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A highly virulent Newcastle disease virus (SA84) was isolated from a large broiler operation in Saudi Arabia. The mean death time of chicken embryos given the minimum lethal dose, the pathogenicity of the isolate for 8-week-old chickens, the plaque characteristics, and the intracerebral pathogenicity index indicated that the isolate is of the viscerotropic velogenic pathotype.
The rate of destruction of hemagglutinins and infectivity of Newcastle disease virus was determined over a temperature range of 37.8 to 60 C. From the calculated values of deltaH and deltaS, it was concluded that inactivation of the hemagglutinating activity and viral infectivity was due to protein denaturation.
Experiments employing the various steps and reagents used in the enzyme-linked immunosorbent assay (ELISA) were conducted to produce an ELISA with the highest sensitivity and specificity possible for detecting Newcastle disease antibodies in chicken sera. Of the four types of antigen tested, crude antigen gave inconsistent results. However, an alcohol-precipitated antigen prepared from infectious allantoic-amniotic fluids was as satisfactory as more highly purified virus preparations. Other factors found to be extremely important were a 0.5M concentration of NaCl in the diluent and wash solutions used in the procedure, and a pH of 13 for sensitizing solution for maximum specific binding of the antigen to the microplate plastic wells. A comparison was made between the hemagglutination-inhibition (HI) titers of 550 known chicken sera and the corresponding ELISA values. Although the ELISA is much more sensitive than the HI test, there was a general but not a direct correlation between the two tests. The ELISA did not give more information than the HI test concerning protection against an NDV-induced drop in egg production. Preliminary observations indicated that this ELISA procedure is also applicable for turkey sera.
A novel disease entity in domestic geese with unusual epidemiology and pathological changes has emerged in some regions of China since the late 1990s. Velogenic Newcastle disease (ND) viruses were isolated from visceral organs and cloacal swabs of diseased birds in the field. Gross lesions of diseased geese were characterized by extensive necrotic foci in intestinal mucosa, spleen and pancreas. Histological examination revealed multisystemic distribution of lesions. Similar disease was reproduced experimentally in geese not only with two field isolates of goose origin, but also with ND viruses belonging to genotypes VI, VII, VIII and IX. These led us to define the novel disease as clinical ND in geese. Additionally, the goose-originated ND virus isolates could be transmitted from geese to specific pathogen free chickens, illustrating the potential threat they posed to the chicken industry. This is the first detailed report of the natural outbreaks and experimental reproduction of ND in geese, providing evidence that geese could play an important role in the epidemiology of ND.
The cell-fusing ability of Newcastle disease virus (NDV) was quantified using flow microfluorometry (FMF). The rate of polykaryocyte formation, fusion dependence on multiplicity of infection, and cell fusion differences for 21 NDV strains were measured using this technique. No correlation was found between the virulence of a virus strain and its cell fusion index calculated from the FMF data.
Newcastle disease virus (NDV) purified by sucrose density gradient centrifugation was chromatographed on a controlled-pore glass (CPG) bead column. By this procedure, the contaminating ribonuclease activity was reduced by 94-96% and the specific viral haemagglutinating activity increased from 3-to 5-fold. Purified NDV moved in two fractions in free electrophoresis.
Nucleotide sequence was determined for the RNA-dependent RNA polymerase (L) gene of 16 Newcastle disease virus (NDV) isolates from diverse geographic and chronological origins. The observed consensus amino acid sequence conformed to the six domains previously identified among paramyxovirus L proteins, and the putative 749QGDNQ753 active site was strictly conserved among all isolates. Analysis of predicted amino acid sequences allowed us to identify a sequencing error in the previously reported L genes for NDV. The correct sequences reported herein provided a more accurate alignment with predicted l-amino acid sequences of other paramyxoviruses. Comparison of L gene coding sequences among isolates revealed that synonymous substitutions dominated non-synonymous substitutions, as observed previously with other NDV genes. However, the overall substitution rate was lower than other genes examined making the L gene the most conserved of the NDV genome. Phylogenetic analysis of L genes among NDV isolates was consistent with previous results that suggested the existence of two major lineages. One group contained strains isolated in North America prior to 1970 and included virulent and vaccine strains, while the second group included virulent viruses isolated worldwide. A comparison of the NDV L coding sequences to other Paramyxoviridae illustrated the unique clustering of the avian-specific paramyxoviruses, further justifying the newly created Avulavirus genus.
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A total of 156 chickens in two villages in Malawi were marked and sampled. One hundred and fifteen of these were vaccinated against Newcastle disease immediately after blood sampling, using the V4 heat-resistant strain applied by eye-drop in one village and the inactivated Newcavac vaccine in the other village. A second blood sample was collected 4 weeks after vaccination. The samples were examined using an indirect ELISA test kit. The titre group median ranged from 2 to 3 before vaccination. Both vaccines led to a positive immune response. Newcavac induced higher and more homogeneous titres compared with the V4 vaccine. There was also an increase in the median of the control group where V4 live vaccine had been applied. The differences between the median titres induced by V4, Newcavac and controls were statistically significant.
The course of persistent infection in Newcastle disease virus (NDV)-infected mouse L cells (LNDV) was analysed. The production of infectious intra- and extracellular virus was markedly reduced (less than 10(1) PFU/ml), 30% of the cells produced virus-specific antigen. Analysis of the synthesis of 3H-nucleocapsid RNA in the LNDV system revealed predominance of low molecular weight RNAs. Virus-specific sequences were shown to be present in fraction of DNA extracted by Hirt's method. Only the "supernatant" Hirt's fraction was infectious. The possibility of the presence of virus-specific sequences in free unintegrated form is discussed.
Isolated envelope proteins of Newcastle disease virus (NDV) were inserted into the lipid bilayer of artificial vesicles to create a viral envelope-like structure. The structure-containing viral antigens at high density elicited a strong immune response, in contrast to purified viral proteins. The artificial envelopes or immunosomes possessed several advantageous properties when used as vaccines. They elicited a faster response and the immunity lasted longer in animals treated with these vesicles than in controls vaccinated with envelope proteins in combination with traditional adjuvants. A further useful feature of the artificial envelopes is that no inflammatory lesions develop at the site of their injection.
Experiments were conducted in chickens to understand the effects of oral immunomodulation. Heat inactivated M phlei, a commensal Mycobacterium and a non-specific immunomodulator, was administered orally prior to live Newcastle disease F (ND F) strain vaccination. In experimental birds it lead to an enhanced cell mediated Immune response (CMI) against the vaccine. There was a reduction in the Haemagglutination inhibiting (HI) antibodies. However, it did not affect the protection against a virulent challenge, as the protection percentage was more or less same in vaccinated birds irrespective of the M.phlei administration. M. phlei administration could not enhance the immune response to inactivated ND F vaccine administered orally. The results indicate that M. phlei favours a CMI response to orally administered live ND F vaccine. It may be of potential use in enhancing CMI against vaccines and a cheaper alternative to costlier recombinant cytokines.
Seventeen Australian strains of Newcastle disease virus were tested for their biological properties: mean death time, heat stability of the hemagglutinin and infectivity of the virus at 56 C, the elution time of virus from chicken erythrocytes, and the ability to hemagglutinate equine red blood cells. The strains differed considerably in their reactions. All had mean-death-time indices of 112 or greater, indicating that all were lentogenic. Strains were identified that had heat-labile and -stable hemagglutinin and infectivity, slow and fast elution, and variable ability to agglutinate equine erythrocytes. The significance of the results is discussed in terms of their usefulness in identifying exotic strains of the virus.
We have isolated a novel class of mutants of Newcastle disease virus which are less cytopathic than their virulent parent but are still capable of infectious virus production. Unlike wild-type virus, the mutants did not form plaques after 2 days of incubation; they did, however, make hemadsorbing spots. The mutants range in production of infectious virus from 10 to 200% of that of the wild type. They were less cytopathic in a single cycle of infection by light microscopy, loss of protein from the plate, and inhibition of total protein accumulation. All of the mutants exhibited extended mean embryo death times, a correlate of virulence in the adult animal.
Meat chickens on commercial broiler farms were vaccinated once at 1 to 15 days of age with a live V4 Newcastle disease virus (NDV) vaccine administered by drinking water, aerosol or coarse spray. Hatchmates were housed and similarly vaccinated in laboratory isolation pens. Samples of birds were bled at weekly to fortnightly intervals and the serums tested for haemagglutination inhibiting antibody to NDV. Log2 mean titres of up to 6.26, and assumed protection levels (based on the percentage of birds with log2 titres of 4 or greater) of up to 89%, were obtained in field trials within 4 weeks of vaccination. Differences were observed between the results obtained from parallel field and laboratory trials. The presence of maternal NDV antibody reduced the response to vaccination. The results show that this V4 vaccine can produce an adequate serological response following mass administration to Australian meat chickens housed under commercial conditions.
Recombinant hemagglutinin-neuraminidase (rHN) and fusion (rF) glycoproteins of virulent and avirulent strains of Newcastle disease virus (NDV) expressed by using baculovirus expression system were used to investigate their protective immunization effects in chickens. The efficacy of immunization with these recombinant proteins was evaluated by challenge infection. The chickens immunized with either rHN or rF protein of a virulent strain or rHN protein of an avirulent strain were completely protected from the lethal infection of virulent NDV. On the other hand, the rF protein of an avirulent strain, in which precursor F protein was not cleaved, showed lower protective effects. Significant levels of specific antibodies against respective proteins were detected in sera from survivors, whereas relatively lower levels of antibodies were found in chickens which were killed by challenge infection. These data indicate that either HN or F protein alone could induce protective immune responses and the cleavage of F protein might be important for its immunological potential.
Temperature-sensitive (ts) mutants of Newcastle disease virus (NDV) were isolated and studied for interferon (IFN) induction in primary chick embryo (CE) cells. At the non-permissive temperature (41 degrees C), there was no viral RNA synthesis or IFN induction by u.v.-treated virions except for ts-3 (RNA+), which did synthesize RNA at 41 degrees C, and whose u.v.-treated virions did induce IFN at this temperature. Another mutant (ts-4) induced IFN without irradiation, at the permissive temperature (37 degrees C). The minimum u.v. target size for IFN inducibility was unaffected by the mutation and corresponded to about 5% of the genome required for the expression of infectivity. These results support the hypothesis that the appearance of NDV RNA immediately after infection (primary transcription) plays a key role in IFN induction.