Browsing by Subject "cell-free expression"

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    ToxIN-mediated resistance to and cell-free production of Salmonella bacteriophages
    (2023-05) McFarlane, John
    Salmonella remains a leading cause of foodborne illnesses in the U.S. and worldwide. Though numerous strategies are implemented to control foodborne Salmonella, many come with limitations that reduce their efficacy or restrict which foods can be treated. An alternative control strategy that addresses these limitations is the application of bacteriophage cocktails, consisting of numerous bacterial viruses which kill the target pathogen. However, the use of phages also comes with two significant drawbacks: phage resistance and manufacturing hurdles. Resistance to phages may reduce the efficacy of phage cocktails, while concerns with at-scale phage production include handling of pathogenic host bacteria and optimization of numerous, complex biological variables. In the first part of this work, I show that ToxIN, a resistance system which aborts phage infection, is present in outbreak strains of Salmonella; confers resistance to the broad host range phage FelixO1 and other phage isolates; and is found in the whole-genome sequences of numerous Salmonella serovars and in other human pathogens. The second half of this work describes the in vitro synthesis of FelixO1 using an Escherichia coli-based cell-free expression system, demonstrating for the first time that a Salmonella phage can be produced in this manner. Taken together, this work illustrates that ToxIN may be of concern for food applications of phages while also establishing an important milestone in advancing Salmonella biocontrol using phages.