Generation of a prophagefree variant of the fast-growing bacterium Vibrio natriegens

The fast-growing marine bacterium Vibrio natriegens represents an emerging strain for molecular biology and biotechnology. Genome sequencing and quantitative PCR analysis revealed that the first chromosome of V. natriegens ATCC 14048 contains two prophage regions (VNP1 and VNP2) that are both inducible by the DNA-damaging agent mitomycin C and exhibit spontaneous activation under standard cultivation conditions. Their activation was also confirmed by live cell imaging of an mCherry fusion to the major capsid proteins of VNP1 and VNP2. Transmission electron microscopy visualized the release of phage particles belonging to the Siphoviridae family into the culture supernatant. Freeing V. natriegens from its proviral load, followed by phenotypic characterization, revealed an improved robustness of the prophage-free variant toward DNA-damaging conditions, reduced cell lysis under hypo-osmotic conditions, and an increased pyruvate production compared to wild-type levels. Remarkably, the prophage-free strain outcompeted the wild type in a competitive growth experiment, emphasizing that this strain is a promising platform for future metabolic engineering approaches.