Establishment of a rescue system for canine distemper virus

Canine distemper virus (CDV) has been rescued from a full-length cDNA done. Besides Measles virus (MV) and Rinderpest virus, a third morbillivirus is now available for genetic analysis using reverse genetics. A plasmid p(+)CD V was constructed by sequential cloning using the Onderstepoort vaccine str ain large-plaque-forming variant. The presence of a T7 promoter allowed tra nscription of full-length antigenomic RNA by a T7 RNA polymerase, which,vas provided by a host range mutant of vaccinia virus (MVA-T7). Plasmids expre ssing the nucleocapsid protein, the phosphoprotein, and the viral RNA-depen dent RNA polymerase, also under control of a T7 promoter, have been generat ed. Infection of HeLa cells with MVA-T7 and subsequent transfection of p(+) CDV plus the helper plasmids led to syncytium formation and release of infe ctious recombinant (r) CDV. Comparison of the rescued virus with the parent al virus revealed no major differences in the progression of infection or i n the shape and size of syncytia. A genetic tag, consisting of two nucleoti de changes within the coding region of the L protein, has been identified i n the rCDV genome. Expression by rCDV of all the major viral structural pro teins has been demonstrated by immunofluorescence.