Anatomic Pathology Readings 11-14

Question 1

How can you get a cDNA (copy of DNA) for PCR of an RNA infectious agent?

Answer 1

Use the enzyme “reverse transcriptase” to promote synthesis of DNA from the RNA template = cDNA (copy of the DNA)

Question 2

How does PCR work?

Answer 2

• Primers (nucleotides) compliment intended target DNA and are added to reagents
• heated to separate DNA to two strands
• cooled to allow primers to bind to DNA
• heated again
• 25-40 cycles
• PCR products (amplicons) read by gel electrophoresis

Question 3

Advantages of molecular approaches:

Answer 3

• Faster and no need for special media
• Diagnose organisms hard/slow to grow or ones that cannot be cultured
• Detect evidence of infection before Ab response
• Clarify Ab + due to infection or maternal Ab
• Document latent infections or carriers
• Diff btwn pathogenic and non-path or vaccinal isolates
• Retrospective analysis and characterization of zoonotic infection

Question 4

PCR can help dx organisms that cannot be cultured like:

Answer 4

many viruses
some mycoplama (haemobart)

Question 5

PCR also good with disease organisms that grow slowly for example:

Answer 5

mycobacteria

Question 6

Good examples of diseases that historically have required seroconversion but now can be diagnosed via molecular assays: name 2

Answer 6

RMSF (rocky mountain spotted fever)

Leptospira

Question 7

Examples of a PCR clarifying infectious dz or maternal Ab

Answer 7

FIV (otherwise can’t tell!)

Question 8

T/F: PCR and RFLP (Restriction fragment length polymorphism) can help select appropriate antimicrobial therapy.

Answer 8

true

Question 9

List Koch’s 3 postulates:

Answer 9

1. Agent found in every case of the disease
2. Agent was not found in other diseases
3. Agent can be isolated and cultured and cause disease in a new host
   (4. Agent could be isolated from the experimentally inoculated host)

Question 10

Why use Koch’s postulates - what do they mean?

Answer 10

They show that a given agent was the cause of clinical disease if the postulates were satisfied

Question 11

What’s considered an additional point of proof of infect dz:

Answer 11

If an agent can be isolated from experimentally inocculated host

Question 12

When trying to find a cause for a disease, what is the most important first question for a clinician when they have detected an infectious agent?

Answer 12

Is it is likely that the infectious organism is truly responsible for clinical disease or other abnormalities?

Question 13

Give an example where the PCR-positive blood sample doesn’t guarantee an infection.

Answer 13

Hemobartonella = IMHA

Hemobartonella can cause IMHA but IMHA can be caused by other causes

Question 14

Are there known pathognomomic or typical gross lesions for anesthetic-associated death (AAD)?

Answer 14

no!

Question 15

Over half of cases with natural disease had ____ lesions which included:

Answer 15

pulmonary:

• Aleurostrongylus abstrusus
• pneumonia
• acute aspiration pneumonia

Question 16

Pulmonary dz was the most common lesion tied to AAD, which two wereafter that?

Answer 16

-heart disease (2 HCM; 4 lymphocytic myocarditis)
-systemic (Panleukopenia; Toxoplasmosis)
(also CNS disease causing one death; noting hepatic lesions in 3 cases)

Question 17

What is the connection between renal disease and anesthetic-associated death (AAD)?

Answer 17

Of the 54 cases of AAD death, none had gross or histo evidence of renal dz! (no connection)

Question 18

What is the life cycle of Aleurostrongylus abstrusus

Answer 18

1. Adult worms in terminal bronchioles
2. Eggs are laid by females, with L1 coughed up and excreted
3. L1 exits in the feces to intermediate host: snail, slug
4. L1 matures to L3 in intermediate host and then either eaten by cat (DH) or paratenic host then to the definitive host

Question 19

t/f cats can never eliminate A. abstrusus on their own?

Answer 19

False! they will eliminate it after 5months and cannot be re-infected

Question 20

What is the gold stnd diagnosis of A. abstrusus

Answer 20

Baerman fecal exam

Question 21

Aleurostrongylus abstrusus infections results in: (2)

Answer 21

• Granulomatous pneumonia
• Hypertrophy and hyperplasia of tunica muscularis in the pulmonary artery = pulmonary hypertension (can persist for up to a year)

Question 22

What parasite other than Aleurostrongylus abstrusus can cause pulmonary hypertension in cats?

Answer 22

Dirofilaria immitis

Question 23

Why do cats die from anesthesia with pulmonary hypertension?

Answer 23

There is a decrease in gas exchange -> less blood back to LA -> LV

Question 24

There were cases with lymphocytic myocarditis- etiology?

Answer 24

Toxoplasma

Possibly Bartonella henselae or a cardiomyopathy

Question 25

Why can't you rule out toxoplasma with a negative IHC?

Answer 25

It only takes a few of tiny T gondii tachyzooited to incite sign inflammation

Question 26

Can you detect a mechanical causes of death eg. soft palate occlusion or improper ET-tube intubation on PM?

Answer 26

no

Question 27

Can you detect respiratory acidosis secondary to hypoventilation on PM?

Answer 27

no

Question 28

Can you detect heart conduction failure on PM?

Answer 28

no

Question 29

What should you sample for histopath on stillborn, or neonatal death? (6 + 1 general)

Answer 29

``` Placenta Liver Kidney Lung Heart Brain Anything specific ```

Question 30

What should you sample fresh* for virology and bacteriology?

Answer 30

``` same as histo except also sample spleen but don't sample heart and brain (Placenta Liver Kidney Lung Spleen Anything specific) ```

Question 31

Who's serum do you sample in fetal necropsy?

Answer 31

Dam and fetus

Question 32

What is most critical tissue to obtain on fetal necropsy:

Answer 32

Placenta!  may have more info than fetus itself

Question 33

In the case of neonatal death, do what before opening up fetus?

Answer 33

- record wt, measure crown to rump, check for hair growth and texture, check for teeth (teeth = close to term) - rinse fetus and fetal membrane and put on wet table to examine

Question 34

In the case of neonatal death, specifically check for these before opening up animal:

Answer 34

- congenital defects (e.g. VSD) - evidence of redness, swelling, improper mobility= fx - umbilicus for swelling (hernia)

Question 35

In the case of neonatal death, what should you do once cavities are open:

Answer 35

Collect samples: 1. virology/bact (1cm+) 2. histo- in formalin (<0.5cm) 3. Sample of blood & stomach contents in a red top tube with new needles

Question 36

When examining placenta on dog, if you see dark red-green band what do you conclude?

Answer 36

It's normal! marginal hematoma on zonary band (zonary placenta) (pale brown and smaller in a cat)

Question 37

Cats placenta differs from dog placentas how?

Answer 37

The marginal hematoma is more narrow with less distinction and is pale brown

Question 38

It's important to do this with placenta to check of evidence of inflammatory cells and bacterial organisms:

Answer 38

Take an impression smear of the chorioallantois

Question 39

What organism loves placenta?

Answer 39

Brucella organisms

Question 40

What if you see a red tinge on internal organs of fetuses?

Answer 40

incidental (unless well demarcated areas of obvious hemorrhage and/or edema are present)

Question 41

What about a clear yellow or red fluid in body cavities of fetuses?

Answer 41

incidental

Question 42

What if you see clotted blood within cavities in a fetus?

Answer 42

This is not normal- may indicate trauma

Question 43

Fetal mummification (dry, shrunken, wrinkled) can mean what?

Answer 43

Non-sepcific. Can point to a viral cause or many other causes in utero

Question 44

What is the cause/name of a fetus which is just a bag of bones with reddish brown uterine fluid?

Answer 44

This is fetal maceration and is commonly due to bacterial infection (specific to bateria*)

Question 45

What is an example of an incidental congenital defect?

Answer 45

hydrancephaly (brain's cerebral hemispheres are absent) | unilateral renal agenesis

Question 46

What is the most common cause of viral abortion in neonatal dogs?

Answer 46

herpes!

Question 47

Clinical signs and PM findings with Herpes

Answer 47

- Sudden death, lethargy, excessive crying | - PM: multi organ hemorrhage (kidney, lungs, liver most obvious)

Question 48

t/f herpes likes warm temp to grow?

Answer 48

FALSE! Herpes virus is optimized at lower temps

Question 49

You can confirm herpes infection with what tests?

Answer 49

histo and PCR

Question 50

How does having herpes affect pregnancy in cats?

Answer 50

Abortion is rare in cats with herpes. It is mostly tied as a secondary cause because of respiratory disease in the queen BUT kittens can have neonatal death due to herpes

Question 51

List other differentials for sporadic abortions and neonatal deaths in dogs, other than the most common (herpes virus)

Answer 51

CPV-1 (canine parvovirus-1) CDV (canine distemper virus) -> rare CAV-1 (canine adenovirus-1) -> not typical; pneumonia in pups BTV (bluetongue virus) (can be secondary to maternal morbidity or due to direct infection)

Question 52

Ddx viral abortion and neonatal death in cats:

Answer 52

- FeLV - FPLV (feline panleukopenia virus/parvo) - FIV - Feline coronavirus - FCaV (feline calicivirus)

Question 53

2 most common bacterial neonatal death in dogs:

Answer 53

Brucella canis (take more than 48 hours to culture) Streptococcus spp Others: salmonella, campylobacter, leptospira, e.coli

Question 54

Protozoal causes for abortion, stillbirth, neonatal death dogs and cats:

Answer 54

Toxoplasma Neospora (dogs and cats are DH for both)

Question 55

This virus is assoc with fatal pneumonia in puppies <4wks

Answer 55

CAV-1

Question 56

How do you tell if there was trauma with dystocia?

Answer 56

regional hemorrhage or edema | +/- fractures

Question 57

What makes you think infanticide?

Answer 57

hemorrhage in brain and broken skull

Question 58

seroconversion detects what?

Answer 58

active immune response 10-14days post inital challenge

Question 59

Serology is best used when?

Answer 59

Serology is best used to monitor for active infections in a group of cattle, rather than specific disease in a single animal

Question 60

"if animal test +, it has dz" =

Answer 60

pos predictive value

Question 61

VNT vs ELISA with serology

Answer 61

ELISA only tells you that Ab binds to Ag- may bind to part of virus that's not nec for infection VNT- demonstrates what amnts of Ab will neutralize the virus

Question 62

serology detection for bacteria: egs

Answer 62

varies:  ELISA, CF, agglutination, leukotoxin inhibition

Question 63

LN vs antileukotoxin

Answer 63

LN measures fxn Ab which interfer with toxin b/c antileukotoxin binds to any leukotoxin (LN better indicator of + predictive immunity)

Question 64

problem with VI at cellular level

Answer 64

not all cells show cytopathy with certain dz's (non-cyto BVDV)

Question 65

main disadv of VI

Answer 65

1. takes 2 wks before actual - 2. cells needed may be hard to get 3. can't find new or emerging viruses

Question 66

use EM for

Answer 66

enteric and sometimes resp viruses

Question 67

disadv of bacterial isolation

Answer 67

1. tech has to pick correct colony 2. bacteria present are all N in resp tract 3. antibiotic therapy can = negative culture

Question 68

direct vs indirect FA

Answer 68

direct detects Ag

Question 69

disav of FA

Answer 69

dye won't stay long without breaking down so can't have another lab consult (IHC helps with this problem)

Question 70

limitation of IHC

Answer 70

lack of specificity to monoclonal Ab, esp. newly recogn agents

Question 71

IHC vs in-situ hybridization

Answer 71

same but doing IHC in-situ is potentially more sensitive and can identify low numbers of copies of the infectious agent nucleic acid

Question 72

RT-PCR also called

Answer 72

q (quantitative) PCR b/c can show if large load of agent if fluor earlier on

Question 73

these study pathogenesis, host-agent interaction and type animal pathogens (req. strict QC)

Answer 73

microarrays

Question 74

should use PCR on secretions in the water?

Answer 74

no- meaningless

Question 75

PCR vs. IHC, in-situ hybrid which is more accurate

Answer 75

IHC, in-situ and PCR as adjunct

Question 76

Definition of stillborn

Answer 76

Born dead at full term

Question 77

Definition of neonatal death

Answer 77

Death within the first 3 weeks after birth