Deletion of Major Nonessential Genomic Regions in the Vaccinia Virus Lister Strain Enhances Attenuation without Altering Vaccine Efficacy in Mice

The vaccinia virus (VACV) Lister strain was one of the vaccine strains that enabled smallpox eradication. Although the strain is most often harmless, there have been numerous incidents of mild to life-threatening accidents with this strain and others. In an attempt to further attenuate the Lister strain, we investigated the role of 5 genomic regions known to be deleted in the modified VACV Ankara (MVA) genome in virulence in immunodeficient mice, immunogenicity in immunocompetent mice, and vaccine efficacy in a cowpox virus challenge model. Lister mutants were constructed so as to delete each of the 5 regions or various combinations of these regions. All of the mutants replicated efficiently in tissue culture except region I mutants, which multiplied more poorly in human cells than the parental strain. Mutants with single deletions were not attenuated or only moderately so in athymic nude mice. Mutants with multiple deletions were more highly attenuated than those with single deletions. Deleting regions II, III, and V together resulted in total attenuation for nude mice and partial attenuation for SCID mice. In immunocompetent mice, the Lister deletion mutants induced VACV specific humoral responses equivalent to those of the parental strain but in some cases lower cell-mediated immune responses. All of the highly attenuated mutants protected mice from a severe cowpox virus challenge at low vaccine doses. The data suggest that several of the Lister mutants combining multiple deletions could be used in smallpox vaccination or as live virus vectors at doses equivalent to those used for the traditional vaccine while displaying increased safety.