Two groups of 3 rabbits each were immunized with either recombinant vaccinia virus, WR-SFB5env, carrying the human T-cell lymphotropic virus type I (HTLV-I) env gene at the site of the hemagglutinin gene of the WR strain, or control vaccinia virus, HA-WR, lacking the functional hemagglutinin gene. All 6 rabbits responded with anti-vaccinia virus antibodies. WR-SFB5env elicited anti-HTLV-I env antibodies but no vesicular stomatitis virus (HTLV-I) pseudotype neutralizing antibodies in all 3 rabbits. After 10 weeks, the animals were challenged by transfusion of blood from an HTLV-I-infected rabbit. Two of the 3 vaccinated rabbits and all 3 control rabbits became infected with HTLV-I, as indicated by seroconversion and detection of HTLV-I proviral sequences by polymerase chain reaction. The rabbit that had been protected from initial challenge became infected with HTLV-I upon rechallenge 12 weeks after the first challenge. In view of the proven prophylactic effect of passive immunization against HTLV-I, our vaccine trial failed because WR-SFB5env was incapable of inducing neutralizing antibodies against HTLV-I in the immunized animals. It remains to be studied whether cell-mediated immunity such as antibody-dependent cellular cytotoxicity was involved in the temporary protection of I vaccinated rabbit.