Tail-Engineered Phage P2 Enables Delivery of Antimicrobials into Multiple Gut Pathogens

Abstract

Bacteriophages can be reprogrammed to deliver antimicrobials for therapeutic and biocontrol purposes and are a promising alternative treatment to antimicrobial-resistant bacteria. Here, we developed a bacteriophage P4 cosmid system for the delivery of a Cas9 antimicrobial into clinically relevant human gut pathogens <i>Shigella flexneri</i> and <i>Escherichia coli</i> O157:H7. Our P4 cosmid design produces a high titer of cosmid-transducing units without contamination by a helper phage. Further, we demonstrate that genetic engineering of the phage tail fiber improves the transduction efficiency of cosmid DNA in <i>S. flexneri</i> M90T as well as allows recognition of a nonnative host, <i>E. coli</i> O157:H7. We show that the transducing units with the chimeric tails enhanced the overall Cas9-mediated killing of both pathogens. This study demonstrates the potential of our P4 <i>cas9</i> cosmid system as a DNA sequence-specific antimicrobial against clinically relevant gut pathogenic bacteria.