Introduction

A 12-year-old 101.5-kg sexually intact male zoo-managed Sumatran tiger (Panthera tigris sumatrae) was evaluated at the University of Illinois Veterinary Teaching Hospital (VTH) for a 3-day history of vomiting, hyporexia to anorexia, and lethargy. The tiger arrived at the zoological institution 15 months prior to the onset of clinical signs and had been otherwise healthy. Routine preshipment testing prior to arrival at the current institution was unremarkable and included a CBC, serum biochemistry panel, infectious disease serology (feline leukemia virus, feline immunodeficiency virus, feline calicivirus, Toxoplasma gondii, and heartworm), and rectal culture. The tiger was housed alone in an exhibit that included indoor and outdoor access with a freshwater pool (3 × 1.5 × 1.5 m) available in the outdoor enclosure, and was fed 3.6 kg of commercial carnivore diet per day.

The tiger was immobilized at the zoological institution by IM dexmedetomidine (0.013 mg/kg) and midazolam (0.05 mg/kg) administration using a plastic dart via CO₂-powered pistol. Seventeen minutes after initial drug delivery, the tiger was heavily sedated, and ketamine (3 mg/kg) was remotely delivered IM. After onset of general anesthesia, the tiger was intubated (20-mm-internal-diameter cuffed endotracheal tube), crated, and transported to the VTH for additional diagnostic tests. During transport, supplemental ketamine administered IM and isoflurane administered via endotracheal tube were provided to maintain an appropriate anesthetic plane. Intermittent positive pressure ventilation was provided as needed to address hypercapnia.

Upon arrival to the VTH, an IV catheter was placed in the left medial saphenous vein during radiograph acquisition and isotonic IV fluids were administered at an open drip rate throughout diagnostic evaluation, surgery, and transport back to the zoological institution for a total volume of 2 L administered. Prior to surgery, an arterial catheter was placed in the left dorsal pedal artery and utilized for direct blood pressure monitoring and serial arterial blood gas analysis. Blood gas analysis revealed hypoglycemia and a trend toward hyperkalemia (potassium concentration, 4.47 mmol/L at 57 minutes and 5.49 mmol/L at 177 minutes); therefore, 5% dextrose was supplemented in the IV fluids (blood potassium concentration, 4.66 mmol/L at 252 minutes). Hypotension was persistent and a dopamine continuous rate infusion (5 to 12 μg/kg/min), partial reversal of α-2 adrenoceptor agonists with atipamezole (0.025 mg/kg, IM, once), and ephedrine (1 mg/kg, IV, once) were required to maintain perfusion. Buprenorphine (0.005 mg/kg, IV, once) and pantoprazole (1 mg/kg, IV, once) were administered postoperatively. The tiger was transported back to the zoological institution and reversed with atipamezole (0.075 mg/kg, IM, once); recovery was unremarkable. Total anesthesia time was 7 hours 27 minutes.

Initial abnormal physical examination findings included a right mandibular canine tooth fracture, moderate dental calculus and gingivitis, and a midcaudal deep abdominal mass approximately 5 cm in diameter. Mucous membranes were initially pink and moist and skin was initially unremarkable. Both mucous membranes and skin became progressively icteric throughout anesthesia.

Three-view abdominal and thoracic radiographs were obtained and reviewed by a board-certified veterinary radiologist. The thorax study was unremarkable. Abdominal radiographs revealed 2 populations of small intestine with gas distended loops up to 4.3 cm in diameter, compared with normal loops up to 1.6 cm in diameter. Abdominal serosal detail, stomach size and placement, and hepatic, splenic, and renal silhouettes were unremarkable. An abdominal ultrasound revealed a small amount of anechoic free fluid between the liver lobes, a moderately distended stomach with decreased motility, and a large amount of layered sediment in the urinary bladder. A CBC and serum biochemistry panel were submitted, revealing a markedly elevated total serum bilirubin concentration of 6.0 mg/dL. The remainder of the values were unremarkable based on reference ranges provided by Species360 Zoological Information Management System (Supplementary Table S1).

Based on initial diagnostic test results, an exploratory laparotomy was recommended to rule out and potentially treat a foreign body obstruction. The tiger was placed in dorsal recumbency, the ventral abdomen was clipped and aseptically prepared using chlorhexidine and sterile saline. A routine 20 cm ventral midline incision was created to expose the abdominal organs. Upon exposure of the abdomen, a moderate amount of effusion was present, and a sample was collected for cytology. The small intestines and colon were thoroughly examined for the presence of a foreign body or other obstructive lesion. None were noted; however, the intestines were diffusely stained yellow from the effusion. The surface of the liver was diffusely covered with small tan foci. The gallbladder, kidneys, and pancreas were grossly normal.

Cytology of the abdominal effusion was consistent with a suppurative exudate, and aspirates of the liver were consistent with mild neutrophilic inflammation (results available on day 0). Based on the liver abnormalities and jaundiced mucous membranes, top differential diagnoses included cholangiohepatitis alone or in conjunction with triaditis, feline infectious peritonitis or neoplasia.

On day 2, the tiger was reimmobilized due to continued anorexia, lethargy, and inability to medicate with oral medications. The tiger was immobilized at the zoological institution with dexmedetomidine (0.013 mg/kg, IM) and midazolam (0.05 mg/kg, IM) using a plastic dart via CO₂-powered pistol. Twenty-two minutes after initial drug delivery, the tiger was heavily sedated, and ketamine (3 mg/kg) was remotely delivered IM. After onset of general anesthesia, the tiger was intubated (20-mm-internal-diameter cuffed endotracheal tube). An IV catheter was placed in the right medial saphenous vein. Physical examination revealed persistent icteric mucous membranes and paw pads as well as marked peri-incisional swelling and bruising. Blood was collected from the IV catheter, and a sterile urine sample was collected via urethral catheterization.

The tiger received 3 L of isotonic crystalloids fluids IV and 4 L of isotonic crystalloid fluids with vitamin B complex SC, buprenorphine (0.05 mg/kg, SC once), N-acetylcysteine (70 mg/kg, IV, once), ampicillin (3 mg/kg, IV, once), pantoprazole (1 mg/kg, IV, once), famotidine (1 mg/kg, IV, once), maropitant (1 mg/kg, SC, once), cefovecin (8 mg/kg, SC, once), and dexamethasone SP (0.1 mg/kg, IM, once). Oral medications administered via orogastric tube included ursodiol (15 mg/kg, once), mirtazapine (0.15 mg/kg, once), and capromorelin (2 mg/kg in 500 mL of water, once). Lactulose (5 mL/kg) was administered rectally with an 18F red rubber catheter. Atipamezole (0.1 mg/kg) was administered IM as reversal 93 minutes after immobilization, and recovery was unremarkable.

A majority of the diagnostic tests of samples collected during surgery became available over the following days. Histopathology of the liver (results available on day 2) revealed acute hepatocellular dissociation, microvesicular degeneration with single-cell necrosis and occasional intrasinusoidal fibrin thrombi, mild lymphoplasmacytic portal hepatitis, and early peribiliary fibrosis. Top differential diagnoses based on these results were hepatic toxicosis, leptospirosis, and feline infectious peritonitis. Concurrent infectious disease testing and examination of the indoor and outdoor enclosure for fungi, mold, necrotic debris, and pesticides occurred. Significant mold growth was found on a log in the enclosure and identified as non-toxic Fuligo septica, otherwise known as the dog vomit slime mold, by the Miller Mycology Lab, University of Illinois.

After the histopathology results were obtained microscopic agglutination testing (MAT) for Leptospira serovar antibodies was requested using serum collected the day of surgery (results available on day 3). Initial titers against Leptospira kirschneri serovar Grippotyphosa were 1:400, and the remaining serovars (Autumnnalis, Bratislava, Canicola, Hardjo, Pomona, and Icterohemorrhagiae) were negative. Immunohistochemistry for Leptospira spp (positive) and feline coronavirus antigen (negative) were also submitted after obtaining the histopathology results (results available on day 21). Aerobic and anaerobic cultures of the liver and gallbladder were negative (results available on day 3).

The blood and urine collected during the second immobilization (day 2) were submitted for additional testing. The Leptospira kirschneri serovar Grippotyphosa titer increased to 1:3,200 supportive of an active infection (results available on day 5). A Leptospira real-time PCR assay capable targeting the rrs (16S) gene¹ was performed and was negative (results available on day 3). Standard aerobic urine culture results were negative; a culture specific for the growth of Leptospira spp was not attempted. Significant urinalysis results included 2+ protein and 3+ bilirubin with a urine specific gravity of 1.044 (results available on day 3); a urine protein-to-creatinine ratio was within normal range per domestic cats (results available on day 3); serum albumin concentrations were also unremarkable (3.2 g/dL, Supplementary Table S1), so further diagnostic evaluation of the proteinuria was not pursued. The results of the CBC and biochemistry panel (results available on day 3) revealed that the tiger had developed a normocytic (mean cell volume, 56.8 fL), normochromic (mean cell hemoglobin concentration, 33.1 g/dL), anemia (Hct, 23.9%; RBC count, 4.2 × 10⁶/μL), and absolute neutrophilia with a left shift (absolute neutrophil count, 22.56 × 10³/μL; absolute band neutrophil count, 0.24 × 10³/μL). At that time, serum total bilirubin concentration remained elevated (5.6 mg/dL).

Based upon the results of the liver biopsy and Leptospira spp titers, the patient’s oral medications were adjusted to amoxicillin–clavulanic acid (10 mg/kg, PO, q 12 h for 5 days), mirtazapine (0.15 mg/kg, PO, q 48 h), ursodiol (15 mg/kg, PO, q 24 h), tramadol (2 mg/kg, PO, q 12 h; discontinued on day 8), and doxycycline (5 mg/kg, PO, q 12 h for 21 days; began on day 6). By day 10, the ursodiol and mirtazapine were discontinued. The tiger’s appetite had returned to normal by day 6, and therefore antimicrobial administration compliance was high for the entire doxycycline course of treatment.

Due to zoonotic concerns, the results of the liver biopsy received on day 2 and subsequent positive Leptospira spp titer received on day 3 prompted immediate implementation of biosecurity protocols for staff and the tiger’s indoor and outdoor enclosures. Masks, gloves, and limiting personnel interactions with the tiger were instituted for animal care staff. Potassium peroxymonosulfate foot baths were already part of routine sanitation procedures. The tiger was not allowed outdoor access to minimize guest exposure. The indoor enclosure, pool, and water bowls were drained and cleaned with 10% bleach daily.

Three negative urine Leptospira real-time PCR¹ assay results were acquired (days 15, 45, and 65) from floor collected urine obtained by zoo personneL After the third negative test, routine cleaning protocols were resumed: cleaning the enclosure twice weekly with a quaternary ammonium disinfectant, once weekly cleaning with a degreaser, and every other week with bleach. On nondisinfecting days, the enclosure was hosed with water. The pool continued to be drained and bleached daily.

During immobilization for an unrelated examination on day 161, an opportunistic urine Leptospira real-time PCR¹ assay, CBC, and serum biochemistry panel were submitted; the results of the CBC and biochemistry were unremarkable and the urine PCR assay was negative. No further follow-up testing was submitted until a wellness examination on day 595. At that time, a CBC, serum biochemistry panel, urinalysis, standard aerobic urine culture and urine protein-to-creatinine ratio were submitted and unremarkable. However, the results of a submitted urine Leptospira real-time PCR¹ assay were positive. After this result was obtained, the Leptospira spp titer panel was submitted from remaining serum collected on day 595; all 7 serovars were negative.

Due to a concern urinary shedding and out of caution for zoonotic disease, on day 599, doxycycline (5 mg/kg, PO, q 12 h for 14 days) was instituted to treat leptospirosis. A recheck urine Leptospira real-time PCR¹ assay was negative on day 602, positive again on day 611 and doxycycline was continued for an additional 14 days. Posttreatment urine Leptospira real-time PCR¹ assay on day 636 was again positive; therefore, amoxicillin (22 mg/kg, PO, q 12 h for 21 days) and doxycycline (5 mg/kg, PO q 12 h for 21 days) were prescribed. Repeat urine Leptospira real-time PCR¹ assays on day 658, 678, and 699 were negative.

From day 595 through day 699, increased biosecurity measures and cleaning protocols were performed as previously described. An isolated episode of hematuria observed by keepers on day 882 prompted rechecking a free-catch urine sample for urine Leptospira real-time PCR assay targeting the 16S RNA gene² that was negative. The tiger remained without clinical signs during the entire second treatment period and remained that way until last follow-up (day 1,071).

The source of Leptospira spp infection in this case was suspected to be a free-ranging wildlife carrier with exposure occurring in the outdoor portion of the exhibit. A trapping program targeting potential Leptospira spp vectors was instituted and animals trapped on zoo grounds were euthanized and submitted for necropsy and Leptospira PCR¹ assay for the 18 months following this tiger’s diagnosis. Animals submitted included raccoons (n = 6), mice (3), and a single woodchuck; only a single raccoon submitted 16 days after the tiger was first diagnosed had a positive Leptospira real-time PCR¹ assay (renal tissue). Titers were not available on the raccoon to further differentiate which serovar was present.

Discussion

This report describes confirmed leptospirosis in a managed large nondomestic felid in North America. The tiger’s clinical disease was successfully treated, and no zoonotic transmission occurred. However, since the initial diagnosis, the tiger has had several positive urine Leptospira PCR assay results, required continued monitoring, an additional treatment cycle and further zoonotic risks.

Leptospirosis is caused by a spirochete within the genus Leptospira spp; of the 22 recognized species, 10 are considered pathogenic and a variety of serovars are reported.³ The infection has a global distribution and is considered to be able to infect all mammals, however many species exhibit subclinical infections.³ Transmission to felids is typically through urine-contaminated water or soil or ingestion of infected prey; the spirochete enters the blood stream via abraded or intact mucous membranes or conjunctiva. Bacteremia lingers for approximately 7 days in dogs and humans, and this information has been extrapolated to felid species. Clinical disease in non-felids occurs most commonly during the bacteremic phase. Hemolytic disease and associated sequelae, including jaundice, occurs most commonly with Leptospira initerrogans serovars Icterohemorrhagiae or Pomona and the liver and kidney are the most common organs affected. After the bacteremic stage, Leptospira spp colonize the renal tubular epithelium approximately 10 days after infection, leading to bacterial shedding into the urine for up to 8 months after infection. The length of shedding can vary with species and individual.³

Among nondomestic felids, clinical disease has not been reported in individuals under managed care; however, seropositivity in free-ranging animals has been shown in several populations. A postmortem review of free-ranging mountain lions (Puma concolor) and bobcats (Lynx rufus) in California identified a prevalence of 46% and 28%, respectively, based upon either seropositive or PCR assay positive status.⁴ A similar percentage of free ranging jaguars (Panthera onca) in Brazil were seropositive for Leptospira spp (13/31).⁵ However, within screening for Leptospira serologic status in 57 managed nondomestic felids, only 1 captive-born ocelot (Leopardus pardalis) and 1 wild-caught margay (Leopardus wiedii) were seropositive.⁶ Recently, urinary shedding of Leptospira DNA has been reported in a captive ocelot without clinical signs.⁷

According to the American College of Veterinary Internal Medicine 2010 consensus statement on leptospirosis, seropositive domestic cats have been reported but clinical disease is rare.⁸ A recent review of the literature identified seroprevalence ranging from 4% to 33.3%, representing over 2,400 domestic cats sampled from multiple global locations and identifying a variety of positive serovars in each population.³ The same review reports prevalence rates spanning from 0% to 67.8% on the urinary shedding of Leptospira DNA from over 1,300 domestic cats from 6 different global locations.³

Multiple laboratory tests were utilized in the diagnosis of leptospirosis in the tiger of the present report, including MAT of the serum, PCR assay of the urine and immunohistochemistry of the liver. Microscopic agglutination testing is the recommended diagnostic test for determination of Leptospira spp antibody titers in domestic canine and feline patients with clinical signs. Infection occurring at least 15 days prior to testing should result in positive IgM and IgG titers on MAT, although little information is available in felids specifically.³ In this case, the rapidly rising titers to Leptospira kirschneri serovar Grippotyphosa between the first (day 0) and second (day 2) immobilizations were strongly supportive of active infection.

Real-time PCR detection of pathogenic Leptospira spp from the urine was the primary diagnostic used to screen and monitor the tiger’s response to antimicrobial therapy during both treatment cycles. In hindsight, submission of serum from the initial immobilization for real-time PCR assay would have been ideal to detect a bacteremic phase, however, was not pursued, and the samples were no longer available. In this particular case, repeated screening of the urine allowed for noninvasive collection from the indoor enclosure floor reducing the need for repeated immobilizations required in this animal for venipuncture. PCR methodology is associated with false-negative results due to transient shedding of the Leptospira organism or recent or current administration of antimicrobials.³ Additionally, PCR assays performed on urine have a lower sensitivity than when performed on serum.¹ Urine PCR assay results were consistently negative throughout the first round of diagnosis and treatment, which may have reflected perioperative antimicrobials administered or that the leptospiral organisms had not yet moved from the bacteremia stage to the renal stage prior to starting antimicrobials, effectively halting organismal shedding.

The tiger of the present report was initially suspected of suffering from foreign body obstruction based upon findings on abdominal imaging. Although no foreign body was found, this procedure allowed for the collection of a liver biopsy specimen and positive immunohistochemistry findings that ultimately led to the diagnosis of leptospirosis. The tiger’s hepatic histopathology findings that included acute hepatocellular dissociation, necrosis, portal hepatitis, and early fibrosis is consistent with the histopathology reported in domestic felines.³ Leptospirosis was not an initially considered differential given that clinical infection is rare in felids and has not been reported in this species.

An isolated incident of hematuria was observed approximately 2.5 years after the tiger’s initial leptospirosis diagnosis and treatment. In 6 domestic cats diagnosed with leptospirosis, 2 had hematuria.³ In the present case, urine Leptospira real-time PCR results were negative when the hematuria occurred indicating the animal was not actively shedding leptospires at the time. The tiger was monitored by animal care staff for repeated hematuria which would prompt immobilization and diagnostic evaluation. Because the hematuria rapidly resolved and the tiger remained clinically and behaviorally normal through to final follow-up, further evaluation has not been pursued. Clinicians suspected that the hematuria may have arisen from a dislodged urolith or prostatitis.

Several other diagnostic tests were performed in the initial and follow-up care of this tiger. Initial 3-view abdominal and thoracic radiographs were obtained and a normal thorax study was noted radiographically. CBC evaluation revealed almost doubling of the neutrophil concentration from day 0 to day 2. The presence of immature neutrophils supported acute inflammation. During these 2 days, the tiger also developed a nonregenerative anemia consistent with anemia of chronic disease. These findings were similar to those reported in domestic felines and canines diagnosed with leptospirosis.^3,8 In a prior report,³ domestic cats with nonregenerative anemia were determined to be chronic carriers, whereas animals with neutrophilia were diagnosed with an acute form of the disease. The authors suspect that the marked hepatic changes (acute hepatocellular degeneration) were consistent with acute inflammation and could explain the left shift noted. In domestic cats, leptospire toxins cause endothelial damage and local ischemia, and the primary organs affected by the toxins include kidneys and liver.³ The tiger reported here never became azotemic, which is common in clinically affected dogs and cats,^3,8 but an elevated serum total bilirubin concentration and clinical jaundice in the absence of hepatocellular enzyme deviation occurred.

Domestic feline antimicrobial directives are based off domestic canine recommendations for the treatment of leptospirosis.⁸ Initial therapy with β-lactam antimicrobials is recommended to treat bacteremia, while follow-up therapy with doxycycline is pursued to clear the bacteria from the renal tubules. A prolonged doxycycline course may be recommended in cats due to their high likelihood of being in a carrier state.³ Initially, the tiger of the present report received amoxicillin–clavulanic acid for 12 days and doxycycline for 21 days based upon the recommendations for canid patients, as the updated consensus statement for felids was not available prior to this tiger’s case. The tiger was negative for Leptospira spp on urine PCR assay after finishing its initial antimicrobial therapy; however, more than a year later had a positive urine PCR result for Leptospira spp, initiating another treatment cycle. It is unknown if the repeated urinary shedding suggested a carrier state in this tiger, which has been reported in domestic cats,³ or reinfection. Although the urine was collected from a clean cement floor from the indoor housing of the tiger, it could not be ruled out that the urine was contaminated by rodent urine.

Vaccination against Leptospira serovars was considered for this tiger and the other mammals in the collection with access to the outdoors. However, there is no commercial domestic felid vaccine. When a commercial canine vaccine was experimentally administered to domestic cats, only 1 animal maintained antibodies for over 1 month.⁹ Due to the lack of response in a related species and unknown risk to nondomestic species in the collection, vaccination was not pursued.

Bodies of water are potential sites of transmission for Leptospira spp as the bacterium is spread between animals through urine or urine-contaminated bodies of water and soil.^3,8 Raccoons, being a species of particular interest in this case, prefer to utilize bodies of water for urination and defecation. While impossible to confirm the originating source of the infection in this case, a raccoon on zoo grounds tested Leptospira positive on necropsy shortly after this clinical case in a tiger. Further, antibodies against Leptospira spp were found in raccoons on grounds at a zoological institution in St Louis, through a serosurvey of 159 animals, documenting 8.9% positivity for Leptospira interrogans serovar Icterohemorrhagiae and 6.3% positivity for Leptospira kirschneri serovar Grippotyphosa.¹⁰ Additionally, opportunistic sampling of urine (premortem) or renal biopsy and urine testing (post mortem) of free-ranging Californian raccoons revealed a 25.6% (22/86) prevalence of Leptospira spp using real-time PCR assay.¹¹

One of the most significant aspects of this case from a hospital, team member, and zoological institution management standpoint is that leptospirosis is a zoonotic disease.^3,8 Dogs presenting with acute or acute-on-chronic renal failure may be treated as suspected leptospirosis cases until proven otherwise. It is recommended that medical and care staff wear gloves and gowns while treating the animal or working with its biologics, house the animal in isolation, and avoid high-power hosing of enclosures.⁸ The tiger of the present report presented as an animal with severe hepatopathy and worsening jaundice. Leptospirosis did not enter the differential list until the liver histopathology was reported on day 2 after presentation. Any member of the veterinary or animal care team that was considered immunosuppressed and came in contact with the tiger once the diagnosis was made was notified and advised to contact their physician for further recommendations. This case brings to light that leptospirosis is a multifaceted disease, and nondomestic felids presenting with jaundice should have leptospirosis included as a potential differential diagnosis.

In conclusion, this is the first reported clinical case of leptospirosis in a large nondomestic felid with the unique presentation of gastrointestinal signs followed by hepatic dysfunction. Domestic cats, free-ranging nondomestic felids, and managed nondomestic felids are more commonly chronic carriers with prolonged shedding up to 8 months, and reports rarely describe clinical disease. However, the tiger in this case epitomized hepatic leptospirosis. Animals belonging to the Felidae order that present for acute vomiting, hepatic, renal, and/or hemolytic disease should have leptospirosis included on the differential list, particularly if in contact with potential vectors or bodies of water. While performing diagnostic evaluation and follow-up testing, it is important to remind care teams of the zoonotic nature of this disease until the animal is no longer shedding the bacterium.

Supplementary Materials

Supplementary materials are posted online at the journal website: avmajournals.avma.org