Identification of freeze-thaw artifact in fresh and decomposed black rockfish (Sebastes melanops) and steelhead trout (Oncorhynchus mykiss).

Identification of freeze-thaw artifact in fish can help to determine whether they have been harvested within the appropriate season and monitor adherence to fishing regulations. Recognition of freeze-specific changes will also prevent potential misinterpretation due to decomposition, disease, injury, or species variation. An initial survey using black rockfish (Sebastes melanops) identified which tissues reliably exhibit freeze artifact. Tissues were exposed to different treatments: immediate formalin fixation; refrigeration or storage at room temperature for 24, 48, or 72 hours; or freezing for 1, 8, or 28 days. Three fish underwent a combination of treatments. Tissue changes in each treatment group were compared macroscopically and microscopically. Macroscopic changes in frozen-thawed and never-frozen fish overlapped somewhat; however, microscopic findings of skeletal myocyte cavitation, lens liquefaction, and brain tissue fractures were consistent findings only in frozen-thawed tissues. A validation study was then done to establish the accuracy of microscopic analysis. Brain and paired ocular and skeletal muscle samples from 61 steelhead trout (Oncorhynchus mykiss) were fixed in formalin either fresh or after being frozen for 4 weeks. Weighted kappa values showed both high observer accuracy and interobserver agreement in the identification of freeze-thaw status. Based on these findings, microscopic changes in the skeletal muscle, eye, and brain are considered consistent and easily identifiable indicators of a previous freeze-thaw cycle and should not be confused with a pathologic process.

ANTICOAGULANT EXPOSURE IN GOLDEN EAGLE (AQUILA CHRYSAETOS) POWER LINE ELECTROCUTION AND WIND TURBINE MORTALITIES.

Golden Eagles (Aquila chrysaetos) are susceptible to anthropogenic mortality factors, including toxic compounds in the environment such as anticoagulant rodenticides (AR) and sources of man-made energy. The physical and behavioral effects of some toxins may predispose eagles to certain causes of death (COD). To investigate the influence of ARs on mortality of Golden Eagles at wind turbine farms, we randomly tested liver samples from 31 eagles found dead on wind farms and submitted to the National Fish and Wildlife Forensic Laboratory from 2013-20. The comparison group was composed of 31 Golden Eagles sampled during the same time frame with a COD of power line electrocution as a proxy for a relatively lower effort and altitude activity. Associations between COD, AR exposure, sex, and life stage were assessed. In each group, 12 birds (35%) were found to have been exposed to brodifacoum or bromadiolone prior to death. Logistic regression showed no significant association between COD and sex (P=0.194) or life stage (P=0.895). Across both mortality types, life stage was not a significant predictor of AR exposure (P=0.725), but males were more likely to have been exposed to ARs (P=0.032). These findings suggest that there is no difference in the influence of anticoagulant exposure on higher and lower altitude activity in Golden Eagles.

Qualitative identification of imidacloprid in postmortem animal tissue by gas chromatography-tandem mass spectrometry.

During an avian mass mortality event investigation at the National Fish and Wildlife Forensic Laboratory in Ashland, OR, imidacloprid became an insecticide of concern. A qualitative analytical toxicology screen of seeds, plucks (tongue, esophagus, and trachea), and ventricular contents was requested. A method for the extraction and qualitative analysis of the insecticide in animal tissues was therefore developed. The procedure relies on a combined Food Emergency Response Network (FERN) and QuEChERS (Quick, Easy, Cheap, Effective, Rugged, and Safe) approach to sample extraction followed by qualitative analysis by gas chromatography-tandem mass spectrometry. Since imidacloprid is not amenable to the conditions of gas chromatography, a trimethylsilyl derivative was created and characterized. Proposed mechanisms for the creation of this derivative and its mass spectrum are described. The imidacloprid-trimethylsilyl (TMS) derivative was detected in all samples submitted.

CHRONICALLY EMBEDDED LEAD PROJECTILES IN WILDLIFE: A CASE SERIES INVESTIGATING THE POTENTIAL FOR LEAD TOXICOSIS.

Research has demonstrated that intramuscularly embedded lead in humans and rats may cause direct plumbism, albeit rarely, and has identified risk factors to this end. To the authors' knowledge, this has not been investigated in wildlife, despite a high incidence of embedded lead in these animals secondary to cynegetic activities. Fourteen wildlife cases submitted to the National Fish and Wildlife Forensics Laboratory for cause-of-death determination had chronically embedded lead projectiles that were unrelated to the cause of death. Tissue lead levels were measured in all cases and revealed clinically significant hepatic lead levels in two cases. The results corroborate comparative literature and suggest that embedded lead fragments carry a low risk for direct plumbism, even in the face of risk factors such as fractures, inflammation, and projectile fragmentation. Wildlife morbidity and mortality from embedded lead is more commonly realized secondary to incidental ingestion and ballistic trauma rather than by direct toxicity.

Using an alternate light source to detect electrically singed feathers and hair in a forensic setting.

Mortality due to electrical injury in wildlife may occur in the form of lightning strike or power line contact. Evidence of electrical contact may be grossly obvious, with extensive singeing, curling, and blackening of feathers, fur, or skin. Occasionally, changes may be subtle, owing to lower current or reduced conductivity, making a definitive diagnosis of electrocution more difficult. We describe the use of an alternate light source in the examination of cases of lightning strike and power line contact in wildlife, and the enhanced detection of changes due to electrical currents in the hair and feathers of affected animals. Subtle changes in the wing feathers of 12 snow geese and 1 wolf that were struck by separate lightning events were made obvious by the use of an alternate light source. Similarly, this technique can be used to strengthen the evidence for power line exposure in birds.

Retrospective analysis of mortalities in elephant shrews (Macroscelididae) and tree shrews (Tupaiidae) at the Smithsonian National Zoological Park, USA.

Investigations into the cause of mortality and other important findings at necropsy were made into two families of small mammals at the Smithsonian National Zoological Park (SNZP; USA). Necropsy reports from 1976 through 2008 were reviewed for all elephant shrews in family Macroscelididae (n = 118) and all tree shrews in family Tupaiidae (n = 90) that lived for greater than 30 days at the SNZP. Causes of mortality were classified by body system and etiology to identify prevalent diseases and trends across demographics for each family. In elephant shrews, gastrointestinal disease (n = 18) and respiratory disease (n = 22) were important causes of mortality with an increased prevalence of pneumonia in adult males. Trauma was a common cause of mortality in tree shrews (n = 22). Cryptococcosis was an important cause of mortality in both families (n = 8 elephant shrews; n = 13 tree shrews). Bacterial infections, often systemic at time of mortality, were also common (n = 16 elephant shrews; n = 17 tree shrews). Arteriosclerosis was a common comorbid pathology noted at necropsy in certain populations, seen only in Elephantulus rufescens in the family Macroscelididae (n = 22) and in only males in the family Tupaiidae (n = 11). Gongylonemiasis was seen commonly in tree shrews (n = 15), as a comorbid finding, or in 5 cases directly leading to mortality. Awareness of the prevalence of these diseases can help guide prevention and intervention strategies.

Uterine adenomyosis in southern three-banded armadillos (Tolypeutes matacus).

Uterine adenomyosis was diagnosed in five southern three-banded armadillos (Tolypeutes matacus) from four different zoological collections in North America between 1995 and 2012. Two cases were diagnosed after ovariohysterectomy and histopathologic evaluation of the uteri, and the remaining cases were identified incidentally at the time of postmortem examination. Animals ranged from 5 to 14 yr of age at the time of diagnosis. Of armadillos diagnosed before postmortem examination, clinical signs included weakness, collapse, anemia, and vulvar discharge. Histopathologic evaluation of the uteri revealed well-developed, irregular endometrial glands extending into the myometrium and occasional hemorrhage within these glands. The two cases diagnosed antemortem were successfully treated with ovariohysterectomy. To the authors' knowledge, this condition has not been previously reported in Xenarthra, including armadillos.

Embryonal rhabdomyosarcoma in a Rothschild's giraffe (Giraffa camelopardalis rothschildi).

Abstract: A 3-yr-old male Rothschild's giraffe (Giraffa camelopardalis rothschildi) presented for acute swelling caudomedial to the left parietal horn. Following initial diagnostics and supportive treatment, the mass was surgically resected and intralesional chemotherapy was administered. Despite treatment, the giraffe's condition worsened and euthanasia was performed. Gross necropsy revealed neoplastic invasion and destruction of underlying parietal bone, adjacent horn base, and sinuses, and metastases in the tracheobronchial and mandibular lymph nodes and lung. Histologically, the tumor was composed of packets of anaplastic round cells. Immunohistochemical studies further characterized the tumor as an embryonal rhabdomyosarcoma. This is the first reported case of rhabdomyosarcoma in a giraffe.

Mortality of captive black-footed ferrets (Mustela nigripes) at Smithsonian's National Zoological Park, 1989-2004.

Black-footed ferret (Mustela nigripes) mortality was investigated retrospectively based on the pathology records of 107 captive animals held at Smithsonian's National Zoological Park from 1989 to 2004. The majority of deaths in neonates were due to cannibalism (n = 42; 64.6%) and maternal trauma (n = 11; 16.9%); both of these causes of mortality decreased during the study period. Prior to 2001, juvenile mortality was most often caused by gastrointestinal disease (n = 11; 52.4%), including coccidiosis, salmonellosis, and clostridium infection. In 2001, improvements in husbandry, hygiene, and medical treatment led to decreases in juvenile mortality associated with gastrointestinal disease. The most common causes of death in adult ferrets were renal or neoplastic disease. The etiology of the high prevalence of renal disease in the last 4 yr of the study is unknown; it was not associated with increasing age or inbreeding. Improved hygiene and vigilant monitoring for signs of gastrointestinal and renal disease will continue to improve the success of the captive propagation of this species.

Uterine leiomyomas in three captive eastern Bongo (Tragelaphus eurycerus isaaci).

Uterine leiomyomas occurred in three eastern bongo (Tragelaphus eurycerus isaaci) 10, 15, and 16 yr of age at three separate zoological institutions. Two animals were nulliparous, and one had given birth to 14 calves over its lifetime. Two of the leiomyomas originated in the cervix. The third originated in the uterine wall and incorporated the body of the uterus and both uterine horns. The tumors in the two animals that had never calved were large and nonresectable. They were associated with gastrointestinal clinical signs including diarrhea, anorexia, and the inability to defecate. The tumor in the animal that had calved was an incidental finding at necropsy. Uterine leiomyomas have not been previously documented in the literature on eastern bongo.