Lecture 6: Erythrocytes- shapes inclusions/infective agents

Define anisocytosis.

Variation in erythrocyte size; often quantified by red cell distribution width (RDW).

Define poikilocytosis.

General term for the presence of abnormally shaped erythrocytes. Normal in goats and young healthy cattle.

Define macrocytosis.

Presence of abnormally large erythrocytes; often associated with regenerative responses (e.g., reticulocytosis) or certain pathologies.

Define microcytosis.

Presence of abnormally small erythrocytes; commonly associated with iron deficiency anemia, portosystemic shunts, or chronic disease.

Define polychromasia.

Presence of erythrocytes with bluish cytoplasm on routine stains, indicating RNA content; reflects circulating reticulocytes and regenerative response.

Define reticulocyte.

Immature erythrocyte containing ribosomal RNA; best visualized with supravital stains; indicates bone marrow regeneration.

What does the term "echinocyte" mean and what is its etymology?

Derived from echinos (Greek: spiney) and crena (Latin: notch). Refers to spiculated erythrocytes, previously called "crenated cells."

What are causes of echinocytes?

- Artifacts (common in pigs, thick blood films)

- Snake bites (coral, rattlesnake, water moccasin)

- Uremia, post-transfusion, pyruvate kinase (PK) deficiency in dogs

- Glomerulonephritis, neoplasia, doxorubicin in dogs

- Total body cation depletion in horses

- Fatty acids, bile salts, drugs, lysophospholipids

- RBC dehydration, ATP depletion, ↑pH, ↑RBC Ca²⁺

What are causes of acanthocytes?

- Altered cholesterol:phospholipid ratio in membrane

- Liver disease

- Hemangiosarcoma, glomerulonephritis, DIC in dogs

- Physiologic in young goats and some young cattle

What are stomatocytes and their significance?

Erythrocytes with a mouth-shaped central pallor.

- Often artifacts in thick blood films

- Can indicate hereditary stomatocytosis: swollen RBCs with ↑MCV and ↓MCHC

- Can result from drugs that bind preferentially to the inner lipid bilayer

Describe the process of spherocyte formation in immune-mediated anemias.

1. Macrophages have receptors for Ig and complement.

2. Antibody/complement-coated RBCs bind these receptors.

3. Entire RBCs may be phagocytized OR macrophages may remove portions of membrane.

4. RBCs reseal but lose biconcave shape → form spherocytes.

5. Despite smaller diameters, volumes are not markedly reduced.

6. MCV may be high due to concurrent reticulocytosis.

What conditions are associated with spherocytes?

- Immune-mediated hemolytic anemia (IMHA; especially dogs)

- Snake bites, bee stingsZinc toxicity

- Erythrocyte parasites

- Transfusion of stored blood

- Dyserythropoiesis in English Springer Spaniels

- RBC band 3 deficiency in Japanese Black cattle

Which erythrocyte changes indicate fragmentation?

Schistocytes (best evidence), keratocytes, acanthocytes.

Do spherocyte-like cells always indicate IMHA?

No. A few spherocyte-like cells can appear with fragmentation and do not indicate immune-mediated disease in this context.

What hematologic findings often accompany erythrocyte fragmentation due to DIC?

Thrombocytopenia, coagulation disorders, and mild to moderate anemia.

What are mechanical causes of erythrocyte fragmentation?

- Disseminated intravascular coagulation (DIC)

- Caval syndrome (heartworm disease)

- Glomerulonephritis

- Cardiac valvular stenosis

What are causes of erythrocyte fragmentation due to endothelial injury?

- Hemangiosarcoma (especially dogs)

- Vasculitis

- Splenic disease

- Hepatic disease

What are causes of erythrocyte fragmentation due to thermal injury?

Heat stroke and severe burns.

Besides mechanical, endothelial, or thermal causes, what other conditions can cause fragmented erythrocytes?

- Severe iron deficiency

- Myelofibrosis

- Hemophagocytic histiocytic disorders

- Congenital or acquired dyserythropoiesis in dogs

- Pyruvate kinase deficiency (post-splenectomy in dogs)

What are eccentrocytes, and in which species are they clinically significant?

RBCs with eccentrically placed hemoglobin, leaving a pale crescent-shaped area; significant in dogs.

What are causes of eccentrocytes in dogs?

Endogenous oxidants: diabetic ketoacidosis, inflammation, neoplasia (≈50% lymphoma), idiopathic causes

Exogenous oxidants: onions, garlic, NSAIDs, propofol, vitamin K

How severe is anemia caused by eccentrocytes?

Ranges from mild (endogenous oxidants) to mild-severe (exogenous oxidants).

What type of injury produces eccentrocytes?

Membrane oxidant injury.

What are causes of eccentrocytes in dogs?

Acetaminophen, onions, garlic, vitamin K, propofol, vitamin K antagonist rodenticides.

What are causes of eccentrocytes in horses?

Red maple (Acer rubrum) toxicity and enzyme deficiencies (G6PD and glutathione reductase).

What is a bovine cause of eccentrocytes?

IV administration of hydrogen peroxide.

Which conditions are associated with multiple erythrocyte shape abnormalities?

- Liver disorders

- Hemangiosarcoma (dogs)

- Glomerulonephritis

- Myeloid neoplasms

- DIC (esp. dogs)

- Chronic iron deficiency anemia (dogs, ruminants)

- Doxorubicin toxicity (dogs & cats)

- Myelofibrosis

- Congenital dyserythropoiesis

Which nucleated erythrocyte stages are typically seen in blood smears?

Rubricytes and metarubricytes.

Are nucleated erythrocytes ever normal?

Occasionally present in low numbers in healthy dogs and cats.

What are physiologic/pathologic causes of nucleated erythrocytes in regenerative anemias?

Marrow response to anemia (reticulocytosis + rubricytosis).

What toxic or environmental conditions can cause nucleated erythrocytes?

Lead toxicity and heat stroke.

What conditions of marrow injury/disease can cause nucleated erythrocytes?

- Septicemia

- Endotoxic shock

- Drugs

- Heat stroke

- Myelodysplasia

- Hematopoietic neoplasms

- Infiltrative disorders

What conditions related to splenic dysfunction cause nucleated erythrocytes?

Splenectomy or impaired splenic filtration.

What systemic disorders can lead to nucleated erythrocytes in dogs?

Cardiovascular disease, inflammation, trauma, hyperadrenocorticism.

What systemic disorders can lead to nucleated erythrocytes in cats?

Hepatic lipidosis, acute trauma, inflammation.

Which hereditary conditions may present with nucleated erythrocytes?

Hereditary dyserythropoiesis in dogs and cattle.

What are Howell-Jolly bodies?

Micronuclei (nuclear remnants) seen within erythrocytes.

In which species can low numbers of Howell-Jolly bodies be normal?

Horses and cats.

What conditions cause increased Howell-Jolly bodies?

- Regenerative anemias

- Splenectomy

- Glucocorticoid therapy

- Vincristine therapy (in regenerative anemia, due to nuclear fragmentation)

What are Heinz bodies?

Denatured hemoglobin precipitates attached to the inner RBC membrane.

In cats, what percentage of Heinz bodies is considered normal?

Up to 5% of erythrocytes.

What diseases in cats are associated with increased Heinz bodies?

Diabetes mellitus, lymphoma, hyperthyroidism.

What are causes of Heinz bodies in dogs?

- Splenectomy

- Zinc toxicity

- Acetaminophen, methylene blue, methionine, phenazopyridine, menadione (vitamin K3)

- Naphthalene

- Skunk musk exposure

What are iatrogenic or dietary causes of Heinz bodies in cats?

- Allium species ingestion (onion, garlic, leek, chive)

- Propylene glycol in soft-moist foods

- Propofol anesthesia

- Hydrogen peroxide as an emetic

What are causes of Heinz bodies in livestock and horses?

- Onions (wild and domestic)

- Kale and other Brassica species (ruminants)

- Red maple leaves (horses, alpacas)

- Copper toxicity (sheep, goats)

- Lush winter rye (Florida cattle)

- Selenium deficiency (St. Augustine grass cattle)

- Post-parturient cattle on perennial ryegrass (New Zealand)

- Phenothiazine therapy (horses)

What is basophilic stippling, and what does it represent?

Diffuse aggregates of ribosomes and polyribosomes within erythrocytes.

In which situations is basophilic stippling seen?

Regenerative anemia (especially in ruminants)

Lead toxicity

What type of organisms are Babesia spp.?

Protozoal organisms (single-celled with nucleus and cytoplasm) that infect erythrocytes.

Name important Babesia species and their hosts.

B. bigemina → cattle

B. vogeli → dogs

B. gibsoni → dogs

B. felis → cats

What is the pathogenesis of Babesia spp. infection?

Protozoa invade and multiply within RBCs → intravascular hemolysis → anemia, icterus, hemoglobinuria; immune-mediated destruction may contribute.

How do Theileria spp. differ from Babesia spp.?

Theileria spp. have both a tissue phase and a blood phase, with schizogony occurring in lymphocytes before RBC infection.

Which Theileria species are important and their hosts?

T. buffeli → cattle (USA, usually non-pathogenic)

T. equi → horses (formerly classified as B. equi)

T. cervi → deer

Pathogenic species in ruminants occur in Africa, Asia, Middle East

What is the pathogenesis of Theileria spp. infection?

Schizogony in lymphocytes → lymphoproliferation and immunosuppression; erythrocytic stage contributes to anemia. Pathogenicity depends on species and host.

Which hemoparasite causes severe disease in domestic cats in the USA?

Cytauxzoon felis.

What life cycle stages of Cytauxzoon felis are important in pathogenesis?

Intermediate schizont stage in macrophages → vascular occlusion and tissue necrosis

Merozoites in RBCs → hemolytic anemia

What type of organisms are Anaplasma spp.?

Rickettsial (bacterial) organisms that divide within vacuoles inside cells.

Name important Anaplasma species and their hosts.

A. marginale → cattle

A. ovis → sheep and goats

What is the pathogenesis of Anaplasma spp. infection?

Colonization of RBC margins → extravascular hemolysis in the spleen → progressive anemia without hemoglobinuria.

What type of organisms are hemotropic Mycoplasmas?

Small bacterial organisms that attach to the outside of erythrocytes.

Name clinically important hemotropic Mycoplasma species and their hosts.

M. haemofelis → cats

M. haemocanis → dogs

M. suis → pigs

M. ovis → sheep and goats

M. haemolamae → camelids

What is the pathogenesis of hemotropic Mycoplasmas?

Adherence to RBC surface → immune-mediated clearance by macrophages → hemolytic anemia (often regenerative); severity depends on species and host immunity.

What is Bartonella spp., and how does it affect RBCs?

Bacterial organisms that parasitize erythrocytes and endothelial cells; may cause persistent bacteremia, vasculitis, and endocarditis in some species.

Which viral infection is considered a "hemoparasite mimic" due to RBC inclusions?

Canine distemper virus.

What are drepanocytes?

Fusiform or spindle-shaped erythrocytes, also called "sickle cells."

In which animals are drepanocytes considered normal?

Members of the deer family and some Angora goats.

What is the mechanism of drepanocyte formation in vitro?

Hemoglobin polymerization induced by increased oxygen tension and increased pH.

What is rouleaux formation?

Stacking of erythrocytes in linear chains resembling coins, due to increased plasma proteins reducing zeta potential between cells.

What is agglutination?

Clumping of erythrocytes due to antibody (usually IgM) binding, cross-linking RBCs.

How can rouleaux be differentiated from agglutination?

Mix blood with physiologic saline:

- Rouleaux disperses (cells separate after washing in saline).

- Agglutination persists (cells remain clumped in saline).

What is a simple method for testing rouleaux vs agglutination microscopically?

Add one part blood to 49 parts saline, place under coverslip → if adherence disappears → rouleaux; if persists → agglutination.