Relationship between red blood cell lifespan and endogenous carbon monoxide in the common bottlenose dolphin and beluga.

Certain deep-diving marine mammals [i.e., northern elephant seal (Mirounga angustirostris), Weddell seal (Leptonychotes weddellii)] have blood carbon monoxide (CO) levels that are comparable with those of chronic cigarette smokers. Most CO produced in humans is a byproduct of heme degradation, which is released when red blood cells (RBCs) are destroyed. Elevated CO can occur in humans when RBC lifespan decreases. The contribution of RBC turnover to CO concentrations in marine mammals is unknown. Here, we report the first RBC lifespans in two healthy marine mammal species with different diving capacities and heme stores, the shallow-diving bottlenose dolphin (Tursiops truncatus) and deep-diving beluga whale (Delphinapterus leucas), and we relate the lifespans to the levels of CO in blood and breath. The belugas, with high blood heme stores, had the longest mean RBC lifespan compared with humans and bottlenose dolphins. Both cetacean species were found to have three times higher blood CO content compared with humans. The estimated CO production rate from heme degradation indicates some marine mammals may have additional mechanisms for CO production, or delay CO removal from the body, potentially from long-duration breath-holds.NEW & NOTEWORTHY This is the first study to determine the red blood cell lifespan in a marine mammal species. High concentrations of carbon monoxide (CO) were found in the blood of bottlenose dolphins and in the blood and breath of belugas compared with healthy humans. Red blood cell turnover accounted for these high levels in bottlenose dolphins, but there may be alternative mechanisms of endogenous CO production that are contributing to the CO concentrations observed in belugas.

Population Pharmacokinetics of Danofloxacin After Single Intramuscular Dose Administration in California Brown Pelicans (Pelecanus occidentalis californicus).

The pharmacokinetics of danofloxacin was investigated in rehabilitated California brown pelicans (Pelecanus occidentalis californicus) after a single intramuscular injection at a dose of 15 mg/kg body weight. The concentration of the drug in plasma was assayed by high-pressure liquid chromatography. A sparse-sampling design was used to reduce the number of samples (1-4 venipunctures) obtained from 24 brown pelicans. A population pharmacokinetic analysis with nonlinear mixed-effects modeling was used to accommodate the sparse-sampling strategy. The nonlinear mixed-effects modeling approach measured both fixed effects (typical values for the population) and random effects (between-subject variability) for this population. A 1-compartment model best represented the concentration-versus-time data after injection. After injection, the elimination half-life, peak concentration, area under the curve, and volume of distribution were 2.76 hours, 2.5 µg/mL, 13.75 µg/h/mL, and 4.35 L/kg, respectively. Rate of absorption was highly variable among the birds. The intramuscular injection of danofloxacin in pelicans at this dose produced plasma concentrations that meet therapeutic targets for bacteria with a minimum inhibitory concentration of ≤0.25 µg/mL. This dose can be used for future studies to evaluate the efficacy of danofloxacin for treating susceptible bacteria.

EVALUATION OF PLASMA 25-HYDROXYVITAMIN D, IONIZED CALCIUM, AND PARATHYROID HORMONE IN GREEN SEA TURTLES (CHELONIA MYDAS) EXPOSED TO DIFFERENT INTENSITIES OF ULTRAVIOLET B RADIATION.

For many reptile species, adequate ultraviolet B (UVB) radiation is essential for proper calcium metabolism. In this study, the effects of UVB radiation on calcium metabolism were evaluated in green sea turtles (Chelonia mydas). 25-Hydroxyvitamin D, parathyroid hormone (PTH), and ionized calcium (iCa) were measured in juvenile (n = 18, 9 indoor, 9 outdoor) and adult (n = 8, 4 indoor, 4 outdoor) turtles. All animals were fed an identical diet. Outdoor animals had access to unfiltered sunlight, whereas indoor animals were housed under artificial lighting without UVB. Mean values for 25-hydroxyvitamin D for the outdoor and indoor groups were 34.33 ± 7.98 nmol/L and 7.11 ± 1.69 nmol/L for juveniles and 73.25 ± 30.34 nmol/L and 14.0 ± 11.52 nmol/L for adults respectively. Mean values for iCa for the outdoor and indoor groups were 0.98 ± 0.07 mmol/L and 0.99 ± 0.06 mmol/L for juveniles and 1.18 ± 0.22 mmol/L and 0.97 ± 0.18 mmol/L for adults respectively. UVB exposure (P < 0.001) and age (P < 0.001) had a significant effect on 25-hydroxyvitamin D as well as a significant interaction between the two variables (P = 0.008), with highest values in adult outdoor turtles. There was a significant interaction between age group and UVB status for iCa (P = 0.036), with greater values in older outdoor turtles. 25-Hydroxyvitamin D and total calcium were positively correlated, rs = 0.39, P = 0.042. iCa and calcium-to-phosphorus ratios were also positively correlated, rs = 0.42, P = 0.027. These results suggest that UVB exposure is an important source of 25-hydroxyvitamin D for green sea turtles and has significant effects on calcium metabolism in this species. PTH values in this study were near the minimum limits of detection and suggest that current mammalian-based PTH assays are not valid for reptiles.

MAMMARY FIBROADENOMATOUS HYPERPLASIA IN A JUVENILE CALIFORNIA SEA LION ( ZALOPHUS CALIFORNIANUS).

A juvenile female California sea lion ( Zalophus californianus) presented with a firm subcutaneous mass in the right caudoventral pelvic region, measuring approximately 15 × 15 cm, and edema in the right hind flipper. The initial diagnostic workup included an ultrasound, blood analysis, and fine-needle aspirate of the mass under manual restraint. While the blood analysis was within normal limits, the ultrasound revealed a large discrete hyperechoic homogeneous mass with anastomosing hypoechoic fluid pockets that, when aspirated, was suggestive of a reactive lymph node. Short-term empiric antibiotic therapy prescribed to address possible lymphadenitis yielded no response, and edema of the right hind flipper and perianal region worsened. The sea lion was anesthetized to facilitate a more thorough exam, repeat blood analysis, obtain survey radiographs, evaluate with ultrasound, and perform biopsy of mass. The thoracic radiographs were normal; however, the large radiopaque mass was observed cranial to the pelvis on lateral abdominal view and appeared solitary on ultrasound. An exploratory surgery was performed to completely excise the mass. On histopathologic review, the mass demonstrated diffuse fibrous hyperplasia of lobular ducts of the mammary gland and mild mastitis, characterizing it as a mammary fibroadenoma. The surgical wound healed within 6 wk, and there was no recurrence of mammary hyperplasia.

Diagnostic assessment of reproductive status in white-spotted bamboo sharks (Chiloscyllium plagiosum).

Relatively little is known about elasmobranch reproductive physiology compared to other species. An increased understanding of elasmobranch reproduction would improve the success of captive breeding and may aid in situ conservation efforts by reducing demand for wild-caught individuals. The reproductive status of seven adult female white-spotted bamboo sharks (Chiloscyllium plagiosum) (WSB) was monitored via coelomic ultrasonography and analysis of plasma biochemistry and steroid hormones over 6 months. Based on ultrasonic findings, females were categorized at each blood collection time point as: no follicular activity (INACTIVE), follicles but no eggs present (ACTIVE-OVARY), and eggs present within the oviduct (ACTIVE-OVIDUCT). Triglyceride concentrations were greater for those with the ACTIVE-OVARY (75.98 mg/dL; CI 61.81-90.15 mg/dL) and ACTIVE-OVIDUCT (87.0 mg/dL; CI 70.20 to 103.81 mg/dL) categories than INACTIVE (51.81 mg/dL; CI 37.07-66.54 mg/dL) category. No significant differences were observed for PCV, total solids, calcium, phosphorus, iron or progesterone. Estradiol concentrations were less for the INACTIVE (0.15 ng/ml; CI 0.08 to 0.25 ng/ml) than ACTIVE-OVARY (0.63 ng/ml; CI 0.42 to 0.88 ng/ml) and ACTIVE-OVIDUCT (0.92 ng/ml; CI 0.64-1.26 ng/ml) category. Testosterone concentrations differed among reproductive states, being greater with the INACTIVE (0.22 ng/ml; CI 0.13 to 0.37 ng/ml) and peaking in the ACTIVE-OVIDUCT (2.12 ng/ml; CI 1.25-3.60 ng/ml) state. The ultrasonic technique was performed in a standardized manner, and the anatomy was validated via opportunistic post-mortem examinations and MRI. Using the described diagnostic techniques, reproductive status in WSB can be routinely monitored, and findings have implications for improving the success of captive breeding efforts in other elasmobranch species.

Cutaneous malignant melanoma in a Haller's round ray Urobatis halleri.

Multiple black raised nodular masses were noted on the dorsal surface of an adult male Haller's round ray Urobatis halleri. Biopsy of 2 masses was performed, and histopathology revealed proliferative sheets of melanocytes exhibiting mild anisocytosis and anisokaryosis, supporting a diagnosis of malignant melanoma. Approximately 2 mo following the biopsy procedure, the round ray became acutely anorexic and was found dead in its enclosure. A full necropsy was performed, and tissues were submitted for histopathology. The black raised nodular masses again exhibited histologic features of a melanoma. In addition to the nodular masses present, multiple flat areas of increased pigmentation were also present throughout the course of the case and were not suggestive of neoplasia histologically. The transformation of benign to malignant neoplasia has been well described in other species and may have played a role in the development of multiple tumors in this case.

MALE REPRODUCTIVE PHYSIOLOGY AND THE DEVELOPMENT OF ARTIFICIAL INSEMINATION IN THE MAGELLANIC PENGUIN (SPHENISCUS MAGELLANICUS) USING CHILLED-STORED SEMEN.

Research was performed to increase our understanding of male Magellanic penguin (Spheniscus magellanicus) reproductive biology and to develop artificial insemination (AI) technology to assist with maintaining the species' genetic diversity. Seminal traits were characterized from seven males with noncontaminated ejaculates (n = 123) displaying high in vitro motion parameters, membrane integrity, and morphology. Seven females were maintained in nest sites that permitted visual, auditory, and tactile contact with their paired male but not copulation for 18.3 ± 2.4 days before egg lay. After cloacal AI (2.6 ± 0.4 inseminations/female) with semen chilled for up to 20.5 hr at 5°C, all females produced one to two fertile eggs, with the first oviposition occurring within 7 days of plasma progesterone concentrations exceeding 0.8 ng/ml. Overall fertility was 91.7%, hatchability was 63.6%, and genetic analyses confirmed that all embryos and hatchlings were sired by AI males. The heterospermic AI design demonstrated that eggs were fertilized by spermatozoa chilled for 1.5-19.8 hr before AI and were laid 4.5-11.5 days post AI. These results contribute new data on Magellanic penguin sperm biology and demonstrate that high fertility rates after AI of chilled semen can be achieved with females remaining in proximity to their paired mate.

APHAKIC PHACOEMULSIFICATION AND AUTOMATED ANTERIOR VITRECTOMY, AND POSTRETURN MONITORING OF A REHABILITATED HARBOR SEAL (PHOCA VITULINA RICHARDSI) PUP.

A female harbor seal pup rescued along the coast of San Diego on 13 June 2012 was diagnosed with bilateral mature cataracts, apparently congenital, in association with vitreal herniation in the anterior chamber of each eye. The cataracts were surgically removed on 1 August 2012 with single-port aphakic phacoemulsification and automated anterior vitrectomy. Postoperative monitoring during the next several weeks indicated that vision had been functionally repaired and that she could visually orient to and capture live fish in three different environments and in the presence of other animals. Consequently, we equipped the seal with a satellite-linked radio transmitter and returned her to the Pacific Ocean on 21 November 2012, and then monitored her movements until radio contact ended on 2 March 2013. She remained along the San Diego coast from 21 November until 5 December 2012 when she relocated to the Coronado Islands and remained there until 26 December. She then traveled directly to San Clemente Island and remained foraging in the near-shore kelp beds there through 2 March 2013, when radio contact ended. To our knowledge, this is the first published report of cataract treatment in a marine mammal using high-frequency ultrasound to emulsify the lenses followed by suction removal of the emulsified microfragments (i.e., phacoemulsification). Moreover, the rapid postoperative recovery of the seal and its quick acclimation, orientation, navigation, and foraging in marine habitats after return to the Pacific Ocean indicates that these surgical procedures can be safe and effective treatments for cataracts in seals, with substantially reduced postsurgical complications relative to other types of lens fragmentation and removal procedures.

CHARACTERIZATION OF A FOLLICULAR CELL CARCINOMA OF THE THYROID IN A YELLOWBAR ANGELFISH (POMACANTHUS MACULOSUS).

Histopathologic diagnosis of thyroid neoplasia in teleosts is complicated, because thyroid tissue is unencapsulated, and normal tissue can have wide ectopic extensions. Assessment of thyroid hormone concentrations in conjunction with histologic evaluation of thyroid carcinoma has not been reported in teleosts, even though routinely performed in other species. A yellowbar angelfish (Pomacanthus maculosus) presented with a mass that extended beyond the gill arches. Partial surgical resection was performed, and a histologic diagnosis of thyroid carcinoma was made. Plasma from the affected fish and two control Pomacanthus were used to assess thyroid hormone concentrations using a domestic mammalian assay. Thyroid-stimulating hormone and throxine were undetectable in two of three fish. Triiodothyronine was lower in the affected fish than in controls. The tumor did not appear to be actively secreting hormone. This is the first characterization of thyroid tumors and corresponding thyroid hormones in teleosts, which may assist in diagnosis of this disease.

Treatment of ureteral calculus obstruction with laser lithotripsy in an Atlantic bottlenose dolphin (Tursiops truncatus).

An adult female bottlenose dolphin (Tursiops truncatus) presented with acute anorexia secondary to progressive azotemia (blood urea nitrogen = 213 mg/dl, creatinine [Cr] = 9.5 mg/dl) and electrolyte abnormalities (K = 7.4 mEq/L). It was later diagnosed with postrenal obstruction secondary to bilaterally obstructing ureteral calculi seen on ultrasound. Treatment of the obstruction required two endoscopic procedures, cystoscopy for ureteral stent placement and ureteroscopy to perform intracorporeal lithotripsy on the obstructing calculi. Before the first procedure, the dolphin's azotemia was stabilized with aggressive fluid therapy, peritoneal dialysis, and treatment for acidosis. Diuresis subsequent to the fluid therapy enabled passage of the right obstructing urolith. For both endoscopic procedures, the dolphin was placed in left lateral recumbency due to the peritoneal dialysis catheter in the right retroperitoneal region. For the first procedure, a 12-French (Fr) flexible cystoscope was inserted retrograde into the bladder via the urethra, whereupon a calculus was seen obstructing the left ureteral orifice. A 4.8-Fr, 26-cm double-pigtail ureteral stent was placed up the left ureter to relieve the postrenal obstruction. Inadvertent proximal migration of the left ureteral stent occurred during the procedure. However, renal parameters (serum Cr = 5.8, K = 5.4) improved significantly by the next day. For the second procedure, 28 hr later, ureteroscopy was performed to treat the calculus and replace the existing stent with a longer stent. The left ureteral calculus was pulverized into tiny fragments by using a holmium:yttrium-aluminum-garnet laser inserted through a 6.9-Fr semirigid ureteroscope. The migrated stent was visualized in the distal left ureter and replaced with a 90-cm single-pigtail ureteral stent that was sutured exterior to the urogenital slit and removed 3 days later. Renal function normalized over the next several days, and the dolphin recovered over the next 2 mo.