Highly Pathogenic Avian Influenza Virus A(H5N1) Clade 2.3.4.4b Infection in Free-Ranging Polar Bear, Alaska, USA.

We report a natural infection with a Eurasian highly pathogenic avian influenza A(H5N1) clade 2.3.4.4b virus in a free-ranging juvenile polar bear (Ursus maritimus) found dead in North Slope Borough, Alaska, USA. Continued community and hunter-based participation in wildlife health surveillance is key to detecting emerging pathogens in the Arctic.

Pathology of short-term dorsal fin tag-attachments in tagged and re-stranded short-beaked common dolphins Delphinus delphis on Cape Cod, MA.

Odontocetes are difficult to study in the wild, making tagging and remote tracking a valuable practice. However, evaluations of host responses at tagging sites have been primarily limited to visual observations in the field. Here we explore the macro- and microscopic pathology of dorsal fin tag attachments in 13 stranded and released short-beaked common dolphins Delphinus delphis from Cape Cod, MA that later re-stranded and died or were euthanized 1-28 d post-tagging. Tags were attached to stranded dolphins' dorsal fins using 2 methods: core biopsy or piercing. Grossly, the piercing method resulted in epidermal compression into the dermis. One tag site had a necrotic border 28 d after application. Grossly, the biopsy method resulted in minimal to no tissue reaction. Two tag sites had granulation tissue accumulation 4 and 12 d after tagging. Histopathologic findings for all tag types and animals consisted of focal epithelial loss, dermal edema, perivascular edema, inflammation and hyperplasia, and inter- and extracellular edema in the adjacent epidermis. Minor expected pathological changes given the procedure were also observed: superficial epidermal necrosis in 3 cases, and superficial bacterial colonization in 2 cases. There was no evidence of sepsis and tagging was not related to cause of re-stranding or death in any case. These gross and histopathologic findings support previous observational conclusions in small delphinids that with appropriate sterile technique, the impacts of single pin dorsal fin tagging on the animal can be minimal and localized. Of the 2 methods, core biopsy may be a better tagging method.

Investigation of SARS-CoV-2 infection and associated lesions in exotic and companion animals.

Documented natural infections with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in exotic and companion animals following human exposures are uncommon. Those documented in animals are typically mild and self-limiting, and infected animals have only infrequently died or been euthanized. Through a coordinated One Health initiative, necropsies were conducted on 5 animals from different premises that were exposed to humans with laboratory-confirmed SARS-CoV-2 infection. The combination of epidemiologic evidence of exposure and confirmatory real-time reverse transcriptase-polymerase chain reaction testing confirmed infection in 3 cats and a tiger. A dog was a suspect case based on epidemiologic evidence of exposure but tested negative for SARS-CoV-2. Four animals had respiratory clinical signs that developed 2 to 12 days after exposure. The dog had bronchointerstitial pneumonia and the tiger had bronchopneumonia; both had syncytial-like cells with no detection of SARS-CoV-2. Individual findings in the 3 cats included metastatic mammary carcinoma, congenital renal disease, and myocardial disease. Based on the necropsy findings and a standardized algorithm, SARS-CoV-2 infection was not considered the cause of death in any of the cases. Continued surveillance and necropsy examination of animals with fatal outcomes will further our understanding of natural SARS-CoV-2 infection in animals and the potential role of the virus in development of lesions.

Brucella ceti sequence type 23, 26, and 27 infections in North American cetaceans.

Brucella ceti infection is associated with a variety of disease outcomes in cetaceans globally. Multiple genotypes of B. ceti have been identified. This retrospective aimed to determine if specific lesions were associated with different B. ceti DNA sequence types (STs). Characterization of ST was performed on 163 samples from 88 free-ranging cetaceans, including common bottlenose dolphin Tursiops truncatus (T.t.; n = 73), common short-beaked dolphin Delphinus delphis (D.d.; n = 7), striped dolphin Stenella coeruleoalba (n = 3), Pacific white-sided dolphin Lagenorhynchus obliquidens (n = 2), sperm whale Physeter macrocephalus (n = 2), and harbour porpoise Phocoena phocoena (n = 1), that stranded along the coast of the US mainland and Hawaii. ST was determined using a previously described insertion sequence 711 quantitative PCR. Concordance with 9-locus multi-locus sequence typing was assessed in a subset of samples (n = 18). ST 26 was most commonly identified in adult dolphins along the US east coast with non-suppurative meningoencephalitis (p = 0.009). Animals infected with ST 27 were predominately perinates that were aborted or died shortly after birth with evidence of in utero pneumonia (p = 0.035). Reproductive tract inflammation and meningoencephalitis were also observed in adult T.t. and D.d. with ST 27, though low sample size limited interpretation. ST 23 infections can cause disease in cetacean families other than porpoises (Phocoenidae), including neurobrucellosis in D.d. In total, 11 animals were potentially infected with multiple STs. These data indicate differences in pathogenesis among B. ceti STs in free-ranging cetaceans, and infection with multiple STs is possible.

Prevalence and genotype of Toxoplasma gondii in stranded Hawaiian cetaceans.

Toxoplasma gondii is a significant threat to endangered Hawaiian wildlife including birds and marine mammals. To estimate the prevalence of T. gondii in stranded cetaceans from 1997 to 2021 in Hawai'i, we tested tissues from 37 stranded spinner dolphins Stenella longirostris and 51 stranded individuals that represented 18 other cetacean species. DNA from cetacean tissue extracts were screened using a nested polymerase chain reaction (PCR) assay targeting the Toxoplasmatinae internal transcribed spacer 1 of the nuclear ribosomal DNA. A positive result was obtained in 9 tissues examined for each of 2 spinner dolphins out of 525 tissue samples analyzed by PCR. The PCR-positive spinner dolphins had disseminated acute toxoplasmosis with necrosis, inflammation, and intralesional protozoal cysts and tachyzoites in multiple organs. Discrete positive immunostaining for T. gondii was observed in all tissues tested including the adrenal gland, brain, liver, and lung. Both positive spinner dolphins were negative for cetacean morbillivirus. The T. gondii genotyping was performed by restriction fragment length polymorphism (PCR-RFLP) based on 10 genetic markers. The PCR-RFLP analysis revealed the T. gondii belonged to PCR-RFLP-ToxoDB genotype #24, previously detected in wild pig Sus scrofa in O'ahu, bobcats Lynx rufus from Mississippi, USA, and chickens Gallus gallus from Costa Rica and Brazil. These cases represent the first report of this genotype in aquatic mammals and the second and third reports of fatal disseminated T. gondii infection in stranded spinner dolphins from Hawai'i. Nearshore species, like spinner dolphins, may be at increased risk of mortality from this parasite in marine coastal waterways via sewage systems, storm water drainage, and freshwater runoff.

Subcutaneous, abdominal, and thoracic encapsulated fat necrosis in bowhead whales Balaena mysticetus from Alaska, USA.

We describe a case series of encapsulated fat necrosis with subcutaneous, abdominal, and thoracic locations in 7 subsistence-harvested bowhead whales Balaena mysticetus. Masses had a variably-dense fibrous capsule surrounding necrotic adipocytes and calcium salts (saponification). One animal also had prior concussive injury, pleural fibrosis, and hepatic lipoma; the other animals had no significant findings. The described condition is uncommon in bowhead whales, with 7/575 (1.2%) observed from 1996 to 2015. The exact mechanisms of development of encapsulated fat necrosis in bowhead whales remain to be determined. Encapsulated fat necrosis has been reported in other baleen whales, humans, and cows. It is usually an incidental finding during post-mortem examination that needs to be differentiated from neoplastic and inflammatory lesions, as the latter may have public health implications. Assessment of further cases in bowhead whales and other baleen whales is warranted to better understand their pathogenesis.

Manatee Trichechus manatus latirostris calf mortality in Florida: a retrospective review of pathology data from 2009-2017.

High maternal investment and extended inter-calving intervals in Florida manatees Trichechus manatus latirostris make calf survivorship critical to overall population growth. However, detailed patterns of causes of mortality in calves have not been reported and state agency statistics report portions of perinatal mortality based on body length rather than actual cause of death (COD). The objectives of this study were to categorize COD based on necropsy data and geographical location in Florida for 1209 manatee calf carcasses (<236 cm total length) examined between January 2009 and December 2017 and to describe factors contributing to calf mortality. Results indicated COD was attributed to natural causes (47%, n = 573), cold stress syndrome (38%, n = 457), watercraft injury (13%, n = 155), or other human-related causes (2%, n = 24). Natural causes were the leading COD for small calves <151 cm, with death due to stillbirth or dystocia most frequent (48%, n = 273/573). Enteric trematodiasis contributed to a large proportion of deaths from natural causes in large calves within the southwest region of Florida, with an increasing annual trend. Brevetoxicosis contributed substantially to natural causes within the southwest region exclusively and was commonly comorbid with enteric trematodiasis. Cold stress syndrome was the leading cause of death for large calves (151-235 cm), with the Atlantic region having the highest proportion of cases. Watercraft injury was a sustained threat to large calves, especially within the southwest region. This report provides details on specific health threats and patterns of mortality among manatee calves.

Rectus sheath catheters-a novel approach to perioperative analgesia for colorectal surgery in an enhanced recovery after surgery (ERAS) protocol: a case series.

INTRODUCTION: Opioids have played a critical role in the management of perioperative pain following abdominal surgery. Increasing attention is being paid to the deleterious side effects and limitations of this practice. This case report offers a novel alternative to opioid-based analgesia in the form of rectus sheath catheters (RSCs) which we employed as part of an enhanced recovery after surgery (ERAS) protocol. METHODS: Three patients underwent laparoscopic- assisted colorectal surgery and were treated intra- and postoperatively with local anesthesia administered via bilateral rectus sheath catheters as well as by multimodal adjuncts. Evaluations of the patients' pain scores, opioid usage, and abdominal sensitivity to sharp stimuli were conducted daily. RESULTS: The patients demonstrated a substantially lessened opioid requirement over their hospital stay with two of them requiring no opioid analgesic medications postoperatively. DISCUSSION: We suggest that the incorporation of these catheters into an ERAS protocol can play an important role in further reducing perioperative opioid usage for procedures in which pain control can be especially challenging.

Persistent Mü̈llerian duct syndrome in a beluga whale Delphinapterus leucas.

This case study describes a persistent Mü̈llerian duct syndrome (PMDS), a rare form of XY disorder of sex development (DSD), in a mature (>15 yr) beluga whale Delphinapterus leucas. The phenotypically and genetically male beluga whale had both Mü̈llerian (paramesonephric) and Wolffian (mesonephric) duct derivatives. A mild hydrometra was present. Gross and histological analyses indicated the testes were atrophied. Histopathologic findings in the testes also included fibrosis in addition to ductus deferens ductular dilation, epididymal ductular dilation, lymphoplasmacytic balanitis, uterine glandular atrophy, and endometrial fibrosis. DSDs are rarely reported in cetaceans, and PMDS has never been described in a cetacean.

Metastatic testicular Sertoli cell tumor in a free-ranging cryptorchid adult spotted seal Phoca largha in North Slope, Alaska, USA.

This case describes a metastatic Sertoli cell tumor (SCT) with lymphatic spread to the abdominal and thoracic lymph nodes, pancreas, and adrenal gland in an adult spotted seal Phoca largha. The neoplasm was composed of tubules lined by palisading neoplastic cells separated by a variably dense fibrous stroma. This pinniped was 1 of 2 cryptorchid seals and the sole case of genital neoplasia among 70 ice seals necropsied by the North Slope Borough from 2012 to 2017. Overall, SCTs are rarely reported in marine mammals.

Habitat use pattern of the giant parasitic nematode Crassicauda magna within the pygmy sperm whale Kogia breviceps.

The giant (>3 m) parasitic nematode Crassicauda magna infects kogiid whales, although only 3 studies to date have provided detailed descriptions of these worms, all based upon fragmented specimens. These fragments were found within the neck region of kogiids, an unusual anatomic site for this genus of parasites. C. magna is a species-specific parasite among kogiids, infecting only pygmy sperm whales Kogia breviceps, and with a primarily cervico-thoracic distribution. To date, however, the pattern of habitat use within the host and transmission path of this parasite remain unknown. We used detailed dissections (n = 12), histological examination of host tissues (n = 2), and scanning electron microscopy of excised nematodes (n = 7) to enhance our understanding of this host-parasite relationship. Results revealed that a critical habitat for the parasite is an exocrine gland in the whale's ventral cervical region. C. magna male and female tails were found intertwined within the glandular lumen, and eggs were observed within its presumed secretion, illuminating the transmission path out of the host. The cephalic ends of these worms were often meters away (curvilinearly), embedded deeply within epaxial muscle. A single worm's complete, tortuous 312 cm course, from the gland to its termination in the contralateral epaxial muscle, is described for the first time. This study also provides the first scanning electron micrographs of C. magna, which illustrate taxonomically important features of the heads and tails of both male and female worms.

Discrimination between bycatch and other causes of cetacean and pinniped stranding.

The challenge of identifying cause of death in discarded bycaught marine mammals stems from a combination of the non-specific nature of the lesions of drowning, the complex physiologic adaptations unique to breath-holding marine mammals, lack of case histories, and the diverse nature of fishing gear. While no pathognomonic lesions are recognized, signs of acute external entanglement, bulging or reddened eyes, recently ingested gastric contents, pulmonary changes, and decompression-associated gas bubbles have been identified in the condition of peracute underwater entrapment (PUE) syndrome in previous studies of marine mammals. We reviewed the gross necropsy and histopathology reports of 36 cetaceans and pinnipeds including 20 directly observed bycaught and 16 live stranded animals that were euthanized between 2005 and 2011 for lesions consistent with PUE. We identified 5 criteria which present at significantly higher rates in bycaught marine mammals: external signs of acute entanglement, red or bulging eyes, recently ingested gastric contents, multi-organ congestion, and disseminated gas bubbles detected grossly during the necropsy and histologically. In contrast, froth in the trachea or primary bronchi, and lung changes (i.e. wet, heavy, froth, edema, congestion, and hemorrhage) were poor indicators of PUE. This is the first study that provides insight into the different published parameters for PUE in bycatch. For regions frequently confronted by stranded marine mammals with non-specific lesions, this could potentially aid in the investigation and quantification of marine fisheries interactions.

Systemic Edwardsiella tarda infection in a Western African lungfish (Protopterus annectens) with cytologic observation of heterophil projections.

This report describes a case of systemic bacterial infection caused by Edwardsiella tarda in a Western African lungfish (Protopterus annectens) exposed to poor environmental and husbandry conditions. The fish presented with a large, external ulcerative lesion and died 2 weeks after developing anorexia. Histological evaluation revealed multifocal areas of necrosis and heterophilic and histiocytic inflammation throughout multiple tissues. Gram stain identified small numbers of intra- and extracellular monomorphic Gram-negative 1 to 2 µm rod-shaped bacilli. Cytology of lung granuloma, kidney and testes imprints identified heterophilic inflammation with phagocytosis of small monomorphic bacilli and some heterophils exhibiting cytoplasmic projections indicative of heterophil extracellular traps (HETs). Initial phenotypic analysis of isolates from coelomic fluid cultures identified E. tarda. Subsequent molecular analysis of spleen, liver and intestine DNA using an E. tarda-specific endpoint PCR assay targeting the bacterial fimbrial subunit yielded a 115 bp band. Sequencing and BLASTN search revealed the sequence was identical (76/76) to E. tarda strain FL95-01 (GenBank acc. CP011359) and displayed 93% sequence identity (66/71) to Edwardsiella hoshinae strain ATCC 35051 (GenBank acc. CP011359). This is the first report of systemic edwardsiellosis in a lungfish with concurrent cytologically identified structures suggestive of HETs.

Gastric pH and Toxin Factors Modulate Infectivity and Disease Progression After Gastrointestinal Exposure to Bacillus anthracis.

Gastrointestinal (GI) anthrax is the most prevalent form of naturally acquired Bacillus anthracis infection, which is associated with exposure to vegetative bacteria in infected meat (carnivores) or to fermented rumen contents (herbivores). We assessed whether key host and pathogen factors modulate infectivity and progression of infection using a mouse model of GI infection. Gastric acid neutralization increases infectivity, but 30%-40% of mice succumb to infection without neutralization. Mice either fed or fasted before exposure showed similar infectivity rates. Finally, the pathogen's anthrax lethal factor is required to establish lethal infection, whereas its edema factor modulates progression and dissemination of infection.

Hepatic lipomas and myelolipomas in subsistence-harvested bowhead whales Balaena mysticetus, Alaska (USA): a case review 1980-2016.

We describe a case series of benign hepatic fatty tumors in 10 subsistence-harvested bowhead whales. Microscopic features included lipomatous and myelolipomatous masses. Extensive atrophy and/or destruction of hepatic parenchyma was not observed. No other significant disease was present except in an animal with unrelated chronic pleuritis. Based on our longitudinal case series (1980-2016) which identified 1-2 hepatic lipomas and myelolipomas in landed whales annually at Barrow, Alaska (USA), since 2012, hepatic lipomas and myelolipomas are occasionally seen in hunter-harvested bowhead whales. A conservative estimate for the percentage of bowhead whales with hepatic fatty tumors in landed whales in Barrow from 2012 to 2016 was 6% (7/111). The pathogenesis and exact cell origin of these benign fatty tumors in bowhead whales is undetermined. Assessment of further cases is warranted to better define the tissue distribution and pathogenesis of these tumors in bowhead whale liver.

Disseminated toxoplasmosis Toxoplasma gondii in a wild Florida manatee Trichechus manatus latirostris and seroprevalence in two wild populations.

Marine mammals are important indicators for ecosystem health and serve as sentinel species for infectious agents including zoonoses. Histological examination of tissues from a stranded Florida manatee Trichechus manatus latirostris revealed protozoal cysts in the cerebrum and intrahistiocytic tachyzoites in the liver and caudal mesenteric lymph node. Disseminated Toxoplasma gondii infection was confirmed by immunohistochemistry and sequencing of the nuclear ribosomal internal transcribed spacer region of formalin-fixed tissues. The lack of baseline information on Florida manatees' exposure to this pathogen prompted a study into the seroprevalence of T. gondii in 2 separate geographic habitats in Florida, USA, during the winters from 2011-2014. Serum was collected during routine health assessments of 44 apparently healthy manatees from Crystal River (n = 26) on the west central coast of Florida and Brevard County (n = 18) on the east coast of Florida. Serum was screened for detection of T. gondii immunoglobulin G (IgG) antibodies via the modified agglutination test. Two animals from Crystal River from 2011 and 2012 (7.7%) and one animal from Brevard County from 2011 (5.6%) tested positive for T. gondii antibodies. Overall seroprevalence for T. gondii was low in the 2 sampled populations and may reflect a low seroprevalence or animal susceptibility. However, continued monitoring of this pathogen in aquatic ecosystems is warranted due to both possible anthropogenic sources and zoonotic potential.

Protozoal-related mortalities in endangered Hawaiian monk seals Neomonachus schauinslandi.

Protozoal infections have been widely documented in marine mammals and may cause morbidity and mortality at levels that result in population level effects. The presence and potential impact on the recovery of endangered Hawaiian monk seals Neomonachus schauinslandi by protozoal pathogens was first identified in the carcass of a stranded adult male with disseminated toxoplasmosis and a captive monk seal with hepatitis. We report 7 additional cases and 2 suspect cases of protozoal-related mortality in Hawaiian monk seals between 2001 and 2015, including the first record of vertical transmission in this species. This study establishes case definitions for classification of protozoal infections in Hawaiian monk seals. Histopathology and immunohistochemistry were the primary diagnostic modalities used to define cases, given that these analyses establish a direct link between disease and pathogen presence. Findings were supported by serology and molecular data when available. Toxoplasma gondii was the predominant apicomplexan parasite identified and was associated with 100% of mortalities (n = 8) and 50% of suspect cases (n = 2). Incidental identification of sarcocysts in the skeletal muscle without tissue inflammation occurred in 4 seals, including one co-infected with T. gondii. In 2015, 2 cases of toxoplasmosis were identified ante-mortem and shared similar clinical findings, including hematological abnormalities and histopathology. Protozoal-related mortalities, specifically due to toxoplasmosis, are emerging as a threat to the recovery of this endangered pinniped and other native Hawaiian taxa. By establishing case definitions, this study provides a foundation for measuring the impact of these diseases on Hawaiian monk seals.

ACUTE PHASE PROTEINS AS A MARKER OF RESPIRATORY INFLAMMATION IN PRZEWALSKI'S HORSE (EQUUS FERUS PRZEWALSKII).

Acute phase proteins are sensitive markers of inflammation, which are highly conserved across taxa. Although the utility of these proteins are becoming well defined in human and domestic animal medical fields, their role in nondomestic species remains unclear. In this communication, a 20-yr-old Przewalski's horse was presented for unresolving aspiration pneumonia, which cultured a unique Actinomyces-like bacteria. Despite waxing and waning clinical signs and minimal changes on baseline hematologic analysis, protein electrophoresis, serum amyloid A, and surfactant protein D serum concentrations showed changes that more accurately reflected the clinical severity of this case.

REPRODUCTIVE NEOPLASMS IN WILD AND LONG-TERM CAPTIVE FEMALE FLORIDA MANATEES (TRICHECHUS MANATUS LATIROSTRIS).

Few reports of neoplastic diseases in manatees exist in the veterinary literature. This case series presents reproductive neoplasia noted in eight wild and long-term captive female Florida manatees (Trichechus manatus latirostris) obtained through carcass recovery and animal rehabilitation programs between April 2009 and May 2014. All cases were evaluated histologically, and diagnoses of uterine carcinoma (n = 1), granulosa cell tumor (n = 2), ovarian adnexal tumor (n = 1), and leiomyoma (n = 5) were made. The underlying cause of tumor development and effects on reproductive success is currently unknown, but possible asymmetric reproductive aging and/or a correlation between obesity and reproductive disorder in long-term nonreproductive female manatees are of interest and warrant further investigation.