So far, there is only fragmentary and unconfirmed information on bacteriophages infecting the genus Bifidobacterium. In this report we analyzed three prophage-like elements that are present in the genomes of Bifidobacterium breve UCC 2003, Bifidobacterium longum NCC 2705, and Bifidobacterium longum DJO10A, designated Bbr-1, Bl-1, and Blj-1, respectively. These prophagelike elements exhibit homology with genes of double-stranded DNA bacteriophages spanning a broad phylogenetic range of host bacteria and are surprisingly closely related to bacteriophages infecting low-G+C bacteria. All three prophage-like elements are integrated in a tRNA(Met) gene, which appears to be reconstructed following phage integration. Analysis of the distribution of this integration site in many bifidobacterial species revealed that the attB sites are well conserved. The Blj-1 prophage is 36.9 kb long and was induced when a B. longum DJO10A culture was exposed to mitomycin C or hydrogen peroxide. The Bbr-1 prophage-like element appears to consist of a noninducible 28.5-kb chimeric DNA fragment composed of a composite mobile element inserted into prophage-like sequences, which do not appear to be widely distributed among B. breve strains. Northern blot analysis of the Bbr-1 prophage-like element showed that large parts of its genome are transcriptionally silent. Interestingly, a gene predicted to encode an extracellular beta-glucosidase carried within the Bbr-1 prophage-like element was shown to be transcribed.