An influence of cytotoxic T lymphocyte (CTL) response over Epstein-Barr virus (EBV) evolution was first suggested by the finding that virus isolates from highly HLA-A11-positive Oriental populations were specifically mutated in two immunodominant A11-restricted CTL epitopes. Here we turn to a second HLA allele, B35.01 and show that B35.01-restricted CTL responses in Caucasian donors reproducibly map to a single peptide epitope, YPLHEQHGM, representing residues 458-466 of the type 1 EBV nuclear antigen 3A protein (B95.8 strain). In this case, however, most EBV isolates from a highly B35.01-positive population (in The Gambia) either retained the CTL epitope sequence or carried a mutation (P-->S at position 2) which conserved antigenicity; changes leading to reduced antigenicity (Y-->N at position 1) were found in only a minority of cases. Furthermore, CTL recognizing the YPLHEQHGM epitope could be reactivated from the blood of some B35.01-positive Gambian donors by in vitro stimulation with the synthetic peptide, indicating that epitope-specific immunity does exist in this population. Possible differences between the A11-based and B35.01-based studies are discussed.