Efficient recovery of attenuated canine distemper virus from cDNA

To provide insights into the biology of the attenuated canine distemper virus (CDV) Onderstepoort (OP) strain (large plaque forming variant), design next-generation multivalent vaccines, or further investigate its promising potential as an oncolytic vector, we employed contemporary modifications to establish an efficient OP-CDV-based reverse genetics platform. Successful viral rescue was obtained however only upon recovery of a completely conserved charged residue (V13E) residing at the N-terminal region of the large protein (L). Although L-V13 and L-V13E did not display drastic differences in cellular localization and physical interaction with P, efficient polymerase complex (P+ L) activity was recorded only with L-V13E. Interestingly, grafting mNeonGreen to the viral N protein via a P2A ribosomal skipping sequence (OP^neon) and its derivative V-protein-knockout variant (OP^neon-V^ko) exhibited delayed replication kinetics in cultured cells. Collectively, we established an efficient OP-CDV-based reverse genetics system that enables the design of various strategies potentially contributing to veterinary medicine and research.