Phocine Distemper Research Articles

The development of a qualitative real-time RT-PCR assay for the detection of hepatitis C virus

Real-time polymerase chain reaction (PCR) represents a favourable option for the detection of hepatitis C virus (HCV). A real-time reverse transcriptase PCR (RT-PCR) assay was developed as a qualitative diagnostic screening method for the detection of HCV using the ABI PRISM 7500 Sequence Detection System. The primers and probe were designed to target the 5'-untranslated region of the hepatitis C viral genome. A second heterologous probe assay was developed for the detection of the haemagglutinin gene of phocine distemper virus (PDV) and was used as an internal control. A semi-automated HCV extraction method was also implemented using the ABI PRISM 6100 Nucleic Acid PrepStation. The HCV assay was optimised as a qualitative singleplex RT-PCR assay with parallel testing of the target and internal control. The assay results (n = 200) were compared to the COBAS AMPLICOR HCV Test v2.0 assay. The assay demonstrated a high rate of sensitivity (99%), specificity (100%) and an acceptable limit of detection (LOD) of 100 IU/ml. The development of a qualitative multiplex assay for the simultaneous detection of HCV and internal control indicates the same high rates of sensitivity and specificity. This sensitive real-time assay may prove to be a valuable method for the detection of HCV.

Age, geographical distribution and taphonomy of an unusual occurrence of mummified crabeater seals on James Ross Island, Antarctic Peninsula

AbstractAn unusually dense collection of some 150 dead crabeater seals (Family Phocidae), in various stages of decay, occurs in the Brandy Bay hinterland, north-western James Ross Island, northern Antarctic Peninsula. Throughout the past 100 years, the presence of shelf ice (no longer present today) and sea ice in Prince Gustav Channel, between James Ross Island and the Antarctic Peninsula, has prevented seals from readily accessing the western side of James Ross Island. However, open water pools, some over one kilometre in diameter, remain accessible throughout the winter months, allowing seals to haul out onto the ice. It is likely that some of these seals may become disorientated as they wander away from the pools and instead head toward Brandy Bay and onto low-lying and snow-covered Abernethy Flats, easily mistaken for sea ice in early winter, where they perish. The large number of variably-decayed animals present suggests that this has probably happened on numerous occasions. However, some of the dead seals also probably perished during a documented mass dying event of crabeater seals in Prince Gustav Channel caused by an unidentified epidemic, possibly phocine distemper virus (PDV), during the spring of 1955.

Phocine distemper virus: characterization of the morbillivirus causing the seal epizootic in northwestern Europe in 2002

In 2002, 14 years after the first devastating die-off in the northwestern European seal population in 1988, a second epizootic was caused by the re-emergence of phocine distemper virus (PDV) in this area. The nucleotide sequence of the surface hemagglutinin (H) gene from the PDV of 2002 and the deduced amino acid sequence were determined and aligned with those of PDV isolates from 1988. There was a 98–99% nucleotide and a 98% amino acid identity of the H protein between the PDV isolates. From the close relationship shown in this study, it was concluded that PDV seems to be a strictly seal-adapted virus with only minor genetic variability over a longer period. In 1988, a fatal phocine distemper epizootic in seals from northwestern European waters led to the death of nearly 23,000 animals [13]. After the recovery of the seal population in the following years, a second mass die-off caused by phocine distemper virus (PDV) occurred in 2002 with the loss of approximately 30,000 seals [13]. Both epizootics showed similarities regarding the clinical and pathological findings, but some distinct features regarding the time of disease outbreak and epidemiologic progress were noticed [13, 22]. Until today, the origin and geographic distribution of PDV in northwestern Europe remain unclear in many aspects. It has been hypothesized that migrating seal species from other regions may act as carriers for PDV in the European seal populations [7]. PDV is a member of the genus Morbillivirus, closely related to canine distemper virus (CDV). In contrast, two other morbilliviruses from marine mammals, i.e. the porpoise and the dolphin morbilliviruses, are more closely related to the rinderpest (RPV) and the peste des petits ruminants viruses, respectively [2]. The surface and core proteins of PDV causing the first epizootic in Europe have been characterized in detail [2, 6, 18]. Both surface proteins, the hemagglutinin (H) and fusion (F) proteins, play an important role in viral attachment to and fusion with the host cell and formation of syncytia [2]. Variation in these proteins may cause changes in virulence and host-cell tropism. Earlier studies revealed a high identity between the F proteins from different CDV and PDV isolates, whereas the H protein was more variable [18, 24]. In order to analyse whether modifications of virulence and host range had occurred between the PDV isolates from both epizootics, the nucleotide and deduced amino acid sequences of the H gene of a PDV isolate from 2002 (PDV/ 02) were determined and compared to sequences from previous isolates. O.C.T-embedded lung tissue samples from three seals that died during the seal distemper epidemic in 2002 at the German North Sea, were obtained for evaluation. The samples were shock-frozen in liquid nitrogen and stored at -80 C until use. For verification of the PDV infection, a set of universal morbillivirus primers based on a conserved sequence of the phosphoprotein P gene was used as G. Muller U. Kaim P. Wohlsein W. Baumgartner Department of Pathology, University of Veterinary Medicine Hannover, Bunteweg 17, 30559 Hannover, Germany

In search of virus carriers of the 1988 and 2002 phocine distemper virus outbreaks in European harbour seals

European harbour seal (Phoca vitulina) populations decreased substantially during the phocine distemper virus (PDV) outbreaks of 1988 and 2002. Different hypotheses have stated that various seals and terrestrial carnivore species might be the source of infection. To further analyse these hypotheses, grey (Halichoerus grypus) and ringed (Phoca hispida) seals, polar bears (Ursus maritimus) and minks (Mustela lutreola) were sampled from the North Sea and East Greenland coasts between 1988 and 2004 and investigated by RT-PCR using a panmorbillivirus primer pair. However, all samples were negative for morbillivirus nucleic acid.

Age- and sex-specific mortality patterns in an emerging wildlife epidemic: the phocine distemper in European harbour seals.

Analyses of the dynamics of diseases in wild populations typically assume all individuals to be identical. However, profound effects on the long-term impact on the host population can be expected if the disease has age and sex dependent dynamics. The Phocine Distemper Virus (PDV) caused two mass mortalities in European harbour seals in 1988 and in 2002. We show the mortality patterns were highly age specific on both occasions, where young of the year and adult (>4 yrs) animals suffered extremely high mortality, and sub-adult seals (1–3 yrs) of both sexes experienced low mortality. Consequently, genetic differences cannot have played a main role explaining why some seals survived and some did not in the study region, since parents had higher mortality levels than their progeny. Furthermore, there was a conspicuous absence of animals older than 14 years among the victims in 2002, which strongly indicates that the survivors from the previous disease outbreak in 1988 had acquired and maintained immunity to PDV. These specific mortality patterns imply that contact rates and susceptibility to the disease are strongly age and sex dependent variables, underlining the need for structured epidemic models for wildlife diseases. Detailed data can thus provide crucial information about a number of vital parameters such as functional herd immunity. One of many future challenges in understanding the epidemiology of the PDV and other wildlife diseases is to reveal how immune system responses differ among animals in different stages during their life cycle. The influence of such underlying mechanisms may also explain the limited evidence for abrupt disease thresholds in wild populations.

Pathological Findings in Harbour Seals ( Phoca vitulina ): 1996–2005

Between 1996 and 2005 the carcasses of 355 harbour seals originating from the coast of Schleswig-Holstein, Germany, were investigated for pathological changes. The animals were collected before (n=280) and after (n=75) the second phocine distemper virus (PDV) epizootic in 2002. The seals were either found dead or were killed due to severe illness. Necropsy was performed in each case, in addition to histopathological, immunohistochemical, microbiological and parasitological examinations. Throughout the period of study, the respiratory and alimentary tracts were the organ systems most consistently affected by pathological change. The most common cause of death was bronchopneumonia caused by parasitic and/or bacterial infection of the lung. Less frequently identified changes included: trauma, gastroenteritis, uterine torsion or dystocia, polyarthritis/polymyositis, intestinal torsion, septicaemia, dermatitis, and keratitis. The most frequent causes of bronchopneumonia, gastroenteritis, polyarthritis, dermatitis and septicaemia were infections with alpha/beta-haemolytic streptococci, Escherichia coli and Clostridium perfringens. A number of changes were more frequently identified after 2002. These included the presence of parasites in the lung, stomach and intestine; bronchopneumonia, gastritis, enteritis, septicaemia and perinatal death. The increased prevalence of these changes may have been related to the preceding PDV epidemic.

Parasites in harbour seals (Phoca vitulina) from the German Wadden Sea between two Phocine Distemper Virus epidemics

Parasites were collected from 107 harbour seals (Phoca vitulina) found on the coasts of Schleswig-Holstein, Germany, between 1997 and 2000. The prevalence of the parasites and their associated pathology were investigated. Eight species of parasites, primarily nematodes, were identified from the examined organs: two anisakid nematodes (Pseudoterranova decipiens (sensu lato), Contracaecum osculatum (sensu lato)) from the stomach, Otostrongylus circumlitus (Crenosomatidae) and Parafilaroides gymnurus (Filaroididae) from the respiratory tract, one filarioid nematode (Acanthocheilonema spirocauda) from the heart, two acanthocephalans, Corynosoma strumosum and C. semerme (Polymorphidae), from the intestine and an ectoparasite, Echinophthirius horridus (Anoplura, Insecta). Lungworm infection was the most prominent parasitological finding and secondary bacterial bronchopneumonia the most pathogenic lesion correlated with the parasites. Heavy nematode burdens in the respiratory tract were highly age-related and more frequent in young seals. A positive correlation was observed between high levels of pulmonary infection and severity of bronchopneumonia. The prevalence of lungworms in this study was higher than in seals that died during the 1988/1989 Phocine Distemper Virus epidemic, and the prevalence of acanthocephalans and heartworms had decreased compared to findings from the first die-off.

Antigenic characterization of phocine distemper virus causing mass mortality in 2002 and its relationship to other morbilliviruses

The antigenic relationship between the phocine distemper virus (PDV) strain causing the epidemic in 2002 and the PDV strain of 1988, canine distemper virus from two dogs and one marten, and one measles virus strain was investigated in vivo and in vitro using monospecific polyclonal and monoclonal antibodies directed against five different proteins of canine or phocine distemper virus (N, P, M, F, H). Epitopic mapping revealed no difference between the PDV strains causing the epidemics in 1988 or 2002. However, the use of these antibodies allowed discrimination between different morbilliviruses including a vaccine strain of canine distemper virus. The major differences among the investigated morbilliviruses were found in the H protein.

Involvement of morbilliviruses in the pathogenesis of demyelinating disease

Two members of the morbillivirus genus of the family Paramyxoviridae, canine distemper virus (CDV) and measles virus (MV), are well-known for their ability to cause a chronic demyelinating disease of the CNS in their natural hosts, dogs and humans, respectively. Both viruses have been studied for their potential involvement in the neuropathogenesis of the human demyelinating disease multiple sclerosis (MS). Recently, three new members of the morbillivirus genus, phocine distemper virus (PDV), porpoise morbillivirus (PMV) and dolphin morbillivirus (DMV), have been discovered. These viruses have also been shown to induce multifocal demyelinating disease in infected animals. This review focuses on morbillivirus-induced neuropathologies with emphasis on aetiopathogenesis of CNS demyelination. The possible involvement of a morbillivirus in the pathogenesis of multiple sclerosis is discussed.

Using sparse survey data to investigate the declining abundance of British harbour seals

AbstractThis study presents an analysis of changes in the regional abundance of harbour seals Phoca vitulina based on repeated aerial surveys of haulouts, and demonstrates the use of sparse data to deliver advice about population status and management. Generalized linear models with negative binomial errors were used to represent these overdispersed data. The shape parameter of the negative binomial distribution was directly estimated from the data where this was possible. Information from time‐series of counts where there were few gaps in the data was used to improve the estimation of this parameter in areas where fewer surveys had been carried out. The results show that the number of harbour seals in eastern England has not increased since the end of the 2002 phocine distemper epidemic. There is also evidence of a general decline in most of the large harbour seal colonies around Britain. The populations in the Inner Hebrides were an exception, with numbers appearing to be stable or increasing. Between 2001 and 2006, the population in Orkney and Shetland declined by 40% (95% confidence interval: 30–50%), indicating harbour seals in these areas experienced substantially increased mortality or very low recruitment over this period. The widespread declines, ranging from Shetland to The Wash, suggest that the causes may have been present over a large part of the North Sea.

Genetic diversity of Hungarian canine distemper virus strains

To achieve proper diagnosis of dogs based on acute clinical symptoms and poorly preserved field samples taken from animals that died due to canine distemper (CD), a new differential diagnostic test has been developed based on polymerase chain reaction (PCR). In this study, more than 150 samples collected from dogs showing respiratory, gastrointestinal and neurological signs suggesting canine distemper virus (CDV) infection were examined. The samples consisted of urine, blood and nasal swabs collected from clinically ill patients, sent to our laboratory by clinicians from various veterinary clinics throughout Hungary. Various organs collected during the necropsy of dogs with pathological changes that suggested CDV infection were also included. Three distinct PCRs were designed. For diagnostic purposes, a primer pair specific to a 409 bases-long segment within the conservative part of the large polymerase region (L) of the CDV genome was designed. Using this test, out of the 150 analyzed samples, 46 (30.66%) proved to be positive for CDV, indicating that CDV still represents a high risk to the canine population in Hungary. For the phylogenetical analysis, a primer pair that completely encompasses the hemagglutinin (H) gene of the CDV genome was designed. The amplicons of this region were sequenced in both directions using the appropriate primers. Our results indicate that several different CDV genotypes are currently present in Hungary. Nine of the analyzed Hungarian strains turned out to belong to the so-called Arctic group of CDVs, and were most closely related to non-European strains from North America, China and Greenland, as well as to the phocine distemper virus 2 (PDV-2) isolated from Baikal seals ( Phoca sibirica). One of the Hungarian strains showed high similarity to other European isolates from Denmark, Germany, Italy and Turkey, as well as to other isolates from geographically more distant regions, such as the USA. Three Hungarian strains seem to join a new cluster that is formed by only a couple of strains, one isolated from a mink in Denmark, and another from a dog in North America. Using a third set of primers, a restriction fragment length polymorphism (RFLP) assay has also been designed for the fast and reliable differentiation of the wild-type CDVs from the vaccine strains.

Does Human Proximity Affect Antibody Prevalence in Marine-Foraging River Otters (Lontra canadensis)?

The investigation of diseases of free-ranging river otters (Lontra canadensis) is a primary conservation priority for this species; however, very little is known about diseases of river otters that forage in marine environments. To identify and better understand pathogens that could be important to marine-foraging river otters, other wildlife species, domestic animals, and humans and to determine if proximity to human population could be a factor in disease exposure, serum samples from 55 free-ranging marine-foraging river otters were tested for antibodies to selected pathogens. Thirty-five animals were captured in Prince William Sound, Alaska (USA), an area of low human density, and 20 were captured in the San Juan Islands, Washington State (USA), an area characterized by higher human density. Of 40 river otters tested by indirect immunofluorescent antibody test, 17.5% were seropositive (titer > or =320) for Toxoplasma gondii. All positive animals came from Washington. Of 35 river otters tested for antibodies to Leptospira interrogans using the microscopic agglutination test, 10 of 20 (50%) from Washington were seropositive (titer > or =200). None of the 15 tested animals from Alaska were positive. Antibodies to Neospora caninum (n=40), Sarcocystis neurona (n=40), Brucella abortus (n=55), avian influenza (n=40), canine distemper virus (n=55), phocine distemper virus (n=55), dolphin morbillivirus (n=55), porpoise morbillivirus (n=55), and Aleutian disease parvovirus (n=46) were not detected. Identifying exposure to T. gondii and L. interrogans in otters from Washington State but not in otters from Alaska suggests that living proximal to higher human density and its associated agricultural activities, domestic animals, and rodent populations could enhance river otter exposure to these pathogens.

Relatedness of Streptococcus equi subsp. zooepidemicus strains isolated from harbour seals ( Phoca vitulina) and grey seals ( Halichoerus grypus) of various origins of the North Sea during 1988–2005

The present study was designed to identify 15 beta-hemolytic streptococci isolated during a period between 1988 and 2005 from nine harbour seals and six grey seals from various origins of the North Sea. All isolates were identified as Streptococcus equi subsp. zooepidemicus. The bacteria were additionally investigated for relatedness by restriction fragment length polymorphism analysis of PCR amplified 16S-23S rDNA intergenic spacer region and gene szp and by macrorestriction analysis of chromosomal DNA of the strains by pulsed field gel electrophoresis. The molecular analysis yielded identical or closely related patterns within the strains of the present study and with the S. equi subsp. zooepidemicus strains isolated from harbour seals of German North Sea which were investigated previously [Akineden, Ö., Hassan, A.A., Alber, J., El-Sayed, A., Estoepangestie, A.T.S., Lämmler, C., Weiss, R., Siebert, U., 2005. Phenotypic and genotypic properties of S. equi subsp. zooepidemicus isolated from harbor seals ( Phoca vitulina) from the German North Sea during the phocine distemper outbreak in 2002. Vet. Microbiol. 110, 147–152]. This indicates that this single or closely related bacterial clone existed during both phocine distemper virus epidemics in 1988 and 2002 and that a direct transmission of the strains has occurred between two seal species and between seal populations of far distant regions possibly with grey seals as a vector.

Continuum description of a contact infection spread in a SIR model

We consider the process of an epidemic spread in a population of individuals with low mobility within the SIR scheme. In a continuum limit such propagation mechanism is described by a non-linear reaction–diffusion equation with a diffusion coefficient being the function of the densities of susceptible. The traveling wave solution of the corresponding system of partial differential equations is obtained and analyzed. We show that the model allows for description of Kendall epidemic waves, and give the dependence of the infection wave’s shape on the parameters of the system. An explicit calculation is done for realistic values of parameters obtained from field epidemiological data for a phocine distemper virus infection among harbor seals in 1988.

Phocine distemper virus in the North and European Seas – Data and models, nature and nurture

Two outbreaks of phocine distemper have severely affected harbour seal ( Phoca vitulina) populations in European and UK waters. The first occurred in 1988 when the causative virus was identified as a new member of the genus morbillivirus. The second outbreak in 2002 was first detected on the same Danish Island of Anholt and involved similar populations and geographical locations. However, despite the obvious similarities between the epidemics, differences in viral transmission and case mortality were found. Harbour seals are highly susceptible to infection while sympatric grey seals ( Halichoerus grypus) are resistant but could be important asymptomatic carriers of the disease. Arctic phocid seals remain the most likely source of the virus and grey seals could be the link between these primary hosts and the harbour seal populations further south. Future epidemiological models should therefore consider including multiple host species. The future conservation and management of harbour seal populations vulnerable to PDV relies on the ability to accurately predict the long-term impact on population abundance and distribution. Although knowledge about the behaviour and pathogenesis of the virus has increased substantially and data on host movements and contact rates are accumulating, studies into the determinants of the host range have lagged behind. The development of more realistic epidemiological models should be combined with studies into the factors controlling species and individual susceptibility. Assessing the risk of infection to endangered but currently unexposed potential host species (such as the Hawaiian monk seal, Monachus schauinslandi) is essential for guiding potential conservation management options, such as vaccination

SEROLOGIC SURVEY FOR BRUCELLA SPP., PHOCID HERPESVIRUS-1, PHOCID HERPESVIRUS-2, AND PHOCINE DISTEMPER VIRUS IN HARBOR SEALS FROM ALASKA, 1976–1999

Harbor seals (Phoca vitulina richardsi) were captured in the coastal regions of Southeast Alaska, Gulf of Alaska, Prince William Sound (PWS), and Kodiak Island during 1976-1999. Blood was collected from 286 seals. Sera were tested for evidence of exposure to Brucella spp., phocid herpesvirus-1 (PhoHV-1), phocid herpesvirus-2 (PhHV-2), and phocine distemper virus (PDV). Antibody prevalence rates were 46% (46/100) for Brucella spp., 93% (225/243) for PhoHV-1, 0% (0/286) for PhHV-2, and 1% (2/160) for PDV. Antibody prevalence for Brucella spp. was directly related to host age. Antibody prevalence for PhoHV-1 was higher in PWS as compared to the other three regions. No evidence of mortality attributable to these four agents was observed during the course of this study. Based on the results of this survey, none of these agents is considered a significant mortality factor in harbor seals from the four regions of coastal Alaska included in the study.

The 1988 and 2002 phocine distemper virus epidemics in European harbour seals

We present new and revised data for the phocine distemper virus (PDV) epidemics that resulted in the deaths of more than 23 000 harbour seals Phoca vitulina in 1988 and 30,000 in 2002. On both occasions the epidemics started at the Danish island of Anholt in central Kattegat, and subsequently spread to adjacent colonies in a stepwise fashion. However, this pattern was not maintained throughout the epidemics and new centres of infection appeared far from infected populations on some occasions: in 1988 early positive cases were observed in the Irish Sea, and in 2002 the epidemic appeared in the Dutch Wadden Sea, 6 wk after the initiation of the outbreak at Anholt Island. Since the harbour seal is a rather sedentary species, such 'jumps' in the spread among colonies suggest that another vector species could have been involved. We discussed the role of sympatric species as disease vectors, and suggested that grey seal populations could act as reservoirs for PDV if infection rates in sympatric species are lower than in harbour seals. Alternatively, grey seals could act as subclinical infected carriers of the virus between Arctic and North Sea seal populations. Mixed colonies of grey and harbour seal colonies are found at all locations where the jumps occurred. It seems likely that grey seals, which show long-distance movements, contributed to the spread among regions. The harbour seal populations along the Norwegian coast and in the Baltic escaped both epidemics, which could be due either to genetic differences among harbour seal populations or to immunity. Catastrophic events such as repeated epidemics should be accounted for in future models and management strategies of wildlife populations.

Phocine Distemper Outbreak, the Netherlands, 2002

During the 2002 phocine distemper epidemic, 2,284 seals, primarily harbor seals (Phoca vitulina), were found stranded along the Dutch coast. Stranding pattern varied with age, sex, state of decomposition, wind, and location. Cumulative proportion of deaths (54%) was comparable to that in the first reported epidemic in 1988.

Acquired Immunity and Stochasticity in Epidemic Intervals Impede the Evolution of Host Disease Resistance

Disease can generate intense selection pressure on host populations, but here we show that acquired immunity in a population subject to repeated disease outbreaks can impede the evolution of genetic disease resistance by maintaining susceptible genotypes in the population. Interference between the life-history schedule of a species and periodicity of the disease has unintuitive effects on selection intensity, and stochasticity in outbreak period further reduces the rate of spread of disease-resistance alleles. A general age-structured population genetic model was developed and parameterized using empirical data for phocine distemper virus (PDV) epizootics in harbor seals. Scenarios with acquired immunity had lower levels of epizootic mortality compared with scenarios without acquired immunity for the first PDV outbreaks, but this pattern was reversed after about five disease cycles. Without acquired immunity, evolution of disease resistance was more rapid, and long-term population size variation is efficiently dampened. Acquired immunity has the potential to significantly influence rapid evolutionary dynamics of a host population in response to age-structured disease selection and to alter predicted selection intensities compared with epidemiological models that do not consider such feedback. This may have important implications for evolutionary population dynamics in a range of human, agricultural, and wildlife disease settings.

Phylogenetic Characterization of Canine Distemper Viruses Detected in Naturally Infected Dogs in North America

In 2004, six puppies and one adult dog from a total of four premises were subjected to necropsy evaluation. For five of the seven dogs, disease caused by canine distemper virus (CDV) infection was suspected based on clinical signs. In all of the dogs, a diagnosis of CDV infection was established by the presence of compatible gross and histologic lesions, immunohistochemical labeling for CDV antigen, and detection of CDV RNA by reverse transcription-PCR. To further characterize the CDV strains detected in the four cases, complete gene sequences were determined for the hemagglutinin (H) and fusion (F) protein genes, while partial gene sequencing was performed for the phosphoprotein gene. A total of 4,508 bases were sequenced for the CDV strains detected from each of the four cases. Two cases were found to have identical sequences except for 2 bases in the intergenic region of the F and H genes. Phylogenetic analysis strongly suggested an evolutionary relationship between sequences detected in these two cases and those of phocine distemper virus 2 and two other strains of CDV not previously detected in the continental United States. Clear phylogenetic relationships were not established for viruses detected in the two additional cases; however, one strain showed similarity to CDV strains detected in a panda from China. Importantly, the three CDV strains detected were demonstrated to be genetically distinct from known vaccine strains and strains previously reported in the continental United States.