Changes in epizoic crustacean infestations during cetacean die-offs: the mass mortality of Mediterranean striped dolphins Stenella coeruleoalba revisited

In the summer and autumn of 1990, a cetacean morbillivirus caused a massive epizootic mortality of striped dolphins Stenella coeruleoalba in the western Mediterranean. Previous circumstantial evidence suggested that the disease could also have increased host susceptibility to infestations with epizoic crustaceans. In this study we provide strong evidence supporting this hypothesis. We examined striped dolphins stranded along the Mediterranean central coast of Spain from 1981 to 2004 (n = 136), and recorded data on prevalence, intensity of infestation, size and reproductive status of 2 sessile crustacean species specific to cetaceans, the phoront cirriped Xenobalanus globicipitis and the mesoparasitic copepod Pennella balaenopterae. Compared with the pre-epizootic (n = 12) and post-epizootic (n = 62) dolphin samples, the following changes were noted in the dolphins stranded during the epizootic (n = 62): (1) the prevalence of both X. globicipitis and P. balaenopterae increased; (2) the intensity of X. globicipitis and P. balaenopterae infestations did not increase; indeed, it was even slightly lower than in the other periods, as was their degree of aggregation; (3) individuals of both species were smaller, and a higher proportion were non-gravid; (4) the 2 species tended to co-occur in the same dolphins, but their numbers did not co-vary. These patterns strongly suggest that, during the epizootic, there was a short-term increase in the probability of infestation of these 2 species because of the sudden rise in the population of susceptible hosts; the growth of the new recruits was limited by the early death of dolphins. The high susceptibility was likely related to the immunosuppressive effects of viral infection and the abnormally heavy loads of polychlorinated biphenyls found in sick dolphins; the level of inbreeding was also higher in dolphins from the 'epizootic' sample. Epizoic crustaceans could be suitable indicators of health in cetacean populations.     

Epidemiological features of the first Unusual Mortality Event linked to cetacean morbillivirus in the South Atlantic (Brazil, 2017–2018)

Since the 1980s, cetacean morbillivirus (CeMV) has caused mass mortality events worldwide. However, no epizootics had been recorded in the South Atlantic, until an unusual mortality event (UME) linked to Guiana dolphin cetacean morbillivirus (GD-CeMV) began in Ilha Grande Bay, southeastern Brazil, in November 2017. In a five-month period, the UME spread to neighboring Sepetiba Bay and accounted for the death of at least 277 Guiana dolphins (Sotalia guianensis). Prevalence of morbillivirus positive dolphins, as estimated from RT-PCR diagnostics, was 92.3% (24/26) in Ilha Grande Bay and 91.9% (57/62) in Sepetiba Bay. Females had higher mortality rates during the UME (1.5:1), in contrast with historical mortality data from both bays that showed a 2:1 male to female death ratio. Calf mortality rates also increased in both bays. These results suggest that females and calves were more vulnerable to morbilliviral infection. Herein, we discuss possible explanations for such sex-biased death pattern during the UME and their implication for the conservation of endangered Guiana dolphins. We also speculate about the origin and spread of morbillivirus in the South Atlantic Ocean.     

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

Sera from free-ranging Atlantic bottlenose dolphins Tursiops truncatus inhabiting the Indian River Lagoon (IRL), Florida were tested for antibodies to cetacean morbilliviruses from 2003 to 2007 as part of a multidisciplinary study of individual and population health. A suite of clinicoimmunopathologic variables were evaluated in morbillivirus-seropositive dolphins (n = 14) and seronegative healthy dolphins (n = 49). Several important differences were found. Serum alkaline phosphatase, creatine phosphokinase, chloride, albumin and albumin/globulin ratios were significantly lower in seropositive dolphins. Innate immunity appeared to be upregulated with significant increases in lysozyme concentration and marginally significant increases in monocytic phagocytosis. Adaptive immunity was also impacted in dolphins with positive morbillivirus antibody titers. Mitogen-induced T lymphocyte proliferation responses were significantly reduced in dolphins with positive morbillivirus antibody titers, and marginally significant decreases were found for absolute numbers of CD4+ lymphocytes. The findings suggest impairment of cell-mediated adaptive immunity, similar to the immunologic pattern reported with acute morbillivirus infection in other species. In contrast, dolphins with positive morbillivirus antibody titers appeared to have at least a partially upregulated humoral immune response with significantly higher levels of gamma globulins than healthy dolphins, which may represent an antibody response to morbillivirus infection or other pathogens. These data suggest that subclinical dolphin morbillivirus infection in IRL dolphins may produce clinicoimmunopathologic perturbations that impact overall health.     

Morbillivirus infection in live stranded, injured, trapped, and captive cetaceans in southeastern Queensland and northern New South Wales, Australia

We report serologic evidence of cetacean morbillivirus (CMV) infection in five of eight cetacean species found live stranded, injured, or trapped along the coast of southeastern Queensland and northern New South Wales, Australia between December 2005 and January 2011. Antibody to CMV was detected in 13 of 27 (48%) wild cetaceans sampled. Antibody prevalence was significantly higher in clinically diseased (69%) compared to nondiseased (18%) animals (P=0.018). There was high antibody prevalence (83%, n=6) in melon-headed whales (Peponocephala electra). Two of 13 (15%) captive cetaceans sampled between November 2005 and January 2011 had CMV antibodies and, as infection was unlikely to have occurred while in captivity, CMV infection appears to have been present in Australian wild cetaceans since at least 1985. These results indicate that morbillivirus infection is occurring without widespread cetacean mortality in this region. However, as the deaths of two immature Australian offshore bottlenose dolphins (Tursiops truncatus) were attributed to CMV infection, morbillivirus infection should be included in the differential diagnosis of disease in cetaceans in Australia. Captive cetacean populations may be prone to significant mortality as a result of CMV introduction, so strict quarantine procedures should be enforced when injured or stranded cetaceans are hospitalized and rehabilitated at Australian zoos and marine parks.     

Characterization of microsatellite loci in Tursiops aduncus

We describe the cloning and characterization of five highly polymorphic microsatellite loci cloned from aduncus dolphins (Tursiops aduncus) from Western Australia. Five polymorphic microsatellite loci were isolated and tested on up to 350 animals, showing 7–23 alleles and expected heterozygosity values from 0.68 to 0.89. We also tested the loci on striped dolphins and franciscana dolphins, where we also found high levels of polymorphism (9–16 alleles in 102 striped, and 4–5 alleles in 13 franciscana dolphins). Considering that the cetacean genome is highly conserved, the characterized markers are likely to be useful in a number of cetacean population studies.     

Post-epizootic chronic dolphin morbillivirus infection in Mediterranean striped dolphins Stenella coeruleoalba

Dolphin morbillivirus (DMV) has caused 2 epizootics with high mortality rates on the Spanish Mediterranean coast, in 1990 and 2006-07, mainly affecting striped dolphins Stenella coeruleoalba. Following the first epizootic unusual DMV infections affecting only the central nervous system of striped dolphins were found, with histological features similar to subacute sclerosing panencephalitis and old dog encephalitis, the chronic latent localised infections caused by defective forms of measles virus and canine distemper virus, respectively. Between 2008 and 2010, monitoring by microscopic and immunohistochemical (IHC) studies of 118 striped dolphins stranded along Catalonia, the Valencia Region and Andalusia showed similar localised DMV nervous system infections in 25.0, 28.6 and 27.4% of cases, respectively, with no significant differences among regions or sex. The body length of DMV-infected dolphins was statistically greater than that of non-infected dolphins (196.5 vs. 160.5 cm; p < 0.001). Molecular detection of DMV was performed by 2 different RT-PCR techniques amplifying a 429 bp fragment and a 78 bp fragment both within the phosphoprotein (P) gene. The 429 bp RT-PCR results contradicted the IHC-DMV results as only 3 of 6 dolphins with positive IHC-DMV had positive PCR results. All 6 cases were positive with the 78 bp RT-PCR. These findings contraindicate the use of the 429 bp RT-PCR protocol based on the P gene to detect this specific form of DMV. DMV localised nervous infection constitutes the most relevant single cause of stranding and death in Mediterranean striped dolphins in the years following a DMV epizootic, and it might even overwhelm the effects of the epizootic itself, at least in 2007.     

Evidence of susceptibility to morbillivirus infection in cetaceans from the United States

Cetacean morbilliviruses (CeMV) are viruses that can cause mass mortalities among various odontocete species. In this study levels of “herd” immunity in cetaceans from the U.S. coast are described from the distribution and prevalence of antibodies against morbilliviruses. Neutralizing antibody titers against dolphin morbillivirus (DMV), porpoise morbillivirus (PMV), phocine distemper (PDV), and canine distemper viruses (CDV) were measured. Positive samples had higher titers against the CeMV than against the other morbilliviruses tested, indicating that although PDV or CDV can be used to investigate exposure their use may result in a higher false negative rate. The results suggest that morbillivirus did not persist in coastal populations of bottlenose dolphins (Tursiops truncatus) after the major outbreaks that occurred in the 1980s and 1990s. Bottlenose dolphins from Beaufort, North Carolina; St. Joseph Bay, Florida; and Cape May, New Jersey had anti‐DMV seroprevalences ranging from between 15% and 33% but those from Charleston, South Carolina and Sarasota Bay, Florida, sampled in recent years were largely negative. These latter groups are therefore now vulnerable to infection and could experience high mortality if exposed to CeMV. Sero‐surveys of this kind are therefore vital for assessing the risk of new and recurring viral outbreaks in coastal cetaceans.     

Predicting receptor functionality of signaling lymphocyte activation molecule for measles virus hemagglutinin by docking simulation

Predicting susceptibility of various species to a virus assists assessment of risk of interspecies transmission. Evaluation of receptor functionality may be useful in screening for susceptibility. In this study, docking simulation was conducted for measles virus hemagglutinin (MV‐H) and immunoglobulin‐like variable domain of signaling lymphocyte activation molecule (SLAM‐V). It was observed that the docking scores for MV‐H and SLAM‐V correlated with the activity of SLAM as an MV receptor. These results suggest that the receptor functionality may be predicted from the docking scores of virion surface proteins and cellular receptor molecules.     

Nearby clusters of hemagglutinin residues sustain SLAM-dependent canine distemper virus entry in peripheral blood mononuclear cells

Signaling lymphocytic activation molecule (SLAM, CD150) is the universal morbillivirus receptor. Based on the identification of measles virus (MV) hemagglutinin (H) amino acids supporting human SLAM-dependent cell entry, we mutated canine distemper virus (CDV) H and identified residues necessary for efficient canine SLAM-dependent membrane fusion. These residues are located in two nearby clusters in a new CDV H structural model. To completely abolish SLAM-dependent fusion, combinations of mutations were necessary. We rescued a SLAM-blind recombinant CDV with six mutations that did not infect ferret peripheral blood mononuclear cells while retaining full infectivity in epithelial cells.     

CETACEAN ABUNDANCE IN HAWAIIAN WATERS ESTIMATED FROM A SUMMER/FALL SURVEY IN 2002

Cetacean abundance is estimated for the U.S. Exclusive Economic Zone (EEZ) around the Hawaiian Islands based on a ship line‐transect survey from August to November, 2002. Sighting detection functions are estimated from this and other NOAA research surveys from 1986 to 2002 using a new, multiple‐covariate approach. Twenty‐four species were seen on this survey, including two species (Fraser's dolphin [Lagenodelphis hosei] and sei whale [Balaenoptera borealis]) that had not been previously documented to occur in Hawaiian waters. The most abundant large whales are sperm whales (Physeter macrocephalus) and Bryde's whales (Balaenoptera edeni). The most abundant delphinids are pilot whales (Globicephala macrorhynchus), rough‐toothed dolphins (Steno bredanensis), Fraser's dolphins, spotted dolphins (Stenella attenuata), and striped dolphins (Stenella coeruleoalba). Dwarf and pygmy sperm whales (Kogia sima and Kogia breviceps) and Cuvier's beaked whales (Ziphius cavirostris) are also estimated to be quite abundant. Some of the migratory baleen whales (fin whales [Balaenoptera physalus], sei whales, minke whales [B. acutorostrata], and humpback whales [Megaptera novaeangliae]) were seen only late in the survey. Abundance is estimated for 19 cetacean species. The overall density of cetaceans is low in the study area, especially for delphinids. The precision of density and abundance estimates is generally low for all species because of the low number of sightings.     

Striped dolphins and short‐beaked common dolphins in the Gulf of Corinth,Greece: Abundance estimates from dorsal fin photographs

Capture‐recapture methods relying on dorsal fin natural markings have never been applied successfully to striped dolphins, Stenella coeruleoalba, and were rarely used to assess abundance of short‐beaked common dolphins, Delphinus delphis. We used digital photo‐identification to obtain abundance estimates of striped and common dolphins living in mixed groups in the Gulf of Corinth, Greece. The proportion of either species was calculated based on the relative number of photographs of adult animals showing relevant portions of their body during conspicuous surfacings. Striped dolphins and common dolphins averaged 95.0% and 3.2% of all individuals, respectively. Animals showing intermediate pigmentation accounted for another 1.8%. Striped dolphin numbers were relatively high, with a point estimate of 835 animals (95% CI = 631–1,106). Common dolphins numbers were low (point estimate 28 animals; 95% CI = 11–73) and individuals were scattered within striped dolphin groups, indicating that this common dolphin population may be nonviable. Within a semiclosed Gulf exposed to considerable anthropogenic impact, the future of both dolphin species is of concern due to their suspected geographic isolation and restricted extent of occurrence. Information provided here can be used to inform timely conservation efforts.     

A review of cetacean occurrence in West African waters from the Gulf of Guinea to Angola

• 1 The cetacean fauna of the west coast of Africa is poorly described. Therefore, literature on the occurrence of cetacean species in the waters of 13 potential West African range states from the Gulf of Guinea to Angola was reviewed, including sighting, stranding, capture, bycatch and whaling records.
• 2 At least 28 species of cetacean were documented in the study region, comprising seven baleen whale species and 21 species of toothed whale (including at least 17 delphinid species).
• 3 Cetaceans could be broadly split into seven ecological categories, based on their distribution. A warm temperate/tropical deep‐water cetacean community dominated the study area. Cooler water from the Benguela Current influenced southern Angola (≤16°S latitude) and at least three cetacean species occurred predominantly in this region.
• 4 Only three or fewer species were confirmed in the waters of Togo, Nigeria, Cameroon and the Democratic Republic of Congo. Seventeen or more species were documented in Ghana, Gabon and Angola, where dedicated cetacean research projects have been initiated in recent years. Angola had the most diverse documented cetacean community: 28 confirmed species.
• 5 The humpback whale Megaptera novaeangliae was the most widely recorded species, and was documented in 11 (85%) countries. Sperm whales Physeter macrocephalus, Bryde's whales Balaenoptera cf. brydei, bottlenose dolphins Tursiops truncatus and Atlantic spotted dolphins Stenella frontalis were recorded in over half of the countries.

     

Characteristics of Fishing Operations,Environment and Life History Contributing to Small Cetacean Bycatch in the Northeast Atlantic

Fisheries bycatch is a key threat to cetacean species globally. Managing the impact requires an understanding of the conditions under which animals are caught and the sections of the population affected. We used observer data collected on an albacore tuna gillnet fishery in the northeast Atlantic, to assess operational and environmental factors contributing to bycatch of common and striped dolphins, using generalised linear models and model averaging. Life history demographics of the captured animals were also investigated. In both species, young males dominated the catch. The age ratio of common dolphins was significantly different from that estimated for the population in the region, based on life tables (G = 17.1, d.f. = 2, p = 0.002). Skewed age and sex ratios may reflect varying vulnerability to capture, through differences in behaviour or segregation in populations. Adult females constituted the second largest portion of the bycatch for both species, with potential consequences for population sustainability. Depth was the most important parameter influencing bycatch of both species and reflected what is known about common and striped dolphin habitat use in the region as the probability of catching common dolphins decreased, and striped dolphins increased, with increasing depth. Striped dolphin capture was similarly influenced by the extent to which operations were conducted in daylight, with the probability of capture increasing with increased operations in the pre-sunset and post-sunrise period, potentially driven by increased ability of observers to record animals during daylight operations, or by diurnal movements increasing contact with the fishery. Effort, based on net length and soak time, had little influence on the probability of capturing either species. Our results illustrate the importance of assessing the demographic of the animals captured during observer programmes and, perhaps more importantly, suggest that effort restrictions alone may not be sufficient to eradicate bycatch in areas where driftnets and small cetaceans co-occur.     

Selectively receptor-blind measles viruses: Identification of residues necessary for SLAM- or CD46-induced fusion and their localization on a new hemagglutinin structural model

Measles virus (MV) enters cells either through the signaling lymphocyte activation molecule SLAM (CD150) expressed only in immune cells or through the ubiquitously expressed regulator of complement activation, CD46. To identify residues on the attachment protein hemagglutinin (H) essential for fusion support through either receptor, we devised a strategy based on analysis of morbillivirus H-protein sequences, iterative cycles of mutant protein production followed by receptor-based functional assays, and a novel MV H three-dimensional model. This model uses the Newcastle disease virus hemagglutinin-neuraminidase protein structure as a template. We identified seven amino acids important for SLAM- and nine for CD46 (Vero cell receptor)-induced fusion. The MV H three-dimensional model suggests (i) that SLAM- and CD46-relevant residues are located in contiguous areas in propeller beta-sheets 5 and 4, respectively; (ii) that two clusters of SLAM-relevant residues exist and that they are accessible for receptor contact; and (iii) that several CD46-relevant amino acids may be shielded from direct receptor contacts. It appears likely that certain residues support receptor-specific H-protein conformational changes. To verify the importance of the H residues identified with the cell-cell fusion assays for virus entry into cells, we transferred the relevant mutations into genomic MV cDNAs. Indeed, we were able to recover recombinant viruses, and we showed that these replicate selectively in cells expressing SLAM or CD46. Selectively receptor-blind viruses will be used to study MV pathogenesis and may have applications for the production of novel vaccines and therapeutics.     

Canine distemper virus epithelial cell infection is required for clinical disease but not for immunosuppression

To characterize the importance of infection of epithelial cells for morbillivirus pathogenesis, we took advantage of the severe disease caused by canine distemper virus (CDV) in ferrets. To obtain a CDV that was unable to enter epithelial cells but retained the ability to enter immune cells, we transferred to its attachment (H) protein two mutations shown to interfere with the interaction of measles virus H with its epithelial receptor, human nectin-4. As expected for an epithelial receptor (EpR)-blind CDV, this virus infected dog and ferret epithelial cells inefficiently and did not cause cell fusion or syncytium formation. On the other hand, the EpR-blind CDV replicated in cells expressing canine signaling lymphocyte activation molecule (SLAM), the morbillivirus immune cell receptor, with similar kinetics to those of wild-type CDV. While ferrets infected with wild-type CDV died within 12 days after infection, after developing severe rash and fever, animals infected with the EpR-blind virus showed no clinical signs of disease. Nevertheless, both viruses spread rapidly and efficiently in immune cells, causing similar levels of leukopenia and inhibition of lymphocyte proliferation activity, two indicators of morbillivirus immunosuppression. Infection was documented for airway epithelia of ferrets infected with wild-type CDV but not for those of animals infected with the EpR-blind virus, and only animals infected with wild-type CDV shed virus. Thus, epithelial cell infection is necessary for clinical disease and efficient virus shedding but not for immunosuppression.     

DISTRIBUTION and NUMBERS OF STRIPED DOLPHINS IN THE WESTERN MEDITERRANEAN SEA AFTER THE 1990 EPIZOOTIC OUTBREAK

A survey was conducted in the summer of 1991 in the western Mediterranean to map the distribution and to estimate the number of striped dolphins surviving the 1990 outbreak of the morbillivirus epizootic. The highest densities of striped dolphins were found in the Alboran Sea and the Ligurian Sea. Total numbers were estimated at 117,880 (SE: 38,962; 95% CI: 68,379–214,800). The mortality resulting from the epizootic is unknown but may have been severe because the mean size of dolphin schools observed during the epizootic outbreak was significantly smaller than that before the event. A return to larger school sizes after the 1990 outbreak is not evidence of a recovery but can be viewed as a regrouping of individuals into schools of a preferred size. Calves were observed in 24.6% of the schools. They were not seen in schools of ten or less but were present in about half the schools of 30 or more dolphins, a result which suggests segregation similar to that seen in populations of this species elsewhere in the world.     

Measles virus (MV) hemagglutinin: evidence that attachment sites for MV receptors SLAM and CD46 overlap on the globular head

Measles virus hemagglutinin (MVH) residues potentially responsible for attachment to the wild-type (wt) MV receptor SLAM (CD150) have been identified and localized on the MVH globular head by reference to a revised hypothetical structural model for MVH (www.pepscan.nl/downloads/measlesH.pdb). We show that the mutation of five charged MVH residues which are conserved among morbillivirus H proteins has major effects on both SLAM downregulation and SLAM-dependent fusion. In the three-dimensional surface representation of the structural model, three of these residues (D505, D507, and R533) align the rim on one side of the cavity on the top surface of the MVH globular head and form the basis of a single continuous site that overlaps with the 546-548-549 CD46 binding site. We show that the overlapping sites fall within the footprint of an anti-MVH monoclonal antibody that neutralizes both wt and laboratory-vaccine MV strains and whose epitope contains R533. Our study does not exclude the possibility that Y481 binds CD46 directly but suggests that the N481Y mutation of wt MVH could influence, at a distance, the conformation of the overlapping sites so that affinity to CD46 increases. The relevance of these results to present concepts of MV receptor usage is discussed, and an explanation is proposed as to why morbillivirus attachment proteins are H, whereas those from the other paramyxoviruses are HN (hemagglutinin-neuraminidase).     

Adrenal Gland and Lung Lesions in Gulf of Mexico Common Bottlenose Dolphins (Tursiops truncatus) Found Dead following the Deepwater Horizon Oil Spill

A northern Gulf of Mexico (GoM) cetacean unusual mortality event (UME) involving primarily bottlenose dolphins (Tursiops truncatus) in Louisiana, Mississippi, and Alabama began in February 2010 and continued into 2014. Overlapping in time and space with this UME was the Deepwater Horizon (DWH) oil spill, which was proposed as a contributing cause of adrenal disease, lung disease, and poor health in live dolphins examined during 2011 in Barataria Bay, Louisiana. To assess potential contributing factors and causes of deaths for stranded UME dolphins from June 2010 through December 2012, lung and adrenal gland tissues were histologically evaluated from 46 fresh dead non-perinatal carcasses that stranded in Louisiana (including 22 from Barataria Bay), Mississippi, and Alabama. UME dolphins were tested for evidence of biotoxicosis, morbillivirus infection, and brucellosis. Results were compared to up to 106 fresh dead stranded dolphins from outside the UME area or prior to the DWH spill. UME dolphins were more likely to have primary bacterial pneumonia (22% compared to 2% in non-UME dolphins, P = .003) and thin adrenal cortices (33% compared to 7% in non-UME dolphins, P = .003). In 70% of UME dolphins with primary bacterial pneumonia, the condition either caused or contributed significantly to death. Brucellosis and morbillivirus infections were detected in 7% and 11% of UME dolphins, respectively, and biotoxin levels were low or below the detection limit, indicating that these were not primary causes of the current UME. The rare, life-threatening, and chronic adrenal gland and lung diseases identified in stranded UME dolphins are consistent with exposure to petroleum compounds as seen in other mammals. Exposure of dolphins to elevated petroleum compounds present in coastal GoM waters during and after the DWH oil spill is proposed as a cause of adrenal and lung disease and as a contributor to increased dolphin deaths.     

PCR 扩增Sry 基因进行鲸类动物性别的鉴定

哺乳动物Y染色体短臂上的Sry 基因决定雄性发育方向。本研究参照哺乳动物Sry 基因保守区序列设计引物, 以非性别特异性的线粒体DNA 细胞色素b 基因作为阳性对照, 用PCR 扩增江豚、长喙真海豚等鲸类动物的Sry 基因片断并对其进行凝胶电泳分析来鉴定鲸类动物的性别。通过此方法对87 个已知性别鲸类动物标本的检验, 结果完全正确, 并进一步应用此方法成功地完成了另外33 个未知性别鲸类标本的性别鉴定。由此建立了一套简单、快速、可靠的鲸类动物的性别鉴定方法。