Avian Hematology

AVIAN VENIPUNCTURE

SPECIAL CONSIDERATIONS PRIOR TO HANDLING AND VENIPUNCTURE

• Weight and health of species determines amount of blood
• Have supplies ready
• Sites vary with species and physical condition
• Provide supportive care
• Make sure patient is stable
• Do blood draw in stages as needed

Any bird that looks sick in their cage is a sick bird, and may be critical. Careful attention to patient while handling is a must to make sure the bird isn't getting stressed by the procedure of drawing blood. A very sick or traumatized bird may need to be placed in a warm incubator with oxygen, and gradually receive fluids and antibiotics before diagnostics are even considered. A bird having difficulty breathing while perched will have much more difficulty when excited and stressed when handled. When in doubt, always put a stressed bird down.

Avian venipuncture: Amount

Asymtomatic patients:

Can safely draw in ml's 1% of the body weight in grams (change 1% to decimal and multiply by weight)

weigh in grams

Sick birds: Take 1/2 that amount

Venipuncture: Amounts

Example #1 sunconure weighs 110 grams 1% = 0.01 x 110 = can take 1.1 ml of blood

Example #2 cockateil weighs 85 grams 1% = 0.01 x 85 = can take 0.85ml of blood

Venipuncture: 

Have supplies ready before you draw

• Know lab requirements
• Amounts, plasma/ serum requirements vary with labs
• Microtainer tubes for small amounts come in red tops (sst), lav tops (edta), and green (heparin: heavy metal, some species cbc)
• syringes sizes vary per species size and volume of blood needed  

Venipuncture site: Jugular vein

The right jugular vein is visualized on a cockateil. In most psittacines there is a featherless area on the right side of the neck. Wet the area with alcohol to visualize the vessel. Enter the skin first and then the vessel. Hold off after venipuncture to prevent hematoma formation.

Right jugular vein visualized on an African Grey parrot while under anesthesia. The right jugular vein is larger than the left and the best site for venipuncture in psittacines.

Dr. Teresa Lightfoot demonstrates the one handed technique on this cockatoo.

 

Ulnar or Brachial Vein

In non psittacine avian patients such as raptors, pelicans, waterfowl, alternative venipuncture sites other than the jugular vein may be necessary. In these patients, the ulnar (brachial) and medial metatarsal veins can be used.

In this seabird, the ulnar or brachial vein will be a better site of collection instead of the jugular vein.

The ulnar vein crosses the ventral surface of the elbow joint. Make sure to hold the vessel off well after the procedure as hematoma formation can occur at this site due to less subcutaneous tissue surrounding the vessel.

 

Medial Metatarsal Vein

The medial metatarsal vein is located on the medial side of the lower leg in most birds. This is the preferred site to use in the waterfowl.

Venipuncture of the medial metatarsal on a chicken.

Avian Hematology: Avian CBC includes the following:

• Pcv and total protein (or total solids)
• note plasma color
• buffy coat check for microfilaria
• granulocyte count on hemacytometer using the eosinophil unopette method
• wbc differential with rbc morphology and platelet estimation.
• note any parasites

Avian Hematology: Pcv/total protein

Hydration Status affects serum /plasma yield

• Normal pcv: 35% to 55%
• Plasma/ serum color :normally slight icterus
• Total protein values : 3.0-6.0 g/dl (may vary with age of patient)
• T.P. < 2.3 g/dl carries a poor prognosis

Eosinophil pipette and unopette; note the pipette size is 25 microliters.

Avian cbc: indirect wbc 

Eosinophil unopette system used to stain heterophils and eosinophils for easy identification when counted on the hemacytometer. This count is later used to calculate the total leukocyte count.

Charging both sides of the hemacytometer using the eosinophil unopette. To avoid staining the erythrocytes do not allow the unopette to stand longer than 5 minutes before charging the hemacytometer. Heterophils and eosinophils appear as round "Refractile pink-red" cells in the hemacytometer using the eosinophil method for obtaining the total leukocyte count.   Calculate wbc indirectly using eosinophil unopettes, 25 microliter pipette Fill/Incubate eosin unopette 5 minutes Eosin stains eosinophils and heterophils (granulocytes) basophils do not stain

Avian cbc: wbc formula

Total WBC = number if cells stained in chamber x 1.1 x 16x100 
                            percentage of heterophils + eosinophils  

 

1. Take Granulocyte ct from hemacytometer

2. multiply by formula [ x1.1 x 16 x 100 ]

This total number is equivalent to [1760]

(Calculates in dilution factor from unopette)

3. Divide this number by percentage of eos and heterophil from differential

See examples below :  

Indirect wbc

Example #1:

hemacytomer granulocyte count = 450

Diff granulocyte % = 65% hets, 4% eos

= 69 ( divide by this #)

450 x 1760 = 792,000

792,000 / 69 = wbc 11,478 mm3

cbc: indirect wbc

Example #2

hemacytomer granulocyte count = 172

Diff granulocyte % = 35% hets, 2% eos

= 37 ( divide by this #)

172 x 1760 = 302,720

302,720/37 = wbc 8,181 mm3 

cbc: differential

• white blood cell counts vary with species
• Some species normally have higher lymph% than heterophil%

Avian differential

• Perform 100 cell count wbc differential
• Perform thrombocyte estimate on smear
• Report rbc morphology
• Note any parasites or report as none seen
• Note any toxic cells , blasts, or atypical cells
• Once differential is done, calculate absolute values by multiplying percentage values by the total white blood cell count.

Blood smear: ID of cells

• red blood cells
• thrombocytes
• granulocytes: heterophils (eq. to neutrophil)
• eosinophils
• basophils
• Agranulocytes: lymphocytes
• monocytes  

 Avian Red Blood Cells

• All nucleated
• Report RBC morphology: Aniso, Poly, Poik as slight, mod, marked or as 1+, 2+, 3+
• Polychromasia and Rubricytes (immature rbc) best sign of regeneration 

Immature red blood cells

• Note immature rbc's as: 1+, 2+, 3+ immature rbc's separate from
• polychromatophilic rbc's
• rubriblast, prorubricyte, rubricytes
• rubricytes = (basophilic, late polychromatophilic, early polychromatophilic), polychromatophilic rbc, mature rbc
• Immature rbc rounder cell, pyknotic nucleus as matures 

Thrombocytes

nucleated, maturation in bone marrow An adequate number is 1-2 thrombocytes per oil immersion field. clumping normal nucleus pyknotic appearance oval cell w/ colorless cytoplasm, may contain 2 - 4 magenta granules

Granulocytes

• Heterophils
• Eosinophils
• Basophils  

Granulocytes: Heterophil

• Heterophil : corresponds to the mammalian neutrophil
• Most common wbc in some avian species
• Round cell with colorless cytoplasm
• Brick red /rod shaped granules
• Nucleus has 2-3 lobes  

Granulocytes: Eosinophil

• eosinophil: round cell with light blue cytoplastm
• granules will be consistent on individual slide but color shape and size of granules can vary from species to species
• Find heterophil to compare: eos granules are brighter or duller than heterophil granules 

Granulocytes: Basophil

Basophil: small round cells with a centrally located nucleus nucleus often obscured by deeply basophilic granules cytoplasm, when seen is colorless to light purple red

Agranulocytes: Lymphs

• Lymphocytes: small, medium, large
• Usually round (to irregular cells) , resembling mammalian lymphocytes.
• round, centrally located nucleus with dense clumped chromatin
• varying amounts of cytoplasm is light blue 

Small lymph vs thrombocyte

Lymph: round cell dense clumped chromatin light blue cytoplasm azurophilic granules if present, throughout cytoplasm

Agranulocytes: Monocytes

Monocytes: largest wbc's cell irregular in size, with bilobed to round nucleus cytoplasm is blue grey, often with granular appearance. similar to mammalian monos, +/- vacuoles

Avian Blood Parasites 

A. Plasmodium (Avian Malaria) B. Haemoproteus C. Leukocytozoon

Blood Parasites: A. Plasmodium

rbc and extracellular Parasite displaces host cell nucleus or "plows" it out of the way Pathogenic in canaries, gyrfalcons, chickens, ducks and pidgeons. Hemolytic anemia, hemoglobinuria, leukocytosis and lymphocytosis

  

Blood Parasites: B. Hemoproteus

Psittacines (Hookbills), pidgeons, and seabirds most commonly affected rbc Parasite occupies 1/2 of cell cytoplasm Intermediate host insect vectors generally considered nonpathogenic

 

Blood Parasites: C. Leukocytozoon

Important dz in ducks, geese fowl, turkeys largest of intracellular blood parasites rbc's and leukocytes parasite distorts host cell Hemolytic anemia and hemoglobinuria

  

The End

 Questions ?
Susan Dempsey CVT