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PubMed2005

Purification of recombinant nucleocapsid protein of Newcastle disease virus from unclarified feedstock using expanded bed adsorption chromatography.

In the present work, a single-step purification of recombinant nucleocapsid protein (NP) of the Newcastle disease virus (NDV) directly from unclarified feedstock using an expanded bed adsorption chromatography (EBAC) was developed. Streamline 25 column (ID = 25 mm) was used as a contactor and Streamline chelating adsorbent immobilized with Ni2+ ion was used as affinity adsorbent. The dynamic binding capacity of Ni2+ -loaded Streamline chelating adsorbent for the NP protein in unclarified feedstock was found to be 2.94 mg ml(-1) adsorbent at a superficial velocity of 200 cm h(-1). The direct purification of NP protein from unclarified feedstock using expanded bed adsorption has resulted in a 31% adsorption and 9.6% recovery of NP protein. The purity of the NP protein recovered was about 70% and the volume of processing fluid was reduced by a factor of 10. The results of the present study show that the IMA-EBAC developed could be used to combine the clarification, concentration and initial purification steps into a single-step operation.

Adsorption↗
PubMed1991

Detection of Newcastle disease virus RNA in infected allantoic fluids by in vitro enzymatic amplification (PCR).

The Polymerase Chain Reaction (PCR) procedure was applied in order to identify the Newcastle disease virus (NDV), an avian paramyxovirus (A-PMV 1). The sequence selected for amplification consists of 238 bp lying in the gene encoding the fusion protein F. A pair of 19-mer and 18-mer oligonucleotides, flanking this sequence, were used as primers. Following RNA extraction by the proteinase K method, a cDNA was prepared using the previous 19-mer oligonucleotide as the primer. The amplification reaction product was analyzed by electrophoresis and ethidium bromide staining, using the restriction enzymes HaeIII, Mbo II, and Nar I. The PCR was performed on cDNA prepared from 30 A-PMV 1 and 3 other strains (A-PMV2, A-PMV3, A-PMV4). It was thereby demonstrated that the selected sequence was highly specific and constant. However, two of the PMV1 strains isolated from feral ducks, are thought to present a deletion of about 25 bp inside this fragment as shown by the smaller length of the corresponding amplified product and the disappearance of the NarI restriction site. The advantages of this technique, as a first step in evaluating virulence by means of molecular biology, is discussed.

Allantois↗
PubMed1984

Monoclonal antibodies identify a strain-specific epitope on the HN glycoprotein of Newcastle disease virus strain Australia-Victoria.

A panel of monoclonal antibodies raised against the hemagglutinin-neuraminidase glycoprotein (HN) of the Australia-Victoria strain of Newcastle disease virus has been used to compare that strain and eight other strains of the virus. The ability of the antibodies to neutralize infectivity, inhibit hemagglutination and neuraminidase, and bind to purified virions in solid-phase radioimmunoassays was determined for each strain. Of the four antigenic sites delineated by these antibodies on the HN of the homologous strain, site 1 (that with the greatest neutralizing susceptibility), is apparently conserved in all the strains tested as revealed by neutralization assays. The least neutralizing site, number 4, is also conserved in most of the strains tested. Site 2, which lies at or near the neuraminidase site, appears to be conserved in the avirulent strains but not in the virulent strains. An antibody to site 3 is unable to bind to a significant extent to any of the heterologous strains tested, and thus recognizes a strain-specific epitope. Inhibition of hemagglutination and neuraminidase by antibodies to each site were also examined and the results suggest that antibodies to sites 1 and 2 may distinguish virulent and avirulent strains at least with respect to these functions.

Antibodies, Monoclonal↗
PubMed1951

The pathogenesis of infection with a virulent (CG 179) and an avirulent (B) strain of Newcastle disease virus in the chicken. I. Comparative rates of viral multiplication.

The comparative growth rates of a virulent (CG 179) and an avirulent (B) strain of NDV in the chicken were analyzed. Following intramuscular inoculation, the CG 179 and B strains both increased at the same rate in the extraneural tissues, i.e. the blood, lung, rectum, and spleen, but the CG 179 strain showed an accelerated growth rate in the brain. The CG 179 strain also multiplied more rapidly in the brain than the B strain following intracerebral administration of minimal inocula. Recovery with the B strain was associated with a decline in virus titer, first in the circulating blood, then in the visceral organs, and lastly in the central nervous system. Certain neuropathological observations were correlated with the pattern of virus growth.

Animals↗
PubMed2003

[Safety, stability and immunogenicity of an oral DNA vaccine against Newcastle disease].

Mice and 3-day-old chickens were orally inoculated with the recombinant attenuated Salmonella typhimurium strain ZJ111 carrying pcDNA3-F expression plasmid encoding the fusion protein of Newcastle disease virus (NDV). The results showed that ZJ111/pcDNA3-F was relatively safe. The recombinant plasmid pcDNA3-F was stable within the host stain ZJ111 in vitro and in vivo as shown by restriction enzyme analysis and PCR identification of the F gene. In an experimental vaccination study, 3-day-old chickens were orally immunized with ZJ111/pcDNA3-F with a dose of 108 cfu per chicken and boosted two weeks later. At week 4 post boosting, all chickens were challenged with a lethal dose of a virulent NDV strain F48 E9. The results showed that oral vaccination with ZJ111/pcDNA3-F induced stronger humoral and cellular immune responses than intramuscular immunization with naked pcDNA3-F plasmid. It also exhibited higher protection rate than the latter (66.7% vs 50%). This study indicates that the DNA vaccine using attenuated Salmonella typhimurium as delivery carrier had good safety, stability and immunogenicity and exhibited good potential of low cost and convenience for poultry disease control.

Animals↗