A review of the 1988 European seal morbillivirus epizootic.

An epizootic of morbillivirus infection killed thousands of common seals (Phoca vitulina) in European seas in 1988. Most of the affected seals had respiratory signs and the main post mortem finding was acute pneumonia. The histopathological changes were similar to those of canine distemper. Six common porpoises (Phocoena phocoena) found stranded on the coast of Northern Ireland in late 1988 had similar lesions. Morbillivirus infection also killed several thousand Siberian seals (Phoca siberica) in Lake Baikal in 1987 and 1988. A morbillivirus (phocine distemper virus) has been isolated from affected seals in several European countries and studies of the antigenicity of the virus indicate that it has several unique epitopes that distinguish it from the other known morbilliviruses. Biochemical studies of the viral proteins, RNA and nucleotide sequence confirm that it is a new morbillivirus. There is seroepizootiological evidence of morbillivirus infection in Greenland harp seals (Pagophilus groenlandicus), ringed seals (Phoca hispida) and Dutch common seals several years before the 1988 epizootic. Antibodies to a morbillivirus have also been found in bottlenose dolphins (Tursiops truncatus) from the eastern coast of the USA. Further studies are required to determine whether these sea mammal populations have been infected with phocine distemper virus.

Immunological relationships of simian virus 41 (SV41) to other paramyxoviruses and serological evidence of SV41 infection in human populations.

Antigenic relationships of simian virus 41 (SV41) to other paramyxoviruses were examined by immunoprecipitation of isotope-labelled SV41-infected cell lysates with specific antisera. SV41 is closely related to the group comprising human parainfluenza virus 2 (HPIV-2), simian virus 5 (SV5), parainfluenza virus 4 and mumps virus. Slight cross-neutralization was detected between SV41, HPIV-2 and SV5. Anti-SV41 activities were detected in 21 of 1116 human serum specimens, indicating that a proportion of the human population is infected with SV41. The haemagglutinin-neuraminidase of SV41 was preferentially immunoprecipitated by anti-SV41 positive sera.

Immunological relationships between phocid and canine distemper virus studied with monoclonal antibodies.

The immunological relationships between distemper viruses, isolated from a seal and mink in Denmark and from a dog in Greenland, were investigated with 39 previously developed monoclonal antibodies (MAbs) directed against four major structural proteins of canine distemper virus (CDV). They were also investigated with 16 newly developed MAbs directed against the fusion (F) and large glycoprotein (named H in analogy with measles virus) of phocid distemper virus (PDV) isolated from a harbour seal (Phoca vitulina). These MAbs were reacted with the three different isolated viruses and with the LEC strain of measles virus, in ELISA and immunofluorescence tests. In addition, immunoprecipitation tests were carried out with some of the cross-reacting antibodies. All 55 MAbs reacted identically with distemper virus isolated from seals or mink. When the MAbs produced against CDV were tested, 37 of 39 antibodies reacted with a virus isolated from a sled dog diseased in an outbreak of distemper in Greenland prior to the epizootic among seals in the North Sea. Of the 39 antibodies, 25 reacted with PDV and distemper virus isolated from mink. Of these antibodies, only three of the nine antibodies directed against the H protein of CDV cross-reacted with PDV and distemper virus from mink. Eleven MAbs, reacting with six epitopes of the H protein of PDV, were produced. All 11 antibodies reacted with distemper virus from mink, two of the antibodies reacted with CDV and none reacted with measles virus. All five antibodies reacting with three different epitopes of the F protein of PDV reacted with distemper virus from mink and CDV. Of these five antibodies three, directed against two epitopes, reacted with measles virus. Of the two envelope proteins, the H protein shows pronounced immunological differences between PDV and CDV. In contrast, immunologically the F protein appears to be well conserved among morbilliviruses. It is concluded that the virus causing the epizootic in seals in the North Sea in 1988 may have infected mink on land, or, alternatively, the virus in the sea may have originated from virus-infected mink.

Morbillivirus group: genome organisation and proteins.

The Paramyxoviridae family is divided into three genera: Paramyxovirus, Pneumovirus and Morbillivirus. In the last group, there are four closely related viruses which are seriously pathogenic for man and animals, and usually cause acute diseases. At least two of them (measles and canine distemper viruses) can cause a persistent infection which leads to a chronic disease of the nervous system that, in the end, is fatal. For a long time, the biochemical analysis of morbilliviruses was hampered by the high susceptibility of some of their proteins to proteolysis. With cloning and sequencing technology, more data on the biology of those viruses are now available.

The structural proteins of a porcine paramyxovirus (LPMV).

The porcine paramyxovirus is a newly identified agent of a fatal disease in piglets, endemic in Mexico since 1980, where it was seen around the town of La Piedad, Michoacan, Mexico (hence LPM virus). At least six [35S]methionine-labelled proteins could be resolved by SDS-PAGE and five of them were clearly immunoprecipitated. Selective labelling of LPMV-infected cells with [3H]glucosamine revealed two bands with an Mr of about 66K and 59K, corresponding to the two viral glycoproteins, the haemagglutinin-neuraminidase protein and the fusion protein. Labelling of virus with [32P]orthophosphate disclosed one band with an Mr of 52K, corresponding to the phosphoprotein. Analysis of nucleocapsids obtained from purified virus or from a permanently infected cell line revealed one major band with an Mr of 68K, the nucleoprotein. Two other proteins were also identified, the large protein and the matrix protein, with apparent Mr of about 200K and 40K, respectively. The protein migration pattern of LPMV was compared, by SDS-PAGE, with that of Newcastle disease virus, bovine parainfluenza 3 virus and Sendai virus. Differences in the Mr of LPMV proteins and the proteins of these paramyxoviruses were observed. We propose that LPMV should be classified as a novel member of the genus Paramyxovirus.

Polypeptides of pneumonia virus of mice. II. Characterization of the glycoproteins.

The kinetics of synthesis and the nature of the oligosaccharides of the glycoproteins of pneumonia virus of mice (PVM) were studied. Tryptic peptide mapping showed that the two major glycosylated polypeptides G1 and G2 were different forms of the same protein. G2 was derived from G1 which in turn appeared to be derived from an unidentified precursor. The G1/G2 protein of PVM is probably a haemagglutinin since a monoclonal antibody directed against it has a high haemagglutination inhibition titre. On the basis of experiments with inhibitors and glycosidases it was deduced that G1 and G2 have both N-linked and O-linked oligosaccharides. The putative fusion protein-equivalent of PVM was shown to possess N-linked oligosaccharides. In the presence of tunicamycin a high mobility form (F1t) appeared to be derived from a precursor (F0t) with the same mobility as the fully glycosylated protein. If by analogy with other paramyxoviruses this represents a cleavage event, the difference in mobility of the precursor and product suggests that the putative F2 product is smaller than the corresponding F2 protein of other paramyxoviruses. However, no F2 candidate protein was detected and evidence for an F1,2 dimer was inconclusive. The glycoproteins of PVM resemble those of respiratory syncytial virus in terms of their pattern of glycosylation, but differ in their processing.

Pneumovirus-like characteristics of the mRNA and proteins of turkey rhinotracheitis virus.

Electronmicroscopy has indicated that turkey rhinotracheitis virus (TRTV), the causative agent of an acute respiratory disease in turkeys, is a member of the Paramyxoviridae family. To determine if TRTV belongs to one of the three defined genera of this family (Paramyxovirus, Morbillivirus and Pneumovirus) we have analysed the RNA and proteins induced during replication of TRTV in Vero cells. Following replication in the presence of actinomycin D 10 polyadenylated RNA bands, ranging in Mr from 0.22 to 2.0 X 10(6), were detected in infected cells; some bands probably contained 2 or more RNA species. Viral proteins were studied after radiolabelling in the presence of [35S]methionine and [3H]glucosamine. Comparison of the polypeptides in mock-infected and infected cells, virions and nucleocapsids and after lentil-lectin chromatography and immunoprecipitation revealed seven virus-specific polypeptides (p), some of which were glycosylated (gp): gp82 (Mr 82K), gp68, gp53, gp15, p43, p40 and p35. These are considered to be analogous to the large glycopolypeptide (HN, H and G), fusion protein precursor F0, the F protein cleavage products F1 and F2, nucleocapsid (N), phosphorylated (P) and matrix (M) polypeptides, respectively, of the Paramyxoviridae. Two other polypeptides (Mr 200K and 22K) were also detected, as was a glycopolypeptide of Mr 97K, probably related to gp82. Tunicamycin inhibited glycosylation of gp53 and gp15 but gp82 was little affected, most glycans still being present on a glycopolypeptide of approximately 79K. This finding, indicating that gp82 has mostly O-linked glycans, considered with the mRNA profile and the molecular weight of the N protein shows that of the three genera in this family, TRTV most closely resembles the Pneumovirus genus.

Comparison of proteins induced in cells infected with rinderpest and peste des petits ruminants viruses.

The two morbilliviruses rinderpest virus (RPV) and peste des petits ruminants virus (PPRV) are closely related and cause severe disease in large and small ruminants, respectively. They show distinct epidemiological patterns and are distinguishable by reciprocal cross-neutralization tests. We have analysed the proteins induced by these viruses in infected cells and have shown that they can be distinguishable by a very marked difference in the apparent mol. wt. of the nucleocapsid (N) protein. The N protein of PPRV is almost identical in mobility on polyacrylamide gels to the N proteins of measles virus and canine distemper virus (60K). Several strains of RPV and PPRV from widespread geographical locations were studied and found to show this difference in the N protein.

Fusion glycoprotein of measles virus: nucleotide sequence of the gene and comparison with other paramyxoviruses.

The sequence of the fusion (F) glycoprotein mRNA of the Hallé strain of measles virus was determined from a cDNA clone representing the entire length of the mRNA. It contained 2384 nucleotides, excluding poly(A), with a 5' consensus sequence typical of paramyxoviruses and a 3' terminus found in measles virus mRNAs. The coding sequence was preceded by an unusually long (580 nucleotide) 5' non-translated region, which contained 44% cytosine. The longest open reading frame coded for a polypeptide of 553 amino acids with a predicted molecular weight of 59.84 K. Comparison of the sequence with that of the Edmonston strain of measles virus showed that the gene is highly conserved. No amino acid differences were observed between the two strains. The F polypeptide had three regions of high hydrophobicity: an N-terminal signal peptide, the N-terminus of F1 and a C-terminal membrane-spanning region. The four potential asparagine-linked glycosylation sites (one in the signal peptide) were all in the F2 subunit. Comparison of the measles virus F amino acid sequence with other paramyxoviruses revealed homologies with these viruses. Certain regions such as the N terminus of F1 and ten cysteine residues which probably impose structural restraints were highly conserved.

Nucleotide sequence of the gene encoding the fusion glycoprotein of Newcastle disease virus.

The nucleotide sequence of the gene encoding the fusion (F) glycoprotein of the Beaudette C strain of Newcastle disease virus (NDV) has been determined from cDNA clones obtained from virion RNA. The gene is 1792 nucleotides long, including mRNA start and polyadenylation signals typical of paramyxoviruses. The single open reading frame encodes a polypeptide of 553 amino acids, with a predicted molecular weight of 59042. The F polypeptide has three regions of high hydrophobicity: an N-terminal signal peptide, the N terminus of F1 (known from protein sequencing) and a C-terminal membrane-spanning region by which the F glycoprotein is anchored to the membrane. The cleavage site of F0 is located in a highly basic region of the F polypeptide. Five potential asparagine-linked glycosylation sites are present in the amino acid sequence, of which one is in F2 and the others in F1. Comparison of the NDV F amino acid sequence to those from other paramyxoviruses reveals homology to Sendai virus, simian virus 5 and human respiratory syncytial virus. There is also limited homology between the N terminus of F1 of NDV and the N termini of HA2 of influenza viruses. Post-translational modifications of the NDV F polypeptide are discussed in the light of information provided by the amino acid sequence.

Sequence analysis of the P and C protein genes of human parainfluenza virus type 3: patterns of amino acid sequence homology among paramyxovirus proteins.

The complete nucleotide sequence of the P + C mRNA of human parainfluenza virus type 3 (PF3) was determined by sequencing cDNA, viral genomic RNA and mRNA. The P + C mRNA is 2009 nucleotides in length, exclusive of poly(A), and contains two overlapping open reading frames (ORFs). The P + C mRNA encodes two proteins, the 602 amino acid nucleocapsid phosphoprotein P and the 199 amino acid non-structural protein C. Peptide mapping confirmed that the two proteins are unrelated. Hybrid-arrest translation experiments assigned each of the two proteins to its respective ORF. These studies showed that the coding strategy of the PF3 P + C mRNA is similar to that of Sendai virus. Amino acid sequence alignment showed that the P and C proteins of PF3 and Sendai virus represent homologous pairs. However, these homologies are represented by high contents of accepted amino acid substitutions and by similarity in hydropathy profiles rather than by high contents of exact amino acid matches. Homology with the P and C proteins of measles, canine distemper and respiratory syncytial viruses was at the threshold of significance. The patterns of amino acid sequence homology among the paramyxovirus HN, F, NP, P and C proteins are compared.

Characterization of a paramyxovirus isolated from the brain of a piglet in Mexico.

A virus morphologically resembling members of the family Paramyxoviridae has been isolated from the brain of a piglet with a central nervous disorder accompanied by pneumonia and corneal opacity. The virus, designated LPM, grows in a large variety of cultured cell types and elicits a cytopathic effect including formation of syncytia and cytoplasmic inclusion bodies. The virus has hemagglutinating, neuraminidase and hemolytic activities. Studies on experimental transmission showed that young pigs are susceptible to infection which induced a disease similar to that in natural cases. The virus killed mice and chicken embryos. The structural proteins of LPM virus, as resolved by polyacrylamide gel electrophoresis are similar to those described for other paramyxoviruses. Serologically the virus proved to be distinct from the paramyxoviruses tested so far.

The structural polypeptides of avian paramyxoviruses.

The structural polypeptides of twenty-three avian paramyxoviruses from five serotypes were examined by polyacrylamide gel electrophoresis in the presence of sodium dodecylsulphate under reducing and non-reducing conditions. All virus polypeptide profiles consisted of 7--10 polypeptides of which two were glycosylated. Some variation was seen in the profiles of viruses from the same serotype, but groups formed on the basis of their serological relationships in haemagglutination inhibition tests were identical to those formed on the basis of similarities in their polypeptide profiles.

Intrinsic fluorescence of some myxo- and paramyxoviruses and of their subviral fractions.

Fluorescence spectra of Sendai and influenza A(H1N1) viruses have different emission maxima; their quantum yield is much lower than that of tobacco mosaic virus. Solubilized envelopes and nucleoproteins of Sendai, influenza and mumps virus have different half-bandwidth and relative quantum yield values of emission, according to the agent used for disruption. Thus emission (as well as excitation and absorption) spectra of Triton X-100-solubilized envelopes show a marked hypsochromic shift as compared with the envelopes obtained by Tween20-disruption. The results are correlated with the different disruption extent achieved with the two agents.

Biochemical characterization of a Yucaipa-like virus.

A Yucaipa-like virus (PLOC/Senegal/273/77) was grown in embryonated chicken eggs and used for biochemical investigations after purification. The genome of the virus is composed of one fragment of single-stranded (ss)RNA with an estimated mol. wt. of 5.6 X 10(6). There are six virus structural polypeptides with mol. wt. of 126 000, 68 000 (major), 60 000, 52 000 (major), 44 000 and 39 000 (major). The fatty acid composition of the virus envelope seems to be very selective since we found only fatty acids containing 14 and 16 carbon atoms.

Antigenic and structural relationships between avian paramyxoviruses isolated from ducks in Hong Kong and Mississippi, U.S.A.

Representative isolates of the paramyxoviruses duck/Hong Kong/75 and duck/Mississippi/75 were shown to be serologically closely related by haemagglutination and neuraminidase inhibition tests. The structural polypeptides of these viruses were also shown to be similar. For each of the isolates tested, polyacrylamide gel electrophoresis in the presence of SDS revealed a similar polypeptide pattern consisting, under reducing conditions, of seven polypeptides with apparent mol. wt. ranging from 46000 to 190000. Each virus had two glycosylated polypeptides with apparent mol. wt. of 56000 and 71000 to 72000 under reducing conditions and 62000 to 63000 and 135000 to 142000 under non-reducing conditions.