AN AVIRULENT ICP34.5 DELETION MUTANT OF HERPES-SIMPLEX VIRUS TYPE-1 IS CAPABLE OF IN-VIVO SPONTANEOUS REACTIVATION

The herpes simplex virus type 1 (HSV-1) ICP34.5 gene is a neurovirulen ce gene in mice. In addition, some ICP34.5 mutants have been reported to have a reduced efficiency of induced reactivation as measured by in vitro explantation of latently infected mouse ganglia. However, since spontaneous: reactivation is almost nonexistent in mice, nothing has been reported on the effect of ICP34.5 mutants on spontaneous reactiva tion in vivo. To examine this, we have deleted both copies of the ICP3 4.5 neurovirulence gene from a strain of HSV-1 (McKrae) that has a hig h spontaneous reactivation rate in rabbits and used this mutant to inf ect rabbit eyes. All rabbits infected with the ICP34.5 mutant virus (d 34.5) survived, even at challenge doses greater than 4 ic 10(7) PFU pe r eye. In contrast, a 200 fold-lower challenge dose of 2 x 10(5) PFU p er eye was lethal for approximately 50% of rabbits infected with eithe r the wild-type McKrae parental virus or a rescued ICP34.5 mutant in w hich both copies of the ICP34.5 gene were restored. In mice, the 50% l ethal dose of the ICP34.5 mutant was over 10(6) PFU, compared with a v alue of less than 10 PFU for the rescued virus. The ICP34.5 mutant was restricted for replication in rabbit and mouse eyes and mouse trigemi nal ganglia in vivo. The spontaneous reactivation rate in rabbits for the mutant was 1.4% as determined by culturing tear films for the pres ence of reactivated virus. This was more than 10-fold lower than the s pontaneous reactivation rate determined for the rescued virus (19.6%) and was highly significant (P < 0.0001, Fisher exact test). Southern a nalysis confirmed that the reactivated virus retained both copies of t he ICP34.5 deletion. Thus, this report demonstrates that (i) the ICP34 .5 gene, known to be a neurovirulence gene in mice, is also important for virulence in rabbits and (ii) in vivo spontaneous reactivation of HSV-1 in the rabbit ocular model, although reduced, can occur in the a bsence of the ICP34.5 gene.