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Retrospective differentiation of canine distemper virus and phocine distemper virus in phocids.

Formalin-fixed paraffin-embedded tissues from one Caspian seal (Phoca caspica), one harp seal (Phoca groenlandica), one hooded seal (Cystophora cristata), and one harbor seal (Phoca vitulina vitulina) were used to compare the utility of immunohistochemistry (IHC) versus that of a novel seminested reverse transcriptase polymerase chain reaction (RT-PCR) to detect and differentiate canine distemper virus (CDV) and phocine distemper virus (PDV). Four antibodies made against PDV were able to detect both viruses. Two antibodies made against cetacean morbillivirus (CMV) did not label antigens from either CDV or PDV. A third anti-CMV antibody inconsistently stained CDV antigens but did not label PDV antigens. The seminested RT-PCR was able to detect RNA of the phosphoprotein gene in all positive cases. Nucleotide sequence analyses of seminested RT-PCR products were used to differentiate CDV RNA from PDV RNA. From these data, it was determined that IHC using antibodies generated against PDV provided a rapid means of detection for both CDV and PDV antigens; however, differentiation between CDV and PDV was achieved only with the RT-PCR assay.

Animals↗

Biological properties of phocine distemper virus and canine distemper virus.

Morbilliviruses constitute a major threat to the health of animal and man. To date the Morbillivirus genus in the Paramyxoviridae family comprises five established members, namely canine distemper virus (CDV), phocine distemper virus (PDV), measles virus (MV), rinderpest virus (RPV), and peste-des-petits-ruminants virus (PPRV). In addition, morbillivirus candidates infecting aquatic mammals were recently discovered. The present review on the biology of morbilliviruses focuses on knowledge gained by our group in studies on PDV and CDV. The aims of these studies were: i) to investigate the biological properties of the recently recognized PDV, which was found to be the primary etiology of epidemics with high mortality in seals in Western Europe, ii) to extend our knowledge of the biological properties of CDV. The morbillivirus particle is enveloped. The helical nucleocapsid core contains a single-stranded, non-segmented RNA genome of negative sense of 15 to 16 kilobases in length. The genome is organized in six transcriptional units or genes. Overall, the studies of the genome of PDV revealed a genetic map principally fitting with that determined for other morbilliviruses. The nucleotide and deduced amino acid sequences have been determined for five PDV genes named in analogy with the encoded structural proteins of other morbilliviruses in the order: 3'N(1683)-P(1644)-M(1443)-F(2206)-H(1952)-L5' (The figures in brackets denote nucleotide lengths of the genes of the Danish PDV isolate). The L gene (covering approximately 8900 nucleotides) remains to be sequenced. The six genes are likely to code for at least eight distinct proteins. The nucleocapsid (N) protein was found to consist of 523 amino acids in PDV. The following gene of the transcription map encoded the P protein of 507 amino acid residues. Similar to other morbilliviruses, the P gene of PDV was shown to have additional coding capacity for two distinct proteins V (299 amino acids) and C (174 amino acids). The results presented provide evidence for editing at transcript of the PDV P gene by insertion of nontemplated G residues at a specific site. The edited version of the mRNA was found to encode the cystein-rich V protein. The three envelope-associated proteins of PDV were predicted to consist of 335 (M), 537 (F0) and 607 (H) amino acid residues. The nucleotide and deduced amino acid sequences of the N, P, M, F, and H genes of PDV were aligned with corresponding sequences of other established members of the genus Morbillivirus.(ABSTRACT TRUNCATED AT 250 WORDS)

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Antibodies to canine distemper and phocine distemper viruses in polar bears from the Canadian arctic.

Serum samples collected from 200 polar bears (Ursus marititnus) from two populations in the Canadian arctic, the western Hudson Bay and Lancaster Sound populations, between 1989 and 1996, were tested for antibodies to canine distemper (CDV) and phocine distemper viruses (PDV) using virus neutralization. Antibodies to CDV and PDV were detected in 48 and six polar bears, respectively. All six bears that tested positive for PDV also tested positive for CDV; in only one case did the antibody titer for PDV exceed that of CDV. Differences in antibody prevalence to CDV were detected between populations and age classes but not sex or year of sampling.

Age Factors↗

Detection of phocine distemper virus using the polymerase chain reaction.

During the fatal seal epizootics in the North and Baltic Seas in summer 1988 a virus was isolated which was shown to be the causal agent. It was subsequently classified as morbillivirus by neutralization assays, reaction with monoclonal antibodies and nucleic acid hybridization studies. The virus (tentatively called Phocine Distemper Virus, PDV) is difficult to grow in culture making rapid diagnosis difficult. We have used the Polymerase Chain Reaction (PCR) as an alternative and fast method to detect the presence of virus-specific nucleic acid and we describe here the amplification of cell culture derived PDV RNA in a "one-tube" reaction using heterologous (Rinderpest Virus cDNA derived) F gene primers. The resulting 370 bp DNA fragment was shown to be morbillivirus derived by Southern blot hybridization using cloned RPV F gene as probe.

Animals↗

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.

Age Factors↗

Neutralizing antibodies to phocine distemper virus in Atlantic walruses (Odobenus rosmarus rosmarus) from Arctic Canada.

The first evidence of phocine distemper virus (PDV) infection in Atlantic walruses (Odobenus rosmarus rosmarus) from Nottingham Island, Northwest Territories, Canada, is reported. Blood samples were collected from three male walruses killed by Inuit hunters in the fall of 1990. Differential virus neutralization test for each animal yielded higher titers against PDV than against other members of the Morbillivirus genus including canine distemper, peste des petits ruminants, rinderpest and measles viruses. Thus, PDV infection may be enzootic in walruses of the eastern Canadian Arctic.

Animals↗

Demonstration of antibodies in archival sera from Canadian seals reactive with a European isolate of phocine distemper virus.

Sera from seals infected during the 1988 European epizootic of phocine distemper virus and sera from Canadian seals collected since 1972 have been tested for the presence of antibodies to morbillivirus. Approximately one third of the Canadian sera have been shown to contain anti-morbillivirus antibodies; the possibility that these populations of seals provided a source of infection for European seals is discussed.

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Identification and real-time PCR quantification of Phocine distemper virus from two colonies of Scottish grey seals in 2002.

The North Sea European harbour seal (Phoca vitulina) population has endured two phocine distemper virus (PDV) epidemics in 1988 and 2002. The grey seal (Halichoerus grypus) is a sympatric seal species that shows little or no mortality from PDV. Two Scottish grey seal breeding colonies were sampled for evidence of PDV infection approximately 2 months after the peak of the 2002 epidemic. In both colonies, a proportion of mothers (13/109) and pups (6/84) tested positive for PDV in their leukocytes. All infected animals were asymptomatic and completed the breeding season successfully. These results illustrate that grey seals come into contact with infectious seals and can become infected themselves without experiencing acute effects. In some seals the virus is able to replicate from the primary site of infection. This study provides evidence that grey seals may have an active role in the spread of PDV during an epidemic.

Animals↗

Molecular evolution of viral fusion and matrix protein genes and phylogenetic relationships among the Paramyxoviridae.

Phylogenetic relationships among the Paramyxoviridae, a broad family of viruses whose members cause devastating diseases of wildlife, livestock, and humans, were examined with both fusion (F) and matrix (M) protein-coding sequences. Neighbor-joining trees of F and M protein sequences showed that the Paramyxoviridae was divided into the two traditionally recognized subfamilies, the Paramyxovirinae and the Pneumovirinae. Within the Paramyxovirinae, the results also showed groups corresponding to three currently recognized genera: Respirovirus, Morbillivirus, and Rubulavirus. The relationships among the three genera of the Paramyxovirinae were resolved with M protein sequences and there was significant bootstrap support (100%) showing that members of the genus Respirovirus and the genus Morbillivirus were more closely related to each other than to members of the genus Rubulavirus. Both F and M phylogenies showed that Newcastle disease virus (NDV) was more closely related to the genus Rubulavirus than to the other two genera but were consistent with the proposal (B. S. Seal et al., 2000, Virus Res. 66, 1-11) that NDV be classified as a separate genus within the Paramyxovirinae. Both F and M phylogenies were also consistent with the proposal (L. Wang et al., 2000, J. Virol 74, 9972-9979) that Hendra virus be classified as a new genus closely related and basal to the genus Morbillivirus. Rinderpest was most closely related to measles and a more derived virus than to canine distemper virus, phocine distemper virus, or dolphin morbillivirus.

Databases, Nucleic Acid↗

Approaches in the understanding of morbillivirus neurovirulence.

Certain members of the morbillivirus genus, canine distemper virus, phocine distemper virus, and the cetacean viruses of dolphins and porpoises exhibit high levels of central nervous system (CNS) infection in their natural hosts. CNS complications are rare for measles virus (MV) and are not associated with rinderpest virus (RPV) and peste des petits ruminants virus (PPRV) infection. However, both RPV and PPRV are neurovirulent in permissive murine strains. Human postmortem tissue, neural cell cultures, and animal models have been used to answer major questions concerning morbillivirus neurovirulence. Studies of the MV CNS complication subacute sclerosing panencephalitis (SSPE) indicate that virus could enter the CNS either by direct infection of endothelial cells or in infected leucocytes, followed by infection of predominately neurones and oligodendrocytes. It has been established that MV neurovirulence in mice is partially determined by the virus-receptor specificity. The two known MV receptors, CD46 and SLAM, have been examined in normal and SSPE brain tissue and the findings suggest that further receptors may be necessary to explain infection of the CNS with wild-type strains of MV. In both humans and mice (and in vitro), once infection of neurones has been established, virus spreads transneuronally. It is possible that all morbilliviruses transiently infect the CNS in their natural hosts, but development of disease is dependent on the efficiency of the immune response. Alternatively, for RPV and PPRV, virus entry may be restricted due either to absence of viral receptors or failure of virus to replicate or spread in the CNS.

Animals↗

Phocine distemper virus, the agent responsible for the 1988 mass mortality of seals.

The biochemical characterisation of phocine distemper virus (PDV) has shown that PDV is related to but clearly distinct from canine distemper virus (CDV) and relative to its relationship with CDV is only remotely related to the other morbilliviruses, namely measles virus (MV) or rinderpest virus (RPV) and peste-des-petits-ruminants virus (PPRV). Comparative studies with monoclonal antibodies indicate that the virus is serologically closely related to CDV with many conserved epitopes, particularly on the internal proteins of the virus, while the external attachment (H) protein shows the greatest level of variability among the distemper virus isolates. The analysis of the viral proteins by electrophoresis indicates molecular weight differences between CDV and PDV in the fusion (F), phosphoprotein (P), H, nucleocapsid (N) and matrix (M) proteins. The RNA profiles of CDV and PDV are indistinguishable and different from those for RPV and MV. Nucleotide sequence analysis of cDNA clones of the virus show approximately 70% homology between CDV and PDV and approximately 48% with MV. These data prove that PDV is a different virus from CDV and co-circulates with it probably primarily in sea mammals.

Animals↗

The genes encoding the phospho- and matrix proteins of phocine distemper virus.

The nucleotide sequences of the phosphoprotein (P)/V/C and matrix (M) protein genes of phocine distemper virus (PDV) have been determined and the deduced amino acid sequences of the proteins derived from these genes compared with those of the other morbilliviruses. The 1655 nucleotides of the P gene encode a phosphoprotein of 507 amino acid residues (from nucleotide numbers 60 to 1583) which is 75% identical to that of canine distemper virus (CDV). The C proteins of the two viruses are 73% identical. The C protein has the same length, 174 amino acid residues, as C of CDV. The nucleotide sequences of the P genes are 78% identical. The editing site in the P gene is present as a stretch of 18 nucleotides, conserved in all other morbilliviruses. A V-like protein can be accessed by insertion of one G residue at this site. The P and M genes are separated from adjacent genes and each other by the CTT trinucleotide sequence which is totally conserved in morbilliviruses. The M gene is 1447 nucleotides long and encodes a typical paramyxovirus M protein of 335 amino acids (identical in length to those of CDV, measles virus and rinderpest virus). The gene is 78% identical to CDV in the coding region (nucleotides 32 to 1039) but only 34% identical to CDV in the 407 nucleotides of the 3' untranslated sequence of the M mRNA. The M protein of PDV is 91% identical to the M protein of CDV. The data demonstrate interesting variations in the level of conservation of various genome areas and prove the distinct nature of PDV.

Amino Acid Sequence↗

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.

Age Factors↗

Nucleotide sequence analysis of the large (L) genes of phocine distemper virus and canine distemper virus (corrected sequence).

This paper corrects the previously published sequence of the L gene of canine distemper virus (CDV). Errors in the published sequence (M. S. Sidhu et al., 1993, Virology 193, 50-65) led to frame shifts between residues 1021-1032, 1190-1219 and 1645-1650; a deletion of 21 amino acids between residues 1684-1705, and a single residue deletion at residue 1478. Residue 237 is now found to be glycine rather than tryptophan and residue 1626 proline instead of threonine. The sequence of the L gene of phocine distemper virus (PDV) was also determined. Alignment of the morbillivirus L proteins showed that PDV and CDV are more closely related to each other than to rinderpest virus and measles virus. Two regions of low identity are proposed to function as hinge regions between three highly conserved domains (I-III) in the morbillivirus L proteins. New sequence motifs have been identified on the basis of conservation in the morbilliviruses and the Paramyxovirinae.

Amino Acid Sequence↗

Modelling the spread of phocine distemper virus among harbour seals.

Data presented in earlier publications on the 1988 epizootic among seals in North West Europe show a pattern that is somewhat inconsistent with the predictions of the standard mathematical model of epidemics. We argue that for animals living in herds or colonies, such as seals, the mutual contact behaviour is such that models for the transmission of infectious diseases should be applied with special care for the distinction between numbers and densities. This is demonstrated by using a mechanistic description of the contacts among seals, which leads to a slightly different formulation of the model. Results of the analysis of this formulation are more in line with the data. The model introduced here can be applied to epidemics among all kinds of animals living in herds and in fact to any species with constant local density, independent of the total population size (i.e., occupying a variable area). Application of the traditional formulation, using different parameters for herds of different sizes, will give equally good results for non-lethal diseases. However, especially for diseases with a low R0 and high death rates, such as the phocine distemper virus (PDV) disease, the two model formulations give quite different results. Further analysis of the model is performed to determine the most important factors influencing such an epidemic. The survival of infected animals turns out to have a disproportionately great influence on the intensity of the epidemic. Therefore in the case of the PDV epizootic we conclude that marine pollution may not only have contributed to the high death rates, but, if so, it has intensified the epizootic as well.

Animals↗

Dolphin and porpoise morbilliviruses are genetically distinct from phocine distemper virus.

The morbilliviruses recently isolated from two cetacean species in the North and Mediterranean Seas have been shown to differ from phocine distemper virus isolated from European seals using monoclonal antibodies. We have identified a "universal" morbillivirus primer set, based on highly conserved regions of the morbillivirus phosphoprotein (P) gene and used this to amplify a region surrounding the RNA editing site from all known members of the group. Sequence analysis of this region of the gene shows that the dolphin and porpoise viruses are related but quite different from all other members of the group, forming a distinct lineage more closely related to the ruminant morbilliviruses than to the carnivore viruses.

Animals↗

Investigations on course and outcome of phocine distemper virus infection in harbour seals (Phoca vitulina) exposed to polychlorinated biphenyls. Virological and serological investigations.

The influence of polychlorinated biphenyls (PCBs) on phocine distemper virus (PDV) infections in harbour seals (Phoca vitulina) was studied. Six out of ten seals had been conditioned with a defined mixture of PCB-congeners for several weeks. Following exposure to the cell culture-propagated PDV isolate 2558/Han 88 the complete clinical picture of "1988 seal plague" was provoked in all ten seals inoculated. Four out of six PCB-conditioned seals and two out of four seals not loaded with PCBs succumbed to the infection within three weeks post inoculation. With regard to the clinical course, duration of cell-associated viremia, PDV-antigen distribution in tissues of fatally infected seals and the humoral immune response to PDV no differences between PCB-loaded and unloaded seals were recognized. Evidence was obtained that the pathogenesis of experimental PDV-infection in harbour seals shares some features with those of canine distemper in terrestrial carnivores. In contrast, however, to experimental distemper infection of gnotobiotic dogs prompt development of high titres of PDV-specific IgG did not correlate with recovery from infection.

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