Fragmented Erythrocytes)

DISTINCTIVE FEATURES: Schistocytes are RBC fragments; they are smaller than normal RBCs, often irregu-
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lar and sometimes triangular in shape with pointed extremities.

DIAGNOSTIC SIGNIFICANCE: Schistocytes are commonly seen with microangiopathic hemolytic disease,
where erythrocytes hit fibrin strands, are cut, and their phospholipid membranes, upon touching, reseal,
resulting in circulating erythrocyte fragments.
Schistocytes occur with many disorders such as disseminated intravascular coagulation (DIC), myelo-
fibrosis, vasculitis, glomerulonephritis, hemangiosarcoma, cardiac disease, and iron deficiency anemia.
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Some of the RBC fragments will round up and become spherocytes; therefore, some spherocytes may be
seen along with schistocytes.
NEXT STEPS: Investigate for microangiopathic disease—especially vascular neoplasms of the liver and
spleen (hemangiosarcoma). Rule out DIC by evaluating a coagulation profile. To investigate iron
deficiency anemia, look for microcytic, hypochromic RBCs and investigate for a source of blood loss
(endoparasites, ectoparasites, hematuria, gastrointestinal blood loss). Cardiac auscultation, electrocar-
diography, heartworm testing, and echocardiography may be considered to assess for cardiac disease.
Plate 2-15 Schistocytes

POIKILOCYTES SCHISTOCYTES (FRAGMENTED ERYTHROCYTES) 53
Plate 2-15 Schistocytes (con’t)

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Plate 2-15b
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Plate 2-15d
Plate 2-15e
Plate 2-15f
Plate 2-15g
Plate 2-15h
Plate 2-15i
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Plate 2-15a


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Plate 2-15j
Keratocytes (Blister Cells and Helmet Cells)

DISTINCTIVE FEATURES: Keratocytes are mature erythrocytes that contain single or multiple, intact or
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ruptured, clear, peripheral circular areas. RBCs containing intact clear circular areas are sometimes called
blister cells, whereas RBC containing clear circular areas that have ruptured resulting in one or two pro-
jections from the RBCs are sometimes called helmet cells.
DIAGNOSTIC SIGNIFICANCE: Keratocytes are formed by RBC trauma, very similar to schistocytes. Kera-
tocyte formation is potentiated in feline blood stored in EDTA anticoagulant. Keratocytes are seen in
various disorders associated with other RBC abnormalities (i.e., acanthocytosis, echinocytosis) in dogs.
Keratocytes may be seen with various other disorders such as DIC, liver disease, vascular neoplasia (i.e.,
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hemangiosarcoma), iron deficiency anemia, and myelodysplastic syndrome.

NEXT STEPS: Many of the same diseases resulting in RBC trauma that form schistocytes also cause
keratocyte formation (refer to above section on schistocytes).
Plate 2-16 Keratocytes
10 um
POIKILOCYTES KERATOCYTES (BLISTER CELLS AND HELMET CELLS) 55
Plate 2-16 Keratocytes (con’t)

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Plate 2-16b
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Plate 2-16d
Plate 2-16e
Plate 2-16f
Plate 2-16g
Plate 2-16h
Plate 2-16i
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Plate 2-16a
10 um


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Eccentrocytes (Hemighosts)
DISTINCTIVE FEATURES: Eccentrocytes are mature erythrocytes in which the hemoglobin is pushed to one
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side of the cell leaving an opposing pale staining region.

DIAGNOSTIC SIGNIFICANCE: Eccentrocytes result from oxidative damage to the erythrocytes from inges-
tion of exogenous oxidants such as onions, garlic, acetaminophen, propylene glycol (some foods and
antifreeze solutions), zinc, and vitamin K antagonist intoxication. They may also be formed from elabora-
tion of endogenous oxidants in some very ill patients with diseases such as diabetes mellitus.
NEXT STEPS: Check for other evidence of oxidative injury to RBCs such as the presence of
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Heinz bodies. Finding eccentrocytes warrants investigation for oxidant ingestion and for underlying
disease.
Plate 2-17 Eccentrocytes
Eccentrocyte with
Heinz Body
Howell-Jolly
Body
Heinz Body
POIKILOCYTES ECCENTROCYTES (HEMIGHOSTS) 57
Plate 2-17 Eccentrocytes (con’t)

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Platelet
Heinz
Body
Eccentrocyte
Polychromatophilc
RBC
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Plate 2-17b
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Plate 2-17d
Plate 2-17e
Plate 2-17f
Plate 2-17g
Plate 2-17h
Plate 2-17i
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Plate 2-17a

Eccentrocyte with
Heinz Body
Howell-Jolly
Body
Heinz Body

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Platelet
Heinz
Body
Eccentrocyte
Polychromatophilc
RBC
Plate 2-17j
Pyknocytes
DISTINCTIVE FEATURES: Dense, irregularly contracted RBCs are formed by oxidative damage. Pyknocytes
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look a lot like spherocytes and are often differentiated by the company they keep (i.e., pyknocytes occur
with eccentrocytes and may even form from eccentrocytes). Some pyknocytes will have a membranous
tag unlike spherocytes, which are smooth and perfectly round. Pyknocytes stain more intensely with
NMB stain compared with spherocytes.
DIAGNOSTIC SIGNIFICANCE: Pyknocytes indicate oxidative injury and generally occur with
eccentrocytes.
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NEXT STEPS: On the basis of history, clinical findings, and laboratory findings, investigate for causes
of oxidative insult.
POIKILOCYTES PYKNOCYTES 59
Plate 2-18 Pyknocytes

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Plate 2-18f
Echinocytes (Crenated Erythrocytes, Burr Cells, Berry Cells)

DISTINCTIVE FEATURES: Echinocytes are spiculated erythrocytes with equally spaced projections over
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their entire surface. Morphology may vary from irregularly spiculated (type I), to equally spaced blunt
projections (type II), to equally spaced pointed projections (type III).
DIAGNOSTIC SIGNIFICANCE: Echinocytes may be an artifact caused by slow drying, excess EDTA,
improper smear preparation, or old blood (prolonged storage before smear preparation). However, they
may occur secondary to hyponatremic dehydration, renal disease, doxorubicin (a chemotherapeutic)
toxicity, certain drugs, and snake bite (rattlesnake and coral snake) and bee sting envenomization, in
which the unique type III echinocytes predominate.
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NEXT STEPS: When significant numbers of echinocytes are seen, rule out the possibility of an artifact.
Make sure these cells are differentiated from acanthocytes (see below). If an artifact is excluded,
investigate for patient dehydration and renal disease, review medications, and, if type III echinocytes
are evident, rule out snake bite and bee sting envenomation.
Plate 2-19 Echinocytes

POIKILOCYTES ECHINOCYTES (CRENATED ERYTHROCYTES, BURR CELLS, BERRY CELLS) 61
Plate 2-19 Echinocytes (con’t)

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Plate 2-19b
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Plate 2-19g
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Acanthocytes (Spur Cells)

DISTINCTIVE FEATURES: Acanthocytes are RBCs with multiple, irregular projections that are randomly
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spaced. Acanthocytes contrast to echinocytes in that acanthocyte projections are randomly spaced, and
echinocyte projections are regularly spaced.
DIAGNOSTIC SIGNIFICANCE: Acanthocytes may occur in dogs and cats with liver, splenic, or renal dis-
orders. Acanthocytes are important, as they are often seen in the peripheral blood of patients with
vascular neoplasms, in particular hemangiomas and hemangiosarcomas. Acanthocytes may also be seen
with DIC and lymphoma.
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NEXT STEPS: Make sure to differentiate acanthocytes from echinocytes. If true acanthocytes are present,
top considerations include an underlying vascular neoplasm, particularly involving the liver and
spleen.
Plate 2-20 Acanthocytes

POIKILOCYTES ACANTHOCYTES (SPUR CELLS) 63
Plate 2-20 Acanthocytes (con’t)

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Plate 2-20b
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Plate 2-20h
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Stomatocytes
DISTINCTIVE FEATURES: Stomatocytes are mature erythrocytes of normal size and color but with an oval
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or stoma (mouth)–shaped area of central pallor.

DIAGNOSTIC SIGNIFICANCE: Stomatocytes most commonly occur as artifacts, especially in the thick
areas of the smear. Stomatocytes occur in high numbers in the peripheral blood of Alaskan Malamutes
with hereditary stomatocytosis. This condition is characterized by short-limb dwarfism, chondrodyspla-
sia, and anemia. The erythrocytes have a shortened life span, and a regenerative response is usually
evident. Hereditary stomatocytosis has also been reported in Miniature and Standard Schnauzers without
clinical signs of disease. Familial stomatocytosis–hypertrophic gastritis is a multiorgan disease affecting
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Drentse Partirjshond dogs (Dutch Partridge dogs). In Abyssinian and Somali cats, a hereditary RBC
defect, characterized by increased osmotic fragility, macrocytosis, and some stomatocytosis, is the sus-
pected cause of recurrent Coombs-negative hemolytic anemia. Small-diameter stomatocytes (termed
imperfect spherocytes and stomatospherocytes) may be seen with some forms of IMHA, particularly those
that are poorly regenerative.
NEXT STEPS: When stomatocytes are evident in significant numbers, first investigate for breed-
associated disorders. If imperfect spherocytes or stomatospherocytes are evident, assess for other
evidence to support the diagnosis of IMHA (i.e., RBC agglutination, ghost RBC, positive Coombs test,
RBC regeneration).
Plate 2-21 Stomatocytes

POIKILOCYTES STOMATOCYTES 65
Plate 2-21 Stomatocytes (con’t)

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Plate 2-21b
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Plate 2-21d
Plate 2-21e
Plate 2-21f
Plate 2-21g
Plate 2-21h
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Crystallized Hemoglobin
DISTINCTIVE FEATURES: Large, diamond, rhomboid, rectangular, or square-shaped, intensely staining
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hemoglobin crystals are observed within RBCs.

DIAGNOSTIC SIGNIFICANCE: Hemoglobin crystals are seen most commonly in cats and rarely in dogs.
No recognized significance has been reported.
Plate 2-22 Crystallized Hemoglobin

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MORPHOLOGIC CHANGES ASSOCIATED WITH DISEASE CRYSTALLIZED HEMOGLOBIN 67
Plate 2-22 Crystallized Hemoglobin (con’t)

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Plate 2-22b
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Plate 2-22e
Plate 2-22f
Plate 2-22g
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Plate 2-22j
Erythrocytic Ghosts (Lysed Red Blood Cells, Fading Erythrocytes)

DISTINCTIVE FEATURES: Erythrocytic ghosts are pale RBC membranes with no or minimal hemoglobin.
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DIAGNOSTIC SIGNIFICANCE: With intravascular hemolysis, the hemoglobin is dispersed, and the eryth-
rocyte membrane is rapidly cleared. Therefore, the presence of erythrocytic ghosts on peripheral blood
smears suggests either very recent intravascular hemolysis or in vitro hemolysis. RBC lysis that occurs
during smear preparation usually appears as RBC smudges. With lipemia, erythrocytes have increased
membrane permeability and fragility, resulting in occasional smudged erythrocytes and erythrocytic
ghosts, especially when smears are not made immediately. Smudged erythrocytes are characterized by
fuzzy, ill-defined cell borders.
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NEXT STEPS: When ghost RBCs are present, evaluate the smear for concurrent RBC agglutination and
spherocytosis to lend support to the diagnosis of intravascular hemolysis. Check the specimen for
lipemia, and rule out in vitro hemolysis by reviewing blood collection and handling techniques.
Plate 2-23 Erythrocytic Ghosts

MORPHOLOGIC CHANGES ASSOCIATED WITH DISEASE ERYTHROCYTIC GHOSTS 69
Plate 2-23 Erythrocytic Ghosts (con’t)

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Plate 2-23b
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Plate 2-23d
Plate 2-23e
Plate 2-23f
Plate 2-23g
Plate 2-23h
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Plate 2-23a


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Plate 2-23j
Nucleated Red Blood Cells (Metarubricytes and Rubricytes)

DISTINCTIVE FEATURES: Metarubricytes and rubricytes are nRBCs that are most commonly seen in
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peripheral blood smears. These cells have small, round, central nuclei with coarse chromatin (rubricyte)
or smooth pyknotic chromatin (metarubricyte), and either polychromatophilic (blue-gray) or orthochro-
mic (orange-red) cytoplasm, which help in differentiating them from small lymphocytes. Additionally,
nuclei in nRBCs are central in location, and a rim of cytoplasm may be traced around the entire nucleus.
In contrast, it is difficult to trace a complete rim of cytoplasm around the nucleus of mature lymphocytes
because of the eccentric placement of the nucleus.
DIAGNOSTIC SIGNIFICANCE: Nucleated RBCs (usually of the metarubricyte or rubricyte stages) are
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occasionally found in small numbers in peripheral blood smears from normal dogs and cats (<2
nRBCs/100 WBCs). Nucleated RBCs are quantitated during the WBC differential count, and reported as
the number of nRBCs per 100 WBCs. Increased numbers of nRBCs may occur with both regenerative
(expected) and nonregenerative anemias (abnormal, suggesting an asynchronous regenerative response).
The most common cause of increased nRBCs is with regenerative anemia (hemorrhage or hemolysis).
It is important to note that the presence of nRBCs alone should not be taken as an indication of bone
marrow response to an anemia, as increased nRBCs may be seen in nonregenerative anemias. Evaluation
of the regenerative response to an anemia should always be based on the degree of reticulocytosis.
Nucleated erythrocytes may be present in low numbers in peripheral blood smears secondary to
conditions such as splenic contraction; splenic disease or dysfunction or splenectomy (decreased nRBC
removal from the circulation); trauma (bone fractures resulting in immature cells of the marrow entering
the circulation); hyperadrenocorticism (Cushing’s disease); steroid administration (increased RBC sur-
vival time in the circulation); and cardiovascular disease (secondary to hypoxia). They may be seen in
certain canine breeds (miniature Schnauzers and toy and miniature Poodles) and in some inflammatory
conditions.
Nucleated erythrocytes may be present in low, moderate, or high numbers with many disease states
such as regenerative anemias, lead poisoning, bone marrow diseases, hemangiosarcoma, septicemia, and
endotoxic shock. Bone marrow neoplasia (primary leukemia or metastatic neoplasia), inflammatory
diseases, lead poisoning, and heat stroke may damage the bone marrow sinusoidal endothelial lining
and put pressure on marrow hematopoietic tissue, causing nRBCs to more easily and more rapidly pass
into the circulation.
More immature stages of nRBCs (back to the rubriblast stage) may be found with some myeloprolif-
erative diseases such as erythroleukemia and sometimes in markedly regenerative anemias, the latter
sometimes observed in cats with Mycoplasma hemofelis infection.
NEXT STEPS: The significance of finding nRBCs on blood smear review is based on the numbers of
nRBCs, ruling out breed differences, and determining if anemia is present. As the differential WBC
count is performed, nRBCs are recorded as “nRBCs/100 WBC.” If greater than 5 nRBCs/100 WBC
are present, most laboratories correct the WBC count for the presence of nRBCs. Some laboratories
report the total nucleated cell count, which includes leukocytes and nRBCs. Nucleated RBCs are not
included in the differential WBC count, and an absolute number is given.
MORPHOLOGIC CHANGES ASSOCIATED WITH DISEASE NUCLEATED RED BLOOD CELLS 71
Plate 2-24 Nucleated Red Blood Cells (nRBCs)

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Small
lymphocyte
nRBC
Plate 2-24 Nucleated Red Blood Cells (nRBCs) (con’t)

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Rubriblast
Rubriblast
MORPHOLOGIC CHANGES ASSOCIATED WITH DISEASE NUCLEATED RED BLOOD CELLS 73
Make sure not to confuse small lymphocytes (A and B) with nucleated

red blood cells (C and D).
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A C
B D
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Plate 2-24b
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Plate 2-24d
Plate 2-24e
Plate 2-24f
Plate 2-24g
Plate 2-24h
Plate 2-24i
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Plate 2-24a

Small
lymphocyte
nRBC

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Rubriblast

Rubriblast
Plate 2-24j
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A C
B D
Howell-Jolly Bodies
DISTINCTIVE FEATURES: Howell-Jolly bodies (H-J bodies) are small, homogeneous, dark purple (same
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color as RBC nuclei), spherical structures within the RBC cytoplasm.

DIAGNOSTIC SIGNIFICANCE: H-J bodies are fragments of RBC nuclei retained after expulsion of the
nucleus (nuclear remnants). One should be careful not to confuse H-J bodies with an erythroparasite.
Low numbers of H-J bodies may be present in the peripheral blood of normal cats and are infrequently
observed in the peripheral blood of normal dogs. The number of erythrocytes containing H-J bodies
tends to increase during regenerative anemias (similar to the increase in nRBCs) as well as other condi-
tions (similar disorders and mechanisms seen with increased nRBCs) such as splenic disease, postsple-
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nectomy state, bone marrow disease (myelophthesis), chemotherapy, radiation, erythroid toxins, and
glucocorticoid therapy.
NEXT STEPS: The number of H-J bodies and association with anemia or normovolemia determine the
significance of H-J bodies found on blood smear review.
MORPHOLOGIC CHANGES ASSOCIATED WITH DISEASE HOWELL-JOLLY BODIES 75
Plate 2-25 Howell-Jolly Bodies

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Neutrophil
Trypanosoma
cruzi H-J body
nRBC
NRC
Small
lymphocyte
Make sure not to confuse Howell-Jolly bodies with: A, Babesia gibsoni

B, Canine Distemper Inclusions C, Basophilic stippling
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Babesia gibsoni
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Howell-Jolly Body
Babesia
gibsoni
A
Basophilic stippling
Canine Distemper
B C
MORPHOLOGIC CHANGES ASSOCIATED WITH DISEASE HOWELL-JOLLY BODIES 77
Make sure not to confuse Howell-Jolly bodies with: D, Cytauxzoon felis

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H-J body
Cytauxzoon
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Cytauxzoon
H-J body
Cytauxzoon
D D
H-J body
Cytauxzoon
D
77.e1
Next Topic Plate 2-25e

Plate 2-25a Plate 2-25f
Plate 2-25b Plate 2-25g
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Plate 2-25c Plate 2-25h

Plate 2-25d
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Neutrophil
Trypanosoma
cruzi H-J body
nRBC
NRC

77.e2
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Small
lymphocyte
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Babesia gibsoni
Howell-Jolly Body
Babesia
gibsoni
A
77.e3
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Canine Distemper
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B C
H-J body
Cytauxzoon
Cytauxzoon
H-J body
Cytauxzoon
D D
77.e4
H-J body
Cytauxzoon
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D

Basophilic Stippling
DISTINCTIVE FEATURES: When ribosomes become unstable, aggregate, and precipitate during drying of
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the blood film, they stain as faint blue-black dots evenly dispersed within erythrocytes and are recog-
nized as basophilic stippling.
DIAGNOSTIC SIGNIFICANCE: Basophilic stippling may occur with regenerative anemias in both dogs
and cats and commonly with lead poisoning. Make sure to distinguish basophilic stippling from siderotic
inclusions (see below). Basophilic stippling is usually finely distributed throughout the cytoplasm,
whereas siderotic inclusions tend to aggregate loosely.
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NEXT STEPS: If basophilic stippling and moderate numbers of nRBCs are present in a nonanemic
patient or in one with a mild anemia, blood lead levels should be determined to rule out lead toxicosis.
Mild anemia may occur with chronic lead exposure. Assess for neurologic signs, gastrointestinal clini-
cal signs, or both, which occur secondary to lead poisoning. If basophilic stippling (primarily within
polychromatophilic RBCs) is evident with a strongly regenerative moderate or marked anemia, the
basophilic stippling is likely secondary to erythroid regeneration.
MORPHOLOGIC CHANGES ASSOCIATED WITH DISEASE BASOPHILIC STIPPLING 79
Plate 2-26 Basophilic Stippling

Make sure to distinguish basophilic stippling from Pappenheimer bodies
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(see Section II, plate 27, for illustrations of Pappenheimer bodies).

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Plate 2-26 Basophilic Stippling (con’t)

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nRBC
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Dog with IMHA
Spherocytes
Basophilic
Stippling
MORPHOLOGIC CHANGES ASSOCIATED WITH DISEASE BASOPHILIC STIPPLING 81
Make sure not to confuse with basophilic stippling. A, Canine distemper inclusions
B, Howell-Jolly bodies C, Babesia gibsoni
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Howell-Jolly bodies
Canine Distemper Babesia gibsoni
A B C
81.e1
Next Topic
Plate 2-26a
Plate 2-26b
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Plate 2-26c
Plate 2-26d
Plate 2-26e
Plate 2-26f
Plate 2-26g
Plate 2-26h
Plate 2-26i
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Plate 2-26j
Plate 2-26k
Plate 2-26l
Plate 2-26a


81.e2
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Dog with IMHA
nRBC
Spherocytes
Basophilic
Stippling

Plate 2-26j Plate 2-26k

81.e3
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Plate 2-26l
Howell-Jolly bodies
A B C
Siderotic Inclusions (Pappenheimer Bodies)

DISTINCTIVE FEATURES: Siderotic inclusions are aggregated basophilic structures generally eccentrically
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placed on one side near the periphery of the RBC.

DIAGNOSTIC SIGNIFICANCE: These are iron-containing inclusions that are occasionally seen in disorders
such as lead poisoning, hemolytic anemia, dyserythropoiesis, and myeloproliferative disorders. Poly-
chromatophilic RBCs and mature RBCs containing these inclusions are called siderocytes.
NEXT STEPS: Distinguish siderotic inclusions from basophilic stippling (see section on basophilic stip-
pling). If evidence of lead poisoning is present (mild to no anemia, basophilic stippling, and increased
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numbers of nRBCs), blood lead levels should be determined. If signs of dyserythropoiesis or myelo-
proliferative disease are evident on blood film evaluation, examination of a bone marrow sample may
be indicated. While seldom done, a Prussian blue stain can be used to confirm the presence of Pap-
penheimer bodies (siderotic inclusions will stain blue).
Plate 2-27 Siderotic Inclusions

MORPHOLOGIC CHANGES ASSOCIATED WITH DISEASE SIDEROTIC INCLUSIONS 83
Plate 2-27 Siderotic Inclusions (con’t)

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Plate 2-27a
Plate 2-27b
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Plate 2-27c
Plate 2-27d
Plate 2-27e
Plate 2-27f
Plate 2-27g
Plate 2-27h
Plate 2-27i
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Plate 2-27a


83.e2
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Heinz Bodies
DISTINCTIVE FEATURES: Heinz bodies appear as singular, round projections from the RBC membrane.
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They may be quite small or very large and prominent. In some cases, a single RBC may have several
small Heinz bodies. Heinz bodies stain the same red-pink color of mature RBCs with routine Wright
stains, and stain pale basophilic with Diff-Quik stain. With Diff-Quik stain, Heinz bodies may be seen
both as round projections from the RBC membrane and as clear round inclusions that have not yet pro-
jected out from the RBC membrane. If the presence of Heinz bodies is in question, or to get a more
accurate determination of the number of Heinz bodies present, a blood smear may be stained with NMB.
With NMB, Heinz bodies stain blue-black, whereas the RBC is pale aqua. In wet mount preparations
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with NMB, Heinz bodies appear as refractile inclusions when out of focus and are sometimes referred
to as erythrocyte refractile bodies (ER bodies).
DIAGNOSTIC SIGNIFICANCE: Heinz bodies occur from oxidative injury resulting in denatured (oxi-
dized), precipitated hemoglobin that tends to adhere to the inner surface of RBC membranes. Heinz
bodies are seen most frequently in peripheral blood smears from cats (up to 5% of RBC in normal cats
have small Heinz bodies). Feline RBCs are more susceptible to Heinz body formation, and the feline
spleen is nonsinusoidal and less efficient at removing Heinz bodies.
Large numbers of Heinz bodies are seen in sick cats, especially those with diabetes mellitus, hyper-
thyroidism, and lymphoma. Heinz bodies are always abnormal in dogs. Some causes of Heinz body
hemolytic anemia secondary to oxidant ingestion include onions, garlic, acetaminophen, methylene blue,
methionine, phenazopyridine, zinc (pennies minted after 1982), naphthalene (chemical in mothballs),
propylene glycol (in some moist foods), and vitamin K3 (menadione).
NEXT STEPS: If moderate to high numbers of Heinz bodies are present in feline blood smears or any
Heinz bodies are detected in canine blood, first rule out oxidant ingestion. In cats, rule out underlying
metabolic disease (diabetes mellitus and hyperthyroidism) and lymphoma. Both enumeration of
Heinz bodies and assessment of the size of the Heinz bodies are useful in determining the extent of
oxidative injury. Also, carefully check RBC morphology for the presence of eccentrocytes, pyknocytes,
and ghost RBCs, which may also be seen with oxidative RBC damage.
MORPHOLOGIC CHANGES ASSOCIATED WITH DISEASE HEINZ BODIES 85
Plate 2-28 Heinz Bodies

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Next Topic
Plate 2-28a
Plate 2-28b
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Plate 2-28c
Plate 2-28d
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Plate 2-28a

Plate 2-28d
Plate 2-29 Pale Staining Heinz bodies

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Heinz
Body
Platelets
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86.e1
Next Topic
Heinz
Plate 2-29a Body
Plate 2-29b
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Plate 2-29c
Plate 2-29d
Platelets
Plate 2-29e
Plate 2-29f
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Plate 2-29a


86.e2
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Plate 2-29f
MORPHOLOGIC CHANGES ASSOCIATED WITH DISEASE HEINZ BODIES 87
Plate 2-30 Heinz Bodies Stained with New Methylene Blue (NMB)
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Reticulocytes
87.e1
Next Topic
Plate 2-30a
Plate 2-30b
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Plate 2-30c
Plate 2-30d
Plate 2-30e
Plate 2-30f
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Plate 2-30a

Reticulocytes

87.e2
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Plate 2-30f
Distemper Inclusions
DISTINCTIVE FEATURES: Distemper inclusions are aggregates of viral nucleocapsids that appear micro-
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scopically as round to oval to irregular, red to blue inclusions. The inclusions tend to occur in either the
monocyte and lymphocyte cell series or the neutrophil and erythrocyte cell series. However, in some
cases of distemper, all cell lines may have inclusions. Distemper inclusions are most easily identified on
Diff-Quik–stained smears, where the inclusions stain a bright, homogeneous eosinophilic color. Distem-
per inclusions may be seen on Wright’s-stained smears; however, they tend to stain pale blue and are
more difficult to identify.
DIAGNOSTIC SIGNIFICANCE: Distemper inclusions are seen only rarely on peripheral blood smears from
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dogs with distemper infection; however, when present, they are diagnostic for infection. Inclusions are
found in a small percentage of dogs during the viremic phase of distemper, which is usually associated
with clinical signs of upper respiratory disease.
Plate 2-31 Distemper Inclusions A. Diff-Quick–stained B. Wright’s stained
A A
A A
A B
MORPHOLOGIC CHANGES ASSOCIATED WITH DISEASE DISTEMPER INCLUSIONS 89
Plate 2-31 Distemper Inclusions (con’t)

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B B
B B
Make sure not to confuse Distemper inclusions with A, Basophilic stippling

B, Howell-Jolly bodies C, Babesia gibsoni
Howell-Jolly bodies
A B C
89.e1

Plate 2-31a Plate 2-31h
Plate 2-31b Plate 2-31i
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Plate 2-31c Plate 2-31j

Plate 2-31d Plate 2-31k
Plate 2-31e
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A A
A A
A B
89.e2
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B B
B B
Howell-Jolly bodies
A B C
Dysplastic Changes
DISTINCTIVE FEATURES: Dysplastic changes of the erythroid cell line may include large RBCs, maturation
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asynchrony (fully hemoglobinized RBCs with retained nuclei), binucleate nRBCs, and nRBCs with frag-
mented nuclei and megaloblasts (extremely large nRBCs).
DIAGNOSTIC SIGNIFICANCE: Very mild erythroid dysplasia may occur in markedly regenerative states
and is associated with accelerated hematopoiesis. The most significant dysplastic changes are found with
myelodysplastic syndromes (MDSs), notably with FeLV-induced myelodysplasia in cats. Myelodysplas-
tic syndromes are associated with cytopenia(s) and ineffective hematopoiesis with evidence of cellular
dysplasia in peripheral blood and bone marrow. Often, an asynchronous regenerative response to the
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anemia exists, as polychromatophilic RBCs or reticulocytes are infrequent to absent, yet increased nRBCs
are present. Some MDSs have evidence of dysplasia in the myeloid and platelet cell lines as well.
NEXT STEPS: When dysplastic nRBCs are found, if the changes are mild and infrequent, and a signifi-
cant, markedly regenerative anemia exists, then, this is likely secondary to accelerated hematopoiesis.
Note that dysplasia may occur secondary to some disease states, toxins, and drugs. If erythropoiesis
is ineffective and many significant dysplastic changes have occurred, bone marrow cytology is advised
to confirm MDS. In cats, FeLV testing should be performed as well.
Plate 2-32 Dysplastic Changes
Binucleate nRBC
Fragmented nucleus
MORPHOLOGIC CHANGES ASSOCIATED WITH DISEASE DYSPLASTIC CHANGES 91
Plate 2-32 Dysplastic Changes (con’t)

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Large nRBC
Large nRBC
91.e1
Next Topic
Plate 2-32a Binucleate nRBC
Plate 2-32b
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Plate 2-32c
Plate 2-32d
Plate 2-32e
Plate 2-32f
Plate 2-32g
Plate 2-32h
Plate 2-32i
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Plate 2-32j
Plate 2-32a
Fragmented nucleus


91.e2
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Large nRBC
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Large nRBC

Plate 2-32j
Erythrocytic Parasites
Mycoplasma hemocanis (previously Hemobartonella canis)
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DISTINCTIVE FEATURES: Mycoplasma hemocanis (Hemobartonella canis) organisms are gram-positive para-
sites of dogs found on the surface of the RBC and generally appear as blue-black, filamentous chains of
small rods or dots. When found on the blood smear, the organisms are often present in high numbers.
DIAGNOSTIC SIGNIFICANCE: Infection with Mycoplasma hemocanis typically only causes significant
disease in dogs that have been splenectomized, have severe splenic disease, or are immunocompromised
and thus have moderate to severe hemolytic anemia. Occasional spherocytes and positive Coombs test
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suggest an immune-mediated component to the anemia.

NEXT STEPS: If the patient is known to be splenectomized, antibiotic treatment is warranted. If the
patient is not splenectomized, assessment for disease of the spleen or causes of immunosuppression
along with antibiotic treatment is recommended. Polymerase chain reaction (PCR) testing on the
blood may be used to confirm infection by identification of the presence of organism-specific nucleic
acid (deoxyribonucleic acid [DNA], ribonucleic acid [RNA]).
Plate 2-33 Mycoplasma hemocanis
Babesia gibsoni
Mycoplasma
hemocanis
Babesia
gibsoni
ERYTHROCYTIC PARASITES MYCOPLASMA HEMOCANIS (PREVIOUSLY HEMOBARTONELLA CANIS) 93
Plate 2-33 Mycoplasma hemocanis (con’t)

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Make sure not to confuse Mycoplasma hemocanis with A, Stain precipitate

B, Extracellular bacteria
A B
93.e1

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Plate 2-33e
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Babesia gibsoni
Mycoplasma
hemocanis

Babesia
gibsoni


93.e2
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A B
Mycoplasma hemofelis (previously Hemobartonella felis)

DISTINCTIVE FEATURES: Mycoplasma hemofelis (Hemobartonella felis) organisms are gram-positive, epicel-
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lular parasites of cats. The organisms form small dots or rods on the surface of the RBC. They stain
blue-black and tend to occur on the periphery of erythrocytes. Occasional chains of dots and ring forms
may be seen.
DIAGNOSTIC SIGNIFICANCE: Infection with Mycoplasma hemofelis may be primary or secondary. Primary
disease results in an acute, moderate to severe anemia with a marked regenerative response. As in dogs
with M. hemocanis infection, an immune-mediated component to the anemia may exist (autoagglutination
and positive Coombs test). Many cats are subclinically infected, and the parasitemia is only recognized
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when the patient becomes immunocompromised by another disease process (i.e., neoplasia, FeLV infec-
tion) resulting in secondary disease. In these cases, the anemia is often nonregenerative or poorly regen-
erative. Some of the most severe anemias occur with concurrent FeLV and Mycoplasma infection. Note
that not finding Mycoplasma organisms during a routine slide evaluation does not rule out the possibility
of infection.
Mycoplasma organisms are present on the outside of the erythrocyte; therefore, blood smears should
be made soon after blood collection, as the parasites may dislodge from the erythrocyte membrane. In
this case, the organisms may be found free in the extracellular space on blood films but are extremely
difficult to distinguish from stain precipitate. Thus, if confirmation of the parasite is sought from an
outside consultant, premade blood smears should be submitted along with an EDTA tube of whole blood.
NEXT STEPS: If Mycoplasma organisms are confirmed on blood smear review, antibiotic treatment is
warranted. If suspected, PCR testing for Mycoplasma species is recommended. Rule out concurrent
disease (i.e., neoplasia, FeLV, and FIV infection) and check the blood film for autoagglutination of
RBC to rule out a concurrent immune-mediated component to the anemia.
ERYTHROCYTIC PARASITES MYCOPLASMA HEMOFELIS (PREVIOUSLY HEMOBARTONELLA FELIS) 95
Plate 2-34 Mycoplasma hemofelis

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Plate 2-34 Mycoplasma hemofelis (con’t)

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platelet
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ERYTHROCYTIC PARASITES MYCOPLASMA HEMOFELIS (PREVIOUSLY HEMOBARTONELLA FELIS) 97
Plate 2-34 Mycoplasma hemofelis (con’t)

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Make sure not to confuse Mycoplasma hemofelis with A, stain precipitate

B, extracellular bacteria
A B
97.e1
Next Topic Plate 2-34g

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Plate 2-34e Plate 2-34k
Plate 2-34f
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platelet

97.e2
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A B
Cytauxzoon felis
DISTINCTIVE FEATURES: Cytauxzoon felis is a protozoal hemoparasite of the cat and is readily identified
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by its “signet ring” appearance. Lower numbers of “safety pin” and “Maltese cross” forms may be
observed. Cytauxzoon felis is an intraerythrocytic parasite that is generally present in only a small percent-
age (approximately 1%) of erythrocytes. In the terminal stages of the disease, organisms may be present
in up to 25% of the erythrocytes. Rarely, large macrophages containing schizonts full of developing
merozoites are observed on the feathered edge of the blood smear.
DIAGNOSTIC SIGNIFICANCE: Cytauxzoonosis is a highly fatal disease of domestic cats. Nonfatal forms
have been identified in some parts of the country. The severe clinical signs (depression, fever, anorexia,
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icterus) are secondary to the tissue phase of infection (infection of the spleen, liver, lung, lymph nodes,
and bone marrow). The associated anemia is usually nonregenerative, and concurrent thrombocytopenia
and leukopenia may exist.
NEXT STEPS: Cytauxzoon felis organisms are differentiated from Mycoplasma hemofelis on peripheral
blood smears by finding signet ring–shaped organisms with generally a single organism per eryth-
rocyte, as opposed to finding multiple, dot- or rod-shaped organisms on the periphery of the RBC,
respectively.
ERYTHROCYTIC PARASITES CYTAUXZOON FELIS 99
Plate 2-35 Cytauxzoon felis

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Cytauxzoon organism free

in background of smear
H-J body
Plate 2-35 Cytauxzoon felis (con’t)

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Macrophage at feathered edge of

blood smear full of developing
cytauxzoon organisms
ERYTHROCYTIC PARASITES CYTAUXZOON FELIS 101
Make sure not to confuse Cytauxzoon felis organisms with

Howell-Jolly (H-J) bodies
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H-J body
Cytauxzoon
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Cytauxzoon H-J body
Cytauxzoon
101.e1
Next Topic Plate 2-35j

Plate 2-35d Plate 2-35k
Plate 2-35e Plate 2-35l
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Plate 2-35f Plate 2-35m

Plate 2-35h Plate 2-35n
Plate 2-35i
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H-J body

Plate 2-35f Plate 2-35h

Macrophage at feathered edge of

blood smear full of developing
cytauxzoon organisms

101.e2
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Plate 2-35k Plate 2-35l

Cytauxzoon organism free

in background of smear
Plate 2-35m Plate 2-35n

H-J body
Cytauxzoon
Cytauxzoon H-J body
Cytauxzoon

Babesia spp.
DISTINCTIVE FEATURES: Babesia organisms are intraerythrocytic protozoal parasites. In dogs, two forms
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are distinguished by size: (1) Babesia canis are large organisms that appear as singular or paired piro-
plasms and may be oval, pear shaped, or ovoid. Currently, three subspecies of Babesia canis (subspecies:
canis, vogeli, and rossi) have been identified. The different subspecies have variable virulence, with clinical
signs ranging from mild to severe. The second form of Babesia organisms are small and form single to
multiple signet rings within infected RBCs. To date, three main species of small Babesia have been identi-
fied: (1) B. gibsoni, (2) B. conradae, and (3) B. microti–like piroplasms (Theileira annae). With both large and
small Babesia, the organism’s cytoplasm tends to stain clear to blue, whereas the nucleus stains red to
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purple.
DIAGNOSTIC SIGNIFICANCE: Babesia organisms are intracellular and cause hemolytic anemia with an
intravascular component, often with a concurrent thrombocytopenia. Spherocytes and a positive Coombs
test may indicate an immune-mediated component. Primary babesiosis generally results in a strongly
regenerative anemia. In some animals, the infection is subclinical until the animal becomes immunosup-
pressed with another disease. In the later circumstance, the anemia may be nonregenerative. Clinical
signs of babesiosis range from subclinical to mild to severe. Acute disease is often associated with signs
of depression, fever, weakness, lymph node enlargement, splenic enlargement, and anemia.
NEXT STEPS: The presence of intraerythrocytic Babesia organisms on blood smear review is diagnostic
for canine babesiosis. The absence of organisms does not rule out infection that may be subclinical.
The two general categories of Babesia, the large and small forms, may be identified microscopically.
However, further categorization of species and subspecies may only be performed by using molecular
techniques (PCR and DNA sequencing).
When Babesia organisms are identified, review the blood film carefully for evidence of concurrent
IMHA (RBC agglutination and spherocytes), and assess for thrombocytopenia as well.
ERYTHROCYTIC PARASITES BABESIA SPP. 103
Plate 2-36 Babesia canis

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Make sure not to confuse Babesia organisms with A, Canine Distemper Inclusions
B, Howell-Jolly bodies C, Basophilic stippling
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A B C
Plate 2-37 Babesia gibsoni
H-J body
ERYTHROCYTIC PARASITES BABESIA SPP. 105
Plate 2-37 Babesia gibsoni (con’t)

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Babesia
gibsoni
H-J body
105.e1
Next Topic
Plate 2-36a
Plate 2-36b
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Plate 2-36c
Plate 2-36d
Plate 2-36e
Plate 2-36h
Plate 2-36i
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Plate 2-36a


105.e2
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A B C
105.e3
Next Topic
Plate 2-37a
Plate 2-37b
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Plate 2-37c
Plate 2-37d
Plate 2-37e
Plate 2-37f
Plate 2-37g
Plate 2-37h H-J body
Plate 2-37i
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Plate 2-37j
Plate 2-37k
Plate 2-37l
Plate 2-37a


105.e4
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Babesia
gibsoni
H-J body


105.e5
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Plate 2-37l
Red Blood Cell Artifacts

Refractile Artifact
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DISTINCTIVE FEATURES: Refractile artifact is more commonly seen in Diff-Quik–stained smears as a

slightly irregular, refractile clear spot seen on RBCs upon adjusting the fine focus of the microscope.
DIAGNOSTIC SIGNIFICANCE: Do not confuse refractile artifact with an RBC parasite or inclusion.
Plate 2-38 Refractile Artifact of Red Blood Cell

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RED BLOOD CELL ARTIFACTS REFRACTILE ARTIFACT 107
Plate 2-38 Refractile Artifact of Red Blood Cell (con’t)

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107.e1
Next Topic
Plate 2-38b
Plate 2-38c
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Plate 2-38d
Plate 2-38e
Plate 2-38f
Plate 2-38g
Plate 2-38h
Plate 2-38i
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Plate 2-38b


107.e2
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Plate 2-38g
Plate 2-38h
Plate 2-38i
Platelet over Red Blood Cell

DISTINCTIVE FEATURES: Occasionally platelets will overlie RBCs and may be readily identified by adjust-
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ing the fine focus of the microscope and comparing the size and morphology to other platelets on the
smear.
DIAGNOSTIC SIGNIFICANCE: Do not confuse platelets overlying RBCs with hemoparasites or inclusions.
Plate 2-39 Platelets Overlying Red Blood Cells

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RED BLOOD CELL ARTIFACTS PLATELET OVER RED BLOOD CELL 109
Plate 2-39 Platelets Overlying Red Blood Cells (con’t)

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109.e1
Plate 2-39a
Plate 2-39b
Plate 2-39c
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Plate 2-39e
Plate 2-39f
Plate 2-39g
Plate 2-39h
Plate 2-39i
Plate 2-39j
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Plate 2-39a


109.e2
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SECTION 3:
WHITE BLOOD CELLS
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Normal Leukocyte Count, 112 Granular Lymphocytes (Large Granular

Lymphocytes), 162
Leukocytosis, 114
Monocytes, 164
Leukopenia, 116
Eosinophils, 166
Neutrophils, 118
Basophils, 172
Mature (Segmented) Neutrophils, 118
Band Neutrophils, 120
Mast Cells, 176
Metamyelocytes, 124 Sideroleukocytes, 180
Myelocytes, 126 Erythrophage (Erythrophagocytosis), 182
Promyelocytes, 128 Macrophage, 184
Myeloblasts, 130 Inclusions, Parasites, and Infectious
Barr Body, 132 Agents, 186
Hypersegmented Neutrophils, 134 Distemper Inclusions, 186
Pelger-Huët and Pseudo-Pelger-Huët Ehrlichia/Anaplasma spp., 188
Neutrophils, 136 Hepatozoon spp., 192
Toxic Changes, 140 Histoplasma capsulatum, 194
Döhle Bodies, 140 Mucopolysaccharidosis, 196
Diffuse Cytoplasmic Basophilia, 144 Chédiak-Higashi Syndrome, 198
Cytoplasmic Vacuolization (Foamy Birman Cat Anomaly, 200
Cytoplasm), 146 GM Gangliosidosis, 202
Toxic Granulation, 148
White Blood Cell Artifacts, 204
Ring (Doughnut) Form, 150
Pyknotic Cells, 204
Giant Neutrophils, 152
Basket or Smudge Cells, 206
Lymphocytes, 154 Platelet over White Blood Cell, 208
Small Lymphocytes, 154 Changes Associated with Delayed
Reactive Lymphocytes, 158 Processing, 210
Plasmacytoid Reactive Lymphocytes, 160 Understained Smears, 212
111
112 SECTION 3 WHITE BLOOD CELLS
Normal Leukocyte Count

DIAGNOSTIC SIGNIFICANCE: A scan of the blood film may determine relative numbers of blood leuko-
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cytes and may be used to provide quality control of complete blood count (CBC) results obtained by
hematology analyzers. The presence of normal numbers of leukocytes does not rule out disease of infec-
tious, inflammatory, or neoplastic origin, emphasizing the need for careful microscopic review of cellular
morphology.
Plate 3-1 Normal Leukocyte Count

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NORMAL LEUKOCYTE COUNT 113
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Leukocytosis
DIAGNOSTIC SIGNIFICANCE: Leukocytosis is defined as the presence of increased leukocyte numbers,
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which may be caused by an increase in the number of one cell type or a combination of cell types. The
significance of a leukocytosis is based on the degree of elevation, the cell types constituting the increase,
and the cellular morphology (normal, dysplastic, toxic, neoplastic). Increased numbers of neutrophils
secondary to inflammation is the most common cause of a leukocytosis.
Plate 3-2 Leukocytosis

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LEUKOCYTOSIS 115
Plate 3-2 Leukocytosis (con’t)

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Leukopenia
DIAGNOSTIC SIGNIFICANCE: Leukopenia is defined as a decrease in leukocyte numbers, which is most
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often caused by a decrease in the number of neutrophils (neutropenia), as neutrophils are the most
numerous of the peripheral blood leukocytes. Careful exam of the entire blood smear is emphasized by
the fact that sometimes during blood smear preparation, the leukocytes can be concentrated on the
feathered edge. In this case, the monolayer will contain very few leukocytes, suggesting an artifactual
leukopenia if the monolayer only, and not the entire slide, is examined for WBC estimation.
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Plate 3-3 Leukopenia

LEUKOPENIA 117
Plate 3-3 Leukopenia (con’t)

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Artifactual leukopenia. A, Monolayer, devoid of leukocytes

B, Same smear where leukocytes are concentrated on the feathered edge
A B
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Neutrophils
Neutrophils are the most abundant of the granulocytic cell types. Maturation stages from least mature
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to most mature are: myeloblasts, promyelocytes, myelocytes, metamyelocytes, band neutrophils, and
mature (segmented) neutrophils. As cells mature, they decrease in size, the cytoplasm becomes less blue
and eventually becomes clear to lightly colored, and the nucleus becomes more condensed and eventu-
ally lobulated.
Neutrophils are released from bone marrow into circulation in an age-related fashion, with the most
mature cells (mature neutrophils) released first. The bone marrow has a storage pool of mature neutro-
phils, so immature forms are not released into the circulation until the demand (severe, acute infection
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and inflammation) for neutrophils exceeds the storage pool’s ability to supply adequate numbers of
mature cells. Increased numbers of immature neutrophils in the peripheral blood is referred to as a “left
shift” and is the hallmark of acute inflammation. Immature neutrophils are also released in an age-related
fashion, and as such, the immature neutrophil stage primarily seen in peripheral blood during a left shift
is the band neutrophil, with less mature stages seen only rarely. In general, the more severe the systemic
inflammation, the higher the number of neutrophils with increased numbers of immature forms present.
The morphology and diagnostic significance of the various neutrophil maturational stages and the
abnormalities that occur in the neutrophil series are discussed below.
Mature (Segmented) Neutrophils
DISTINCTIVE FEATURES: Mature neutrophils of dogs and cats have similar morphologies. Neutrophils are
round and approximately 10 to 12 micrometers (µm) in diameter. The cytoplasm is clear, moderately
abundant, and may contain a few very indistinct, faint pink granules, and/or small clear vacuoles, or
both. The nucleus is highly condensed and lobulated, generally containing three or four lobules. Nuclear
lobes may be connected by a thin strand of chromatin, but more commonly, simply a narrowing of the
chromatin exists between lobes to about one third the diameter of the thick portion of the lobules.
DIAGNOSTIC SIGNIFICANCE: Mature neutrophil numbers are increased (neutrophilia) in conditions such
as epinephrine release (from excitement), endogenous steroids (stress resulting in glucocorticoid release),
steroid administration, inflammation (infectious and noninfectious causes), tissue necrosis, and neopla-
sia. Mature neutrophilia caused by inflammation is often associated with an increase in band neutrophils
(left shift). Leukemoid response is a term used to describe a marked neutrophilia, often with a left shift,
which may be seen with severe, acute, focal inflammation (i.e., pyometra, peritonitis, pyothorax, severe
abscessation). A persistently increasing, moderate to marked neutrophilia, in which a source of inflam-
mation, infection, tissue destruction, or necrosis cannot be found, should raise the suspicion for chronic
granulocytic leukemia.
Mature neutrophil numbers are decreased (neutropenia) by conditions such as overwhelming bacte-
rial infections, viral infections (parvovirus, panleukopenia, feline leukemia virus [FeLV]), certain bacte-
rial infections (Salmonella), rickettsial infections (i.e., chronic ehrlichiosis), endotoxemia (infection with
gram-negative bacteria), chemotherapeutic agents, bone marrow necrosis, myelophthesic diseases (bone
marrow infiltration with neoplastic cells), and bone marrow suppression or failure (i.e., drug-induced
neutropenia, immune-mediated neutropenia, cyclic hematopoiesis).
Plate 3-4 Normal Mature Segmented Neutrophils

NEUTROPHILS MATURE (SEGMENTED) NEUTROPHILS 119
Plate 3-4 Normal Mature Segmented Neutrophils (con’t)

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Band Neutrophils
DISTINCTIVE FEATURES: Band neutrophils are similar in size and appearance to mature neutrophils except
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that the nuclei are nonsegmented (band shaped) or only slightly constricted, and nuclear chromatin is
not as tightly clumped. The classic band neutrophil has a plump C-shaped or S-shaped nucleus, with
parallel sides, a smooth nuclear margin, and a fairly constant width. If slight constricted areas are present,
they are more than one third the width of the rest of the nucleus, distinguishing the bands from mature
neutrophils, which have constricted areas of less than one third the width of other areas of the nucleus.
The nuclear chromatin is less dense than that of a mature neutrophil. The cytoplasm remains clear unless
toxic change is present.
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DIAGNOSTIC SIGNIFICANCE: Band neutrophils are either absent or present in very low numbers in
healthy dogs and cats. A left shift is the hallmark of acute inflammation. Systemic inflammatory media-
tors travel to bone marrow, resulting in the production and release of neutrophils. Despite this systemic
response, the inflammation may be focal (i.e., pyometra) or diffuse (i.e., bacterial sepsis). Additionally,
not all focal areas of inflammation induce a systemic response. For example, cystitis (inflammation of
the bladder) is not generally associated with an increased number of band neutrophils. Chronic granu-
locytic leukemia may also cause an increase in band neutrophil numbers. Also, pseudo–band neutrophils
(hyposegmented mature neutrophils) may be seen in Pelger-Huët anomaly (discussed later).
A left shift may be regenerative or degenerative. A regenerative left shift is a predominance of mature
neutrophils (neutrophilia) with fewer bands, indicating a normal response to inflammation. A degenera-
tive left shift occurs when bands (and often more immature forms of neutrophils) exceed the number of
mature neutrophils. Often, a concurrent neutropenia exists. This indicates neutrophil production is not
able to meet demand and is associated with a guarded or poor prognosis. Toxic changes are frequent
with degenerative left shifts.
As band neutrophils are the most common immature neutrophil released into peripheral blood and
cannot be distinguished from mature neutrophils with current hematology analyzers, blood smear evalu-
ation is extremely important in identifying and quantitating a left shift.
NEUTROPHILS BAND NEUTROPHILS 121
Plate 3-5 Band Neutrophils

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Large
platelet
Plate 3-5 Band Neutrophils (con’t)

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Stain
precipitate
NEUTROPHILS BAND NEUTROPHILS 123
A, Band neutrophils. Note the open chromatin

B, Neutrophils with Pelger-Huët Anomaly. Note the condensed chromatin
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platelet


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Metamyelocytes
DISTINCTIVE FEATURES: Metamyelocytes are about the same size to slightly larger than mature and band
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neutrophils. The nuclei have one margin indented more than 25% into the nucleus and have an indented
or kidney-shaped nucleus. The nuclear chromatin is readily recognized as condensed but generally less
so than that of band neutrophils. The cytoplasm is nearly colorless but more basophilic than the nontoxic
band neutrophil.
DIAGNOSTIC SIGNIFICANCE: Metamyelocytes are rarely seen in the peripheral blood of healthy dogs
and cats. Metamyelocytes are sometimes seen in peripheral blood during severe inflammation along
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with band neutrophils as part of a left shift. Granulocytic leukemia may also cause an increase in meta-
myelocytes but occurs rarely. Also, pseudometamyelocytes (hyposegmented mature neutrophils) may
be seen in Pelger-Huët anomaly (discussed later).
Plate 3-6 Metamyelocytes

NEUTROPHILS METAMYELOCYTES 125
Plate 3-6 Metamyelocytes (con’t)

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Myelocytes
DISTINCTIVE FEATURES: Myelocytes are about the same size to slightly larger than metamyelocytes. The
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nuclei are often slightly indented, but the indentation is less than 25% the thickness of the nucleus. The
chromatin of metamyelocytes is only mildly condensed and often appears more open. The cytoplasm is
clear, pale blue, and may contain a few visible magenta-staining granules.
DIAGNOSTIC SIGNIFICANCE: Myelocytes are rarely seen in the peripheral blood of healthy dogs and
cats. A few myelocytes may be found in peripheral blood during severe inflammation along with band
neutrophils and metamyelocytes as part of a left shift. Chronic granulocytic leukemia may also cause an
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increase in myelocytes. Also, pseudomyelocytes (hyposegmented mature neutrophils) may be seen in

Pelger-Huët anomaly (discussed later).
NEUTROPHILS MYELOCYTES 127
Plate 3-7 Myelocytes

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Promyelocytes
DISTINCTIVE FEATURES: Promyelocytes are slightly larger than myelocytes. The nuclei are round to oval,
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with lacy to coarse chromatin without distinct clumps of condensed chromatin. Nuclei of a few promy-
elocytes may contain visible nucleoli or nuclear rings, but most do not. The cytoplasm is light blue and
contains numerous magenta-staining granules, the latter being a hallmark feature of promyelocytes.
DIAGNOSTIC SIGNIFICANCE: Promyelocytes are rarely seen in the peripheral blood of healthy dogs and
cats. A few promyelocytes may occasionally be seen in peripheral blood during severe inflammation
along with bands, metamyelocytes, and myelocytes as part of a left shift. Chronic granulocytic leukemia
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may also cause an increase in promyelocytes.
Plate 3-8 Promyelocytes

NEUTROPHILS PROMYELOCYTES 129
Plate 3-8 Promyelocytes (con’t)

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Myeloblasts
DISTINCTIVE FEATURES: Myeloblasts are difficult to distinguish from the primitive blasts of other hema-
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topoietic cell lines. They are about the same size as, or slightly smaller than, promyelocytes. They have
round to oval, centrally located nuclei that have finely stippled chromatin and one to several visible
nucleoli. A small to moderate amount of moderately basophilic cytoplasm may contain a few fine azu-
rophilic granules (primary granules).
DIAGNOSTIC SIGNIFICANCE: It is extremely rare to see myeloblasts in the peripheral blood smears of
healthy dogs and cats. Finding a myeloblast on a peripheral blood smear always raises the possibility
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of granulocytic leukemia; however, a myeloblast may rarely be seen on a peripheral blood smear during
severe inflammation, along with bands, metamyelocytes, myelocytes, and promyelocytes as part of a
left shift.
Plate 3-9 Myeloblasts

NEUTROPHILS MYELOBLASTS 131
Plate 3-9 Myeloblasts (con’t)

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Plate 3-9d Plate 3-9f

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Barr Body
DISTINCTIVE FEATURES: The nucleus of some neutrophils of females may have a single visible Barr body.
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A Barr body is a small, well-defined, “drumsticklike” nuclear appendage attached to the neutrophil
nucleus by a slender chromatin strand. Barr bodies are usually located near one end of the nucleus.
DIAGNOSTIC SIGNIFICANCE: Barr bodies represent the inactive X chromosome of the female. Rare Barr
body–like projections may be seen in males because of chance segmentation in the neutrophil. Barr bodies
have no diagnostic significance.
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Plate 3-10 Barr Bodies

NEUTROPHILS BARR BODY 133
Plate 3-10 Barr Bodies (con’t)

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Hypersegmented Neutrophils
DISTINCTIVE FEATURES: Neutrophils with five or more distinct nuclear lobes are classified as
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hypersegmented.
DIAGNOSTIC SIGNIFICANCE: Hypersegmentation is an aging change, which may occur as an in vitro
artifact secondary to delayed blood smear preparation (several hours) from ethylenediaminetetraacetic
acid (EDTA)–anticoagulated blood; a cellular aging change caused by prolonged blood transit time (i.e.,
chronic excess glucocorticoids), as an acquired defect (i.e., B12 or folic acid deficiency, FeLV myelodys-
plasia), or as an inherited maturation defect (i.e., macrocytosis of poodles).
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Plate 3-11 Hypersegmented Neutrophils

NEUTROPHILS HYPERSEGMENTED NEUTROPHILS 135
Plate 3-11 Hypersegmented Neutrophils (con’t)

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Pelger-Huët and Pseudo-Pelger-Huët Neutrophils

DISTINCTIVE FEATURES: Pelger-Huët and pseudo-Pelger-Huët neutrophils are mature neutrophils that
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are hyposegmented because of an inherited (Pelger-Huët) or acquired (pseudo–Pelger-Huët) defect in

granulocyte segmentation, which creates the appearance of a persistent left shift. The heterozygous form
of Pelger-Huët anomaly is seen in dogs and cats and is clinically benign as neutrophils function normally.
The homozygous state in utero is generally fatal. Hyposegmentation is recognized by the mature (coarsely
clumped) nuclear chromatin in relation to the lack of segmentation of the cell. Hyposegmentation occurs
in other peripheral blood leukocytes (i.e., eosinophils) but is most easily recognized in neutrophils
because of their predominant number and normally segmented nuclei. Nuclei of mature Pelger-Huët
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neutrophils may be round, oval, band-shaped, dumb-bell shaped, or bilobed. Lack of toxic change and
lack of illness or clinical signs in what appears to be a marked left shift are important in the identification
of Pelger-Huët anomaly. Pelger-Huët neutrophils are easily recognized if the animal is not sick; however,
if the animal is ill and Pelger-Huët anomaly has not been previously documented, then recognition of
Pelger-Huët neutrophils may be very difficult. If the patient becomes well following treatment, and the
left shift fails to resolve, then Pelger-Huët anomaly is most likely.
With resolving chronic inflammatory disease, an acquired pseudo-Pelger-Huët disorder is likely to
occur. This is very rare and is differentiated from true Pelger-Huët anomaly, as the nuclear hyposegmen-
tation resolves together with the inflammation.
DIAGNOSTIC SIGNIFICANCE: Careful morphologic assessment of cytoplasmic and nuclear features is
important in distinguishing inherited or acquired Pelger-Huët anomaly from a true left shift. Correlation
of the findings with the health status of the patient is very important. If the patient is sick, continued
assessment of the blood film over time may be necessary to rule out Pelger-Huët anomaly.
NEUTROPHILS PELGER-HUËT AND PSEUDO-PELGER-HUËT NEUTROPHILS 137
Plate 3-12 Pelger-Huët Anomaly

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Plate 3-12 Pelger-Huët Anomaly (con’t)

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NEUTROPHILS PELGER-HUËT AND PSEUDO-PELGER-HUËT NEUTROPHILS 139
A, Band neutrophils. Note the open chromatin

B, Neutrophils seen with Pelger-Huët Anomaly. Note the condensed chromatin
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Plate 3-12f
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A B
Toxic Changes
Systemic inflammation, most notably secondary to bacterial infections, may cause microscopic toxic
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changes in neutrophils. The most common toxic changes are cytoplasmic and include Döhle bodies, toxic
granulation, diffuse cytoplasmic basophilia, and cytoplasmic vacuolation. Toxic neutrophils may contain
only one or two of these changes, or all may be evident in severely toxic cells. During rapid neutrophil
production (neutropoiesis) in bone marrow in response to systemic inflammatory mediators, neutrophils
undergo specific maturation defects, which results in toxic changes. As with a left shift, the presence of
toxic neutrophils indicates systemic inflammation. Less common toxic changes involve the nucleus (i.e.,
ring forms) or affect cell size (i.e., giant neutrophils) and are rarely seen except with severe inflammation,
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often with a significant left shift.

Like band neutrophils and other immature neutrophil forms, toxic neutrophils are not recognized by
automated hematology analyzers and must be identified on blood film review. The different toxic
changes discussed below are generally graded as slight, mild, moderate, or marked; or on a 1 to 4+ scale.
In general, the more pronounced the toxic change and the higher the percentage of toxic neutrophils,
the higher is the grade.
Döhle Bodies
DISTINCTIVE FEATURES: Döhle bodies are variably sized, variably shaped, light blue to slate gray struc-
tures in the cytoplasm of neutrophils. These foci are clumps of endoplasmic reticulum that are usually
degraded during maturation, yet retained during accelerated neutropoiesis.
DIAGNOSTIC SIGNIFICANCE: Döhle bodies are the most easily produced and, therefore, the most
common form of toxic change observed in neutrophils. Regardless of the number of neutrophils contain-
ing Döhle bodies, when occurring alone without other toxic changes, Döhle bodies should never be
interpreted as meaning anything more than a mild toxic change in dogs. Döhle bodies may occur in
clinically healthy cats and, thus, are often not reported as a toxic change in cats unless these bodies are
numerous or present with other toxic changes. Make sure not to confuse Döhle with Ehrlichia morulae
and Distemper inclusions or platelets overlying WBCs.
NEXT STEPS: When Döhle bodies are evident in the dog or seen in high numbers in cats, assess for
other toxic changes and for the presence of immature neutrophils (left shift). Be careful to not confuse
Döhle bodies with Ehrlichia morulae, distemper inclusions, or parasites.
TOXIC CHANGES DÖHLE BODIES 141
Plate 3-13 Döhle Bodies

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Plate 3-13 Döhle Bodies (con’t)

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TOXIC CHANGES DÖHLE BODIES 143
A, Döhle bodies B, Canine neutrophil with Ehrlichia morulae

C, Distemper inclusions D, Platelet over neutrophil
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C D
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A B
C D
Diffuse Cytoplasmic Basophilia

DISTINCTIVE FEATURES: With diffuse cytoplasmic basophilia, the neutrophil cytoplasm stains basophilic
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(blue). Diffuse basophilia occurs when ribosomes remain scattered throughout the cytoplasm (as opposed
to being clumped into Döhle bodies), instead of being degraded during maturation. Diffuse cytoplasmic
basophilia is graded as a mild, moderate, or severe, depending on the number of neutrophils affected
and the severity of the basophilia.
DIAGNOSTIC SIGNIFICANCE: The presence of diffuse cytoplasmic basophilia indicates systemic inflam-
mation, regardless of whether a neutrophilia or a left shift exists (although it usually is associated with
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both).
Plate 3-14 Cytoplasmic Basophilia

TOXIC CHANGES DIFFUSE CYTOPLASMIC BASOPHILIA 145
Plate 3-14 Cytoplasmic Basophilia (con’t)

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Cytoplasmic Vacuolization (Foamy Cytoplasm)

DISTINCTIVE FEATURES: Neutrophils containing foamy appearing vacuoles within their cytoplasm are
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considered toxic. Toxic vacuolization generally is recognized in association with diffuse cytoplasmic
basophilia, as the basophilic background enhances the ability to discern the vacuoles. Toxic vacuolization
is graded as mild, moderate, or severe, depending on the number of neutrophils affected and the degree
of vacuolization.
Cytoplasmic vacuolization should not be confused with the discrete clear vacuoles that develop in
neutrophils exposed to EDTA for several hours. The rate of EDTA-induced cytoplasmic vacuolization is
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variable, depending on temperature and other factors, but may occur rather rapidly. Therefore, if cyto-
plasmic vacuolization is present without other evidence of inflammation (i.e., other toxic changes, left
shift), the possibility of artifactual cytoplasmic vacuolization should be considered.
DIAGNOSTIC SIGNIFICANCE: Toxic vacuolization occurs secondary to systemic inflammation or
artifactually.
NEXT STEPS: Determine how quickly blood smears were made from the EDTA blood tube. Prolonged
exposure of peripheral blood to EDTA before smear preparation is the most common cause of arti-
factual cytoplasmic vacuolization within neutrophils.
TOXIC CHANGES CYTOPLASMIC VACUOLIZATION (FOAMY CYTOPLASM) 147
Plate 3-15 Cytoplasmic Vacuolization

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Toxic Granulation
DISTINCTIVE FEATURES: Neutrophils containing low to high numbers of magenta-staining cytoplasmic
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granules are toxic. These are retained primary granules, which are not normally present in later stages
of neutrophil development (usually seen up to the progranulocyte stage). Depending on the number of
neutrophils affected, toxic granulation is interpreted as a mild, moderate, or severe.
DIAGNOSTIC SIGNIFICANCE Toxic granulation indicates systemic inflammation.
NEXT STEPS: If neutrophilic granulation is observed without evidence of other toxic changes, a left
shift, or both, and a source of inflammation or infection is not evident, consider uncommon hereditary
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diseases associated with neutrophil inclusions, for example, Chédiak-Higashi syndrome in blue-
smoke Persian cats (usually abnormal granules are also seen in eosinophils and basophils); GM2
gangliosidosis (German Shorthaired Pointers and some cats [dark blue granules are seen in neutro-
phils and azurophilic granules in lymphocytes]); Birman cat anomaly (seen in purebred Birman cats);
mucopolysaccharidosis (MPS) type VI (Siamese and Domestic Shorthair cats and Dachshunds); and
MPS type VII (neutrophilic inclusions are seen in dogs and cats with this disorder).
Plate 3-16 Toxic Granulation

TOXIC CHANGES TOXIC GRANULATION 149
Plate 3-16 Toxic Granulation (con’t)

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A, Toxic granulation B, Mucopolysaccharidosis C, Birman Cat Anomaly

D, Chédiak-Higashi Syndrome E, Primary neutrophil granules
A B C
D E
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Plate 3-16e
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A B C
D E
Ring (Doughnut) Form

DISTINCTIVE FEATURES: Immature neutrophils containing a round nucleus with a hole in the center
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(doughnut-shaped nucleus) without significant indentations in the nucleus are a rarely observed toxic
change involving the nucleus.
DIAGNOSTIC SIGNIFICANCE: Ring-form neutrophils are seen occasionally in cats and infrequently in
dogs with severe systemic inflammation or toxemia. This finding is considered a toxic change, and the
neutrophils are best classified as band neutrophils.
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Plate 3-17 Ring-Form Neutrophils

TOXIC CHANGES RING (DOUGHNUT) FORM 151
Plate 3-17 Ring-Form Neutrophils (con’t)

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Giant Neutrophils
DISTINCTIVE FEATURES: Giant (larger than normal) neutrophils and bands are considered a toxic change
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that affects cell size.

DIAGNOSTIC SIGNIFICANCE: Skipped cell divisions during accelerated neutrophil production and mat-
uration process may lead to the formation of large neutrophils and bands in the peripheral blood. This
is generally associated with other toxic changes.
NEXT STEPS: Giant neutrophils and bands are usually associated with other toxic changes. If other
evidence of toxicity is not present, then other causes of giant neutrophils, for example, poodle mac-
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rocytosis (poodle marrow dyscrasia syndrome), FeLV myelodysplasia in cats, and administration of
certain chemotherapeutics, should be investigated.
Plate 3-18 Giant Neutrophils

TOXIC CHANGES GIANT NEUTROPHILS 153
Plate 3-18 Giant Neutrophils (con’t)

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Lymphocytes
Lymphocytes vary in size in the peripheral blood of dogs and cats. Small lymphocytes predominate with
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a few lymphocytes that are slightly to moderately larger. Larger lymphocytes in peripheral blood have
less densely staining, but still clearly clumped, nuclear chromatin. Lymphocytes are typically the second
most common peripheral blood leukocyte (mature neutrophil being the most common).
Small Lymphocytes
DISTINCTIVE FEATURES: Small lymphocytes are mature cells, approximately 10 µm in diameter with
densely staining (dark purple), round to oval nuclei, which are sometimes slightly indented and usually
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have large, well-defined chromatin clumps (heterochromatin) with intermixed smooth glassy areas
(euchromatin). Occasionally, small lymphocytes may have nuclear chromatin, which appears smudged,
especially with quick stains. The nucleus is eccentric, and a scant amount of moderately blue cytoplasm
exists. Because of the eccentric placement of the nucleus, it is difficult to completely trace the rim of
cytoplasm around the nucleus.
DIAGNOSTIC SIGNIFICANCE: Small lymphocytes are the most abundant form of lymphocyte in canine
and feline blood. Increased numbers (lymphocytosis) may occur with epinephrine release secondary to
excitement (mild lymphocytosis is common in young dogs and cats); chronic immune stimulation, and
lymphoid neoplasia (chronic lymphocytic leukemia). Decreased numbers (lymphopenia) of lymphocytes
may occur with acute inflammation or, commonly, as a “stress” leukogram secondary to glucocorticoid
release.
NEXT STEPS: Be careful to not confuse small lymphocytes with nucleated red blood cells such as
rubricytes and metarubricytes. A rim of cytoplasm may usually be traced entirely around the nucleus
of nucleated RBCs but not in mature lymphocytes.
Distinguishing a lymphocytosis secondary to immunostimulation from chronic lymphocytic leukemia
may be difficult. Careful assessment for underlying infectious disease (especially ehrlichiosis in dogs),
correlation of the age of the patient and clinical signs (chronic lymphocytic leukemia is uncommon in
young animals), serial monitoring of lymphocyte absolute numbers and morphology, and, in some cases,
the use of adjunct diagnostics (immunostains and flow cytometry) should be employed.
LYMPHOCYTES SMALL LYMPHOCYTES 155
Plate 3-19 Small Mature Lymphocytes

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Lymphocyte
nRBC
Plate 3-19 Small Mature Lymphocytes (con’t)

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LYMPHOCYTES SMALL LYMPHOCYTES 157
A, Small mature lymphocyte. Note eccentric nucleus and

scant rim of cytoplasm B, Nucleated red blood cell. Note cytoplasm surrounding
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the more central nucleus

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nRBC
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Reactive Lymphocytes
DISTINCTIVE FEATURES: The primary features that identify a reactive lymphocyte are cytoplasmic
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changes. In general, reactive lymphocytes are larger because of a mild to moderate increase in the amount
of cytoplasm, and the cytoplasm stains more deeply basophilic compared with the cytoplasm of small,
mature lymphocytes. Additionally, the nuclear chromatin is less condensed. The deep cytoplasmic baso-
philia of reactive lymphocytes represents an abundance of polyribosomes associated with increased
protein synthesis and usually contrasts with the pale, perinuclear Golgi region.
DIAGNOSTIC SIGNIFICANCE: Reactive lymphocytes are immune-stimulated lymphocytes with upregu-
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lated synthesis of inflammatory mediators, and/or immunoglobulins (antibodies), or both. Reactive

lymphocytes in peripheral blood suggest active, systemic antigenic stimulation secondary to both infec-
tious and noninfectious disorders.
NEXT STEPS: It is important to differentiate reactive lymphocytes from neoplastic lymphocytes. In
general, reactive lymphocytes occurring secondary to systemic immune stimulation, are present
together with a wide range of lymphocyte morphology (small, mature lymphocytes; reactive lympho-
cytes; plasmacytoid lymphocytes; intermediate-sized lymphocytes; and occasionally also granular
lymphocytes). Neoplastic lymphocytes, as discussed in Section V: Hematopoietic Neoplasia, may be
small, intermediate sized, and blastic. If in high numbers, neoplastic lymphocytes are morphologically
similar, forming a uniform population. Further testing such as immunostaining and flow cytometry
may be needed to identify a neoplastic population of lymphocytes.
LYMPHOCYTES REACTIVE LYMPHOCYTES 159
Plate 3-20 Reactive Lymphocytes

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Plasmacytoid Reactive Lymphocytes

DISTINCTIVE FEATURES: With strong, chronic inflammation, reactive lymphocytes may become plasma-
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cytoid (plasma cells in peripheral blood). Plasma cells are well differentiated B lymphocytes. They are
larger than small lymphocytes and have an eccentrically placed round nucleus, with condensed nuclear
chromatin and deeply basophilic cytoplasm because of the abundant polyribosomes associated with
increased protein synthesis. Also, a perinuclear clear zone (Golgi region) is generally visible.
DIAGNOSTIC SIGNIFICANCE: Plasmacytoid reactive lymphocytes indicate strong, chronic, systemic
inflammation but are etiologically nonspecific.
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NEXT STEPS: If significant numbers of plasmacytoid lymphocytes are present, investigate for infec-
tious or inflammatory disease, rule out recent vaccination, and, if associated with hyperglobulinemia,
consider serum electrophoresis to characterize the immunoglobulin pattern, which aids in categoriz-
ing the inflammatory response and may also be used to check for lymphoid malignancy (lymphoid
leukemia, plasma cell neoplasia, lymphoma).
Plate 3-21 Plasmacytoid Reactive Lymphocytes

LYMPHOCYTES PLASMACYTOID REACTIVE LYMPHOCYTES 161
Plate 3-21 Plasmacytoid Reactive Lymphocytes (con’t)

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Proteinaceous
background
Large platelet
clumps
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Proteinaceous
background
Large platelet
clumps
Plate 3-21h Plate 3-21j

Granular Lymphocytes (Large Granular Lymphocytes)

DISTINCTIVE FEATURES: Large granular lymphocytes have a slightly larger nucleus and moderate amount
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of cytoplasm, which ranges from light to moderately basophilic and contains few, variably sized, eosino-
philic intracytoplasmic granules that may be dispersed throughout the cytoplasm or concentrated in a
perinuclear region of the cytoplasm.
DIAGNOSTIC SIGNIFICANCE: Large granular lymphocytes may be present in low numbers in the periph-
eral blood of clinically normal dogs and cats. Increased numbers may occur with chronic immune stimu-
lation (especially chronic ehrlichiosis in dogs). When moderate to high numbers of granular lymphocytes
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are present in peripheral blood and infectious disease has been ruled out, granular lymphocytic leukemia
or circulating neoplastic cells from lymphoma of granular lymphocyte origin should be investigated.
Plate 3-22 Granular Lymphocytes

LYMPHOCYTES GRANULAR LYMPHOCYTES (LARGE GRANULAR LYMPHOCYTES) 163
Plate 3-22 Granular Lymphocytes (con’t)

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Monocytes
DISTINCTIVE FEATURES: Canine and feline monocytes are larger than neutrophils and similar in size or
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slightly larger than eosinophils and basophils. Their nuclei vary greatly in morphology, ranging from
elongated U-shapes that resemble band neutrophils to irregular multilobulated forms. The nuclear chro-
matin is characteristically lacy to ropy, with only a few small isolated clumps of heterochromatin. The
cytoplasm is moderate to abundant, gray-blue with a ground-glass texture, often sparsely dusted with
minute eosinophilic granules, and occasionally lightly vacuolated. Cytoplasmic borders are usually
irregular, sometimes with fine, filamentous, pseudopodia-like extensions. Because of their relatively large
size, monocytes may be concentrated along the feathered edge, and their proportion underestimated in
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blood smear differential white blood cell (WBC) counts.

DIAGNOSTIC SIGNIFICANCE: A few monocytes are present in the peripheral blood of healthy dogs and
cats. Increased numbers of monocytes (monocytosis) may occur with disorders such as subacute and
chronic inflammation, excess endogenous or exogenous glucocorticoids (common in the dog but incon-
sistent in the cat), autoimmune hemolytic anemia (part of a leukemoid response to accelerated hemato-
poiesis), FIV infection (cat), and neoplasia of the monocytic cell line (rare). Decreased numbers of
monocytes (monocytopenia) is not diagnostically significant because of the normally low numbers of
monocytes in healthy dogs and cats.
In conditions such as severe bacterial infections or septicemia, some of the circulating monocytes may
have transformed into macrophages. Macrophages are often highly vacuolated with an increased amount
of lightly basophilic cytoplasm and convoluted nuclei. Sometimes, these cells are referred to as reactive
or toxic monocytes and yet are very rarely identified in peripheral blood.
Plate 3-23 Monocytes

MONOCYTES 165
Plate 3-23 Monocytes (con’t)

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Eosinophils
DISTINCTIVE FEATURES: Eosinophils are slightly larger than neutrophils, with nuclei that are less lobu-
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lated, often divided into only two distinct lobules, and with less condensed chromatin than those of
mature neutrophils. The cytoplasm of eosinophils is clear to faintly basophilic and contains large numbers
of prominent pink granules. Feline eosinophil granules are uniformly small and rod shaped. Canine
eosinophil granules are round and vary widely in number and size. Canine eosinophils occasionally
contain a single, large granule that may be mistaken for an inclusion body or organism. Eosinophils of
Greyhounds (and few other Sight Hound breeds) are peculiar because they may degranulate during
staining and appear vacuolated on smears (the so-called gray eosinophils of Greyhounds).
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DIAGNOSTIC SIGNIFICANCE: Eosinophils may usually be found in low numbers in the blood smears of
healthy dogs and cats. Increased numbers of eosinophils (eosinophilia) may occur with many hypersen-
sitivity diseases of allergic or parasitic origin, both of which could involve the skin (ectoparasites, flea
bite dermatitis), lungs (heartworm infection, lungworm infection, feline asthma), and gastrointestinal
tract (inflammatory bowel disease, intestinal parasites). Rarely, a paraneoplastic eosinophilia secondary
to the presence of a solid tumor (especially mast cell neoplasia) may be seen. A moderate to marked
eosinophilia, which is unexplained, may be seen with eosinophilic leukemia (rare). Excess glucocorti-
coids, either endogenous or exogenous, may cause a decrease in circulating eosinophils (eosinopenia).
Otherwise, due to low numbers of eosinophils normally in the dog and cat, an eosinopenia is not diag-
nostically significant.
NEXT STEPS: If an eosinophilia is detected, further assessment for underlying allergic, parasitic, or
neoplastic disease is warranted. Check the blood film carefully for any circulating mast cells and for
hemoparasites (microfilaria).
EOSINOPHILS 167
Plate 3-24 Canine Eosinophils

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Plate 3-25 Feline Eosinophils

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EOSINOPHILS 169
Plate 3-25 Feline Eosinophils (con’t)

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Basophil
eosinophil
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Basophil
eosinophil
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Plate 3-26 Grey Eosinophils of Sight Hounds

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eosinophil
Neutrophil
EOSINOPHILS 171
Plate 3-26 Grey Eosinophils of Sight Hounds (con’t)

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eosinophil
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Plate 3-26l
Basophils
DISTINCTIVE FEATURES: Basophils are the largest and least numerous of the mature granulocytic cell
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types and are often not seen on a standard 100-cell differential cell count in healthy dogs and cats. Baso-
phil nuclei are less densely staining, have fewer lobulations, and have a more elongated, ribbonlike
appearance compared with the nuclei of neutrophils. The basophil cytoplasm is pale to moderately blue-
gray to purple and typically contains granules. Occasionally, no granules may be visible. Granule size
and color vary between dogs and cats. In dogs, basophil granules are usually low in number (typically
<50 granules per cell) and stain dark blue to metachromatic. Canine basophils occasionally lack obvious
granules but are recognizable by their size, nuclear morphology, and cytoplasmic staining. In cats, baso-
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phils contain abundant oval, pale lavender to gray granules, although immature basophils may also
contain a few primary, dark purple granules.
DIAGNOSTIC SIGNIFICANCE: Basophils are rare in the peripheral blood of healthy dogs and cats.
Increased numbers of basophils (basophilia) are often associated with a concurrent eosinophilia, and
similar etiologies of hypersensitivity disease as for increased eosinophils apply for a basophilia. A mod-
erate to marked unexplained basophilia may occur secondary to basophilic leukemia (extremely rare).
Decreased numbers of basophils (basopenia) are not diagnostically significant because of the low numbers
of basophils in the peripheral blood of healthy dogs and cats.
NEXT STEPS: Be careful to not confuse mast cells with basophils. Mast cells are larger than basophils
and often will be found concentrated on the feathered edge of the smear. Mast cells usually contain
large numbers of granules compared with basophils, and mast cell nuclei are round to ovoid, whereas
basophil nuclei are ribbonlike.
Plate 3-27 Canine Basophils

BASOPHILS 173
Plate 3-27 Canine Basophils (con’t)

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A, Canine basophil B, Canine mast cell

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A B
Plate 3-28 Feline Basophils

BASOPHILS 175
Plate 3-28 Feline Basophils (con’t)

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A, Feline basophil B, Feline mast cell
A B
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A B
Mast Cells
DISTINCTIVE FEATURES: Mast cells are large mononuclear cells that have a moderate to abundant amount
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of pale blue cytoplasm. The nucleus is round to oval, and the cytoplasm contains many small, round,
intensely staining, intracytoplasmic metachromatic (reddish-purple) granules. Granule numbers range
from few to many, but typically high numbers of granules are present and may be so abundant as to fill
the cytoplasm, obscuring the nucleus. Poorly differentiated mast cells may have low numbers of intra-
cytoplasmic granules.
DIAGNOSTIC SIGNIFICANCE: Rarely, a mast cell may be seen on the peripheral blood smear of a healthy
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dog or cat. Mast cells have been observed in the peripheral blood of dogs and cats with mast cell neo-
plasia and in very low numbers on blood smears from dogs with severe inflammatory conditions. Large
numbers of mast cells may be seen with mast cell leukemia (very uncommon).
NEXT STEPS: Be careful to not confuse mast cells with basophils. Mast cells are larger than basophils
and often will be found concentrated on the feathered edge of the smear. Mast cells usually contain
large numbers of granules compared with basophils, and mast cell nuclei are round to ovoid, whereas
basophil nuclei are ribbonlike.
Finding circulating mast cells on peripheral blood smears from dogs and especially cats warrants a
thorough search for underlying mast cell neoplasia of the skin, liver, spleen, and gastrointestinal tract.
MAST CELLS 177
Plate 3-29 Mast Cells

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Plate 3-29 Mast Cells (con’t)

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MAST CELLS 179
A, Feline basophil B, Feline mast cell

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A B
C, Canine basophil D, Canine mast cell
C D
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A B
C D
Sideroleukocytes
DISTINCTIVE FEATURES: Sideroleukocytes are neutrophils or monocytes containing intracytoplasmic,
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yellowish-brown to blue-black hemosiderin pigment.

DIAGNOSTIC SIGNIFICANCE: Sideroleukocytes are rarely observed on peripheral blood smears with
disorders such as immune-mediated hemolytic anemia (IMHA) and following blood transfusions.
Hemosiderin is an iron-containing pigment, and sideroleukocytes may be confirmed by staining a blood
film with an iron stain such as Prussian blue (rarely done).
NEXT STEPS: If sideroleukocytes are identified and a blood transfusion has not been performed, top
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consideration is IMHA and warrants close review of the blood film for ghost RBCs, spherocytes, RBC
agglutination, and hemoparasites.
Plate 3-30 Sideroleukocytes

SIDEROLEUKOCYTES 181
Plate 3-30 Sideroleukocytes (con’t)

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Erythrophage (Erythrophagocytosis)
DISTINCTIVE FEATURES: An erythrophage is a leukocyte (usually a neutrophil or monocyte) containing
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engulfed RBCs (erythrophagocytosis).

DIAGNOSTIC SIGNIFICANCE: Erythrophagocytosis is uncommonly identified on peripheral blood
smears and may be observed with immune-mediated diseases and some leukemias.
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Plate 3-31 Erythrophages

ERYTHROPHAGE (ERYTHROPHAGOCYTOSIS) 183
Plate 3-31 Erythrophages (con’t)

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Macrophage
DISTINCTIVE FEATURES: Macrophages are inflammatory cells found in tissue and only very uncommonly
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in the peripheral blood of healthy dogs and cats. Macrophages are large mononuclear cells with abundant
clear cytoplasm and round nuclei with coarse chromatin and are often vacuolated and sometimes cyto-
phagic. They may be uniform but, in some neoplastic conditions, may have criteria of malignancy (large
cell and nuclear size, multiple nuclei, prominent nucleoli).
DIAGNOSTIC SIGNIFICANCE: Monocytes are the peripheral blood precursors to tissue macrophages.
Monocytes leave peripheral blood secondary to inflammatory mediators and develop into tissue
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macrophages. Macrophages have phagocytic activity and may engulf foreign material, cells, infectious
agents (especially fungal organisms), and debris. If macrophages are found in peripheral blood smears,
they are usually on the feathered edge because of their large size.
NEXT STEPS: Macrophages are rarely found in the peripheral blood of healthy animals; however, when
collecting blood from a patient, if a regional source of inflammation or infection is present, concurrent
sampling of subcutaneous tissue may occasionally result in low numbers of tissue macrophages in
blood. If macrophages demonstrate criteria of malignancy, then histiocytic neoplasia should be
strongly suspected and would warrant assessment for histiocytic neoplasia of the lymph nodes, liver,
spleen, lungs, and bone marrow.
MACROPHAGE 185
Plate 3-32 Macrophages

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Inclusions, Parasites, and Infectious Agents

Distemper Inclusions
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DISTINCTIVE FEATURES: Distemper inclusions are aggregates of viral nucleocapsids that appear as round
to oval to irregular, red to blue, cytoplasmic structures. The inclusions tend to occur in either the mono-
cyte and lymphocyte cell series or the neutrophil and erythrocyte cell series. Distemper inclusions are
most easily identified on Diff-Quik–stained smears as bright, homogeneous, eosinophilic, round to ovoid
cytoplasmic structures. Distemper inclusions may be seen on Wright-stained smears; however, they tend
to stain pale blue and are somewhat more difficult to identify.
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DIAGNOSTIC SIGNIFICANCE: Distemper inclusions are seen only rarely on peripheral blood smears from
dogs with distemper; however, when present, they are diagnostic for infection. These inclusions are
found in a small percentage of dogs during the acute viremic phase of distemper, which is usually associ-
ated with clinical signs of upper respiratory disease. The lack of identifiable inclusions does not rule out
distemper infection, and if suspected clinically, warrants further testing. Make sure to distinguish Dis-
temper inclusions from other inclusions and artifacts such as Döhle bodies, Ehrlichia morulae and
platelets overlying WBCs.
Plate 3-33 Distemper Inclusions

Plate 3-33 Distemper Inclusions (con’t)
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A B
C D
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A B
C D
Ehrlichia/Anaplasma spp.
DISTINCTIVE FEATURES: Ehrlichiosis is a group of tickborne diseases caused by a gram-negative intracel-
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lular bacterium that infects blood cells. The disease is categorized by the host and the type of blood cell
infected. On light microscopy, Ehrlichia organisms form aggregates within a cytoplasmic vacuole called
a morula. Morulae are large, round to oval clusters composed of small dotlike organisms, which stain as
a stippled dark blue inclusion. One or more morulae may be present in a cell.
Canine monocytic ehrlichiosis is caused primarily by Ehrlichia canis and less commonly by Ehrlichia
chaffeensis. Morulae are found primarily within the cytoplasm of monocytes. The disease may be acute
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or chronic. Clinical signs may be mild to severe and may include fever, lethargy, anorexia, enlarged
lymph nodes and spleen, bleeding disorders, vomiting or diarrhea, ocular changes, and neurologic signs.
CBC abnormalities may include one or all of the following: nonregenerative anemia, thrombocytopenia,
and neutropenia.
Canine granulocytic ehrlichiosis is caused by Ehrlichia ewingii and Anaplasma phagocytophilum (for-
merly Ehrlichia equi and Ehrlichia phagocytophilia). Morulae are found primarily in the cytoplasm of
neutrophils. Disease, clinical signs, and CBC changes are essentially the same as in canine monocytic
ehrlichiosis; however, suppurative polyarthritis is more commonly seen with granulocytic ehrlichiosis.
Rare cases of cats infected with Anaplasma phagocytophilum have been documented.
DIAGNOSTIC SIGNIFICANCE: Identification of morulae is diagnostic for ehrlichiosis or anaplasmosis.
However, morulae are not seen in the vast majority of cases and may be present in low numbers of cells.
If infection is suspected, further testing (serology, polymerase chain reaction [PCR]) may be needed for
diagnosis. Also, further testing for other tickborne disease is recommended to rule out co-infections.
NEXT STEPS: Be careful to not confuse artifacts (i.e., platelet overlying leukocyte) or other inclusions
(i.e., Döhle bodies) with morulae.
INCLUSIONS, PARASITES, AND INFECTIOUS AGENTS EHRLICHIA/ANAPLASMA SPP. 189
Plate 3-34 Ehrlichia Morula

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Plate 3-34 Ehrlichia Morula (con’t)

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INCLUSIONS, PARASITES, AND INFECTIOUS AGENTS EHRLICHIA/ANAPLASMA SPP. 191

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A B
C D
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Plate 3-34b
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Plate 3-34f
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191.e2
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A B
C D
Hepatozoon spp.
DISTINCTIVE FEATURES: Hepatozoon spp. are protozoan parasites of dogs, with infection resulting from
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ingestion of infected ticks. The organism forms large (up to 11 µm) gametocytes, which are oval to ellipti-
cal, clear to pale blue structures within the cytoplasm of neutrophils, monocytes, or both. The organism
typically displaces the nucleus to one side of the cell.
DIAGNOSTIC SIGNIFICANCE: Hepatozoonosis is caused by Hepatozoon canis and Hepatozoon americanum
and is a severe disease that often results in death. Clinical signs are related to severe cardiac and skeletal
muscle infection (myositis), with clinical signs that include fever, weight loss, muscle loss or wasting,
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and skeletal pain. CBC findings are often remarkable, as hepatozoonosis is frequently associated with a
marked mature neutrophilic leukocytosis with or without a left shift. Gametocytes of H. canis are seldom
found in peripheral blood leukocytes; however, gametocytes of H. americanum are frequently present.
NEXT STEPS: Identification of gametocytes in the cytoplasm of canine peripheral blood neutrophils
and monocytes is diagnostic for hepatozoonosis. Lack of identification of the parasite does not rule
out infection. If appropriate history of tick exposure, geographic location of the patient (most often
seen in the Gulf Coast states of North America), appropriate clinical signs, and marked neutrophilia
are present, further testing such as PCR should be performed.
Plate 3-35 Hepatozoon Gametocytes

INCLUSIONS, PARASITES, AND INFECTIOUS AGENTS Hepatozoon spp. 193
Plate 3-35 Hepatozoon Gametocytes (con’t)

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Plate 3-35b
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Plate 3-35d
Plate 3-35e
Plate 3-35f
Plate 3-35g
Plate 3-35h
Plate 3-35i
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Plate 3-35a


193.e2
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Plate 3-35j
Histoplasma capsulatum
DISTINCTIVE FEATURES: Histoplasma capsulatum organisms are small (2 to 4 µm in diameter), round to
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oval yeast organisms found in the cytoplasm of peripheral blood monocytes, neutrophils, and occasion-
ally eosinophils in infected dogs and cats. The yeasts have an eccentrically placed, reddish, crescent-
shaped nucleus; clear to light blue cytoplasm; and a thin, clear halo (cell wall) surrounding the organism.
The leukocytes typically show marked toxic changes. A cell may contain one or more yeast organisms,
which may be seen extracellularly (free in the background of the smear) when leukocytes containing the
yeast organisms have ruptured (during smear preparation).
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DIAGNOSTIC SIGNIFICANCE: The presence of Histoplasma yeast organisms in peripheral blood indicates
disseminated histoplasmosis and carries a poor prognosis. Localized histoplasmosis such as gastrointes-
tinal histoplasmosis may become disseminated, especially if immunosuppressive doses of glucocorti-
coids are given.
Plate 3-36 Histoplasma Yeast

INCLUSIONS, PARASITES, AND INFECTIOUS AGENTS HISTOPLASMA CAPSULATUM 195
Plate 3-36 Histoplasma Yeast (con’t)

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Plate 3-36b
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Plate 3-36e
Plate 3-36f
Plate 3-36g
Plate 3-36h
Plate 3-36i
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Plate 3-36a


195.e2
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Plate 3-36j
Mucopolysaccharidosis
DISTINCTIVE FEATURES: Mucopolysaccharidoses (MPSs) are a group of uncommon hereditary lysosomal
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storage disorders, which result in leukocyte inclusions. With MPS, the vast majority (generally ≥90%) of
neutrophils, and occasionally also lymphocytes, contain reddish to purple intracytoplasmic granules.
Basophils generally have larger granules, and in cats, basophil granules stain basophilic instead of the
typical mauve color. Affected individuals often have significant physical abnormalities such as dished
faces, small head and ears, and skeletal deformities of the spine and long bones, which are noted at a
young age.
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DIAGNOSTIC SIGNIFICANCE: MPS is generally associated with severe medical consequences in the
homozygous state and a shortened life span. The cytoplasmic granules need to be distinguished from
other granule abnormalities and from toxic granulation in neutrophils, normal neutrophil primary gran-
ules (usually very faint), Birman cat anomaly and Chédiak-Higashi syndrome.
Plate 3-37 Cytoplasmic Granules Associated with Mucopolysaccharidosis
Basophil
Plate 3-37 Cytoplasmic Granules Associated with Mucopolysaccharidosis (con’t)
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A B C
D E
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Plate 3-37b
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Plate 3-37e
Plate 3-37f
Plate 3-37g
Plate 3-37h
Plate 3-37i
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Plate 3-37a

Basophil

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197.e3
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A B C
D E
Chédiak-Higashi Syndrome
DISTINCTIVE FEATURES: Chédiak-Higashi syndrome (CHS) is a rare inherited disease found in smoke
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blue Persian cats, in which abnormal formation of lysosomal granules and abnormal degranulation
occurs. Affected cats typically have a diluted smoke blue coat color and yellow-green irises and may
have slight neutrophil function defects and bleeding disorders. With CHS, eosinophilic intracytoplasmic
granules are found in neutrophils and less commonly in monocytes and lymphocytes. Leukocytes often
contain a single granule but may contain two or three granules. Eosinophil and basophil granules are
enlarged and eosinophil granules may appear roundish instead of having the typical rod shape. To
confirm if the granules are Döhle bodies or granules associated with CHS, a blood smear may be stained
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using peroxidase staining, and CHS granules will stain black.

NEXT STEPS: CHS is diagnosed on the basis of finding characteristic eosinophilic leukocyte granules
in smoke blue Persian cats and ruling out other granule abnormalities and toxic change of
neutrophils.
Plate 3-38 Cytoplasmic Granules Associated with Chédiak-Higashi Syndrome

Plate 3-38 Cytoplasmic Granules Associated with Chédiak-Higashi Syndrome (con’t)
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A B C
D E
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Plate 3-38b
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Plate 3-38f
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Plate 3-38i
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199.e2
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199.e3
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A B C
D E
Birman Cat Anomaly

DISTINCTIVE FEATURES: Birman cat anomaly is an inherited neutrophil granulation defect of Birman cats
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with no clinical signs or evidence of neutrophil dysfunction. Fine eosinophilic granules are found in the
cytoplasm of neutrophils.
DIAGNOSTIC SIGNIFICANCE: Birman cat anomaly must be differentiated from toxic granulation
and MPS.
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Plate 3-39 Cytoplasmic Granules Associated with Birman Cat Anomaly

Plate 3-39 Cytoplasmic Granules Associated with Birman Cat Anomaly (con’t)
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A B C
D E
201.e1
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Plate 3-39a
Plate 3-39b
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Plate 3-39d
Plate 3-39e
Plate 3-39f
Plate 3-39g
Plate 3-39h
Plate 3-39i
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Plate 3-39a


201.e2
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Plate 3-39j
201.e3
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A B C
D E
GM Gangliosidosis
DISTINCTIVE FEATURES: GM gangliosidosis, which is an uncommon inherited lysosomal storage disorder
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of dogs and cats, occurs in two forms. In GM1 gangliosidosis, small, punctate, clear cytoplasmic vacuoles
are found in lymphocytes. In GM2 gangliosidosis, azurophilic granules are found in the cytoplasm of
lymphocytes, and dark purple granules are found in neutrophils.
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INCLUSIONS, PARASITES, AND INFECTIOUS AGENTS GM GANGLIOSIDOSIS 203
Plate 3-40 Cytoplasmic Vacuoles Associated with GM1 Gangliosidosis

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Plate 3-40d
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Plate 3-40h
White Blood Cell Artifacts

Pyknotic Cells
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DISTINCTIVE FEATURES: Pyknosis occurs in senescent (old) leukocytes and results from preprogrammed
cell death (apoptosis). With pyknosis, the nucleus becomes dense and compact and begins to fragment
(karyorrhexis) resulting in spheres of dark-staining nuclear chromatin. Therefore, pyknotic cells have an
intact cytoplasmic membrane with one or more, variably sized, dense, round, dark nuclear fragments.
DIAGNOSTIC SIGNIFICANCE: The presence of pyknotic cells generally indicates a delay (hours) in
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making the blood smear after blood collection. Although pyknotic cells are often neutrophils because of
their short life span and because they are typically the most numerous leukocyte, the identity of pyknotic
cells cannot be determined. Preparation of blood smears as soon as possible after blood collection will
help avoid such artifacts.
Plate 3-41 Pyknotic Cells

WHITE BLOOD CELL ARTIFACTS PYKNOTIC CELLS 205
Plate 3-41 Pyknotic Cells (con’t)

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Plate 3-41b
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Plate 3-41j
Basket or Smudge Cells

DISTINCTIVE FEATURES: Bare or free nuclei occur when WBCs rupture during blood smear preparation.
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The free nucleus may retain a similar appearance to the intact cell nucleus, or the nuclear chromatin may
spread out and form an amorphous, netlike, eosinophilic structure (basket cell).
DIAGNOSTIC SIGNIFICANCE: Free nuclei and basket cells represent ruptured nucleated cells, and a few
are commonly seen on many blood smears. Increased numbers may be seen with leukocytosis because
of the increased number of leukocytes and increased chances of rupture. Also, conditions with high
numbers of nucleated RBCs may increase the number of basket cells.
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Plate 3-42 Basket/Smudge Cells

WHITE BLOOD CELL ARTIFACTS BASKET OR SMUDGE CELLS 207
Plate 3-42 Basket/Smudge Cells (con’t)

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Plate 3-42b
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Plate 3-42d
Plate 3-42e
Plate 3-42f
Plate 3-42g
Plate 3-42h
Plate 3-42i
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Platelet over White Blood Cell

DISTINCTIVE FEATURES: Occasionally platelets will overlie WBCs and may be readily identified by adjust-
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ing the fine focus of the microscope.

DIAGNOSTIC SIGNIFICANCE: Platelets over WBCs (or RBCs) are incidental findings of no clinical sig-
nificance and must be distinguished from parasites or inclusions.
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Plate 3-43 Platelets Overlying White Blood Cells

WHITE BLOOD CELL ARTIFACTS PLATELET OVER WHITE BLOOD CELL 209

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A B
C D
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Plate 3-43d
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Plate 3-43g
Plate 3-43h
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Plate 3-43d Plate 3-43f


209.e2
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A B
C D
Changes Associated with Delayed Processing

DISTINCTIVE FEATURES: If blood smears are not made in a timely fashion, the cells age and degenerate.
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Neutrophil nuclei become poorly defined and appear to have smudged borders and homogenized
chromatin.
DIAGNOSTIC SIGNIFICANCE: Recognize cellular aging changes associated with increased storage of
blood prior to making smears.
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Plate 3-44 Neutrophils Affected by Delayed Blood Smear Preparation

WHITE BLOOD CELL ARTIFACTS CHANGES ASSOCIATED WITH DELAYED PROCESSING 211
Plate 3-44 Neutrophils Affected by Delayed Blood Smear Preparation (con’t)

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Plate 3-44b
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Plate 3-44e
Plate 3-44f
Plate 3-44g
Plate 3-44h
Plate 3-44i
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Plate 3-44a


211.e2
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Plate 3-44j
Understained Smears
DISTINCTIVE FEATURES: Understained smears have pale-staining leukocytes with ill-defined nuclear
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features.
DIAGNOSTIC SIGNIFICANCE: Recognize smears that are understained, and avoid false identification of
inclusions and intracellular organisms.
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Plate 3-45 Understained Blood Films and Subsequent

Poor Staining of Leukocytes
Eosinophil
Eosinophil Neutrophil Neutrophil
Neutrophil Eosinophil
Understained
leukocyte
WHITE BLOOD CELL ARTIFACTS UNDERSTAINED SMEARS 213
Plate 3-45 Understained Blood Films and Subsequent

Poor Staining of Leukocytes (con’t)
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Plate 3-45a
Plate 3-45b
Plate 3-45c
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Plate 3-45e
Plate 3-45f
Plate 3-45g
Plate 3-45h
Plate 3-45i
Plate 3-45j
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Eosinophil Neutrophil
Plate 3-45a
Eosinophil
Neutrophil
Neutrophil

Eosinophil Understained
leukocyte

213.e2
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Plate 3-45j
SECTION 4:
PLATELETS
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Normal Morphology, 216

Thrombocytopenia, 218
Thrombocytosis, 222
Platelet Clumps, 224
Activated Platelets, 226
Macroplatelets (Megaplatelets), 228
Megakaryocytes, 230
Parasites, 232
Anaplasma platys (formerly Erhlichia platys), 232
215
216 SECTION 4 PLATELETS
Normal Morphology
DISTINCTIVE FEATURES: Platelets are cytoplasmic fragments that form from bone marrow megakaryo-
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cytes (platelet progenitors) and are the smallest formed element in peripheral blood. As platelets are
cytoplasmic fragments, they have no nucleus and generally appear as light blue to light pink, round to
oval structures, with few to many small reddish granules.
DIAGNOSTIC SIGNIFICANCE: One of the most important functions of platelets is the role they play in
hemostasis, requiring both adequate platelet numbers and normal platelet function. Blood smear review
is important in assessing the number of platelets as well as evaluation of platelet size, shape, and granu-
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larity. Canine and feline blood smears should have between 7 and 35 platelets on the average of ten 100×
oil objective fields in the monolayer to be considered adequate. Feline platelets tend to become activated
easily during collection, handling of the blood sample, or both; thus, feline platelets often clump, result-
ing in falsely decreased platelet counts and estimates.
Plate 4-1 Normal Platelets

NORMAL MORPHOLOGY 217
Plate 4-1 Normal Platelets (con’t)

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Plate 4-1b
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Plate 4-1d
Plate 4-1e
Plate 4-1f
Plate 4-1g
Plate 4-1h
Plate 4-1i
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Plate 4-1a


217.e2
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Plate 4-1j
Thrombocytopenia
DISTINCTIVE FEATURES: Thrombocytopenia is the absence of platelet clumps and less than seven platelets
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per oil power field (100× objective) on an average for 10 or more fields.
DIAGNOSTIC SIGNIFICANCE: Be sure that platelet clumps are not present, as this will falsely decrease
the platelet estimate, leading to an erroneous diagnosis of thrombocytopenia. If platelet clumps are not
present, then each platelet seen on average in the 100× (oil objective) field equals approximately 15,000
to 20,000 platelets per microliter (µL).
Feline platelets are small and often similar in size to erythrocytes and may not be counted as platelets
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by automated hematology analyzers. Mild thrombocytopenia (100,000 to 200,000 platelets/µL) is not

recognized on evaluation of a blood film. Only moderate and marked thrombocytopenias (platelet
counts below 100,000/µL) are recognized. This is not a problem, since increased bleeding secondary to
thrombocytopenia alone does not occur until platelet numbers are below 50,000/µL, and spontaneous
bleeding secondary to thrombocytopenia alone does not occur until platelet numbers are below
25,000/µL.
NEXT STEPS: If platelet clumps have been ruled out and a true thrombocytopenia is evident, inves-
tigation for the underlying pathologic mechanisms for thrombocytopenia should be performed.
Three main mechanisms of thrombocytopenia occur secondary to many different etiologies: (1)
decreased platelet production (certain drugs and toxins, immune-mediated destruction of platelets,
megakaryocytes, or both; neoplasia of the bone marrow); (2) decreased platelet survival (blood loss,
disseminated intravascular coagulation [DIC], immune-mediated destruction); and (3) platelet
sequestration (secondary to enlarged spleen). If a cause cannot be established, bone marrow cytol-
ogy is often very helpful in determining if disease of the marrow is the cause of persistent, unex-
plained thrombocytopenia).
When evidence of clinical bleeding is present, and platelet numbers and coagulation times (prothrom-
bin time [PT], partial thromboplastin time [PTT]) are normal, then investigation into platelet function
defects should be considered with platelet function tests. Diseases resulting in altered platelet function
may be both hereditary (Basset Hound thrombopathia, Chédiak-Higashi syndrome, cyclic hematopoiesis
in Grey Collies, Glanzmann’s thrombasthenia of Great Pyrenees and Otterhounds), and acquired (drugs,
toxins, infections, neoplasia, hyperglobulinemia).
THROMBOCYTOPENIA 219
Plate 4-2 Thrombocytopenia

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Plate 4-2 Thrombocytopenia (con’t)

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One platelet
THROMBOCYTOPENIA 221
Artifactual thrombocytopenia. A, Monolayer, devoid of platelets

B, Feathered edge large platelet clumps
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A B
221.e1

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Plate 4-2e
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221.e2
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One platelet


A B
Thrombocytosis
DISTINCTIVE FEATURES: Greater than 35 platelets per oil power field (100×) objective on an average of 10
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or more fields.
DIAGNOSTIC SIGNIFICANCE: Physiologic thrombocytosis (increased platelet production) is the most
common cause of thrombocytosis, occurring secondary to a reactive process (i.e., inflammation, iron
deficiency anemia in dogs, rebound from thrombocytopenia, chronic blood loss anemia), with platelet
counts typically fewer than 1,000,000 platelets/µL (<60 platelets per oil power field [100× objective] on
an average of 10 or more fields). If the platelet count is persistently greater than 1,000,000 platelets/µL,
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then essential thrombocythemia and acute megakaryocytic leukemia should be considered, both of which
are very rare hemic neoplasms of dogs and cats.
Plate 4-3 Thrombocytosis

THROMBOCYTOSIS 223
Plate 4-3 Thrombocytosis (con’t)

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Plate 4-3a
Plate 4-3b
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Plate 4-3d
Plate 4-3e
Plate 4-3f
Plate 4-3g
Plate 4-3h
Plate 4-3i
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Plate 4-3a


223.e12
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Plate 4-3j
Platelet Clumps
DISTINCTIVE FEATURES: Platelet clumps may appear as groups of distinct platelets, but degranulated
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platelet clumps may appear as blue blobs, which are difficult to recognize as platelet clumps. Because
of their large size, platelet clumps may be concentrated on the feathered edge, but smaller clumps may
also be present in the monolayer.
DIAGNOSTIC SIGNIFICANCE: Platelet clumping is typically an artifact secondary to platelet activation
during collection, handling of peripheral blood, or both. Platelet clumping is a common problem in cats.
Platelet clumps result in platelet counts (manual and machine generated) being falsely decreased, and
severe clumping may result in the platelet count being falsely reported as thrombocytopenia (pseudo-
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thrombocytopenia). Also, some conditions cause platelets to be hyperactive, so they are more prone to
being activated, resulting in platelet clumps. If many platelet clumps are present in a patient that has a
low platelet count, it should be considered pseudothrombocytopenia. When many platelet clumps are
present, the patient is considered to have sufficient numbers of platelets to not spontaneously bleed from
thrombocytopenia alone, regardless of the platelet count generated by the hematology analyzer.
Plate 4-4 Platelet Clumps

THROMBOCYTOSIS PLATELET CLUMPS 225
Plate 4-4 Platelet Clumps (con’t)

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Plate 4-4b
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Plate 4-4d
Plate 4-4e
Plate 4-4f
Plate 4-4g
Plate 4-4h
Plate 4-4i
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Plate 4-4a


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Plate 4-4j
Activated Platelets
DISTINCTIVE FEATURES: Activated platelets have thin cytoplasmic projections. The platelet granules may
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become condensed in the center of the platelet and should not be mistaken for a nucleus. Also, activated
platelets may degranulate, and few or no reddish intracytoplasmic granules may be present. Degranu-
lated platelet clumps may appear as bluish blobs, which are difficult to recognize as platelet clumps.
DIAGNOSTIC SIGNIFICANCE: Platelet activation is typically an artifact that occurs during collection,
and/or handling of peripheral blood, or both. Feline platelets activate easily, and activated platelets and
platelet clumps are common findings. Make sure not to confuse activated platelets with infectious agents
such as trypanosomes and sphirochetes.
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Plate 4-5 Activated Platelets

THROMBOCYTOSIS ACTIVATED PLATELETS 227
Plate 4-5 Activated Platelets (con’t)

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Make sure not to confuse activated platelets with infectious organisms such as
trypanosomes and spirochetes. A, Trypanosoma spp. B, Spirochetes spp.
C, Activated platelets
A B C
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Plate 4-5b
Plate 4-5c
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Plate 4-5e
Plate 4-5f
Plate 4-5g
Plate 4-5h
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Plate 4-5j
A B C
Macroplatelets (Megaplatelets)
DISTINCTIVE FEATURES: Platelets that are greater than 5 µm in diameter (as large as or larger than feline
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red blood cells [RBCs]).

DIAGNOSTIC SIGNIFICANCE: Low numbers of megaplatelets may be seen on peripheral blood smears
from normal cats. Increased numbers of megaplatelets in thrombocytopenic dogs suggests increased
thrombopoiesis; however, this also may be seen in myeloproliferative and myelodysplastic disorders.
Megaplatelets may also be seen on blood smears from healthy animals with hereditary platelet function
defects. Some Cavalier King Charles Spaniels may have congenital macrothrombocytopenia, which is a
benign inherited giant platelet disorder. These patients may be thrombocytopenic and have megaplate-
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lets. The platelets of these dogs function normally, and despite low numbers of platelets, bleeding
abnormalities are absent, as the platelet mass is normal.
Plate 4-6 Macroplatelets

THROMBOCYTOSIS MACROPLATELETS (MEGAPLATELETS) 229
Plate 4-6 Macroplatelets (con’t)

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Plate 4-6a
Plate 4-6b
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Plate 4-6d
Plate 4-6e
Plate 4-6f
Plate 4-6g
Plate 4-6h
Plate 4-6i
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Plate 4-6a


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Plate 4-6k
Megakaryocytes
DISTINCTIVE FEATURES: Megakaryocytes are bone marrow platelet progenitors, which undergo endomi-
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tosis rather than mitosis and cell division. Megakaryocytes are extremely large cells (generally 50 to
150 µm), which have a single nucleus with multiple lobes (2–16). The cytoplasm is blue to pink and
moderate to abundant, both dependent on the maturity of the cell. Megakaryocytes also contain few to
many reddish intracytoplasmic granules.
DIAGNOSTIC SIGNIFICANCE: Megakaryocytes are an extremely rare finding on peripheral blood smears
from normal animals. If present, secondary to their large size, megakaryocytes are usually found at the
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feathered edge. Megakaryocytes may be observed in the peripheral blood with megakaryocytic leukemia
(very rare), which may be associated with either thrombocytosis or thrombocytopenia and morphologi-
cally abnormal platelets.
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MEGAKARYOCYTES 231
Plate 4-7 Megakaryocytes (con’t)

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Parasites
Anaplasma platys (formerly Erhlichia platys)
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DISTINCTIVE FEATURES: Anaplasma platys (formerly Ehrlichia platys) is a tickborne intracellular bacterium
that infects platelets, resulting in infectious cyclic thrombocytopenia in dogs. The organisms appear as
a blue-black cluster within platelets.
DIAGNOSTIC SIGNIFICANCE: Finding intracellular Anaplasma platys organisms in the cytoplasm of
canine platelets is diagnostic. Infection is associated with thrombocytopenia. If diagnosed, assessment
for other tickborne diseases is advised to rule out co-infection.
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PARASITES ANAPLASMA PLATYS (FORMERLY ERHLICHIA PLATYS) 233
Plate 4-8 Anaplasma platys (con’t)

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SECTION 5:
HEMATOPOIETIC NEOPLASIA
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Overview, 235
Lymphoid Leukemia, 235
Acute Lymphoblastic Leukemia, 235
Chronic Lymphocytic Leukemia, 238
T-Cell Large Granular Lymphocytic
Leukemia, 240
Acute Myeloid Leukemia, 242
Acute Undifferentiated Leukemia, 242
Acute Myeloblastic Leukemia, 242
Acute Promyelocytic Leukemia, 242
Acute Myelomonocytic Leukemia, 243
Acute Monocytic Leukemia, 243
Acute Erythroleukemia, 243
Acute Megakaryoblastic Leukemia, 246
Chronic Myeloid Leukemia, 248
Chronic Granulocytic Leukemia, 248
Chronic Monocytic Leukemia, 252
Chronic Myelomonocytic Leukemia, 254
Polycythemia Vera, 254
Essential Thrombocythemia, 254
Mitotic Figures, 256
234
LYMPHOID LEUKEMIA ACUTE LYMPHOBLASTIC LEUKEMIA 235
Overview
Leukemia is defined as a clonal proliferation of neoplastic cells originating in bone marrow. Leukemia of
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every peripheral blood cell type has been identified in dogs and cats. Lymphoid leukemia (both acute
and chronic forms) is common in dogs and cats, whereas leukemias of the other cell lines are quite
uncommon. Diagnosis of leukemia is based on clinical presentation (clinical signs and physical examina-
tion), and laboratory data, including complete blood count (CBC) and serum chemistry analysis. The
findings of large numbers of neoplastic cells on CBC and blood smear microscopic evaluation are one
important part of the diagnosis and classification of the various leukemias. Definitive diagnosis may
often require bone marrow evaluation.
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Many different leukemia classification schemes have been proposed and utilized. Various schemes
are based on a mix of information such as clinical behavior (response to therapy, survival time), cellular
morphology (microscopic evaluation of the neoplastic cells), histopathology (neoplastic pattern on tissue
biopsy), topography (extent of neoplastic involvement), phenotype (genetic expression), and genotype
(genetic characteristics and aberrations) of the neoplastic cell population.
Although blood smear evaluation remains an important part of diagnosis and classification of leuke-
mias, it is important to recognize that morphology alone is not sufficient to classify various leukemias.
This is especially true with the acute forms of leukemia, as blasts of various lineages may appear mor-
phologically identical.
Two large categories of leukemia exist: (1) lymphoid leukemias and (2) myeloid (nonlymphoid) leu-
kemias, with acute and chronic forms of each category. Acute leukemias are composed of blast cells. This
type of leukemia manifests acutely and is aggressive, often associated with large numbers of circulating
blasts, peripheral blood cytopenias, occasional secondary organ infiltration, and a poorer prognosis
compared with that for chronic leukemias.
With regard to acute leukemias, identification of the lineage of the blast cell population may be dif-
ficult, especially if it is solely based on light-microscopic morphologic features with routine hematologic
stains. Blast cells may sometimes have distinct features to suggest phenotype; however, most appear
similar microscopically, and further tests are needed to identify ontogeny. Such tests aiding in differentia-
tion may include cytochemical staining using enzymes and immunocytochemistry or flow cytometry,
which utilize labeled antibodies targeted against specified antigens found in various cell lines (immuno-
phenotyping). Furthermore, clonality tests are available for identifying lymphoid leukemias, to assess
whether a population of lymphoid cells are clonal in origin (suggestive of neoplasia) or nonclonal (sug-
gestive of inflammation or reactivity). Identification of genetic abnormalities (cytogenetics) is increas-
ingly playing a role in the classification of leukemias in veterinary medicine.
In contrast to acute leukemias, chronic leukemias develop more slowly, are not often associated with
cytopenias until the late stage of disease, often do not have organ involvement, and are associated with
a more favorable prognosis. The neoplastic cell population in chronic leukemias comprises mature, well-
differentiated cells that are often morphologically identical to normal blood leukocytes. Thus, the mature
morphology of the neoplastic cells may make it very difficult to distinguish leukemia from inflammatory
processes. Differentiation is based on ruling out sources of infection, inflammation, or immunostimula-
tion and continued monitoring of the CBC over time to document the persistence, and increasing
numbers, of one type of leukocyte.
A brief overview of the different types of leukemias in dogs and cats is presented in this chapter, with
an emphasis on morphologic features.
Lymphoid Leukemia
Acute Lymphoblastic Leukemia
DISTINCTIVE FEATURES: Acute lymphoblastic leukemia (ALL) is a neoplastic proliferation of lympho-
blasts and may be either B-cell or T-cell phenotype. Lymphoblasts are large cells, often 1.5 to 2 times the
diameter of neutrophils. Lymphoblasts contain slight to moderate amounts of basophilic cytoplasm, a
round to oval eccentrically placed nucleus, stippled to slightly coarse nuclear chromatin, and often either
faint or, more commonly, prominent nucleoli. Nucleoli are typically singular, but occasionally, two
nucleoli may be present.
DIAGNOSTIC SIGNIFICANCE: Lymphoblasts are rarely seen in the peripheral blood of normal dogs and
cats. Occasional lymphoblasts may be seen in peripheral blood with strong immune stimulation. Low
to high numbers of lymphoblasts may be seen in peripheral blood with lymphoblastic leukemia. Classic
236 SECTION 5 HEMATOPOIETIC NEOPLASIA
CBC findings with ALL include large numbers of circulating blasts and, often, one or multiple
cytopenias.
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NEXT STEPS: Light microscopy alone cannot reliably distinguish T-cell ALL from B-cell ALL, nor
can it reliably distinguish lymphoblasts from myeloblasts or monoblasts. Other methods such as
immunocytochemistry and flow cytometry are needed to define exact phenotype. B-cell leukemia is
associated with a more favorable prognosis compared with T-cell leukemia.
Low to occasional to moderate numbers of circulating lymphoblasts may be seen with stage V lym-
phoma, indicating neoplastic seeding of bone marrow and the subsequent leukemic phase. Distinguish-
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ing stage V lymphoma from late stage lymphoid leukemia may be difficult. Determining the location of
the bulk of neoplastic involvement (marrow or blood versus solid tissue) may be used to distinguish the
conditions.
Finding low to high numbers of blasts in peripheral blood warrants assessment for involvement of
lymphoreticular organs for staging purposes.
Plate 5-1 Acute Lymphoblastic Leukemia
nucleolus
mitotic figure
nucleolus
LYMPHOID LEUKEMIA ACUTE LYMPHOBLASTIC LEUKEMIA 237
Plate 5-1 Acute Lymphoblastic Leukemia (con’t)

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Chronic Lymphocytic Leukemia

DISTINCTIVE FEATURES: Chronic lymphocytic leukemia (CLL) is a neoplastic proliferation of small,
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mature lymphocytes. Morphology is identical to that of nonneoplastic mature lymphocytes. The lym-
phocytosis in CLL may be mild in early stages and marked in later stages. Cytopenias are uncommon
and typically only seen when the neoplastic cells are present in very high numbers.
DIAGNOSTIC SIGNIFICANCE: CLL is most often a leukemia found in older dogs and cats. When a mild
to moderate lymphocytosis is present, distinguishing it from a reactive population of lymphocytes sec-
ondary to immunostimulation may be difficult and relies on monitoring of the CBC over time (persistent
lymphocytosis, often with a slowly progressive increase in numbers). Immunophenotypic analysis and
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documentation of a monoclonal population may also lend support to the diagnosis of leukemia. In addi-
tion, clonality testing is also available to help identify clonal populations of lymphocytes. It is important
to recognize that clonality by itself is not diagnostic of neoplasia. As an example, clonal populations of
T lymphocytes have been identified in dogs with chronic ehrlichiosis.
NEXT STEPS: As for ALL, distinguishing between B-cell and T-cell phenotypes is not possible with
light microscopy alone, and further techniques are required. When CLL is diagnosed, assessment for
involvement of the lymphoreticular organs is advised to further stage the disease.
A diagnosis of CLL warrants monitoring of the CBC, despite treatment, as evolution into a blast crisis
may occur, culminating in an acute leukemia.
Plate 5-2 Chronic Lymphocytic Leukemia

LYMPHOID LEUKEMIA CHRONIC LYMPHOCYTIC LEUKEMIA 239
Plate 5-2 Chronic Lymphocytic Leukemia (con’t)

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T-Cell Large Granular Lymphocytic Leukemia

DISTINCTIVE FEATURES: T-cell large granular lymphocytic leukemia (T-cell LGL leukemia) is composed
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of large, immature T lymphocytes, with variable numbers of round to irregular, variably sized, magenta-
staining cytoplasmic granules.
DIAGNOSTIC SIGNIFICANCE: Low numbers of nonneoplastic circulating granular lymphocytes may be
seen in dogs and cats secondary to immunostimulation. In particular, low to moderate numbers of
granular lymphocytes may be seen in dogs, with or without a mild lymphocytosis and with ehrlichiosis.
Thus, as with CLL, ruling out etiologies of immunostimulation and monitoring of the CBC over time
are needed to help confirm a leukemic process.
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NEXT STEPS: As for all of the lymphoid leukemias, assessment for lymphoreticular organ infiltration
is suggested for staging. Although commonly referred to as lymphocytic leukemia of granular lympho-
cytes, in dogs, this disorder actually arises in the spleen rather than in bone marrow.
Plate 5-3 T-Cell Large Granular Lymphocytic Leukemia

LYMPHOID LEUKEMIA T-CELL LARGE GRANULAR LYMPHOCYTIC LEUKEMIA 241
Plate 5-3 T-Cell Large Granular Lymphocytic Leukemia (con’t)

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Acute Myeloid Leukemia

Acute myeloid leukemia (AML) is a group of acute leukemias composed of blast cells of one, or more
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than one, of the myeloid cell lines: granulocytic (neutrophils, eosinophils, basophils); monocytic; ery-
throid and megakaryocytic. Finding 20% or more blasts in blood or bone marrow is useful in diagnosis.
In overt AML, blasts are present in blood and bone marrow in very high numbers, with cytopenias and
possible lymphoreticular organ infiltration.
As with all acute leukemias, lineage is best determined through a combination of light-microscopic
evaluation and assessment of phenotype by using cytochemical stains or immunophenotyping by flow
cytometry.
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AML must be distinguished from myelodysplastic syndromes (MDS), especially with feline leukemia
virus (FeLV)–induced MDS in cats. Dysplastic changes may be seen with AML but are much more
common in MDS; the presence of dysplasia warrants FeLV testing in cats and careful monitoring of the
CBC, as MDS may progress to AML.
Acute Undifferentiated Leukemia

DISTINCTIVE FEATURES: Acute undifferentiated leukemia (AUL) is composed of blast cells of uncharacter-
ized lineage (lymphoid or myeloid), where the cells lack any defining morphologic features and may
have features of blasts of various lineages. The cells also lack cytochemical, structural, and immunologic
markers to aid in determination of ontogeny.
AUL is diagnosed on the basis of the presence of greater than 20% of nucleated cells in blood or
bone marrow constituting blast. Indeed, blasts are often in very high numbers both in blood and bone
marrow.
Acute Myeloblastic Leukemia

DISTINCTIVE FEATURES: The three subtypes of acute myeloblastic leukemia are AML-M0, AML-M1,
AML-M2, which are categorized by the degree of maturation and cellular differentiation of the neoplastic
cells performed by microscopic evaluation and immunophenotypic assessment.
All three subcategories consist of 20% or more myeloblasts in blood, bone marrow, or both (usually
large numbers of blasts are present, often approaching 100% of all nucleated cells).
Myeloblasts are difficult to distinguish from primitive blasts of other hematopoietic cell lines, and
sometimes even from lymphoblasts. Myeloblasts are about the same size, or slightly smaller than, pro-
myelocytes. They have round to oval, centrally located nuclei, which contain finely stippled chromatin
and one to several visible nucleoli. A small to moderate amount of moderately basophilic cytoplasm is
present and, at the later blast stage, may contain a few to several magenta-staining granules. Very imma-
ture myeloblasts may lack granules altogether.
DIAGNOSTIC SIGNIFICANCE: It is extremely rare to see myeloblasts in peripheral blood smears from
healthy dogs and cats. Finding a myeloblast in a peripheral blood smear always raises the concern of
granulocytic leukemia (although it must be kept in mind that it is difficult, if not impossible, to reliably
distinguish myeloblasts from lymphoblasts on the basis of morphology alone). However, rarely, a myelo-
blast may be seen in peripheral blood with severe inflammation, as part of an orderly left shift with toxic
change. In addition, rare immature hematopoietic precursors may be found in peripheral blood second-
ary to toxic or hypoxic damage to bone marrow.
NEXT STEPS: The persistent (progressive) presence of a few to large numbers of blasts in blood war-
rants bone marrow examination. Serial CBC and bone marrow examinations may be needed for
diagnosis, especially if the leukemia is in an early stage (low numbers of blasts).
Acute Promyelocytic Leukemia

DISTINCTIVE FEATURES: Acute promyelocytic leukemia (AML-M3) is an extremely uncommon leukemia
in animals. Diagnosis is based on finding 20% or more atypical-appearing promyelocytes in peripheral
blood, bone marrow, or both. Part of this count includes any identified myeloblasts as well.
The atypical-appearing promyelocytes may be hypergranular or poorly granular. Promyelocytes are
of the same size or slightly larger than myeloblasts, with a slight increase in pale blue cytoplasm. When
granules are present, they are azurophilic, small, and evenly distributed. Nuclei are round and slightly
smaller than those of the myeloblast. Lacy to coarse chromatin is seen, with faint or visible nucleoli
ACUTE MYELOID LEUKEMIA ACUTE ERYTHROLEUKEMIA 243
sometimes evident. Usually, nucleoli are fewer and more difficult to visualize compared with
myeloblasts.
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DIAGNOSTIC SIGNIFICANCE: AML-M3 is very uncommon and must be differentiated from a left shift
caused by severe inflammation or infection and from myelodysplastic syndromes.
Acute Myelomonocytic Leukemia
DISTINCTIVE FEATURES: In acute myelomonocytic leukemia (AML-M4), the neoplastic cells are of mixed
lineage, consisting of both immature myeloid cells and monocytic cells.
Bone marrow cytology is needed for diagnosis and is based on 20% or more of all nucleated bone
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marrow cells constituting myeloblasts, monoblasts, or promonocytes; 20% or more of all nucleated bone
marrow cells having to be of monocytic lineage; and 20% or more of all nucleated bone marrow cells
having to be of granulocytic lineage.
DIAGNOSTIC SIGNIFICANCE: Bone marrow cytology and cytochemical stains are helpful in distinguish-
ing myeloid cells from cells of monocytic lineage when cell counts are performed.
Acute Monocytic Leukemia
DISTINCTIVE FEATURES: Acute monocytic leukemia (AML-M5) is diagnosed on the basis of 80% or more
of all nucleated cells being of monocytic origin, with 20% or more of all nucleated cells consisting of
myeloblasts, monoblasts, or promonocytes.
Of the two subtypes, AML-M5a is composed of more immature monocytic cells (acute monoblastic
leukemia) and is more common in young animals. Here, monoblasts comprise 50% or more of monocytic
cells. The second subtype, AML-M5b, is composed of more mature monocytic cells (acute monocytic
leukemia) and is seen most often in older animals, and 50% or more of the monocytic cells are
promonocytes.
Monoblasts are roundish cells with a moderate amount of basophilic cytoplasm and a round to oval
nucleus that has an undulating outline. The nuclear chromatin ranges from stippled to stringy, and one
or more nucleoli are visible. The irregular outline to the nuclear membrane is one of the more distin-
guishing features of the monoblast.
DIAGNOSTIC SIGNIFICANCE: Bone marrow cytology and cytochemical stains are helpful in the diagnosis
of AML-M4.
Acute Erythroleukemia
DISTINCTIVE FEATURES: Acute erythroleukemia (AML-M6) is diagnosed by the presence of greater than
50% erythroid precursors in bone marrow. The two subtypes are AML-M6a, in which 20% or more of all
bone marrow nucleated cells are myeloblasts. AML-M6b is characterized by 20% or more of all bone
marrow nucleated cells consisting of myeloblasts and rubriblasts. The peripheral blood picture often
contains large numbers of immature erythroid and myeloid cells, with dysplastic erythroid elements
often seen in both subtypes.
Rubriblasts are round cells with dark blue cytoplasm and a roundish nucleus that is often centrally
located but may be eccentrically placed. The nuclear chromatin is stippled, and one or more nucleoli are
visible.
DIAGNOSTIC SIGNIFICANCE: AML-M6b is the subtype most commonly seen in animals, especially in
FeLV-positive cats. AML-M6 must be differentiated from myelodysplastic syndromes, also frequently
associated with FeLV infection in cats.
Rubriblasts are generally not seen in peripheral blood smears from healthy dogs and cats. Rubriblasts
may rarely be seen with strongly regenerative anemia such as immune-mediated anemia in dogs and
Mycoplasma hemofelis infection in cats. Also, rubriblasts may be seen in peripheral blood smears from
animals with repopulating bone marrow after severe insult.
Plate 5-4 Acute Erythroleukemia

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Plate 5-4 Acute Erythroleukemia (con’t)

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Acute Megakaryoblastic Leukemia

DISTINCTIVE FEATURES: Acute megakaryoblastic leukemia (AML-M7) is diagnosed on the basis of the
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presence of 20% or more blasts found in bone marrow, with 50% of these cells consistent with
megakaryoblasts.
Megakaryoblasts are larger than other blast cells and may have a single nucleus or a lobulated nucleus
from endomytotic divisions. The cytoplasm may be scant to moderate and often stains basophilic with
cytoplasmic blebbing.
DIAGNOSTIC SIGNIFICANCE: Megakaryoblasts are extremely uncommon in the peripheral blood of
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normal dogs and cats. Although megakaryoblasts may have unique features, allowing differentiation
from other myeloid blasts, cytochemistry and immunomarkers are often needed to define lineage.
Plate 5-5 Acute Megakaryoblastic Leukemia

ACUTE MYELOID LEUKEMIA ACUTE MEGAKARYOBLASTIC LEUKEMIA 247
Plate 5-5 Acute Megakaryoblastic Leukemia (con’t)

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Chronic Myeloid Leukemia

All forms of chronic myelogenous leukemia are uncommon in dogs and cats and, when diagnosed, are
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most often a disease of older animals. The chronic leukemias are composed of mature, well-differentiated
cells that are morphologically similar to their nonneoplastic counterparts. Diagnosis is based on finding
large numbers of mature cells in peripheral blood that cannot be explained by underlying etiologies,
which may be challenging at times. Often, serial CBC examinations are needed to document persistently
high and, often, slowly increasing numbers of cells.
In contrast to acute leukemias, which consist of rapidly dividing blasts, which quickly efface the
marrow and result in significant and often multiple cytopenias at the time of diagnosis, chronic leukemias
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are composed of mature, slowly developing and dividing cells. Thus, typically, cytopenias evolve only
when high numbers of leukemic cells are present in peripheral blood.
Similar to chronic lymphocytic leukemia, monitoring of the CBC over time is needed to evaluate for
an evolving blast crisis, signaling a transition from chronic leukemia to acute leukemia.
Chronic Granulocytic Leukemia
DISTINCTIVE FEATURES: Chronic granulocytic leukemia is diagnosed by the presence of large numbers
of neutrophils, eosinophils, or basophils in peripheral blood and bone marrow. Mature forms predomi-
nate, with a left shift composed of lower numbers of immature forms (bands to myeloblasts). Dysplastic
changes may be present.
DIAGNOSTIC SIGNIFICANCE: Chronic granulocytic leukemia may be difficult to distinguish from a
severe neutrophilia or leukemoid response. Evidence of neutrophil toxic change (suggesting infection)
and careful assessment of the patient for underlying inflammation, infection, and tissue necrosis, with
close monitoring of the CBC over time, are helpful to differentiate the conditions. Chronic granulocytic
leukemia may be associated with nonregenerative anemia and thrombocytopenia in the later stages of
disease, when large numbers of circulating neoplastic cells are present.
CHRONIC MYELOID LEUKEMIA CHRONIC GRANULOCYTIC LEUKEMIA 249
Plate 5-6 Chronic Granulocytic Leukemia (Neutrophilic)

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Plate 5-7 Chronic Granulocytic Leukemia (Eosinophilic)

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Chronic Monocytic Leukemia

DISTINCTIVE FEATURES: Chronic monocytic leukemia is diagnosed on the basis of large numbers of
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mature monocytes in peripheral blood and bone marrow. It may be associated with cytopenias in later
stage of disease, when large numbers of circulating neoplastic cells are present.
Plate 5-8 Chronic Myeloid Leukemia (Monocytic)

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Plate 5-8 Chronic Myeloid Leukemia (Monocytic) (con’t)

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Chronic Myelomonocytic Leukemia

DISTINCTIVE FEATURES: Chronic myelomonocytic leukemia is diagnosed on the basis of large numbers
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of mature neutrophils and monocytes in peripheral blood and bone marrow. It may be associated with
cytopenias in the later stages of disease.
Polycythemia Vera
DISTINCTIVE FEATURES: Polycythemia vera (PV) is a chronic leukemia of dogs and cats, in which persis-
tent, unexplained polycythemia exists.
DIAGNOSTIC SIGNIFICANCE: Diagnosis of PV is based on excluding two much more common causes of
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polycythemia: (1) relative polycythemia and (2) nonneoplastic absolute polycythemia.

Relative polycythemia is secondary to dehydration or splenic contraction (catecholamine release
caused by excitement). Dehydration may be determined by physical examination and by the presence
of panhyperproteinemia or hyperalbuminemia and prerenal azotemia.
Nonneoplastic etiologies of absolute polycythemia may be appropriate physiologic responses such as
production of erythropoietin (EPO) secondary to hypoxia from cardiac disease, respiratory disease, or
both, resulting in an increase in RBC production. Assessment to rule out disease of the heart and lungs
(auscultation of the thorax, thoracic radiography, ultrasonography, electrocardiography), blood gas
assessment to confirm hypoxia, and measurement of EPO levels aid in diagnosis.
Inappropriate causes of absolute polycythemia are from increased EPO production in the absence of
hypoxia, as in EPO-secreting neoplasms of the kidney or EPO secretion secondary to nonneoplastic renal
disease. These are diagnosed when hypoxia is ruled out through blood gas assessment and measurement
of EPO.
Additionally, some endocrinopathies such as hyperadrenocorticism (Cushing disease) and hyperthy-
roidism may result in polycythemia (hormonal stimulation of erythropoiesis) and may be diagnosed
with a combination of serum chemistry tests, physical examination findings, and patient history.
Essential Thrombocythemia
DISTINCTIVE FEATURES: Essential thrombocythemia (ET) is a rare, chronic leukemia in dogs and cats.
Diagnosis is based on the identification of sustained, chronic, unexplained, marked thrombocytosis with
megakaryocytic hyperplasia of bone marrow. Platelets are usually normal in appearance but may be of
variable size. Dysplastic changes and associated cytopenias are not commonly seen.
DIAGNOSTIC SIGNIFICANCE: Diagnosis of ET is based on excluding all causes of nonneoplastic throm-
bocytoses, the most common of which is a reactive thrombocytosis secondary to underlying inflamma-
tion, infection, neoplasia, and iron deficiency anemia. Other less common causes of a nonneoplastic
thrombocytosis are secondary to certain drugs and splenic contraction (epinephrine release).
Additionally, thrombocytosis may also be seen with some forms of leukemia such as basophilic leu-
kemia, PV, and acute megakaryoblastic leukemia.
Plate 5-9 Essential Thrombocythemia

CHRONIC MYELOID LEUKEMIA ESSENTIAL THROMBOCYTHEMIA 255
Plate 5-9 Essential Thrombocythemia (con’t)

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Plate 5-9i
Mitotic Figures
DISTINCTIVE FEATURES: Mitotic figures are cells undergoing cell division. A well-defined nucleus is not
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seen; instead, chromatids are seen as dark purple (same color as the nucleus), linear structures forming
a spindle. Mitotic cells may be found in any stage of mitoses, and thus, chromatids may be concentrated
in the center of the cell, forming a spindle shape or pulled apart from the center and found in two polar
groups. If the cell undergoing division is neoplastic, aberrant mitoses, in which the chromatids are dis-
persed randomly throughout the cytoplasm of the cell and appear disorganized, may be seen.
DIAGNOSTIC SIGNIFICANCE: Mitotic figures are not normally seen in the peripheral blood of normal
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dogs and cats. When present, they are often found at the feathered edges of the blood smear. Rarely,
mitotic figures are seen in the peripheral blood of nonleukemic animals, in cases of reactive lymphocy-
tosis, and rarely in cats with strongly regenerative anemias. Usually, when mitotic figures are found on
peripheral blood smears, they are part of a leukemic cell population. Therefore, when mitotic figures are
identified in peripheral blood smears, assessment for underlying leukemia is warranted.
Plate 5-10 Mitotic Figures

MITOTIC FIGURES 257
Plate 5-10 Mitotic Figures (con’t)

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Plate 5-10a
Plate 5-10b
Plate 5-10c
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Plate 5-10e
Plate 5-10f
Plate 5-10g
Plate 5-10h
Plate 5-10i
Plate 5-10j
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SECTION 6:
EXTRACELLULAR ORGANISMS
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Microfilaria, 260
Trypanosomes, 262
Spirochetes, 264
Bacteria, 266
259
260 SECTION 6 EXTRACELLULAR ORGANISMS
Microfilaria
DISTINCTIVE FEATURES: Microfilariae from Dirofilaria immitis or Acanthocheilonema reconditum (Dipetalo-
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nema reconditum) are large, extracellular filarial larvae, which have an elongated “wormlike” body. In
general, D. immitis (L1) larvae are usually present in high numbers, have a stationary or nonprogressive
movement in wet preps, a straight body, a straight tail, and a tapered head. In contrast, A. reconditum
larvae generally are few in number, have a progressive movement in wet preps, a curved body, and a
blunt head. Differentiation based on microscopic evaluation is difficult, and mixed infections may occur.
DIAGNOSTIC SIGNIFICANCE: The presence of filarial larvae indicates infection with either or both
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D. immitis or A. reconditum.
Plate 6-1 Microfilaria

MICROFILARIA 261
Plate 6-1 Microfilaria (con’t)

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A, Fibrin strand B, Two microfilaria
A B
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Next Topic
Plate 6-1a
Plate 6-1b
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Plate 6-1d
Plate 6-1e
Plate 6-1f
Plate 6-1g
Plate 6-1h
Plate 6-1i
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Plate 6-1a


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A B
Trypanosomes
DISTINCTIVE FEATURES: Trypomastigotes, the flagellated stage of trypanosomes found in peripheral
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blood, are large, extracellular protozoa that have an elongated or “blade-shaped” body with an undulat-
ing membrane, a tapering posterior end, and a short flagellum directed anteriorly.
DIAGNOSTIC SIGNIFICANCE: The presence of trypanosomes in peripheral blood indicates trypanoso-
miasis. Trypanosomiasis is a zoonotic disease, and dogs and cats are important reservoirs of infection.
In the United States, trypanosomal infection is most often caused by Trypanosoma cruzi (Chagas disease)
and is transmitted by biting triatomine bugs, resulting in systemic infection and many variable clinical
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signs. The predominant clinical manifestation is cardiac disease secondary to myocarditis.
Plate 6-2 Trypanosomes
Platelets
Trypanosomes
TRYPANOSOMES 263
Plate 6-2 Trypanosomes (con’t)

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Platelet
Trypanosome
Make sure not to confuse curve shaped platelets with trypanosomes.

A, Platelets B, Trypanosomes
Platelets
A B
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Plate 6-2d
Plate 6-2e
Plate 6-2f
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Platelets
Trypanosomes

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Platelet
Trypanosome

Platelets
A B
Spirochetes
DISTINCTIVE FEATURES: Spirochetes are rarely seen in peripheral blood and are bacteria of the order
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Spirochaetales. Spirochetes appear as small, thin, corkscrew-shaped, extracellular organisms.

DIAGNOSTIC SIGNIFICANCE: The presence of spirochetes in peripheral blood suggests borreliosis or
lyme disease, which is a tickborne disease caused by the bacterium Borrelia burgdorferi. Finding spiro-
chetes on peripheral blood films warrants further testing, empirical antibiotic therapy, or both.
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Plate 6-3 Spirochetes
Clusters of
spirochetes
SPIROCHETES 265
Plate 6-3 Spirochetes (con’t)

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Spirochete on
surface of RBC
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Make sure not to confuse activated platelets with spirochetes and platelet
fragments. A, Activated Platelet B, Platelet Fragment C, Spriochetes
Activated
platelets
Platelet fragment
A B C
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Plate 6-3e
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Clusters of
spirochetes

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Spirochete on
surface of RBC
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Activated
platelets
Platelet fragment
A B C

Bacteria
DISTINCTIVE FEATURES: Moderate numbers of extracellular bacterial rods or cocci may occur.
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DIAGNOSTIC SIGNIFICANCE: Bacterial overgrowth occurs occasionally in ethylenediaminetetraacetic

acid (EDTA) tubes and must be differentiated from true septicemia. Bacterial septicemia usually has low
numbers of intracellular organisms and markedly toxic neutrophils and must be correlated with clinical
signs and history of the patient. Bacterial contamination usually has only extracellular bacteria (the
neutrophils may be toxic or have no toxic change, depending on the health status of the patient during
blood sampling).
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Plate 6-4 Extracellular Bacteria
Clusters of
bacteria
Rod bacteria on
RBC surfaces
BACTERIA 267
Plate 6-4 Extracellular Bacteria (con’t)

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Make sure to distinguish bacteria (rare) from clumps of stain precipitant

(common). A, Stain precipitation. Note globular to amorphous appearance
B, Bacteria. Note distinct uniform rod shape
A B
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Plate 6-4e Plate 6-4j
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Clusters of
bacteria

Rod bacteria on
RBC surfaces


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A B
APPENDIX
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Reticulocyte Count
The reticulocyte count, which is used to assess bone marrow response to anemia, is a more sensitive
measurement of bone marrow regenerative response than is polychromasia because a greater concentra-
tion of ribosomes in red blood cell (RBC) cytoplasm is required for RBCs to stain polychromatophilic
with Romanowsky-type (Wright) stains than is required for RBC to stain as reticulocytes with supravital
stains (new methylene blue [NMB]). Thus, all polychromatophilic cells stain as reticulocytes with supra-
vital stains, whereas some reticulocytes do not stain polychromatophilic with Romanowsky-type stains.
The reticulocyte count may be performed in-house or by referral laboratories. Box 1 gives the procedure
for performing in-house reticulocyte counts. Referral laboratories may perform reticulocyte counts on
ethylenediaminetetraacetic acid (EDTA) blood samples submitted for hemogram analysis or for other
reasons. Relative reticulocyte counts are reported as the percentage of RBCs that are reticulocytes. Abso-
lute reticulocyte counts are reported as the number of reticulocytes per volume of blood. When possible,
the absolute reticulocyte count should be used to evaluate the regenerative response rather than the rela-
tive reticulocyte count.
Reticulocytes are young RBCs containing a sufficient concentration of polyribosomes so that after
incubation in supravital stains such as NMB, polyribosomes aggregate forming blue-black dots or aggre-
gates. Reticulocytes do not increase to what is considered adequate levels to immediately classify an
anemia as regenerative. It takes approximately 2 days for the number of young RBCs to increase in
the circulation and 5 to 7 days for them to reach a maximum level with most acute anemias that are
BOX 1 New Methylene Blue Stain for Reticulocytes
A. Principle
New methylene blue is a cationic dye capable of penetrating the living cell and precipitating
the ribosomes of the immature red blood cell (RBC). In the reticulocyte, new methylene
blue (NMB) stain precipitates with the remnants of the endoplasmic reticulum (ER), which
is primarily the ribosome and messenger ribonucleic acid (mRNA) for hemoglobin synthesis.
As the RBC matures, the ER is lost, so only cells recently released from bone marrow into
the circulation will contain the ER. Therefore, the reticulocyte count estimates the bone
marrow response to anemia.
B. Procedure
1. Place equal amounts of NMB stain and ethylenediaminetetraacetic acid (EDTA) blood
in a test tube for 20 minutes.
2. From this mixture, make a blood smear.
3. Count 1000 RBCs, recording the number of reticulocytes observed in these 1000 cells.
C. Calculation of Reticulocyte Count
number of reticulocytes counted
per 1000 RBC
= % reticulocytes
10
269
270 APPENDIX
regenerative (hemorrhage or hemolysis). Therefore, an acute anemia may appear nonregenerative by a

reticulocyte count performed before young RBCs have increased in the circulation.
In dogs and cats, the reticulocyte count increases proportionately to the severity of anemia if the
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anemia is regenerative (i.e., bone marrow regeneration is occurring appropriately in response to the
anemia). As erythropoiesis increases in response to anemia, young RBCs are released into the circulation
1 to 2 days earlier than normal. As a result, they are within the circulation but still stainable as reticulo-
cytes for 1 or 2 days longer than under normal conditions. Also, the increase in erythropoiesis causes
many more young RBCs to be produced and released. Hence, increased erythropoiesis is accompanied
by an increase in peripheral blood reticulocyte count proportional to the magnitude of increased eryth-
ropoiesis, which is, in turn, proportional to the severity of anemia.
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In cats, reticulocytes may be divided into two morphologic classes: (1) punctate reticulocytes and (2)
aggregate reticulocytes. Punctate reticulocytes are RBCs containing a few small, dotlike (punctate), blue-
black structures (small polyribosome aggregates) but no large aggregates of ribosomes. Aggregate reticu-
locytes are RBCs containing one or more medium to large, blue-black structures that may appear as a
cluster or network of aggregated structures. Punctate and aggregate reticulocytes should be differenti-
ated and counted separately. Counting the number of punctate reticulocytes may be very time consum-
ing, as they can reach very high numbers. Therefore, in moderate to severe anemias, punctate reticulocytes
are frequently not counted, since bone marrow regenerative response is interpreted solely on the basis
of aggregate reticulocyte numbers in moderate to severe anemias.
Feline punctate reticulocytes represent a later maturational stage of feline aggregate reticulocytes. The
feline aggregate reticulocyte circulates only about 1 day before sufficient ribosomes are degraded for it
to stain as a punctate reticulocyte. Feline aggregate reticulocyte counts are interpreted similarly to reticu-
locyte counts in dogs and cattle. However, in mild anemias, feline aggregate reticulocytes may be held
in bone marrow until they reach the punctate stage, resulting in little or no increase in peripheral blood
aggregate reticulocyte numbers. Feline punctate reticulocytes may occur in high numbers (up to 10%)
in health, increase during regenerative anemia, and may be elevated during mild regenerative anemia
without concurrent elevation of feline aggregate reticulocytes. Also, punctate reticulocytes may be ele-
vated during severe nonregenerative or poorly regenerative anemias without an appropriate concurrent
elevation of feline aggregate reticulocytes. Because feline punctate reticulocytes may be increased during
mild anemia without concurrent increase in feline aggregate reticulocytes, feline punctate reticulocytes
may be helpful in determining whether mild anemias are regenerative or nonregenerative. For example,
high numbers of punctate reticulocytes in a cat with mild anemia of sufficient duration to allow bone
marrow response suggests regenerative anemia, even if the aggregate reticulocyte count is not increased.
Whereas, an absence of an increase in punctate reticulocytes in the peripheral blood of a cat with mild
anemia of sufficient duration to allow bone marrow response suggests nonregenerative anemia unless
an increase in aggregate reticulocytes occurs. In cats, an increase in the aggregate reticulocytes without
concurrent increase in punctate reticulocytes suggests early bone marrow response to anemia. In a few
days, the increased numbers of aggregate reticulocytes will mature, causing an increase in punctate
reticulocyte numbers. Cats with moderate to severe anemia of sufficient duration to allow bone marrow
response should have an increased aggregate reticulocyte count. If they do not, the anemia is very likely
nonregenerative or poorly regenerative regardless of the punctate reticulocyte count. For example, a cat
that has had a packed cell volume (PCV) of 10 for a week and has an aggregate reticulocyte count of
0.1% and a punctate reticulocyte count of 25% has a nonregenerative anemia. If punctate reticulocytes
are counted as aggregate reticulocytes, nonregenerative or poorly regenerative anemias may be misclas-
sified as regenerative anemias; however, failure to count punctate reticulocytes may cause some ade-
quately regenerative mild anemias to be misclassified as inadequately regenerative.
Absolute Reticulocyte Count
The absolute reticulocyte count is the number of reticulocytes per microliter (reticulocytes/µL) of blood.
The absolute reticulocyte count more accurately reflects the bone marrow’s response compared with the
percent reticulocyte count because it automatically adjusts for the decrease in mature RBC numbers,
preventing a misimpression of increase in reticulocytes that may be given by percent reticulocyte count
during anemia. For example, a dog that normally has a PCV of 40%, an RBC count of 6,000,000
reticulocytes/µL, and a percent reticulocyte count of 1% would have an absolute reticulocyte count of
60,000/µL (1% of 6,000,000 = 60,000). If the dog becomes severely anemic with the hematocrit (HCT)
and RBC count dropping to one fourth of the original values (10 and 1,500,000 reticulocytes/µL, respec-
tively), a reticulocyte count of 3% would appear to indicate increased bone marrow reticulocyte produc-
tion, when, in reality, a 3% reticulocyte count with a 1,500,000/µL RBC count represents only 45,000
reticulocytes/µL. Hence, although the percent reticulocyte count was increased, the actual number of
APPENDIX 271
BOX 2 Calculations for Determining the Basal Reticulocyte Production Rate
A. The basal reticulocyte production rate (BRPR) is used to make adjustments for
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decreased sequestration of reticulocytes and increased pore size of marrow

endothelial cells during anemia and to adjust for the decreased number of mature
erythrocytes present during anemia.
1. Basal production rate is useful in dogs only.
2. Basal production rate greater than 1 is considered a responding anemia.
B. Calculation of BRPR:
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observed packed
cell volume (PCV ) 1
reticulocyte coun × × = BRPR
normal PCV correction factor
(usually considered [see below ]
to be 40)
C. Correction factors for the various hematocrit (HCT) values are given below:
HCT Correction Factor
≥35% 1.0
25–35% 1.5
15–25% 2.0
≤15% 2.5
D. Sample calculation of BRPR for a dog with an HCT of 10% and a reticulocyte count
of 15%:
10 1
15 × × = 15 × 0.25 × 0.4 = 1.5 BRPR
40 2.5
reticulocytes in peripheral blood was decreased, indicating a nonregenerative, or poorly regenerative,

anemia.
Basal Reticulocyte Production Rate
In dogs, a basal reticulocyte production rate (BRPR) has been determined empirically and may be par-
ticularly useful for evaluating a regenerative response when only HCT and percent reticulocytes are
known. The BRPR adjusts for the decrease in numbers of mature RBCs, the increased time that reticu-
locytes circulate as reticulocytes because of their early release from bone marrow during anemia, and
the expected proportional increase in erythropoiesis as the severity of anemia increases. To determine
the BRPR, the patient’s percent reticulocyte count is multiplied by the patient’s actual HCT divided
by the patient’s expected normal HCT (usually predicted as 40), and the resulting value is divided by
the appropriate BRPR correction factor. BRPRs above 1 indicate adequate bone marrow response to the
anemia, whereas those less than 1 indicate inadequate bone marrow response to the anemia. Thus far,
BRPR correction factors have been determined only for dogs. The formula and the correction factors
used for calculating the BRPR are given in Box 2 along with a sample calculation.
Saline Dilution or Dispersion Test

RBC autoagglutination and rouleaux formation may be differentiated by saline dilution. Saline dilution
may be performed by mixing equal amounts of physiologic saline and EDTA-anticoagulated blood,
placing a drop of the mixture on a microscope slide, then placing a coverslip over the saline–blood
mixture, and examining the unstained mixture microscopically for clumping. If the clumping is caused
by autoagglutination, cell clumps will persist after saline dilution, and an unorganized or random
(agglutinated) association of RBCs will be recognized. If clumping is caused by rouleaux formation, the
cells will either disperse into individual cells, or an organized side-to-side rouleaux pattern (resembling
a roll of coins) will easily be recognized.
INDEX
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Page numbers followed by “f” indicate figures, “t” indicate tables, and “b” indicate boxes.
A Acute myeloid leukemia (AML), Anaplasma phagocytophilium, 189

Absolute polycythemia 242–246 Anaplasma platys, 232
causes, 254 acute erythroleukemia, 243 appearance, 232f–233f
nonneoplastic etiologies, 254 acute megakaryoblastic leukemia, diagnostic significance, 232
Absolute reticulocyte count, 26, 270 246 features, 232
Abyssinian cats, hereditary RBC acute monocytic leukemia, 243 Anaplasma spp., 188
defect, 64 acute myeloblastic leukemia, 242 diagnostic significance, 188
Acanthocheilonema reconditum, 260 acute myelomonocytic leukemia, features, 188
Acanthocytes (spur cells), 62 243 Anemia, 26
appearance, 62f–63f acute promyelocytic leukemia, blood smears, appearance, 27f
diagnostic significance, 62 242–243 bone marrow, effectiveness, 36
DIC/lymphoma, appearance, acute undifferentiated leukemia, diagnosis, 26
62 242 diagnostic significance, 26
features, 62 dysplastic changes, 242 differentials, 26
in vitro artifactual changes, 46 MDS, contrast, 242 features, 26
occurrence, 62 Acute myelomonocytic leukemia immune-mediated component,
Acanthocytosis, 54 (AML-M4), 243 existence, 94
Activated platelets, 226 diagnostic significance, 243 mechanisms, 26
appearance, 226f–227f features, 243 regenerative characteristic, 34, 36
diagnostic significance, 226 Acute promyelocytic leukemia Anisocytosis, 40
features, 226 (AML-M3), 242–243 diagnostic significance, 40
Acute blood loss, 26 diagnostic significance, 243 features, 40
Acute erythroleukemia (AML-M6), features, 242–243 Antigen, RBC binding, 32
243 Acute undifferentiated leukemia Artifacts, hypochromic RBCs
appearance, 244f–245f (AUL), 242 (differentiation), 38
diagnostic significance, 243 features, 242 Artifactual leukopenia, appearance,
features, 243 Agglutination, 32 116f
subtypes, 243 absence, 50 Artifactual thrombocytopenia, 221f
Acute leukemia, blast cell appearance, 30f–31f AUL. See Acute undifferentiated
population lineage diagnostic significance, 32 leukemia
(identification), 235 features, 32 Azurophilic granules (primary
Acute lymphoblastic leukemia stack-of-coins formation, 32 granules), presence, 130
(ALL), 235–236 Aggregate reticulocytes, 36
appearance, 236f–237f count, increase, 36 B
diagnostic significance, 235–236 Alaskan Malamutes (peripheral B-cell leukemia, 236
features, 235 blood), stomatocytes Babesia canis, 102
Acute megakaryoblastic leukemia (occurrence), 64 Babesia conradae, 102
(AML-M7), 246 ALL. See Acute lymphoblastic Babesia gibsoni (appearance), 76f–77f,
appearance, 246f–247f leukemia 81f, 89f, 104f–105f
diagnostic significance, 246 AML. See Acute myeloblastic Babesia microti-like piroplasms, 102
features, 246 leukemia; Acute myeloid Babesia spp., 102
Acute monocytic leukemia leukemia appearance, 104f
(AML-M5), 243 AML-M3. See Acute promyelocytic diagnostic significance, 102
diagnostic significance, 243 leukemia features, 102
features, 243 AML-M4. See Acute PCR/DNA sequencing, 102
subtypes, 243 myelomonocytic leukemia Babesia vogeli, 102
Acute myeloblastic leukemia AML-M5. See Acute monocytic Background, 10–20
(AML), 242 leukemia basophilia
diagnostic significance, 242 AML-M6. See Acute erythroleukemia increase, 10
features, 242 AML-M7. See Acute increase, appearance, 10f–11f
subtypes, 242 megakaryoblastic leukemia cytoplasmic fragments, 12
273
274 INDEX
Background (Continued) Basset Hound thrombopathia, 218 Canine basophils, appearance,

features, 10 Bee sting envenomation, 60 173f–174f, 179f
fibrin, 18 Berry cells (echinocytes), 60 Canine distemper, appearance, 76f,
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infectious agents, 14 Birman cat anomaly, 200 81f

skin contaminants, 20 appearance, 149f, 196f Canine eosinophils, appearance,
stain precipitation, 16 cytoplasmic granules, association, 166f–167f
Bacteria, 266 200f–201f Canine erythrocytes, appearance, 24
appearance, 15f diagnostic significance, 200 Canine mast cell, appearance,
diagnostic significance, 266 features, 200 173f–174f, 179f
features, 266 mucopolysaccharidosis, 201f Canine monocytes, characteristics,
stain precipitation, 266f primary granules, 201f 164
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Bacterial contamination, toxic granulation, 201f Canine monocytic ehrlichiosis,

extracellular bacteria Blast cell population, lineage cause, 188
(presence), 266 (identification), 235 Canine neutrophil
Bacterial infections, 118 Blister cells (keratocytes), 54 distemper inclusion, Diff-Quik
Bacterial overgrowth, occurrence, Blood chemistry parameters, stain (usage), 189f
266 checking, 28 Ehrlichia morulae, presence, 187f
Band neutrophils, 120 Blood smears, 3–20 Canine red blood cell (canine RBC),
appearance, 121f–123f, 139f anemic blood smears, appearance, 24f–25f
diagnostic significance, 120 appearance, 27f Canine reticulocytes, release, 36
features, 120 assessment, 28 Cardiac auscultation, usage, 52
left shift, 120 background, 10–20 Cardiac diseases, 52
Barr body, 132 body, 8 exclusion, 28
appearance, 132f–133f delayed blood smear preparation, Cavalier King Charles Spaniels,
diagnostic significance, 132 impact, 210f–211f congenital
features, 132 diagnostic significance, 3 macrothrombocytopenia, 228
X chromosome representation, differential WBC counts, 164 Chagas disease (Trypanosoma cruzi),
132 evaluation, importance, 235 262
Basal reticulocyte production rate, feathered edge, 6 Chédiak-Higashi syndrome (CHS),
271 features, 3 148, 198, 218
determination, calculations, 271b microscopic evaluation, usage, cytoplasmic granules, association,
Basket cells (smudge cells), 206 235 198f–199f
appearance, 206f–207f monolayer, 4 features, 198
diagnostic significance, 206 preparation, delay, 134 Chemotherapeutic agents, usage, 118
features, 206 review, usage, 94 Chromatic clumps
Basopenia, 172 Blood urea nitrogen (BUN), 28 (heterochromatin), presence,
Basophilia Blood, myeloblasts (presence), 154
cytoplasmic basophilia, 242 Chronic blood loss, 26
appearance, 144f–145f Blue-smoke Persian cats, Chediak- anemia, 269
increase, 10 Higashi syndrome, 148 Chronic ehrlichiosis, 118
appearance, 10f–11f Body, 8 Chronic granulocytic leukemia, 126,
diagnostic significance, 10 appearance, 8f–9f 248
features, 10 diagnostic significance, 8 appearance, 249f–251f
Basophilic cytoplasm, presence, features, 8 diagnostic significance, 248
130 Bone marrow features, 248
Basophilic leukemia, 172 cellular dysplasia, evidence, 90 Chronic leukemias, development,
Basophilic staining cellular cytology, 28 235
organelles (ribosomes), disease (myelophthesis), 74 Chronic lymphocytic leukemia
presence, 34 disorders, 44 (CLL), 238
Basophilic stippling, 78 effectiveness, 36 appearance, 238f–239f
appearance, 76f, 79f–80f, 89f infiltration, 118 diagnosis, CBC monitoring, 238
diagnostic significance, 78 megakaryocytes (platelet diagnostic significance, 238
features, 78 progenitors), 216 features, 238
occurrence, 78 monocytes, presence, 252 Chronic monocytic leukemia, 252
siderotic inclusions, contrast, 78 myeloblasts, presence, 242 appearance, 256
Basophils, 172 necrosis, 118 features, 252
canine basophils, appearance, neoplasia, 218 Chronic myeloid leukemia, 248–254
172f–174f, 179f platelet progenitors, 230 chronic granulocytic leukemia,
diagnostic significance, 172 Borrelia burgdorferi (impact), 264 248
features, 172 Breed-associated disorders, 42 chronic monocytic leukemia, 252
feline basophils, appearance, 172f, Burr cells (echinocytes), 60 chronic myelomonocytic
174f–175f leukemia, 254
granules, numbers, 172 C polycythemia vera, 254
numbers, decrease (basopenia), C-shaped nucleus, 120 serial CBC examinations,
172 Calculi, 38 requirement, 248
INDEX 275
Chronic myelomonocytic leukemia, Cytoplasmic vacuolization (foamy Echinocytes (Continued)

254 cytoplasm), 146 crenated erythrocytes, 60
features, 254 appearance, 147f diagnostic significance, 60
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CHS. See Chédiak-Higashi syndrome diagnostic significance, 146 features, 60

Circulating lymphoblasts, presence, features, 146 in vitro artifactual changes, 46
236 Echinocytosis, 54
Circulating neoplastic cells, D Echocardiography, usage, 52
checking, 50 Delayed processing, changes, 210 Ectoparasites, 52
Clinical bleeding, evidence, 218 diagnostic significance, 210 Ehrlichia chaffeensis (impact), 188
CLL. See Chronic lymphocytic features, 210 Ehrlichia equi, 188
leukemia Diabetes mellitus, Heinz bodies Ehrlichia morulae
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Codocytes (target cells), 48 (presence), 84 appearance, 189f–190f

Complete blood count (CBC) Diff-Quik presence, 189f, 209f
monitoring, 238, 240 stain, usage, 84 Ehrlichia phagocytophilia, 188
results, 112 usage, 34 Ehrlichia platys, 232
serial CBC examinations, Diff-Quik-stained smears, usage, Ehrlichia spp., 188
requirement, 248 186 diagnostic significance, 188
usage, 235 Differential cell counts, estimation, features, 188
Congenital disorders, 44 4 Ehrlichial infections, 26
Congenital dyserythropoiesis, 48 Diffuse cytoplasmic basophilia, 144 Ehrlichiosis, characteristics, 188
Congenital macrothrombocytopenia diagnostic significance, 144 Electrocardiography, usage, 52
(Cavalier King Charles features, 144 Endogenous glucocorticoids, excess,
Spaniels), 228 Dipetalonema reconditum, 260 164
Coombs test, 50 Dirofilaria immitis, 260 Endogenous oxidants, elaboration,
Coombs-negative hemolytic anemia, Discocytes, morphology, 24 56
recurrence, 64 Disseminated intravascular Endoparasites, 52
Crenated erythrocytes (echinocytes), coagulation (DIC) Endotoxemia, 118
60 appearance, 62 Eosinophilic chronic granulocytic
Crystallized hemoglobin, 66 schistocytes, occurrence, 52 leukemia, appearance, 251f
appearance, 66f–67f Distemper inclusions, 88, 186 Eosinophilic leukocyte granules,
diagnostic significance, 66 appearance, 88f–89f, 186f–187f presence, 198
features, 66 diagnostic significance, 88, 186 Eosinophils, 166
Cushing’s disease Diff-Quik stain, usage, 189f canine eosinophils, appearance,
(hyperadrenocorticism), 70, 254 features, 88, 186 166f–167f
Cyclic hematopoiesis, 118 identification, 88 diagnostic significance, 166
Grey Collies, 218 peripheral blood smears, usage, features, 166
Cytauxzoon felis, 98 186 feline eosinophils, appearance,
appearance, 99f–101f Wright-stained smears, 88, 186 168f–169f
diagnostic significance, 98 DNA sequencing, 102 EPO. See Erythropoietin
features, 98 Döhle bodies, 140 Erythrocyte refractile (ER) bodies,
organisms, Mycoplasma hemofelis appearance, 141f–142f 84
organisms (contrast), 98 diagnostic significance, 140 Erythrocytic ghosts
safety pin/Maltese cross forms, features, 140 appearance, 68f–69f
numbers (decrease), 98 granules, 198 diagnostic significance, 68
signet ring appearance, 98 Doughnut form (ring form), 150 fading erythrocytes, 68
Cytauxzoon, appearance, 77f Drentse Partirjshond dogs (Dutch features, 68
Cytauxzoonosis, impact, 98 Partridge dogs), multiorgan lysed red blood cells, 68
Cytoplasm, RBC membrane excess disease, 64 Erythrocytic parasites, 92–102
(comparison), 46 Drug-induced neutropenia, 118 Babesia spp., 102
Cytoplasmic basophilia, Dyserythropoiesis, 82 Cytauxzoon felis, 98
appearance, 144f–145f Dysplastic changes, 90 Mycoplasma hemocanis, 92
Cytoplasmic fragments, 12 appearance, 90f–91f Mycoplasma hemofelis, 94
appearance, 13f diagnostic significance, 90 Erythroid production, response, 28
diagnostic significance, 12 features, 90 Erythroid toxins, 74
features, 12 Dysplastic nRBCs, presence, 90 Erythroleukemia, 42
Cytoplasmic granules Erythrophage
Birman cat anomaly, appearance, E (erythrophagocytosis), 182
200f–201f Eccentrocytes (hemighosts), 56 appearance, 182f–183f
Chédiak-Higashi syndrome, appearance, 56f–57f diagnostic significance, 182
association, 198f–199f diagnostic significance, 56 features, 182
mucopolysaccharidosis, features, 56 identification, peripheral blood
association, 196f–197f Echinocytes, 60 smears (usage), 182
Cytoplasmic vacuoles, GM1 appearance, 60f–61f Erythropoietin (EPO)
gangliosidosis (association), berry cells, 60 levels, measurement, 28
202f–203f burr cells, 60 production, 254
276 INDEX
Essential thrombocythemia, 254 Fibrin (Continued) Helmet cells (keratocytes), 54

appearance, 254f–255f microfilaria, confusion Hemangiosarcoma, 52
diagnostic significance, 254 (avoidance), 18 Hematocrit
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features, 254 strand, appearance, 260f estimation, 3

Ethylenediaminetetraacetic acid Fingerprints, skin contaminants measurement, 3
(EDTA), 44 (collection), 20 Hematopoiesis
anticoagulant, feline blood FIV. See Feline immunodeficiency ineffectiveness, 90
storage, 54 virus leukemoid response, acceleration,
bacterial overgrowth, occurrence, Foamy cytoplasm (cytoplasmic 164
266 vacuolization), 146 Hematopoietic neoplasia, overview,
EDTA-anticoagulated blood, 134 Folded red blood cells, 48 235
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EDTA-induced cytoplasmic Fragmented erythrocytes Hematuria, 52

vacuolization rate, 146 (schistocytes), 52 Hemighosts (eccentrocytes), 56
Exogenous glucocorticoids, excess, Hemobartonella canis, 92
164 G Hemobartonella felis, 94
Exogenous oxidants, 56 Gastrointestinal blood loss, 52 Hemoglobin crystals, presence,
External hemorrhage, 36 Ghost RBC, diagnosis, 64 66
Extracellular bacteria Giant neutrophils, 152 Hemoglobin deficient mature
appearance, 17f appearance, 152f–153f erythrocytes, 38
presence, 266 diagnostic significance, 152 Hemolysis, 26
stain, distinction, 16f–17f features, 152 Hemolytic anemia, 82
Extracellular infectious agents, Glanzmann’s thrombasthenia, Hemoparasites
appearance, 15f 218 checking, 50
Glomerulonephritis, 52 investigation, 26
F Glucocorticoid therapy, 74 stain, confusion, 16
Fading erythrocytes (erythrocytic GM. See Granulocyte macrophage Hemosiderin, iron-containing
ghosts), 68 Gram-negative bacteria, infection, pigment, 180
Familial stomatocytosis- 118 Hepatozoon americanum (presence),
hypertrophic gastritis, 64 Granular lymphocytes (large 192
Feathered edge, 6 granular lymphocytes), 162 Hepatozoon canis, 192
appearance, 7f appearance, 162f–163f Hepatozoon gametocytes,
diagnostic significance, 6 diagnostic significance, 162 appearance, 192f–193f
features, 6 features, 162 Hepatozoon spp., 192
leukocyte concentration, 116f lymphocytic leukemia, 240 diagnostic significance, 192
thrombocytopenia, 219f Granulocyte macrophage (GM) features, 192
Feline aggregate reticulocytes gangliosidosis, 202 Hepatozoonosis, cause, 192
maturational stage, 269 cytoplasmic vacuoles, association, Heterochromatin (chromatin
release, 36 202f–203f clumps), presence, 154
Feline basophils, appearance, 172f, features, 202 Histoplasma capsulatum, 194
174f–175f Granulocytic leukemia, possibility, diagnostic significance, 194
Feline blood storage, 54 130 features, 194
Feline eosinophils, appearance, Great Pyrenees, Glanzmann’s organisms, characteristics, 194
168f–169f thrombasthenia, 218 Histoplasma yeast
Feline erythrocytes, appearance, 24 Grey Collies, cyclic hematopoiesis, appearance, 194f–195f
Feline immunodeficiency virus 218 organisms, presence, 194
(FIV) infection, 164 Grey eosinophils (sight hounds), Howell-Jolly bodies (H-J bodies),
Feline leukemia virus (FeLV), 26, appearance, 170f–171f 74
118 Greyhounds, gray eosinophils, appearance, 75f–77f, 81f, 89f
FeLV associated myelodysplastic 166 diagnostic significance, 74
syndrome, 42 features, 74
FeLV-induced myelodysplasia, H Hyperadrenocorticism (Cushing’s
90 Heartworm testing, usage, 52 disease), 70, 254
myelodysplasia, 152 Heavy metal toxicities, 36 Hyperalbumenemia, presence,
Feline mast cell, appearance, 172f Heinz bodies, 84 254
Feline monocytes, characteristics, appearance, 85f Hyperglobulinemia, 160, 218
164 canine blood, relationship, 46 Hypersegmented neutrophils,
Feline platelets, characteristics, diagnostic significance, 84 134
218 Diff-Quik stain, usage, 84 appearance, 134f–135f
Feline red blood cells (feline RBCs) features, 84 diagnostic significance, 134
appearance, 24f–25f hemolytic anemia, causes, 84 features, 134
pallor, absence, 50 NMB, usage, 84 Hyperthyroidism, Heinz bodies
Fibrin, 18 appearance, 87f (presence), 84
appearance, 19f pale staining Heinz bodies, Hypochromia, 38
diagnostic significance, 18 appearance, 86f diagnostic significance, 38
features, 18 Wright stains, 84 features, 38
INDEX 277
Hypochromic RBCs Leukemia (Continued) Macroplatelets (megaplatelets),

appearance, 38f–39f chronic lymphocytic leukemia 228
artifacts, differentiation, 38 (CLL), 238 appearance, 228f–229f
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Hypochromic RBCs, development, classification schemes, proposal, diagnostic significance, 228

38 235 features, 228
Hyposegmentation, occurrence, 134 lymphoid leukemia, 235 Mast cells, 176
Hypoxia, response, 28 myeloid leukemia (nonlymphoid appearance, 177f
leukemia), 235 canine mast cell, appearance,
I T-cell large granular lymphocytic 173f–174f, 179f
Immature neutrophils, release, 120 leukemia, 240 feline mast cell, appearance,
Immune-mediated anemia, 243 Leukemoid response, 118 174f
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Immune-mediated hemolytic Leukocyte count, 112 Mature neutrophils (segmented

anemia (IMHA), 44 appearance, 112f–113f neutrophils), 118
diagnosis, 50 diagnostic significance, 112 diagnostic significance, 118
evidence, 64 Leukocytes features, 118
disorder, 180 pyknosis, occurrence, 204 numbers, decrease (neutropenia),
support, 32 staining problems, 212f–213f 118
Immune-mediated neutropenia, 118 Leukocytosis, 114 Mature segmented neutrophils,
Immune-mediated RBC destruction, appearance, 114f–115f appearance, 119f–121f
26 diagnostic significance, 114 Megakaryoblasts, characteristics,
Immunoglobulin G (IgG) antibody Leukopenia, 116 246
coating, 32 appearance, 116f–117f Megakaryocytes, 230
Immunostimulation, etiologies, 240 diagnostic significance, 116 appearance, 230f–231f
Imperfect spherocytes, 64 Lipemic blood samples, impact, 30 diagnostic significance, 230
In vitro artifactual changes, 46 Liver disease, 48 features, 230
Inclusions, 186–202 Lymphoblastic leukemia, peripheral Megaplatelets (macroplatelets),
Infectious agents, 14, 186 blood (presence), 235–236 228
extracellular infectious agents, Lymphocytes, 154–162 Membrane loss (spherocytes), 46
appearance, 15f clonal populations, identification, Metamyelocytes, 126
features, 14 238 appearance, 127f
Inherited disorders, 42 examples, 73f diagnostic significance, 126
Internal hemorrhage, 36 granular lymphocytes (large features, 126
Intracytoplasmic granules, presence, granular lymphocytes), 162 nuclear chromatin, recognition,
196 plasmacytoid reactive 126
Iron deficiency, 36 lymphocytes, 160 Metarubricytes (nucleated red blood
anemia, 38, 48, 52 reactive lymphocytes, 158 cells), 70
small lymphocytes, 154 Microcytes, appearance, 44f–45f
K Lymphocytosis, distinguishing, Microcytosis, 44
Keratocytes (blister cells/helmet 154 diagnostic significance, 44
cells), 54 Lymphoglandular bodies, 12 features, 44
appearance, 54f–55f Lymphoid leukemias, 235–240 Microfilaria, 260
diagnostic significance, 54 acute lymphoblastic leukemia, appearance, 15f, 260f–261f
features, 54 235–236 diagnostic significance, 260
formation, RBC trauma (impact), chronic lymphocytic leukemia, features, 260
54 238 fibrin, confusion (avoidance), 18
T-cell large granular lymphocytic Miniature Schnauzers, hereditary
L leukemia, 240 stomatocytosis, 64
Large granular lymphocytes, 162 Lymphoma Mitoses, aberrations, 256
presence, 162 appearance, 62 Mitotic figures, 256
Large-diameter RBCs, impact, 40 Heinz bodies, presence, 84 appearance, 256f–257f
Lead poisoning, 82 Lysed red blood cells (erythrocytic diagnostic significance, 256
Leptocytes ghosts), 68 features, 256
appearance, 48f–49f Monocytes, 164
diagnostic significance, 48 M appearance, 164f–165f
features, 48 Macrocytes, appearance, 42f–43f characteristics, 243
folded red blood cells, 48 Macrocytosis, 42 diagnostic significance, 164
pathologic states, 48 diagnostic significance, 42 Ehrlichia morulae, presence,
target cells (codocytes), 48 features, 42 209f
Leukemia poodles, 136 features, 164
acute leukemia, blast cell reticulocytosis, relationship, 42 intracytoplasmic hemosiderin
population lineage Macrophages, 184 pigment, presence, 180
(identification), 235 appearance, 185f peripheral blood precursors, 184
categories, 235 diagnostic significance, 184 presence, 252
chronic leukemias, development, features, 184 Monocytic cell line, neoplasia, 164
235 phagocytic activity, 184 Monocytosis, 164
278 INDEX
Monolayer, 4 Mycoplasma organisms New methylene blue (NMB)

appearance, 4f–5f confirmation, blood smear review appearance, 87f
diagnostic significance, 4 (usage), 94 stain, usage, 36
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features, 4 presence, 94 usage, 84, 269b

leukocytes, absence (appearance), Myeloblasts, 130 Nonanemic patient, basophilic
116f appearance, 130f–131f stippling (presence), 78
platelets, absence, 221f diagnostic significance, 130 Nonlymphoid leukemias (myeloid
RBCs, apposition, 3 features, 130 leukemias), 235
Morphologic changes, disease presence, 242 Nonneoplastic absolute
(association), 30–108 Myelocyte, 126 polycythemia, 254
acanthocytes (spur cells), 62 Myelocytes Normocytes, morphology, 24
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agglutination, 32 appearance, 127f Nuclear chromatin, recognition,

anisocytosis, 40 diagnostic significance, 127 126
eccentrocytes (hemighosts), 56 features, 127 Nucleated erythrocytes, presence,
echinocytes (crenated magenta-staining granules, 70
erythrocytes/burr cells/berry 128 numbers, increase (occurrence),
cells), 60 Myelodysplastic syndrome (MDS), 70
folded red blood cells, 48 42 Nucleated red blood cells (nRBCs)
hypochromia, 38 AML, contrast, 242 appearance, 71f–72f, 155f
keratocytes (blister cells/helmet dysplastic changes, 90 diagnostic significance, 70
cells), 54 Myelofibrosis, 52 dysplastic nRBCs, presence, 90
leptocytes, 48 Myeloid leukemias (nonlymphoid features, 70
macrocytosis, 42 leukemias), 235 immature stages, 70
microcytosis, 44 Myelophthesic diseases, 118 increase, 70, 74
poikilocytes, 46–64 Myelophthesis (bone marrow metarubricytes, 70
polychromasia, 34 disease), 74 rubricytes, 70
pyknocytes, 58 Myeloproliferative disorders, 82 significance, 70
reticulocytes, 36
rouleaux, 30 N O
schistocytes (fragmented Neoplasia, 38 Open chromatin, appearance, 139f
erythrocytes), 52 diagnosis, absence, 238 Otterhounds, Glanzmann’s
spherocytes, 50 Neoplastic lymphocytes, thrombasthenia, 218
stomatocytes, 64 characteristics, 158 Oxidative RBC damage, 26
Morphology Nephrotic syndrome, 48
discocytes/normocytes, 24 Neutropenia, decrease, 118 P
diagnostic significance, 24 Neutrophilic chronic granulocytic Packed cell volume (PCV)
features, 24 leukemia, appearance, aggregate reticulocyte count,
platelets, 216 249f–250f 269
Morphology variation, 60 Neutrophils, 118–139 measurement, 3
Morula, 188 abundance, 118 Pale staining Heinz bodies,
Mucopolysaccharidosis (MPS), 196 band neutrophils, 120 appearance, 86f
appearance, 196f Barr body, 132 Panhyperproteinemia, presence,
Birman cat anomaly, appearance, delayed blood smear preparation, 254
200f impact, 210f–211f Paraneoplastic eosinophilia, 166
cytoplasmic granules, association, giant neutrophils, 152 Parasites, 186–202, 232
196f–197f hypersegmented neutrophils, Anaplasma platys, 232
diagnostic significance, 196 136 Partial thromboplastin time (PTT),
features, 196 inclusion, 148 218
Mycoplasma hemocanis intracytoplasmic hemosiderin Pelger-Huët anomaly
appearance, 17f, 92f–93f, 266f pigment, presence, 180 appearance, 137f–139f
diagnostic significance, 92 mature neutrophils (segmented identification, 136
features, 92 neutrophils), 118 pseudomyelocytes, appearance,
Hemobartonella canis, 92 metamyelocyte, 126 126
infection, impact, 92 morphologies, similarities, Pelger-Huët neutrophils, 136
polymerase chain reaction (PCR) 118 appearance, 130f, 139f
testing, usage, 92 myeloblast, 130 diagnostic significance, 136
Mycoplasma hemofelis, 94 myelocyte, 126 features, 136
appearance, 17f, 95f–97f, numbers, increase, 114 Perinuclear clear zone (Golgi
266f–267f Pelger-Huët neutrophils, 136 region), visibility, 160
Cytauxzoon felis organisms, platelets, 189f Perinuclear Golgi region, contrast,
contrast, 98 promyelocyte, 130 158
diagnostic significance, 94 Pseudo-Pelger-Huët neutrophils, Peripheral blood
Hemobartonella felis, 94 136 cellular dysplasia, evidence, 90
infection, 243 release, 118 EDTA exposure, 146
organisms, dislodging, 16 toxic changes, evidence, 50 monocytes, presence, 252
INDEX 279
Peripheral blood (Continued) Polychromatophils, 40 Recurrent Coombs-negative

presence, 235–236 Polycythemia, 28 hemolytic anemia, 64
smears appearance, 29f Red blood cell artifacts,
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myeloblasts, presence (rarity), diagnostic significance, 28 106–108

242 features, 28 platelet over red blood cell,
rubriblasts, absence, 243 identification, 28 108
usage, 182, 186 nonneoplastic absolute refractile artifact, 106
Plasmacytoid reactive lymphocytes, polycythemia, 254 Red blood cells (RBCs)
160 RBCs, increase, 28 agglutination
appearance, 160f–161f relative polycythemia, 254 diagnosis, 64, 102
diagnostic significance, 160 Polycythemia vera (PV), 254 presence, 32
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features, 160 diagnostic significance, 254 canine red blood cells,

Platelet clumps features, 254 appearance, 24f–25f
analysis, 218 impact, 28 cytoplasm
appearance, 224f–225f Polymerase chain reaction (PCR) ribosomes, concentration,
diagnostic significance, 224 sequencing, 102 269
discovery, 6 testing, usage, 92 spherical structures, 74
features, 224 Polyribosomes, concentration feline red blood cells, appearance,
Platelet over neutrophil, (impact), 269 24f–25f
appearance, 189f, 191f Poodles folded red blood cells, 48
Platelet over red blood cell, 108 macrocytosis, 136 fragmentation, 26, 46
appearance, 108f–109f marrow dyscrasia, 42 immune-mediated RBC
diagnostic significance, 108 Postsplenectomy state, 74 destruction, 26
features, 108 Prerenal azotemia, presence, 254 increase, 28
Platelet over white blood cells, 208 Primary granules linear stacks, formation, 30
appearance, 208f–209f appearance, 148f, 197f membranes
diagnostic significance, 208 Birman cat anomaly, appearance, abnormalities, 46
features, 208 200f hemoglobin, absence/
Platelets Promyelocytes, 130 minimum, 68
activated platelets, 226 appearance, 130f–131f immunoglobulin G (IgG)
appearance, 216f–217f atypical appearance, 242–243 antibody coating, 32
clumping, feline problem, 224 diagnostic significance, 130 monolayer, apposition, 3
feline platelets, characteristics, features, 130 morphologic abnormalities,
218 Propylene glycol, exogenous 50
function, requirement, 216 oxidants, 56 morphology, 40
morphology, 216 Prothrombin time (PT), 218 nuclei, fragments (retention),
diagnostic significance, 216 Pseudo-Pelger-Huët neutrophils, 74
features, 216 136 numbers, decrease, 26
numbers, evaluation, 26 diagnostic significance, 136 oxidative injury, 56
progenitors (bone marrow features, 136 oxidative RBC damage, 26
megakaryocytes), 216 Pseudomyelocytes, appearance, production, increase, 28, 254
Poikilocytes, 46–64 127f projections, 62
appearance, 46f–47f Pseudothrombocytopenia, 224 regeneration, 64
diagnostic significance, 46 Pulmonary diseases, exclusion, smudges, appearance, 68
features, 46 28 survival time, increase, 70
keratocytes, 54 PV. See Polycythemia vera trauma, 54
leptocytes, 48 Pyknocytes, 58 Refractile artifact, 106
pyknocytes, 58 appearance, 59f diagnostic significance, 106
schistocytes, 52 diagnostic significance, 58 features, 106
spherocytes, 50 features, 58 Refractile artifact of red blood cell,
stomatocytes, 64 membranous tag, 58 appearance, 106f–107f
Polychromasia, 34 Pyknosis, occurrence, 204 Regenerative anemias, 48
absence/minimum, 42 Pyknotic cells, 204 macrocytosis, expectation,
appearance, 34f–35f appearance, 204f–205f 42
correlation, 36 diagnostic significance, 204 rubriblasts, absence, 243
diagnostic significance, 34 features, 204 Relative polycythemia, 254
Diff-Quik, usage, 34 presence, indication, 204 Renal diseases, 48
features, 34 exclusion, 28
usage, 34 R Reticulocytes, 36
Polychromatophilic erythrocytes, Reactive lymphocytes, 158 appearance, 36f–37f
number (increase), 34 appearance, 159f basal reticulocyte production rate,
Polychromatophilic RBCs diagnostic significance, 158 calculations, 271b
basophilic stippling, evidence, features, 158 canine reticulocytes, release,
78 plasmacytoid reactive 36
differentiation, difficulty, 34 lymphocytes, 160 diagnostic significance, 36
280 INDEX
Reticulocytes (Continued) Small mature lymphocytes, Thrombocytopenia (Continued)

features, 36 appearance, 155f–157f diagnostic significance, 218
new methylene blue (NMB), Small-diameter RBCs, impact, feathered edge, 219f
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usage, 269b 40 features, 218

punctuate reticulocytes, 36 Smear preparation, storage pseudothrombocytopenia,
Reticulocytosis, 34 (prolongation), 60 224
absence/minimum, 42 Smear stains, background, 10 true thrombocytopenia, evidence,
Ribosomes (basophilic staining Smoke blue Persian cats, 218
cellular organelles), presence, eosinophilic leukocyte granules Thrombocytosis, 222–228
34 (presence), 198 activated platelets, 226
Rickettsial infections, 118 Smudge cells (basket cells), appearance, 222f–223f
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Ring form (doughnut form), 150 206 diagnostic significance, 222

diagnostic significance, 150 Snake bit envenomation, 60 features, 222
features, 150 Somali cats, hereditary RBC defect, macroplatelets (megaplatelets),
Ring-form neutrophils, appearance, 64 228
150f–151f Spherocytes, 40, 102 platelet clumps, 224
Romanowsky-type stains, usage, appearance, 50f–51f Tissue macrophages, monocytes
269 diagnostic significance, 50 (peripheral blood precursors),
Rouleaux, 30 features, 50 184
appearance, 30f–31f membrane loss, 46 Torocytes, 38
diagnostic significance, 30 Spiculated erythrocytes, 60 Total protein concentration, 24
features, 30 Spirochetes, 262 Toxic changes, 140–152
formation appearance, 15f, 226f, 264f–265f cytoplasmic vacuolization (foamy
increase, observation, 30 diagnostic significance, 264 cytoplasm), 146
Rubriblasts, absence, 243 features, 264 diffuse cytoplasmic basophilia,
Rubricytes (nucleated red blood Splenic disease, 74 144
cells), 70 Spreader smear, pressure Döhle bodies, 140
(application), 6 giant neutrophils, 152
S Spur cells (acanthocytes), 62 ring form (doughnut form),
S-shaped nucleus, 120 Stain precipitant, appearance, 150
Salmonella (bacterial infections), 16f–17f toxic granulation, 148
118 Stain precipitation, 16 Toxic granulation, 148
Schistocytes (fragmented appearance, 266f appearance, 148f–149f, 197f
erythrocytes), 52 diagnostic significance, 16 Birman cat anomaly, appearance,
appearance, 52f–53f features, 16 200f
diagnostic significance, 52 Standard Schnauzers, hereditary diagnostic significance, 148
features, 52 stomatocytosis, 64 features, 148
occurrence, 52 Steroid administration, 70 Trypanosoma cruzi (Chagas disease),
Schistocytosis, 44 Stomatocytes, 64 262
Segmented neutrophils (mature appearance, 64f–65f Trypanosoma spp., appearance,
neutrophils), 118 diagnostic significance, 64 226f
Serial complete blood count (CBC) features, 64 Trypanosomes, 262
examinations, 34 Stomatospherocytes, 64 appearance, 15f, 262f–263f
requirement, 248 Systemic inflammation, impact, diagnostic significance, 262
Sideroleukocytes, 180 140 features, 262
appearance, 180f–181f Trypomastigotes, characteristics,
diagnostic significance, 180 T 262
features, 180 T-cell acute lymphoblastic Type III echinocytes,
Siderotic inclusions, 82 leukemia, light microscopy predomination, 60
appearance, 82f–83f (usage), 236
basophilic stippling, contrast, 78, T-cell large granular lymphocytic U
82 leukemia (T-cell LGL Understained blood films,
diagnostic significance, 82 leukemia), 240, 240f–241f 212f–213f
features, 82 diagnostic significance, 240 Understained smears, 212
iron-containing inclusions, 82 features, 240 diagnostic significance, 212
Sight hounds, grey eosinophils T-cell leukemia, 236 features, 212
(appearance), 170f–171f Target cells (codocytes), 48 Urinalysis (urine specific gravity),
Skin contaminants, 20 Thrombocytopenia, 102, 218 28
appearance, 21f appearance, 219f–221f Urinary tract infection, 38
diagnostic significance, 20 artifactual thrombocytopenia, Urine, blood loss, 38
features, 20 219f
Small lymphocytes, 154 congenital V
diagnostic significance, 154 macrothrombocytopenia Vasculitis, 52
example, 73f (Cavalier King Charles Venipuncture, skin contaminants
features, 154 Spaniels), 228 (collection), 20
INDEX 281
Viral nucleocapsids, aggregates, 88, White blood cell (WBC) artifacts White blood cells (WBCs)
186 (Continued) (Continued)
Vitamin K antagonist intoxication, platelet over white blood cell, line, usage, 26
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56 209 rupture, 206

pyknotic cells, 204 Wright stains, 84
W understained smears, 212 usage, 269
White blood cell (WBC) artifacts, White blood cells (WBCs)
204–212 count, estimates, 6 X
basket cells (smudge cells), 206 differential count, 70 X chromosome, Barr body
delayed processing, changes, 210 estimation, 4 representation, 132
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CANINE AND FELINE

Amy C. Valenciano, DVM, MS, DACVP

3251 Riverport Lane

ATLAS OF CANINE AND FELINE PERIPHERAL BLOOD SMEARS

Copyright © 2014 by Mosby, an imprint of Elsevier Inc.

Library of Congress Cataloging-in-Publication Data

Valenciano, Amy C., author.

Vice President and Publisher: Linda Duncan

Last digit is the print number: 9 8 7 6 5 4 3 2 1

To my parents who taught me the value of honesty

To Deb, for your enduring love and support.

To my mother who gave me my core tenants of faith, honesty and respect; my

Equipment and Supplies

counts, which may be affected by overfilling and underfilling the tube.

monolayer, precluding reliable microscopic examination.

Hematologic Reference Intervals

may be readily identified.

Plate 1-1 Normal Monolayer

Plate 1-1 Normal Monolayer (con’t)

Plate 1-1b Plate 1-1c

Plate 1-1d Plate 1-1e

Plate 1-1f Plate 1-1g

Plate 1-1h Plate 1-1i

point of application of the blood drop.

Plate 1-2 Normal Feathered Edge

Plate 1-2b Plate 1-2c

Plate 1-2d Plate 1-2e

Plate 1-2f Plate 1-2g

space exists between cells.

Plate 1-3 Normal Body

Plate 1-3 Normal Body (con’t)

Plate 1-3b Plate 1-3c

Plate 1-3d Plate 1-3e

Plate 1-3f Plate 1-3g

Plate 1-3h Plate 1-3i

Plate 1-4 Increased Background Basophilia

Plate 1-4 Increased Background Basophilia (con’t)

Plate 1-4b Plate 1-4c

Plate 1-4d Plate 1-4e

Plate 1-4f Plate 1-4g

Plate 1-4h Plate 1-4i

Plate 1-5 Cytoplasmic Fragments

Plate 1-5b Plate 1-5c

Plate 1-5d Plate 1-5e

Plate 1-6 Extracellular Infectious Agents

Plate 1-7 Stain Precipitant

Plate 1-7 Stain Precipitant (con’t)

Make sure not to confuse stain precipitate with A, Mycoplasma hemocanis;

Next Topic Plate 1-7f

Plate 1-7c Plate 1-7i

Plate 1-7a Plate 1-7b

Plate 1-7c Plate 1-7d

Plate 1-7e Plate 1-7f

Plate 1-7g Plate 1-7h

Plate 1-7i Plate 1-7j

Plate 1-8 Fibrin

Plate 1-8b Plate 1-8c

Plate 1-8d Plate 1-8e

ate superficial squamous epithelial cells.

Plate 1-9 Skin Contaminants

Plate 1-9b Plate 1-9c

Normal Morphology (Discocytes and Echinocytes (Crenated Cells, Burr Cells,

Normal Morphology (Discocytes and Normocytes)