Project Description

Since the discovery of penicillin in 1928 and the development of sulfa drugs in the 1930s, many classes of antibiotics have been developed to control and eradicate bacterial infections in humans, animals, and plants. These small molecule compounds interfere with specific pathways involved in the structure and function of the bacterial cell wall and membrane, DNA, RNA, protein, and folic acid synthesis. Subsequent to their widespread misuse and overuse, resistance and cross-resistance to these antibiotics has emerged and expanded over the decades so that drug resistant bacteria now pose a significant health risk. An alternative to conventional antibiotics is phage therapy. Investigated as a therapeutic since the 1920s, phage therapy research and utilization was diminished in the West where antibiotics were extensively used. Bacteriophages have host specificity and are either lytic or temperate in their life cycle. Lytic phage infect a bacteria, reproduce, and lyse the cell to release more phage. Temperate phages infect the bacterial host and may insert their genome into the host to be replicated and passed on to future generations without killing the host, a drawback for therapeutic applications.

Our approach to antibiotic resistance is to engineer a temperate phage, Lambda, with two distinct methods of killing: the phage’s lytic cycle and the light-activated production of superoxide by KillerRed expressed by the prophage. KillerRed is a phototoxic fluorescent protein which produces reactive oxygen species (ROS) when photobleached. Having two methods of killing decreases the probability of developing resistance. By engineering Lambda with KillerRed, our system overcomes the prior limitations of using wild-type temperate phages and provides a phage inactivation mechanism. Since approximately half of all phages are temperate and lysogeny is a defense mechanism, our system promotes localized cell death even under the condition of lysogeny. This project describes the characterization and modeling of our system.