The widely administered Mycobacterium bovis BCG is an attractive live vector for the development of AIDS vaccines. We explored immune responses induced in cynomolgus macaques tor BCG-SIV3, a mixture of three recombinant BCG strains expressing the SIVmac251 nef, gag and env genes. After a single intradermal (ID) inoculation, circulating blood cells from rBCG-SIV3-vaccinated monkeys exhibited CTL responses targeted against the three antigens and interferon-gamma (IFN-gamma) secretion was observed. A rectal or oral boosting dose of rBCG-SIV3 elicited anti-SIV IgAs in the rectum of vaccinated monkeys and increased IFNgamma secretion by circulating blood cells. Despite a good response against the vector, rBCG-SIV3 administration did not induce IgG antibody responses or lymphoproliferation against the SIV antigens in blood. This could be due to the lack of in vivo persistence of the recombinant BCG strains that were used. Rectal challenge with fully pathogenic SIVmac251-infected all animals. However, after viral challenge, anti-SIV cellular and antibody responses were higher in rBCG-SIV3 monkeys than in controls indicating that the vaccine induced anti-SIV CD4(+) T-cell memory. (C) 2003 Elsevier Ltd. All rights reserved.