Clin Path 3

What is the function of neutrophils in mammals? What are some of the toxic changes that occur?

1. Phagocytic

2. Destroy microorganisms (especially bacteria)

3. Increase with inflammation

• Increased bone marrow production

• Shortened maturation + basophilic Dohle bodies + increased cytoplasmic basophilia

• Decrease in nuclear segmentation

What is the function of eosinophils w/in mammals?

1. Respond to metazoan infections (especially helminth larvae)

2. Phagocytic (less so than neutrophils)

3. Increase in allergic inflammation (mast cell + basophil degranulation especially)

4. Antigen-antibody complexes

What is the function of basophils w/in mammals?

1. Participate in allergic and delayed hypersensitivity reactions (lobed in mammals)

What is the function of monocytes w/in mammals?

1. Engulf + degrade microorganisms/abnormal cells/cell debris

2. Regulate immune responses and myelopoiesis

What are some environmental things that could cause hematologic values to become variable?

1. Collection methods

2. Difficulty in collection

3. Circadian rhythm

4. Age/sex of animal

5. Different species

What values can anesthesia affect?

1. Glucose

2. Potassium

3. RBC’s (quantity)

What are the rules for safe maximum collection volume for mammals?

1. 7.5% total blood volume is which restored w/in 48 hours

2. 1% of animal’s body weight

How does the circadian rhythm affect rodents? How does stress affect them?

1. Leukocytosis in light phase + decrease in dark phase

2. Stress = leukopenia, and chronic stress = increase in neutrophils

What animals are lymphocytes dominant in?

Rats, mice (60-80%), rabbits, hamsters, gerbils, and birds

What three factors are common w/in rats/mice?

1. Polychromasia (rbc’s have high turnover rate; 2-7% reticulocytes)

2. Howel jolly bodies

3. Neutrophils w/ ringed nuclei can be common

What is the RBC life span of a rat/mouse?

45-68 days

What are the Neutrophils of gerbils/chinchillas like?

1. Hyposegmented

What is one large characteristic unique to guinea pigs pertaining to white blood cells?

Large lymphocytes can have Kurloff bodies (large cytoplasmic inclusion); hormone influenced in young

What trends in white blood cells will you see for bacterial infections in rabbits?

Increase in absolute neutrophils, but not leukocytosis which is a normal human response

How can you tell guinea pig eosinophils apart from neutrophils (heterophils)?

Eosinophils are more lobular w/in the nucleus and have larger rod shaped granules

What is the relationship between neutrophils and lymphocytes for rabbits experiencing inflammatory conditions?

Neutrophils become more dominant compared to lymphocytes which are normally dominant in rabbits

True/false

Rabbits have lower eosinophil counts and higher basophil counts than most mammals

True

What are some causes of non-regenerative anemia in ferrets?

1. Malignant neoplasm (lymphoma)

2. Systemic infection and/or

3. Hyper-estrogen is

4. GI ulcers

What white blood cell is usually dominant in ferrets?

Neutrophils rather than lymphocytes

What is there a lack of with inflammatory disease in ferrets? Bacterial infections?

1. Lack of leukocytosis

2. Lack of lymphocytosis

What RBC morphology is common in mini pigs?

RBC speculation

What RBC morphology is normal in camelids? Why?

Ovalocytosis makes easier passage through capillaries and rapid gas exchange (O2 binds better)

What is a normal WBC response to inflammation for camelids?

1. Neutrophilia (normal)

2. Hyperfibrinogenemia (normal)

3. Non-regenerative anemia (unique)

4. Immature neutrophils

What is a normal WBC response to stress for camelids?

Neutrophilia and lymphopenia

Why is it that deer blood films will appear sickle called?

Occurs during blood film prep (exposed to O2 and affected by pH)

What does new methylene blue stain?

Reticulocytes, Heinz bodies, and howel jolly bodies

What is common during inflammation for deer?

Rouleaux (abnormal protein allows for stacking)

What is normal RBC morphology for birds?

Elliptical + nucleated + large

(Makes automatic count difficult due to nucleation)

What is the gold standard for obtaining HGB values in birds?

HGB-cyanmethemoglobin method

1. Lyse red cells to release HGB

2. Spin off free nuclei

Heme proteins are different, HGB has lower affinity for O2 and diffuses readily into tissues

What is the usual RBC life span of a bird?

28 to 45 days depending on bird

What is stippled basophilia associated with when it comes to birds? What does it signify normally?

1. RBC regeneration, response to anemia

2. Lead poisoning

What is considered a countable reticulocyte?

Distinct ring of aggregated reticulum that encircles nucleus (less than 4 specks not counted)

In response to anemia, regenerative

How do you get a reticulocyte count?

1. Count 500 red blood cells (including retics). Divide retics by 500 and multiply by 100 which will give you a percentage.

2. Count 1,000 red blood cells and proceed normally.

What are the two “halves” of an automated blood machine for a CBC?

Smaller dilution will lyse RBCs to get HGB levels and WBC differential; larger dilution will get RBC count/morphology/sizing

Descriptors for heterophils in birds

1. Most abundant, colorless cytoplasm w/ eosinophilic elongated granules

2. Nucleus = lobed (2-3)

3. Inflammatory response = increased cytoplasm basophilia, vacuoles, abnormal granulation

Descriptors for avian eosinophils

1. Round, strongly eosinophilic granules (stain intensely, lack central, refractile body, large, swollen, and round in Romanowsky stain)

2. Cytoplasm = clear blue

3. Nuclei lobed and darker

Descriptors for avian basophils

1. Smaller size

2. Deeply metachromic granules

3. Obscure nucleus

4. Cytoplasm = clear blue

5. Nuclei = non-lobed, resemble mast cells

6. Frequently found in blood

Descriptors of avian lymphocytes

1. Chromatin is heavily clumped/reticulated in mature lymphs

2. Occasionally contain azurophilic granules/irregular cytoplasmic projections (cellular degeneration, significant if large #)

3. Round cells often showing cytoplasmic irregularity around adjacent RBC in smear

4. Slightly indented, centrally or slightly eccentrically nucleus

What are the two categories of abnormal lymphs in birds?

1. Reactive lymphs - develop when antigenically stimulated; small-medium w/ heavily clumped nuclear chromatin and deeply basophilic cytoplasm

2. Blast transformed lymphs - anaplastic features, may be neoplasticism but may be caused by immunologic stimulation; large w/ dispersed, smooth nuclear chromatin

Descriptors of avian monocytes

1. Abundant, blue-gray cytoplasm, may appear opaque

2. Contain vacuoles or fine/dust-like eosinophilic granules

3. Nucleus vary in shape

What is the normal range for thrombocytes for avians? Couple fun facts?

1. 20-30k cells/mm3 or 10-15/1,000 RBCs

2. Vacuoles can occur in activated/phagocytic thrombocytes

3. Contain large amounts of serotonin (can be phagocytic)

4. Young birds = larger #

5. Thrombocytopenia = decreased marrow production or increased peripheral utilization/destruction (associated w/ severe septicemia possibly diffuse intravascular coagulation)

What is the number one thing that you would see when it comes to hemo-parasites w/in hematology?

1. Spherocytes

2. Agglutination (antibodies can cause due to excess proteins)

Differentiate between Hemobartonella and Eperythrozoon mycoplasmas. What was this category originally classified as?

1. Hemobartonella = ring forms/free floating

2. Eperythrozoon = ring forms and free floating

• Formerly classified as rickettsial, now mycoplasma 16S RNA gene

What is the most important and common strain of Hemobartonella felis? What is the second most important?

1. Mycoplasma haemofelis (Ohio strain that is most pathogenic)

2. Candidatus mycoplasma haemomunutum (California strain)

What is the morphology, spread, and clinical signs of mycoplasma haemofelis? If there is a disease name, list it.

1. Small blue rods/ring forms (attached to surface of cell, parasite can come off red cell if delay in slide prep)

2. Infected blood = blood feeding arthropods (fleas/ticks), cat bites, iatrogenic exposure, queens to kittens

3. Regenerative anemia, lethargy, splenomegaly, fever, icterus

4. Feline infectious anemia (often related to FeLV)

What is the morphology, spread, and clinical signs of mycoplasma haemocanis? If there is a disease name, list it.

1. Small chains of cocci across surface of RBC that can appear Y-shaped

2. Infected blood = blood feeding arthropods (fleas/ticks), cat bites, iatrogenic exposure

3. Anemia, icterus (rare)

4. Formerly known as Hemobartonella canis

What are the 3 other types of mycoplasmas?

1. Mycoplasma wenyonii

• Immunocompromised/splenectomized cattle

• Iatrogenic (same needle in multiple animals)

• Syndrome = dependent edema and lymphadenopathy

2. Mycoplasma ovis

• Subclinical in sheep/goats

3. Mycoplasma haemosuis

• Pathogen of young and splenectomized pigs (causes severe hemolytic anemia)

4. Candidatus mycoplasma haemolamae

• Llamas and alpacas

• Opportunistic

• Mild anemia, similar to cattle

What does intravascular hemolysis versus extravascular hemolysis cause?

1. Intravascular = hemoglobinuria, ghost cells, pink plasma, acute anemia (RBCs destroyed in the vessels which causes hemoglobin to be filtered through the kidneys if haptoglobin is saturated)

2. Extravascular = unconjugated bilirubin in the blood, splenomegaly, gradual anemia, icterus (RBCs are removed and destroyed by macrophages in the spleen, liver, and bone marrow which results in unconjugated bilirubin that the liver has to filter, and if it can’t it builds up and causes icterus)

What are some traits of anaplasma sp?

1. Intraerythrocytic rickettsial parasite

2. Anaplasmosis + regenerative anemia

3. Similar in size to HJ bodies, but often more than 1 occurring in the RBC

4. Early infection shows 50% RBCs infected, later less than 5%

Anaplasma marginale morphology, spread, clinical signs

1. 0.5-1um dark blue inclusion on periphery of RBC

2. Transmitted by ticks (Boophilus annulatus) + horse and deer flies (most prevalent tick borne disease in cattle worldwide)

3. Infected cells destroyed by liver/spleen = severe anemia, immune mediated anemia (spherocytes/agglutination)

What are the other two anaplasmas covered in lecture?

1. Anaplasma Centrale - less pathogenic and used as vaccine

2. Anaplasma ovis - transmitted via ticks, biting flies, iatrogenically

How is canine babesiosis diagnosed? What are the two categories?

Observation of basophilic, pear-shaped trophozoites in RBCs on stained smears

1. Large Babesia sp: 3-7um, single/pair tear drop forms

2. Small Babesia sp: 1-3um, signet ring form

What are some methods of transmission for canine babesiosis?

1. Tick infestations/exposure

2. Recent dog bites (B. Gibsoni especially)

3. Blood transfusions

4. Splenectomy

5. Immunosuppresion

6. Transplacental transmission

What are some of the clinical findings for canine babesiosis? Pathology?

1. Thrombocytopenia - most common regardless of species

2. Leukogram is highly variable

• Pronounced leukemoid response, reticulocytosis, leukopenia are possible

3. Anemia (Intra/extravascular hemolysis)

4. Icterus (Intra/extravascular hemolysis)

5. Splenomegaly (Intra/extravascular hemolysis)

Babesia alters RBC membrane antigens and results in intravascular + extravascular hemolysis or can cause immune mediated hemolytic anemia (B. gibsoni) or blockage of capillaries due to sticky RBC membrane (severe case) or cytokine driven systemic inflammation (severe case)

What species does B. Bigemina/B. Bovis infect and what is its disease name?

1. Infects cattle

2. Texas fever/redwater fever/cattle tick fever

What is the morphology of B. Bigemina/B. Bovis?

• Lighter staining

• Pleomorphic

• Tear drop/pear shaped

• Large (3-5um long, 2um wide)

• Intracellular organism often in pairs that forms an acute angle

What are some of the clinical signs of B. Bigemina/B. Bovis?

1. Acute leukemia

2. High fever

3. Enlargement of spleen/liver

4. Results in death up to 90% of susceptible naive cattle

5. Attacks and destroys RBC’s

What are some of the differences between B. Bigemina and B. Bovis?

B. Bigemina

• Infects large RBC’s

• Causes intravascular hemolysis

• Clinical = high fever, jaundice, hemoglobinuria, severe anemia

• More acute, but less fatal

B. Bovis

• Infects smaller RBC’s

• Sequestration (alters RBC surface, becomes sticky, capillary blockage, causes neurologic signs, shock etc)

• Slower onset, but much more fatal

How is B. Bigemina/B. Bovis prevented/treated?

• Dipping/spraying cattle immediately w/ acaracide Coumpaphos

• Cattle must be dipped at the same time to prevent re-infection

What is another name for Babesia equi? What are some key morphology aspects?

Theileria equi is new classified name

1. 4 organisms may be joined forming a maltese cross (sometimes can see single or paired, but not common appearance)

2. Organimsms can infect concurrently

3. Can parasitize lymphocytes initially

4. Destroys RBC’s like babesiosis, but infects lymphs like theileriosis (schizonts)

How is Theileria different in transmission than B. caballi?

1. Zygotes do not multiply in tick

2. Can be infected as larvae and transmit infection as nymphs, or nymphs to adults

3. Lose parasites after transmission

4. Only stimulated to complete maturation after tick attaches to feed

5. Usually transmission occurs after a few days of attachment to host

6. Infection can persist for life (chronic carrier)

How is B. caballi different in transmission than Theileria?

1. Zygotes multiply as “vermincules” (invade many of tick’s organs, esp. ovaries)

2. Pass Babesia to egg

3. Infected larvae, nymphal/adult tick of infected generation attach to new host - parasite is stimulated to undergo final maturation - infecting host

4. Usually transmission occurs after a few days of attachment to host

5. Infection typically cleared w/in a few years, horses may recover

What animals are infected with T. equi/B. caballi?

Equids = horses, mules, donkeys, zebras

Zebras = important recevoir for infection in Africa (does not develop disease, but carries)

What are some of the main clinical signs for T. equi/B. caballi? Which usually causes more severe disease?

• Fever

• Anemia

• Icterus

• Hemoglobinuria

• Inappetence

• Edema

T. equi is usually more severe + fatal, can cause cerebral babesiosis (from capillary blockage) + more pronounced hemolysis. B. caballi is usually a milder disease and most equids usually become temporary carriers

• Both causes RBC destruction

What is common morphology for B. caballi?

1. 2-4um long in pairs at right angles, can be hard to find in smear or acute disease

2. Both organisms can infect concurrently

3. Restricted to RBCs

What is the morphology of theileriosis and what species does it infect?

Morphology:

• Small - 1um

• Signet ring/comma shaped

Species:

• Cattle = Theileria parva, Theileria annulata

• Sheep + goats = Theileria lestoquardi

What is the pathology of theileriosis?

Lymphocytes infected initially + monocytes/macrophages (form schizonts) which creates uncontrolled proliferation of these cells (like cancer) which leads to massive lymphoid hyperplasia. Leukocytes die off and release merozoites which infect RBC’s but immunosuppression has already occurred and leads to secondary infections.

What are some clinical signs?

1. Severe anemia

2. Pulmonary edema

3. Wasting

4. Wasting

5. High mortality

What is the pathology of cytauxzoon felis?

1. Macrophages/mononuclear phagocytes infected first

2. Massive replication occurs

3. Become engorged/sticky and block vascular endothelium (mainly spleen, liver, lungs, bone marrow; schizonts in tissue + macrophages)

4. Obstruct small vessels (ischemia, organ failure)

5. Merozoites infected after and cause severe non-regenerative hemolytic anemia

6. Classified w/ theileria

What are some clinical signs for C. felis?

1. Thrombocytopenia + leukopenia

2. Acute lethargy

3. Lethargy

4. Anorexia

5. Fever

6. Icterus

How is C. felis diagnosed?

1. Signet rings in RBC’s - late in course of DZ

2. Schizonts in macrophages in cytology

3. Schizonts in spleen, liver, lymph node, bone marrow

4. PCR - more sensitive + specific

What blood cell shape is common with immune mediated hemolytic anemia?

Spherocytes

What is the disease name for plasmodium? Morphology?

Plasmodium is responsible for malaria and is usually asymptomatic, but if not, can cause hemolytic anemia, depression, and acute death. Intravascular and extravascular hemolysis common.

1. Schizont is commonly seen in RBC’s and sometimes platelets/WBC’s (12-24 merozoites, maturing stage)

2. Trophozoite stage is signet ring appearance and can be seen in same cells (early stage)

3. Gametocyte can displace nucleus and can be seen in the same cells (sexual stage picked up by mosquitoes)

4. All can contain yellow/brown granules = iron

How can plasmodium be differentiated from haemoproteus?

1. Schizogony occurs mainly in endothelial cells of blood vessels and organs, not in RBC’s

2. Only gametocytes are seen in circulating RBC’s vs. all stages like Plasmodium and are sausage shaped

3. Usually does not cause severe hemolysis

What is the morphology of Leukocytozoon?

1. Dark staining macrogametocytes

2. Light staining microgametocytes

3. Do not contain stainable iron usually (not usually in RBC’s, depends on species)

4. Found in peripheral blood (thought to infect young RBC’s) + WBC’s + macrophages (depending on species)

5. Large distorted cells with grossly misplaced nucleus

What animals is Leukocytozoon highly pathogenic for? What cells are mainly infected in these species?

1. Waterfowl: ducks, geese, swans (infects RBC’s + leukocytes)

2. Turkey (infects RBC’s and macrophages)

What tissues does Leukocytozoon infect?

Invades endothelia and parenchyma of liver/heart/kidneys and mature initially and then spreads to RBC’s/WBC’s since spread by flies

What is the morphology of Trypanosoma?

1. Much smaller than Dirofilaria (2-3 RBCs vs. 8 RBCs for Dirofilaria)

2. Extracellular in blood

3. Ribbon-like, banana shaped, lateral, undulating membrane

4. Thin tail (flagellum)

5. Rarely seen in direct smear, more often seen in stained smears

What are the 3 species of Trypanosoma discussed in class?

1. Trypanosoma cruzi - causes disease in dogs/people (Chagas’ disease/sleeping sickness)

• 16-20um in length

1. T. theileri - nonpathogenic in cattle

• 25-120um in length

1. T. melophagium - nonpathogenic in sheep

How long does it take for Dirofilaria immitis to develop to young adults in the tissues? Sexual maturity in the heart? Where do they grow in the heart?

1. 3-4 days to go from L3-L4 larvae

2. 45-65 days to go from L4-L5 (prevention effective up to this point, ~2 months)

3. Mature to adults in the heart for another 60-70 days and then start depositing microfilaria

4. Adults found in right ventricle and pulmonary artery

What is the other worm that Dirofilaria immitis must be distinguished from? What are 2 key differences used to ID?

Dipetalonema reconditum

1. Head in Dirofilaria = tapered versus reconditum = blunt

2. Tail in Dirofilaria = straight versus reconditum = hooked

What is the intermediate host for D. reconditum?

Cat/dog flea

What are 5 tests to ID microfilaria? Which ones can differentiate between Immitis and Reconditum?

1. Direct smear/stained smear (can differentiate usually)

2. Microhematocrit method (Buffy coat)

3. Knotts test (can differentiate)

4. Filter technique

5. Immunodiagnostic tests (ELISA, can differentiate)

What are 3 clinical signs for Ehrlichia?

1. Splenomegaly

2. Fever

3. Nasal discharge/ocular discharge

What is Ehrlichia canis transmitted by? Morphology?

Transmitted by brown dog tick

• Small cluster of morulae (3-6um) appear as coccoid shaped structure

• Infects monocytes/lymphocytes

What species does E. Playtys and E. Ewingii infect? What cells do they infect?

E. Playtys

• Infects platelets w/ multiple morulae

• Infects dogs

E. Ewingii

• Infects neutrophils/eosinophils

• Humans can contract

• Infects dogs

What cells does Ehrlichia equi infect? What species?

• Infects horses

• Neutrophils + eosinophils

What species does Ehrlichia risticii + Ehrlichia sennetsu infect? What cells?

E. risticii

• Horses

• Monocytes/macrophages

E. sennetsu

• Horses/humans

• Monocytes