Developing a Broad-Host-Range Modular Microcin Secretion System to Protect Plants from Bacterial Pathogens

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2024

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Agricultural bacterial pathogens present a major threat to the global food supply. New methods of controlling these pathogens are badly needed, as existing methods such as applying chemical pesticides or razing contaminated crops are not sustainable. Though traditional small-molecule antibiotics have received some use in this capacity, they are hardly ideal for wide-scale application due to the ever-looming threat of antibiotic resistance. One emerging solution has been the use of biocontrol strains, beneficial bacteria that naturally kill or otherwise inhibit pathogenic strains. However, the search for new biocontrol strains relies on serendipitously finding bacteria that naturally have anti-pathogen properties. To expedite the production of new biocontrol strains, I helped develop a pipeline for engineering biocontrol strains to secrete antimicrobial peptides known as microcins. Characterized microcins have a greater specificity than other types of antibiotics, primarily targeting close phylogenetic relatives of the bacteria that naturally produce them. In order to rapidly engineer microcin-secreting biocontrol strains, I refactored a two-plasmid microcin secretion system to increase its modularity. Promoters, microcin coding sequences, signal peptides, replication origins, and even secretion systems can be interchanged in this design, allowing for an array of permutations to the system according to the user’s needs. I show that my refactored system can secrete a known microcin, inhibiting growth of its target bacterium, and I map out how broad-host-range plasmid versions of my system can be used to engineer Pseudomonas and Pantoea biocontrol strains to secrete microcins. In the future, this system can be used to test novel microcins against a multitude of pathogen targets and deploy them in diverse bacterial biocontrol chassis to protect susceptible crops.

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