Engineering a Temperate P. aeruginosa Bacteriophage to Carry an Inducible Toxin to Serve as an Antibacterial Agent

Bacteria are becoming resistant to antibiotic treatment, threatening doctors’ ability to successfully treat infections. This led researchers to discover alternative treatments like phage therapy, based on viruses infecting bacteria to treat bacterial infections. Pseudomonas aeruginosa regularly infects compromised individuals and is often the target of phage therapy. In our study, we attempt to create an engineered phage to carry a toxin for a more effective phage therapy treatment. We plan to use a "double kill" approach. First, the phage will infect the bacteria, next lysogens will secrete an effector via the T6SS to reduce surrounding WT bacteria, and lastly we will induce a "self-destruct" toxin to kill the lysogen.

First, we established the infection dynamics of four candidate phages in two P. aeruginosa strains. We then quantified what percentage of cells remain WT or become lysogens post-infection and what percentage of the WT population gained resistance to phages.

We next focused on toxins and their effect on various P. aeruginosa strains. We determined the toxicity of two genes over-expressed in four strains. We then created a mutant with a knockout of a Toxin-Immunity protein to determine if effectors encoded in a prophage can utilize the T6SS. The next stage of our study is to engineer the phage to carry the self-destruct toxin. Once we have established its efficacy we will add the T6SS effector.

We believe that this study can produce a highly effective tool in the ongoing battle with bacteria amidst the struggle with overcoming bacterial resistance to antibiotics.