The comparative analysis of the pathogenicity of a parental herpes simplex virus type 1 strain and its phosphonoacetic acid (PAA)-resistant and acyclovir (ACV)-resistant mutants showed marked differences among them. After orofacial skin inoculation of hairless mice the parental and PAA-resistant viruses were detected during the first 4 days after infection at high and increasing titers in the trigeminal ganglia; the ACV-resistant mutant was present at low and decreasing titers in the ganglia. Severe and slow-healing skin lesions were produced by the parental and PAA-resistant viruses; mild and rapidly healing lesions were produced by the ACV-resistant mutant. Virus titers in ganglia and the intensity of skin lesions were related to the virus dose used in the primary infection. Latent infections became established in trigeminal ganglia of mice inoculated with 10(6.0) plaque-forming units of the parental or PAA-resistant virus; no latent infections were detected in ganglia of mice inoculated with 10(7.0) plaque-forming units of the ACV-resistant mutant. Serum antibody titers attained similar values 4 weeks after primary infection with both mutants and the parental virus. Mice infected with the ACV-resistant mutant were reinfected with the parental and PAA-resistant viruses; the degree of protection against development of skin lesions, mortality, and latency was related to the dose of ACV-resistant virus used in the primary infection. Mortality was prevented by a dose of 10(6.0) plaque-forming units, skin lesions were prevented by a dose of 10(6.5) plaque-forming units, and latency was prevented by a dose of 10(7.0) plaque-forming units of the ACV-resistant mutant. Protection against reinfection with the PAA-resistant mutant was achieved with lower doses than protection against the parental virus. Serum antibody titers showed a 4- to 15-fold increase after reinfection. The results suggest that the ACV-resistant, latency-negative mutant has many attributes of a live attenuated herpes simplex virus vaccine.