SARS-CoV-2 morbidity, treatment interventions, and vaccination practices in tigers (Panthera tigris ssp) in North American zoos.

OBJECTIVE: Evaluate SARS-CoV-2 morbidity, mortality, clinical signs, treatment interventions, and vaccination practices in tigers under professional care. ANIMALS: Amur (Panthera tigris altaica), Sumatran (Panthera tigris sumatrae), and Malayan (Panthera tigris jacksoni) tigers managed under the Tiger Species Survival Plan (SSP). METHODS: A retrospective, voluntary online survey was sent to all North American zoos holding SSP tigers between January 2020 and June 2023. RESULTS: Responses were received from 55 of 108 institutions (51%) housing 162 tigers in total, and SARS-CoV-2 infection was diagnosed in 39 tigers from 15 institutions (20 Amur, 8 Sumatran, and 11 Malayan [1 to 18 years old; 17 males and 22 females]). This corresponds to a minimum study group infection incidence of 24% over 42 months. Clinical signs included dry cough (82%), inappetence (64%), lethargy (62%), nasal discharge (46%), wheezing (31%), wet cough (18%), and ocular discharge (15%). Most cases were characterized as mild (n = 22) or moderate (14). A single case was characterized as severe. Two cases were asymptomatic. Seventeen positive tigers had been vaccinated once (n = 8) or twice (9) for SARS-CoV-2 prior to infection. No deaths due to SARS-CoV-2 were reported in the study group. Treatment interventions included antibiotics (49%), NSAIDs (18%), antiemetics (15%), and fluids (13%). No treatments were administered in 19 of 39 cases (49%). Amongst participating institutions, 69% reported fully vaccinating tigers for SARS-CoV-2 (≥ 2 doses). CLINICAL RELEVANCE: Most SARS-CoV-2-infected tigers presented with mild to moderate clinical signs and recovered with limited to no treatment interventions. Asymptomatic SARS-CoV-2 infections can occur in tigers and may be underreported. Tigers vaccinated for SARS-CoV-2 remain susceptible to infection.

DEVELOPMENT OF A HEMOLYMPH COLLECTION TECHNIQUE AND REPORT OF BIOCHEMICAL PARAMETERS OF HEMOLYMPH IN A MANAGED POPULATION OF FEMALE THORNY DEVIL STICK INSECTS (EURYCANTHA CALCARATA).

Insects are increasingly common in households, zoological collections, research, and food industries. Increased knowledge of insect health parameters is necessary to ensure proper welfare. The study goal was to develop a hemolymph collection technique and report hemolymph serum biochemical parameters for the thorny devil stick insect (Eurycantha calcarata). Clinically healthy adult stick insects (5 males and 14 females, 15.1-24.7 g) were enrolled. Four collection techniques were evaluated. Hemolymph collection was unsuccessful in males, but was successful in females by using a single technique. The insect was manually restrained in an elevated position and an 18-ga × 2.54-cm needle was used to puncture the membrane just caudal to the third pair of legs. With the puncture site directed ventrally, ≤1 ml of hemolymph was collected via gravity and allowed to visibly clot. The sample was then centrifuged, and the serum was separated. Serum samples were individually analyzed (Avian/Reptilian Profile Plus, VetScan VS2, Abaxis, Inc, Union City, CA 94587, USA). Fourteen samples (0.2-1.0 ml) were collected from 14 females. Median (minimum-maximum) parameters included the following: aspartate aminotransferase 12 (0-45) U/L, creatinine kinase 25 (0-76) U/L, uric acid 7.5 (3.1-13.7) mg/dl, glucose 12 (8-22) mg/dl, calcium 18.6 (17.2-19.4) mg/dl, phosphorus 15.0 (n = 1) or >30.0 (n = 13) mg/dL, total protein 2.7 (1.6-2.9) g/dL, albumin 0.9 (0.2-1.2) g/dL, globulin 1.7 (1.6-1.8) g/dL (n = 6) or not quantified (n = 8), potassium 10.6 (9.0-11.8) mmol/L, sodium < 100 mmol/L, and bile acids 0 lmol/L. This is the first report of biochemistry parameters in clinically healthy female stick insects. Larger sample sizes are needed to establish statistically valid reference ranges. Hemolymph collection techniques for male stick insects warrant further investigation.

INVESTIGATION OF A POINT-OF-CARE VISCOELASTIC COAGULATION MONITOR AND ITS COMPARISON TO THROMBOELASTOGRAPHY IN CLINICALLY HEALTHY AFRICAN ELEPHANTS (LOXODONTA AFRICANA).

Elephant endotheliotropic herpesvirus (EEHV) can induce fatal hemorrhagic disease (HD) in African elephants (Loxodonta africana). Once clinical signs develop, progression is rapid, even with aggressive treatment. There is a critical need to develop point-of-care diagnostic tests to aid in identification of EEHV-HD prior to the onset of overt clinical signs. Study objectives were to investigate a novel, point-of-care viscoelastic coagulation monitor (VCM Vet), compare the results to thromboelastography (TEG), and report traditional hemostatic analytes in adult African elephants. Whole blood was collected from seven clinically healthy elephants (four females and three males, 18-47 yr) and analyzed in duplicate via VCM Vet and kaolin-activated TEG 1-3 and 30 min following collection, respectively. Separated plasma was frozen for ancillary coagulation testing. Both analyses generated quantifiable clotting reactions with variables (median [range]) describing clot formation rate (VCM Vet, clot time = 682 s [530-987 s], clot formation time = 244 s [186-744 s], Alpha = 40° [14-47°]; TEG, reaction time = 6.2 min [3.7-11.8 min], kinetic time = 1.3 min [0.9-2.6 min], Alpha = 70° [57-77°]), clot strength (VCM Vet, maximum clot formation = 34 units [20-45 units]; TEG, maximum amplitude = 75 mm [69-80 mm], shear elastic modulus strength = 14.7 Kdynes/s [11.3-19.5 Kdynes/s]), and clot lysis (VCM Vet, lysis index at 30 min = 100% [100-99%], lysis index at 45 min = 98% [95-100%]; TEG, lysis index at 30 min = 0% [0-0.4%], lysis index at 60 min = 1.4% [0-2.6%]) recorded. Additional testing (median [range]) included D-dimer concentration (33 ng/ml [28-94 ng/ml]), prothrombin time (12.4 s [12.2-13.2 s]), activated partial thromboplastin time (17.2 s [14.2-18.8 s]), and fibrinogen concentration (297 [282-383] mg/dL). Tracings generated by VCM Vet and TEG were clinically similar, and there was visual agreement and minimal difference between quantitative variables for duplicate tests. VCM Vet is a promising, user-friendly tool for use in identification and management of coagulopathies in African elephants.

Anesthetic efficacy of dexmedetomidine-ketamine in eastern box turtles (Terrapene carolina carolina) is enhanced with the addition of midazolam and when administered in the forelimb versus the hindlimb.

OBJECTIVE: To compare dexmedetomidine-ketamine (DK; 0.1 and 10 mg/kg, respectively) with midazolam (M; 1.0 mg/kg) or 0.9% sodium chloride (S; 0.2 mL/kg) administered IM in the forelimb (F) or hindlimb (H) in eastern box turtles (Terrapene carolina carolina). ANIMALS: 20 clinically healthy, captive adult eastern box turtles. METHODS: In a randomized, blinded, complete crossover study with 1-week washout periods, turtles were administered each of 3 treatments: F-DKS, F-DKM, or H-DKM. Palpebral reflex, muscle tone, and withdrawal responses were serially assessed and used to calculate cumulative sedation scores at each 5-minute time point. The ability to intubate was evaluated. At 60 minutes, atipamezole (1.0 mg/kg) and either flumazenil (F-DKM, H-DKM; 0.05 mg/kg) or 0.9% sodium chloride (F-DKS; 0.5 mL/kg) were administered IM. RESULTS: All treatments resulted in clinically relevant anesthetic effects. F-DKM produced significantly higher sedation scores than H-DKM or F-DKS at all time points between 10 and 60 minutes (P < .05). Sedation score variability was observed with all treatments with significantly higher variability for H-DKM (P < .05). Intubation was successful in 32, 89, and 11% of turtles in F-DKS, F-DKM, and H-DKM, respectively. Median (range) recovery time was 10 (5-22), 16 (7-45), and 12 (4-28) minutes for F-DKS, F-DKM, and H-DKM, respectively. CLINICAL RELEVANCE: In eastern box turtles, forelimb dexmedetomidine-ketamine resulted in clinically relevant anesthetic effects that were heightened with the addition of midazolam. Hindlimb administration of midazolam-dexmedetomidine-ketamine resulted in reduced and more variable anesthetic effects compared to forelimb administration, supporting a hepatic first-pass effect.

POTASSIUM CHLORIDE ADMINISTERED VIA FOUR ROUTES FOR EUTHANASIA OF ANESTHETIZED GOLDFISH (CARASSIUS AURATUS).

Immersion overdose in tricaine methanesulfonate (MS-222) is ineffective for euthanasia of adult goldfish (Carassius auratus), and investigation of alternative techniques is warranted. This study evaluated potassium chloride (KCl) administered via four routes for goldfish euthanasia. Thirty clinically healthy adult goldfish (17.2-41.9 g) were anesthetized via immersion in buffered MS-222 (300 mg/L) for 5 min beyond anesthetic induction. Fish were then randomly administered one of five treatments: KCl (333 mg/mL, 4.5 mmol/mL, 4.5 mEq/mL) via intracardiac injection at 10 mEq/kg (IC), intracoelomic injection (ICe) or bilateral topical delivery over the gill filaments (T) at 100 mEq/kg, or 90-min immersion at 4,500 mEq/L (saturated solution) dissolved in induction solution (water [W]), or no treatment (control [C]). Following treatment, all fish were moved to anesthetic-free freshwater. Serial heart rates were collected via Doppler device until sound cessation or recovery. Median (range) time to perform treatments was 5.3 (1.2-8.0) min, 3 (3-3) s, and 10 (10-10) s in IC, ICe, and T, respectively. Doppler cessation occurred in 6/6, 6/6, 6/6, 6/6, and 0/6 fish in median (range) times of 3 (0-210), 18 (10-45), 118 (90-390), and 150 (60-240) min in IC, ICe, T, W, and C, respectively; this was significantly different between groups (P = 0.002). Following or during treatment, 1/6, 2/6, 6/6, and 4/6 fish in IC, ICe, T, and W, respectively, exhibited intermittent transient jerking movements. Median (range) time to recovery in C was 5.5 (3.5-6.5) min. All administration routes were effective, but time to Doppler cessation varied and transient movements were noted. Preliminary follow-up research determined that return to MS-222 induction solution following ICe KCl treatment abolished movements. ICe KCl at 100 mEq/kg was technically simple and resulted in Doppler sound cessation in a clinically relevant time frame in 6/6 anesthetized goldfish. Return to MS-222 immersion following KCl administration is recommended.

EVALUATION OF INTRACARDIAC ADMINISTRATION OF POTASSIUM CHLORIDE, IVERMECTIN, OR LIDOCAINE HYDROCHLORIDE FOR EUTHANASIA OF ANESTHETIZED BLUE CRABS (CALLINECTES SAPIDUS).

Methods to anesthetize and euthanize aquatic invertebrates have proven unreliable in decapods; thus studies to optimize euthanasia techniques for crustaceans are needed. Study objectives were to evaluate efficacy of intracardiac potassium chloride (KCl), ivermectin, or lidocaine hydrochloride (HCl) for euthanasia of anesthetized blue crabs (Callinectes sapidus). Twenty adult male crabs (n = 5/group) were immersed in 500 mg/L eugenol for 5 min beyond loss of the righting reflex and then randomly administered intracardiac 10 mEq/kg KCl (333 mg/mL), 5 mg/kg ivermectin (10 mg/ml), 100 mg/kg lidocaine hydrochloride (HCl) (20 mg/ml), or 5 ml/kg saline (0.9%). Serial heart rate assessments were made using a Doppler probe placed over the dorsum, and times to loss of righting reflex, Doppler sound cessation, and/or recovery were recorded. Median (range) time to loss of righting reflex was 32 (17-57) min. One crab in all groups, except lidocaine HCl, had no detectable Doppler sounds prior to injection. In the remaining crabs, Doppler sound cessation occurred in 4/4, 4/4, 4/5, and 0/4 crabs administered KCl, ivermectin, lidocaine HCl, and saline, respectively. Median (range) time to Doppler sound cessation was 30 (0-55), 18 (16-28), and 50 (0-90) s in KCl, ivermectin, and lidocaine HCl groups, respectively. Tonic limb movements were observed in 5/5 KCl-treated crabs. Median (range) time to recovery was 180 (115-345) and 300 m in four saline-treated crabs and one lidocaine HCl-treated crab, respectively. Intracardiac KCl at 10 mEq/kg and ivermectin at 5 mg/kg were effective, rapid methods for euthanasia of anesthetized blue crabs.

Cutaneous epitheliotropic lymphosarcoma in a captive white catfish (Ameiurus catus Linnaeus).

A wild caught white catfish (Ameiurus catus Linnaeus) developed multiple cutaneous masses. Cytology revealed neoplastic lymphocytes and microscopy confirmed dermal infiltration with epitheliotropism in the epidermis, oral mucosa, and cornea, without internal organ involvement. Transmission electron microscopy did not identify viral particles. Histopathology supported cutaneous epitheliotropic lymphosarcoma, a condition most commonly reported in mammals. This is the first reported case of cutaneous epitheliotropic lymphosarcoma in an ictalurid and one of the few published cases of this condition in any fish species.

Development and Evaluation of a Serum Transfusion Process in the Thorny Devil Stick Insect (Eurycantha calcarata).

The thorny devil stick insect (Eurycantha calcarata) is a common invertebrate maintained under human care. Blood (hemolymph) transfusions are a widely used therapeutic tool in other species, but investigation in terrestrial arthropods remains scarce. Study objectives were development and evaluation of a serum transfusion process in the thorny devil stick insect. Twenty-five clinically healthy adult insects (9.9-23.0 g) were enrolled and baseline body weights were collected. Hemolymph collection was not successful in males, thus, all were recipient only (n = 12, MR). Females were divided into donor (n = 4, FD) or donor and recipient (n = 9, FDR) groups. Females were manually restrained and up to 1 mL of hemolymph was collected from the abdominal segment caudal to the proximal hindlimb using an 18 g hypodermic needle and passive collection via gravity. Hemolymph was quantified, centrifuged, and the serum separated. Insects were then injected superficially at the lateral aspect of the abdomen with 0.01 mL/g FD serum (MR), lactated Ringer's solution (LRS) equal to collected hemolymph volume (FD), or a combination of FDR serum and LRS equal to collected hemolymph volume (FDR). Response to stimulation, surface temperature, and righting reflex and mortality were serially assessed for up to 24 h and 7 days, respectively, following injection. In FD, median (range) injected LRS dose was 0.04 (0.03-0.06) mL/g. In FDR, median (range) injected serum, LRS, and combined serum and LRS dose was 0.03 (0.02-0.04), 0.01 (0-0.04), and 0.04 (0.02-0.06) mL/g, respectively. A mild temperature increase (maximum +2.9°C) (MR n = 10, FD n = 3, FDR n = 8) and delayed righting reflex (MR n = 4, FD n = 3, FDR n = 7) occurred in a subset of insects following injection. Two deaths occurred at 2 min (n = 1, FDR) and 96 h (n = 1, FD) post-injection. This is the first report of serum transfusions in thorny devil stick insects, and while largely successful, minor to severe transfusion reactions may occur.