RBCs Pt. 3 and WBCs Pts 1 and 2

Question 1

what magnification do we need to use to visualize parasitic RBC inclusions? why?

Answer 1

100x oil magnification because these are really small!!

Question 2

describe hemotropic mycoplasma spp. inclusions in RED blood cells (4)

Answer 2

1. epicellular: small bacteria on the surface of RBCs
2. present in many species; pathogenicity varies (most pathogenic in cats and pigs)
3. parasitemia will wax and wane, so diagnosis on blood smear has LOW sensitivity and specificity (these guys like to hide in organs like the spleen) so test of choice is PCR
4. be careful to distinguish from stain precipitate: look for these guys at the outer edge of the cell; will look like basophilic dots (cocci), in small rings or linear chains

Question 3

describe cytauxazoon felis parasitic inclusion in RED blood cells (5; it’s a lot sorry)

Answer 3

1. this is a tick-transmitted, intracellular protozoal organism closely related to Babesia whose reservoir species is bobcats and has only been identified in cats for domestic species; has 2 stages: tissue (shizonts) and RBC stage (prioplasms)
2. tissue stage (schizonts): primary cause of clinical disease; occurs BEFORE organisms can be seen on a bloodsmear; schizonts = parasites in macrophages and occlude the vessels of most organs
3. RBC stage: prioplasms: second phase of infection, variable parasitemia, very small, round to oval with a dot-like nucleus (looks like a ring with a tiny blue dot and a little skirt inside the RBC); resembles Babesia gibsoni
4. initial clinical signs are nonspecific, with rapid progression within a few days (early diagnosis is critical); can present as severe non-regenerative anemia and/or sepsis-like signs (leukopenia with toxic change, thrombocytopenia, shock-like signs)
5. primary method of diagnosis is checking on a bloodsmear, but if you suspect this and the smear is negative, do a fine needle aspirate to look for the tissue phase! PCR is also available and confirmatory, but takes a few days to send off and get results so if suspect just start treating

Question 4

describe anaplasma spp. parasitic inclusions in RED blood cells

Answer 4

1. obligate, tick-transmited, intracellular bacterium found in tropical and subtropical regions worldwide; two kinds
2. A. marginale: in cattle, is high virulent but age-dependent as calves are resistant but naive young adults are highly susceptible; causes severe extra-vascular hemolytic anemia, parasitemia peaks prior to anemia, can confirm via serology of PCR
3. A. ovis: in sheep, deer, and goats: variable disease, mild to severe
4. small, 0.5-1 um, basophilic structures at periphery of cell: MUST differentiate from Howell-Jolly bodies and basophilic stippling, do this because you may 1-2 anaplasma spp. per cell (HJ only one, and basophilic stippling many per cell)

Question 5

describe babesia spp. parasitic inclusions in RED blood cells

Answer 5

1. tick-transmitted, intraerythrocytic protozoan parasites that only have a RBC stage (merozoite); can be large (B.canis) or small; primarily in dogs, but also in cattle, horses, and sheep
2. B. canis (large-form): low virulence in US, but more virulent international; often subclinical but if clinical disease will be more likely in puppies (sick or immunosuppressed), causing mild anemia and mold to moderate thrombocytopenia; 2-4um, piriform/tera-drop shaped with a dot-like red nucleus and may be paired or up to 4 together
3. B. gibsoni (small form): worldwide, moderately virulent; increased incidence in pit bulls and dog fighting/dog bits increase infection risk; cause moderate-marked regenerative anemia, intra and extravascular hemolytic anemia, +/- spherocytes due to secondary IMHA, and moderate-marked thrombocytopenia
   round to oval, with a dot-like red nucleus and thin cytoplasmic membrane, >2um
4. general Babesia diagnosis:
   -blood smear: critical for virulent B. gibsoni, low sensitivity unless high numbers but using capillary blood (from ear vein or nail clip) may concentrate organisms
   -serology: titers, sensitive screening test but only documents exposure, cannot confirm acute infection
   -PCR: sensitive and specific but longer turnaround time

Question 6

describe microfilaria inclusions in RED blood cells

Answer 6

1. can be from dirofilaria immitis (heartworm) or dipetlonema reconditum
2. these are big! but may only see at feathered edge or in deep body of blood smear so CHECK THE WHOLE SMEAR
3. to differentiate between dif types of microfilaria, need Knott’s test or a heartworm antigen test because is difficult with just a blood smear

Question 7

describe the leukogram changes in the corticosteroid (stress) response (4)

Answer 7

neutrophilia, lymphopenia, monocytosis, eosinopenia

Question 8

describe the leukogram changes in the epinephrine response (AKA physiologic leukocytosis) (2) give 5 species variations

Answer 8

neutrophilia and lymphocytosis

1. young, healthy cats: common!
2. healthy foals: also common!
3. pigs: can see 2-3 times the upper RI for neutrophils and lymphocytes
4. uncommon in dogs: more likely in puppies and excitable breeds (toy breeds)
5. cattle: parturition or exercise induced

Question 9

describe the leukogram changes in the inflammatory leukogram and consider species variations

Answer 9

variable; classic is neutrophilia with a left shift and monocytosis; more common in young animals

Question 10

describe a left shift of neutrophils (3)

Answer 10

1. immature neutrophils in peripheral blood
2. typically consists of band neutrophils and should be orderly! where most immature stages seen in lowest numbers and more mature stages seen in highest numbers
3. if not orderly, suggests bone marrow disorder

Question 11

what do band neutrophils look like? (3)

Answer 11

1. horse-shoe shaped nucleus with parallel sides and a smooth outline
2. coarse chromatin (distinguish from monocyte)
3. slightly larger with slightly bluer cytoplasm than a segmented neutrophil

Question 12

how to distinguish band neutrophils from monocytes?

Answer 12

1. monocytes have bulbous or knob like ends of their nucleus
2. monocytes have deeper blue-grey cytoplasm, often with vacuoles
3. monocytes have smoother chromatin and are typically bigger than band neutrophils

Question 13

what are the 2 types of neutrophils-left shift?

Answer 13

1. regenerative left shift: orderly (mature»immature); seen with normal or increased segmented neutrophils and lower numbers of immature cells; better prognosis
2. degenerative: immature»mature OR neutropenia with ANY left shift!!; is a sign of overwhelming inflammation and things going downhill

Question 14

describe toxic changes to neutrophils generally (3)

Answer 14

1. morphologic abnormalities of the cytoplasm that occur during maturation, seen with severe infection/sepsis, severe inflammation, and/or excessive tissue damage
2. mild changes can be seen potentially with any condition with inflammation, infectious or non-infectious
3. moderate-marked changes indicate bacteria infections, other infections, and sepsis

Question 15

list and describe 4 toxic changes to neutrophils

Answer 15

1. Dohle bodies: the mildest of the toxic changes, blue-grey aggregates of retained ER; rare sightings are normal in healthy cats!
2. basophilic cytoplasm
3. vacuolization: when seen in neutrophils WITHOUT other toxic changes (like basophilia or Dohle bodies) is likely artifact due to prolonged storage or old sample
4. toxic granulation” purple red granules; rare in cats, dogs, and LAs but is the best clue for inflammation in exotics!

Question 16

describe neutrophilia

Answer 16

1. the CLASSIC response to inflammation
2. may also see a let shift and toxic changes
3. with chronicity, left shift may go away as the bone marrow outpaces the demand/consumption in tissue
4. the degree of neutrophilia depends on the intensity of the stimulus and the species’ bone marrow reserve/responsiveness (ex. dogs have the largest storage pool, so neutrophilia is normal and can get really high, but cows have a small storage pool so any increases indicate inflammation, horses and cats are in the middle)

Question 17

how do you tell neutrophilia from stress?

Answer 17

1. they often present together, so it is important to clinically identify inflammation (if an animal is sick due to inflammation, they will also be stressed because they are sick)
2. inflammation is more likely than just stress if: neutrophilia is 2-3x the upper RI, left shift and/or toxic changes are present, or if there is other evidence of inflammatory disease present (fever, increases acute phase proteins)

Question 18

describe extreme neutrophilia (3)

Answer 18

1. on the highest end of the RI; also called a leukomoid response
2. can find localized purulent pus pockets (pyometra, walled off abscesses, endocarditis, etc.)
3. less common causes include:
   -hepatozoon americanum infection: a tick-borne protozoal diseae
   -neoplastic proliferation of neutrophils

Question 19

describe neutropenia due to inflammation (6)

Answer 19

1. occurs when tissue demand/consumption exceeds production in the bone marrow, usually due to an acute, severe, overwhelming demand
2. can also be due to endotoxemia: which causes neutrophils to migrate from the circulating pool to the marginating pool (stick to endothelial walls = can’t sample)
3. some infectious agents can destroy neutrophil precursors in the bone marrow (canine and feline parvovirus)
4. typically associated with inflammation, a left shift and +/- toxic changes
5. species variations:
   -dogs have the largest storage pool so neutropenia suggests severe, overwhelming inflammation (BAD)
   -cats and horses have a moderate storage pool so neutropenia is still severe but happens easier than in dogs
   -ruminants have a very little storage pool, so neutropenia is a normal response to significant acute inflammation
   -calves have a higher storage pool though!! so respond more like cats and dogs
6. can be due to gram negative bacterial infection/septicemia, or salmonellosis (BIG in horses)

Question 20

describe what is happening with a normal neutrophil count during inflammation

Answer 20

1. can be seen with mild inflammation, recovery from neutropenia (regenerating), or progression towards neutropenia (degenerating)
2. detect the inflammation by:
   -looking for left shift and/or toxic changes
   -other indications of underling inflammation (fever)
   -changes in acute phase proteins

Question 21

summarize the 4 main species differences regarding neutrophils

Answer 21

1. species with less responsive bone marrow/lower BM storage pool: ruminants and horses to some degree, neutropenia develops more rapidly with acute inflammation and neutrophil changes are not as prominent as in cats and dogs
2. fibrinogen: increases with inflammation and also with dehydration
3. other acute phase proteins can detect inflammation: like serum amyloid A in horses
4. ruminants neutrophil to lymphocyte ratio: healthy adult ruminants are lymphocyte predominant (N:L = 1:2); inversion of this ration (predominance of neutrophils) suggests inflammation

Question 22

list some causes of non-inflammatory neutropenia and neutrophilia and explain how to differentiate these from inflammation (5)

Answer 22

1. iatrogenic: chemotherapy, cytotoxic drugs
2. pancytopenia: decrease in all 3 cell lines (neutrophils, red cells, and platelets): due aplastic anemia
3. immune-mediated neutropenia: present really sick and febrile (disease of exclusion, no confirmatory tests)
4. breed variations: border collies (trapped neutrophil syndrome) or grey collies (cyclic hematopoiesis affects neutrophils more because shorter half life)
5. idiopathic: we don’t know but we ruled everything else out

Question 23

list and describe 6 acquired disorders that affect neutrophil function

Answer 23

1. glucocorticoids: decreased adhesion molecules
2. systemic diseases: diabetes mellitus, renal failure, liver cirrhosis
3. nutritional deficiencies
4. some viral diseases: FeLV, BVD
5. burn patients
6. periparturient neutrophil dysfunction in cattle: poorly understood

Question 24

identify and state the significance of various neutrophil inclusions (4)

Answer 24

1. granulocytic ehrlichiosis and anaplasmosis: primarily in dogs, rarely in cats, can see in large animals; cause morulae in neutrophils (pale blue/lavender, round, stippled to clumped structure looks like a raspberry, but using this method to detect this disease sucks; use serology or PCR instead); thrombocytopenia, anemia, and polyarthritis
2. monocyte ehrlichiosis: causes morulae in MONOCYTES and NOT neutrophils but are rarely seen in blood and only present during acute infection in hella low numbers so again use PCR or serology
3. distemper inclusions: aggregates of viral particles, transiently visible in blood of dogs with acute infection that are various shades of purple and blue but affect all types of leukocytes and erythrocytes because this virus infects hematopoietic precursor cells in the bone marrow
4. bacteria: usually not seen in peripheral blood, but located in tissue