PRIMARY HEPATIC NEOPLASIA IN THREE ADULT BAMBOO SHARKS (CHILOSCYLLIUM PLAGIOSUM AND CHILOSCYLLIUM PUNCTATUM) IN AN AQUARIUM POPULATION.

Neoplasia in elasmobranchs is uncommonly documented. This report describes primary hepatic neoplasia in three adult female bamboo sharks under managed care: biliary adenocarcinoma in a whitespotted bamboo shark (Case 1; Chiloscyllium plagiosum), biliary adenocarcinoma in a brownbanded bamboo shark (Case 2; Chiloscyllium punctatum), and hepatocellular carcinoma in a whitespotted bamboo shark (Case 3). Case 1 presented with extensive cutaneous papillomas and was electively euthanized. At necropsy, a 4-cm-diameter, pale-tan, firm hepatic mass was identified and diagnosed histologically as a biliary adenocarcinoma. Case 2 presented with decreasing body condition despite normal food intake. Coelomic ultrasound and exploratory surgery revealed several large masses in both hepatic lobes, and the patient was euthanized. At necropsy, nine, 1-6-cm-diameter, black to tan, firm hepatic masses were identified and diagnosed histologically as biliary adenocarcinoma with branchial intravascular neoplastic emboli. Case 3 presented for routine health examination and was euthanized for diagnostic purposes after coelomic ultrasound revealed multiple hepatic masses. At necropsy, two 1-3-cm-diameter, brown- and-black mottled, firm hepatic masses were identified and diagnosed histologically as hepatocellular carcinoma. Immunohistochemistry was performed in two of these cases and was noncontributory, likely because of a lack of cross reactivity between antibodies (antipancytokeratin) and elasmobranch tissues.

Bacterial abscesses in whitespotted bamboo sharks (Chiloscyllium plagiosum) in managed care.

Sixteen whitespotted bamboo sharks (Chiloscyllium plagiosum) with histologically similar bacterial abscesses were submitted to Northwest ZooPath from nine zoo and aquarium institutions over a 17-year period. These abscesses were characterized by inflammatory cell infiltrates and necrosis with intralesional small, Gram-positive, acid-fast negative, cocci bacteria. The clinical presentation, histologic findings, and culture results indicate that Enterococcus faecalis is a relatively common cause of these lesions in whitespotted bamboo sharks. This organism also provides a treatment challenge due to its inherent antibiotic-resistant properties and ability to form biofilms, confounding the host's immune response. Enterococcus faecalis represents an important cause for abscess formation and cellulitis in captive whitespotted bamboo sharks.

DEXMEDETOMIDINE AND MIDAZOLAM INTRAMUSCULAR SEDATION IN BROWNBANDED BAMBOO SHARKS (CHILOSCYLLIUM PUNCTATUM).

Bamboo sharks are some of the most common elasmobranch species in zoos and aquaria and are frequently sedated for medical exams, treatments, and research. This study investigated the use of an IM sedation protocol of a single dose of dexmedetomidine (0.05 mg/kg) and midazolam (2.0 mg/kg) in brownbanded bamboo sharks (Chiloscyllium punctatum). Sharks were serially monitored every 5 min for heart rate, branchial beats, righting reflex, coelomic response, cloacal response, pelvic fin reflex, response to noxious stimulus, voluntary movement, and ability to swim. This sedation dose was effective at rapidly and significantly decreasing responses to tactile and noxious stimuli with minimal respiratory depression and was quickly reversible with atipamezole (0.5 mg/kg) and flumazenil (0.05 mg/kg). Sedated sharks developed a mild metabolic acidosis evidenced by a significant increase in lactic acid (mean < 0.37 mmol/L presedation, 4.2 mmol/L after reversal) and decrease in blood pH (mean 7.464 presedation, 7.277 after reversal); however, clinical intervention was not required. This protocol should be further investigated in different elasmobranch species but is promising for providing sedation for noninvasive procedures in brownbanded bamboo sharks.

Exploration of serum cardiac troponin I as a biomarker of cardiomyopathy in southern sea otters (Enhydra lutris nereis).

OBJECTIVE: To compare serum cardiac troponin I (cTnI) concentrations between sea otters with and without cardiomyopathy and describe 2 cases of cardiomyopathy with different etiologies. ANIMALS: 25 free-ranging southern sea otters (Enhydra lutris nereis) with (n = 14; cases) and without (11; controls) cardiomyopathy and 17 healthy managed southern sea otters from aquariums or rehabilitation centers (controls). PROCEDURES: Serum cTnI concentration was measured in live sea otters. Histopathologic and gross necropsy findings were used to classify cardiomyopathy status in free-ranging otters; physical examination and echocardiography were used to assess health status of managed otters. Two otters received extensive medical evaluations under managed care, including diagnostic imaging, serial cTnI concentration measurement, and necropsy. RESULTS: A significant difference in cTnI concentrations was observed between cases and both control groups, with median values of 0.279 ng/mL for cases and < 0.006 ng/mL for free-ranging and managed controls. A cutoff value of ≥ 0.037 ng/mL yielded respective sensitivity and specificity estimates for detection of cardiomyopathy of 64.3% and 90.9% for free-ranging cases versus free-ranging controls and 64.3% and 94.1% for free-ranging cases versus managed controls. CONCLUSIONS AND CLINICAL RELEVANCE: Cardiomyopathy is a common cause of sea otter death that has been associated with domoic acid exposure and protozoal infection. Antemortem diagnostic tests are needed to identify cardiac damage. Results suggested that serum cTnI concentration has promise as a biomarker for detection of cardiomyopathy in sea otters. Serial cTnI concentration measurements and diagnostic imaging are recommended to improve heart disease diagnosis in managed care settings.

TRANSTHORACIC ECHOCARDIOGRAPHIC EVALUATION AND SERUM CARDIAC TROPONIN VALUES IN ANESTHETIZED HEALTHY FEMALE SOUTHERN SEA OTTERS (ENHYDRA LUTRIS NEREIS).

Information about antemortem cardiac evaluation in sea otters (Enhydra lutris) is limited, despite well-established clinical care and rehabilitation procedures and a reported elevated risk of cardiac disease for this species. Serum cardiac troponin I (cTnI) concentration and echocardiographic assessment are two ways of screening for and diagnosing cardiac disease. However, no baseline data or reference intervals for either evaluation are published for sea otters. The objectives of this prospective study were to establish serum cTnI concentrations and echocardiographic technique and quantitative measurements in anesthetized healthy female southern sea otters (Enhydra lutris nereis) (n=15). Serum cTnI values were assessed by a high-sensitivity assay. Serum cTnI concentration ranged from <0.006 to 0.038 ng/ml. A complete echocardiogram, including two-dimensional and M-mode modalities, was performed. Echocardiographic measurements for left atrial size, aorta size, left ventricular structure, and left ventricular function were reported. The median left atrial size to aorta ratio was 1.22 (range 0.80-1.59) in short-axis and 1.70 (range 1.39-2.15) in long-axis. The median left ventricular internal dimension was 3.53 cm (range 2.87-4.92 cm) when assessed in two dimensions and 3.58 cm (range 2.80-4.48 cm) by M-mode. Serum concentrations of cTnI and transthoracic echocardiography may represent valuable tools for the antemortem diagnosis of cardiac disease in sea otters.

Artificial insemination and parthenogenesis in the whitespotted bamboo shark Chiloscyllium plagiosum.

Non-lethal methods for semen collection from elasmobranchs to better understand species reproduction has accompanied the development of artificial insemination. Ejaculates (n = 82) collected from whitespotted bamboo sharks Chiloscyllium plagiosum (n = 19) were assessed and cold-stored raw or extended at 4 °C. Females (n = 20) were inseminated with fresh or 24-48 h cold-stored raw or extended semen and paternity of offspring determined with microsatellite markers. Insemination of females with fresh semen (n = 10) resulted in 80 hatchlings and 27.6% fertility. Insemination of females with semen cold-stored 24 h (n = 4) and 48 h (n = 1) semen resulted in 17 hatchlings and fertilization rates of 28.1% and 7.1% respectively. Two females inseminated with fresh or cold-stored semen laid eggs that hatched from fertilization and parthenogenesis within the same clutch. Parthenogenesis rate for inseminated females was 0.71%. Results demonstrate artificial insemination with cold-stored semen can provide a strategy for transport of male genetics nationally and internationally, precluding the need to transport sharks. Production of parthenotes in the same clutch as sexually fertilized eggs highlights the prevalence of parthenogenesis in whitespotted bamboo sharks and poses important considerations for population management.

Systemic phaeohyphomycosis in splitnose rockfish (Sebastes diploproa) caused by Devriesia sp.

A novel pathogen was documented after two wild-caught, juvenile, splitnose rockfish presented with buphthalmia, grey corneal endothelial plaques and evidence of uveitis. Cytologic evaluation of ocular contents revealed fungal hyphae. Histologic evaluation identified multiple fungal granulomas and granulomatous inflammation in the globes, periocular tissue and heart. Fungi were slender, hyphenated and branched at angles, had parallel cell walls and had brown pigmentation in haematoxylin- and eosin-stained sections. Both fish were diagnosed with phaeohyphomycosis. Culture with nuclear ribosomal RNA internal transcribed spacer (ITS) segment identification further classified the fungus as Devriesia sp., which has not been previously documented as a cause of disease in animals.

Testosterone and semen seasonality for the sand tiger shark Carcharias taurus†.

Understanding the fundamental reproductive biology of a species is the first step toward identifying parameters that are critical for reproduction and for the development of assisted reproductive techniques. Ejaculates were collected from aquarium (n = 24) and in situ (n = 34) sand tiger sharks Carcharias taurus. Volume, pH, osmolarity, sperm concentration, motility, status, morphology, and plasma membrane integrity were assessed for each ejaculate. Semen with the highest proportion of motile sperm was collected between April and June for both in situ and aquarium sand tiger sharks indicating a seasonal reproductive cycle. Overall, 17 of 30 semen samples collected from aquarium sharks from April through June contained motile sperm compared to 29 of 29 of in situ sharks, demonstrating semen quality differences between aquarium and in situ sharks. Sperm motility, status, morphology, and plasma membrane integrity were significantly higher (P < 0.05) for in situ compared to aquarium sand tiger sharks. Testosterone was measured by an enzyme immunoassay validated for the species. Testosterone concentration was seasonal for both aquarium and in situ sharks with highest concentrations measured in spring and lowest in summer. In situ sharks had higher (P < 0.05) testosterone concentration in spring than aquarium sharks. This study demonstrated annual reproduction with spring seasonality for male sand tiger sharks through marked seasonal differences in testosterone and semen production. Lower testosterone and poorer semen quality was observed in aquarium sharks likely contributing to the species' limited reproductive success in aquariums.

HEMATOLOGIC AND PLASMA BIOCHEMICAL REFERENCE VALUES FOR JUVENILE GREEN-NAPED LORIKEETS ( TRICHOGLOSSUS HAEMATODUS HAEMATODUS).

The green-naped lorikeet ( Trichoglossus haematodus haematodus) is a small, brightly colored psittacine that is one of many subspecies of the rainbow lorikeet popular in captivity. Overall, the rainbow lorikeet population is declining but the wide range of subspecies means the population is classified as "least concern" by the International Union for Conservation of Nature. The goal of this study was to establish normal hematologic and plasma biochemical analyte reference intervals for juvenile green-naped lorikeets. Hematology and plasma biochemistry variables were determined for 102 clinically healthy, captive-born, juvenile (2-6 mo) green-naped lorikeets house at a single institution. This study is the first report for hematologic and plasma biochemical values for any Trichoglossus species.

Distribution and clinical impact of functional variants in 50,726 whole-exome sequences from the DiscovEHR study.

The DiscovEHR collaboration between the Regeneron Genetics Center and Geisinger Health System couples high-throughput sequencing to an integrated health care system using longitudinal electronic health records (EHRs). We sequenced the exomes of 50,726 adult participants in the DiscovEHR study to identify ~4.2 million rare single-nucleotide variants and insertion/deletion events, of which ~176,000 are predicted to result in a loss of gene function. Linking these data to EHR-derived clinical phenotypes, we find clinical associations supporting therapeutic targets, including genes encoding drug targets for lipid lowering, and identify previously unidentified rare alleles associated with lipid levels and other blood level traits. About 3.5% of individuals harbor deleterious variants in 76 clinically actionable genes. The DiscovEHR data set provides a blueprint for large-scale precision medicine initiatives and genomics-guided therapeutic discovery.

The use of spirometry to evaluate pulmonary function in olive ridley sea turtles (Lepidochelys olivacea) with positive buoyancy disorders.

This study utilized computed spirometry to compare the pulmonary function of two stranded olive ridley sea turtles (Lepidochelys olivacea) presenting with a positive buoyancy disorder with two healthy captive olive ridley sea turtles held in a large public aquarium. Pulmonary function test (PFT) measurements demonstrated that the metabolic cost of breathing was much greater for animals admitted with positive buoyancy than for the normal sea turtles. Positively buoyant turtles had higher tidal volumes and significantly lower breathing-frequency patterns with significantly higher expiration rates, typical of gasp-type breathing. The resulting higher energetic cost of breathing in the diseased turtles may have a significant impact on their long-term survival. The findings represent a method for clinical respiratory function analysis for an individual animal to assist with diagnosis, therapy, and prognosis. This is the first study, to our knowledge, to evaluate objectively sea turtles presenting with positive buoyancy and respiratory disease using pulmonary function tests.

Gene transcription in sea otters (Enhydra lutris); development of a diagnostic tool for sea otter and ecosystem health.

Gene transcription analysis for diagnosing or monitoring wildlife health requires the ability to distinguish pathophysiological change from natural variation. Herein, we describe methodology for the development of quantitative real-time polymerase chain reaction (qPCR) assays to measure differential transcript levels of multiple immune function genes in the sea otter (Enhydra lutris); sea otter-specific qPCR primer sequences for the genes of interest are defined. We establish a 'reference' range of transcripts for each gene in a group of clinically healthy captive and free-ranging sea otters. The 10 genes of interest represent multiple physiological systems that play a role in immuno-modulation, inflammation, cell protection, tumour suppression, cellular stress response, xenobiotic metabolizing enzymes, antioxidant enzymes and cell-cell adhesion. The cycle threshold (C(T)) measures for most genes were normally distributed; the complement cytolysis inhibitor was the exception. The relative enumeration of multiple gene transcripts in simple peripheral blood samples expands the diagnostic capability currently available to assess the health of sea otters in situ and provides a better understanding of the state of their environment.

Serum vitamin A concentrations in captive sea otters (Enhydra lutris).

Individual dietary preferences and difficulty with animal training create challenges and nutritional concerns when evaluating a captive sea otter (Enhydra lutris) diet. The importance of vitamin A within the body reflects the necessity that it be ingested in adequate amounts to ensure optimal health. To compare levels of serum vitamin A concentrations from captive sea otters on daily oral vitamin A supplementation, serum samples from eight adult sea otters from three institutions were evaluated for serum vitamin A concentrations. The eight animals were fed a total of four different diets and received oral supplementation via three different methods. Multiple diet items were analyzed for vitamin A content and were found to have low to nondetectable levels of vitamin A. Oral vitamin A supplementation, as a slurry with dietary items, was shown to be effective and a mean serum concentration of approximately 170 +/- 51 microg/L was obtained for serum vitamin A concentrations in captive sea otters. Captive diets can be modified to increase vitamin A concentration and supplementation and, if accepted, can be used as a means to ensure adequate vitamin A intake.

Use of laser rhinoscopy to treat a nasal obstruction in a captive California sea lion (Zalophus californianus).

Laser rhinoscopy was used to treat a nasal obstruction in a captive California sea lion (Zalophus californianus). The rehabilitated, adult, female sea lion developed mucopurulent, intermittent, bilateral nasal discharge and functional nasal obstruction 20 mo after acquisition by the Aquarium of the Pacific in Long Beach, California. A 3-mm-thick soft tissue structure spanning the region between the soft and hard palates, a deviated nasal septum, and several nasopharyngeal polyps were identified. Biopsies and cultures of the obstructive web showed ulcerative granulation tissue with suppurative inflammation, bacterial infection, and a partial section of an arthropod larva (not speciated). Laser rhinoscopy was performed to relieve the caudal nasopharyngeal obstruction and ablate the polyps. The sea lion appeared to breathe through the nares with lessened nasal discharge for a period of 6 wk after laser therapy, but within 8 wk the mucopurulent nasal discharge returned, the obstruction had reformed, and the sea lion was euthanized. Postmortem examination confirmed antemortem diagnoses of caudal nasopharyngeal obstruction secondary to inflammatory tissue; however, no additional sections of arthropod parasites were located microscopically.