Proboscidea - Elephants Flashcards

Question 1

Q

Describe the treatment of digital osteitis in African Elephants using regional limb perfusion.

What are the predisposing factors for elephants developing septic arthritis and osteitis?

Discuss acute v chronic osteitis.

Answer

A

Case Description:

In this case, initial tx included corrective trimming, cryotherapy to remove exuberant soft tissue, soaking in Epsom salts, flushing with antiseptics, oral potentiated sulfonamide abx. Reoccurred, tried same thing but became severely anemic on the sulfonamide.
Trained for application of tourniquet (motor cycle inner tube), IVRP. Did for 70 treatments over 6 months, also flushed with metronidazole.
Ceftiofur – broad spectrum third gen cephalosporin, time dependent, active metabolites have strong affinity for proteins. Higher initial concentration, longer drug will stay around.

Predisposing factors include malnutrition, lack of exercise, obesity, pre-existing disease, old age, improper enclosure surface, wet and dirty conditions, inappropriate foot trimming. Early and regular rads should be routine for any foot care program.

Acute V Chronic Osteitis

Chronic – numerous inflammatory cells, release of cytokines stimulate osteoclastic bone resorption and ingrown fibrous tissue – radiolucent lesions.
- Radiolucent lesions not always indicative of infection.
- Difficult to manage medically because of vascular thrombosis, ischemia, tissue necrosis, reduced activity of antibiotics.
- Surgical management usually necessary to debride necrotic tissue.
Acute – abx may be sufficient for infection control.

Reference: Dutton, CJ et al. SUCCESSFUL TREATMENT OF DIGITAL OSTEITIS BY INTRAVENOUS REGIONAL PERFUSION OF CEFTIOFUR IN AN AFRICAN ELEPHANT (LOXODONTA AFRICANA). JZWM 48.2 (2017): 554-558.

Question 2

Q

Discuss the particle size of elephant aerosols and the impact this may have on Mycobacterium transmission.

Answer

A

STUDY:

Cultured trunk wash samples; based on sequencing results, biochemical profile for a proxy bacteria (Rothia spp) was established. Similar thing for expired air samples.
It was found that elephants are capable of producing aerosolized bacterial particles of a size small enough to remain airborne for prolonged periods and penetrate the lower regions of the human respiratory tract.

Mycobacterium Transmission:

Result of close, freq, prolonged contact with shedding elephant.

Inhalation of aerosolized droplet nuclei.
Several risk factors for contracting TB from shedding elephant:
- Close or frequent contact.
- Use of high-pressure hosing when cleaning.
- Necropsy of elephant with TB.
- Close contact does not appear necessary in all cases.
High risk for transmission for transmission of aerosolized bacteria from elephants – breathing, vocalizing, blowing from trunk.

Reference: Burke, Sophie M., et al. “DETECTION OF AEROSOLIZED BACTERIA IN EXPIRED AIR SAMPLES FROM ASIAN ELEPHANTS (ELEPHAS MAXIMUS).” Journal of Zoo and Wildlife Medicine 48.2 (2017): 431-439.

Question 3

Q

Discuss the use of the iStat portable clinical analyzer in Elephant samples.

Answer

A

iStat overall acceptable in elephants for iCa, pH, glucose, Na, HCO3, TCO2, PCO2.

Differences small at 10 min and 4 hours post sample collection except K+.
Release of K from intracellular stores may cause false increases with time.
Poor correlation present for K between analyzers.
iStat should provide acceptable measures of iCa and blood gas analytes.

Discussion points:

Other study showed PCO2 decreases and pH increases after 15 min lateral recumbency.
Values that fall outside of the reference range using the iSTAT would warrant re-evaluation on a benchtop analyzer prior to taking clinical action.
Treatment of hypocalcemia-related dystocia warranted if iCa < 1.2 mmol/L

Tarbert, Danielle K., Behling-Kelly E. Priest H, Childs-Sanford S. EVALUATION OF THE I-STAT PORTABLE CLINICAL ANALYZER FOR MEASUREMENT OF IONIZED CALCIUM AND SELECTED BLOOD CHEMISTRY VALUES IN ASIAN ELEPHANTS (ELEPHAS MAXIMUS). Journal of Zoo and Wildlife Medicine 48.2 (2017): 319-327.

Question 4

Q

Discuss the use of the lancet and swab technique in EEHV testing of Elephants.

Is this technique superior to others? What benefits are there to using this technique?

Answer

A

EEHV Background:

EEHV-HD most common in Asian elephants < 8yo, peak 1-4 yrs. Sudden onset, rapid death. Tx aggressive supportive care and antivirals.
Viremia detectable days to weeks prior to clinical signs, routine monitoring weekly is recommended for all institutions holding Asian elephant calves.

STUDY:

Study aim – evaluate blood sampling using lancing device, assess efficacy of commercially available sampling matrices (filter papers, swabs) for EEHV diagnosis.
Conclusion: Both foam and flocked swabs present higher analytical sensitivity for detection of EEHV1 compared to Whatman FTA cards, filter papers, or cotton-tipped swabs. Not significantly different vs EDTA blood tube.
Lancet and swab technique satisfactory in detection of both subclinical and clinical EEHV1 viremia. Suitable alternative for EEHV screening when venipuncture is not feasible.
Disadvantage: small amount of blood produced through lancing adversely affects detection sensitivity of EEHV viremia. Insufficient for ancillary testing other than platelet and WBC smear.

REFERENCE: Lopez, Javier, et al. ASSESSMENT OF A LANCET-AND-SWAB BLOOD SAMPLING TECHNIQUE FOR SURVEILLANCE OF ELEPHANT ENDOTHELIOTROPIC HERPESVIRUS INFECTION. Journal of Zoo and Wildlife Medicine 48.3 (2017): 659-667.

Question 5

Q

Discuss the use of composite materials in repair elephant tusk fractures.

Answer

A

TAKE HOME: The composite cap and wrap both allowed rads for evaluation of pulp canal after placement.

Tusk Repair Background:

Current recommended therapies for pupitis include pulpotomy, pulpectomy, extraction.
Conservative therapy – abx, flushing.
Metal caps, circumferential bands prevent wear and stabilize fractures.
Heavy, easily removed, not able to see on rads.
Composite material – matrix and reinforcement combination. Durable, light, rads.

Elephants in This Case Series:

Elephant #1: Fabricated composite cap using fiberglass and Kevlar cloths layered and laminated with epoxy resin. After 4mos wore through, replaced with a thicker cap. Came off after 15 mos. Eventually fractured the other tusk and made a similar cap.
Elephant #2: Used composite band of seven layers of material to stabilize a longitudinal tusk crack. Carbon fiber materials, braided fiberglass, polyurethane resin. Evidence of infection/foul odor, started cleaning with chlorohex, cultured pseudomonas. 6 mos later extraction of the tusk was attempted but failed. Transected at drainage hole to facilitate conservative therapy. In theory could place wrap to stabilize fracture, let it grow out and then trim. Didn’t do in this case.

Sim, R. et al. USE OF COMPOSITE MATERIALS AS A COMPONENT OF TUSK FRACTURE MANAGEMENT IN AN ASIAN ELEPHANT (ELEPHAS MAXIMUS) AND AN AFRICAN ELEPHANT (LOXODONTA AFRICANA). Journal of Zoo and Wildlife Medicine 48.3 (2017): 891-896.

Question 6

Q

Discuss the normal cyclicity and breeding recommendations of the Asian elephant.

Answer

A

Female elephants should conceive as soon as cycling regularly. Acyclicity and repro tract pathology when females repeatedly exposed to repro hormones without being bred.

Current recommendations are to breed females before age 24y, but typically 6-15 years.
656 day mean gestation length comparable to other Asian herds.
4-6 yr intercalf interval appears typical of Asian elephants.
Large uterine blood clots, normal following calving, may block implantation for up to 2 years after birth.

Kiso, Wendy K., et al. REPRODUCTIVE PARAMETERS AND BIRTH STATISTICS FOR A HERD OF ASIAN ELEPHANTS (ELEPHAS MAXIMUS) IN NORTH AMERICA OVER A 20-YEAR PERIOD. Journal of Zoo and Wildlife Medicine 48.4 (2017): 987-996.

Question 7

Q

What are the periparturient events in Elephus maximus?

Answer

A

Periparturient events fell within previously described ranges.

P4 dropping to baseline 3-10 d before birth.
Most births at night.
Loss of mucosal plug may signal imminent birth but was rarely observed.
Delays greater than 24h between membrane rupture and delivery have been associated with increased calf mortality.
Appearance of tail bulge occurred within 10 min of birth in most cases.
Prolonged labors may result from oversized calves, twinning, calf malposition, malpresentation or malformation, uterine inertia, hypocalcemia, maternal obesity.
Elephants may have contractions for several days to weeks before birth, can discontinue labor for long periods of time.

Kiso, Wendy K., et al. REPRODUCTIVE PARAMETERS AND BIRTH STATISTICS FOR A HERD OF ASIAN ELEPHANTS (ELEPHAS MAXIMUS) IN NORTH AMERICA OVER A 20-YEAR PERIOD. Journal of Zoo and Wildlife Medicine 48.4 (2017): 987-996.

Question 8

Q

What are the three stages of labor described in elephants?

Answer

A

Three stages of labor described in elephants – different than other hoofstock!

Stage 1 – fetus already positioned dorsosacrally without cervical dilation.
Stage 2 – dilation of the cervix, fetal entry into birth canal, appearance of bulge under the tail, rupture of fetal membranes.
Stage 3 – expulsion of the placenta, may take up to 10 hours.

Rectal massage triggers Ferguson’s reflex, initiates release of endogenous oxytocin. Requires confirmation of appropriate calf positioning and full cervical dilation.

Most passed placenta within 4 hrs, similar to other herds.

Kiso, Wendy K., et al. REPRODUCTIVE PARAMETERS AND BIRTH STATISTICS FOR A HERD OF ASIAN ELEPHANTS (ELEPHAS MAXIMUS) IN NORTH AMERICA OVER A 20-YEAR PERIOD. Journal of Zoo and Wildlife Medicine 48.4 (2017): 987-996.

Question 9

Q

What risk factors are associated with decreased neonatal survival in Elephus maximus?

Answer

A

older cows, primiparous mothers, longer labors.

Kiso, Wendy K., et al. REPRODUCTIVE PARAMETERS AND BIRTH STATISTICS FOR A HERD OF ASIAN ELEPHANTS (ELEPHAS MAXIMUS) IN NORTH AMERICA OVER A 20-YEAR PERIOD. Journal of Zoo and Wildlife Medicine 48.4 (2017): 987-996.

Question 10

Q

Describe the neonatal exam of an Asian elephant.

What findings are normal?

What findings may be concerning?

What vital parameters differ from adults?

What clinical pathology differences exist between neonatal and adult elephants?

Answer

A

Normal:

Raspy thoracic auscultation at birth common, resolved in most after walking.
Normal calves sternal within 2 min, stood with assistance 6 min, walked within 2 hrs.
Vocalization, ear flapping, moving of trunk shortly after birth

Abnormal:

Mean weight of abnormal calves higher than normal calves.
Heart murmurs in newborn elephants do not appear to be normal and have been associated with EEHV and congenital abnormalities. However, a heart murmur in this case resolved spontaneously, may not always be clinically significant.
Elephants that fail to stand within 24h of birth typically poor prognosis.
Clinicians should check mouths of calves at birth for teeth and suckle reflex. Absence or loss within first 24h after birth warrants evaluation.
Umbilical abnormalities – short umbilical cord of elephants, sudden rupture between cord and placenta results in bleeding and skin tears. Evaluate umbilicus for several days after birth.

Vital Parameters:

Neonatal HR higher than adults 100-150 HR v 25-35 HR
Neonatal RR higher vs adults (45-70 RR vs 10-12 RR).
Rectal temps similar to adults, did not differ between normal and abnormal calves.

Clinical Pathology:

increased RBC count, Hct, Hbg in calves.
Heterophils predominate vs monocytes in adults.
Platelets lower in neonates.
High ALP may reflect osteoblastic activity and intestinal development. Caution in overinterpretation of liver enzymes in newborn elephants.
High serum bilirubin likely normal in calves.
Bilirubinuria found in calves, assoc with NI and sepsis in foals but neither was present in any of the calves.

Wiedner, Ellen, et al. VITAL SIGNS AND FIRST OCCURRENCES IN NORMAL AND ABNORMAL NEWBORN ASIAN ELEPHANT (ELEPHAS MAXIMUS) CALVES. Journal of Zoo and Wildlife Medicine 48.4 (2017): 997-1015.

Question 11

Q

What important behavioral milestones occur in newborn elephant calves and what time do they typically occur?

Answer

A

Wiedner, Ellen, et al. VITAL SIGNS AND FIRST OCCURRENCES IN NORMAL AND ABNORMAL NEWBORN ASIAN ELEPHANT (ELEPHAS MAXIMUS) CALVES. Journal of Zoo and Wildlife Medicine 48.4 (2017): 997-1015.

Question 12

Q

Discuss thromboelastrography in healthy Asian elephants.

What do the various aspects of the trace represent?

How does sample storage affect results?

How do elephants compare to other hoofstock species?

Answer

A

TEG

TEG – thromboelastography; complete analysis of in vivo hemostasis because both cellular and plasma components of the blood are present.
PT/aPTT not consistently predictive of bleeding risk, does not account for hemostatic contribution of cellular elements (platelets, leukocytes, erythrocytes).

TEG parameters: Results presented as a trace and quantitative analysis.

SP – split point, time for initial clot to form.
R – reaction time, time from analysis start to when the TEG reaches ampliture of 2mm, represents clotting factor initiation of hemostasis resulting in thrombin generation.
- This is what PT, aPTT measure.
K – clotting time, time for clot formation measured as time R until amplitude of 20 mm.
Alpha angle – together with K time represents speed of clot formation.
- Cross-linking of fibrin.
MA – maximum amplitude, measures maximal platelet-fibrin interaction in the clot, represents maximal clot strength.
G – calculated measure from amplitude giving indication of the shear elastic modulus strength.

Storage:

All TEG parameters were significantly different after 24h storage vs fresh samples at 60 minutes. Reduced SP, R, K in the 24h sample.
Delayed analysis of whole blood not recommended – hypercoagulable TEG.
Plasma samples may be frozen and stored to allow retrospective analysis.

Comparison: similar to pigs, hypercoagulable to horses

larger maximum amplitude of trace (similar to pig)
steeper alpha angle (similar to pigs, rats)
generally slower reaction times, quicker coagulation times.
R and K similar to pigs.
Hypercoagulable vs horses.

Perrin, KL, Krogh, AK, Kjelgaard-Hansen, Bertelsen MF et al. THROMBOELASTOGRAPHY IN THE HEALTHY ASIAN ELEPHANT (ELEPHAS MAXIMUS): REFERENCE INTERVALS AND EFFECTS OF STORAGE. Journal of Zoo and Wildlife Medicine 2018 49(1): 54-63.

Question 13

Q

Coinfection with what bacterial pathogen has been documented in wild Asian elephants infected with EEHV?

Answer

A

EEHV1a & EEHV4 with Clostridium perfringens - α, β, and ϵ toxins.

Boonsri K, Somgird C, Noinafai P et al. ELEPHANT ENDOTHELIOTROPIC HERPRESVIRUS ASSOCIATED WITH CLOSTRIDIUM PERFRINGENS INFECTION IN TWO ASIAN ELEPHANT (ELEPHAS MAXIMUS) CALVES. Journal of Zoo and Wildlife Medicine 2018 49(1): 178-182

Question 14

Q

Discuss the corrective shoeing of elephant with unequal leg lengths.

Answer

A

STUDY: The current version is made of two types of shoe rubber, glued together and attached to the pad of the shorter leg with a liquid adhesive. The first elephant also has bilateral wedge pads to offload pressure from the fourth nails. The shoes are removed each month for foot care, then replaced.

Additional considerations:

Training elephants to stand still for shoe application.
Staff time, material costs.
Monthly removal for foot trimming is necessary because shoes preclude natural wear of feet and nails.
Shoes should be rinsed if muddy or wet.
Expect to spend several months building prototypes and trying different shoes before settling on final products.

Johnson G, Smith J, Peddie J, Peddie L, DeMarco J, Wiedner E. USE OF GLUE-ON SHOES TO IMPROVE CONFORMATIONAL ABNORMALITIES IN TWO ASIAN ELEPHANTS (ELEPHAS MAXIMUS). Journal of Zoo and Wildlife Medicine 2018 49(1): 183-188

Question 15

Q

What effects did administration of degralex acetate have on musth and serum testosterone concentration in Asian elephant bulls?

What is the mechanism of action for this drug?

What are normal signs of musth?

Answer

A

Goal of study: Assess effect of degarelix acetate administration on suppression of musth and temporal gland secretion and changes in serum testosterone in male Asian elephants.

Safety of DA was demonstrated for SQ administration.
Tx effective in suppressing musth and temporal gland secresion completely in bull 1, partially in bull 2, and postponing musth for 4-8 mos in all three musth cycles tested.
High circulating testosterone significantly reduced on day following administration.
Limitations of use – relatively difficult administration, high cost. Failure of complete cessation of musth on following day and recurrent transient musth in one bull in this study.
Could be individual variation or dose insufficiency.

Degarelix acetate – potent synthetic long-acting GnRH antagonist.

Results in reduction in LH and testosterone synthesis.
Formation of a gel depot of the drug after SQ administration.
Rapid suppression of testosterone in rats, humans, goats.

Musth

Natural feature of life cycle in males, associated with heightened aggressive and sexual behavior, temporal gland swelling and secretions, urine dribbling, elevated androgen production.
May result in injury or death of handlers, dangers to other animals.
There is a relationship between aggressive behavior and elevated androgen levels in musth.

Pathirana IN, Rajapaksa C, et al. EFFECTS OF GONADOTROPIN-RELEASING HORMONE ANTAGONIST DEGRALEX ON MUSTH AND SERUM TESTOSTERONE CONCENTRATIONS IN ASIAN ELEPHANTS (ELAPHAS MAXIMUS). Journal of Zoo and Wildlife Medicine 2018 49(3): 779-783.

Question 16

Q

What drugs are commonly used to treat Elephant TB?

Answer

A

Isoniazid

Rifampin

Pyrazinamide

Ethambutol

(Fluoroquinolones)

Question 17

Q

What drug side effects are common while treating TB in elephants?

What additional issues exist with administration of drugs for treating TB in elephants?

Answer

A

Reported effects in elephants – inappetence, lethargy, epiphora, blepharitis, hepatitis, diarrhea, anemia, pica, leukopenia, trunk paralysis, elevated LDH, stiff/sore limbs

Most are administered orally – dosing becomes difficult when clinical signs occur – rifampin does not read appropriate levels rectally and ethambutol causes mucosal irritation and expulsion when given rectally

Question 18

Q

What is the most common site of foot disease in Asian elephants?

Answer

A

Lateral nail (N5) of both front feet - due to the peak pressure of an elephant’s foot along lateral digits

Question 19

Q

What is the most common abnormality of captive elephant feet?

Answer

A

Nail abnormalities - 67-80% of lesions

Question 20

Q

What is the relationship between bone mineral density and serum Ca & Phos levels?

Answer

A

Negatively correlated

No similar correlation with ALP due to its osteoblastic activity in bone formation and activity in other tissues

Question 21

Q

How did age and sex affect bone mineral density in Asian elephants?

Answer

A

BMD lower in females

Decreased with age

Tuskless males less than tusked males

Question 22

Q

What relationship has been documented between diet and season in Asian elephant vitamin D physiology?

Answer

A

No serum or diet parameters were affected by time or season.

25(OH)D3 levels were nondetectable in all samples despite supplementation of the diet with recommended levels of vitamin D3, and UV exposure was at sufficient levels for cutaneous vitamin D synthesis for 6 mo of the year.

Elephants may not be able to utilize dietary cholecalciferol and appear to not rely on cutaneous vitamin D synthesis

Question 23

Q

What species can only get vitamin D from the diet?

What species synthesize vitamin D from UV light?

What species cannot synthesize it from UV light?

Answer

A

Species that only get D from diet – carnivores (dog, cat, polar bear)

Species that synthesize it from UV – cattle, goats, sheep, alpacas

Species that don’t synthesize it from UV - horses

Question 24

Q

Discuss the unique opthalmic anatomy of the elephant.

Answer

A

No lacrimal puncta, lacrimal glands, or nasolacrimal ducts - this is what casues tears (produced in adnexal glands) to run down their face.

Question 25

Q

Discuss the unique dental anatomy of elephants.

What type of teeth are the tusks?

What type of dentition are elephant cheek teeth?

How are teeth replaced?

Answer

A

Tusks - modified incisors - have enamel when they erupt which is quickly shed once it erupts which exposes the dentine (ivory) - can grow 18 cm/year up to 3 feet long and 100 kg in weight

Cheek teeth - Dental formula I 1/0, PM 3/3, M 3/3 - new molars formed in alveolar pockets at back of jaw - composed of compressed plates of enamel-wrapped dentine joined by cemetnum - as teeth push forward, they dissolve their roots and break off

Six sets of molars throughout life

· Dental formula – I 1/0, C 0/0, PM 3/3, M 3/3.

· Tusks extension of maxillary incisors, prominent in male and female Africans.

· Asian bulls also have tusks.

· Asian cows lack tusks but have small vestigial structures (tushes).

· Elephants will have 24 cheek teeth (PM and M) over course of lifespan.

Shed in sections, replaced by next tooth pushing forward from behind

Question 26

Q

Describe the cardiovascular anatomy of elephants.

How many vena cava do elephants have?

Why is catheter placement sometimes difficult in elephants?

Answer

A

Two cranial, one caudal (Fowler 8)

Cardiovascular system

Two cranial vena cava, single caudal vena cava

Numerous arteriovenous anastomoses

Veins have valves which make venipuncture and catheter placement challenging but assist in hydraulics

Question 27

Q

Describe the unique anatomy of the elphant thorax.

Answer

A

Pleura consists of thick connective tissue that connects lungs to ribs and diaphragm. Still slides and acts as a pleural membrane. (Fowler 8)

· Lungs (ZP).

· Thick visceral pleura (lung capsule) continuous with intraseptal network of elastic tissue, supports pulmonary parenchyma.

· Lack cartilage except proximal intrapulmonary primary bronchi.

· Thorax lacks pleural space.

· Pulmonary visceral pleura connected to parietal pleural surfaces of the thorax, attaches lungs to all surfaces of inner thorax and diaphragm.

· Collagenous connective tissue with fluid pockets and some capillaries.

Question 28

Q

Describe the unique elements of the elephant biliary system.

Answer

A

No gallbladder. Produce bile alcohols rather than bile acids - predisposes them to cholelith formation. (Fowler 8)

Question 29

Q

Describe the anatomy of the elephant foot.

Answer

A

Five digits

Metacarpals arranged vertically with equal weight distrubtion

Metatarsals arranged horizontally - most weight on 2,3,4

Bones sit on digital cushion made of fat and connective tissue - expands when compressed pushing blood back up to heart

Cartilagionous prepollux and prehalux creates subcompartments within the cushion

Slipper - sole (underside of toes) and pad (underside of foot) - keratimized material grows from undelrying germinal epithelium - 4-12 mm thick, smooth in juveniles, corregate in adults

Question 30

Q

What is an ankus?

Answer

A

The bullhook - used for guiding elephants in free contact situations. (Fowler 8).

Question 31

Q

What is the most common complication with etorphine anesthesia in wild African elephants?

Answer

A

Pink foam syndrome - pulmonary hypertension (Fowler 8)

Question 32

Q

What are common complications of anesthesia in Elephants?

Answer

A

Tusk & bone fractures

Respiratory & lactic acidosis

Apnea

Bloat

Hypo/Hypertension

Nerve damage

cardiac arrythmias

Hypoxemia

(Fowler 8)

Question 33

Q

What is unique about the osmolarity of elephants?

Answer

A

It is very low - all fluids are hypertonic. (Folwer 8)

Question 34

Q

What mammal has the largest red blood cells?

Answer

A

Elephant (Folwer 8, Terio)

Question 35

Q

Do elephants have blood types?

Answer

A

Yes - cross match before transfusion (Fowler 8)

Question 36

Q

Describe the procedure for performing a trunk wash.

Answer

A

60 mL of sterile saline are placed in one nostril, the trunk is lifted and held there briefly, lowered and the elephant then exhales into a plastic collection bag.

Repeat the process for the other nostril.

Question 37

Q

Identify three venipuncture sites in the elephant.

Answer

A

Auricular veins are the primary site. Cephalic (proximal medial forelimb) and medial sapheous (distal medial hindlimb) can also be used with the aid of a tourniquet to raise the veins.

(Fowler 8)

Question 38

Q

What three OIE reportable diseases have been documented in both species of elephants?

What are the etiologic agents?

Answer

A

Foot and Mouth Disease - Apthovirus - Picornaviridae

Tuberculosis - Mycobacterium tuberculosis

Anthrax - Bacillus anthracis (primarily a wild disease)

Terio ZP

Question 39

Q

What clinical signs have been observed in elephants with foot and mouth disease?

Answer

A

Anorexia, vesicles on mucosal surfaces, trunk exudate, swelling of feet, sloughing of nails and slipper.

Young elephants can have systemic infection with lymphocytis myocarditis and erosive enteritis

(Fowler 8, Terio ZP)

Question 40

Q

Two differentials for vesicular disease in elephants.

Answer

A

Foot and mouth disease. Elephantpox.

(Fowler 8)

Question 41

Q

What is floppy trunk syndrome in elephants? Is there a suggested cause of the disease?

Answer

A

Flaccid trunk paralysis - seen in wild African elephants in Zimbabwe - thought to be due to Heliotropium ovalifolium or lead toxicity.

Question 42

Q

What factors predispose managed elephants to foot disease?

Answer

A

Neglect of regular nail and sole trimming and care

Prolonged exposure to wet or soiled environments

Artificial substrates

Sedentary lifestyle

Conformational defects

Trauma

(Terio ZP)

Question 43

Q

What factors predispose elephants to arthritis?

Answer

A

Multifactorial – age-related, mechanical trauma, repetitive loading stress due to hard environmental substrates lack of appropriate exercise, excessive body weight, poor confirmation.

More common and severe in forelimbs.

(Terio ZP)

Question 44

Q

What noninfectious diseases of the heart are documented in elephants?

Are there any predisposing factors?

Answer

A

Arteriosclerosis & atherosclerosis - cause is unknown

Possible contributing factors – mechanical stress in affected vessels, medial anoxia, high serum calcium, advancing age.
No correlation with serum lipid levels.
Aorta, coronary arteries, aortic branch arteries most commonly affected.
Thickened, rough, discolored endothelial surface with discrete, white, firm, raised plaques.

Question 45

Q

Identify two degenerative renal conditions of managed elephants. Are there potential causes or unique findings on necropsy?

Answer

A

Chronic interstitial nephritis.

Aged captive elephants, presumed degenerative/age-related.
Red/brown to pale grossly with rough, granular, or slightly pitted subcapsular cortical contours.
Cortical parenchyma may contain pale cortical to medullary streaking or clear fluid-filled cystic structures.
Tubules in affected areas decreased in number, atrophic, som may be cystic.

Renal medullary/papillary necrosis.

NSAIDS.
Other causes – renal hypoxia, dehydration, pyelonephritis, chronic urinary tract obstruction.
Gross – thin yellow/white streaks in medulla to radiating areas of necrosis or infarction in medullary tissue.

Question 46

Q

What are the most likely causes of this lesion found on necropsy of an elephant?

Answer

A

Gastric ulcers.

Possibly related to NSAIDS, stress, combinations of NSAIDS and steroids, endogenous glucocorticoid release, infectious agents, chronic renal disease.

Terio ZP

Question 47

Q

What is the most common neoplasm of the elephant? Is there a particularly affected demographic?

Answer

A

Myometrial leiomyomas.

Nulliparous, ages, Asian elephants.

(Terio ZP)

Question 48

Q

What is the most significnat infectious disease threat to Elephants?

What age groups of elephants are most susceptible to this disease?

How prevalent is this diease?

Answer

A

EEHV Impact and Epidemiology

All adults carry and shed one or more strain
Adults that survived exposure and viremia are now latent carriers
Exposure is natural but Asian elephants between 1-8 yrs are at high risk of developing infection (most commonly due to EEHV1)
Asian elephants under human care in NA, Europe, and Asia have succumbed
Deaths also reported in wild Asian elephants

Asian Elephants

Cause of death in 53% of all Asian elephants born in NA since 1980
May 2017, 35 cases of EEHV in Asian elephants born in NA since 1980 – ¼ (25%) developed disease
- 11 elephants survived and 24 died (overall NA mortality of 68%)
In Europe, > 200 born since 1995, 26 died (60% of deaths) from EEHV
- Greatest cause of death of elephants born in NA and Europe
First published PCR positive case in 2006 in Cambodia
- Since, fatal EEHV1 in Laos, EEVH1 and EEHV4 in Thailand, & 9 EEHV1 in India
Healthy Asian elephants in India shed EEHV1, EEHV4, and EEHV5 from trunks

African Elephants

EEHV impact unknown
Multiple EEHVs have been identified on necropsy from asymptomatic elephants
Very sporadic fatalities exist (EEHV2 in 11mo male and 13yr female, 10yr with EEHV6)
Two elephants survived (15mo – EEHV6 & 5yr – EEHV3B)
Two NA elephants with asymptomatic shedding of EEHV6 and EEHV3 in trunk washes
More recent mortalities - elephants under 15 years appear susceptible

Question 49

Q

What demographic of elephants is most susceptible to EEHV?

Answer

A

Asian elephants ages 1-8

Also documented in african elephant calves - under 15

Question 50

Q

Describe the pathogenesis of EEHV infections.

Answer

A

Endotheliotropic.

Vascular compromise results in hemorrhage, edema, coagulopathy.
Hypocoagulability, DIC.
Most types primarily affect capillary endothelium, EEHV3 may damage larger vessels.
Death most likely due to hypovolemic shock due to cardiac and circulatory collapse.
Terio ZP

Question 51

Q

Describe the lesions associated with EEHV that could be found on necropsy.

Question 52

Q

What are the EEHV strains?

Which strains are endemic in each species?

Which strains have produced hermorrhgic disease?

What are the classic presentations of the various strains?

Answer

A

Asian Elephants - EEHV1a, EEHV1b, 4, 5 - most HD cases are due to 1a (1b, 3, 4, 5 also documented)

African Elephants - EEHV 2, 3, 6, 7 - HD cases reported in

Cutaneous papillomas - EEHV 2, 3, 6, 7

Question 53

Q

Describe the clinical signs, pathogenesis, and lesions of elephant pox.

What inclusion bodies are observed?

Answer

A

Elephantpoxvirus - Orthopoxvirus - Poxviridae

Infected rodents may be definitive hosts - but they are definitely reservoirs

Transmission via direct contact, rodents, or fomites

CS vary from mild cutaneous infection to fatal systemic illness

Lesions often on trunk, temples, perineum, vulva, can develop on conjunctiva, tongue, oral cavity, pharynx, esophagus, trachea, intestinal mucosa, pleural and peritoneal linings.

Skin – multiple 1-2 cm diameter, coalescing, gray-yellow papules with central reddening that progress to vesicles, pustules, ulcerations.

Ulcers common on MM. Toenail and slipper corneum may be affected, necrosis, sloughing.

Intracytoplasmic eosinophilic inclusions.

Question 54

Q

What differential diagnoses should be excluded in a potential EEHV case?

Answer

A

Bacterial sepsis, other systemic infections, coagulopathies i.e. toxins, autoimmune dz, parasitic infection. Encephalomyocarditis virus EMCV important rule out due to cardiovascular lesions in both infections.

Terio ZP

Question 55

Q

What type of virus is encephalomyocarditis virus?

What are the clinical signs of infection in elephants?

How is this disease transmitted?

What are teh typical lesions on necropsy?

What is this disease a primary differential for?

Answer

A

Encephalomyocarditis virus EMCV

ssRNA, Picornavirus.
Captive and wile, Asian and African.
Sudden death without promontory signs.
Encephalitis, myocarditis.
Rodents primary host, reservoir.
Infected carcasses and feces.
Principally cardiotropic, acute myocarditis with cardiac dysfunction leading to death.
Gross – Pericardial and/or peritoneal transudate effusion, epicardial and myocardial hemorrhage, multifocal myocardial pallor, multicentric mucosal and serosal petechial hemorrhage.
Primary differential for EEHV.
Dx requires ID of EMCV via PCR or VI in tissues with necrotizing myocarditis and related lesions.

Question 56

Q

How is elephant TB transmitted?

Which tuberculous organisms have been isolate?

Which species is more commonly affected?

What lesions are found on necropsy?

Answer

A

Transmission via aerosolized particles - infections have occurred both ways between people and elephants.

Mostly M. tuberculosis, M. bovis and others rarely isolated.

Asian elephants are more commonly affected but both can be

Gross – Lung, thoracic LN variably sized coalescing, firm, yellow-white nodular granulomas. Also in trachea. Soft casseous, cavitated, or hard mineralized centers.

Terio ZP

Question 57

Q

What two bacterial pathogens have been implicated in Elephant enteritis?

Answer

A

Salmonella & Clostridium

Terio ZP

Question 58

Q

You come across a deceased wild elephant with a lack of rigor mortis and hemorrhagic fluid coming out of its trunk.

What is your primary differential? What precautions need to be taken during this necropsy? How can you confirm your diagnosis?

Answer

A

Anthrax - Bacillus anthracis

Do not open carcass - will aerosolize. Draw blood - smears will show high numbers of gram positive sporulated bacteria.

Question 59

Q

Describe the tracheal anatomy and intubation technique for elephants.

Answer

A

Hypoxemia common complication of anesthesia due to pressure on the diaphragm from abdominal contents

Elephants have a large fleshy epiglottis that lacks cartilage allowing for a tight seal, but can be easily pulled forward to allow access to the glottis

Thick vocal chords occlude the entrance to the trachea ventrally, but thin out dorsally

Intubation is blind, guided by breathing – displace the epiglottis, manually guide the tube to the dorsal aspect of the glottis where the chords are thinner.

West

Question 60

Q

Describe the clinical signs and potential causes of colic in elephants.

Answer

A

Less dramatic signs than in horses – postural changes, straining, off feed, lethargy

Surgical colics – uniformly fatal due to surgical issues
Medical colics – sand/feed/fecal impactions, esophageal obstructions (choke), FB ingestion
Cholelithiasis, pancreatic malabsorption, carcinomatosis also produces GI disease

Fowler 8

Question 61

Q

What agents are commonly used in the deliberate poisonings of wild elephants?

Answer

A

Toxicities

Deliberate poisoning of wild elephants – strychnine, arsenic, other heavy metals, poison coated arrows, insecticide-contaminated well water, organophosphate-spiked watermelons
Many of these do not have antidotes or wild animals present too debiliatated

Fowler 8

Question 62

Q

Describe the reproductive anatomy of the male elephant.

Answer

A

Male reproductive tract

Testes sit caudal to kidneys, supplied by central artery that radiates smaller arteries outwardly – still maintains testicular temps at 94-97
No epididymis – ampulla is the primary site of sperm storage
Accessory sex glands – ampulla, prostate, seminal vesicle (viewd rectally on US), bulborethral gland (viewed below anus on US)
S-shaped penis elevates vulva into position prior to penetration (levator muscles)

Question 63

Q

Describe the reproductive anatomy of female elephants.

Answer

A

Female Reproductive tract

Vestibule – urogenital canal – is 1-1.4 m long, curves into the pelvis at the level of the anus
Clitoris located distally
Urethral opening and vagina found in cranial portion of urogenital canal
Two blind pouches on lateral sides of vagina
Ovaries – small
Uterus bicornuate and short bodies

Fowler 8

Question 64

Q

What is musth?

Answer

A

Musth

Complex hormonal phenomenon
Not rut – it isn’t seasonal and males don’t breed during this period
Testosterone increases 20-fold, androstenedione to testosterone ratio changes
Musth still occurs in castrated males
CS – dark liquid from temporal gland, perineal swelling, dribbling of urine, fetid odor, at height of musth male stop eating and lose weight
Young males may have a swell-smelling liquid – honey musth

Fowler 8

Answer 64

A

Estrous cycle

15-16 weeks (8-12 week luteal phase, 4-6 week follicular phase)
Progestogen levels high during luteal phase, low during follicular phase
During the follicular phase, two LH peaks occur – typically 3 weeks apart – ovulation occurs with the second peak
Female is inseminated or paired the day of that second peak

Answer 65

A

5-alpha-reduced-pregnane is the primary protestagen in elephants – but standard progesterone assays have been used because of cross-reactivity

Fowler 8

Answer 66

A

Placentation is zonary & endotheliochorial – calves are born immune competent

Fowler 8

Answer 67

A

· Erythrocytes among largest of mammals (~ 10 microns diameter).

· Erythrocyte count lower.

· Large erythrocyte size, high fibrinogen and globulin levels and low plasma albumin contribute to fast erythrocyte sedimentation rate.

· Lymphocyte – predominant circulating leukocyte.

· Asian elephants relatively high monocytes normally.

· Segmented monocytes 4x more than monocytes without segmentation.

· Heterophils.

Answer 68

A

· Thick skin; dermis has abundant dense dermal collagen.

· Have sepaceous glands, sweat glands sparse except above toenails.

· Temporal gland – Modified apocrine gland in SQ of temporal fossa between ocular lateral canthus and external auditory meatus.

· Present in African and Asian, both sexes.

· Chemical communication.

Answer 69

A

Housing Requirements (Fowler 8)

Social animals – spaces needed for multiple animals and the ability to separate for medical or behavioral reasons
Smooth floors, roughened floors, sand pits, polymers for flooring
Need access to dirt and water for skin protection
Thirty minutes of exercise is a reasonable goal
Enrichment items need to be strong – large plastic balls, fire hoses, tires, fake trees
Additional considerations for withstanding the aggressive behavior of bulls in musth
ERD for procedures

Answer 70

A

Feeding (Fowler 8)

Wild – browsers and grazers, high fiber and low protein (8-12%), eating about 1.2-2% of BW per day
Asia – palm leaves, bark, rice, tamarind, salt, green fodder, browsing in forest
NA – hay, browse, pellets, grain, fruits, vegetables
Caretakers should be educated on hay quality, nontoxic species of browse, and appropriate storage
Obesity is a common problem – leading to reproductive abnormalities, heart disease, osteoarthritis
1. Diets by feeding lower energy forage, increased browse and eliminated pellets and other concentrates can be successful
Elephants can take up to 100 L of water at a time – usually much less than this varying day by day. They can go several days without drinking – but refusal to drink is often an early sign of illness

Answer 71

A

Preventive Medicine (Fowler 8)

Daily inspection – skin, eyes, mouth, feet, feces, appetite, & drinking
Foot trimming as needed
Vaccines – rabies & tetanus toxoid
CBC/Chem/Fecal/UA yearly
Annual trunk wash mandated by USDA
EEHV screening

Answer 72

A

Tracheal Anatomy & Intubation

Hypoxemia common complication of anesthesia due to pressure on the diaphragm from abdominal contents
Elephants have a large fleshy epiglottis that lacks cartilage allowing for a tight seal, but can be easily pulled forward to allow access to the glottis
Thick vocal chords occlude the entrance to the trachea ventrally, but thin out dorsally
Intubation is blind, guided by breathing – displace the epiglottis, manually guide the tube to the dorsal aspect of the glottis where the chords are thinner.

Answer 73

A

Free-Ranging Immobilization

Etorphine – nonselective opiate agonist – binds mu, kappa, delta receptors, activates G-proteins, with effects at low fractional receptor occupancy
1. Can be reversed
2. Doses at 1.5-3 mcg/kg
Drug delivery
1. 65-mm needles – hind leg, back, or shoulder as preferred sites
Elephants that go down sternally, need to be rotated to lateral
At risk for severe hypoxemia & hypertension (pink foam syndrome)
1. Correct with megavertebrate demand ventilator
2. Pink foam more common the more stress there is (azaperone & acepromazine may help)
Recovery – naltrexone at 100x the dose of the etorphine
1. First signs – trunk and ears – within 2 minutes
2. Recovery complete within 10 minutes

Answer 74

A

Managed Care Immobilizations

Hand injected, darted, or pole syringe
Standing sedation – xylazine/butorphanol produced deeper sedation than azaperone
General anesthesia – etorphine (6 mg may be sufficient compared to 10-12 mg commonly used in the field)
Maintenance – inhalants or 1-2 mg doses of etorphine

Answer 75

A

Physical Examination (Fowler 8)

Gait – four-beat walk, lame elephants do not head nod or hip hike
Lumps & bumps, verrucous growths on Asian elephant trunks are normal, vulvar papillomas are also common
Blepharospasm rather than tearing is a more reliable indicator of ocular disease
Temporal gland drainage is normal, but may indicate musth in an adult male
ACS should inspect the pad & sole daily
Temperature can be obtained from the middle of freshly passed fecal ball
Auscultation usually only possible in elephants less than 2270 kg (5000 lbs) – lift front leg, place stethoscope behind the elbow
Trunk odor may indicator pneumonia
Oral examination helpful for EEHV monitoring
Venipuncture
1. Auricular veins, warming the ear up may help identify them
2. Cephalic (proximal medial forelimb) or medial saphenous (distal medial hindlimb)– tourniquet may help raise the veins

Answer 76

A

Surgery (Fowler 8)

Fraught with complication – conservative medical management, even of large wounds, often is sufficient
Cesarian sections no longer performed – no dam survived, dead calves have been retained for multiple years without dams becoming septic
Techniques and tools are available for laparoscopic vasectomies of wild elephants
Vestibulotomy has been done to remove a dead calf from birth canal, but issues healing are common
Dental surgeries are the most common – tusk extraction, pulpotomy, pulpectomy, molar extraction (impaction)
Surgical castration does not decrease aggression of bulls in musth
Necropsies
1. Team approach
2. Thorax should be entered last to reduce expose to TB, N95s not surgical masks should be worn, if granulomas are seen, minimize personal and put on additional PPE before proceeding

Answer 77

A

Laparoscopy in the elephant

Animal positioning
- Standing is best- sedated and restrained or general anesthesia and craned
  - Typically can be performed with torb/detomidine and local blocks
- Platform to put surgeons at level of paralumbar fossa
- If craned, ropes around each leg in axillary/inguinal area- less pressure on thorax improves respiration, less on abdomen assists sx
- Intubation and assisted ventilation when insufflation is used

Surgical anatomy of the elephant

Flank approach, just rostral to tuber coxae and ventral to caudal ribs
- Ribs extend into area of paralumbar fossa and difficult to palpate- probe w/5in, 18G spinal needles to identify ribs
- Incision for primary cannlua 15cm and made ~10cm rostral to tuber coxae and immediately ventral to ribs
- Cut through muscle layers, then make window in fibroelastic peritoneum and fat
- Primary cannula placed and held w/purse-string suture in peritoneum
- Second site on contraleteral flank same way
- Accessory site made in same way w/2cm incision 10cm rostral to midlle of primary incision
  - Obturator introduced through in direction of testis
Exploration w/visualization of kidney, ureter, testes, ovary, uterus, ductus deferens, colon, portions of small bowel possible

Elephant vasectomy procedure

Intra-abdominal testes located caudal and lateral to kidneys
Vas deferens is better termed ductus deferens -> more like long epididymis
- Originates at caudal pole of testis and courses toward bladder in mesoductus
- Traumatic grasping forceps through telescope grab ductus deferens
- Scissors through accessory portal resect 4-8cm section (removed through primary cannula)
- Abdomen desufflated, equipment removed, and sites closed
- As soon as ductus deferens identified on first side, second group of surgeons begin similar approach on contralateral side
  - Peritoneum not incised until first side is closed for gas-tight seal
  - Second side repeated like first side
Closure
- Peritoneum, external abdominal oblique fascia and muscle all closed in simple continuous pattern with 0 polyglyconate
- Skin closed w/horizontal mattress sutures of 5 stainless steel w/plastic stents
  - Accessory portals closed in same way w/one suture
  - Ends of wire suture twisted together (like cerclage wire), not tied
- Easier to close when animal is not recumbent
- Scrub perisurgical area before placing in lateral recumbency for reversal

Answer 78

A

· Salmonellosis

o Primarily enteritis, may result in sepsis - motile gram negative bacilli.

o S. typhimurium, S. Dublin, S. enteriditis reported in elephants.

o Damage to ileal and proximal colonic enterocytes, binding to GALT, results in infection of tissue MP.

o Gross – Distal small intestine, colonic mucosa reddened or discolored, friable fetid material. Viscera i.e. liver and spleen may have pinpoint to larger pale foci of necrosis and inflammation.

o Multifocal coalescing erosions and ulceration of mucosa overlying GALT.

o Dx on culture of feces or infected tissue.

Answer 79

A

· Anthrax

o OIE reportable, zoonotic, highly fatal.

o Aerobic gram positive endospore-forming.

o Bacillus anthracis.

o Free-ranging elephants, more rare in zoos.

o Seasonal epizootics assoc with hot, dry weather following heavy rain or floods.

o Routes on finfection – aerosol, ingestion, wound contamination.

o MP phacotysose vegetative or spore forms, bacterial replication results in MP lysis and dissemination.

o Potent exotoxin causes increased capillary permeability and coagulopathy.

§ Peracute, acute disease without other CS.

o Lesions reflect toxin-mediated vasculopathy.

o Hemorrhagic fluid exuding from body orifices.

o Marked edema of SQ tissues of head and neck.

o Blood is dark, thicks, clots poorly.

o Widespread hemorrhage internally.

o Anthrax exotoxin inhibits rigor mortis, carcass may be flaccid or have delayed rigor.

o Do not open carcass.

o Blood smears contain high numbers of sporulated gram positive bacilli.

o Dx can be presumed based on history and carcass presentation, detection of sporulated bacilli in blood smears.

Answer 80

A

· Clostridium spp.

o Toxemia from Clostridium septicum may be fatal, contamination of cutaneous wounds.

o Anthrax important ddx since both are spore forming.

o C. perfringens and C. difficile may result in hemorrhagic enteritis, fatal sepsis.

Answer 81

A

EEHV Preparedness

Plan for viremia, treatment of ill animals, necropsy guidelines, and supplies needed
Famciclovir – large amount needed not usually available on short notice
Cultivate and maintain open communication with vet staff, elephant care team, administrators, and public relations personnel

EEHV Vigilance

Most up-to-date resource is the EEHV Advisory Group www.eehvinfo.org
Elephants can die within 24 hours of showing signs
- Can be viremic upto 2 weeks prior to onset of clinical signs
Mild to moderate leukopenia (monocytopenia), and thrombocytopenia
Compare to individual elephants own CBC (not general reference intervals)
Recommendations: monitor at-risk elephants (1-8 yr Asian) weekly for EEHV viremia via whole blood qPCR and CBC

Answer 82

A

Early, Aggressive Treatment for EEHV (F9)

All ill young elephants as possible cases until proven negative by whole blood qPCR
Treatment initiated rapidly and often before confirmation of qPCR
Famciclovir most often used in NA and Europe (also acyclovir and ganciclovir)
- Efficacy has not been proven but also no data saying it doesn’t work
Rectal fluids immediately (can be q2h, minimum 3-4x daily)
Fresh and frozen-thawed plasma and crystalloids given IV every 1-3 days
Antibiotics for secondary infections, anti-inflammatories, and opioids
Consider steroids when DIC is observed (not evaluated treatment)
Difficult to predict if viremia will remain subclinical or cause clinical disease

Fowler 7

TREATMENT

Anti-viral Drugs and Other Medications:

•Most survivors were started early on high doses of anti-viral medications.
- -current recommendations are to start young animals immediately if nonspecific mild clinical signs are present (depression/lethargy) and before diagnosis is confirmed.
•Anti-viral drugs can be hard to acquire quickly: some zoos keep antivirals on site to treat an elephant for 2-3 days until new supplies are available.
- -may need to work with your local human hospital/pharmacy for supplies.
•3 Anti-virals have been used to treat EEHV: acyclovir, famciclovir, ganciclovir.
- -aciclovir and famciclovir are effective against alpha herpesvirus in humans (effective against beta-herpes like EEHV are unclear)
- -ganciclovir is effective against beta herpesvirus
- -famciclovir is most commonly used antiviral in EEHV treatment. Loading dose of 16mg/kg q 6 hours for first day, 12mg/kg q 6 hours on day 2, continued for 2 weeks.

Fluid Therapy:

•Fluid therapy may be lifesaving in EEHV patients via treatment of early shock.
- -methods of administration: oral, rectal, IV.
- -bolus 20mL/kg dose and then reassess the elephant. Usually max of 3 boluses. Maintenance rate = 40ml/kg/day
- -IV access: ear vein, saphenous vein, etc.
•Fresh plasma, fresh-frozen plasma, and fresh whole blood are colloids that have been administered to elephants.
- -Cross match of donor and recipient should be done.
- -A very slow infusion rate initially to observe patient for reaction
•Synthetic colloids such as hetastart and dextrans and pxyglobin have not been evaluated in elephants but may be a good option.
•Almost all species benefit from blood transfisiopn at a HCT <12%, and many clinicians elect to transfuse at a HCT<20%.
- -1% body weight (in kg) is suggested as max amount of blood to withdraw

Answer 83

A

In addition to the chart, monocytosis, leukocytosis, and thrombocytosis occur mid disease along with anemia and elevations in APP.

Answer 84

A

Fowler 9

Introduction – Mycobacterium tuberculosis (Mtb)

Important primary respiratory tract disease of Asian elephants, rare in African elephants
Between 1997 and 2001, prevalence in NA was 5.1% in Asians and 0% in Africans
- Close working relationship between Asian elephants and people
Two reported Mtb in free-ranging Asian elephants and one unconfirmed African elephant case

Transmission

Requires close, prolonged aerosol contact with infected individuals
Fomites and manure are sources for M. bovis but not of importance of Mtb

Fowler 6

Definition

· The term tuberculosis in mammals defines disease caused by M tuberculosis complex which includes M tuberculosis, M bovis, M africanum, M microti, and M. canetti, M caprae and M pinnipipedia

· Mycobacteriosis describes disease caused by non tuberculous mycobacteria (NTM) also called “atypical mycobacteria” or “mycobacteria other than TB” (MOTT)

· M. elephantis – newly described myco in elephant (n=1), also seen in human sputum

Etiology

· M. tb is the primary agent in elephants, but dz has been caused by M. bovis

· M. szulgai is an uncommon NTM but has been isolated from 2 African elephants

· M. avium is often isolated from elephants but NOT been assoc with disease

Epidemiology

· Both African and Asian are susceptible

o Asian more often affected – (More Asians used in performances with closer human contact, and more often females)

· There are no reports to date of TB in wild elephants

· Reservoirs

o humans = M tb

o cattle = M bovis

· Transmission – resp or alimentary routes

o Feces, urine, genital, milk, feed and water may have contaminated droplets

· M. avium intracellulare group can survive in the environment and doesn’t need a host to survive, as opposed to M. tb

Answer 85

A

Diagnosis (Fowler 9)

Screening should be included in preventative medicine
- Individual and herd health
Thoracic radiographs and lung/gastric washes are difficult due to their large body size and poorly studied immune system
Intradermal tuberculin testing causes non specific reactions and is contraindicated
Direct tests - culture and qPCR confirms the presence of Mtb organisms
- Culture is the gold standard and only tests that confirms active infection
- Trunk washes are submitted for culture as it contains material from the lower respiratory tract
  - Training elephants to allow 60mL of saline into its trunk and exhaling into a sterile container
Indirect tests – serology, interferon gamma tests (IFN-γ), and cytokine stimulation assay demonstrates exposure
- Serology identifies antibodies to mycobacterial antigens
- Cytokine stimulation assays and IFN-γ detect components of cell-mediated immunity
- Cannot confirm active infection
If positive on indirect tests, confirmatory direct test is recommended (culture, qPCR)
- Positive TW culture, confirm by submitting a duplicate specimen or a new TW
- Multiple TWs performed over weeks to months may be necessary due to low bacterial numbers or intermittent shedding
- Positive cultures should be tested for antibiotic susceptibility

Answer 86

A

Treatment

Key is to prevent shedding into the environment
- Use a combination of first-line antitubercular medications
- Avoid serious drug-related adverse events
- Achieving and maintaining adequate serum drug levels throughout treatment
Should be started as soon as culture is positive, before antibiotic susceptibilities are complete
Primary drugs – isoniazid, rifampin, pyrazinamide, ethambutol, and levofloxacin
- Isoniazid – causes early rapid killing of actively replicating Mtb organisms within a few days of starting treatment; not as monotherapy due to developing resistance
- Rifampin – resolves cavitary lesions and activity against latent Mtb organisms
- Pyrazinamide and Ethambutol – synergistic with other drugs and good at preventing resistance but should not be used instead
- Levofloxacin – can be substituted for the other three except isoniazid
Enrofloxacin has been used but effectiveness is questionable due to resistance to ciprofloxacin
- Oral or rectal administration
Appropriate drug doses and concentrations are unknown but need to achieve blood concentrations
Adverse reactions – depression, colic, inappetence, black fetid manure, blepharospasms, ocular tearing, and lethargy
Treated with a short intensive phase and a longer continuation phase
- Intensive phase – high and frequent doses of 3-4 drugs for 8-10 weeks
- Continuation phase – antibiotics at lower frequency (not lower doses) over months
TW monitoring for continued non detectable Mtb shedding and drug levels exceed MIC (q6 months)
Treatment failures – intolerance to medications or development of resistance

Answer 87

A

Male reproductive tract

Testes sit caudal to kidneys, supplied by central artery that radiates smaller arteries outwardly – still maintains testicular temps at 94-97
No epididymis – ampulla is the primary site of sperm storage
Accessory sex glands – ampulla, prostate, seminal vesicle (viewd rectally on US), bulborethral gland (viewed below anus on US)
S-shaped penis elevates vulva into position prior to penetration (levator muscles)

Answer 88

A

Female Reproductive tract

Vestibule – urogenital canal – is 1-1.4 m long, curves into the pelvis at the level of the anus
Clitoris located distally
Urethral opening and vagina found in cranial portion of urogenital canal
Two blind pouches on lateral sides of vagina
Ovaries – small
Uterus bicornuate and short bodies

Fowler 8

Answer 89

A

Estrus cycle

wild – maturity @ 10-12yrs
captive – early – 3.5-5yrs
spontaneous ovulation, polyestrous, non-seasonal (births all year long)
- may have a peak in rainy season
captive – natural estrous sync has been observed (may be artificial)
estrous cycle is 13-18wks – longest spontaneous cycle of any mammal
- wild may have seasonal diff in length of folic and luteal phases
- can monitor progestagens weekly via serum, urine, or feces
- biphasic profile of interchanging periods of elevated (luteal phase, 6-12wks) and baseline (interluteal or follicular phase, 4-6wks) progestagen concentations (see fig. 66-2 on pg 505)
2 distinct luteinizing hormone (LH) peaks – big diff from other mammals
- during follicular phase, only 2__^nd induces ovulation
- 2 peaks are 19-22d apart, each one is preceded by estrogen surge
- both terminate a follicular wave, but only 2^nd is ovulatory
- 2^nd one causes dominant follicle (~20mm) to rupture 12-24hr later
- 1^st (anovulatory) LH peak induces luteinization of larger follicles of 1^st follicle wave
- Luteinized follicles (LUFs) – seen on US ~10d post 1^st LH peak
  - Coincident w/ sharp rise in inhibin level
  - Luteal cells of CLs appear to produce inhibin and might be a source of this hormone in addition to dominant follicle
  - Inhibin is important factor for selection of dominant follicle
1st LH peak may be to induce luteinization of granulosa cells in larger follicles which secrete inhibin, which promotes deviation of a single dominant follicle in 2nd wave.
- Lutenized follicles are source of accessory CLs
- There are ~2-10 of these on each ovary of cycling and preg elephants, in addition to CL from ovulation.
Accessory CLs have a diff growth curve from ovulatory CL (they arise 8-10d earlier)
- They are usually more numerous on ovary of ovulation
- Max diam @25-30d post 2nd LH surge & 10 d earlier than CL from dom follicle
Ovulatory CL takes ~ 10d to be seen US after ovulation
- Ovulatory CL is largest of all CL @33mm (30-41mm) @40d post ovul (vs access. CL avg 25mm, 10-30mm range)
CL secretory capacity is diff from other mammals
- Only minute quantities of progesterone found (this is main progestagen in other sp)
- elephant CLs produce:
  - 5α -pregnane-3,20-dione (5α-DHP)
  - 3α -hydroxy-5α-prengnane-20-one (5α -P-3α -OH)
- corresponding receptors have been found in endometrium
- measured in blood
  - African = 5α -P-3α-OH predominantly
  - Asian = 17α-hydroxyprogesterone (17α -OH) is found in urine and feces but NOT in African species

Answer 90

A

US monitoring during estrous cycle (good pics pg 504)

Transrectal, low (4-2MHz) to med (<7.5) freq probes
may need extension handle to visualize cranial uterus & ovaries
luteal phase – may see anechoic mucous plug w/in vagina (protects uterus from contamination)
- excreted during follicular phase (non-conceptive cycle, 1^st LH peak) but maintained if elephant is pregnant
- endometrium – thin, 23.7mm cross sect diameter (vs. 35.4mm avg in follicular phase)
- ovaries – multiple, large CLs
  - accessory CLs maintain fluid-filled central cavity during 1^st 3wks post ovulation
  - antral follicles never occur during luteal phase – thus presence of multiple follicles is good indicator that female is in follicular phase
follicular phase
- presence of multiple follicles on ovaries
- higher fluid content in endometrium
  - due to low progestagen and increased estrogen
  - small amounts of free fluid might accumulate w/in uterus lumen 1-2wks before ovulation.
- uterus becomes more toned toward LH peak
  - horns – convoluted & easier to see b/c start to retract into pelvis
  - tissues are denser (due to contraction of myometrium) – increased brightness in image
  - after ovulation, endometrial diameter decreases progressively
  - reaches smallest thickness of 17mm in the early folicular phase of next cycle

Answer 91

A

Hormone cycle induction

Prostaglandin F2α (PGF2α) – doesn’t work in elephants
- Used in domestic ruminants and horses to induce luteolysis and initiate a new cycle
- African elephants – 2 cycling females, dose of 80-125mg/elephant did not affect serum progestagen secretion
- PGF2α given for 3 consec. days to young preg female (5mo gestation) to induce abortion
  - Urinary progesterone decreased to baseline but returned to pregnancy levels 10d after last injection
  - Healthy calf was delivered
Human Choriogondatotropin (hCG) – may work in elephants
- 5000 IU injected at time of last AI (to ensure ovulation)
  - Subsequent ovulation seen (6 cases) but may be due to natural timing of 2^nd LH peak.
  - May not be reliable: one case suggests that follicle needs to reach a certain size or stage of maturation to be receptive for ovulation induction
Gonadotropin-Releasing Hormone (GnRH) – may work in elephants but imprecise.
- May induce release of LH at lower levels than normal LH surges
- One study uses GnRH agonist (buserelin acetate) to induce 2^nd LH peak around time of 1^st LH peak
  - Give buserelin IV 13-43d after progest reaches baseline ->anovulatory LH surge
  - This was followed by 2^nd LH surge 15-22d later; 1 female conceived

Answer 92

A

Pregnancy

Gestation (African & Asian): 620-640 +/- 14d
Progesterone levels during 1^st 6wks conception are same as normal luteal phase.
- Transient progestagen decrease @ 8wks (conc. nears baseline)
- If pregnant, gradual increase to above non-conceptive luteal phase (2-3x higher)@ 10-12wks (esp during 1^st half of pregnancy)
- Levels may vary during pregnancy and at some points may be similar to those seen during luteal phase
Diagnosis of Pregnancy
- Progestagen:
  - Elevation of progestagen beyond normal luteal phase length, more than 10 wks above baseline level (wkly samples)
  - Asian: ratio of progesterone to 17α –OH is lower during 1^st 3-8 wks of conception vs nonconceptive cycle
- Prolactin
  - Single sample >6months
  - Prolactin rises 200-600 times higher than nonpreg values
- US
  - From day 50 post ovulation, embryonic vesicle may be detected in uterine horn via transrectal US
  - From day 250, transabdominal US may be successful to visualize fetus
Source of progestagen during pregnancy = conception preventing CL regression (aka “luteal rescue”), thus prolonging progesterone secretion.
- Functional CLs present throughout entire pregnancy
  - Mostly active 3-15months of gestation
  - Studies suggest that elephant placental tissue has no endocrinologic competence
  - Unknown what causes prolong of luteal lifespan and initiation of growth (“luteotropic factor”)
  - Unknown what endocrine events trigger parturition

Answer 93

A

Embryonic and fetal development

·embryonic fetal parameters have been measured to develop a growth curve for 1^st 200d gestation (chart and pics pg 510)
implantation seems to be delayed, 1^st fluid accumulation seen at 42-46d post conception
embryonic vesicle (EV) seen on day 50, (8-10mm)
blastocyst (formed after fertilization of oocyte) attaches in ipsilateral horn of ovulaton by replacement of endometrium and formation of endotheliochorial zonary (girdle) placenta
embryo visible w/in embryonic vesicle @62-65d
yolk sac distinct @ 74d, also see placental band (echodense protrusion into EV)
allantois visible @ 85-95d
heartbeat @ 70-80d
head and rump differentiation @83-85d
midgut herniation seen @ 95-120d
formation of hind/forelimb, straightening of cervical flexure, formation of brain w/ ventricles, organization of lung, liver, kidney, gastric vesicle all seen @ 100-120d
beyond 200d gestation, transrectal visualization difficult d/t drop of uterus into abdominal cavity
- Can do transcutaneous US from behind, paramedian to vestibule or from flank and inguinal regions w/ running water from a garden hose to increase coupling

Answer 94

A

Normal Parturition Process

Majority of births occur at night
May be born anterior or posterior position; 70% are born hindlimbs first
Progress of events during parturition
1. Onset of increased discomfort 7d to 48h prior to birth
2. Restlessness and first abdominal contractions – first stage of labor
3. Loss of mucus plug that seals the vagina
  - White, yellow to brown, stick and mucoid, or bloody
4. Increased signs of pain stronger contractions, straining, agitation – second stage of labor
5. Rupture of allantois
6. Appearance and disappearance of bulge under the tail below the anus and advancement of fetal feet
7. Strong contractions and final entrance of the fetus into the vestibulum – third stage of labor
8. Continued visibility of the bulge as soon as the calf’s body enter the female’s pelvic canal (and breaks hymen in nulliparous cows)
9. Movement of the calf through the vaginal vestibulum and expulsion from the mother’s body
10. Delivery of afterbirth 45 minutes to a few hours later
  1. Entire process may be finished within less than an hour but can take upto 48 hours

Answer 95

A

Prediction of Parturition

Most likely time of parturition as 620-660 days from last observed mating or rise in progestagens
Gestational length can vary by upto 3 months
Individual observation of physiologic or behavioral changes or daily serum progesterone
- Progestagen levels drop below 2/3 of the mean pregnancy value a few days prior to parturition
Physical changes predictive of birth (breast development, milk secretion, ventral abdominal or vulva edema) are not always as obvious as in other species
Loss of the mucus plug
- Usually occurs a few hours prior to birth, but can be several day prior or middle of gestation
Other signs: frequent defecation and urination, small fecal ball size, reduced appetite, restlessness, nervousness, beating the vulva with the tail, and labor
- Labor – freezing intermittently, spread legs, moving flanks, tail flagging, straining, kneeling, lateral recumbency, and repeatedly lying down and standing back up
Most reliable indicator is daily measurement serum progesterone
- A sudden >50% drop in progesterone levels around due date indicates birth may occur soon
Transrectal ultrasound allows evaluation of the cervix, parturition timing, and fetus position

Answer 96

A

Abnormal Pregnancy

Early embryonic loss or resorption may go unnoticed, uterine pathologies
- One case due to implantation next to a uterine leiomyoma
Infectious diseases - Poxvirus, Salmonella, EEHV (suggested but no proof)
Death of fetus may result in long-term retention without problems for the dam
- May remain in the uterus or be expelled days to years after death

Answer 97

A

Complications During Parturition

Common in elephants under human care, may be fatal for mother and/or calf
- 12% of calves (Asian or Africa) are stillborn
- Asian timber elephants in Myanmar only have 4% stillbirths
Asian elephants seem more prone to dystocia than African elephants
- Dystocia is significantly associated with stillbirths (50% in Africans and 86% in Asians)
Reasons for Dystocia
- Oversized fetus
  - More problems in Asians than Africans
  - Asian vs African birth height – 93.7cm vs 89.9cm
  - Asian vs Africa birth weight – 116.5kg vs 103.5kg
  - May be related to GL and body condition of cow (link b/w obesity and dystocia)
- Malposition/Malformation of Fetus
  - Dystocia linked to breech and anterior positions
  - 60% of calves head first experienced dystocia, compared to 16% born hindlimb first
  - 40% born head first were stillborn compared to 12% born hindlimbs first
  - Deformities – ankylosis, encephalomeningocele, hydrocephalus, spina bifida, cleft palate, missing maxilla and trunk, and tetralogy of Fallot
  - Malpositions – head tilt, twisted legs and entanglement with the umbilical cord
- Weak Labor/Uterine Inertia
  - Linked to fitness, calcium deficiency, and insufficient labor due to external stressors
  - Normal plasma total calcium concentrations – 3.6mmol/L
  - Normal plasma iCa concentrations – 1.25mmol/L
  - Should be fed calcium and cholecalciferol-rich diets at all times, particularly when close to parturition
  - Wild elephants, allomothering and herd-supported births are normal
    - A subdominant female under human care with no relationship with the rest of the herd may be stressed by feeling vulnerable and intimidated
      - Separation from the herd may alleviate stress or may increase stress
- Difficulties in Passage Through the Birth Canal
  - Inadequate dilation of the cervix, poor softening of the vagina and vaginal vestibulum (older, primiparous females), fibrosis of the hymen, lower reproductive tract pathologies, and obesity
  - Parity is a primary factor for passage problems
  - Fitness may play a role in successful calf expulsion
    - 4% stillbirths in working Myanmar timber elephants vs 12% in zoo elephants
  - In Myanmar timber elephants, stillbirths dropped from 11.3% for first-born calves to 1.8% for later-born calves
    - First born calves were 6.83x more likely to be stillborn than subsequent calves from the same mother
    - Similar for Asian elephants under human care in Europe
  - The hymen that “seals” the entrance from the vaginal vestibulum into the vagina breaks during first parturition (not during mating) and may be fibrotic with age adding an obstacle for older first-time mothers

Answer 98

A

Obstetrics

Veterinary intervention is limited due to size and special reproductive anatomy
- Rectal palpation and transrectal ultrasonography are valuable tools
Triggering the Ferguson Reflex
- Manual massage of rectal floor to stimulate the underlying vagina and cervix
  - Pull knuckles along the rectal wall towards the anus causes stimulation of the underlying vaginocervical receptors which will trigger the release of endogenous oxytocin
Calcium
- If total serum calcium and iCa are low, administer oral or slow IV calcium
  - IV Ca-gluconate (23%) at a dose of 400-2000mL has been reported in a dystocic Asian
Oxytocin
- Elephant uterus is very sensitive to oxytocin
  - Assess for degree of cervical dilation prior to administration
- Consider if cow is multiparous, cervix is open, parts of the fetus have entered the vagina, and birthing has stopped for atleast an hour
  - If not met, uterine rupture has occurred
- 20-60 IU IM every 2 hours (takes ~20 minutes from injection until onset of full effect)
- Carbetocin
  - Longer and smoother acting metabolite
  - Single dose of 200-400μg
Denaverine
- Antispasmodic drug to help relax the cervix and birth canal while not affecting labor
- Not much experience in elephants but worth considering in nulliparous females with inadequate cervical opening
- 8000-1600mg IM, repeated once an hour
- No effect on early cervical dilation
Estradiol and Prostaglandin E
- Estradiol gel (600-800mg) applied by transrectal massage
  - May be repeated at 300-400mg q3-4h
- Prostaglandin E1 and E2 used in humans and mares for cervical ripening and induction of labor
- Misoprostol (synthetic PGE1)
  - Dilates the cervix and induce uterine contractions
  - 1000mg orally or 500mg transrectally q12h (for a 4000kg elephant)
- Dinoprostone (PGE2 analogue)
  - 1.5-2.5mg transrectally above the cervical region
Surgical Approaches for Fetal Delivery
- Surgery as a last resort
- Cesarean section is not an option in elephants
  - 6 C-sections have occurred, all resulting in death of mother and calf
  - Weight of intestines, large incision, mechanical forces, slow healing of skin
- Episiotomy/Vaginal Vestibulotomy and Fetotomy
  - No direct access to the vagina and cervix
  - Direct access to vaginal vestibulum via vertical incision (15cm x 8cm) ventral to the anus
  - Extraction forces applied physiologically not horizontally
  - Local anesthesia over stand sedation for the cow to assist with fetal expulsion
  - Fetotomy as a last option (but needed to prevent ascending infections)
    - Attempts have results in dam death due to uterine trauma and infection
    - Success in 1 Asian elephant (9 hour procedure)
  - Persistent vestibular fistula
    - Common postsurgical complication
    - Risk of ascending infection due to fecal contamination
    - Second intention healing of skin while suturing the mucous membranes and muscular layer of the vaginal vestibulum are mandatory to prevent problem
- Fetal Retention
  - If the fetus never enters the birth canal, no membrane rupture, and the cervix remains closed, no interventions should be done (safest option)
  - Will expulse calf (sometimes years later) and may cycle and become pregnant again after fetal expulsion

Answer 99

A

Fowler 6 Ch 44 – Neonatal Elephant Mortality

· Higher stillbirths and infant mortality in Asian elephants in US and Europe than in SE Asia
· Most significant factors
o Multifactorial dystocias and stillbirths
§ Primarily older nulliparous
§ Overweight and physically unfit females at higher risk
§ Pelvic fusion, uterine abnormalities, hormonal fluctuations, fetal malposition
§ Infectious salmonellosis and Herpesvirus infections have caused stillbirths
o Herpesvirus infections
§ High mortality in calves
o Maternal rejection and aggression
§ Calves commonly pulled from dam at birth, gradually reintroduced to avoid this problem
o Trauma (herdmates, exhibit design)
o Poor-survivability in hand-reared calves
§ Hand-rearing should be an absolute last resort
§ FPT
§ Diarrhea from inappropriate formula
§ Diarrhea from infectious causes
§ Sepsis
§ Nutritional malabsorption
§ Metabolic bone disease
o Infectious diseases and sepsis
§ One report of fatal umbilical infection
§ Sepsis secondary to other medical problems or injuries
o GI disorders
§ Impactions have been reported
§ Enteritis, volvulus, and torsions reported
§ Clostridial diseases and salmonellosis
· Misc. info
o Umbilical hernias reported
§ surgical correction in one case
§ Medical resolution in two cases (publication pending) by repeated, long-term manual reduction
§ (Medical resolution also successful in a current case at STL Zoo)
o AI no responsible for 50% of pregnancies in North America

Answer 100

A

Neonates

Protocol for neonatal elephant examinations
- When calf is first separated from dam
  - Immediately after birth for uniparous dams, w/in 24-48h for multiparous dams
  - Exam includes- BW (avg. 105.5kg for Asian), height (avg. 88.9cm for Asian), length, girth measurement, PE (esp. periocular, oral, anogential areas), ausculatation (thoracic and abdominal), assessment of limb conformation, evaluation of umbilicus (treatment w/iodine)
  - Blood collection- CBC, biochemistries, assessment of passive transfer
    - Prior to nursing, passive transfer negative- no in utero placental immunoglobulin transfer
    - EDTA whole blood and placental sample to National EEHV Lab
  - Regular monitoring crucial
  - 92% of time meconium passage w/in 7h of birth
    - If not passed by 2 days of age- potential problem (e.g., congenital defect [e.g., atersia ani/coli], constipation, dehydration, GI stasis, insufficient milk intake)
Ongoing assessment of calf health
- Diluted chlorhexidine solution on umbilicus- QID x 1 day, then TID x 3 days, when calf sleeping or dam restrained
- BW- daily or when possible for first 2wk , gain should be ~0.45-1.4kg/day for first year
- Oral exam- BID for ulcers, swelling, cyanosis of tongue (late signs of EEHV in some)
- Respiratory rate- BID; normal for 1wk old is 20-22bpm
- Heart rate- BID; normal for first week is 115bpm (range 100-128)
- Rectal temp- BID; normal 36.3-37C (97-99F)
  - CBC and chem PRN
  - Keep first occurrence log to identify and monitor behavioral and developmental signs to help determine health status
Nursing
- Neonates often clumsy and may need assistance in learning
- Train nulliparous dams to allow mammary gland manipulation prior to birth
- 74% of calves attempt to nurse w/in 7h of delivery (but can be delayed up to 24h)
- If none by 8h, review calf’s condition and behavior
- If none by 12-24h, need to decide whether to supplement
  - Based on calf’s vitality, strength of nursing efforts, dam’s receptivity to calf
- Supplements
  - Can be colostrum or milk from the dam, milk replacer, plasma
  - Can be offered from a bottle and nipple (bovine type) or stomach tube
  - May be reluctant to take a bottle if already nursed from dam
Failure of passive transfer
- Zonary, endotheliochorial placenta barrier to in utero transfer of immunoglobulins
- Time from for absorption via gap junctions in intestinal mucosa is probably 12-36h
- Colostrum consumption is typically 2-10L during early nursing
- Supplementation with colostrum milked from dam, banked elephant plasma, commercial equine hyperimmune plasma
- Even after absorption window, may provide local intestinal immunity
- Oxyocin (30-60 IU IM) may facilitate milking of dams (yield is 300-1000ml/milking)
- IV plasma can be used w/failure of passive transfer or particularly weak/at risk calves
  - 40-80ml/kg IV over 2-4 days
  - Single bolus of 10-20ml/kg IV over 30-60 minutes can be given
  - Monitor response to treatment (TP, gamma globulins, FPT tests) after transfusion

Answer 101

A

Elephant calves

Training
- Start socialization w/humans quickly
- Consistent interactions that don’t allow undesirable behaviors
- Formal training as matures- present all parts of body for exam, sternal/lateral recumbency on request, present feet/limbs for rads, open mouth for oral exam, step on scale, stand for urine collection, allow blood collection, accept medications (PO, rectal, parenteral)
Preventive health
- Monthly CBC and chems for first 2-3y
  - e.g., HCT decreases may be early sign of EEHV
- Urine and feces
  - Monitor urine volume, frequency, signs of stranguria, discoloration
  - Feces may be lighter and less formed on milk
  - Sign to perform more diagnostics- excess mucus, blood, or sand in feces
  - Salmonella culture/PCR for any calf with diarrhea, may be fatal
- Weight
  - Poor weight gain caused by calf or dam illness
  - Monitor production of dam and intake of calf
  - Domperidone (500-5000mg/day PO) may be used for agalactia
- Vaccination
  - Tetanus toxoid at 3-4mo, at 1y, then q5y
  - Rabies at 6-8mo, then annually in endemic area or if high risk of exposure
Hand rearing
- 11 cases of reported completely hand reared calves- 5 survived passed infancy
- May hand rear due to intractable dam aggression, calf/dam illness
- Grober ElephantGro African and Asian milk replacement formulas
  - Elephants have much less lactose than cow milk and different fatty acid profile
  - Do not use formulas based on cow milk- likely induce diarrhea
- Commercial bovine bottles and nipples
  - Wash after every use, sterilize q3-4 days
- Newborns nurse on demand (up to 12X in 24h)
  - Gradually increase interval to 6-7X/day by 1y of age
  - Calves consume 10-15% BW daily, volumes depend on formula caloric density and fluid requirements
  - Estimated caloric requirements: 8000kcal/day for 100kg calf, 16000-20000kcal/day for 200kg calf
- NSHP (and bone fractures) has occurred in formula-fed calves
  - Improperly balanced formulas or malabsorption of nutrients in intestinal tract
  - Ca and iCa monitoring relatively insensitive monitoring method
  - Urinary Ca and P may be better to detect early deficiencies or imbalances
    - Fracture treatment not successful
- Solid foods introduced at 1-2mo for calf experimentation
  - Gradual weaning by 3-5y
  - Gradual decrease in milk intake starts at 12-14mo
  - Try not to wean before 2y

Answer 102

A

Management of sick calves
- Sedation is likely needed for diagnostic testing and treatment- Torb/detomidine
- Fluids rectally, IV, IP, but not SQ
  - Auricular arteries best place for catheter, but difficult to maintain and nothing can be administered into them but balanced electrolyte solutions
- Trauma
  - From herdmate aggression/physical encounters, accidental falls, drowning
  - Prevention- assess cow for aggressive behavior, nonslip surfaces, limit access to pols/moats until calf is familiar, proactive plan for introduction to herd
Sepsis and MRSA
- Sepsis following maternal rejection, FPT, transport, trauma, umbilical infection
- Acute septicemia from Salmonella after stressful event
- Antimicrobial use reported, but studies lacking
- MRSA
  - One case, hand-reared calf, acquired from human caretakers
  - Discharge from skin wounds at 7wk of age
  - Skin lesions resolved w/antibiotic therapy, but calf failed to thrive
  - Enterococcal septicemia
Umbilical hernias
- Reported in Asian calves
- May be detected at neonatal exam, but may not be apparent until 2-4wk
- Larger ones require surgical correction
- Nonsurgical treatment (daily manual reduction) possible when small in size, completely reducible, no incarcerated viscera, no infection
- Early detection important- repeated palpation and ultrasound
Diarrhea and other GI problems
- Establish normal parameters for color, consistency, frequency for each individual
- Diarrhea a common problem
  - Differentials- dietary intolerance, dietary indiscretion, imbalanced/abnormal bacterial flora, parasitism, septicemia, salmonellosis
  - Diagnostics- PE, CBC, chem, and fecal culture, cytology, parasitology
  - Treatment- fluid therapy, targeted antibiotics, antiparasitics
- Calves ingesting sand at young age may develop habitual intake- intestinal impaction
  - Management- change substrate, fluids, enemas, psyllium or other fiber supplements
  - Severity can require surgery
Dental problems
- Tusk fracture- risk of pulp exposure greater than in adults due to tusk length
  - Rads to see pulp canal
  - Sx removal of necrotic or exposed pulp tissue, sealing of open canal w/dental synthetic bone graft material and chemical composite
  - Metal crown to prevent excessive wear
- Molar eruption time varies among individuals
  - Common problems- malalignment, abnormal wear, abscessation

Answer 103

A

Diagnostics (Fowler 8)

Clinical pathology
1. CBC
  1. Largest RBC of any mammal, lower cell counts
  2. Heterophils not neutrophils
  3. Lymphocyte the dominant white cell
  4. Two types of monocytes – unsegmented and bi- or tri-lobed
  5. Inflammation – monoctyes increase, few band heterophils may indicate severe inflammation
2. Chemistry
  1. Lower sodium, chloride, higher proteins (mostly globulins)
  2. Elephants have blood groups – cross match before transfusion
  3. Elephants don’t produce bile acids
  4. BUN & creatinine are generally low
3. Urinalysis
  1. Transient glycosuria & ketonuria can occur in healthy animals
  2. Low specific gravity – neonates have more concentrated urine, but that changes within 24 hours
4. Trunk was for respiratory evaluation
  1. 60 mL sterile saline in one nostril, trunk lifted, lower and exhaled into a bag

Answer 104

A

Elephant Foot Radiography-

Anatomy:

· Elephants are ungulates with modified digitigrade stance on forefoot, and semiplantigrade on the hindfoot

· Both species has five digits in each foot

· Asian elephants (Elephas maximus)-

o Forefoot-

§ Nails associated with D1 to D5

§ Nails intimately connected to distal phalanx

§ D1 has only two phalanges (P1 and P2)

· Hindfoot- typically nail associated with D2 to D5 (can vary)

§ A single phalanx without sesamoids on D1

· African elephants (Loxodonta Africana)-

· Forefoot-

§ Four toenails (actual number may vary)

§ D1 has one phalanx and sesamoid

§ D5 has one phalanx and paired sesamoids

§ D2 to D4 have three phalanges each with paired sesamoids

· Hindfoot-

§ Three toenails (may vary)

§ D1 is represented by a sesamoid

§ D2 and D5 each have two phalanges

§ D3 and D4 each have three phalanges

· Pathology in the nail, sole, and/or pad may lead to pathologic changes in the bones of the digits- most commonly P3 of the weight-bearing digits D2 to D5.

Patient Preparation-

· Proper training and animal/human safety measures are essential

Equipment Setup-

· Image plate and lead shield to minimize artifact are housed within a hexcelite wooden tunnel for protection

· Radiographs are collimated to evaluate each digit individually

o Center on the toe at a 45-degree angle for a dorsoproximal-palmarodistal oblique projection

o “Foot-o-grams: are not recommended

o Orthogonal views are not recommended

o Center x-ray beam at cuticle for images of P3

o Center beam 6 to 7.5cm above cuticle for images of P1-P2

· A technique chart should be developed (example- Table 67-1)

· A tripod should be used to minimize human exposure

· Manual restraint of radiographic plates not recommended for safety reasons

· Use of proper PPE is essential

Conclusion-

· Use of consistent technique across institutions will facilitate case consultation with radiography and zoological medicine specialists.

Answer 105

A

AJVR 2022 83(8):ajvr.22.01.0016
Potential diagnostic biomarkers for pulmonary tuberculosis in humans are not elevated in Mycobacterium tuberculosis culture-positive Asian elephants (Elephas maximus)
Coughlin LL, Sanchez CR, Monti MI, Griffioen JA, Nutter FB, Beamer GL

ABSTRACT:
OBJECTIVE: To determine (1) if chemokine (C-X-C motif) ligand 1 (CXCL1), matrix metalloproteinase 8 (MMP8), interleukin-10 (IL-10), interferon-γ (IFN-γ), and tumor necrosis factor-α (TNF-α) can be detected in serum from Asian elephants, and (2) if their concentrations are significantly elevated in Mycobacterium tuberculosis (M.tb) culture–positive elephants compared to –negative elephants. CXCL1, MMP8, IL-10, IFN-γ, and TNF-α were recently identified as potential diagnostic biomarkers for pulmonary tuberculosis in experimental studies in animals and humans. Therefore, we hypothesized that they would be detectable and significantly elevated in M.tb culture–positive elephants compared to M.tb culture–negative elephants.
SAMPLE: 101 Asian elephant serum samples, including 91 samples from 6 M.tb-negative elephants and 10 samples from 5 M.tb-positive elephants (none of which exhibited clinical signs of disease). M.tb status was determined by trunk wash culture.
PROCEDURES: Commercially available ELISA kits were used to determine the concentrations of each biomarker in serum samples.
RESULTS: Biomarker concentrations were below the limit of detection for the assay in 100/101 (99%) samples for CXCL1, 98/101 (97%) samples for MMP8, 85/101 (84%) samples for IL-10, 75/101 (74%) samples for IFN-γ, and 45/101 (45%) samples for TNF-α. Multiple M.tb culture–positive elephants did not have detectable levels of any of the 5 biomarkers.
CLINICAL RELEVANCE: CXCL1, MMP8, IL-10, IFN-γ, and TNF-α were not elevated in M.tb culture–positive Asian elephants compared to M.tb culture–negative Asian elephants. This may be related to disease state (ie, clinically asymptomatic). More sensitive assays are needed to better understand the role of these biomarkers in M.tb infection in Asian elephants.

Background:
- Mycobacterium tuberculosis infection is a threat to elephant health and conservation
– Humans are the natural host for TB and main source of transmission to elephants
– Infection can spread from between elephants, other mammals, and back to humans
– Asian elephants with TB develop weight loss, inappetence, lethargy, coughing
– Similar pulmonary pathology as humans (e.g., granulomas with necrosis)
- Current gold standard for TB in Asian and African elephants is trunk wash culture
– Limitations: time required to train animals, time required to obtain culture results intermittent shedding, and the possibility of bacterial and fungal contamination
- Serologic tests, including the Dual Path Platform VetTB, seem more advantageous
– Demonstrated high sensitivity and specificity for TB in Asian and African elephants
- CXCL1, MMP8, IL-10, IFN-γ, & TNF-α reported to be elevated TB infected mice
– CXCL1 = chemokine, involved in neutrophil recruitment and activation
– MMP8 = collagenase, contributes to TB pulmonary granulomas/necrosis
– IL-10 = inhibitory cytokine, suppresses host immune response, promotes TB survival
– IFN-γ & TNF-α =inflammatory cytokines, macrophage activation, granuloma formation

Key Points:
- Multi-center retrospective serosurvey (6 TB-negative & 5 TB-positive elephants)
- CXCL1, MMP8, IL-10, IFN-γ, and TNF-α were below the limit of detection in most samples
– However, a concentration above the limit of detection was present in at least 1 sample
– While detected in Asian elephants, biomarkers unlikely to have utility for elephant TB

TLDR: CXCL1, MMP8, IL-10, IFN-γ, and TNF-α were not elevated in TB culture–positive Asian elephants

Answer 106

A

CHANGES IN SERUM CARDIAC TROPONIN I IN ASIAN ELEPHANTS (ELEPHAS MAXIMUS) WITH ELEPHANT ENDOTHELIOTROPIC HERPESVIRUS INFECTION
Kailey B. Anderson, DVM, James C. Steeil, DVM, DACZM, Erin Latimer, MS, Victoria Hall, DVM, MS, DACVPM, Lee-Ann C. Hayek, PhD, and Joao Brandao, LMV, MS, DECZM (Avian)
Journal of Zoo and Wildlife Medicine 53(2): 249–258, 2022

Abstract: Elephant endotheliotropic herpesvirus (EEHV) is one of the most important causes of mortality in Asian elephants (Elephas maximus). The unusual tropism of EEHV for endothelial cells of capillaries can lead to catastrophic vascular dysfunction, hemorrhage, cardiac damage, and death. Cardiac troponin I (cTnI) is an intracellular protein of cardiomyocytes that is released into circulation in levels directly correlated to the severity of cardiomyocyte damage. The purpose of this study was to assess if cTnI could be used to distinguish when EEHV viremia leads to clinical disease versus subclinical infection. Thirty-seven individual Asian elephants contributed 53 blood samples that were evaluated for EEHV viremia using quantitative polymerase chain reaction and analyzed for cTnI using a high-sensitivity assay. Viremia was categorized as none (24/53), low (< 20,000 vge/ ml, 12/53) and high (20,000 vge/ml, 17/53). Seven of the nonviremic samples had detectable cTnI. Nine low viremia samples were positive for EEHV1 (1A and 1B combined) and lacked a detectable cTnI. Fourteen high viremia samples were positive for EEHV1 and had detectable cTnI. There was statistical significance between having viremia and having a detectable cTnI value (P=0.0001), and animals with EEHV1 viremia were more likely to have a positive cTnI value (P=0.04). The presence of cTnI was associated with the presence of clinical signs, with higher values of cTnI in the presence of clinical signs versus subclinical viremia (P=0.0001). In addition, four elephants contributed multiple samples from a single viremic event and results displayed a trend of elevation in troponin values with progression of EEHV viremia. The association of EEHV viremia with cTnI suggests these markers might be used in conjunction to help predict when EEHV viremia is likely to progress to EEHV-HD for an individual.

Key Points:
- EEHV targets endothelial cells lining capillaries – leads to catastrophic vascular dysfunction, hemorrhage, edema and often death
– Often have petechia and ecchymosis of the epicardium, endocardium and myocardium, pericardial effusion, vasculitis, and changes to other internal organs
– Gold standard diagnosis – qPCR of EDTA whole blood
– Low level viremia – may be subclinical primary infection, transient recrudescence that won’t progress to EEHV-HD or subclinical EEHV-HD that can occur up to 30d before clinical signs
- Cardiac troponin 1 – not present in the serum unless destruction of cardiomyocytes
– Prev report of increase in cTn1 in elephant with clinical EEHV which then decreased after recovery from EEHV - suggests lack of persistent viral damage to the heart
- Objective – determine if CTn1 can be used as early detection for EEHV HD
- A significant relationship was seen between level of viremia and an elevated cTn1
– The presence of detectable cTn1 was associated with having clinical signs of EEHV HD
– cTn1 is associated with EEHV viremia regardless of the level of viremia (low or high) indicating that even in subclinical cases there is likely some level of cardiac damage
– Consider low level viremia with elevated cTn1 as a clinical case due to potential for cardiac damage
– Animals with EEHV1 viremia were more likely to have a detectable cTn1 value
– Higher EEHV1 viremia values were associated with higher cTn1 values
– No association of dying of EEHV and cTn1 level
– Elephants with a prior episode of EEH HD were more likely to experience elevated cTn1 during subsequent viremic episode
- No clear association between EEHV5 and cTn1 levels – no other strains detected in this study
- cTn1 may also be useful for cardiac disease in non-EEHV viremic elephants

Take-Home Message:
- cTn1 increases with EEHV viremia and decreases with resolution of viremia.
- May be useful to follow up with cTn1 when low-level EEHV viremia is identified through routine monitoring

Answer 107

A

POINT-OF-CARE AND STANDARD LABORATORY REFERENCE INTERVALS FOR COAGULATION VALUES IN ASIAN ELEPHANTS (ELEPHAS MAXIMUS) VARIATION BY AGE CLASS, SEX, AND TIME TO CENTRIFUGATION
JZWM 2022 53(2) 291-301

Abstract: In Asian elephants (Elephas maximus), elephant endotheliotropic herpesvirus causes significant calf mortality. Coagulation testing may aid veterinarians in early identification and management of hemostatic disorders. This study sought to establish reference intervals for select coagulation and platelet values. Blood was collected from clinically healthy Asian elephants (n=63) in juvenile (15 yr old, n =9), adult (.15 to 50 yr old, n = 41), and geriatric (.50 yr old, n = 13) age classes at seven institutions in Kanchanaburi Province, Thailand. Activated clotting time (ACT) was immediately assessed with a handheld analyzer, whereas remaining blood was stored at 58C in sodium citrate and potassium EDTA collection tubes and transported to a central laboratory. Coagulation values were assessed on an automated blood coagulation analyzer, and platelet values were assessed on a hematology analyzer. Reference intervals were established for ACT, prothrombin time, activated partial thromboplastin time, thrombin time, fibrinogen, platelet count, mean platelet volume, platelet distribution width, and plateletcrit according to the American Society for Veterinary Clinical Pathology guidelines. No significant differences were observed for any value when comparing sex and time to centrifugation. Plasma fibrinogen (P = 0.002) and platelets (P = 0.003) varied significantly by age class, with adults displaying the highest fibrinogen concentrations and geriatric individuals displaying the lowest platelet counts. The ACT kaolin cartridges resulted in high success rates (84.3% feasibility) compared with celite cartridges (4.8% feasibility). Further studies are warranted to stratify reference intervals in accordance with age class trends.

Intro
- Fifty-three percent of elephant calf mortality under human care in North America since 1980 is attributed to elephant endotheliotropic herpesvirus (EEHV), a disease causing acute fatal hemorrhage.
- Rapid assessments of hemostatic function, as well as laboratory analyses, may assist veterinarians in monitoring case progression and identifying at-risk conspecifics
- This study sought to determine reference intervals for point-of-care activated clotting time (ACT), as well as laboratory-measured prothrombin time (PT), activated partial thromboplastin time (aPTT), thrombin time, fibrinogen, platelet count, mean platelet volume (MPV), platelet distribution width (PDW), and plateletcrit (PCT)
- Also evaluating between sex and age classes

M&M
- Asian elephants under human care from seven institutions in Kanchanaburi Province, Thailand
- n=63. 3 age classes: juvenile, adult, geriatric
- ACT measured with point of care analyzer
- All other samples analyzed 24–31 h after venipuncture at the central laboratory
- PT, aPTT, thrombin time, fibrinogen, platelets, MPV, PDW, and PCT were measured

Results
- 21 samples for ACT were taken and all but one had cartridge failure, so they discontinued that measurement and switched to ACT kaolin cartridge which had a higher rate of success
- Reference intervals for all analytes are in the table below
- Differences between sexes for any coagulation value were not significantly different
- Differences in fibrinogen concentration, platelet counts, and PCT were deemed significant between age classes
– Fibrinogen was different between juveniles and adults as well as adults and geriatrics. Adults had the highest concentration
– Human and dog literature suggests fibrinogen increases with age
– Acute phase protein, may indicate low level inflammation not detected on PE
– Platelets were different between juveniles and geriatrics. Lowest in the geriattric cohort
– In humans and laboratory mice, platelet count decreases in geriatric individuals, in agreement with the presented data from Asian elephants.
– Although differences in PCT were deemed significant on initial statistical evaluation, adjustment for multiple comparisons did not reveal any significant differences between age classes
- Time between collection and centrifugation was assessed and no significant differences were found

Discussion
- In this study, celite cartridges for measurement of ACT in a subset of animals resulted in a low rate of feasibility (4.8%), whereas ACT kaolin cartridges resulted in a high rate of feasibility (84.3%).
– In Asian elephants, aPTT is significantly shorter than reported values for domesticated large animals and more closely resembles canine values
- This study establishes point-of-care and laboratory coagulation reference intervals by sampling a large population of Asian elephants in human care

Answer 108

A

JZWM 2022 53(4) 654-660
EVALUATION OF PLASMA CARDIAC TROPONIN I IN ASIAN ELEPHANTS (ELEPHAS MAXIMUS) USING TWO CLINICAL ANALYZERS

Abstract: Cardiac troponin I (cTnI) is specific to myocardial tissue, highly conserved across taxa, and a reliable indicator of myocardial disease in human and veterinary medicine. Biomarkers, like cTnI, may be useful for cardiac evaluation of elephants because the application of other modalities is complicated by the size of the animal. The goal of this study was to establish observed ranges for plasma cTnI in Asian elephants (Elephas maximus) measured by two point-of-care analyzers. Blood was collected from captive juvenile (15 yr; n=9), adult (16–50 yr; n = 42), and geriatric (.50 yr; n = 16) elephants. Following centrifugation, heparinized plasma was stored at 58C prior to and in between analyses on iSTAT (Abbott Point of Care Inc, Princeton, NJ 08540, USA) and HUBI-QUANpro (Humiasis Co, Ltd, Anyang-si 14042, South Korea) analyzers. With the exception of two results, plasma concentrations of cTnI were below the limit of quantification (LOQ , 0.05 ng/ml) for the HUBIQUANpro (n = 64), which prohibited comparison between the two analyzers. Observed ranges were determined for plasma cTnI concentrations reported by the iSTAT for the entire population sampled (n = 58; mean 0.011 ng/ ml; SD 6 0.013 ng/ml; range 0.00–0.07 ng/ml; 95% CI 0.008–0.015 ng/ml; median 0.01 ng/ml) and with outliers excluded (n = 50; mean 0.007 ng/ml; SD 6 0.007 ng/ml; range 0.00–0.02 ng/ml; 95% CI 0.005–0.009 ng/ml; median 0.01 ng/ml). No significant differences were observed between age classes (P = 0.70) or sexes (P = 0.34). Higher cTnI concentrations were significantly correlated with increasing age (Pearson’s r = 0.426; P = 0.002). Future studies are warranted to investigate the diagnostic potential of plasma cTnI in Asian elephants

Intro
- Cardiac troponin I (cTnI) is one of three proteins in the troponin complex, which binds to myocardiocyte actin molecules to regulate calcium binding and actin-myosin interaction
– Damage to myocardiocytes causes cTnI to dissociate from actin molecules, leak into the extracellular space, and enter circulation
– Because of its specificity to myocardial tissue, rapid release, and low basal levels in circulation in healthy patients, cTnI is a reliable indicator of myocardial disease
- Several etiologies of cardiac disease have been documented in elephants, including elephant endotheliotropic herpesvirus (EEHV), encephalomyocarditis virus (EMCV), and West Nile virus (WNV), all of which can induce myocardial damage.
- The objectives of this study were to evaluate the agreement between two POC analyzers for the measurement of plasma cTnI concentrations in Asian elephants and to determine the observed range of plasma cTnI concentrations for a captive population of Asian elephants in Thailand

M&M
- n= 64 asian elephants, apparently healthy, blood taken and cTnI measured on each analyzer (HUBIQUANpro and iSTAT)
- Compared between analyzers, age groups and sexes

Results and discussion
- With the exception of two results, plasma concentrations of cTnI were below the lower limit of quantitation of the HUBIQUANpro analyzer, which prohibited investigation of agreement between the two analyzers
- Descriptive statistics were determined for plasma cTnI concentrations reported by the iSTAT
- No significant differences were observed between age classes(??), sexes, or time between centrifugation and analysis
- However higher cTnI concentrations were significantly correlated with increasing age
- With outliers excluded, cTnI concentrations were below the lower limit of detection (below the analytical range) of the iSTAT device and below the LOQ of the HUBIQUANpro device
- For future studies, ultrasensitive assays are recommended as a reference standard against which POC devices may be compared.
- Although all elephants were evaluated by physical examination, hematology, and biochemistry, a comprehensive cardiac examination was not performed.

Takeaway:
- This study measured plasma cTnI concentrations in healthy Asian elephants on two analyzers and provides observed ranges as reported by the iSTAT device.
- cTnI appeared to increase with age, though the underlying cardiac status of all subjects was not fully evaluated

Answer 109

A

J Zoo Wildl Med. 2023;53(4):661-669
DETECTION OF ELEPHANT ENDOTHELIOTROPIC HERPESVIRUS 1A IN ARCHIVAL TISSUE USING RNASCOPE® IN SITU HYBRIDIZATION
Cook KA, Ling PD, Terio KA, Baumgartner WA, Howard LL, Landolfi JA

ABSTRACT: Hemorrhagic disease due to elephant endotheliotropic herpesvirus infection (EEHV-HD) is an important cause of calf mortality in managed and free-ranging Asian (Elephas maximus) and African elephant (Loxodonta spp.) populations. Consequently, infection has profound implications for elephant population growth and sustainability. The mechanisms of disease caused by EEHV (i.e., infection, dissemination, shedding, latency) are relatively undefined, in part because of a lack of robust validated assays for detecting viral gene products in relevant samples. To address this issue, we used RNAscope® in situ hybridization (ISH) based on EEHV1A DNA polymerase and terminase genes to detect EEHV1A RNA in archival formalin-fixed, paraffin-embedded Asian elephant heart and tongue from PCR-confirmed cases (n = 4) of EEHV-HD and Asian elephants (n = 2) that died from other causes. EEHV1A-positive cases had positive hybridization signal in endothelial cell nuclei of both tissues for both DNA polymerase and terminase. EEHV-negative cases lacked signal. In positive cases, the number of positive nuclei was manually assessed to provide an estimate of the viral load and compare sensitivity of the two probes. In all cases, heart had greater signal than tongue for both probes (Wilcoxon rank test; P ≤ 0.01). Overall, terminase hybridization signal was greater than DNA polymerase signal (Wilcoxon rank test; P ≤ 0.01). Results indicate RNAscope ISH is a valuable tool for detection of EEHV in archival samples and for confirming infection. Additionally, the terminase gene is the optimal target and heart is preferable to tongue for detection in cases of EEHV-HD. Results will inform future investigations of viral tropism in EEHV-HD cases due to EEHV1A.

Background:
- EEHV = double-stranded DNA herpesviruses, genus Proboscivirus, subfamily Deltaherpesvirinae
– Seven species of EEHVs have been identified
– Types 1A/1B, 4, and 5 are endemic to Asian elephants
– Types 2, 3, 6, and 7 are endemic to African elephants
– Host adapted and ubiquitous among adult, clinically normal Asian and African elephants
- Disease occurs most often in calves and presents as EEHV-hemorrhagic disease (EEHV-HD)
– Asian calves between 1-8yo & African calves between 6-13yo are most susceptible
– Majority of EEHV-HD mortalities in Asian calves are due to EEHV1A
– Deaths in Asian calves have also occurred with types 1B, 3, 4, and 5
– Similarly, in African calves with types 2, 3, and 6
- Lack of EEHV-HD prior to 1yo suggests some maternal antibody protection
– Cases are likely the result of primary infection rather than reactivation
- PCR = gold standard for detecting viruses in biological samples, most specific and sensitive
– Because samples first undergo digestion to extract nucleic acids, tissue architecture is destroyed, limiting application of PCR for determination of viral tropism
- IHC & in situ hybridization (ISH) visualize protein or nucleic acid, respectively, in intact tissues
– IHC may be limited by species-specific reagent availability
– Similarly, labeling of protein antigen does not necessarily indicate pathogen replication
– Both IHC and conventional ISH can be challenging to apply in archival samples
- RNAscope = multiplex ISH technique with superior specificity and sensitivity vs. conventional ISH
– Uses several short probes complementary to the target sequence
– Can be used with archival samples and ensures selective amplification

Key Points:
- Despite unknown and variable formalin-fixation time, successful hybridization with the positive control probe was achieved in all samples and confirmed RNAscope viability
- Viral DNA polymerase & terminase are highly conserved EEHV genes essential for viral replication
– DNA polymerase is expressed early in viral replication during the lytic phase of infection
– Terminase is expressed later in replication to package the new double-stranded DNA
– These are also the genes targeted in PCR assays for EEHV
- Intranuclear signal for terminase was significantly more abundant than DNA polymerase
– The timing of gene expression may account for this difference
– Expression of late-expressed terminase genes could dominant during end-stage disease
- Vascular lesions in EEHV1A infections are found in the tongue, heart, and liver
– Characterized by endothelial intranuclear inclusion bodies
- Potential explanations for the greater signal in heart tissue vs. tongue include:
– Differences in number of endothelial cells, chronological differences in organ involvement during infection, and disease course
- The pathogenesis of EEHV-HD remains to be elucidated
– The current study reinforced that endothelial cells are the known target in EEHV-HD
– However, endothelial cells are not viable targets to facilitate initial infection, dissemination, latency, or shedding
– Salivary glands and the GI tract may be potential sources of viral transmission
– Salivary glands may be a site of latency
– Mononuclear phagocytic cells may be a method of dissemination

TLDR: RNAscope ISH is a valuable tool for confirming EEHV infection and detection of EEHV in archival samples

Answer 110

A

JZWM 2022 53(4) 670-678
PHARMACOKINETICS OF RECTALLY AND ORALLY ADMINISTERED LEVOFLOXACIN IN ASIAN ELEPHANTS (ELEPHAS MAXIMUS)

Abstract: Appropriate and effective antibiotic use is a critical component of veterinary medicine, but there are variations across species regarding dosage and administration of these drugs. Oral or rectal routes of administration are typically used in elephants, but not all medications can achieve adequate concentrations rectally. The fluoroquinolone antimicrobials are used in elephants because of their favorable antimicrobial spectrum and pharmacokinetics compared with other oral agents. They are commonly used as part of multiple antibiotic regimens for the treatment of tuberculosis (Mycobacterium tuberculosis). The objective of this study was to determine the pharmacokinetic profile of levofloxacin after oral and rectal administration in Asian elephants (Elephas maximus). Dosages of 5 mg/kg orally and 15 mg/kg rectally were evaluated in 13 Asian elephants. Blood was collected at various time points from 0 to 72 h for pharmacokinetic analysis. Pharmacokinetic parameters were determined and reached concentrations above minimum inhibitory concentrations of various bacterial organisms via both routes. A pharmacokinetic-pharmacodynamic assessment was used to estimate appropriate minimal inhibitory concentrations for bacteria that could be potentially treated with this antimicrobial.** Based on these findings, levofloxacin may be a consideration for administration orally (5 mg/kg) and rectally (15 mg/kg) in Asian elephants**. Antimicrobial stewardship principles, culture and susceptibility of suspected pathogens, and blood level monitoring should be used to tailor administration of levofloxacin in this species.

Key Points:

Intro
- Antibiotics in elephants – oral or rectal routes
– Some meds not good for rectal route (poor solubility or bioavailability)
- Mycobacterium tuberculosis – one of most important diseases in elephants
– First line: rifampin, isoniazid, and pyrazinamide
– Secondary therapeutics include the fluoroquinolones (enro historically the most commonly used)
– Oral enro reaches adequate plasma concentrations in elephants, but anecdotally has some side effects (decreased appetite, lethargy)
- Levofloxacin (third generation fluoroquinolone) = L-isomer of ofloxacin
– Undergoes limited metabolism; eliminated almost entirely in urine
– Less side effects than other fluoros, less effect on cardiac QT interval, and safer for patients with liver dz
– Broader spectrum than 2nd generations (enro, cipro) with activity against gram-positive aerobes, some gram-negative aerobes, some anaerobes, and other organisms (Chlamydia, Mycoplasma, Mycobacteria)
- Objective: determine pK properties of levo via oral and rectal routes and potential to tx bacterial infections in Asian elephants

Study
- Thirteen female Asian elephants from 3 different facilities
- Levofloxacin at 5mg/kg (oral) or 15mg/kg (rectal); blood collected 0 – 72 hours
– At least one month between doses

Results
- Peak concentrations: 2 +/-1.8 hours (oral) and 1 +/- 1.5 hours (rectal)
- Half-life: 12 +/- 1.45 hours (oral) and 10 +/- 1.4 hours (rectal)
- Blood concentrations for 72 hours in most elephants from both routes

Discussion
- First time levofloxacin PK reported in this species
- Advantages of levo – excellent activity with wide bacterial spectrum and high oral absorption
– Better than ciprofloxacin (variable and incomplete oral absorption)
– Primary metabolite or enro, also associated with resistance for TB
- More interindividual variability after the rectal dose
– Many factors can affect rectal administration (leakage, inconsistent solubility of drug in vehicle, inconsistent absorption, interindividual variation of rectal environment)
– Despite variability, mean concentrations from rectal were similar to oral concentrations
– Recommend oral > rectal given potential variability
- Times to peak concentration and half-life were similar between routes (1-2 hours; 10-12 hours)
- MIC concentrations from study at these doses in elephants may be sufficient to treatment many Enterobacterales, but not sufficient to treat P. aeruginosa isolates
– Doses attained PK=PD targets effective for MIC values <0.12ug/mL (most of Enterobacterales) and some reported isolates of M. tb, but not all
- No adverse effects. Some elephants passed some blood after rectal administration

Take-Home Message:
- Both oral levofloxacin (5mg/kg) and rectal levofloxacin (15mg/kg) are viable antibiotic options for Asian elephants
- Oral had less variability than rectal route
- Doses attained targets effect for most of Enterobacterales, some M. tb variants, but not P. aeruginosa

Answer 111

A

Journal of Zoo and Wildlife Medicine 53(4): 791–796, 2022
ADAPTATION OF A POINT-OF-CARE CANINE PROGESTERONE TEST FOR USE OF PARTURITION PREDICTION IN CAPTIVE ASIAN ELEPHANTS (ELEPHAS MAXIMUS): PROOF OF CONCEPT
Fieke M. Molenaar, DVM, MSc, DZooMed, MRCVS, Marcus Rowcliffe, PhD, and Andrew Lakey, PhD – Rev by AJC

Abstract: In the Asian elephant (Elephas maximus), the levels of progesterone products 5α-pregnane, 3α-hydroxypregnane, and 17α-progesterone are elevated during pregnancy. Detection of a sudden decrease in blood progesterone product levels in the final days of pregnancy is considered an objective way of predicting impending parturition. Point-of-care (POC) tests eliminate the cost involved in transporting samples to an external laboratory and provide an almost instant result, facilitating decision-making for animal monitoring and management. This proof-of-concept study aims to investigate the ability of the AgPlus POC immunoassay system to measure 4-pregnen-3,20-dione in pregnant elephant serum samples and adapt the method for detection of the preparturient progesterone decrease. Frozen serum samples of two pregnant elephants (N = 82) and fresh serum samples of one pregnant elephant (N = 10) were analyzed using both the POC method and a radioimmunoassay in a reference laboratory. Statistical analysis of the data showed that there was no significant difference between the two methods for detection of the progesterone drop, indicating that the POC method can be considered appropriate for use in elephant parturition prediction. Refinement of the methodology, an increase of sample size, and temporal tandem radioimmunoassay would be required to further validate this method for use in elephant reproductive management.

Key Points:
- Parturition prediction based on gestation in Asian elephants not reliable (published 637-686 days; actually 630-700 at this institution) 🡪 most accurate/objective is measuring blood progesterone products (drop of 5α-pregnane & 17α-progesterone to baseline) & visualizing transrectal US dilation of cervix (12-24 hr warning)
- No POC test for elephant progesterone products 🡪 comparing AgPlus canine progesterone POC w/ validated laboratory assay (for dogs) by using frozen and fresh serum samples of 3 pregnant elephants
- Both AgPlus POC & laboratory assays poorly correlated, but both measured substantial drop in progesterone products w/in 5 days of birth
– Both analyzers able to establish an “average” of progesterone products prior to the drpo
– Measurements showed variation and some early drops in progesterone products long before parturition, which rapidly returned to previous levels (“false alarms”)
– “False alarm” measurements count be from natural hormonal fluctuations, sample storage, handling error, technical artefact 🡪 should obtain at least 1 more sample after a single low measurement
- AgPlus POC overall helpful to observe progesterone trends, but does require serial blood samples

Take Home Message:
- POC progesterone analyzers should be able to monitor progesterone trends and detect a progesterone drop prior to parturition.
- However, it requires serial blood samples and some early progesterone drops (false alarms) were detected.

Answer 112

A

JZWM 2023 53(4):801-810
Relationship Between Reproductive And Bone Biomarkers And Osteoarthritis In Zoo Asian (Elephas maximus) And African (Loxodonta africana) Elephants
Chusyd DE, Brown JL, Golzarri-Arroyo L, et al

ABSTRACT: Osteoarthritis (OA) is common in zoo Asian (Elephas maximus) and African (Loxodonta africana) elephants. This study investigated the relationship between confirmed or suspected OA with ovarian cyclicity, gonadotropins, progestagens, luteinizing hormone (LH), follicle-stimulating hormone (FSH), and collagen type I (CTX-I) in zoo elephants. In Asian elephants, odds of having confirmed or suspected OA decreased with cycling (OR = 0.22, P = 0.016; OR = 0.29, P = 0.020, respectively), however, not when adjusted for age (odds ratio [OR] = 0.31, P = 0.112; OR = 0.58, P = 0.369, respectively). In African elephants, none of the models between confirmed OA and cycling status were significant (P > 0.060), while the odds of having suspected OA decreased with cycling (OR = 0.12, P = 0.001), even after adjusting for age (OR = 0.15, P = 0.005). Progestagens (Asian elephants P > 0.096; African elephants P > 0.415), LH (Asian P > 0.129; African P > 0.359), and FSH (Asian P > 0.738; African P > 0.231) did not differ with confirmed or suspected OA status, unadjusted. CTX-I concentrations were not related to OA status (P > 0.655). This study concluded hormonal changes may not have a strong impact on OA, so additional investigation into other serologic biomarkers is warranted.

Background:
- 36% of institutions in North America that hold elephants had at least one elephant with arthritis
- Repro status (esp. menopause) has been consistently associated with arthritis in women
– Endogenous sex hormones may play a protective role against OA
– Many zoo elephants exhibit abnormal ovarian cycles with baseline progestagen patterns similar to those of postmenopausal women
- Hard surfaces have been associated with overall poor elephant foot and joint health
- Serum CTX-I generated during bone loss represent a potential marker of bone catabolism

Key Points:
- In both species, noncycling elephants were more likely to exhibit OA
– This relationship was largely explained by age, as elephant OA increased with age
– Older elephants also are more likely to be acyclic
- No association between CTX-I with OA status in either elephant species
– CTX-I levels higher in African elephants, likely due to larger stature and weight

TLDR:
- CTX-I was not useful in identifying elephants with OA in this study.
- No associations were observed between serum progestagens, LH, or FSH with OA status

Answer 113

A

Zoo Biology 41(5): doi: 10.1002/zoo.21737, 2022
Predictors of testosterone in zoo‐managed male African elephants (Loxodonta Africana)
Kaitlyn M Campbell 1 2, James A Wilson 1, Kari A Morfeld 2 – Reviewed by AJC

Abstract:
Reproductive complications for both male and female zoo-managed African elephants (Loxodonta africana) contribute to the rapidly declining population. In zoo-managed bull elephants, few studies have explored the potential physiological, physical, social, and environmental factors that influence bull fertility, particularly, androgen production. Testosterone is the essential steroid hormone for male sexual maturation and inadequate concentrations can be detrimental for spermatogenesis. Testosterone, fecal glucocorticoid metabolites, leptin, glucose, insulin, and triglycerides were analyzed from weekly fecal and blood serum samples taken over 6 months from six zoo-managed African elephant bulls (10-19 years of age). Testosterone levels were compared to endocrine factors, weekly social and environmental variables, daily musth signs, and body condition scores (BCS). The glucose-to-insulin ratio (G:I) was the only physiological biomarker found to be positively associated with testosterone. Predictive physical variables included Musth Score (+) and Moderate Exercise (+). Bulls with BCS signifying overweight (BCS 4) had lower testosterone (36.6 ± 1.6 ng/g fecal extraction [FE]) than bulls with healthy BCS 3; 51.2 ± 4.9 ng/g FE). Numerous social variables influenced testosterone concentrations, including Total Contact Day (+), Female Interaction Day (+), Indirect Contact Day (+), Indirect Contact Night (+) and Total No Contact (-). Both percentage of Time Outdoor and Time Mixed positively influenced testosterone, whereas testosterone decreased for percentage of Time Indoors.** Each additional daily browse opportunity increased testosterone by approximately 7 ng/g FE**. In managed care, the emphasis should be placed on optimizing these predictors of testosterone production to promote bull reproductive health.

Key Points:
- Without extensive efforts to increase African elephant breeding success, the zoo-managed population will not be sustainable beyond 50 years.
- Failure of male breeding potentially due to lack of opportunity & history of poor semen quality
- Testosterone – steroid hormone, produced by Leydig cells
- Acts on nuclear receptors 🡪 seminiferous tubules to promote spermatogonia differentiation & growth
- Aim of study = explore psychological, physical, social and environmental factors that influence testosterone
- Six male African elephants from four institutions (10-19 years) = adolescents

Results
- HIGHER testosterone levels seen with
– Higher glucose:insulin ratio
– Higher Musth score
– Hours of moderate intensity exercise
– More female interaction time
– More total elephant (M + F) contact time
– More time spent outside
– Increase in daily browse
- LOWER testosterone levels seen with
– Higher BCS
– More time spent inside
- No significant change in testosterone levels with
– Triglycerides, fecal corticosteroids, and leptin (hormone released by adipose, suppresses food intake)
– Hours of sleep per night
– Hours of high intensity exercise

Discussion
- In contrast to previous study, age and weight were not predictors of testosterone concentration in this study
- All bulls were adolescents (10-20 years), considered sexually mature but may not reproduce until closer to 30 years of age in free-ranging populations
- Limitation – small sample size
- Indirect contact also positively correlated with testosterone
- Indoor spaces tend to be smaller than outdoor spaces, which may explain that result
- High testosterone does not necessarily assure fertility, but provides info on optimizing bull health

Take-Home Message:
- Healthy body condition, both visual and metabolic fitness, is correlated with testosterone production
- Musth scores rise in relation to testosterone levels
- Social interaction positively influences androgen concentrations
- Percentage of time spent in indoor space negatively impacts testosterone, whereas time outside positively impacts testosterone
- Positive relationship between daily browse and testosterone

Answer 114

A

The historical development of zoo elephant survivorship.
Scherer L, Bingaman Lackey L, Clauss M, Gries K, Hagan D, Lawrenz A, Müller DW, Roller M, Schiffmann C, Oerke AK.
Zoo Biology 2023;42(2):328-338

In the discussion about zoo elephant husbandry, the report of Clubb et al. (2008, Science 322: 1649) that zoo elephants had a “compromised survivorship” compared to certain non-zoo populations is a grave argument, and was possibly one of the triggers of a large variety of investigations into zoo elephant welfare, and changes in zoo elephant management. A side observation of that report was that whereas survivorship in African elephants (Loxodonta africana) improved since 1960, this was not the case in Asian elephants (Elephas maximus). We used historical data (based on the Species360 database) to revisit this aspect, including recent developments since 2008. Assessing the North American and European populations from 1910 until today, there were significant improvements of adult (≥10 years) survivorship in both species. For the period from 1960 until today, survivorship improvement was significant for African elephants and close to a significant improvement in Asian elephants; Asian elephants generally had a higher survivorship than Africans. Juvenile (<10 years) survivorship did not change significantly since 1960 and was higher in African elephants, most likely due to the effect of elephant herpes virus on Asian elephants. Current zoo elephant survivorship is higher than some, and lower than some other non-zoo populations. We discuss that in our view, the shape of the survivorship curve, and its change over time, are more relevant than comparisons with specific populations. Zoo elephant survivorship should be monitored continuously, and the expectation of a continuous trend towards improvement should be met.

Key Points
- Found improvement of survival of animals > 10 yo over time for both species
– Suggests continuous progress in zoo elephant husbandry for adult survivorship, not limited to African as previously suggested
- Improvement in survivorship was more evident from 1910-1960 but from 1960 on the improvement was mostly just in African elephants even though overall adult Asian elephant survivorship is higher than African
– Probably an effect of Asian starting higher so less progress to be made
- Asian elephants had significantly higher survivorship but there was significant interaction with species and birth year
- Juvenile survival was not found to have changed from 1960-2019
– Juvenile Asian elephant survivorship significantly lower than African, likely reflects recent impact of EEHV

Conclusions
- Retrospective article looking to refute a previous article’s claim that zoo elephants have compromised survivorship
- Found that adult survivorship of both species in zoos is improving over the last century but juvenile appears static

Answer 115

A

Fowler 10 Chapter 91 EEHV Update

Abstract
- Elephant endotheliotropic herpesvirus (EEHV) = seven types (1 to 7) and subtypes of viruses in the order Herpesvirales, subfamily Betaherpesvirinae, genus Proboscivirus
- EEHVs may cause infection without apparent illness or may cause acute, often fatal hemorrhagic disease (EEHV HD), with high levels of virus in the blood.
– EEHV HD is seen mostly in young (1 to 8 years) Asian elephants (Elephas maximus); morbidity and mortality have also been seen in older Asian elephants and recently in both young (6 to 13 years) and older African elephants (Loxodonta africana).
- EEHV1A has caused the most deaths and disease in Asian elephants, followed by EEHV1B
- EEHV types 2 to 6 have each caused a smaller number of deaths

New EEHV cases in African elephants
- (2) African elephant deaths due to EEHV2 in the United States & (1) from EEHV6 in Thailand
- (2) known survivors of EEHV HD in African elephants, one each from EEHV3B and EEHV6 in the US
- Early indications: range of affected ages is greater in African elephants (up
- to 13 years, as well as one adult [37 years]) compared to Asian (typically 1 to 8 years of age)
– More low level viremias and shedding (both frequency of shedding & individuals seen shedding) in African elephants than in Asian
Therapy
- In addition to historical use of antiviral drugs (eg cyclovirs), new avenues of therapeutics include use of mesenchymal stem cells, fresh & lyophilized platelets, & innate immunity immunostimulants

Monitoring program
- Early aggressive treatment = increase survival due to EEHV HD
- Key components = routine testing by polymerase chain reaction (PCR), daily observations, and routine blood work
– Presence of a low level of EEHV viremia detected by PCR does not necessarily mean that EEHV HD will result
PCR
- Early detection of viral DNA in blood: routine monitoring by either quantitative PCR (qPCR-preferred) or two rounds of conventional PCR (cPCR)
– Low levels may be detected in blood up to a month before clinical signs appear or even be present for up to a year without clinical signs
- If possible, weekly or twice weekly monitoring by PCR is recommended in both Asian and African elephant calves up to the age of 15 years
- Common to detect low levels (<1000 viral genome equivalents/mL or vge/mL) of viral DNA in elephant blood, especially in the African elephants
– Repeat sample may be helpful to determine transient viremia or a rising viremia
- In addition, serostatus, if known, and history of previous EEHV viremias are important pieces to consider when deciding on a schedule of repeat testing.
– Viremia >1000 vge/mL should be tested by qPCR more frequently, up to daily, especially if no known prior history with that EEHV type
- Treatment decisions: based on viral levels and if/how rapidly they are increasing, clinical signs, and hematology
– Do not wait for clinical signs to appear before starting therapy, especially if EEHV1 is present.
– If clinical signs or hematological changes are present, start therapy even if viral levels are low
– If viral levels stabilize at a low level, or as they decline during treatment, testing by PCR may be decreased
- If blood collection is difficult in elephants that have not been well-trained for venipuncture, standing sedation or the use of lancets to collect small amounts of blood should be considered and planned for in advance

Hematology & serum biochemistry
- Individual reference ranges for each animal in the herd should be established, with weekly (young) or monthly (adult) routine complete blood counts (CBC) and differential
- Decreases in leukocytes, monocytes, and thrombocytes may be seen before other outward indications
– Levels of the leukocytes, monocytes, and platelets and the monocyte:heterophil ratio may be used to make treatment decisions and serve as a prognostic indicator
- Anemia and hypoproteinemia may be seen later in the disease course
- Serum amyloid A (SAA) but not haptoglobin (HP) was increased in elephants with EEHV1 >10,000 vge/mL
– Not specific to EEHV–just an indication of an inflammatory process
- Thromboelastography may be useful to determine prognosis
– Done onsite on citrated whole blood
– Not predictive of whether a viremia may progress to EEHV HD or remain as a subclinical infection
- Detectable cardiac troponin I in an elephant with an EEHV viremia may be useful as a predictor of early cardiac damage/EEHV HD and could be useful in treatment decisions

Behavioral/clinical observations
- Calves (up to 15 years) should be observed daily, and even slight changes should be noted and acted on
- Typical early behavioral indications: lameness, changes in sleep patterns (either more or less sleep), changes in mentation or training compliance, inappetence, and lethargy
- Later clinical findings: cyanosis of the tongue, scleral injection, ventral edema, ascites, and tachycardia

Prevalence
- Young Asian elephant shown to shed high levels of EEHV in their trunk secretions about 2 to 4 weeks after recovering from EEHV HD
- Asian elephants (any age) may shed intermittently throughout their lives
- Noninvasive methods to determine shedding = trunk washes, oral swabs, and fecal sampling
– Oral swabs were not as effective as TW for detecting EEHV shedding

Serology
- Specific and sensitive EEHV serology test may identify signs of a previous infection
- Three EEHV specific serology assays for Asian elephants have been described to date.
- Antigen-capture based enzyme-linked immunosorbent assay (ELISA) using an EEHV1A strain glycoprotein B (gB) expressed in bacteria
– Captive Asian elephants in Europe and Thailand revealed high seroprevalence in these populations
– gB proteins from EEHVs known to be endemic within Asian elephant populations, which include EEHV1A, 1B, 4 and 5, are 62% to 87% identical to each other, with many potential common epitopes shared between them
– Most likely a general indication of prior EEHV infection but is unable to
discriminate infection among the four EEHV types
- ELISA assays using gB and glycoproteins H and L (gH/gL) expressed in mammalian cells have been described, which appear to have improved specificity and sensitivity to first generation bacterially expressed antigens
- Luciferase Immunoprecipitation System (LIPS)
– Several advantages compared to other platforms
– Antigens are expressed in mammalian cells and tested in a liquid phase assay potentially preserving native protein conformation
– Detection of IgG doesn’t require species specific reagents
– Preparation of multiple antigens for testing may be done rapidly
– Performs equally or even better than all of these platforms in several systems including parasites, SARS CoV2, and herpesviruses
– EEHV1 species encode several unique antigens with potential for providing specific biomarkers to distinguish prior infections with EEHV1 from 4 or 5
– Protein ORF Q was consistently highly immunoreactive in EEHV1 experienced adult elephants from several captive herds in North America
– Immunoreactivity to ORF Q in the LIPS assay may reveal whether or not an elephant has been infected previously to this EEHV species, which is associated with the highest number of deaths
– ORF Q proteins from EEHV1A and 1B species are sufficiently diverged from each other such that immunoreactivity to each of them can, in some cases, distinguish between infection with these subspecies
— Requires matching the correct protein with the strain known to circulate within the elephant herd being screened, which may not always be known, especially for semicaptive or wild elephants
- Significant antibody levels are passed transplacentally and maintained up to 2 years of age, confirming an earlier report

Sequencing
- Results thus far indicate these viruses
- Form two branches of AT-rich and GC-rich genomes
- Fall into a unique and novel genus known as the Proboscivirus genus
- Encode genes that represent nearly half of an estimated 118 genes in their genomes that are unique to this genus
- EEHV cases found in elephants in human care in Western countries were caused by viruses carried from elephants imported from Asian range countries
- Disease has a sporadic pattern rather than epidemic or zoonotic

Answer 116

A

Fowler 10
Chapter 94: Artificial Insemination in Elephants
Pp 655-659

Successful repro of both Asian and African elephants in captivity is compromised by lack of suitable breeding bulls, decreasing genetic diversity, and increasing female infertility triggered mainly by prolonged nonrepro cycle activity
Semen collection and cryopreservation, monitoring female sexual cycle activity via hormone profiles and US assessments, ID treating pathologies of females repro tract, and AI, pregn monitoring and vet assistance during parturition assist captive breeding
AI under US and endoscopic guidance nonsurgical technique that may be entirely performed without anesthesia in conditioned animals

Introduction
- Total numbers of African and Asian elephants declining both in wild and in captivity, including zoo pops
- African- vulnerable, Asian- endangered
- Keeping elephant bulls more costly, risky, and logistically challenging compared to females, sex ratios in zoos biased towards females
- Only few captive males produce sperm of suitable quality, mainly due to age and management factors
- Sperm motility, conc, viability and morphology may substantially fluctuate b/t days for unknown reasons
- Results in low birth rates and shrinking genetic diversity
- Translocation of elephants logistically challenging, etc. and disruptive to social bonds
- AI advantageous in this regard
- Can do procedures without chemical restraint in well trained animals
- Standing sedation can be done

Methods

Reproductive Assessment of Male and Female Elephants
- Sexual maturity in males Asian- 6yrs, african- 8yrs
- In wild, males do not sire offspring before manage of 25 dye to strong hierarchy which implies social suppression
- Sexual maturity of female may be reached at age of less than 4yrs in captivity, but not before 10-12yrs in wild (access to high quality food in captivity)
- Transrectal ultrasound to see ovarian processes such as follicular development and CL formation
- Usually require probe extension
- Male repro- intra abdominally located tests and bulbourethral glands, prostate, seminal vesicles and ampullae
- Nulliparous female tract about 3m long
- Vestibulum average length of 1.3mm, curves from vertically to horizontally, terminating in cranial sac above the bony pelvis into vaginal os- measuring merely 4x2mm- ends flanked by two blind pouches of similar outer appearance that may complicate AI
- Architecture vaginal os changes permanently in primiparous females- two blind pouches disappear, vaginal opening is dilated to 1-5cm depending on time since previous birthing
- Ensuring vagina serves as natural site for semen and oustide estrus as mucus-filled, protective barrier.
- 15cm long cervix followed by 0.8-1.5-m long bicornuate uterus with a very short corpus uteri and elongated horns leading to the comparably small ovaries
- Twins rare, bear high risk of complications
– Located in separate uterine horns, they may show different developmental stages, derived from two independent cycles
- Repeated noncycling normally does not occur in wild, usually either pregnant or lactating with an interbirthing interval of 3-7yrs
- Frequent cycling activity in captive females associated with pathology, compromising fertility- vestibular cysts, endometrial hyperplasia both Asian/African
– Nulliparous >30yo, vaginal and cervical cysts and neoplastic alterations may fill entire vaginal lumen and interfere with semen transport/mating
– Vestibular polyps occur mainly in nulliparous African in 70% of captive females at age 30 or older, leiomyomata found only in Asian elephants
– Paraovarian cysts, ovarian cysts, with or without hormonal activity, oviductal cysts may block passage of ovulated oocyte to uterus
– Causally linked to nonrepro cycles, frequently encountered in captivity
* Rec to put postpuberty female into suitable breeding or consider AI

Hormone Profiles
- Anovulatory luteinizing hormone (LH) peak → results in formation accessory CLs, followed by ovulatory LH pak
- Delay b/t both averages about 20 d, useful for preparing for an AI
- Ovulation of single Graafian follicle takes place about 12-24 hr about the ovulatory LH peak.
- Most suitable hormones for cycle determination serum LH and progestogens, whereas estrogens, inhibit, FSH and PRL are less informative
- 5alpha-reduced pregnanes major circulating luteal steroid as compared to progesterone in most other mammalian species

Semen collection
- Obtain from vestibule after copulation, stimulation with an estrous female, artificial vagina, electroejaculation, manual rectal stimulation
- Standing sedation protocols
- Electroejaculation requires anesthesia
- Sperm sex sorting- small difference in DNA content b/t X- and Y- chromosome bearing spermatozoa in many species
- Attempts in elephants have been hampered by scarcity of sorting machines= small insemination doses

Semen cryopreservation
- Fresh, chilled, frozen-thawed semen
- TEST-fructose-lactose-egg yolk extender Berliner Cryomedium yielded best results for chilled storage and cryopreservation of sperm from both species
- Best preservation of post thaw sperm motility achieved by cryopreservation using directional freezing technique after seminal fluid removal by centrifugation and action of 7% glycerol
- Extends flex in distance, time, and optimal genetic pairing- removes risk of faulting to obtain donor when needed
- First successful AI in captive Asian after 55 attempts resulted in pregnancy, but not in live birth
- First calf born after AI with frozen-thawed semen of a wild African bull resulted in live birth of calf at Vienna Zoo in 2013

Artificial Insemination
- First attempts surgical approach via vestibulum, etc.
- Current method- hormonal or US ID of ovulatory Graafian follicle
- 1-4 consecutive inseminations per cycle may be performed over course of 48 hours if sedation not required
- Probability of insemination success decline after ovulation has taken place- ultrasound to ID the disappearance of the Graafian follicle from ovary, appearance of early luteinized follicle (corpus hemorrhagicum)
- Insert catheter into vestibule, via endoscopy with 3m flex scope and transrectal US
- Ai catheter used for insemination
- Large proportion of male calves born from AI, excessive use of high-caloric treats given to elephant during Ai prep and procedure resulting in high BG conc maybe favoring males
- Rec maintain in lean body condition

Pregnancy Monitoring
- Gestation 20-23 months
- Diagnose via ultrasound, at earlier at 8 weeks after conception
- Elevant hormones monitoring in serum, urine, or feces
- Decline progesterone at 8-9 wks and 60 wks gestation, avoid stress during this time
- Sexing of calf possible at 12 months of pregnancy either on testosterone in maternal blood serum in Asian or in both African/Asian by transrectal US of pregnant cow in lateral recumbency
- Birth expected to occur at night within 2-5 days of drop of 5alpha-reduced pregnane levels to baseline and within 24hr after loss of complete vaginal mucus plug

Birth management
- Peripartal fatalities in both elephant cows and offspring frequent
- Disruption of matriarchic herd structure by frequent exchange of elephants b/t facilities
- Experienced females help to avoid fatal attacks of the mother towards its calf shortly after painful birth experience, which may occur primarily in primiparous females
- Absence of stable social support- role taken on by veterinarians and keepers

Conclusion
- Need AI right now to be sustainable populations

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