We showed previously that WKA (rat MHC, RT-1k) and DA (RT-1a) rat CTL specific for human T cell leukemia virus type-I (HTLV-I) recognized the gag and pX gene-encoded Ag. In the present study, we explored HTLV-I Ag recognized by CTL from other MHC genotype (RT-1l) rats, LEW and F344, and examined whether HTLV-I Ag expressed by recombinant vaccinia viruses (rVV) could prime WKA and LEW rats for HTLV-I-specific CTL. Upon priming in vivo and repetitive stimulation in vitro with HTLV-I+ syngeneic T cells, HTLV-I-specific CD8+ CTL were demonstrated in LEW and F344 rat spleen cell cultures. Interestingly, these CTL were directed against HTLV-I env and pX gene products. Immunization of LEW and WKA rats with the env and gag gene-expressing rVV, respectively, resulted in generation of HTLV-I-specific CD8+ CTL. However, a pX-gene expressing rVV failed to prime either rat strain. In addition, HTLV-I-specific CD8+ CTL from F1 hybrid (WKA x LEW) rats generated by immunization and restimulation with HTLV-I+ syngeneic T cells showed gag and pX Ag specificity on WKA rat cells and env and pX Ag specificity on LEW rat cells. These results suggest that immunogenicity of HTLV-I gag and env Ag in induction of HTLV-I-specific CTL varies depending on MHC, and that the pX Ag expressed by rVV is not a potent immunogen for rats.