Rapid pathotyping of Newcastle disease virus (NDV) using fluorogenic probes in a PCR assay.
Hybridisation of PCR fragments with fluorogenic probes specific for pathotype allowed an estimation of pathogenicity of Newcastle disease virus (NDV) isolates using a modified TaqMan procedure. Six probes were used, designed to recognise nucleotide sequences in the fusion protein gene sequence corresponding to the precursor protein F0 cleavage site of both virulent and avirulent viruses. Forty-three of the 45 isolates tested, including 18 examined in a blind study were pathotyped successfully and rapidly, with close correlation between cleavage site nucleotide sequences, TaqMan results and intracerebral pathogenicity index (ICPI) values. One isolate, which could not be pathotyped by nucleotide sequencing, was shown using the TaqMan system to be a mixture of virulent and avirulent NDV. The results of this study suggest that using this modified TaqMan protocol, the likely virulence of most ND isolates can be determined rapidly and reproducibly.
Aldous
E W
EW
Virology Department, Central Veterinary Laboratories Agency, Weybridge, New Haw, Addlestone, Surrey KT15 3NB, UK. e.aldous@vla.maff.gov.uk
Collins
M S
MS
McGoldrick
A
A
Alexander
D J
DJ
D000818:Animals; D002642:Chick Embryo; D002645:Chickens / Q000821:virology*; D005456:Fluorescent Dyes / Q000737:chemistry; D009521:Newcastle Disease; D009522:Newcastle disease virus / Q000737:chemistry / Q000235:genetics / Q000472:pathogenicity*; D011201:Poultry Diseases / Q000175:diagnosis / Q000821:virology; D012367:RNA, Viral / Q000737:chemistry / Q000302:isolation & purification; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000662:veterinary; D017422:Sequence Analysis, DNA; D013047:Specific Pathogen-Free Organisms; D013050:Spectrometry, Fluorescence / Q000662:veterinary; D014774:Virulence
D005456:Fluorescent Dyes; D012367:RNA, Viral