By selectively regulating the expression of the trans-dominant-negative mutant polypeptide UL9-C535C, of herpes simplex virus type 1 (HSV-1) origin binding protein UL9 with the tetracycline repressor (tetR)-mediated gene switch, we recently generated a novel replication-defective and anti-HSV-specific HSV-1 recombinant, CJ83193. The UL9-C535C peptides expressed by CJ83193 can function as a potent intracellular therapy against its own replication, as well as the replication of wild-type HSV-1 and HSV-2 in coinfected cells. In this report, we demonstrate that CJ83193 cannot initiate acute productive infection in corneas of infected mice nor can it reactivate from trigeminal ganglia of mice latently infected by CJ83193 in a mouse ocular model. Given that CJ83193 is capable of expressing the viral ␣, , and ␥1 genes but little or no ␥2 genes, we tested the vaccine potential of CJ83193 against HSV-1 infection in a mouse ocular model. Our studies showed that immunization with CJ83193 significantly reduced the yields of challenge HSV in the eyes and trigeminal ganglia on days 3, 5, and 7 postchallenge. Like in mice immunized with the wild-type HSV-1 strain KOS, immunization of mice with CJ83193 prevents the development of keratitis and encephalitis induced by corneal challenge with wild-type HSV-1 strain mP. Delayed-type hypersensitivity (DTH) assays demonstrate that CJ83193 can elicit durable cell-mediated immunity at the same level as that of wild-type HSV-1 and is more effective than that induced by d27, an HSV-1 ICP27 deletion mutant. Moreover, mice immunized with CJ83193 developed strong, durable HSV-1-neutralizing antibodies at levels at least twofold higher than those induced by d27. The results presented in this report have shed new light on the development of effective HSV viral vaccines that encode a unique safety mechanism capable of inhibiting the mutant's own replication and that of wild-type virus.Herpesviruses can cause both an acute productive infection and a long-term latent infection characterized by periodic recurrences (54). Among the human herpesviruses, herpes simplex virus types 1 and type 2 (HSV-1 and -2, respectively) are most closely related and share significant DNA sequence homology (15,31,32). Although HSV infections are often asymptomatic, their clinical manifestations include oral-facial infections, genital herpes, neonatal herpes, keratoconjunctivitis, and herpes encephalitis (48,54). Despite the availability of various effective antiviral therapies, the spread of genital herpes infection continues to increase worldwide and has increased by 30% in the United States in the past two decades (19). The failure of antiviral therapy in effectively controlling the spread of HSV disease and the life-long nature of the infection present a strong need for developing safe and efficacious vaccines against HSV infections (48, 54).HSV replicates in epithelial cells and establishes life-long latent infection in neuronal cell bodies within the sensory ganglia of infected individuals. During productive i...