[Problems of virus disinfection as exemplified with the Newcastle disease virus].
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Differential effect of various metabolic inhibitors on the development of hemadsorption activity and virus formation in cells infected with Newcastle disease virus (NDV) was investigated. It was found that, in BHK-21 cells infected with NDV, cycloheximide did not prevent the development of hemadsorption activity, whereas protein synthesis and virus formation by the cell were rapidly inhibited by the drug. When the drug was added to the culture at 4.5 h after infection or later, hemadsorption activity of the cell continued to develop normally for about 1 h. Similar increase in hemadsorption activity was found in cells which were treated with anti-NDV serum (to neutralize their hemadsorption activity) and then washed and incubated with cycloheximide. However, when cells were treated with the drug early in the infection (1.5 or 3.0 h), they did not show any detectable hemadsorption reaction throughout the infection. In contrast to cycloheximide, iodoacetate added to the culture together with sodium azide inhibited completely both the development of hemadsorption activity and the formation of progeny virus. These results suggest that the change of cell surface to become hemadsorptive may depend upon the energy generating system but not upon de novo synthesis of protein, whereas production of infectious virus may require continuous synthesis of protein.
The Cg strain of Newcastle disease virus (NDV) produces neurologic signs and death in mice. This illness is unusual because of the lack of typical features of a viral encephalitis. Specifically, there is a paucity of infectious virus, detectable cellular inflammatory reaction, cytopathic effect, and viral antigen by immunofluorescence. We previously showed an elevation of alpha-aminoisobutyric acid in the CNS of moribund NDV-infected mice, indicating cellular membrane dysfunction. In an attempt to further our understanding of the pathogenesis of the illness, we evaluated CNS concentrations of sodium, potassium, iron, copper, zinc, magnesium, selenium, and rubidium. Elemental analysis revealed no difference between infected and control mice for all elements except for rubidium, which was significantly elevated in infected mice. Elevation in rubidium was detected in infected mice by X-ray fluorescence and atomic absorption spectrophotometry, whereas rubidium concentrations for control mice were similar by both methods. Neurologic symptoms correlated directly with rising rubidium concentrations. Our data suggest that abnormal trace element levels during viral infection may be one mechanism responsible for the clinical symptoms.
The virions of Newcastle disease virus (NDV) contained an enzyme that catalyzed the incorporation of ribonucleotides into ribonucleic acid (RNA). Optimal conditions for this polymerase activity were identical to the conditions for the vesicular stomatitis virus (VSV) polymerase, and both enzymes were active for longer times at 32 C than at 37 C. However, the specific activity of the NDV polymerase was less than 3% that of the VSV polymerase. Product RNA species from the NDV and VSV polymerase reactions annealed specifically to the homologous virion RNA species. Transcriptive intermediates containing product RNA attached to the respective virion RNA could be identified in both systems.
The effects of single and combined vaccination of turkeys against hemorrhagic enteritis virus (HEV) and Newcastle disease virus (NDV) were investigated. Dual vaccination of turkeys with NDV-B1 and HEVp30 or marble spleen disease virus (MSDV) enhanced white mottling of the spleens and the apoptosis rate in spleen cells (P < 0.05). In addition, simultaneously vaccinated turkeys had fewer HEV-infected spleen cells at 4 days postvaccination than turkeys given HEVp30 or MSDV alone. The anti-HEV antibody response was significantly reduced at 14 days postvaccination (P < 0.05), whereas the anti-NDV antibody response was enhanced (P < 0.05) in turkeys vaccinated with HEVp30 + NDV-B1. Further, the effect of dual vaccination on macrophage function was studied. Spleen cells from NDV-B1-vaccinated turkeys were primed to produce nitric oxide (NO) after stimulation in vitro with lipopolysaccharide. Spleen cells from HEVp30- or MSDV-vaccinated turkeys did not produce NO after in vitro stimulation. In dual-vaccinated turkeys, the priming effect of NDV-B1 was reduced in comparison with single-inoculated birds.
Deoxycholate (DOC) disrupted virions of Newcastle disease virus (NDV), releasing viral nucleocapsids. The nucleocapsids sedimented at about 200S in sucrose gradients and measured from 1.3 to 1.4 mu long by electron microscopy. NDV nucleo-capsids were resistant to pancreatic ribonuclease. These nucleocapsids contained all the 50S ribonucleic acid (RNA) in NDV virions, while virus-associated RNA sedimenting at less than 50S was external to the virions.
Forty Newcastle disease virus strains and isolates could be placed in eight distinct antigenic groups on their ability to induce binding of nine mouse monoclonal antibodies, raised against strain NDV-Ulster 2C, to infected MDBK cells as assessed by an indirect immuno-peroxidase test. Viruses placed in each group appeared to share both biological and epizootiological properties.
Twenty-seven chicken red blood cell agglutinating agents were isolated from 187 tracheal swabbings of apparently healthy migratory mallard ducks (Anas platyrhynchos) in the Mississippi flyway. Twenty-four of the isolants were type A influenza virus; 3 lentogenic Newcastle disease viruses were isolated. Isolations were not made from either 65 giant Canada geese (Branta canadensis) or 60 Franklins' gulls (Larus pipixcan).
For the construction of transfer vector pFGHN1175-1, the gene encoding haemagglutinin-neuraminidase(HN) glycoprotein of newcastle disease virus(NDV) strain F48E8 was removed from plasmid pGEMHN, and inserted into the HindIII site of insertion vector pFG1175-1, downstream of P7.5 promotor. Chicken embryo fibroblast (CEF) cell cultures which had been infected with fowlpox virus (FPV) Chinese vaccine strain 282E4 for 3-4 hours were transfected with pFGHN1175-1 plasmid DNA by liposomal transfection. Recombinant FPV with blue plaques were selected and purified 3 times in CEF cell cultures overlaid agarose containing X-gal. PCR analysis and DNA dot-blotting hybridization assay indicated that the HN gene had inserted into the FPV genomic DNA. The expression of the NDV HN gene in recombinant FPV was confirmed by indirect immunofluorescence assay with specific monoclonal antibody.
Antiviral activity on Newcastle disease virus was examined with some respiration inhibitors including ascochlorin, rotenone, antimycin A(3), piericidin A, dicoumarol, 2, 4-dinitrophenol, pentachlorophenol, and fatty acids. Of the chemicals tested, ascochlorin and rotenone showed significant inhibitory effect on the viral growth in cultured cells as determined by the plate and tube assay methods. Dose response of ascochlorin and rotenone was observed in rate and final yield of hemagglutinin synthesis. Ratio of infectivity for hemagglutinin was nearly equal at any test concentrations of ascochlorin. Thus, the possibility of inhibition of virus maturation by the antibiotic was excluded. Ascochlorin had no activity on virus particles and on viral adsorption onto host cells.
The hemagglutinin-neuraminidase (HN) gene from the Beaudette C strain of Newcastle disease virus (NDV) has been expressed in a recombinant fowlpox virus vector. The HN gene, under the control of the vaccinia p7.5 promoter, was inserted into a nonessential gene in the terminal inverted repeats of fowlpox virus. Expression was demonstrated in tissue culture, a protein of the correct size for fully glycosylated HN protein being recognized by an HN-specific monoclonal antibody on Western blots. When the recombinant fowlpox virus was inoculated into chickens by intravenous or wing-web routes, antibody which recognizes HN from purified NDV virions was produced. Protective immunity to NDV was generated in the chickens; at the highest dose of vaccine 100% of the chickens tested were protected against challenge with a virulent strain of NDV.
The polypeptide composition of Newcastle Disease Virus (NDV) virions grown in two host cell cystems--chorioallantlic membrane (CAM) and BHK-21 cells--was studied. Two strains of virus were compared, one highly virulent, the other completely avirulent. No significant differences in the polypeptide composition of the two strains of virus could be detected. However, differences were found in virions grown in different hosts, the same differences being found in both strains. An additional polypeptide is found in BHK grown virus which is not present in CAM grown virus and this is associated with a decreased relative amount of nucleocapsid protein in BHK grown virus. The possibility of this new polypeptide being a degradation product of the nucleocapsid protein is discussed. BHK grown virions also contain increased amounts of a polypeptide migrating to a position which might be expected of the FO precursor glycoprotein. However, in contrast to the FO polypeptide, this polypeptide does not appear to be glycosylated.
Chickens 3 wk old, inoculated intranasally with a mesogenic (moderately virulent) strain of Newcastle disease virus, developed necrotic lesions of the mucous acini, predominantly of the middle turbinates. The infection subsequently spread to involve much of the rest of the mucosa, including mucous and ciliated epithelial cells, and other acini. The early phase of adsorption of a virulent strain of the virus to the middle turbinates of chicks 5-21 days of age was studied by giving a standard inoculum intranasally to unanesthetized animals. Variation in amounts adsorbed by individual chickens was large, but was minimized by making measurements on pools of turbinates from three chicks at intervals of 1, 3, and 5 hr after exposure of the excised turbinates to antibody, by washing, and by trypsinization. The virus released from the cells into the trypsin was designated as adherent virus, and the infectious virus in the cells after destruction of the cells by water grinding, as cell virus. Paralysis of ciliary action by cocaine increased the number of infected cells in the turbinates about 10-fold at all three time intervals. Pilocarpine injection before virus inoculation caused a large increase in the amount of infected cells 1 hr after virus administration, but was followed by a sharp drop in infected cells by 3 or 5 hr. Pilocarpine given after the virus decreased the number of infected cells and changed the relationship of infected cells to adherent virus. Exposure of chicks to sustained or severe cold caused a similar but less marked effect. The drop in infected cells was restored to control values if chicks were returned to brooder temperatures. The marked drop of infected cells produced by pilocarpine and cold in living chicks, and in cultures of chicken trachea (previous study), is consonant with the idea that virus has been adsorbed on mucus granules in the mucous cells of the turbinates and then has been reexcreted, as unincorporated virus, into the moving mucous sheet. A series of accessory data support this interpretation.
Newcastle disease virus (NDV) was concentrated and purified by microfiltration and exclusion liquid chromatography (ELC) on macroporous glass (MPG). The purified NDV eluted as one peak; its yield was strain-dependent, the degree of purification was 95% to 99%.
Inactivation of Newcastle Disease Virus (NDV) by binary ethylenimine (BEI) is reported. The activity of an oil vaccine prepared with BEI-inactivated NDV was compared to a vaccine prepared with formalin-inactivated NDV. The BEI inactivated vaccine had almost twice the efficacy.
A local virulent strain, VLT, of Newcastle disease virus formed 3- to 4-mm plaques on monolayers of primary chicken embryo cultures on the 4th day after inoculation. It agglutinated chicken and human 0 erythrocytes. Its hemagglutinin was stable at 56 C when compared with those of Komarov (K) and F vaccinal strains of the same virus. The viral titer of infected allantoic fluid dropped from 10(8.1) plaque-forming units to 10(1.0) plaque-forming units/ml within 2 hours when incubated at 56 C. The strain was ether-sensitive; it adsorbed readily on monolayers of chicken embryo cells and did not diffuse through agar. Its intracerebral pathogenicity index, chicken dose LD50, and embryo mean death time (hours) were 1.8, 9.0, and 48, respectively. Two virulent strains isolated in 1974 and 1975 were found to be identical to the VLT strain in terms of certain physical and biological properties. On the basis of plaque morphologic characteristics, hemagglutination spectrum, and hemagglutinin inactivation at 56 C, it was possible to identify readily the field isolate when it was compared with vaccinal strains (K and F) commonly used in Lebanon.
Newcastle Disease Virus (NDV) has interesting anti-neoplastic and pleiotropic immune stimulatory properties. The virus preferentially replicates in and kills tumor cells and appears to be safe and to varying degrees effective in phase II-clinical studies in the US and in Europe. Here we have compared various lytic and non-lytic strains of NDV with regard to their antitumor effects after local or systemic application. As tumor models we used human metastatic melanoma xenotransplants in nude mice and murine metastatic colon carcinoma (CT26), renal carcinoma (Renca) and lymphoma (ESb) cell lines. Intra or peri-tumoral application of NDV or NDV infected tumor cells showed more pronounced antitumor activity than systemic application even when in the latter case much higher dose ranges were used. In the CT26 colon carcinoma model the non-lytic strain Ulster showed stronger antitumor activity than the lytic strain 73T. In the human MeWo melanoma xentransplant model strong antitumor bystander effects were observed by 20% admixture of melanoma cells pre-infected in vitro with NDV (either strain Ulster or Italien). Virus therapy of pre-established human melanomas by intra-tumoral injection of NDV was effective with the lytic strain Italien but not with the non-lytic strain Ulster. Systemic anti-metastatic effects were never observed with NDV alone in contrast to previous results obtained with NDV modified tumor vaccines.
Antigenic variants of D26 strain of Newcastle disease virus (NDV) were selected with monoclonal antibodies directed to the three nonoverlapping antigenic sites on the hemagglutinin-neuraminidase (HN) protein, and their HN genes were sequenced to identify the amino acids important for the integrity of each site. Seven variants for site I, which is immunodominant and conserved among NDV strains, had a change of glutamic acid at position 347, mostly to lysine, and in a single case, to glycine. In the second group of two variants for site IV, a change of asparagine to aspartic acid was found at position 481. This resulted in elimination of the oligosaccharide attached to this asparagine residue of the parental virus. Together with the finding that the site IV was destroyed by treatment with endoglycosidase F, it was suggested that the oligosaccharide is important for maintaining the structure of site IV. The oligosaccharide appeared to contribute to exposing a nearby determinant by conferring hydrophilicity on it. A variant for site II had also a nonconservative mutation resulting in the change of glutamic acid to valine at position 495. The site I recognized by antibodies which inhibit neuraminidase activity with a small substrate neuraminlactose was located closer to the predicted sialic acid-binding site than to the other sites recognized by antibodies lacking the enzyme-inhibiting capacity. The sequence of the parental virus HN gene revealed that the HNo precursor for the HN protein is an extra-long protein whose C terminus is elongated by 45 amino acids, compared with the usual HN protein sequenced in parallel.