The identification of "asymptomatic" (i.e., protective) epitopes recognized by T cells from herpes simplex virus (HSV)-seropositive healthy individuals is a prerequisite for an effective vaccine. Using the PepScan epitope mapping strategy, a library of 179 potential peptide epitopes (15-mers overlapping by 10 amino acids) was identified from HSV type 1 (HSV-1) glycoprotein B (gB), an antigen that induces protective immunity in both animal models and humans. Eighteen groups (G1 to G18) of 10 adjacent peptides each were first screened for T-cell antigenicity in 38 HSV-1-seropositive but HSV-2-seronegative individuals. Individual peptides within the two immunodominant groups (i.e., G4 and G14) were further screened with T cells from HLA-DRgenotyped and clinically defined symptomatic (n ؍ 10) and asymptomatic (n ؍ 10) HSV-1-seropositive healthy individuals. Peptides gB [161][162][163][164][165][166][167][168][169][170][171][172][173][174][175] ؉ T cells from symptomatic patients preferentially recognized gB 661-675 (P < 0.0001). Thus, we identified three previously unrecognized CD4؉ CTL peptide epitopes in HSV-1 gB. Among these, gB 166-180 and gB 666-680 appear to be "asymptomatic" peptide epitopes and therefore should be considered in the design of future herpes vaccines.Herpes simplex virus types 1 and 2 (HSV-1 and HSV-2) are ubiquitous viruses that infect a majority of people worldwide (3,22,68). Shedding of reactivated HSV is estimated to occur at rates of 3 to 28% in adults who harbor latent HSV in their sensory neurons (32,(66)(67)(68). However, the vast majority of these individuals do not experience recurrent herpetic disease and are designated "asymptomatic patients" (22,40,68). In contrast, in some individuals (symptomatic patients), reactivation of latent virus leads to induction of "pathogenic" HSVspecific CD4 ϩ and CD8 ϩ T cells (22, 68) and recurrent disease, ranging from rare episodes occurring every 5 to 10 years to outbreaks occurring monthly or even more frequently among a small proportion of subjects (22,26,60). Interestingly, for genital herpes, symptomatic and asymptomatic patients have similar virus shedding rates (68). It is likely that the same is true for ocular and oro-facial herpes, since shedding rates in tears and saliva of asymptomatic individuals have been reported to be as high as 33.5% (22,29,40,42). Latently infected patients are at risk of developing severe immunopathology, such as blinding herpetic stromal keratitis (HSK) (primarily due to HSV-1), painful genital ulcerations (primarily due to HSV-2), and in rare cases, fatal HSV encephalitis (reviewed in reference 72).Considering the wealth of data addressing the role of T cells in animal models, it is surprising how few reports explore the immunologic basis of symptomatic and asymptomatic HSV infections in humans. The HSV-1-specific CD4 ϩ T-cell responses in the cornea are much more likely to cause pathology than those in the genital tract or anus/buttocks region. Indeed, the involvement of CD4 ϩ T cells that produce Th1 cyt...