Disorders of blood

and blood forming

organs-equine
 Anaemia
Investigation of anaemia in the horse
Introduction
Anemia is defined as an absolute reduction in
the circulating RBC mass.
It is a frequently encountered clinical problem in
the horse.
Differences in the rate of onset and degree of
anemia, coupled with variety in etiology, present
a diagnostic challenge.
Furthermore, unique features of the physiologic
response to anemia in the horse do not lend to
accurate classification of the anemia on the
basis of regeneration, unless a bone marrow
aspirate is obtained.
Hence, it is often helpful to use the clinical signs,
physical findings, and basic laboratory tests to
initially classify the anemia on the basis of
cause, such as blood loss, increased RBC
destruction, or anemia of inadequate
erythropoiesis to further pursue a specific
etiologic diagnosis.
If this approach is unavailing, a bone marrow
aspirate can be performed to determine whether
the anemia is regenerative (i.e., caused by blood
loss or increased RBC destruction) or non-
regenerative due to inadequate erythropoiesis.
Clinical signs and physical findings
The underlying etiology, rate of development,
and extent of reduction of RBC mass determine
the clinical signs.
Whether acute or chronic, significant anemia is
accompanied by pallor of the mucous
membranes and lack of visible episcleral blood
vessels.
Decreased blood viscosity may lead to turbulent
blood flow in the heart and result in a low-grade
systolic murmur typically audible with greatest
intensity over the semilunar valves.
The main physiologic response to anemia
directly reflects the degree of impairment in the
oxygen-carrying capacity of the blood.
Thus with chronic anemia of gradual onset,
signs often are subclinical.
However, the compensatory response to chronic
anemia often is overwhelmed by the challenge
of exercise, whereupon weakness, reduced
performance, or sudden collapse may occur.
In contrast, the physiologic response to acute
anemia is manifested by immediate attempts to
increase oxygen delivery to the vital organs
This is characterized by tachycardia, tachypnea,
and peripheral vasoconstriction.
When these physiologic adaptations are
insufficient, weakness, collapse, blindness,
ataxia, dementia, colic from ileus, laminitis, or
oliguria may ensue as oxygen deprivation to the
respective tissues progresses.
 Acute increase in metabolic demand on the
cardiovascular and hematopoietic systems may
cause a low-grade fever, especially typical of
hemolytic and blood loss anemias.
*****
Blood Loss
Additional clinical signs or physical findings may
provide clues into the etiology.
Signs of hypovolemic shock frequently accompany
acute, severe blood loss.
Evidence of external blood loss may be obvious,
such as the presence of epistaxis, melena,
hematochezia, hematuria, or trauma.
Signs of internal hemorrhage often are less
obvious.
Intrathoracic hemorrhage may cause tachypnea,
shallow rapid breathing, and dyspnea.
Acute intra-abdominal hemorrhage may cause
ileus, colic, or abdominal distention.
Coagulopathy as a cause of blood loss may be
manifested clinically as frank or occult bleeding,
petechial or ecchymotic hemorrhages, or
prolonged bleeding following minor invasive
procedures or trauma.
Because bleeding results in loss of both red
blood cells and protein, symmetric dependent
edema may be an important clinical clue of
hypoproteinemia.
Hemolysis
Icterus is an important clinical feature of
increased RBC hemolysis when the total
bilirubin concentration exceeds 3 mg/d.
Icterus must be carefully evaluated in horses, as
it also is a clinical feature of anorexia and liver
disease.
Furthermore, because the presence of icterus
depends on the rate and extent of RBC
destruction in relation to hepatic clearance of
bilirubin, lack of icterus does not preclude a
diagnosis of hemolysis.
Nonetheless, intense icterus in the horse is
usually the result of hemolysis-more typically
intravascular than extravascular.
Signs of hypovolemia may accompany acute
hemolysis.
Hemoglobinuria is often the first important
clinical clue of intravascular hemolysis but will
not occur unless the renal threshold for
hemoglobin is exceeded.
Both intravascular and extravascular hemolysis
result in icterus of the sclerae, whereas the
reduction in RBCs causes a lack of episcleral
vessels.
Icterus of mucous membranes may overshadow
the pallor of anemia.
Inadequate Erythropoiesis
The clinical signs of nonregenerative anemia are
more obscure because of:
– the slow onset of development,
– the wellestablished compensatory responses,
and
– the fact that this anemia is usually secondary
to a chronic underlying primary disease.
Pallor of the mucous membranes, lack of
episcleral vessels, lethargy, or exercise
intolerance are typical.
Icterus and edema are not features, unless they
are associated with the underlying primary
disease.
*****
Diagnosis of anemia
General
The diagnosis of anemia is confirmed by the
laboratory demonstration of reductions in the
PCV, hematocrit, hemoglobin concentration, and
RBC count.
A reduction in these indices that is not
associated with clinical signs warrants re-
assessment.
Several conditions may complicate accurate
determination of the RBC count and indices and
should be considered in evaluation of the
erythron.
– The PCV and RBC count vary with age, breed, or use
of the horse.
– For example, the PCV of neonates less than 2 days of
age is similar to that of the adult.
– However, the PCV of foals decreases rapidly
in the first week of life, and erythrocytes are
normochromic to hypochromic, normocytic to
hypochromic.
– The PCV typically remains below normal adult
values until approximately 18 months of age.
– In general, well-conditioned or "hot-blooded"
horses, such as the Thoroughbred have a
greater PCV (32%-53%) than poorly
conditioned, "coldblooded" draft breeds or
Miniature Horses (24%-44%).
– Finally, dehydration and splenic contraction
may conceal anemia and should be taken into
account when evaluating the erythron.
RBC morphology may provide some clues to the
cause of the anemia.
Heinz bodies, precipitates of hemoglobin that
are best seen with New Methylene blue stain,
indicate oxidative injury.
Spherocytes and schizocytes indicate immune-
mediated anemia and DIC respectively.
Small nuclear remnants called Howell Jolly
bodies may normally be present in small
numbers in the circulation of horses.
Rouleaux, or a "stacking" appearance to the
peripheral RBCs, is normally extensive in
horses.
Once anemia is confirmed by an absolute
decrease in the RBC count, an etiologic
diagnosis is needed to guide therapy and
prognosis.
Although history, clinical signs, or physical
findings may be sufficient for accurate etiologic
diagnosis, additional testing is often needed
One classic pathophysiologic scheme for
categorizing anemia depends on determination
of whether regeneration is evident.
A regenerative anemia is one in which the bone
marrow responds by increasing RBC production
in the bone marrow with subsequent release into
the circulation.
Blood loss anemia and hemolysis are types of
regenerative anemias.
A feature unique to the horse is the fact that their
erythrocytes remain in the bone marrow until
fully mature, even in the face of severe anemia
and intense erythropoiesis.
Therefore peripheral evidence of erythrocyte
regeneration typically found in other species-
such as reticulocytosis, macrocytosis,
anisocytosis, polychromasia, Howell-Jolly
bodies, basophilic stippling, and nucleated
RBCs is rarely found in the horse.
Because of the lack of obvious evidence of
regeneration in the peripheral blood of the
horse, the RBC indices are not highly useful
for classification of anemia.
However, a mild to moderate increase in the
MCV and anisocytosis are the most common
hints of regeneration in the horse.
These latter changes are more likely to occur
with hemolytic anemia rather than with blood
loss and usually do not occur until 2 weeks
after the onset of anemia.
An increased MCH or MCHC indicates the
presence of free hemoglobin subsequent to
either in vitro or in vivo hemolysis.
Microcytic, hypochromic, and macrocytic states
are seldom observed in the horse, although
moderate to severe iron deficiency may cause
microcytic, hypochromic anemia.
The RBC volume distribution width, an indicator
of variation of the RBC volume that may be
provided by some automated blood cell
counters, increases with regeneration.
Increases in RBC creatine and glucose-6-
phosphate dehydrogenase concentrations may
correlate with increased MCV and immature
RBCs in the circulation in horses.
The most reliable measure of the bone
marrow's responsiveness to anemia in horses
requires evaluation of a bone marrow aspirate.
 The diagnosis of blood loss or hemolytic anemia
typically can be made without bone marrow
evaluation.

Bone Marrow Aspirate

In horses, bone marrow may be collected from
the sternum, rib, or ileum; however, the sternal
site is generally preferred.
The sternal skin on the ventral midline between
or slightly behind the olecranons is clipped and
aseptically prepared.
A subcutaneous block with 2% lidocaine and/or
sedation may facilitate patient compliance.
A stab incision through the skin may be
necessary when using large biopsy needles.
A 3.5 inch 18G spinal needle with stylet in place
or a core biopsy needle may be used.
Use of a spinal needle seems to be well
tolerated and is quicker.
The needle is briskly inserted through the skin-
perpendicular to the flat sternal plate, on the
midline in the cleavage between the pectoral
muscle and advanced until the tip reaches bone.
At this point, the needle should be rotated back
and forth until it is firmly embedded into bone.
The stylet then is removed and a 20-ml syringe
attached.
The syringe plunger should be pulled gently to
the 10-ml mark and released while the needle is
stabilized in the bone.
This maneuver should be repeated two to three
times.
If marrow or blood is seen in the hub of the
needle, suction should be released immediately
to prevent blood from diluting the marrow
aspirate.
If marrow or blood is not obtained, reintroduce
the stylet, advance the needle, and repeat the
aspiration.
Smears should be made immediately after
collection and air-dried.
Aspiration of bone marrow from the sternum is
generally easy, quick, well tolerated, and without
complication.
Rare reports have documented the needle
slipping into the thoracic cavity and nicking the
heart.
Proper patient restraint and/or ultrasound-guided
aspirates should reduce this risk.
Bone marrow core biopsies can be collected
with an infant marrow biopsy needle with the
same procedure as is used for collecting
aspirates.
Once the needle is inserted, the stylet is
removed, and the needle is rotated
unidirectionally.
No aspiration is applied.
The core biopsy is removed from the needle by
inserting the stylet into the beveled end of the
needle to push the core out the hub.
The core is then should be inserted into 10%
formalin fixative for histopathology.
The normal myeloid to erythroid ratio (M: E) in
horses ranges from 0.5 to 2.4.
M: E ratios of less than 0.5 and the presence of
at least 50% reticulocytes are generally
consistent with an appropriate bone marrow
response to anemia in the horse.
The maturity of the erythroid compartment
should also be assessed.
In normal horses, 70% to 90% of erythroid cells
are rubricytes and metarubricytes.
 Prussian blue stain can be used to assist in the
evaluation of bone marrow iron stores.
‫٭‬

Blood loss anemia

a) Acute blood loss

The pathophysiologic effects of acute blood loss

result from decreased circulating blood volume
and associated hypovolemic shock.
Hemorrhage becomes clinically significant when
over 30% of the body's blood is lost rapidly.
Because blood volume represents
approximately 8% of body weight, this loss
constitutes 10.8 liters in a 450-kg horse.
Unfortunately, accurate quantification of blood
loss is rarely possible.
Thus assessment of the presence and severity
of hemorrhage is based on clinical signs and
sequential measurement of PCV and plasma
protein.

Severe hemorrhage usually is due to loss of

vascular integrity, which may result from
iatrogenic (surgery) or accidental trauma or from
erosion of vessel walls by neoplastic, infectious,
or parasitic lesions.
Rarely, coagulopathies such as an inherited or
acquired factor deficiency, thrombocytopenia, or
disseminated intravascular coagulopathy cause
clinically significant acute blood loss.
Common causes of external hemorrhage
caused by large vessel rupture include
castration complication or accidental trauma.
Epistaxis associated with nasal surgery or
guttural pouch mycosis also can result in severe
blood loss.
Rarely, exercise-induced pulmonary
hemorrhage, ethmoid hematoma, or nasal
neoplasia cause serious epistaxis.
Internal hemorrhage occurs most often into large
muscle masses, the abdominal cavity, or the
thorax.
In muscle, fracture fragments or severe soft
tissue trauma can lacerate large vessels.
 Traumatic splenic rupture or spontaneous
middle uterine artery hemorrhage in postpartum
mares can cause hemoperitoneum.
Less often, mesenteric vessels may be
damaged by large strongyle migration,
mesenteric abscessation, or neoplasia.
Similar erosive processes, such as a pulmonary
abscess or neoplasia, as well as trauma, can
cause hemothorax.

Clinical Signs and Diagnosis

Consistent signs include tachycardia, tachypnea,
pale mucous membranes, and prolonged jugular
filling.
With continued bleeding, progressive exercise
intolerance, muscular weakness, and collapse
ensue.
Decreased organ perfusion can cause oliguria
and ileus.
With internal hemorrhage, signs associated with
the site of blood accumulation may be present.
These include dyspnea and pleurodynia in
cases of hemothorax, low-grade colic associated
with hemoperitoneum, or lameness due to
hemarthrosis or muscle hemorrhage.
The source of massive extracorporeal bleeding
usually is obvious.
However, internal or multifocal bleeding may
require a careful physical examination to discern
clinical signs of hemorrhage and to differentiate
them from other signs of abdominal, thoracic, or
musculoskeletal disease.
When a typically non-hemorrhagic emergency
such as colic, pleuritis, or acute musculoskeletal
trauma is encountered, internal bleeding should
always be considered if tachycardia, tachypnea,
and mucous membrane blanching persist after
the patient is stabilized.
Auscultation, percussion, centesis, and
percutaneous ultrasonography are useful
ancillary diagnostic aids.
If no source of vascular damage to account for
the hemorrhage can be identified, then the
diagnostic work-up should include a platelet
count and clotting profile that includes
prothrombin time, activated partial
thromboplastin time, and fibrin degradation
products.
In the first 12 to 24 hours after acute
hemorrhage, the severity of blood loss must be
estimated on the basis of clinical signs, unless
the actual volume of hemorrhage has been
quantified.
This estimation is necessary because several
physiologic alterations render the PCV and
plasma protein unreliable indicators of bleeding.
First, increased sympathetic nervous system
activity triggers splenic contraction, thus
releasing stored red blood cells that temporarily
support the PVC.
Concurrently, decreased hydrostatic pressure
within capillaries causes interstitial fluid to move
into the vascular space.
This begins within 30 min of acute hemorrhage
and can restore 20% to 50% of blood volume
within 6 hours.
Subsequently, further translocation of interstitial
fluid, absorption of fluid from the gastrointestinal
tract, and renal water resorption all act to
support intravascular volume but dilute the PCV.
Thus the PCV continues to decrease until the
true severity of blood loss can be assessed in 24
to 36 hours after hemorrhage ceases.
Plasma protein is reclaimed from lymph during
fluid shifts after bleeding.
As a result, plasma protein, which is a more
useful indicator of severity of hemorrhage in the
first hours after bleeding, increases more rapidly
than PCV there after.
In cases of internal hemorrhage, the
interpretation of PCV and plasma protein is
further complicated by the recycling of up to two
thirds of the erythrocytes and most of the protein
lost into extravascular spaces within the body.
The bone marrow response to hemorrhage is
not reflected by any increase in PCV until 3 to 5
days after the hemorrhage.
Normal red blood cell mass may not be restored
for 4 to 6 weeks.
In general, the clinicopathologic indicators of
response to hemorrhage seen in other species
are not present in horses.
Leukocytosis and thrombocytosis are rare.
Occasionally, the mean corpuscular volume
(MCV) will increase above 60 fl by 4 to 7 days
after hemorrhage
Treatment

The two immediate goals of treatment for acute

blood loss are:
(1) to stop the bleeding and
(2) to reverse hypovolemic shock.
External hemorrhage is best controlled with
direct pressure or large vessel ligation.
With internal hemorrhage, physical and
environmental stress are minimized.
Surgical intervention to identify and control sites
of internal hemorrhage usually is not practical.
If clinical signs of hypovolemic shock are
present, an intravenous crystalloid solution
should be administered rapidly at 40 to 60 ml!kg
to provide cardiovascular support.
Other potential fluid choices include hypertonic
saline and colloid solutions.
The use of hypertonic saline in the face of
uncontrolled hemorrhage is controversial
because some experimental evidence
demonstrated increased blood loss and mortality
caused by the increased blood pressure and
cardiac output that follow hypertonic saline
administration.
If bleeding has been stopped, 7% saline at 4
ml/kg IV (2 L to a 450-kg horse) may be
beneficial, particularly if high-volume crystalloid
solution administration is not practical in a field
setting.
Following the hypertonic saline with appropriate
isotonic fluids within two hours is important.
Although colloid solutions such as dextran,
hydroxyethyl starch, or plasma often are cost-
prohibitive or unavailable, the use of hydroxyethyl
starch solutions (Hespan) to treat hypovolemic
shock in equine practice is increasing.
When administered intravenously at 8 to 10 ml/kg,
6% hydroxyethyl starch produced a significant
positive oncotic effect for 24 hours in 11
hypoproteinemic horses.
The use of hydroxyethyl starch as a resuscitative
fluid in the face of acute blood loss should be
reserved to cases in which bleeding has been
controlled, because a dose-dependent decrease in
prothrombin time, activated partial thomboplastin
time, and fibrinogen concentrations is associated
with its administration.
After stabilization, the necessity for whole blood
transfusion must be assessed.
The decision for transfusion is based on
the overall clinical assessment of the
patient.
b) Chronic blood loss

Chronic blood loss is uncommon in the horse

But, when present, is most often secondary to
gastrointestinal parasite infestation.
Other sources of gastrointestinal blood loss
include infiltrative diseases such as gastric
squamous cell carcinoma or ulcers secondary to
nonsteroidal antiinflammatory drug toxicity.
Urogenital bleeding secondary to neoplasia or
infection is possible, and idiopathic urethral
hemorrhage in male horses have been reported.
Thrombocytopenia or coagulopathy should
always be considered when chronic blood loss is
suspected.
Clinical Signs and Diagnosis
Overt clinical signs of anemia are not associated
with gradual loss of red cell mass until the PCV
falls below 12%.
Subtle signs such as lethargy and pale mucous
membranes may be noted earlier, as may
clinical signs related to the underlying disease-
such as weight loss in the case of parasitism or
neoplasia.
The primary rule-outs for anemia due to chronic
blood loss are anemia of chronic disease and
low-grade hemolysis.
Diagnostic work-up is aimed at characterizing
the anemia, identifying a source of blood loss,
and ruling out other causes of anemia.
Therefore complete blood count, bone marrow
evaluation, fecal occult blood, fecal examination
for parasite ova, urinalysis, total and indirect
serum bilirubin, and a Coggins test for equine
infectious anemia all are indicated.
Chronic blood loss should result in a
regenerative anemia; however, peripheral signs
of erythrocyte regeneration are inconsistent in
the horse and at best include an increase in
mean corpuscular volume and mild anisocytosis.
Bone marrow evaluation with a myeloid/erythroid
ratio of less than 0.5 indicates a regenerative
response.
 Depletion of iron stores by chronic bleeding can
be confirmed by the presence of hypoferremia,
increased total iron binding capacity, decreased
marrow iron, and a poor regenerative marrow
response.
The associated non-regenerative anemia
typically is microcyctic, hypochromic.

Treatment

Treatment is based on identification and

elimination of the source of blood loss.
Because patients with chronic anemia have
often had sufficient time to adjust physiologically,
transfusion is rarely needed.
If evidence of decreased iron stores exists,
ferrous sulfate can be supplemented orally at 2
mg/kg daily.
Administration of parenteral iron is not
recommended because of the possibility of
serious adverse reactions.
Anemia Secondary
to Inadequate Erythropoiesis
The average lifespan of the equine erythrocyte
is 150 days.
Therefore anemia caused by inadequate
erythropoiesis is an insidious process
associated with scant, nonspecific clinical signs.
Causes include anemia of chronic disease,
nutritional deficiency, bone marrow aplasia, or
myelophthisic disease.
Of these, anemia of chronic disease (ACD)
associated with infectious, inflammatory, or
neoplastic disorders is most commonly
encountered in horses.
Anemia of chronic disease (ACD)

Three mechanisms have been incriminated in

the disease process:
– shortened erythrocyte life span,
– insufficient bone marrow response to demand for red
blood cells, and
– decreased release of iron from the reticuloendothelial
system.
Accelerated red cell destruction may be the
result of activation of the mononuclear
phagocyte system in response to inflammation.
Also, the intravascular response to inflammation
may cause increased erythrocyte damage
during passage through small vessels with
subsequent removal by the reticuloendothelial
system.
Normally, the bone marrow would respond to
such increased consumption with an appropriate
increase in red blood cell production.
The rate-limiting factor in the marrow's failure to
respond to anemia of chronic disease may be
inadequate erythropoietin production.
Erythropoietin is the hormone primarily
responsible for regulation of erythropoiesis in the
bone marrow.
Although erythropoietin production is increased
in people and in animal models of anemia of
chronic disease, the hormone is deficient
relative to the anemia.
Administration of pharmacologic doses of
recombinant erythropoietin will reverse ACD.
In addition to the relative erythropoietin
deficiency, erythroid progenitor cell activity is
inhibited in ACD, as is iron release from the
reticuloendothelial system.
It appears likely that the abnormal bone marrow
and iron metabolism responses in ACD are
mediated by cytokines produced in response to
inflammatory conditions, including infection and
neoplasia.
Interleukin-l, tumor necrosis factor, and
interferon all have been shown experimentally to
play roles in ACD.
Diagnosis, Clinical Signs, and Treatment
ACD is diagnosed by the presence of a chronic
disease process accompanied by a mild to
moderate, nonregenerative, normochromic, and
normocytic anemia.
Serum iron and total iron-binding capacity are
decreased, but normal to increased iron stores
can be demonstrated by normal serum ferritin
concentration or positive Prussian blue stain for
marrow iron.
The clinical signs of anemia of chronic disease
are those related to the primary disease, and
treatment is solely related to eliminating the
underlying disease condition and to ensuring
that the anemia is not caused by blood loss or
hemolysis.
Diseases associated with anemia of chronic
disease in horses include:
– pleuropneumonia,
– internal abscessation,
– peritonitis,
– chronic organ failure,
– immune-mediated or granulomatous diseases,
– neoplasia, and
– chronic viral disease such as EIA.
Although hypoferremic, horses with ACD have
normal iron stores and do not require iron
supplementation.
Nutritional deficiency

Inadequate erythropoiesis caused by dietary

inadequacy is rare in horses.
Vitamin B12 and folic acid are important
cofactors in erythrocyte maturation.
In horses, gastrointestinal bacteria synthesize
vitamin B12 thus eliminating the need for
exogenous consumption.
Deficiencies in other micronutrients involved in
erythrocyte production-such as copper, cobalt,
and iron-are rare.
Iron-deficiency anemia in horses is almost
invariably the result of chronic blood loss.
Iron-deficiency anemia is diagnosed on the
basis of hypoferritinemia, hypoferremia, normal
to increased total iron binding capacity, and
detection of decreased iron stores on
examination of bone marrow aspirate.
In horses, normal serum ferritin has been
reported as 152±54.6 ng/ml, normal iron
concentration as 120±5.0 fLg/dl, and normal
total iron binding capacity as 388±8.1 fLg/dl.

Laboratory values improve after oral iron

supplementation.
Anemia due to bone marrow aplasia

Aplastic anemia results from congenital or

acquired developmental failure of hematopoietic
progenitor cells in the bone marrow.
In other species, acquired aplastic anemia has
been associated with bacterial and viral
infections, chronic renal or hepatic failure,
irradiation therapy, and drug administration, but
the majority of cases are considered idiopathic.
Aplastic anemia had only been reported in a few
horses.
No definitive cause is identified, although the
following are implicated:
– An immune-mediated process.
– Phenylbutazone use.
– Unapproved administration of a human
erythropoietin to racehorses resulted in a
drug-induced immune-mediated anemia
characterized by potentially fatal bone marrow
erythroid suppression.
Clinical Signs, Diagnosis, and Treatment
Horses with advanced anemia caused by bone
marrow hypoplasia show nonspecific clinical
signs such as poor performance and weight loss
as well as pale mucous membranes.
Definitive diagnosis of aplastic anemia is based
on bone marrow assessment.
In horses with anemia after administration of
recombinant human erythropoietin, bone marrow
myeloid/erythroid ratios were 6.7 and 3.2
(normal 0.5-1.5), thus indicating severe,
nonregenerative anemia.
These horses also have increased serum iron
concentrations, normal total iron binding
capacities, and increased serum ferritin
concentrations.
Recombinant human erythropoietin can be
detected in the plasma of horses for only 72
hours after dosing.
Use of recombinant human erythropoietin can
be suspected in horses if anti-EPO antibodies
are detected in the patient's serum or if
endogenous erythropoietin levels are abnormally
low.
Treatment of aplastic anemia is focused on
identification of underlying cause and on
corticosteroid administration.
Steroids stimulate erythropoiesis by increasing
erythropoietin production and the sensitivity of
stem cells to this hormone's action.
No successful treatment currently is available for
myelophthisic diseases associated with
pancytopenia.
Hemolytic Anemia
This is a pathologic condition that results from
accelerated erythrocyte removal and can be
intravascular and extravascular.
Clinical signs associated with intravascular
hemolysis are typically acute in onset and
classically include icterus and hemoglobinuria.
Extravascular hemolysis results from
accelerated erythrocyte removal by
macrophages in the spleen or liver and is
characterized by icterus without hemoglobinuria.
Diseases that primarily cause intravascular
hemolysis
1) Oxidative Erythrocyte Damage
This can develop following exposure to a variety of
oxidizing agents such as phenothiazines, onions, or
wilted red maple (Acer rubrum) leaves.
The latter cause is by far the most common.
The oxidizing agent causes hemoglobin to become
denatured with subsequent disulfide bond
formation.
Oxidized hemoglobin forms precipitates referred to
as Heinz bodies that are visible with
Romanowsky's-stained blood smears.
 These cell changes result in increased fragility of
cells with subsequent intravascular hemolysis
and enhanced removal by the mononuclear
phagocytic system in the spleen and liver.
In addition, oxidative damage causes increased
permeability of the membrane, thereby altering
ion transport mechanisms and osmotic
gradients.

Clinical Signs and Diagnosis

Clinical signs result from the combined effects of
tissue hypoxia and hemolysis that results in
fever, tachycardia, tachypnea, lethargy, intense
icterus, and hemoglobinuria, with characteristic
brown coloration of skin, mucous membranes.
Hematologic abnormalities include anemia,
increased MCH concentration and mean
corpuscular hemoglobin, free plasma
hemoglobin, anisocytosis, poikilocytosis,
eccentrocytes, lysed erythrocytes that produce
fragments or membrane ghosts, agglutination,
increased RBC fragility, variable presence of
Heinz bodies, and neutrophilia.
Serum chemistry abnormalities include
increased total and indirect bilirubin, serum
creatinine, and serum urea nitrogen
concentrations, reduced RBC glutathione and
increased aspartate aminotransferase, sorbitol
dehydrogenase, creatinine phosphokinase, and
gamma-glutamyl trans peptidase activities.
Additional abnormalities may include
hypercalcemia and hyperglycemia, with a
variable degree of metabolic acidosis.
Urinalysis findings could include any combination
of the following: hemoglobinuria,
methemoglobinuria, proteinuria, bilirubinuria, and
urobilinogenuria.
Methemoglobin can be quantified
spectrophotometrically.
Red maple leaf, wild onion, or phenothiazine
toxicosis would be strongly suspected if a history
of exposure or opportunity for exposure exists.
Diagnosis is based on clinical signs of an acute
onset primarily of intravascular hemolytic crisis
supported by laboratory evidence of oxidative
damage that is, Heinz bodies or
methemoglobinemia.
Treatment
Treatment for oxidative injury involves reducing
the fragility of erythrocytes, maximizing tissue
oxygenation, maintaining renal perfusion, and
providing supportive care.
The horse needs to be removed from the
environmental source of the toxin and treated
with activated charcoal (8-24 mg/kg up to 2.2 kg
PO) via nasogastric tube to reduce further
absorption of red maple toxin.
Dexamethasone (0.05-0.1 mg/kg IV q12-24h)
may be helpful to stabilize cellular membranes
and reduce extravascular removal of
erythrocytes by phagocytes.
Ascorbic acid (10-20 g PO q24h) is often used
as a scavenger of free radicals.
Prognosis
Complications are a major component of morbidity
following severe hemolysis.
Poor tissue oxygenation may lead to cerebral
anoxia and altered mentation, renal failure, and
myocarditis.
Laminitis is always a concern in horses with
severe illness and may be compounded by the
use of corticosteroids.
DIC may develop secondary to severe hemolysis.
In conclusion, horses should not be housed in
areas with access to red maple trees or other
potential oxidants.
Good quality forage should be available at all
times to reduce the likelihood of horses ingesting
leaves as they fall or blow into pastures.
Diseases that primarily cause extravascular
hemolysis
Neonatal isoerythrolysis
The primary differential diagnosis for a neonatal
foal observed to have evidence of anemia and
intense icterus during the first week of life is
neonatal isoerythrolysis (NI).
Incompatible blood group antigens between the
mare and stallion combined with maternal
exposure to fetal erythrocytes during gestation
results in maternal production of antigen specific
antibodies targeted against the foal's red blood
cells, if the foal inherited the sire's red blood cell
phenotype.
The most common blood types associated with
NI in foals are Aa and Qa.
Foals with NI most commonly suffer from
extravascular hemolysis, although rare
intravascular hemolysis occurs concurrently.
A positive direct Coombs' test provides a
presumptive diagnosis of NI.
However, the most sensitive diagnostic
technique for identification of surface-associated
immunoglobulin (Ig) molecules on suspect foal
red blood cells involves a new direct
immunofluorescence (DIF) assay.
In this assay, isotype specific antibodies bind to
red blood cells that contain surface-associated
antibodies, thus providing a quantitative
measure of erythrocytes in the circulation that
are bound with antibody.
Immune-Mediated Hemolytic Anemia
This results from cross-reacting antibodies that
induce enhanced red blood cell removal.
Autoimmune (primary IMHA) hemolysis results
from loss of self-tolerance and is relatively rare
in horses.
Most commonly, hemolysis results from
adherence of cross-reacting antibodies to
erythrocyte surface antigens (secondary IMHA).
The presence of these molecules on red blood
cells causes intravascular destruction by
complement activation (IgM-mediated) or most
commonly extravascular removal by
macrophages.
It is important to consider that any infectious
agent especially equine infectious anemia,
Babesia organisms, and Anaplasma
phagocytophila; exogenous substances such as
penicillin and phenylbutazone; or neoplasia may
cause alterations in epitopes of the erythrocyte
membrane or neoantigens that contribute to
enhanced removal by immune mechanisms.
Therefore identification of the inciting cause is
important for complete resolution of the
hemolytic crisis.
Several possibilities may explain the onset of
cellular destruction.
The basic mechanisms involve a change in the
red blood cell or an alteration in immunologic
control of self-recognition.
E.g. a change in the red blood cell membrane
may form an antigen that evokes an immune
response.
Drugs, neoplasia, or infection may induce
changes in red cell antigens.
Infectious agents that express similar antigens
as host red blood cell antigens result in
pathogen-induced immune-mediated hemolysis,
termed molecular mimicry.
Genetic predispositions may cause a failure of
self-tolerance.
Failure of autoregulation has been suggested to
result from reduced suppressor lymphocyte
control.
 Finally, failure of appropriate erythropoiesis may
result from precursor erythrocytes being
targeted by the immune response in a manner
similar to that of circulating red blood cells.
The goal of the clinician should be to focus on
identifying any potential inciting causes because
this will allow for appropriate case management
with the best prognosis for efficient and
complete disease resolution.

Clinical Signs and Diagnosis

Horses with IMHA most commonly present with
signs of extravascular hemolysis.
Spherocytes may be present on cytology of
peripheral blood smears.
Diagnosis based on autoagglutination will
suggest surface-bound antibody.
Dilution of the sample with saline (1: 1) will
indicate whether true agglutination is present.
If erythrocytes still agglutinate after dilution, they
can be considered positive for surface-bound
antibody molecules. Tests that are used for the
diagnosis when autoagglutination is absent are
the direct and indirect Coombs' test; the direct
test is more sensitive.
 The direct Coombs' test can be used but may
yield a false negative result if an incomplete set
of reagents is used, if blood is not tested at both
4° C and 37" C, or if severe hemolysis has
resulted in removal of the majority of antibody-
coated RBCs from circulation.
A new direct immunofluorescence assay
therefore has an increased sensitivity to detect
red cell antibodies for the diagnosis of IMHA.

Treatment
Therapy will be determined based on the level of
anemia.
In severe cases, whole blood transfusion may be
indicated.
Current drug administration should be
discontinued.
If, based on confirmed sepsis, antimicrobial
therapy is required, drug therapy should be
continued with a molecularly dissimilar drug.
After blood samples for diagnostic tests (Le.,
Coombs' test or direct immunofluorescence
assay) are obtained, immunosuppressive
therapy may be considered.
Most affected horses require
immunosuppression with corticosteroids.
Because immunosuppression carries the risk of
potentiating infectious agents, underlying
infectious disease conditions such as equine
infectious anemia should be ruled out.
Glucocorticoid therapy benefits the patient in the
short term by reducing the function of
macrophages to recognize antibodies
complexed to red blood cells and in the long
term by altering antibody production by B-
lymphocytes
Dexamethasone used at 0.05 to 0.2 mg/kg IV
q24h has the greatest efficacy in treating IMHA
in horses.
The PCV should be monitored carefully during
the course of steroid therapy, and if the patient
does not respond quickly, the frequency of
administration may be increased to twice daily.
In some instances, it may take up to a week for
the full effect of steroid therapy to be reflected by
a rise in PCV.
Once the PCV is stable at greater than 20%, the
steroid therapy should be carefully tapered by
0.01 mg/kg/day, while the horse is closely
monitored for recurrence of hemolytic crisis.
The major adverse reactions to long-term
administration of corticosteroid in horses are
laminitis, tendon laxity or weakness, and
immunosuppression that leads to secondary
infections.
Therefore the goal is to reduce to the lowest
effective dose as soon as possible.
Alternate day therapy should be administered for
the last week of therapy.
Some individuals may require therapy for
several weeks until disease resolution occurs.
Equine Infectious Anemia
EIA is a retroviral disease characterized by
chronic episodic pyrexia and hemolytic anemia.
The acute form of the disease has a course of
fever, depression, and thrombocytopenia that is
clinically represented by mucosal petechiation.
Horses infected for more than 30 days have the
subacute to chronic form of the disease and
show anorexia, ventral edema, and episodic
fever spikes.
Occasionally, horses may have abdominal pain,
ataxia, abortion, or infertility induced by
inflammation of the affected organ system.
Although not common, deaths may be observed
during the subacute to chronic phases of
disease.
Many horses recover from the initial phases of
the disease, but episodic flare-ups will continue
throughout the first year of infection.
Persistence of virus in the face of detection of an
antibody response indicates reduced immune
clearance.
Therefore it is logical to assume that immune
suppression can occur during times of
environmental stress and thus can lead to
recrudescence of disease.
In some individuals, clinical signs of disease are
not apparent, and diagnosis is incidentally made
on annual serologic testing.
This population of horses represents a carrier
state and is a significant threat to uninfected
horses.
Hematologic evidence of disease includes
thrombocytopenia during the acute phase of
disease with anemia and icterus developing
during the subacute to chronic phases of
disease.
Additional changes may include leukopenia with
lymphocytosis and monocytosis.
During episodes of hemolysis, increased serum
antibody-coated erythrocytes may be detected.
Hyperglobulinemia, increased serum liver
enzyme activities, and proteinuria may develop
during the chronic stages of disease.
Pathogenesis
The virus is tropic for macrophages.
After viral replication the viral particles are released
from the macrophage into the surrounding
extracellular milieu, thus introducing a cycle of viral
amplification within the host.
Serologic detection of disease is best performed
after 16 to 42 days of infection.
Hemolysis is considered to result from immune
complex attachment to RBCs via a viral
hemagglutinin, with subsequent extravascular
removal of cells by the mononuclear phagocytic
system.
Additional mechanisms for virusinduced anemia
involve reduced bone marrow erythrocyte
production through induction of suppressive
cytokines.
 Episodic flare-ups of EIA are believed to result
from antigenic drift of the virus with stimulation of
cycles of immune responses.

Diagnosis, Management, and Prevention

The preferred diagnostic test for EIA is the
Coggins test, which is an agar gel
immunodiffusion assay.
An alternative test is the enzyme-linked
immunosorbent assay (ELISA).
The Coggins test has the advantage of good
sensitivity for chronically infected horses,
whereas the ELISA has fast turn-around time but
lower specificity.
Early in the course of disease the humoral
response may be low, and false-negative test
results can occur with the Coggins test.
A small percentage of chronically infected
horses may test negative.
Alternatively, passive transfer of colostral
antibodies in foals may result in false-positive
results.
Repeat testing in both cases will indicate the
true incidence of disease in the host.
These examples support the suggestion made
by many authors that repeat evaluation of EIA
status is recommended at a minimum of a 30-
day interval before introduction into a herd.
Testing is required for national or international
transport, either with the Coggins test or the
ELISA.
No treatment is available for viral clearance in
positively infected horses.
These animals remain a constant threat for viral
infection to other horses; euthanasia is generally
recommended.
Chronically infected horses are in most cases in
poor condition and unable to remain athletic.
However, some horses remain subclinically
affected and, although they pose a potential
threat to other horses, they are able to maintain
a good quality of life.
In such cases or when valuable breeding
animals are maintained, veterinarians must
adhere to specific precautions.
Absolute attention to restriction of blood transfer
or blood-related material is crucial.
Additional control measures include isolation of
no less than 200 yards from any other horses
and double screening in a stable environment.
These precautions must be approved by the
governing agencies of the specific location;
different states have specific laws regarding
these specifications.
Piroplasmosis
Piroplasmosis is a tick borne disease of horses
resulting from infection with one of two
protozoan hemoparasites of the Apicomplexa
subphylum Babesia caballi or Theileria equi
(formerly Babesia equi).
These are the only recognized intraerythrocytic
diseases of horses.
Natural transmission of disease is a result of tick
infestation.
Piroplasmosis is most commonly reported to
occur in tropical or subtropical regions, although
it is reported to occur rarely in temperate
regions, which reflects the habitat of the natural
tick vectors.
Babesia caballi is transmitted transovarially from
one tick generation to the next.
 Theileria equi is transmitted horizontally by species of
Dermacentor, Hyalomma, and Rhipicephalus, which are
rarely vectors for B. caballi.
The primary vector is Dermacentor nitens.
All horses can be infected with these parasites, but
disease tends to be more severe in elderly horses.
The history of infected horses often includes a recent
episode of transport to an endemic region.

Clinical Signs and Pathogenesis

Clinical signs are apparent in the first 1 to 4 weeks of
infection and include fever depression, anorexia,
weakness, ataxia, lacrimation, mucoid nasal discharge,
chemosis, icterus, hemoglobinuria, and potentially death.
Fatalities may occur within 48 hours of infection,
but chronic disease often develops.
Inapparent carrier horses are common found in
endemic regions.
Hematology may reveal parasitized RBCs early
in the course of disease.
Both B. caballi and T. equi species have been
observed in the Americas, Europe, Asia, Africa,
Middle East, and Russia.
Babesia and Theileria organisms infect and
multiply within red blood cells.
The majority of these infected cells are removed
by the mononuclear phagocytic system, but
some severely affected animals will suffer from
intravascular hemolysis.
Clinical disease induced by T. equi is more
severe than that induced by B. caballi and has a
higher mortality rate.
Untreated horses will remain persistently
infected for an undetermined period of time.
Factors such as transport, inclement weather,
pregnancy, or concomitant disease will have the
potential for induction of clinical disease in
carrier animals.
Diagnosis and Treatment
Diagnosis may be based on observation of
parasitized erythrocytes with Giemsa stained blood
smears, isolation of the organism from the blood, or
positive serology.
Parasitemia precedes hemolysis so diagnosis is
most commonly based on serology.
Babesia- or Theileria-specific antibodies will be
produced approximately 14 days after infection
and can be determined with the CFT or IFAT.
Animals imported into negative regions must
have a negative CFT.
A PCR test that has greater sensitivity for
detection of positive horses has also been
developed.
Treatment is most successful with administration of
imidocarb diproprionate to reduce the parasite
load.
Clearance of B. caballi is achieved with two doses of
2.2 mg/kg intramuscularly, administered 24 hours
apart, whereas T. equi is more difficult to clear and
requires four to six doses of 4 mg/kg
intramuscularly, administered 72 hours apart.
In some cases, B. caballi will spontaneously resolve.
Because of the anticholinesterase properties of the
drug, patients should be carefully monitored for
colic, hypersalivation, diarrhea, or potentially, death.
Donkeys have an increased sensitivity to imidocarb
and should be treated at the lower dose (2.2 mg/kg
1M q24h x 2 doses) and receive close monitoring of
clinical signs.
Equine Ehrlichiosis
This is caused by the rickettsial organism
Ehrlichia equi belonging to the phagocytophilia
complex of ehrlichial agents.
Equine ehrlichiosis is most common in the
United States.

Clinical Signs and Diagnosis

Clinical signs include fever, depression,
petechiae, ventral edema, and ataxia, with
reluctance to move.
Differential diagnoses for these signs include
purpura hemorrhagica, EVA, and encephalitis.
Granulocytopenia, anemia, and
thrombocytopenia are common abnormalities
detected on hematology.
Horses that are younger than 3 years of age
exhibit a less severe form of the disease.
Diagnosis of E. equi may be made from
identification of morula in circulating
granulocytes.
Seroconversion (fourfold rise in serum
antibodies in a convalescent sample) can be
detected with an indirect fluorescent antibody
test.
A PCR assay has been developed for E. equi
and has improved sensitivity and specificity in
comparison to conventional diagnostic tests.
Supportive care for affected patients should
include NSAID therapy, intravenous fluids, and
lower limb sweat wraps.
Intravenous oxytetracycline (7 mg/kg diluted in 1
liter of saline IV q24h x 5-7 days) is effective for
clearance of equine granulocytic ehrlichiosis.
Response to therapy is extremely rapid, thus
supporting the diagnosis in suspect cases.
Carrier phases have not been recognized; in
fact, once-infected horses acquire strong
immunity for up to 2 years, thus providing
resistance to re-infection.
*****