Swapping of orthologous poxvirus intermediate transcription factor genes promotes adaptive evolution. Vaccinia virus WR strain:WR

In contrast to RNA viruses, large DNA viruses such as Vaccinia virus (VACV) have much lower mutation rates yet rapidly adapt to changing environment that makes them particularly suitable for experimental evolution studies. Until now, mutants with deletions of non-essential host range genes were used in experimental evolution of vaccinia virus (VACV). VACV in which the A8 or A23 intermediate transcription factor subunit gene was replaced by the myxoma (MYX) virus ortholog exhibited decreased replication. During three parallel serial passages of the hybrid virus with the MYXA8 gene in BSC-1 cells, plaque sizes and virus yields increased. DNA sequence analyses of virus populations after ten passages revealed high frequencies of five different single nucleotide mutations in the two largest RNA polymerase subunits RPO147 and RPO132, and two different Kozak consensus sequence mutations predicted to increase translation of the MYXA8 mRNA. Both, mutations in the viral RNA polymerase and translation-enhancing mutations, were sufficient to provide a fitness advantage compared to the original hybrid virus. This approach might be used to interrogating additional essential proteins of poxviruses as well as other viruses.