Gregory D. Bossart

Initial evaluation of the effects of aerosolized Florida red tide toxins (brevetoxins) in persons with asthma

Florida red tides annually occur in the Gulf of Mexico, resulting from blooms of the marine dinoflagellate Karenia brevis. K. brevis produces highly potent natural polyether toxins, known as brevetoxins, that activate voltage-sensitive sodium channels. In experimental animals, brevetoxins cause significant bronchoconstriction. A study of persons who visited the beach recreationally found a significant increase in self-reported respiratory symptoms after exposure to aerosolized Florida red tides. Anecdotal reports indicate that persons with underlying respiratory diseases may be particularly susceptible to adverse health effects from these aerosolized toxins. Fifty-nine persons with physician-diagnosed asthma were evaluated for 1 hr before and after going to the beach on days with and without Florida red tide. Study participants were evaluated with a brief symptom questionnaire, nose and throat swabs, and spirometry approved by the National Institute for Occupational Safety and Health. Environmental monitoring, water and air sampling (i.e., K. brevis, brevetoxins, and particulate size distribution), and personal monitoring (for toxins) were performed. Brevetoxin concentrations were measured by liquid chromatography mass spectrometry, high-performance liquid chromatography, and a newly developed brevetoxin enzyme-linked immunosorbent assay. Participants were significantly more likely to report respiratory symptoms after Florida red tide exposure. Participants demonstrated small but statistically significant decreases in forced expiratory volume in 1 sec, forced expiratory flow between 25 and 75%, and peak expiratory flow after exposure, particularly those regularly using asthma medications. Similar evaluation during nonexposure periods did not significantly differ. This is the first study to show objectively measurable adverse health effects from exposure to aerosolized Florida red tide toxins in persons with asthma. Future studies will examine the possible chronic effects of these toxins among persons with asthma and other chronic respiratory impairment.

Protein electrophoresis as a diagnostic and prognostic tool in raptor medicine

Abstract Plasma proteins of 139 healthy adult birds of prey from 10 species were separated by electrophoresis to characterize and document normal reference ranges and species-specific electrophoretic patterns and to evaluate the value of this technique for health screening, disease diagnosis, and prognostic indication. Species studied included bald eagle (Haliaeetus leucocephalus), red-tailed hawk (Buteo jamaicensis), barn owl (Tyto alba), great horned owl (Bubo virginianus), turkey vulture (Cathartes aura), Harris hawk (Parabuteo unicinctus), Stellars sea eagle (Haliaeetus pelagicus), barred owl (Strix varia), screech owl (Otus asio), and black vulture (Coragyps atratus). Several clinical cases show the diagnostic/therapeutic value of protein electrophoresis in raptors. This study establishes species-specific reference ranges for several birds of prey and discusses the benefit of electrophoresis as a diagnostic technique in health screens, as a diagnostic aid in conjunction with other tests, and as a prognostic indicator in clinical evaluation of raptors.

Antibiotic-Resistant Organisms Cultured from Atlantic Bottlenose Dolphins (Tursiops truncatus) Inhabiting Estuarine Waters of Charleston, SC and Indian River Lagoon, FL

Bottlenose dolphins (Tursiops truncatus) from estuarine waters of Indian River Lagoon, FL (IRL) and Charleston, SC (CHS) were cultured to screen for microorganism colonization and to assess antibiotic sensitivity. Swabs (nxa0=xa0909) were collected from the blowhole, gastric fluid, and feces of 171 individual dolphins The most frequently cultured organisms were Plesiomonas shigelloides (nxa0=xa0161), Aeromonas hydrophila (nxa0=xa0144), Escherichia coli (nxa0=xa085), and Pseudomonas fluorescens (nxa0=xa082). In descending frequency, organisms demonstrated resistance to erythromycin, ampicillin, and cephalothin. Human and animal pathogens resistant to antibiotics used in human and veterinary medicine were cultured. Escherichia coli (E. coli) more often was resistant in IRL dolphins. Three cases of methicillin-resistant Staphylococcus aureus (MRSA) were found at CHS. Emergence of antibiotic resistance is not confined to humans. Bottlenose dolphins may serve as sentinels for transfer of resistance from humans and animals or indicate that antibiotics are reaching the marine environment and causing resistance to emerge through selective pressure and genetic adaptation.

ALTERED IN VITRO IMMUNE RESPONSES IN GREEN TURTLES (CHELONIA MYDAS) WITH FIBROPAPILLOMATOSIS

Abstract The immune competence of green sea turtles (Chelonia mydas) with fibropapillomatosis was assessed using in vitro techniques to measure lymphocyte proliferation in response to mitogens. In comparison with captive, healthy green sea turtles, those afflicted with fibropapillomas demonstrated diminished proliferation with Concanavalin A, phytohemagglutinin (T-cell mitogens), and lipopolysaccharide (B-cell mitogen). Also, markedly decreased proliferative responses to the lymphocyte polyclonal stimulator combination of ionomycin and phorbol myristate acetate were observed. Total circulating white blood cell counts were not statistically different between the two groups, although an overall decrease in lymphocyte number was observed in the papilloma group. The albumin/globulin ratio was decreased in the papilloma group because of decreased albumin and increased gamma globulins.

Lacaziosis in bottlenose dolphins Tursiops truncatus along the coastal Atlantic Ocean, Florida, USA

This study represents the first systematic study of lacaziosis (lobomycosis) in bottlenose dolphins Tursiops truncatus in the Atlantic Ocean along the east-central coast of Florida, USA. Lacaziosis is a chronic infection of the skin caused by the fungus Lacazia loboi, which affects only dolphins and humans. Previous studies have shown a high prevalence (6.8 to 12.0%) of lacaziosis in resident dolphins from the adjacent Indian River Lagoon Estuary (IRL), where the disease is endemic. We examined the prevalence of lacaziosis in this coastal area using photo-identification data collected between 2002 and 2008 to determine the prevalence of lacaziosis in coastal dolphins using photographic methodology shown to have high sensitivity and specificity in prior research. The prevalence of skin lesions compatible with lacaziosis estimated from photographic data was 2.1% (6/284), approximately 3 times lower than that described for the estuarine population using similar methods. To exclude potential bias introduced by differences in study duration and survey effort among areas, an 18 mo period when effort was most equal (January 2006 to June 2007) was chosen for statistical comparison. The prevalence of lacaziosis estimated from photographic data was significantly lower (3.8%: n = 6/160) in the Atlantic Ocean compared to the IRL (12.0%: n = 20/167) (risk ratio = 3.19, 95% CI 1.32 to 7.75, p < 0.01 by chi-square analysis). The lower prevalence of lacaziosis in dolphins found in the Atlantic Ocean and the overall lack of movement of dolphins between these habitats suggests that environmental conditions within the estuary may favor viability of L. loboi, and/or that immune compromise in resident estuarine dolphins is a precursor to the disease.

Cutaneous Granulomas in Dolphins Caused by Novel Uncultivated Paracoccidioides brasiliensis

Our findings could stimulate study of public health implications of diseases caused by this fungus.

Clinicoimmunopathologic findings in Atlantic bottlenose dolphins Tursiops truncatus with positive Chlamydiaceae antibody titers

Sera from free-ranging Atlantic bottlenose dolphins Tursiops truncatus inhabiting the Indian River Lagoon (IRL), Florida, and coastal waters of Charleston (CHS), South Carolina, USA, were tested for antibodies to Chlamydiaceae as part of a multidisciplinary study of individual and population health. A suite of clinicoimmunopathologic variables was evaluated in Chlamydiaceae-seropositive dolphins (n = 43) and seronegative healthy dolphins (n = 83). Fibrinogen, lactate dehydrogenase, amylase, and absolute numbers of neutrophils, lymphocytes, and basophils were significantly higher, and serum bicarbonate, total alpha globulin, and alpha-2 globulin were significantly lower in dolphins with positive Chlamydiaceae titers compared with seronegative healthy dolphins. Several differences in markers of innate and adaptive immunity were also found. Concanavalin A-induced T lymphocyte proliferation, lipopolysaccharide-induced B lymphocyte proliferation, and granulocytic phagocytosis were significantly lower, and absolute numbers of mature CD 21 B lymphocytes, natural killer cell activity and lysozyme concentration were significantly higher in dolphins with positive Chlamydiaceae antibody titers compared to seronegative healthy dolphins. Additionally, dolphins with positive Chlamydiaceae antibody titers had significant increases in ELISA antibody titers to Erysipelothrix rhusiopathiae. These data suggest that Chlamydiaceae infection may produce subclinical clinicoimmunopathologic perturbations that impact health. Any potential subclinical health impacts are important for the IRL and CHS dolphin populations, as past studies have indicated that both dolphin populations are affected by other complex infectious and neoplastic diseases, often associated with immunologic perturbations and anthropogenic contaminants.

Asphyxiation in a Bottlenose Dolphin ( Tursiops truncatus ) from Puerto Rico Due to Choking on a Black Margate ( Anisotremus surinamensis )

Bottlenose dolphins (Tursiops truncatus) are found in the coastal and offshore waters of Puerto Rico. However, little is known about causes of their mortality in the Caribbean. On 18 February 2002, a female bottlenose dolphin was found dead in Bahía de San Juan, Puerto Rico. Remarkably, a black margate (Anisotremus surinamensis) was firmly lodged in the dolphin’s oral cavity and the pharynx. The throat of the dolphin was markedly swollen; the larynx was dislocated; and signs of agonal death were evident. Grossly, the cause of death was determined to be asphyxiation due to choking. Fifty strandings of bottlenose dolphins have been reported between 1937 and 2006 in Puerto Rico and the Virgin Islands. In those for which a cause of death was determined, four were human related and 11 died of natural causes (including this case). The present case study fits the definitions of “choking” and “asphyxiation.” The evolved respiratory anatomy of cetaceans in which the larynx is inserted into the nasal passages leading to the blowhole makes asphyxiation due to choking unlikely in odontocetes. However, if the larynx is irreversibly dislodged from its normal position during swallowing, this may cause the dolphin to stop breathing or even drown. Thus, respiratory blockage from a natural prey item represents an uncommon mortality factor associated with piscivorous cetaceans and is the first record of such marine mammal mortality in the Caribbean.

The environment as a driver of immune and endocrine responses in dolphins (Tursiops truncatus)

Immune and endocrine responses play a critical role in allowing animals to adjust to environmental perturbations. We measured immune and endocrine related markers in multiple samples from individuals from two managed-care care dolphin groups (n = 82 samples from 17 dolphins and single samples collected from two wild dolphin populations: Indian River Lagoon, (IRL) FL (n = 26); and Charleston, (CHS) SC (n = 19). The immune systems of wild dolphins were more upregulated than those of managed-care-dolphins as shown by higher concentrations of IgG and increases in lysozyme, NK cell function, pathogen antibody titers and leukocyte cytokine transcript levels. Collectively, managed-care care dolphins had significantly lower levels of transcripts encoding pro-inflammatory cytokine TNF, anti-viral MX1 and INFα and regulatory IL-10. IL-2Rα and CD69, markers of lymphocyte activation, were both lower in managed-care care dolphins. IL-4, a cytokine associated with TH2 activity, was lower in managed-care care dolphins compared to the free-ranging dolphins. Differences in immune parameters appear to reflect the environmental conditions under which these four dolphin populations live which vary widely in temperature, nutrition, veterinary care, pathogen/contaminant exposures, etc. Many of the differences found were consistent with reduced pathogenic antigenic stimulation in managed-care care dolphins compared to wild dolphins. Managed-care care dolphins had relatively low TH2 lymphocyte activity and fewer circulating eosinophils compared to wild dolphins. Both of these immunologic parameters are associated with exposure to helminth parasites which is uncommon in managed-care care dolphins. Less consistent trends were observed in a suite of hormones but significant differences were found for cortisol, ACTH, total T4, free T3, and epinephrine. While the underlying mechanisms are likely multiple and complex, the marked differences observed in the immune and endocrine systems of wild and managed-care care dolphins appear to be shaped by their environment.

Mercury concentrations in Atlantic bottlenose dolphins (Tursiops truncatus) inhabiting the Indian River Lagoon, Florida: Patterns of spatial and temporal distribution

Bottlenose dolphins (Tursiops truncatus) inhabiting the Indian River Lagoon, FL (IRL) have tissue mercury concentrations among the highest reported worldwide. Analysis of total mercury (THg) concentrations in blood collected between 2003 and 2012 showed a significant linear decrease over time (p=0.04). Significant differences in the spatial distribution of THg in resident IRL dolphins were also observed with a general gradient in concentration from north to south. Evaluation of local biogeochemistry and accumulation of mercury in prey species is needed to better understand factors influencing the distribution of Hg in the apex predator. Analyses of temporal and spatial patterns of exposure to THg in this sentinel species may have implications for both ecosystem and public health in the region.