Rescuing eGFP-Tagged Canine Distemper Virus for 40 Serial Passages Separately in Ribavirin- and Non-Treated Cells:...

Area of Science:

• Virology
• Molecular Biology
• Genetics

Background:

• RNA viruses exhibit high mutation rates due to error-prone RNA-dependent RNA polymerases (RdRp).
• Canine distemper virus (CDV), a Morbillivirus, replicates via its L protein acting as an RdRp.
• Viral quasispecies evolve rapidly, especially under mutagenic pressure like ribavirin treatment.

Purpose of the Study:

• To investigate the mutagenic effect of ribavirin on CDV replication.
• To compare the quasispecies diversity of ribavirin-treated and untreated CDV.
• To explore potential high-fidelity replication mechanisms in CDV.

Main Methods:

• Rescue of recombinant enhanced green fluorescence protein-tagged CDV (rCDV-eGFP).
• Serial passage of rCDV-eGFP in VDS cells, with and without ribavirin, for 40 passages.
• Next-generation sequencing (NGS) of viral progenies (rCDV-eGFP-R and rCDV-eGFP-N) at passage 47.

Main Results:

• Ribavirin-treated CDV (rCDV-eGFP-R) accumulated 62 single-nucleotide mutations (SNMs), compared to 23 in untreated CDV (rCDV-eGFP-N).
• The majority of mutations in rCDV-eGFP-R were missense (26) and silent (36).
• No nonsense mutations were observed in either progeny.

Conclusions:

• Ribavirin demonstrably increases the mutation frequency of CDV.
• The rCDV-eGFP-R progeny may harbor sub-progenies with high-fidelity replication capabilities.
• Purification of such sub-progenies could elucidate CDV's high-fidelity replication mechanisms via its L protein.