Sequence determination of the hemagglutinin-neuraminidase (HN) gene of human parainfluenza type 2 virus and the construction of a phylogenetic tree for HN proteins of all the paramyxoviruses that are infectious to humans

doi:10.1016/0042-6822(90)90081-2

Virology

Volume 174, Issue 1, January 1990, Pages 308-313

https://doi.org/10.1016/0042-6822(90)90081-2 Get rights and content

Abstract

The nucleotide sequence of the hemagglutinin-neuraminidase (HN) gene of human parainfluenza type 2 virus (PIV2) was determined. The PIV-2 HN gene was 2112 nucleotides excluding poly(A) tail. There was a single large open reading frame in the mRNA which encoded a protein of 571 amino acids with a calculated molecular weight of 63,262. Analysis of the deduced amino acid sequence revealed that there were fourteen potential glycosylation sites and a major hydrophobic region near the N-terminus, which would anchor the protein in the viral membrane. Comparisons of the HN protein sequences of PIV-2 with those of Simian virus 5 (SV5), Sendai virus (SV, parainfluenza virus type 1), human parainfluenza virus type 3 (PIV-3), type 4 (PIV-4), bovine parainfluenza virus type 3 (BPIV-3), mumps virus (MuV), and Newcastle disease virus (NDV) showed definite amino acid sequence relatedness, indicating a common ancestor for these viruses. Furthermore, statistical analysis of the protein sequences suggested a possible evolutionary relatedness among the paramyxoviruses. This is the first time that a phylogenetic tree has been constructed for all the parainfluenza viruses and mumps virus which are infectious to humans. In addition, amino acid sequences involved in hemag-glutinating and neuraminidase activities of paramyxovirus were discussed.

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Characterization of naturally occurring parainfluenza virus type 2 (hPIV-2) variants
2008, Journal of Clinical Virology
Citation Excerpt :
Glycosylation of a residue could likely mask an epitope, which could explain the absence of reaction with a specific monoclonal antibody. The other potential glycosylation sites remain unchanged (Precious et al., 1990; Kawano et al., 1990). For hPIV-4, it was shown that the appearance or suppression of glycosylation sites resulting from specific changes was an antigenic source of diversity and, possibly, constitution of sub-types (Komada et al., 2000).
Human parainfluenza viruses (hPIV) are respiratory pathogens responsible for upper and lower respiratory tract infections. In most labs, the clinical diagnosis of hPIV is routinely done using techniques based on the detection of viral antigens such as immunofluorescence assay or/and viral isolation.
Five hPIV-2 isolated from respiratory samples exhibited unusual phenotypic and antigenic characteristics. These isolates showed important syncytial cytopathic effect and failed to react with one specific monoclonal antibody. These variant strains were subsequently compared with hPIV-2 prototype strain by cellular and molecular techniques.
Both variant and prototype strains showed similar growth kinetics. Observation of plaque formation and syncytia assay indicated a more important fusogenic activity for the variant strains. Sequencing of fusion (F) and hemagglutinin-neuraminidase (HN) genes showed differences between the “atypical” hPIV-2 isolates and the Greer hPIV-2 prototype strain. These differences were analyzed with molecular modelling and structure prediction soft wares. A potential new glycosylation site in HN, in addition to minor changes observed in the predicted structure for the variant strains could explain their antigenic variation. Genetic changes in the fusion peptide and the cleavage site of F could also explain the difference observed in the fusion activity.
Continuous global viral surveillance is essential to monitor antigenic changes that may occur in nature particularly with regards to the implementation of diagnostic assays. The differences observed in F and HN between the prototype strain and clinical hPIV-2 variants could also provide new data for the analysis of Paramyxovirus fusion mechanisms and their pathogenesis.
Nucleotide sequence of the matrix protein gene of avian paramyxovirus, serotype 3b: Evidence on another member of the suggested new genus of the subfamily Paramyxovirinae
2005, Comparative Immunology, Microbiology and Infectious Diseases
The complete nucleotide sequence of the gene encoding the matrix protein (M) of the avian paramyxovirus, serotype 3b (APMV-3b), has been determined by means of the direct sequencing of viral RNA using reverse transcriptase reaction. The adjacent portions of the neighboring phosphoprotein (P) and fusion (F) protein genes were also sequenced that permitted to determine the consensus sequence of the viral genome, the poly(A) tract, downstream and upstream non-coding portions of the P and F genes, respectively, as well as the corresponding intergenic regions. The gene is 1478 nucleotides long with a protein-coding sequence of 1194 nucleotides. The deduced protein consists of 398 amino acids with a calculated MW 44,465. According to the multalignment and phylogenetic analyses, the APMV-3b M protein has shown the closest relatedness towards Newcastle disease virus (NDV) which has recently been suggested to be excluded from the Rubulavirus genus and assigned (together with APMV-6) to a new Avulavirus genus within the subfamily Paramyxovirinae of the Paramyxoviridae family. On the basis of the M protein genetic multalignment, phylogenetic relationships, bipartite nuclear localization signal identification in combination with the cysteine residues distribution, and by the degree of intrageneric heterogeneity, the APMV-3b is proposed to be another member (together with NDV and APMV-6) of the new genus.
La séquence nucléotidique complète du gène codant pour la protéine de la matrice d'un paramyxovirus aviaire, sérotype 3b (APMV-3b) a été déterminée par séquençage direct de l'ARN viral a l'aide d'une réaction ‘reverse transcriptase’. Les régions adjacentes des gènes de la phosphoprotéine (P) et de la protéine de fusion (F) ont aussi été séquençée afin de déterminer la séquence consensuelle du génome viral, de la zone poly(A), des régions non codantes en amont et en aval des gènes P et F respectivement, ainsi que les régions correspondantes entre ces différents gènes. Le gène est constitué de 1478 nucléotides dont une région codante de 1194 nucléotides. La protéine déduite est formée de 398 amino acides et à un poids moléculaire estimé de 44,465. Selon l'alignement multiple et l'analyse de l'arbre phylogénétique, la protéine M de l'APMV-3b montre la plus grande proximité avec le virus de la maladie de Newcastle (NDV) qui a été récemment proposé pour être transféré du genre Rubulavirus au nouveau genre Avulavirus (avec APMV-6) au sein de la sous-famille Paramyxovirinae de la famille Paramyxoviridae. Basée sur l'alignement multiple, l'arbre phylogénétique, l'identification du signal de localisation nucléaire ‘bipartite’ en combinaison avec la distribution de résidus cystéine et le degrée d'hétérogénéité interne au genre, le APMV-3b est donc proposé comme autre membre du nouveau genre (avec NDV et APMV-6).
Recovery of infectious human parainfluenza type 2 virus from cDNA clones and properties of the defective virus without V-specific cysteine-rich domain
2001, Virology
A full-length cDNA clone was constructed from the genome of the human parainfluenza type 2 virus (hPIV2). First, Vero cells were infected with recombinant vaccinia virus expressing T7 RNA polymerase, and then the plasmid encoding the antigenome sequence was transfected into Vero cells together with polymerase unit plasmids, NP, P, and L, which were under control of the T7 polymerase promoter. Subsequently, the transfected cells were cocultured with fresh Vero cells. Rescue of recombinant hPIV2 (rPIV2) from cDNA clone was demonstrated by finding the introduced genetic tag. As an application of reverse genetics, we introduced one nucleotide change (UCU to ACU) to immediate downstream of the RNA-editing site of the V gene in the full-length hPIV2 cDNA and were able to obtain infectious viruses [rPIV2V(−)] from the cDNA. The rPIV2V(−) possessed a defective V protein that did not have the unique cysteine-rich domain in its carboxyl terminus (the V-protein-specific domain). The rPIV2V(−) showed no growth in CV-1 and FL cells. Replication of the rPIV2V(−) in these cells, however, was partially recovered by adding anti-interferon (IFN)-β antibody into the culture medium, showing that the rPIV2V(−) is highly sensitive against IFN and that no growth of rPIV2V(−) in CV-1 and FL cells is mainly due to its hypersensitivity to endogenously produced IFN. These findings indicate that the V-protein-specific domain of hPIV2 is related to IFN resistance. On the other hand, the rPIV2V(−) efficiently replicated in Vero cells, which are known as a IFN-non-producers. However, the virus yields of rPIV2V(−) in Vero cells were 10- to100-fold lower than those of control rPIV2, although syntheses of the viral-specific proteins and their mRNAs in rPIV2V(−)-infected Vero cells were augmented up to 48 p.i. in comparison with those of rPIV2. Furthermore, the rPIV2V(−) virions showed anomalous in size as compared with rPIV2 virions. These results suggest that the V protein plays an important role in the hPIV2 assembly, maturation, and morphogenesis.
A link between chronic asthma and chronic infection
2001, Journal of Allergy and Clinical Immunology
Background: Asthma is a prevalent disease with marked effects on quality of life and economic societal burden. However, the cause of asthma and its pathophysiology are not completely defined. Recently, the possibility that chronic infection may play a role has been suggested. Objective: We sought to define the association between Mycoplasma and Chlamydia species and chronic asthma. Methods: We performed a comparison study of asthmatic patients and normal control subjects. Fifty-five patients with chronic stable asthma were compared with 11 normal control subjects by using PCR, culture, and serology for Mycoplasma species, Chlamydia species, and viruses from the nasopharynx, lung, and blood. Bronchoalveolar lavage cell count and differential, as well as tissue morphometry, were also evaluated. Computer-generated scoring for the degree of chronic sinusitis in asthmatic patients was additionally evaluated. Results: Thirty-one of 55 asthmatic patients had positive PCR results for Mycoplasma (n = 25) or Chlamydia species (n = 6), which were mainly found on lung biopsy specimens or in lavage fluid. Only 1 of 11 normal control subjects had positive PCR results for Mycoplasma species. The distinguishing phenotype between asthmatic patients with positive and negative PCR results was the significantly greater number of tissue mast cells in the group with positive results. Conclusion: A significant number of patients with chronic stable asthma demonstrate the presence of Mycoplasma species, Chlamydia species, or both in their airways, with the distinguishing feature of increased mast cell number. These findings need further delineation but may help us to understand the pathophysiology of asthma and new treatment options. (J Allergy Clin Immunol 2001;107:595-601.)
Molecular evolution of viral fusion and matrix protein genes and phylogenetic relationships among the Paramyxoviridae
2001, Molecular Phylogenetics and Evolution
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.
Rapid molecular epidemiologic studies of human parainfluenza viruses based on direct sequencing of amplified DNA from a multiplex RT-PCR assay
2000, Journal of Virological Methods
Sequencing studies of limited regions of the human parainfluenza viruses (HPIVs) genomes have helped describe patterns of virus circulation and characterize institutional outbreaks of HPIVs-associated respiratory illness. In this study, we sequenced reverse transcription polymerase chain reaction (RT-PCR)-amplified HPIVs RNA obtained from a multiplex RT-PCR assay described previously for simultaneous detection of HPIV-1, 2 and 3. Differences in the nucleotide sequences of limited regions of the HN gene allowed us to distinguish temporally and geographically diverse HPIV isolates (43 HPIV-1, 7 HPIV-2, 12 HPIV-3 isolates from this and previously published studies). In addition, an outbreak of HPIV-3-associated illness among infants on a pediatric ward was investigated by comparing sequences of three ward isolates with three matched community controls. Sequences of all ward isolates were identical and differed from those of the community controls, suggesting a single introduction and nosocomial transmission of the virus. Combining multiplex reverse transcription polymerase chain reaction (RT-PCR) assays with direct sequencing of the PCR products can provide an integrated system for rapid diagnosis and characterization of HPIVs.

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Short communicationSequence determination of the hemagglutinin-neuraminidase (HN) gene of human parainfluenza type 2 virus and the construction of a phylogenetic tree for HN proteins of all the paramyxoviruses that are infectious to humans

Abstract

Virology

Virology

Cell

Gene

Cell

FEBS Lett.

J. Biol. Chem.

FEBS Lett.

Virology

Intervirology

J. Gen. Virol.

J. Gen. Virol.

Arch. Virol.

Curr. Top. Microb. Immunol.

Rev. Infect. Dis.

Short communication
Sequence determination of the hemagglutinin-neuraminidase (HN) gene of human parainfluenza type 2 virus and the construction of a phylogenetic tree for HN proteins of all the paramyxoviruses that are infectious to humans