A recombinant newcastle disease virus with low-level V protein expression is immunogenic and lacks pathogenicity for chicken embryos.
Newcastle disease virus (NDV) edits its P-gene mRNA by inserting a nontemplated G residue(s) at a conserved editing site (3'-UUUUUCCC-template strand). In the wild-type virus, three amino-coterminal P-gene-derived proteins, P, V, and W, are produced at frequencies of approximately 68, 29, and 2%, respectively. By applying the reverse genetics technique, editing-defective mutants were generated in cell culture. Compared to the wild-type virus, mutants lacking either six nucleotides of the conserved editing site or the unique C-terminal part of the V protein produced as much as 5, 000-fold fewer infectious progeny in vitro or 200,000-fold fewer in 6-day-old embryonated chicken eggs. In addition, both mutants were unable to propagate in 9- to 11-day-old embryonated specific-pathogen-free (SPF) chicken eggs. In contrast, a mutant (NDV-P1) with one nucleotide substitution (UUCUUCCC) grew in eggs, albeit with a 100-fold-lower infectious titer than the parent virus. The modification in the first two mutants described above led to complete abolition of V expression, whereas in NDV-P1 the editing frequency was reduced to less than 2%, and as a result, V was expressed at a 20-fold-lower level. NDV-P1 showed markedly attenuated pathogenicity for SPF chicken embryos, unlike currently available ND vaccine strains. These findings indicate that the V protein of NDV has a dual function, playing a direct role in virus replication as well as serving as a virulence factor. Administration of NDV-P1 to 18-day-old embryonated chicken eggs hardly affected hatchability. Hatched chickens developed high levels of NDV-specific antibodies and were fully protected against lethal challenge, demonstrating the potential use of editing-defective recombinant NDV as a safe embryo vaccine.
Mebatsion
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Department of Virology, Intervet International B.V., 5830 AA Boxmeer, The Netherlands. teshome.mebatsion@intervet.com
Verstegen
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De Vaan
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Römer-Oberdörfer
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Schrier
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D000818:Animals; D002642:Chick Embryo; D018076:DNA, Complementary / Q000096:biosynthesis; D009154:Mutation; D009522:Newcastle disease virus / Q000235:genetics / Q000276:immunology* / Q000472:pathogenicity; D017393:RNA Editing; D014611:Vaccination; D014613:Vaccines, Attenuated / Q000276:immunology; D014614:Vaccines, Synthetic / Q000276:immunology*; D015678:Viral Structural Proteins / Q000502:physiology*; D014765:Viral Vaccines / Q000276:immunology*; D014779:Virus Replication
D018076:DNA, Complementary; C097019:V protein, Simian parainfluenza virus 5; D014613:Vaccines, Attenuated; D014614:Vaccines, Synthetic; D015678:Viral Structural Proteins; D014765:Viral Vaccines