Brucella-induced abortions and infection in bottlenose dolphins (Tursiops truncatus).

Two bottlenose dolphins (Tursiops truncatus) aborted fetuses that died as a result of Brucella infection. Brucella placentitis occurred in both cases. Infected placenta and vaginal/uterine fluids may transmit Brucella species to other cetaceans. In a third case, an identical organism was cultured from lung necropsy tissue of an adult female T. truncatus. Microbiology, specific polymerase chain reaction, and pulsed-field gel electrophoresis results supported the designation of an additional genomic group(s), Brucella delphini, for isolates adapted to T. truncatus. Current serologic diagnostic tests reliable for known Brucella species are unreliable in detecting dolphin brucellosis. Our findings, together with previous reports, suggest that dolphin brucellosis is a naturally occurring disease that can adversely impact reproduction in cetaceans. The zoonotic significance of cetacean brucellosis is unknown, although the disease has not been reported in people who have frequent contact with dolphins. Further studies on the zoonotic aspects, distribution, prevalence, virulence, and impact of this disease in cetaceans and other marine mammal species are needed.     

Proliferative dermatitis associated with a novel alphaherpesvirus in an Atlantic bottlenose dolphin (Tursiops truncatus).

Herpesviruses and herpes-like viruses have been reported in only a small number of species of cetaceans, and, to date, clinical manifestations have been either as a life-threatening, disseminated infection or as a non-life-threatening dermatitis. A stranded juvenile Atlantic bottlenose dolphin, Tursiops truncatus, was admitted to the Dolphin and Whale Hospital for rehabilitation. On initial physical examination, the rostral skin had multifocal regions of hyperplasia, and the skin of the dorsum contained a large number of small papules. Histologically, epithelial hyperplasia was evident, and clusters of epithelial cells contained 5-15-microm intranuclear inclusion bodies. Transmission electron microscopic investigation revealed numerous 170-190-nm enveloped virions in both the intracellular spaces and the cytoplasm of epithelial cells, with numerous nucleocapsids noted in epithelial cell nuclei. Consensus primer polymerase chain reaction identified the presence of a novel herpesvirus associated with the lesions. Phylogenetic analysis of the deduced amino acid sequences of the herpesvirus DNA polymerase gene fragment showed it to align with alphaherpesvirus sequences from humans and domestic animals. Although clearly distinct, it was most closely related to two previously described alphaherpesviruses of dolphins. This case represents the first documentation of herpesvirus dermatitis in the Atlantic bottlenose dolphin.     

Fatal necrotizing fasciitis and myositis in a captive common bottlenose dolphin (Tursiops truncatus) associated with Streptococcus agalactiae.

A common bottlenose dolphin (Tursiops truncatus) was presented for necropsy after acute onset of gastrointestinal signs and cutaneous lesions that rapidly progressed to death. Gross and microscopic findings were characterized by locally extensive severe necrohemorrhagic fasciitis and cellulitis, and severe necrotizing myositis in the head and dorsocranial thorax, with numerous disseminated gram-positive cocci. Streptococcus agalactiae was isolated from the lesions and from visceral organs (liver and lung), and it was identified by standard microbiology techniques. This communication is the first report of necrotizing fasciitis in a marine mammal associated with S. agalactiae.     

Phylogenetic analysis of marine mammal herpesviruses

Five novel DNA-dependent DNA polymerase (Dpol) herpesviral sequences were generated using nested consensus polymerase chain reaction (PCR) in clinical samples from a harbor seal (Phoca vitulina), bottlenose dolphin (Tursiops truncatus), orca (Orcinus orca), California sea lion (Zalophus californianus), and a Phocid herpesvirus 2 (PhHV-2) isolate from a harbor seal (used as positive control). These novel sequences and other representative herpesvirus sequences were included in Bayesian and Maximum Likelihood analyses to illustrate the phylogeny of herpesviruses amongst the marine mammal host species and in comparison to those of other animals. All 19 novel and known marine mammal herpesviruses included in the analyses aligned with members of the Alphaherpesvirinae or Gammaherpesvirinae subfamilies. The novel harbor seal herpesvirus clustered with members of the Macavirus genus, subfamily Gammaherpesvirinae. The novel bottlenose dolphin herpesvirus clustered together in a monophyletic group with another delphinid alphaherpesvirus but could not be associated with an established genus. The orca herpesvirus also clustered with a delphinid alphaherpesvirus and formed a separate clade. The sea lion herpesvirus clustered with PhHV-2. PhHV-1 clustered with varicelloviruses and PhHV-2 clustered strongly in the Gammaherpesvirinae genus Percavirus. All cetacean gammaherpesviruses formed a monophyletic clade and could not be associated with an established gammaherpesviral genus.     

Malignant seminoma with metastasis, Sertoli cell tumor, and pheochromocytoma in a spotted dolphin (Stenella frontalis) and malignant seminoma with metastasis in a bottlenose dolphin (Tursiops truncatus)

Seminoma with metastasis was diagnosed in a spotted dolphin (Stenella frontalis) and an Atlantic bottlenose dolphin (Tursiops truncatus). Sertoli cell tumor and pheochromocytoma were also diagnosed in the spotted dolphin. The spotted and bottlenose dolphins were adult males that stranded and died on the coasts of northwest Florida and southeast North Carolina, respectively. Neoplasia is infrequently reported in cetaceans. This is the first report of seminoma, Sertoli cell tumor, and pheochromocytoma in a dolphin, the first report of three distinct neoplasms in a dolphin, and one of the few reports of malignant neoplasia in dolphins.     

Sublingual squamous cell carcinoma in an Atlantic bottlenose dolphin (Tursiops truncatus).

A 22-yr-old captive-born Atlantic bottlenose dolphin (Tursiops truncatus) presented with a nonhealing sublingual mucosal ulcer that was diagnosed histologically as a squamous cell carcinoma, the first such report in a dolphin. The lesion was excised completely and has not recurred.     

Morphology of the lymph nodes in bottlenose dolphin (Tursiops truncatus) and striped dolphin (Stenella coeruleoalba) from the Adriatic Sea

Morphology of the lymph nodes was examined in six bottlenose dolphins (Tursiops truncatus) and three striped dolphins (Stenella coeruleoalba) from the Adriatic Sea. All animals had been found dead in nature. One group of the nodes was taken from the tracheal branching area and was marked as bifurcational lymph node, and the other group was taken from the mesenteric root and was marked as mesenteric lymph node. Microscopic analysis showed that the lymph nodes in both dolphin species were surrounded by a connective tissue capsule comprising smooth muscle cells. The parenchyma of the mesenteric and bifurcational lymph nodes in bottlenose dolphin was divided into the peripherally situated cortex with the lymphatic nodules and diffuse lymphatic tissue, and the centrally situated medulla structured of the medullary cords separated by the medullary sinuses. These lymph nodes structurally correspond to the lymph nodes in the majority of terrestrial mammals. The mesenteric lymph node of striped dolphin also had a peripherally situated cortex and a centrally positioned medulla as the majority of terrestrial mammals. In the bifurcational lymph nodes of striped dolphin, there was a central dense lymphatic tissue with the lymphatic nodules and a peripheral less dense lymphatic tissue structured of the cell cords and sinuses. The bifurcational lymph node in striped dolphin resembled porcine lymph nodes and belonged to the inverse lymph nodes.     

Congenital diffuse hyperplastic goiter associated with perinatal mortality in 11 captive-born bottlenose dolphins (Tursiops truncatus).

Diffuse hyperplastic goiter was diagnosed by histopathology in 11 perinatal bottlenose dolphins (Tursiops truncatus) that died at four separate zoos and aquaria. Thyroid morphology of these animals was compared with the histologically normal thyroids of two stranded wild bottlenose dolphin calves, a neonate and a 2-mo-old calf. Histologic changes included reduced follicular luminal diameter, markedly reduced or absent luminal colloid, hypertrophy of follicular epithelium, and follicular dysplasia. The etiology of the thyroid gland lesion was not identified. Cause of death was not determined for most of these animals, but they were presumed to have died from metabolic derangements associated with the thyroid lesion, drowning, or dystocia.     

Bronchoscopic and serologic diagnosis of Aspergillus fumigatus pulmonary infection in a bottlenose dolphin (Tursiops truncatus).

A 4-yr-old male bottlenose dolphin (Tursiops truncatus) developed an Aspergillus fumigatus pneumonia. Fungal elements were identified by cytology and microbiology from endoscopic bronchoalveolar lavage and brushings of a raised yellow endobronchial lesion. The results of qualitative immunodiffusion serology, a technique that identifies specific circulating antibodies to Aspergillus fumigatus, were suggestive of an active infection. The dolphin was treated with itraconazole for over 2 yr, which resulted in remission of clinical signs. Pneumonia caused by Aspergillus sp. accounts for the large majority of pulmonary mycoses in marine mammals. Bronchoscopy facilitated an early definitive diagnosis, accurate treatment, and remission.     

An insight into the epidemiology of dolphin morbillivirus worldwide

Serum samples from 288 cetaceans representing 25 species and originating from 11 different countries were collected between 1995 and 1999 and examined for the presence of dolphin morbillivirus (DMV)-specific antibodies by an indirect ELISA (iELISA) (N = 267) or a plaque reduction assay (N = 21). A total of 35 odontocetes were seropositive: three harbour porpoises (Phocoena phocoena) and a common dolphin (Delphinus delphis) from the Northeastern (NE) Atlantic, a bottlenose dolphin (Tursiops truncatus) from Kent (England), three striped dolphins (Stenella coeruleoalba), two Risso's dolphins (Grampus griseus) and a bottlenose dolphin from the Mediterranean Sea, one common dolphin from the Southwest (SW) Indian Ocean, three Fraser's dolphins (Lagenodelphis hosei) from the SW Atlantic, 18 long-finned pilot whales (Globicephala melas) and a bottlenose dolphin from the SW Pacific as well as a captive bottlenose dolphin (Tursiops aduncus) originally from Taiwan. The presence of morbillivirus antibodies in 17 of these animals was further examined in other iELISAs and virus neutralization tests. Our results indicate that DMV infects cetaceans worldwide. This is the first report of DMV-seropositive animals from the SW Indian, SW Atlantic and West Pacific Oceans. Prevalence of DMV-seropositives was 85.7% in 21 pilot whales from the SW Pacific and both sexually mature and immature individuals were infected. This indicates that DMV is endemic in these animals. The same situation may occur among Fraser's dolphins from the SW Atlantic. The prevalence of DMV-seropositives was 5.26% and 5.36% in 19 common dolphins and 56 harbour porpoise from the NE Atlantic, respectively, and 18.75% in 16 striped dolphins from the Mediterranean. Prevalence varied significantly with sexual maturity in harbour porpoises and striped dolphins; all DMV-seropositives being mature animals. The prevalence of seropositive harbour porpoise and striped dolphins appeared to have decreased since previous studies. These data suggest that DMV is not endemic within these populations, that they are losing their humoral immunity against the virus and that they may be vulnerable to new epidemics.     

Saksenaea vasiformis and Apophysomyces elegans zygomycotic infections in bottlenose dolphins (Tursiops truncatus), a killer whale (Orcinus orca), and pacific white-sided dolphins (Lagenorhynchus obliquidens).

During a 10-yr period, a killer whale (Orcinus orca), two Pacific white-sided dolphins (Lagenorhynchus obliquidens), and two bottlenose dolphins (Tursiops truncatus), all housed at SeaWorld of Texas from 1991 to 2001, were infected with fungi from the class Zygomycetes. In four out of five cases, the fungi were identified as either Saksenaea vasiformis or Apophysomyces elegans. All fungi in the class Zygomycetes aggressively invade the vascular system. Death occurred within 23 days after the initial clinical signs. The primary site of infection involved the s.c. tissue and skeletal musculature. In one case, infection originated in the placenta and uterus of a periparturient animal. All cases exhibited systemic spread of the organisms, including two to the central nervous system. The fifth and most recent case, a bottlenose dolphin, was treated with liposomal nystatin, an antifungal formulation with reduced nephrotoxicity. This animal initially responded to therapy; however, 14 days after cessation of therapy, fungal growth reoccurred. Thus, the animal was euthanatized 39 days after the initial clinical signs. This drug represents a promising treatment option if combined with early disease detection and aggressive tissue resection.     

Development and validation of monoclonal and polyclonal antibodies for the detection of immunoglobulin G of bottlenose dolphins (Tursiops truncatus).

Antibodies directed against species-specific immunoglobulin G (IgG) have a broad range of applications in serologic and immunologic research and in the development of clinical assays. Validated anti-IgG antibodies for marine mammal species are in short supply. The objective of this study was to produce and validate antibodies with specificity for IgG of the common bottlenose dolphin (Tursiops truncatus). Bottlenose dolphin IgG was purified using protein G. Two mouse monoclonal antibodies and a rabbit polyclonal antibody were developed from mice and rabbits immunized with bottlenose dolphin IgG. The specificity of the monoclonal antibodies and the polyclonal antibody for bottlenose dolphin IgG was first verified by Western blot analysis and enzyme-linked immunosorbent assay (ELISA). For further validation, both monoclonal antibodies and the polyclonal antibody were incorporated in an indirect ELISA for the detection of the immune response of bottlenose dolphins to a vaccine antigen. Three bottlenose dolphins were immunized with a commercial Erysipelothrix rhusiopathiae vaccine, and serial blood samples were collected from all dolphins for measurement of levels of circulating antibodies. Seroconversion was observed in all 3 dolphins by use of both monoclonal antibodies and the polyclonal antibody. Circulating antibodies were detectable as early as 6 days after immunization in 1 dolphin. Peak antibody levels were detected 14 days after the immunization. The ability to detect seroconversion in all 3 immunized bottlenose dolphins firmly establishes the specificity of the monoclonal antibodies and the polyclonal antibody for IgG of the common bottlenose dolphin.     

Histological Structure of the Adrenal Gland of the Bottlenose Dolphin (Tursiops truncatus) and the Striped Dolphin (Stenella coeruleoalba) from the Adriatic Sea

The structure of the adrenal gland was studied in 11 bottlenose dolphins ( Tursiops truncatus ), and five striped dolphins ( Stenella coeruleoalba ). These species are legally protected in Croatia. All examined animals died of natural causes and were found stranded along eastern Adriatic coast. In both species the adrenal gland consists of a cortex and a medulla; the cortex is divided into three zones. Whereas in the bottlenose dolphin, there is a zona arcuata which contains columnar cells arranged in the form of arches; in the striped dolphin this zone is replaced by zona glomerulosa containing rounded clusters of polygonal cells. In both species, the zona fasciculata consists of radially oriented cords of polygonal cells, whereas in zona reticularis cells are arranged in branching and anastomosing cords. The adrenal medulla in both species contains dark, epinephrine-secreting cells and light norepinephrine-secreting cells. Epinephrine-secreting cells are localized in the outer part of the medulla, whereas norepinephrine-secreting cells are found in the inner part, arranged in clusters and surrounded by septa of thin connective tissue. The gland is surrounded by a thick connective-tissue capsule, from where thick trabeculae extend towards the interior. In the bottlenose dolphin, group of cells resembling both medullar and cortical cells can be seen within the capsule; whereas only groups of cells resembling cortical cells are found within the capsule of the striped dolphin. In the bottlenose dolphin invagination of the adrenal cortex into the medulla is obvious as well as medullary protrusions extending through cortex to the connective tissue capsule.     

Involvement of microbial respiratory pathogens in acute interstitial pneumonia in feedlot cattle

OBJECTIVE: To evaluate viral and bacterial respiratory pathogens and Mycoplasma spp isolated from lung tissues of cattle with acute interstitial pneumonia (AIP) and cattle that had died as a result of other causes. SAMPLE POPULATION: 186 samples of lung tissues collected from cattle housed in 14 feedlots in the western United States. PROCEDURE: Lung tissues were collected during routine postmortem examination and submitted for histologic, microbiologic, and toxicologic examinations. Histologic diagnoses were categorized for AIP, bronchopneumonia (BP), control samples (no evidence of disease), and other disorders. RESULTS: Cattle affected with AIP had been in feedlots for a mean of 1272 days before death, which was longer than cattle with BP and control cattle. Detection of a viral respiratory pathogen (eg, bovine respiratory syncytial virus [BRSV], bovine viral diarrhea virus, bovine herpesvirus 1, or parainfluenza virus 3) was not associated with histologic category of lung tissues. Bovine respiratory syncytial virus was detected in 8.3% of AIP samples and 24.0% of control samples. Histologic category was associated with isolation of an aerobic bacterial agent and Mycoplasma spp. Cattle with BP were at greatest risk for isolation of an aerobic bacterial agent and Mycoplasma spp. CONCLUSIONS AND CLINICAL RELEVANCE: Analysis of these results suggests that AIP in feedlot cattle is not a consequence of infection with BRSV. The increased, risk of isolation of an aerobic bacterial agent from cattle with AIP, compared with control cattle, may indicate a causal role or an opportunistic infection that follows development of AIP.     

Replication of parainfluenza type-3 virus and bovine respiratory syncytial virus in isolated bovine type-II alveolar epithelial cells.

The objectives of our research were to determine whether bovine pulmonary type-II alveolar epithelial cells could be isolated from bovine lung and maintained in tissue culture and to determine whether isolated bovine type-II alveolar epithelial cells would support productive viral replication of bovine parainfluenza type-3 virus and bovine respiratory syncytial virus. Type-II alveolar epithelial cells were isolated from lungs of 4- to 7-day-old male Holstein calves by enzymatic dissociation of pulmonary tissue with trypsin and by separation of cells with the use of filtration and centrifugation on continuous Percoll gradients. Cells were further separated by panning on IgG-coated plastic plates and by lectin binding. Isolated type-II alveolar cells were maintained on basement membrane-coated tissue cultured plates. In culture, type-II cells formed alveolar structures and maintained other cytologic features of type-II cells, including osmiophilic lamellar inclusions. Cell cultures were inoculated with and supported productive replication of bovine parainfluenza type-3 virus and bovine respiratory syncytial virus. This was determined by recovery of infectious viruses from inoculated cell cultures and by identification of viral structures in type-II alveolar epithelial cells by transmission electron microscopy.     

Calf pneumonia

Infectious calf pneumonia is a high-mortality pneumonia of housed dairy-type calves. Viruses are important etiologic agents and infection with bovine respiratory syncytial virus (RSV) and parainfluenza type 3 virus (PI-3 virus) may result in extensive, and sometimes fatal, lung damage. Respiratory viral infections are frequently followed by mycoplasmal and secondary bacterial invasion of the lower respiratory tract, which increases the extent and severity of lung damage. Bad housing, particularly bad ventilation, will increase the severity of pneumonia outbreaks. Although the source of respiratory viral infections is not always obvious, it is likely that a proportion of calves acquired infection from their dams early in life. The possibility of cross-infections from other domestic animals and from humans must also be considered. Diagnosis of respiratory virus infections necessitates submission of suitable respiratory tract specimens that are taken at an early stage in the outbreak together with paired sera. Various therapeutic and prophylactic regimens for the control of calf pneumonia are described.     

Scanning Electron Microscopy of the Orbital Harderian Gland in the Male Atlantic Bottlenose Dolphin (Tursiops truncatus)

The ultrastructure of the Harderian gland of Atlantic bottlenose dolphin ( Tursiops truncatus ) was examined by scanning electron microscopy (SEM). We found the following surface features: the typical round appearance of the ascinar glandular unit with a finely granular surface, a thin cortex and immediately below two types of cells: type I cells (characterized by small lipid vacuoles) and type II cells (characterized by large lipid vacuoles). It has been suggested that different cells forms represent a single cell type in varying activity states. Additionally, a coalescent tubular complex, a small balloon-like structures and large globular structures were observed. These structures may be reservoirs of secretion products.     

Dermatitis with invasive ciliated protozoa in dolphins that died during the 1987-1988 Atlantic Bottlenose Dolphin morbilliviral epizootic

Dermatitis with intradermal cilated protozoa was identified in 18 of 95 (19%) Atlantic Bottlenose Dolphins (Tursiops truncatus) that died during the 1987-1988 Atlantic-dolphin morbillivirus epizootic. The lesions were characterized by focally extensive suppurative and histiocytic dermatitis and cellulitis with ulceration and variable numbers of dermal and hypodermal ciliates. Vasculitis, thrombosis, and/or intravascular ciliates were rarely present. In one dolphin, there was an associated lymphadenitis with ciliates, and in another, bronchopneumonia with rare intrabronchiolar ciliates. Ten of the dolphins were female, and eight were male. The animals ranged in length from 148 to 260 cm. Eleven were from Virginia, four were from New Jersey, and three were from Florida. In 13 dolphins, results of immunohistochemical and/or polymerase chain reaction (PCR) tests were positive for morbillivirus infection. Results of immunohistochemical tests were negative in four dolphins that were not also tested with PCR. Results were also negative in one dolphin tested using both methods. Nine dolphins had concomitant bacterial, fungal, and/or other protozoal infections. Fourteen other dolphins with ciliate-associated dermatitis were identified from 414 Atlantic bottlenose dolphin cases (3%) archived at the Armed Forces Institute of Pathology. The incidence of dermatitis with invasive ciliates is much greater in dolphins that died during the 1987-1988 epizootic.     

Escherichia coli septicemia associated with lack of maternally acquired immunity in a bottlenose dolphin calf

Stillbirth and neonatal mortality are substantial problems in captive bottlenose dolphins (Tursiops truncatus). The cause of these problems often is unknown. We report a case of Escherichia coli septicemia in a male 3-day-old bottlenose dolphin calf. Lesions included omphalitis, synovitis, and hepatic necrosis associated with the presence of Gram-negative bacilli. E. coli was isolated in pure culture from multiple organs. A serum gammaglobulin level of 1.5 g/L indicated a lack of maternally acquired immunity. The observed failure to nurse may have resulted from brain injury due to perinatal asphyxia. Evidence for perinatal asphyxia was the diffuse presence of a moderate amount of meconium in the lungs.