Pyrethroid-associated nephrotoxicity in channel catfish, Ictalurus punctatus, and blue catfish, I. furcatus, at a public aquarium.

Over the course of an approximately 11-month period, an outdoor, freshwater, mixed species, recirculating, display system at a public aquarium experienced intermittent mortalities of channel catfish (Ictalurus punctatus) and blue catfish (I. furcatus). Catfish acutely presented for abnormal buoyancy, coelomic distention, and protein-rich coelomic effusion. Gross lesions typically involved massive coelomic distension with protein-rich effusion, generalized edema, and gastric hemorrhage and edema. Microscopically, primary lesions included renal tubular necrosis, gastric edema with mucosal hemorrhages, and generalized edema. Aerobic culture and virus isolation could not recover a consistent infectious agent. Intracoelomic injection of coelomic effusion and aspirated retrobulbar fluid from a catfish into naïve zebrafish (bioassay) produced peracute mortality in 3 of 4 fish and nervous signs in the fourth compared with 2 saline-injected control zebrafish that had - no mortality or clinical signs. Kidney tissue and coelomic effusion were submitted for gas chromatography tandem mass spectrometry by multiple reaction monitoring against laboratory standards, which detected the presence of multiple pyrethroid toxins, including bioallethrin, bifenthrin, trans-permethrin, phenothrin, and deltamethrin. Detection of multiple pyrethroids presumably reflects multiple exposures with several products. As such, the contributions of each pyrethroid toward clinical presentation, lesion development, and disease pathogenesis cannot be determined, but they are suspected to have collectively resulted in disrupted osmoregulation and fluid overload due to renal injury. Pesticide-induced toxicoses involving aquarium fish are rarely reported with this being the first description of pyrethroid-induced lesions and mortality in public aquarium-held fish.

Establishment of a 2-dimensional echocardiographic protocol and reference parameters for clinically healthy southern stingrays (Hypanus americanus).

OBJECTIVE: To establish an echocardiographic technique and 2-dimensional reference parameters for southern stingrays (Hypanus americanus). A second objective was to compare echocardiographic measurements obtained from animals of different sex, size, environment, handling technique, and position. ANIMALS: 84 presumed healthy, wild, semiwild, and aquarium-housed southern stingrays. PROCEDURES: Animals, anesthetized and manually restrained, were positioned in dorsal recumbency, and echocardiography was performed. A subset of this population was also imaged in ventral recumbency for comparison. RESULTS: Echocardiography was feasible, and reference parameters were established for this species. While some standard measurements could not be assessed due to body habitus, all valves, chambers, and the conus were clearly visualized in the majority of animals. Statistical significance was reached for some variables when comparing animals from different environments and handling methods, but these differences were not considered clinically relevant. The data were therefore separated into 2 subsets of echocardiographic reference parameters based on disc width since some of the measurements were dependent on body size. This approach mostly separated the sexes due to strong sexual dimorphism. CLINICAL RELEVANCE: Limited information is available regarding cardiac disease in elasmobranchs; most of the available information on cardiac physiology focuses on a few shark species. Two-dimensional echocardiography is a noninvasive tool utilized to evaluate cardiac structure and functionality. Southern stingrays are one of the most commonly displayed elasmobranchs in public aquaria. This article expands on the growing body of information regarding veterinary care in elasmobranchs and provides clinicians and researchers with another diagnostic modality to utilize in screening for health/disease.

Seasonal patterns in stable isotope and fatty acid profiles of southern stingrays (Hypanus americana) at Stingray City Sandbar, Grand Cayman.

Ecotourism opportunities in the marine environment often rely heavily on provisioning to ensure the viewing of cryptic species by the public. However, intentional feeding of wildlife can impact numerous aspects of an animals' behavior and ecology. Southern stingrays (Hypanus americana) provisioned at Stingray City Sandbar (SCS) in Grand Cayman have altered diel activity patterns and decreased measures of health. This study looked at seasonal changes in stable isotope (SI) and fatty acid (FA) profiles of provisioned stingrays at SCS. Plasma δ15N was higher in male stingrays (11.86 ± 1.71‰) compared to females (10.70 ± 1.71‰). Lower values for δ15N in males and females were measured in October during low tourist season, suggesting stingrays may be forced to rely on native prey items to supplement the decreased amount of provisioned squid available during this time. Plasma FA profiles were significantly different between sexes and across sampling time points, with FAs 22:6n3, 16:0, 20:5n3, 18:1n3C, 18:0 and 18:1n9T contributing to dissimilarity scores between groups. Dietary FAs primarily contributed to differences between males and females lending further evidence to differences in foraging patterns at SCS, likely due to intraspecific competition. Further, canonical analysis of principal coordinates (CAP) analysis of FA profiles suggest similar diets during peak tourist season and differences in diet between males and females during the low season. This study demonstrates alterations in feeding ecology in stingrays at SCS which is of critical importance for effective management of the SCS aggregation.

DIAGNOSIS AND TREATMENT OF AN ENTEROCOCCUS FECALIS ABSCESS IN THE CRANIAL VAULT OF A SPOTTED EAGLE RAY (AETOBATUS NARINARI).

An adult female spotted eagle ray (Aetobatus narinari) presented for medical evaluation due to a swelling located on the dorsal head. Ultrasound revealed that the swelling originated from a large pocket of fluid in the cranial vault. The swelling was aspirated, and purulent discharge was obtained; Enterococcus faecalis was cultured. An incision was made over the swelling in an attempt to drain fluid but was unsuccessful. Multiple aspirates were performed to drain the abscess, and the animal was treated with oxytetracycline injections. The initial incision sloughed and resulted in a large defect in the cranium that allowed exhibit water to come into the cranial vault and come in contact with the protective membrane of the brain. Forty-two days after initial presentation, the defect in the cranium was healed; fluid from the cranial vault was sampled and appeared normal. During and after treatment, the ray exhibited no abnormal neurologic signs.

Labeled quantitative mass spectrometry to study the host response during aspergillosis in the common bottlenose dolphin (Tursiops truncatus).

Aspergillosis is a fungal infection caused by Aspergillus molds that can affect both humans and animals. Despite advances in diagnostics and therapy, medical management of this disease remains difficult. Expansion of the basic knowledge regarding its pathophysiology in animals is critical to aid in the identification of new biomarkers of infection for diagnosis and therapeutic targets. For such a purpose, proteomics can be used by addressing protein changes during various disease processes. In the present study, a mass spectrometry analysis based on isobaric tagging for relative and absolute quantitation (iTRAQ®) was applied for direct identification and relative quantitation of proteins in blood collected from 32 Aspergillus-diseased common bottlenose dolphins (Tursiops truncatus, 32 samples) in comparison with blood from 55 other dolphins (55 samples from 41 clinically-normal controls and from 14 cetaceans with miscellaneous non-Aspergillus inflammation diseases) and ten convalescent dolphins (28 samples). Sixty-six and 40 proteins were found to be ≥2.0-fold over- and underrepresented versus miscellaneous non-Aspergillus inflammatory dolphins, respectively, and most were confirmed vs. clinically-normal controls and convalescents. Many proteins which play a role in the adaptive immune response were identified, including MHC proteins and others involved in catalytic activity like the NADPH-ubiquinone oxido-reductases. Overall, iTRAQ® appears to be a convenient proteomic tool greatly suited for exploratory ex vivo studies focusing on pathophysiology. This technique should be considered as a preliminary step before validation of new diagnostic markers.

HEMATOLOGIC AND BIOCHEMICAL SUMMARY STATISTICS IN AQUARIUM-HOUSED SPOTTED EAGLE RAYS ( AETOBATUS NARINARI).

During this retrospective study, 18 plasma blood chemistry and 17 complete blood count (CBC) samples were analyzed from clinically healthy spotted eagle rays ( Aetobatus narinari) at Georgia Aquarium in order to generate hematological ranges for complete blood count (CBC) and biochemical profiles. Summary statistics were generated according to the American Society for Veterinary Clinical Pathology guidelines for the determination of reference intervals in veterinary species. 4 The mean packed cell volume (PCV) was 28.09% with a range of 23-35%. Mean total solids were 5.72 g/dl with a range of 5-7.0 g/dl. Lymphocytes were the dominant leukocyte observed on differential (67.35%), followed by fine eosinophilic granulocytes (FEGs) (15.41%), coarse eosinophilic granulocytes (CEGs) (10.24%), monocytes (1.88%), and basophils (1.24%). Chemistry samples were analyzed at two diagnostic laboratories, Michigan State University (MSU) and University of Miami (UMiami), and the results were compared. Both labs have the capacity to run blood chemistries on zoo and aquatic species, but utilize different methods to obtain chemistry analyte values. UMiami uses a thin-film dry-slide technology, whereas MSU uses an ion-selective electrode (ISE) and Beckman Coulter AU 640 analyzer. There is poor agreement between the analyzers used by the two laboratories for both alkaline phosphatase and BUN, because of proportional error. Establishing hematological ranges in spotted eagle rays and in elasmobranchs in general may enhance the understanding of the species and their health. This information may aid clinicians in deciding when and how to treat elasmobranchs.

Evaluation of a genus-specific ELISA and a commercial Aspergillus Western blot IgG® immunoblot kit for the diagnosis of aspergillosis in common bottlenose dolphins (Tursiops truncatus).

Aspergillosis is a fungal infection with high mortality and morbidity rates. As in humans, its definitive diagnosis is difficult in animals, and thus new laboratory tools are required to overcome the diagnostic limitations due to low specificity and lack of standardization. In this study of common bottlenose dolphins (Tursiops truncatus), we evaluated the diagnostic performance of a new commercial immunoblot kit that had been initially developed for the serologic diagnosis of chronic aspergillosis in humans. Using this in a quantitative approach, we first established its positive cutoff within an observation cohort of 32 serum samples from dolphins with "proven" or "probable" diagnosis of aspergillosis and 55 negative controls. A novel enzyme-linked immunosorbent assay (ELISA) test was also developed for detecting anti-Aspergillus antibodies, and results were compared between the two assays. Overall, the diagnostic performance of immunoblot and ELISA were strongly correlated (P < .0001). The former showed lower sensitivity (65.6% versus 90.6%), but higher specificity (92.7% vs. 69.1%), with no cross-reaction with other fungal infections caused by miscellaneous non-Aspergillus genera. When assessing their use in a validation cohort, the immunoblot kit and the ELISA enabled positive diagnosis before mycological cultures in 42.9% and 33.3% subjects addressed for suspicion of aspergillosis, respectively. There was also significant impact of antifungal treatment on the results of the two tests (P < .05). In all, these new serological methods show promise in aiding in the diagnosis of aspergillosis in dolphins, and illustrate the opportunity to adapt commercial reagents directed for human diagnostics to detect similar changes in other animals.

WHOLE BLOOD TRANSFUSIONS TO TREAT SEVERE ANEMIA IN SEVEN COWNOSE RAYS ( RHINOPTERA BONASUS) AND ONE SHORT-TAIL STINGRAY ( DASYATIS BREVICAUDATA).

Blood transfusions can provide life-saving treatment to severely anemic animals. Due to limited availability and the difficulty of storing whole blood and blood products, such as fresh frozen plasma and packed red blood cells, exotic animals often receive fresh whole blood transfusions. Little is known about elasmobranch blood types and transfusions. Conspecific cross-matches within several different elasmobranch species were negative, indicating that in an emergency situation a single whole blood transfusion may be possible without causing a transfusion reaction. Experimental transfusions between healthy conspecific Atlantic rays ( Dasyatis sabina) showed no adverse reactions and autotransfusions in marbled electric rays ( Torpedo marmorata) were successful. There are no published reports of blood transfusions performed on clinically abnormal elasmobranchs. The following case series documents blood transfusions performed on seven cownose rays ( Rhinoptera bonasus) and one short-tail stingray ( Dasyatis brevicaudata). All rays were treated with the same protocol, which included pretreatment with steroids and antibiotics followed by an intravenous transfusion of freshly collected, heparinized, whole blood. Three animals survived and currently exhibit no abnormal clinical signs. Two animals died 55 days and 100 days post transfusion. Three animals died 2-22 days post transfusion. Although complications from blood transfusions could not be ruled out, all five animals that died had other health problems that likely contributed to their demise. All eight animals would almost certainly have died without a blood transfusion as they were severely anemic and moribund at the time of presentation. The methods described in this paper may be useful in the treatment of severely anemic elasmobranchs and this is the first report of blood transfusions in clinically abnormal elasmobranchs.