The order Mononegavirales--current status.
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Conserved Sequence↗
Biomedical subjects
C R Pringle
Publications and source records attributed to C R Pringle.
Analysis of the human serological immune response to a variable region of the attachment (G) protein of respiratory syncytial virus during primary infection.
The serum antibody responses of babies to the variable carboxy-terminal region of the attachment (G) protein of respiratory syncytial virus (RSV) have been analysed using paired acute and convalescent sera from infants experiencing their first RSV infection with viruses of known genotype. The variable 84-85 carboxy-terminal amino acids of the G protein of six recent isolates of group A RSV were expressed in Escherichia coli as fusion proteins with glutahione S-transferase. About half the infants developed antibodies which recognised these fusion proteins. The patterns of response obtained in enzyme linked immunosorbant assays and immunoblotting assays were closely related to the infecting genotype.
Acute Disease↗
Nucleotide sequence of the matrix protein gene of a subgroup B avian pneumovirus.
The nucleotide sequence of the gene encoding the matrix protein of a subgroup B avian pneumovirus has been determined. The gene shows 73.5% homology with that of a subgroup A virus, with most differences occurring in the third codon position. Comparison with pneumovirus matrix proteins shows that the APV matrix protein retains the hydrophobic domain common to the others. The analysis indicates that the matrix protein gene can be used to differentiate the two APV subgroups.
Amino Acid Sequence↗
Virus taxonomy 1996 - a bulletin from the Xth International Congress of Virology in Jerusalem.
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Animals↗
Sequence of the nucleocapsid protein gene of subgroup A and B avian pneumoviruses.
The nucleocapsid protein (N) gene of two subgroup A and one subgroup B strains of avian pneumovirus has been cloned and sequenced. The gene of all three isolates comprised 1197 nucleotides (nt), which formed a single major open reading frame, potentially encoding a protein of 391 amino acid residues. The N gene of the two subgroup A isolates differed by only 1 nt but differed by 282 (24%) nt and 35 (11%) amino acids from the B isolate. The predicted protein was identical in length to that of human, bovine and ovine respiratory syncytial viruses, the amino acid identity being approximately 41% overall but with some regions of identity > 90%.
Amino Acid Sequence↗
Expression and characterisation of the NS1 and NS2 proteins of respiratory syncytial virus.
The NS1 and NS2 proteins of human respiratory syncytial virus (RSV) were expressed using baculovirus. Antisera to these expressed proteins and to synthetic peptides were raised in rabbits and used to characterise the proteins. Multiple forms of both NS1 and NS2 proteins were detected in RSV infected cells by both immunoblotting and radioimmunoprecipitation when non-reducing, but not reducing, conditions were used. In pulse-labelling experiments the monomeric form of NS1 was stable, while that of NS2 was unstable with a half life of about 30 min. The NS1 protein associated with the matrix (M) protein and could be co-precipitated by a monoclonal antibody to M protein. The NS2 protein did not show any detectable association with RSV structural proteins. These results indicate that the NS1 and NS2 proteins have distinct roles in the viral life cycle.
Amino Acid Sequence↗
Identification of mutations contributing to the reduced virulence of a modified strain of respiratory syncytial virus.
The nucleotide sequences of the genome of the RSS-2 wild type strain of respiratory syncytial (RS) virus, which is known to induce upper respiratory tract infection in adults, and that of the attenuated ts1C candidate vaccine derived from it by three cycles of mutagenesis and selection of temperature-sensitive (ts) mutants, have been determined. Comparison of the sequences has located the genetic changes which contribute to the reduced pathogenicity in adults of the candidate vaccine. Thirty-seven nucleotide changes distinguish the wild type and ts1C, 13 of which confer amino acid substitutions; no mutations are present in extragenic regions. Partial nucleotide sequencing of the genomes of the first stage ts mutant (ts1A) and the second stage ts mutant (ts1B), which were intermediates in the derivation of the third stage mutant ts1C, established that five mutations resulting in amino acid substitutions had been induced in the first cycle of mutagenesis, one in the second cycle, and seven in the third cycle. The unique mutation differentiating ts1B from ts1A substitutes an alanine for a threonine at residue 736 in the polymerase (L) protein. The occurrence of a mutation in ts1C inducing substitution of a phenylalanine for a serine residue at an adjacent site (731) suggests that mutations in this region of the polymerase can have significant attenuating effects. The data suggest also that a mutation in the F gene may contribute to the attenuated phenotype.
Adult↗
Nucleotide sequence of the gene encoding the viral polymerase of avian pneumovirus.
We report here the nucleotide sequence of the L gene of avian pneumovirus (APV). This is the second pneumovirus L gene and the second avian paramyxovirus L gene, following that of Newcastle disease virus, to be sequenced. The APV L gene is 6099 nucleotides long and encodes a single large ORF of 2004 amino acids. This makes the APV L protein the smallest to be described for any nonsegmented, negative-strand RNA virus. The protein contains six linear non-contiguous domains, a putative ATP-binding site and four polymerase motifs previously described for the L proteins of negative-strand RNA viruses. Phylogenetic analysis of domain III of 14 different L proteins suggests the pneumoviruses to be as distant in evolutionary terms from the other members of the Paramyxoviridae as are the Filoviridae.
Amino Acid Sequence↗
Nucleotide sequences of the genes encoding the putative attachment glycoprotein (G) of mouse and tissue culture-passaged strains of pneumonia virus of mice.
The sequences of the genes encoding the putative attachment (G) proteins of pathogenic (strain J3666) mouse lung-passaged and nonpathogenic (strain 15) tissue culture-passaged strains of pneumonia virus of mice (PVM) have been determined. In both cases the major polypeptide was synthesised from the second open reading frame (ORF), a feature also found in the G gene of respiratory syncytial (RS) virus, another pneumovirus. However, the ORFs of the G genes of the two PVM strains were initiated at different nucleotide positions in the mRNA and comparison of hydrophobicity profiles revealed the presence of the putative amino-terminal cytoplasmic domain in the strain J3666 G protein and its absence in the predicted G protein of PVM strain 15. In common with the G protein of RS virus, the gene product of both PVM strains contained a high serine, threonine, and proline content. Indirect immunofluorescence analysis of BSC-1 cells expressing the G gene products confirmed the surface location of the proteins. Thus, the absence of a cytoplasmic domain does not interfere with the translocation of the G protein of PVM strain 15. In vitro translation of mRNA from the two PVM genes directed the synthesis of a larger polypeptide with the G gene of PVM strain J3666 than was seen with strain 15 G gene. In addition, a second protein was seen with strain J3666 mRNA which was the same size as the strain 15 G protein.
Amino Acid Sequence↗
Molecular epidemiology of respiratory syncytial virus: a review of the use of reverse transcription-polymerase chain reaction in the analysis of genetic variability.
Respiratory syncytial virus (RSV) is the major viral cause of lower respiratory tract disease (bronchiolitis and pneumonia) in babies and infants. Infections with the virus occur as annual winter epidemics in temperate climates, placing considerable pressure on the provision of hospital beds. The virus is unusual in that it can reinfect individuals and it can infect babies despite the presence of maternal antibody. RSV has a negative sense nonsegmented RNA genome and as such is liable to high levels of mutation. This paper describes methods developed to determine the degree of genetic variability of the virus both during individual epidemics and worldwide. It is necessary for these methods to be quick, easy and cheap so that large numbers of samples can be analysed readily. They are based on extraction of viral RNA directly from clinical samples or from viral cultures, reverse transcription of the viral RNA, and then amplification of selected regions of the genome by the polymerase chain reaction (PCR). PCR products are then analysed by restriction mapping, or, if necessary, direct nucleotide sequencing. In this way isolates of RSV have been shown to fall into a number of genotypes, with epidemics being made up of cocirculating genotypes whose relative proportions vary with each epidemic. An understanding of the molecular epidemiology of this important human pathogen will be of significance in the search for an effective vaccine.
Genetic Variation↗
International Committee on Taxonomy of Viruses (ICTV) activity report.
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Classification↗
Variation in the fusion glycoprotein gene of human respiratory syncytial virus subgroup A.
Six different genotypes (designated lineages SHL1-6) of human respiratory syncytial (RS) virus have been defined by partial nucleotide sequence analysis of the variable SH and the hypervariable G membrane protein genes, and by restriction fragment analysis of the conserved N protein gene of viruses isolated in south Birmingham. Viruses of very similar genotype appear to be present worldwide at the present time. We have determined the nucleotide sequences of the fusion protein genes of five viruses isolated in south Birmingham in the same year, but belonging to different lineages, and have compared them with the sequences of four subgroup A viruses isolated at earlier times from diverse localities. The sequence diversity of the F genes of these five viruses, as measured by nucleotide (94.5-98.5%) and inferred amino acid (97.0-99.3%) identifies, is comparable with that of the nine subgroup A viruses considered as a whole. No sequence changes occur in any of the sites of known epitopes. Comparison of the nine subgroup A sequences with the published sequences of a subgroup B strain and three bovine RS viruses confirms that the F protein sequences are most divergent in the F2 region.
Amino Acid Sequence↗
Sequence and in vitro expression of the phosphoprotein gene of avian pneumovirus.
The phosphoprotein (P) gene of two subgroup A strains of avian pneumovirus comprised 855 nucleotides containing only one substantial open reading frame encoding a protein of 278 amino acids, with a predicted M(r) of 30,323. In vitro translation of P mRNA in a wheat germ system resulted in the synthesis of two polypeptides of M(r) 35,000. Comparison of the deduced P protein sequence with that of the known mammalian pneumoviruses revealed overall amino acid identities ranging from 31 to 34.5%, suggesting a distant relationship. However, there was a much higher identity (63.2-68.4%) in a region of 57 residues, which included a heptad repeat sequence.
Amino Acid Sequence↗
Sequence variation within an outbreak of measles virus in the Coventry area during spring/summer 1993.
Measles virus (MV) was isolated from throat swab samples collected during the spring/summer 1993 in the Coventry area. Viral RNA was reverse transcribed and cDNA prepared using oligo- T primer. Using MV-specific primers the area encoding the external region of the haemagglutinin glycoprotein was amplified using nested PCR and cycle sequenced. Comparisons were made with the Edmonston strain and current MMR vaccine strain. It was found that a high degree of homology existed between all strains examined, but that a majority of clinical samples shared a premature termination signal that potentially shortened the haemagglutinin protein by 35 amino acids. The single clinical sample that lacked this early termination signal appeared to be closely related to the MMR strain and may result form a vaccine-related illness. Truncation of the haemagglutinin protein may have allowed MV to escape the immune response induced by vaccination with the current MMR vaccine.
Adult↗
Evolution of subgroup A respiratory syncytial virus: evidence for progressive accumulation of amino acid changes in the attachment protein.
The variability of the attachment (G) proteins of 48 subgroup A isolates of respiratory syncytial virus (RSV) isolated over 38 years has been examined. Nucleotide sequences of two variable regions of the G protein genes were determined following amplification by PCR. The isolates showed temporal rather than geographical clustering, and there was evidence for progressive accumulation of amino acid changes at an average rate of approximately 0.25% per year estimated over the entire protein. The cocirculation of lineages of RSV at present appears to be the result of a process of evolution and survival of particular genotypes and the extinction of others. Analysis of reactivity of the isolates with monoclonal antibodies showed that their antigenic profiles closely paralleled their relatedness by nucleotide sequence, suggesting that antigenic drift due to immune selection may be occurring.
Amino Acid Sequence↗
Antigenic diversity of respiratory syncytial virus subgroup B strains circulating during a community outbreak of infection.
The epidemiological characteristics and relationship between respiratory syncytial virus (RSV) subgroup and virulence during an outbreak of RSV infection occurring in Southeast Texas in the winter season 1991/92 are described. Fifty-two infants and children were diagnosed with RSV infection by rapid viral antigen detection and/or viral isolation. Subgrouping of the isolates was carried out using 11-monoclonal anti-bodies. Ten isolates were found to be subgroup B, and 8 isolates were subgroup A. The subgroup B strains showed 3 different patterns of reaction with monoclonal antibodies; one of these subgroups was examined further by restriction analysis of parts of its nucleocapsid and attachment protein genes. The peak of RSV outbreak was in December 1991. Both subtypes A and B circulated simultaneously in the same territory, and caused lower respiratory tract infections in similar proportions. The more frequent occurrence of the B subgroup and the diversity of its simultaneously circulated RSV strains have made this outbreak unusual.
Antigens, Viral↗
Analysis of respiratory syncytial virus strain variation in successive epidemics in one city.
The variability of respiratory syncytial virus isolates from five successive epidemics in an urban population was determined. A total of 187 isolates of respiratory syncytial virus from the southern part of Birmingham, United Kingdom, were classified into subgroups A and B and were then further assigned to genetic lineages. Allocation of isolates into lineages was achieved by reverse transcription of infected cell RNA and then PCR amplification of selected regions of the genome; PCR products were examined by restriction mapping or nucleotide sequencing of parts of the nucleoprotein gene, the small hydrophobic protein gene, and the attachment protein gene. Previous work has shown that estimations of genetic diversity by analysis of genes coding for proteins likely (attachment protein) and unlikely (nucleoprotein and small hydrophobic protein) to be under immune pressure gave concordant results. Six genetic lineages of subgroup A isolates have been defined by this procedure; these isolates differ by up to 20% in the amino acid sequences of their attachment proteins; likewise, subgroup B isolates can be divided into two categories by restriction mapping of parts of their nucleoprotein and attachment protein genes. The same genetic lineages appeared to be present worldwide during the same period. The analysis of isolates from successive epidemics showed that different lineages predominated in each epidemic and that not all lineages were present in every epidemic. Some lineages appeared to increase in numbers over several years and then decline, possibly indicating a buildup of resistance in the community to a particular genotype.
Capsid↗
Sequence of the phosphoprotein gene of pneumonia virus of mice: expression of multiple proteins from two overlapping reading frames.
The gene encoding the phosphoprotein of the pneumovirus pneumonia virus of mice (PVM) has been cloned and sequenced. The gene is 903 nucleotides in length and contains a long open reading frame (ORF) capable of encoding a polypeptide of 295 amino acid residues. A smaller, second, overlapping ORF encoding a polypeptide 137 amino acids in length was also present. The large ORF directed the synthesis of a 39-kDa polypeptide and four additional polypeptides with masses of 37 kDa, 26 kDa, 23 kDa, and 16 kDa in vitro. The smaller polypeptides were generated by internal initiation on in-frame AUG initiation codons to generate carboxy co-terminal products. Western immunoblot analysis indicated that at least two of these proteins and several other related polypeptides are present in infected cells, and the possible origins of these are discussed. Western blot analysis using antiserum raised against a synthetic peptide and specific for the predicted second ORF product identified a polypeptide of 23 kDa in PVM-infected cells. The pattern of PVM P gene expression is unlike that of the closely related respiratory syncytial virus and is reminiscent of that of paramyxoviruses such as Sendai virus. This is the first example of a pneumovirus encoding multiple polypeptide products from a single mRNA in vivo.
Amino Acid Sequence↗