Marine Mammal Diagnostics & Clinical Pathology

Question 1

Where to obtain bone marrow samples in cetaceans?

Answer 1

Vertebral bodies, ribs (long bones lack medullary cavity) (F8)

Question 2

Phlebotomy sites in cetaceans

Answer 2

Periarterial vascular retes (PAVR) on dorsal and ventral midlines of fluke blades, peduncle, dorsal fin, hemal arch, common brachiocephalic vein (CBV, purely venous and for BP measurement), hepatic vein (PICC). (F8, West)

Question 3

Clin path analytes helpful as indicators of inflammation in cetaceans

Answer 3

Reticulocyte count, WBC count, differential count, ESR, plasma fibrinogen, serum albumin, serum globulin, ALP (low), serum iron (low); Hb and PCV decrease without blood loss anemia based on reticulocyte count (F8)

Question 4

Most reliable indicator of inflammatory disease in cetaceans

Answer 4

Plasma fibrinogen (photo-optical test), elevation as little as 20% above high normal important. (F8)

Question 5

Which blood tubes are needed for clotting profiles, whole blood sedimentation, plasma fibrinogen in cetaceans?

Answer 5

Orange (thrombin) tube induces clotting, light blue (sodium citrate) tube coag assays, fibrinogen; Potassium EDTA tube for hematology and ESR; green top lithium hep tube for genomic assays. (F8)

Question 6

Collection and retention of marine mammal tissues is regulated by what agency?

Describe your initial approach to a large whale necropsy.

Describe the classification of carcass condition.

Answer 6

Necropsy

• Collection and retention of marine mammal tissues regulated by Marine Mammal Health and Stranding Response Program of National Oceanic and Atmospheric Administration (NOAA)
• Helpful hint for moving larger animals: inject compress air into the abdomen to refloat the animal
• First step in necropsy exam: carcass code (see table 13.1 below)
  
  • Large whales: postmortem gas may cause extrusion of tongue, rectum, umbilicus, penis, vagina, uterus; use small incision to gently release gas

Question 7

Describe the technique for post-mortem marine mammal necropsy.

Describe the technique for extracting ears. Why is this important?

What does blubber condition tell you about the animal?

Answer 7

• Dissection tips
  
  • Reflect blubber layer first, create a checkerboard-like pattern
  • Incise abdominal wall along costal arch and dorsally to the level of rectum
  • Thoracic cavity – transect costochondral junctions, incise intercostal muscles, releases/removes individual ribs
    
    • To avoid transecting the ribs in some animals- go through diaphragm
  • Heart/Lungs of larger animals – create a window in the intercostal muscles
  • Can also create 1 x 3 m windows in abdomen for sampling in larger whales
    
    • Good expedited method for tissue collection (harmful algal blooms, infectious disease, histopath)
  • Extracting ears (important for when acoustic is suspected as the cause of stranding in cetaceans, pinnipeds) – ultrastructural evaluation
    
    • Collect ASAP after death because cochlear hair cells sensitive to autolysis
    • Organ of Corti + associated innervations: shown to have structural alterations in terrestrial mammals that undergo auditory trauma
    • Ideally collect within 5 hours post-mortem, but can still collect up to 30 hours
    • Extracting tympanoperiotic complex: 1) separate periotic from tympanic bone 2) remove stapes with tissue forceps 3) perforate round and oval window 4) perfuse fixative through opening
• Blubber
  
  • Focal blubber hematoma and edema – indicative of blunt trauma
  • Animals with robust body condition ooze oil when incising blubber
  • Suboptimal or catabolic body condition: laminar pale to dark red discoloration of basal ½-1/3 of the blubber layer because increase vascular perfusion and mobilization of fat
  • Blubber thickness measured from epidermal-dermal junction to base of blubber

Question 8

What samples or measurements need to be collected for necropsy of a stranded cetacean?

What additional tissues should be evaluated for examination of a fetal cetacean?

WHat tissues should be sampled for organic pollutants?

What about element evaluation (heavy metals)?

What about polycyclic aromatic hydrocarbons?

Answer 8

• Sample collection
  
  • Vitreous humor & blood collection for analyzing CK and K⁺: increased values indicate degree of muscle/renal injury
    
    • Increase in S100 B in vitreous humor: entangled northern fur seals, important information for drowning cases
  • Stomach contents:
    
    • Fish otoliths used to identify prey (buffered neutral formalin may dissolve otoliths so don’t use formalin to fix stomach contents of fish eating animals)
    • Plant-eating animals – do not freeze contents because fragmentation of sea grass and algae will occur
  • Age: estimated from teeth and bone
    
    • Teeth growth layers – odontocetes, pinnipeds; composed of dentine in odontocetes; dentine and cement for pinnipeds
      
      • Use canines for dentinal counts unless pulp cavity occluded (then use cement in post-canines)
      • Can identify if tetracycline has been administered (binds to calcium) – visible light (yellow-brown coloration), fluorescent light (yellow-gold)
    • Tympanoperiotic bones have annual growth layers in periosteal bone – sirenians and mysticetes
    • Balaenopterid whales (blue whale, fin whale, common minke whale)– ear plugs
• Fetus examination:
  
  • amniotic & allantoic fluids, fetal skin folds (cetacean; number and orientation)
  • number & location of rostral hairs
  • tooth eruption
  • blubber thickness
  • eyelids (open/closed, extent of eye development)
  • umbilicus examination
  • patency of umbilical veins and artery
  • ductus arteriosis & foramen ovale
• Contaminants:
  
  • Analyze lipophilic tissues (blubber, milk, blood, liver) for organic pollutants
  • Analyze kidney, liver, blood, epidermis for element evaluation
  • Polycyclic aromatic hydrocarbons: rapidly metabolized in marine mammals; serum, bile, liver can be analyzed in acute cases
• Mysticetes
  
  • Record pleat lengths and proximity to umbilicus
  • Plate arcades: note number, color, maximal length
  • Collect intact plate and earplugs for isotope analyzing

Question 9

Gas bubble disease and ship strikes are some of the commonly described causes of anthropogenic trauma in marine mammals.

What findings would be seen on necropsy?

What family of whales are commonly affected by gas bubble disease?

Answer 9

• Gas bubble disease (intravascular gas emboli, multisystemic hemorrhage)
  
  • Particularly described in beaked whales (Ziphiidae), associated with development of military/industrial sonar
  • Look for gas bubbles in renal capsule; mesenteric, epicardial, pulmonary vasculature
  • Can also cause auditory trauma
  • Acute form: gas bubbles seen in major organs and blood vessels; systemic – brain, spinal cord, mesenteric veins, coronary vasculature, renal capsule, liver
  • Chronic form: large gas-filled spherical cavities, typically seen in liver, kidney, spleen, gastric wall; reported bone lesions in sperm whales (dysbaric osteonecrosis)
  • Gas scoring: use subcutaneous, mesenteric or coronary veins, lumbocaudal venous plexus
    
    • Different scoring systems
    • Grade I: few bubbles; Grade II: abundant bubbles
    • Grade 0: none; Grade I: subcapsular gas in 1-2 organs; Grade II: gas widely distributed throughout body
  • Sample gas ideally within 12 hours
    
    • Nitrogen supersaturation: N₂ approximately 70%, CO₂ approximately 30% = decompression sickness
  • Fat emboli accompany severe and acute GBD; most readily seen in lung tissue; Oil-Red-O stain on histopath to diagnose
• Ship strike muscle findings: hemorrhage, edema; flocculent, discoid, granular or hyalinized myocellular degeneration with contraction bands