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

The evolution of virulent Vibrio cholerae It has been known since the 1990s that the cholera toxin genes in Vibrio cholerae are found in the integrated bacteriophage CTXΦ, located in the V. cholerae genome adjacent to toxin-linked cryptic (TLC), a chromosomal DNA element of unknown function. TLC is now shown to correspond to the genome of TLCΦ, a satellite filamentous phage that uses the morphogenesis genes of a third filamentous phage (fs2Φ) to form infectious particles. By reconstructing the events that lead to the acquisition of phage DNA and comparing these to the genome of pandemic strains, Hassan et al . obtain a model of how virulent V. cholerae strains evolve to become successful human pathogens. Bacterial chromosomes often carry integrated genetic elements (such as plasmids and prophages) that contribute to the evolutionary fitness of the host bacterium. In Vibrio cholerae , a prophage encodes cholera toxin. Here, the events that led to the acquisition of phage DNA have been reconstructed, revealing the cooperative interactions between multiple filamentous phages that contributed to the emergence of virulent V. cholerae strains. Bacterial chromosomes often carry integrated genetic elements (for example 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 cholerae 1 , is known to be adjacent to a chromosomally integrated element of unknown function termed the toxin-linked cryptic (TLC) 2 . Here we report the 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 recombinases 3 . The dif sequence is also exploited by lysogenic filamentous phages (for example CTXφ) for chromosomal integration of their genomes. Bacterial cells defective in the dimer resolution often show an aberrant filamentous cell morphology 3 , 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 with dif − filamentation phenotypes. Furthermore, lysogeny of a dif − non-toxigenic V. ch