Satellite phage TLCϕ enables toxigenic conversion by CTX phage through dif site alteration

Bacterial chromosomes often carry integrated genetic elements (e.g., plasmids, transposons, prophages, and islands) whose precise function and contribution to the evolutionary fitness of the host bacterium are unknown. The CTXϕ prophage, which encodes cholera toxin in Vibrio cholerae1, is known to be adjacent to a chromosomally integrated element of unknown function termed the toxin-linked cryptic (TLC)2. Here we report characterization of a TLC-related element that corresponds to the genome of a satellite filamentous phage (TLC-Knϕ1) which uses the morphogenesis genes of another filamentous phage (fs2ϕ) to form infectious TLC-Knϕ1 phage particles. The TLC-Knϕ1 phage genome carries a sequence similar to the dif recombination sequence which functions in chromosome dimer resolution using XerC and XerD recombinases3. The dif sequence is also exploited by lysogenic filamentous phages (e.g., CTXϕ) for chromosomal integration of their genomes. Bacterial cells defective in the dimer resolution often show an aberrant filamentous cell morphology3,4. We found that acquisition and chromosomal integration of the TLC-Knϕ1 genome restored a perfect dif site and normal morphology to V. cholerae wild type and mutant strains that displayed dif(-) filamentation phenotypes. Furthermore, lysogeny of a dif(-) nontoxigenic V. cholerae with TLC-Knϕ1 promoted its subsequent toxigenic conversion through integration of CTXϕ into the restored dif site. These results reveal a remarkable level of cooperative interactions between multiple filamentous phages in the emergence of the bacterial pathogen that causes cholera.