DETECTION OF NEWCASTLE-DISEASE VIRUS IN POULTRY VACCINES USING THE POLYMERASE CHAIN-REACTION AND DIRECT SEQUENCING OF AMPLIFIED CDNA

In order to develop an in vitro method for the control of poultry vacc ines for identity and the absence of extraneous agents, reverse transc ription-polymerase chain reaction (RT-PCR) was applied for the detecti on of Newcastle disease virus (NDV). NDV vaccines were employed for th e establishment of the method, using two primer pairs spanning the cle avage site of the FO fusion protein coding sequence. Amplification of a specific cDNA segment was possible from live and inactivated oil-adj uvanted NDV vaccines without prior treatment. The cDNA was characteriz ed by restriction endonuclease digestion as well as by dir ect nucleot ide sequencing. The RT-PCR was able to detect between 5 x 10(2) EID(50 ) (in live vaccine preparations) and 10(5) EID(50) or 0.056 haemagglut inating units of NDV (in inactivated vaccine preparations). In additio n, live vaccine preparations were inactivated with beta-propiolactone (beta-PL). Amplified cDNA was obtained after treatment with 0.1% beta- PL, whereas at a concentration of 1% or 10% no specific bands were vis ible in the agarose gel. These results demonstrate the applicability o f the method for the control of poultry vaccines for identity and for the absence of extraneous agents, and additionally allow a rapid chara cterization of the respective NDV strain.