An outbreak of rubella affected 460 individuals in 2004 and 2005 in the community of Madrid, Spain. Most of the patients were nonvaccinated Latin American immigrants or Spanish males. This study presents the first data on rubella virus genotypes in Spain. Forty selected clinical samples (2 urine, 5 serum, 3 blood, 2 saliva, and 28 pharyngeal exudate samples) from 40 cases were collected. The 739-nucleotide sequence recommended by the World Health Organization obtained from viral RNA in these samples was analyzed by using the MEGA v4.0 software. Seventeen isolates were obtained from 40 clinical samples from the outbreak, including two isolated from congenital rubella syndrome cases. Only viral RNA of genotype 1j was detected in both isolates and clinical specimens. Two variations in amino acids, G253C and T394S, which are involved in neutralization epitopes arose during the outbreak, but apparently there was no positive selection of either of them. The origin of the outbreak remains unknown because of poor virologic surveillance in Latin America and the African countries neighboring Spain. On the other hand, this is the first report of this genotype in Europe. The few published sequences of genotype 1j indicate that it comes from Japan and the Philippines, but there are no epidemiological data supporting this as the origin of the Madrid outbreak.