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PubMed1981

Temperature-sensitive mutant of Newcastle disease virus which has an altered nucleocapsid-associated protein.

Analysis of six temperature-sensitive (ts) mutants of Newcastle disease virus (NDV) representing each of six complementation groups by both SDS-polyacrylamide gel electrophoresis (SDS-PAGE) and two-dimensional gel electrophoresis revealed that in only one mutant was there an alteration in the isoelectric point of a protein. This altered protein was the nucleocapsid-associated protein, NAP. In addition, the mobility of the haemagglutinin-neuraminidase protein, HN, was decreased on non-reduced SDS-PAGE in this mutant. All independent ts+ clones derived from this mutant had normal NAP but HN protein migrated at the decreased rate. Haemagglutinating activity of wide-type (ts+) and ts virions was equally thermostable. Wild-type and ts+ clones derived from this ts mutant were RNA(+) at both permissive and non-permissive temperatures, whereas the ts mutant was RNA(-) at the non-permissive temperature. This ts mutant appears to be a double mutant in both HN and NAP genes, the latter only being a temperature-sensitive lesion which affects virus-directed RNA synthesis.

Capsid↗

PubMedDate not supplied

Effect of genetic selection for increased body weight and sex of poult on antibody response of turkeys to Newcastle disease virus and Pasteurella multocida vaccines.

Primary and secondary antibody responses of 671 turkeys of two genetic lines to Newcastle disease virus (NDV) and Pasteurella multocida vaccines were examined. The randombred control line (RBC2) and a subline (F) of RBC2 had been selected for increased 16-week body weight. Poults were vaccinated at 6 and 12 weeks of age, and serum samples were collected 3 weeks after each vaccination. Antibody titers were determined using an enzyme-linked immunosorbent assay. Line F turkeys had significantly higher 9-week and 15-week serum antibody titers to NDV than line RBC2. However, line RBC2 had significantly higher serum antibody titers to P. multocida at 15 weeks of age than line F. The 9-week and 15-week serum antibody titers to NDV were significantly higher in females than males, but males had significantly higher 15-week serum antibody titers to P. multocida than females. Sex of poults did not contribute significantly to variation in serum antibody response to P. multocida at 9 weeks of age.

Analysis of Variance↗

PubMed1987

Detection of polycistronic transcripts in Newcastle disease virus infected cells and identification of their sequence content.

The synthesis of six to seven polycistronic transcripts of Newcastle disease virus (NDV) in BHK cells was detected by Northern hybridization using cDNA clones generated by reverse transcription of five NDV mRNAs. Within the molecular weight range resolved by the gel electrophoresis system employed, four of the transcripts were suggested to be distronic, containing sequences of two genes, NP-P, P-M, M-F0 and F0-HN, respectively. In addition, tricistronic molecules of M-F0-HN and possibly of NP-P-M as well as P-M-F0 appeared to develop, although they were very low in amount. These data suggest a gene order of NP-P-M-F0-HN on the NDV genome. The polycistronic as well as monocistronic transcripts were generated with an almost constant proportion in amount throughout the virus replication. Further, at least several of them were also generated under the conditions where only the primary transcription was allowed by inhibiting de novo protein synthesis. Therefore, it appears likely that there is no distinct temporal control in NDV genome expression.

Amino Acid Sequence↗

PubMed1989

Location of neutralizing epitopes on the fusion protein of Newcastle disease virus strain Beaudette C.

A panel of eight neutralizing monoclonal antibodies (MAbs) against the fusion (F) protein of Newcastle disease virus (NDV) has been shown to locate a major antigenic site on the basis of competitive binding assay and additivity index studies. Five epitopes (A1 to A5) have been located within this site on the F protein of the Beaudette C strain of NDV on the basis of cross-resistance plaque assays of MAb-resistant mutants raised against these MAbs. Epitopes A1, A4 and A5 are distinct; epitope A2 partially overlaps epitope A3. Nucleotide sequence analysis of the F genes of MAb-resistant mutants showed that each predicted single amino acid substitutions ranging from amino acid residues 157 to 171 for epitope A4 and at residues 72, 78, 79 and 343 for epitopes A1, A2, A3 and A5 respectively. These locations indicate that both the F1 and F2 fragments are involved in the formation of a single antigenic site and suggest the involvement of extensive protein folding in the active form of this F protein.

Amino Acid Sequence↗

PubMed1986

Sequence of 2,617 nucleotides from the 3' end of Newcastle disease virus genome RNA and the predicted amino acid sequence of viral NP protein.

DNA fragments complementary to the Newcastle disease virus genome (strain D26) were cloned and sequenced. The sequence of 2,617 nucleotides from the 3' end of the genome was determined and an open reading frame (OP-1) consisting of 1,467 nucleotides, most likely encoding NP protein, was found in this region. This was followed by a second unfinished open reading frame (OP-2) of at least 729 nucleotides which continued beyond the 2,617th nucleotide. Another relatively short (312 nucleotides long) open reading frame (OP-2') was found overlapping with OP-2, but its significance is still unclear. The amino acid sequence deduced from the nucleotide sequence of OP-1 showed a moderate homology to that of the NP protein of Sendai virus in the central portion of the peptide. The leader sequence of 53 nucleotides was also identified. The 5' end of mRNAs synthesized in the infected cells was analyzed and found to be m7GpppA, suggesting that the transcription of viral mRNAs starts with A, but not with G residue.

Amino Acid Sequence↗

PubMed1989

Isolation and characterization of mouse FM3A cell mutants which are devoid of Newcastle disease virus receptors.

A method was developed to select host cell mutants which did not permit the replication of Newcastle disease virus (NDV), and 14 isolates of NDV-nonpermissive mutants of mouse FM3A cells were obtained. All these isolates were judged to be deficient in NDV receptors, since their ability to adsorb 3H-labeled NDV virions was markedly decreased. They were tested for genetic complementation in pairs by cell fusion and shown to fall into a single recessive complementation group, which was designated as Had-1. Vesicular stomatitis virus was able to replicate in this mutant to produce infectious progeny, but the glycoprotein of the released virion was abnormal in size, suggesting a defective processing of the asparagine-linked carbohydrate chains in the mutant cell. The Had-1 mutant was resistant to wheat germ agglutinin, but sensitive to a Griffonia simplicifolia lectin, GS-II, which recognizes terminal N-acetylglucosamine residues. The altered sensitivity to these plant lectins compared with that of the parental FM3A cells indicates that sialylated sugar chains on the cell surface are almost absent from the Had-1 cells, thereby rendering the cells NDV receptor deficient.

Animals↗

PubMed1999

The 1992 epizootic of Newcastle disease in double-crested cormorants in North America.

In the summer of 1992, morbidity and mortality in juvenile double-crested cormorants (Phalacrocorax auritus; DCC) attributable to Newcastle disease virus (NDV) was observed for the first time in seven northern USA states and one Canadian province, and recurred in three western Canadian provinces. Based on clinical signs and laboratory diagnostic findings, DCC mortality from NDV occurred in 59 of the 63 nesting colonies and two of three non-colony sites investigated. An estimate of in excess of 20,000 DCC died, with mortality rates ranging from < 1 to 37% in Great Lakes colonies to 20 to 92% in Minnesota (USA) and North and South Dakota (USA) colonies. Sick juvenile white pelicans (Pelecanus erythrorhynchos) exhibiting signs similar to sick cormorants, and dead pelicans were observed in Minnesota and North Dakota. Mortality rates in pelican colonies were as high as in the adjacent cormorant colonies, but no cause for the mortality of an estimated 5,000 pelicans was determined. No evidence of NDV was found in other species nesting in proximity to affected cormorants. Although the source of the NDV infection is unknown in cormorants, the simultaneous onset of the epizootics in juvenile birds over a wide geographic area implies that the virus was acquired by adults prior to migration and was carried back to nest sites, exposing susceptible nestlings. The possible transmission of this virus from free-ranging wild birds to domestic poultry is a concern. Based on repeated epizootics in cormorants since 1990, NDV seems to be established in DCC.

Animals↗

PubMed1986

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.

Amino Acid Sequence↗

PubMed1994

Isolation and characterization of a novel mutant mouse cell line resistant to Newcastle disease virus: constitutive interferon production and enhanced interferon sensitivity.

In our attempt to isolate mutant cell lines resistant to Newcastle disease virus (NDV) we developed an improved procedure for enrichment of NDV-resistant cells from mouse FM3A cells and isolated a novel NDV-resistant mutant cell line, Had-2, with characteristics different from Had-1, a previously reported NDV-receptor-deficient mutant strain. Had-2 cells adsorbed NDV normally but the accumulation of viral mRNAs and proteins was inhibited. Had-2 cells had to be grown at higher cell densities in order to be NDV-resistant, and it was revealed that they did not exhibit NDV-resistance when grown at lower cell densities. A conditioned medium prepared from a culture of Had-2 cells grown at high cell density was able to make a low-density culture NDV-resistant. The activity of the conditioned medium to induce NDV-resistance was completely neutralized by addition of both anti interferon (IFN)-alpha and anti IFN-beta antibodies, indicating that Had-2 cells were constitutively releasing IFNs, though their levels were rather low. Had-2 cells were also characterized by an increased sensitivity to IFNs as compared with the parental FM3A cells, since the conditioned medium containing IFNs did not render FM3A cells resistant to NDV.

Animals↗

PubMed2002

Fusogenic activity of reconstituted newcastle disease virus envelopes: a role for the hemagglutinin-neuraminidase protein in the fusion process.

Enveloped viruses, such as newcastle disease virus (NDV), make their entry into the host cell by membrane fusion. In the case of NDV, the fusion step requires both transmembrane hemagglutinin-neuraminidase (HN) and fusion (F) viral envelope glycoproteins. The HN protein should show fusion promotion activity. To date, the nature of HN-F interactions is a controversial issue. In this work, we aim to clarify the role of the HN glycoprotein in the membrane fusion step. Four types of reconstituted detergent-free NDV envelopes were used, on differing in their envelope protein contents. Fusion of the different virosomes and erythrocyte ghosts was monitored using the octadecyl rhodamine B chloride assay. Only the reconstituted envelopes having the F protein, even in the absence of HN protein, displayed residual fusion activity. Treatment of such virosomes with denaturing agents affecting the F protein abolished fusion, indicating that the fusion detected was viral protein-dependent. Interestingly, the rate of fusion in the reconstituted systems was similar to that of intact viruses in the presence of the inhibitor of HN sialidase activity 2,3-dehydro-2-deoxy-N-acetylneuraminic acid. The results show that the residual fusion activity detected in the reconstituted systems was exclusively due to F protein activity, with no contribution from the fusion promotion activity of HN protein.

Animals↗

PubMed2003

Recombinant Newcastle disease virus as a viral vector: effect of genomic location of foreign gene on gene expression and virus replication.

Newcastle disease virus (NDV) was examined for its suitability as a vector for the expression and delivery of foreign genes for vaccination and gene therapy. A reporter gene encoding human secreted alkaline phosphatase (SEAP) was inserted as an additional transcription unit at four different positions in the NDV genome, between the NP and P, M and F, and HN and L genes and behind the L gene. Eight infectious recombinant NDV (rNDV) viruses, four in the non-virulent strain NDFL and four in the virulent derivative NDFLtag, were generated by reverse genetics. SEAP expression levels, replication kinetics and virus yield were examined. Replication kinetics of the rNDV viruses in primary chicken embryo fibroblasts showed that the insertion of an additional gene resulted in a delay in the onset of replication. This effect was most prominent when the gene was inserted between the NP and P genes. With the exception of the strain that carried the SEAP gene behind the L gene, all recombinant strains expressed high levels of SEAP, both in cell culture and in embryonated chicken eggs. In embryonated eggs, the rNDV viruses showed a 2.6- to 5.6-fold (NDFL) or 2.1- to 8.1-fold (NDFLtag) reduction in yield compared with the parent strains. These results show that foreign genes can be inserted at different positions in the NDV genome without severely affecting replication efficiency or virus yield.

Alkaline Phosphatase↗

PubMed2004

Survival of exotic Newcastle disease virus in commercial poultry environment following removal of infected chickens.

During the first weeks of 2003, after exotic Newcastle disease (END) was confirmed in commercial layer flocks in Southern California, it became apparent that the virus survival information in the literature varied widely and was difficult to extrapolate to current local conditions. The END Task Force used the information available in the literature and the recommendations of research scientists to establish protocols for safely handling manure from infected and depopulated premises. In an attempt to gain more applicable knowledge in the management of contaminated poultry manure in the course of the END outbreak, this virus survival study was designed and implemented. Environmental drag swabs were tested for END virus from two of the early-infected commercial ranches that consisted of several houses following immediate removal of the infected flocks. A total of 293 samples, composed of 168 manure drag swab pools, 72 dropping board swab pools, and 38 compost swab pools from 3 houses (ranch 1), and 180 manure belt scraper swab pools from ranch 2 were analyzed for ND virus isolation and characterization for 21 consecutive days postdepopulation. Thirteen manure drag swab pools (from houses 1 and 3) and two manure dropping board swab pools (from house 3) collected from ranch 1 were positive for END virus at 0, 1, 2, 3, 4, 7, 10, 12, and 16 days postdepopulation. No END virus was isolated after the 16th day following depopulation from any of the samples. All samples from ranch 2 were negative during the entire observation period.

Animals↗

PubMed2006

[Construction and characterization of a recombinant fowlpox virus co-expressing F, HN genes of Newcastle disease virus and gB gene of infectious laryngnotracheitis virus].

The Fusion (F) and Haemagglutinin-Neuraminidase (HN) genes of Newcastle disease virus (NDV) and the glycoprotein B (gB) gene of infectious laryngothracheitis virus (ILTV) as well as a LacZ reporter gene were all inserted into a nonessential gene of fowlpox virus (FPV) 017 strain by homologous recombination. The NDV and ILTV genes were each under the control of a fowlpox virus immediate early/late promoter (LP2EP2) while the LacZ reporter gene expression cassette was regulated by a P11 late promoter. A recombinant FPV harboring the F, HN and gB genes as well as the LacZ gene, designated as rFPV-F/HN/gB/LacZ, was obtained after ten cycles of blue plaque purification. The presence of the NDV and ILTV genes was confirmed by PCR. The expression of the recombinant proteins in rFPV-F/HN/gB/LacZ were characterized by Western blot (F and gB proteins) and indirect immunofluorescence test (F, HN and gB proteins). The results demonstrated that all four foreign proteins, which were encoded within a 10 kb gene fragment, could be expressed authentically and efficiently. Compared to the parental virus, rFPV-F/HN/gB/LacZ showed no obvious difference with respect to virus replication and cytopathogenic effects in chicken embryo fibroblasts (CEF) cell culture. Overall, our work suggests that FPV can be a useful live virus vector for the expression of multi- foreign genes against multiple avian pathogens.

Animals↗

PubMed2000

Newcastle disease virus infection induces B7-1/B7-2-independent T-cell costimulatory activity in human melanoma cells.

Viral oncolysates of Newcastle disease virus (NDV) have been widely used for the treatment of malignant melanoma. Apparently, this nononcogenic and apathogenic paramyxovirus can alter the immunogenicity of tumor cells. To determine the influence of NDV infection on a tumor-specific T-cell response on a functional level, we used autologous primary melanoma cells infected with the NDV-strain Ulster. Therefore, melanoma cells and tumor-infiltrating lymphocytes were prepared from a freshly resected tumor, and tumor-infiltrating lymphocytes were subjected to limited dilution cloning. Proliferation assays of the T-helper cell clone sTS3 (CD4+) showed that the T-cell clone was rendered nonreactive against its autologous major histocompatibility complex II+, B7-1/B7-2- melanoma SMS, even remaining unresponsive to subsequent stimulation by interleukin-2. NDV infection of the SMS melanoma cell line not only completely restored the proliferative response of sTS3 to SMS, comparable with stimulation by cross-linking of anti-CD3/anti-CD28 monoclonal antibodies, but also inhibited the induction of anergy. Electrophoretic mobility shift assays of sTS3 cell lysates revealed the induction of the CD28-responsive complex by coincubation with NDV-infected melanoma cells. Because the induction of this complex of nuclear proteins shows specificity for the activation of the CD28 pathway but functional B7-1/B7-2 expression was not detectable on SMS melanoma cells at any timepoint, we propose the induction of a costimulatory factor different from B7 by NDV viral proteins.

Antigens, CD↗

PubMed1962