Ascocotyle longa (Digenea: Heterophyidae) infecting dolphins from the Atlantic Ocean.

We report for the first time the infection of dolphins with Ascocotyle longa found in the intestines of three different species, Sotalia guianensis, Steno bredanensis, and Tursiops truncatus gephyreus, which were found washed ashore along the southeastern and southern Brazilian coast. The worms were identified based on morphological and molecular data using the 28S rDNA gene and the COI gene. Specimens of A. longa from the pinniped Otaria flavescens were also analyzed. As the first isolation of A. longa from cetaceans, the present study increases the distribution area and range of definitive hosts of this trematode, and provides new molecular data to complement the phylogeny of the group in future studies, thus contributing to the scientific knowledge of this potentially zoonotic parasite.

Dolphin-derived NETosis results in rapid Toxoplasma gondii tachyzoite ensnarement and different phenotypes of NETs.

Toxoplasma gondii is a cosmopolitan zoonotic parasite and nowadays considered as an emerging neozoan pathogen in the marine environment. Cetacean innate immune reactions against T. gondii stages have not yet been investigated. Thus, T. gondii tachyzoites were utilized to trigger neutrophil extracellular traps (NETs) in bottlenose dolphin (Tursiops truncatus) polymorphonuclear neutrophils (PMN). Scanning electron microscopy unveiled T. gondii tachyzoites as potent and rapid inducers of cetacean-derived NETosis. Co-localization of extracellular chromatin with global histones, granulocytic myeloperoxidase and neutrophil elastase confirmed classical characteristics of NETosis. Interestingly, different phenotypes of NETs were induced by tachyzoites resulting in spread, diffuse and aggregated NET formation and moreover, 'anchored' and 'cell free' NETosis was also detected. Current data indicate that cetacean-derived NETosis might represent an early, ancient and well-conserved host innate defense mechanism that not only acts against T. gondii but might also occur in response to other closely related emerging apicomplexan parasites affecting marine cetaceans.

Skin marks in bottlenose dolphins (Tursiops truncatus) interacting with artisanal fishery in the central Mediterranean Sea.

Skin marks occur frequently in many cetacean species across the globe revealing a broad spectrum of causes, including social interactions, infectious diseases and injuries produced by anthropogenic factors. The current study used photo-id data from 2005-2014 to estimate the skin mark pattern on resident bottlenose dolphins (Tursiops truncatus) from the Aeolian Archipelago (Italy). Thirteen skin mark types were identified and their origin, prevalence and permanence time were examined. The pattern of skin marks was assessed for the abundance, richness, distribution and severity in six body regions and compared among age classes, sex and degree of dolphins' interaction with trammel nets (DIN). Our results showed higher prevalence, abundance, richness and distribution of skin marks in adults than in the younger age classes, with the exception of black marks and white ring lesions. The prevalence and abundance of skin marks were higher in males than females, with the exception of scratches and white patches. Moreover, gunshot wounds, mutilations and irregular dorsal fin edges were found only on adult males. Since males showed higher DIN than females and, in dolphins with higher DIN, skin marks were more abundant and frequently distributed in different body regions, the skin mark pattern in regard to DIN seems to be sex-related. The more severe marks were observed on adults, males and dolphins with higher DIN, namely skin disorder, tooth rake marks, small shallow indentations, deep indentations and mutilations. On the contrary, the severity of scratches, white patches and dark ring lesions was higher in females than males, but not significantly related to DIN and age of the individuals. Our results showed that photo-id data provide an efficient and cost-effective approach to document the occurrence of skin marks in free-ranging bottlenose dolphin populations, a critical step toward understanding the cause and supporting the conservation strategies.

Detection of Toxoplasma gondii in three common bottlenose dolphins (Tursiops truncatus); A first description from the Eastern Mediterranean Sea.

Toxoplasma gondii has been described in several marine mammals around the world including numerous species of cetaceans, yet infection and transmission mechanisms in the marine environment are not clearly defined. The Israel Marine Mammal Research and Assistance Center has been collating a database of all marine mammal stranding events along the country's national coastlines since 1993. In this study, we describe the molecular detection and characterisation of T. gondii in three common bottlenose dolphins (Tursiops truncatus) including one case of coinfection with herpesvirus. The animals were found stranded on the Mediterranean coast of Israel in May and November 2013. In one of the three cases, the dolphin was found alive and admitted to intensive care. To our knowledge, this is the first report of T. gondii infection of marine mammals in the Eastern Mediterranean Sea. As this parasite acts as an indicator for marine pollution and marine mammal health, we believe these findings add important information regarding the state of the environment in this region.

A new Synthesium species (Digenea: Brachycladiidae) from the bottlenose dolphin Tursiops truncatus (Cetacea: Delphinidae) in Southwestern Atlantic waters.

A new species of Synthesium from the bottlenose dolphin Tursiops truncatus in South Brazilian waters is described. Morphological and molecular identification was performed, and phylogenetic analyses were carried out using the ribosomal small subunit and internal transcribed spacer 1 and the mitochondrial NDH dehydrogenase subunit 3 and cytochrome c oxidase subunit 1 genes. The main characteristics of the new species are the subterminal round-shaped oral sucker, the anterior distribution of vitellaria reaching the level of the ovary and the oval-shaped testes. The results obtained with the molecular markers supported the inclusion of the specimens into the genus Synthesium. The nucleotide divergence detected for the mitochondrial genes among the new species and others of the same genus supported the erection of a new species. This is the ninth species assigned to the genus and the third Synthesium species recorded in the South Atlantic Ocean.

Acute fatal sarcocystosis hepatitis in an Indo-Pacific bottlenose dolphin (Tursiops aduncus) in Hong Kong.

Unlike most species in the genus Sarcocystis, Sarcocystis canis has a broad intermediate host range. Its life cycle is incompletely known and most reports are from the USA. Here we report fatal hepatitis in a 4year old male Indo-Pacific bottlenose dolphin (Tursiops aduncus) from Hong Kong associated with a S. canis-like infection. Diagnosis was made based on clinical presentation, histopathology, transmission electron microscopy (TEM), and molecular characterization. Microscopically, S. canis-like like infection was confined to the liver. Immature and mature schizonts were found in hepatocytes and the parasite was associated with generalized hepatic necrosis. By TEM, schizonts divided by endopolygeny, and merozoites lacked rhoptries. Molecular characterization of parasites present in liver and brain tissues at the cox1 gene showed a high degree of identity (97-98%) and clustered together with Sarcocystis canis, S. lutrae, S. arctica, S. speeri, S. turdusi, and S. rileyi in a phylogenetic study. This is the first report of S. canis-like infection from Asia.

Life-history trade-offs in a generalist digenean from cetaceans: the role of host specificity and environmental factors.

BACKGROUND: Adults and larvae of generalist parasites are exposed to diverse hosts and local environmental conditions throughout their life cycles, thus local adaptation is expected to occur through phenotypic plasticity and/or natural selection. We investigated how the combined effect of cryptic host specificity and local selective pressures could shape reproductive traits of a putative generalist parasite in the oceanic realm. METHODS: The LSU rDNA, ITS2 and the mt-COI of individuals of the digenean Pholeter gastrophilus (Kossack, 1910) Odhner, 1914 (Heterophyidae Leiper, 1909) from oceanic striped dolphins, Stenella coeruleoalba Meyen, and coastal bottlenose dolphins, Tursiops truncatus Montagu, in the western Mediterranean were used to elucidate whether worms were conspecific. Infection parameters were compared between both dolphin species. General Linear Mixed Models were used to analyse the influence of host species on four reproductive traits of P. gastrophilus: body size, maturity stage (non-gravid/gravid), egg size, and number of eggs in utero. AIC values were used to rank competing models, and p-values to assess the effect of specific predictors. RESULTS: Evidence indicated that worms collected from both dolphin species were conspecific. All worms collected were gravid and infection parameters did not differ between dolphin species. However, body size and egg size of individuals of P. gastrophilus were significantly larger in striped dolphins. The number of eggs in utero did not significantly differ between dolphin species but, for a given body size, worms in bottlenose dolphins harboured more eggs. A trade-off between egg size and egg number was found in worms from both dolphin species, with a higher slope in striped dolphins. CONCLUSIONS: Apparently, striped dolphin is a more suitable host for P. gastrophilus, but reproductive investment seems to be adapted to the habitat where the life-cycle develops. Worms from striped dolphins likely face the problem of finding intermediate hosts in the oceanic realm and apparently invest into offspring size to enhance the early survival of larvae and the potential to multiply asexually within the first intermediate host. The small-sized worms from bottlenose dolphins would be adapted to reproduce early because of higher adult mortality, generating smaller and numerous eggs in a coastal habitat where chances of transmission are presumably higher.

A systematic health assessment of indian ocean bottlenose (Tursiops aduncus) and indo-pacific humpback (Sousa plumbea) dolphins incidentally caught in shark nets off the KwaZulu-Natal Coast, South Africa.

Coastal dolphins are regarded as indicators of changes in coastal marine ecosystem health that could impact humans utilizing the marine environment for food or recreation. Necropsy and histology examinations were performed on 35 Indian Ocean bottlenose dolphins (Tursiops aduncus) and five Indo-Pacific humpback dolphins (Sousa plumbea) incidentally caught in shark nets off the KwaZulu-Natal coast, South Africa, between 2010 and 2012. Parasitic lesions included pneumonia (85%), abdominal and thoracic serositis (75%), gastroenteritis (70%), hepatitis (62%), and endometritis (42%). Parasitic species identified were Halocercus sp. (lung), Crassicauda sp. (skeletal muscle) and Xenobalanus globicipitis (skin). Additional findings included bronchiolar epithelial mineralisation (83%), splenic filamentous tags (45%), non-suppurative meningoencephalitis (39%), and myocardial fibrosis (26%). No immunohistochemically positive reaction was present in lesions suggestive of dolphin morbillivirus, Toxoplasma gondii and Brucella spp. The first confirmed cases of lobomycosis and sarcocystosis in South African dolphins were documented. Most lesions were mild, and all animals were considered to be in good nutritional condition, based on blubber thickness and muscle mass. Apparent temporal changes in parasitic disease prevalence may indicate a change in the host/parasite interface. This study provided valuable baseline information on conditions affecting coastal dolphin populations in South Africa and, to our knowledge, constitutes the first reported systematic health assessment in incidentally caught dolphins in the Southern Hemisphere. Further research on temporal disease trends as well as disease pathophysiology and anthropogenic factors affecting these populations is needed.

Gastrointestinal parasites of free-living Indo-Pacific bottlenose dolphins (Tursiops aduncus) in the Northern Red Sea, Egypt.

The present study represents the first report on the gastrointestinal parasite fauna infecting the free-living and alive Indo-Pacific bottlenose dolphins (Tursiops aduncus) inhabiting waters of the Red Sea at Hurghada, Egypt. A total of 94 individual faecal samples of the examined bottlenose dolphins were collected during several diving expeditions within their natural habitats. Using classical parasitological techniques, such as sodium acetate acetic acid formalin method, carbol fuchsin-stained faecal smears, coproantigen ELISA, PCR and macroscopical analyses, the study revealed infections with 21 different parasite species belonging to protozoans and metazoans with some of them bearing zoonotic and/or pathogenic potential. Four identified parasite species are potential zoonotic species (Giardia spp., Cryptosporidium spp., Diphyllobothrium spp., Ascaridida indet.); three of them are known to have high pathogenic potential for the examined dolphin species (Nasitrema attenuata, Zalophotrema spp. and Pholeter gastrophilus) and some appear to be directly associated with stranding events. In detail, the study indicates stages of ten protozoan species (Giardia spp., Sarcocystis spp., Isospora (like) spp., Cystoisospora (like) spp., Ciliata indet. I and II, Holotricha indet., Dinoflagellata indet., Hexamita (like) spp., Cryptosporidium spp.), seven trematode species (N. attenuata, Nasitrema spp. I and II, Zalophotrema curilensis, Zalophotrema spp., Pholeter gastrophilus, Trematoda indet.), one cestode species (Diphyllobothrium spp.), two nematode species (Ascaridida indet, Capillaria spp.) and one crustacean parasite (Cymothoidae indet.). Additionally, we molecularly identified adult worms of Anisakis typica in individual dolphin vomitus samples by molecular analyses. A. typica is a common parasite of various dolphin species of warmer temperate and tropical waters and has not been attributed as food-borne parasitic zoonoses so far. Overall, these parasitological findings include ten new host records for T. aduncus (i.e. in case of Giardia spp., Sarcocystis spp., Cryptosporidium spp., Nasitrema spp., Zalophotrema spp., Pholeter gastrophilus, A. typica, Capillaria spp., Diphyllobothrium spp. and Cymothoidae indet.). The present results may be used as a baseline for future monitoring studies targeting the impact of climate or other environmental changes on dolphin's health conditions and therefore contribute to the protection of these fascinating marine mammals.

Development of an indirect immunofluorescence technique for the diagnosis of toxoplasmosis in bottlenose dolphins.

The diagnosis of toxoplasmosis is often complicated by the lack of specific clinical symptoms or postmortem features, in humans and other animals. The only diagnostic test described so far for the serological diagnosis of Toxoplasma gondii in marine mammals is the modified agglutination test (Dubey et al., Am J Vet Res 48(8):1239-1243, 1987). The development of more sensible and specific immunological techniques requires specific antibodies, which are currently unavailable in the scientific market. Indirect immunofluorescence (IIF) is one of the most widely used methods for the diagnosis of toxoplasmosis in humans (Auer et al., Parasitol Res 12:965-970, 2000). In order to develop and apply this technique to the bottlenose dolphin (Tursiops truncatus), immunoglobulins were firstly purified using ion-exchange chromatography. The purified immunoglobulins were then injected in New Zealand rabbits in order to obtain polyclonal antibodies. These antisera were validated by the IIF technique, using as controls serum samples of dolphins infected by Toxoplasma. The results were visualized using antirabbit IgG labeled with fluorescein. This newly developed and specific serological assay was then tested with the dolphin collection of Loro Parque, Tenerife, Spain (group I), and L'Oceanogràfic of Valencia, Spain (group II). The obtained results in this study showed that none of the dolphins from group 1 were infected by T. gondii and two animals were positive in group 2. Furthermore, we conclude that this study has produced antibodies with high specificity against dolphin immunoglobulins and an IIF method which may be used as immunological diagnostic tools, especially for the serological diagnosis of toxoplasmosis.

Intestinal helminth fauna of bottlenose dolphin Tursiops truncatus and common dolphin Delphinus delphis from the western Mediterranean.

We report on the intestinal helminth fauna of 15 bottlenose dolphins Tursiops truncatus and 6 short-beaked common dolphins Delphinus delphis from the western Mediterranean. Eight helminth species were found in bottlenose dolphin, i.e., the digeneans Synthesium tursionis, Brachycladium atlanticum, and Pholeter gastrophilus, the nematode Anisakis sp., and the cestodes Tetrabothrius forsteri, Diphyllobothrium sp., Strobilocephalus triangularis, and tetraphyllidean plerocercoids. Brachycladium atlanticum, S. triangularis , and tetraphyllidean plerocercoids are new host records. No T. forsteri had previously been reported in Mediterranean bottlenose dolphins. Three species of helminths were recorded in the common dolphin, i.e., the digenean Synthesium delamurei (which was a new host record), and the cestodes T. forsteri and tetraphyllidean plerocercoids. The intestinal helminth communities of bottlenose and common dolphins are depauperate, similar to that of other cetacean species, but those from bottlenose dolphins harbored a higher number of helminth species. This study supports the notion that oceanic cetaceans, such as common dolphins, have a comparatively poorer helminth fauna than that of neritic species, such as bottlenose dolphins, because the likelihood of parasite recruitment is decreased.

Barnacle bonding: morphology of attachment of Xenobalanus globicipitis to its host Tursiops truncatus.

Xenobalanus globicipitis, a unique type of small pseudo-stalked barnacle occurs on the appendages of cetaceans, including the common bottlenose dolphin Tursiops truncatus. In this study, we examined attachment structures of X. globicipitis and modifications to the skin of T. truncatus in areas of attachment compared to skin nearby an attachment site. Barnacles and their six calcareous footplates were measured for their length and width. There was a positive correlation of barnacle width and length to footplate width and length. The thickness of the stratum corneum increased significantly in areas of attachment compared to skin nearby a footplate. The mitotic stratum germinativum at the base of the dermal papillae did not change significantly in areas of attachment compared to skin nearby a footplate. The stratum germinativum lining the lateral walls of the dermal papillae was significantly thicker in areas of skin nearby a footplate compared to in areas of attachment. Skin of T. truncatus nearby a footplate, displayed dermal papillae extending from the dermis and pointing roughly perpendicular to the epidermal stratum corneum. At sites of X. globicipitis attachment, the dermal papillae were forced to extend laterally, parallel to the stratum corneum, and the dermal papillae length to width ratio at an attachment site was significantly higher than on skin near an attachment site. Our results show that attachment of X. globicipitis through production of footplates organized into calcareous rings, leads to a thickened stratum corneum of the epidermis, a thinner lateral mitotic stratum germinativum and displaced structures of the upper dermis. These resulting modifications to the epidermis and dermis of the host may add to securing barnacle attachment to its host.

Serological evidence of Toxoplasma gondii infection in captive marine mammals in Mexico.

Toxoplasma gondii infection in marine mammals is important because they are considered as a sentinel for contamination of seas with T. gondii oocysts, and toxoplasmosis causes mortality in these animals, particularly sea otters. Serological evidence of T. gondii infection was determined in 75 captive marine mammals from four facilities in southern and central geographical regions in Mexico using the modified agglutination test (MAT). Antibodies (MAT, 1:25 or higher) to T. gondii were found in 55 (87.3%) of 63 Atlantic bottlenose dolphins (Tursiops truncatus truncatus), 3 of 3 Pacific bottlenose dolphins (Tursiops truncatus gillii), 2 of 4 California sea lions (Zalophus californianus), but not in 3 West Indian manatees (Trichechus manatus), and 2 Patagonian sea lions (Otaria flavescens). Seropositive marine mammals were found in all 4 (100%) facilities sampled. All marine mammals were healthy and there has not been any case of clinical toxoplasmosis in the facilities sampled for at least the last 15 years. The seroprevalence of T. gondii infection in marine mammals of the same species did not vary significantly with respect to sex and age. This is the first report on the detection of antibodies to T. gondii in marine mammals in Mexico.

Eighteen-year study of South Australian dolphins shows variation in lung nematodes by season, year, age class, and location.

Between 1990 and 2007, carcasses of opportunistically collected short-beaked common dolphins (Delphinus delphis; n=238), Indo-Pacific bottlenose dolphins (Tursiops aduncus; n=167), and common bottlenose dolphins (Tursiops truncatus; n=15) were examined for parasites and life history data. Three species of lung nematodes (Halocercus lagenorhynchi, Stenurus ovatus, Pharurus alatus) were identified in surface nodules, subsurface lesions, or airways. Nematode burdens were light to heavy and, in many cases, would have compromised the dolphins' health. The number of dolphins infected was related to species, year, season, age class, and geographic region. Nematodes were found in all three species but were more prevalent in short-beaked common dolphins (mean annual prevalence=26%) than in bottlenose dolphins (12%). There was a significant increase in prevalence of nematodes in short-beaked common dolphins in 2005-06 (63%) compared to 1990-2004 (14%), with a peak in April-June. More young short-beaked common dolphins were infected than subadults and adults and, during the unusual infection event, there were more dependent calves (<130 cm) than juveniles. There were also more infections in dependent bottlenose dolphin (Tursiops spp.) calves but no increase in overall prevalence was detected during 2005-06. Because neonates of both short-beaked common dolphins and bottlenose dolphins were infected, mother-to-calf transmission is suspected for these species in South Australia. Numbers of infections in short-beaked common dolphins were higher in Gulf St Vincent than elsewhere in South Australia, particularly in 2005-06. The cause of the unusual infection event in short-beaked common dolphins is unknown. We discuss the influence of dolphin diet, life history, and external factors.

Toxoplasmosis in captive dolphins (Tursiops truncatus) and walrus (Odobenus rosmarus).

Toxoplasma gondii infection in marine mammals is intriguing and indicative of contamination of the ocean environment and coastal waters with oocysts. Toxoplasma gondii infection was detected in captive marine mammals at a sea aquarium in Canada. Antibodies to T. gondii were found in all 7 bottlenose dolphins (Tursiops truncatus) tested. Two of these dolphins, as well as a walrus (Odobenus rosmarus) at the facility, died. Encephalitis and T. gondii tissue cysts were identified in histological sections of the brain of 1 dolphin (dolphin no. 1). Another dolphin (dolphin no. 2) had mild focal encephalitis without visible organisms, but viable T. gondii was isolated by bioassay in mice and cats from its brain and skeletal muscle; this strain was designated TgDoCA1. The PCR-RFLP typing using 11 markers (B1, SAG1, SAG2, SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1, and Apico) identified a Type II strain. The DNA sequencing of B1 and SAG1 alleles amplified from TgDoCA1 and directly from the brains of dolphin no. 1 and the walrus showed archetypal alleles consistent with infection by a Type II strain. No unique polymorphisms were detected. This is apparently the first report of isolation of T. gondii from a marine mammal in Canada.

Prevalence and pathology of lungworm infection in bottlenose dolphins Tursiops truncatus from southwest Florida.

Parasitism of the respiratory system is a relatively common finding in stranded cetaceans; however, no systematic investigations regarding the severity, distribution, and clinical consequences of these infections in bottlenose dolphins Tursiops truncatus have been conducted previously. The present study determined the prevalence of lungworm infections in dead stranded (n=22) and live bottlenose dolphins (n=44) from southwestern Florida, USA, during the period from 2003 to 2005. Dead stranded bottlenose dolphins were necropsied and lungs were examined visually, by palpation, and histologically for lesions consistent with verminous pneumonia. When present, nematodes were counted, measured, and identified to species based upon their morphology. Dolphin feces and blowhole swabs were collected and examined for nematode larvae. Lungworm prevalence was 77% in dead animals (n=22). The lesions in most cases were mild, chronic, and not the primary cause of death. Only 13% of dead animals examined had patent infections, with larvae present in blowhole and fecal cytology, and only 18% of animals had intact worms present at necropsy, with a geometric mean intensity of infection of 22.6 worms animal(-1). Intact worms were identified as either Halocercus lagenorhynchi or Skrjabinalius cryptocephalus. The highest prevalence of active infections was found in neonates and calves, including 1 stillborn calf. For free-ranging animals, all blowhole swabs (n=44) were negative, and fecal cytology (n=22) showed a 3% prevalence of patent infection. Findings from the present study support the theory that bottlenose dolphins can be infected transplacentally by lungworms. The impact that such infections may have on neonatal survival is unknown; however, these infections could increase neonatal mortality.

Redescription of Synthesium pontoporiae n. comb. with notes on S. tursionis and S. seymouri n. comb. (Digenea: Brachycladiidae Odhner, 1905).

Synthesium pontoporiae n. comb. is redescribed, together with Synthesium tursionis and Synthesium seymouri n. comb.; the parasites were obtained from stranded and accidentally caught cetaceans. The sucker ratio (ratio between widths of the oral and ventral suckers) in S. pontoporiae was 1:1.8-3.0 (mean 1:2.2); in S. tursionis was 1:0.8-1.2; and in S. seymouri was 1:0.5-0.7. Synthesium pontoporiae differed from its congeners by additional diagnostic characters, including: oval to lobed testes; small cirrus with pyriform proximal region and flexible, tubular distal region formed by evagination of ejaculatory duct; and vitellarium in small follicles extending from the level of the seminal vesicle to the posterior extremity of the body and not forming dendritic radial bunches. Data on the morphology of adult S. pontoporiae and S. tursionis were inferred from confocal laser microscopical observations.

Isolation of Toxoplasma gondii from bottlenose dolphins (Tursiops truncatus).

Toxoplasma gondii infection in marine mammals is intriguing and indicative of contamination of the ocean environment and coastal waters with oocysts. In previous serological surveys, >90% of bottlenose dolphins (Tursiops truncatus) from the coasts of Florida, South Carolina, and California had antibodies to T. gondii by the modified agglutination test (MAT). In the present study, attempts were made to isolate T. gondii from dead T. truncatus. During 2005, 2006, and 2007, serum or blood clot, and tissues (brain, heart, skeletal muscle) of 52 T. truncatus stranded on the coasts of South Carolina were tested for T. gondii. Antibodies to T. gondii (MAT 1:25 or higher) were found in 26 (53%) of 49 dolphins; serum was not available from 3 animals. Tissues (heart, muscle, and sometimes brain) of 32 dolphins (26 seropositive, 3 seronegative, and 3 without accompanying sera) were bioassayed for T. gondii in mice, or cats, or both. Tissues of the recipient mice were examined for T. gondii stages. Feces of recipient cats were examined for shedding of T. gondii oocysts, but none excreted oocysts. Toxoplasma gondii was isolated from hearts of the 3 dolphins (2 with MAT titers of 1:200, and 1 without accompanied serum) by bioassay in mice. Genotyping of these 3 T. gondii isolates (designated TgDoUs1-3) with the use of 10 PCR-RFLP markers (SAG1, SAG2, SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1, and Apico) revealed 2 genotypes. Two of the 3 isolates have Type II alleles at all loci and belong to the clonal Type II lineage. One isolate has a unique genotype. This is the first report of isolation of viable T. gondii from T. truncatus.

Examination of naturally exposed bottlenose dolphins (Tursiops truncatus) for Microsporidia, Cryptosporidium, and Giardia.

Bottlenose dolphins (Tursiops truncatus) captured in the estuarine waters off the coasts of South Carolina and Florida were examined for the presence of Microsporidia, Cryptosporidium sp., and Giardia sp. DNA extracted from feces or rectal swabs was amplified by polymerase chain reaction using parasite-specific small subunit ribosomal RNA gene primers. All positive specimens were subjected to gene sequence analysis. Of 83 dolphins, 17 were positive for Microsporidia. None was positive for Cryptosporidium or Giardia. Gene sequence data for each of the positive specimens were compared with data in GenBank. Fourteen specimens were found similar to, but not identical to, the microsporidian species Kabatana takedai, Tetramicra brevifilum, and Microgemma tinca, reported from fish, and possibly represent parasites of fish eaten by dolphins. Gene sequence data from 3 other specimens had approximately 87% similarity to Enterocytozoon bieneusi, a species known primarily to infect humans and a variety of terrestrial mammals, including livestock, companion animals, and wildlife. It is not clear if these specimens represent a species from a terrestrial source or a closely related species unique to dolphins. There were neither clinical signs nor age- or gender-related patterns apparent with the presence of these organisms.

Pulmonary vascular proliferation and lungworm (Stenurus ovatus) in a bottlenose dolphin (Tursiops turncatus).

A female adult bottlenose dolphin suddenly died at 17 days after the capture. Macroscopically, severe pulmonary congestive edema was found. Histopathology revealed many lungworms in the bronchioli and the worms were identified as Stenurus ovatus. Variously sized vessels proliferated around the lesioned bronchioli. Based on these findings, chronic bronchopneumonia due to the lungworm was diagnosed and vascular proliferation was similar to angiomatosis recently reported in Atlantic bottlenose dolphin.