GOLDSCHMIDT AWARD LECTURE

License to Kill: Bioengineering a bacterium with Antimicrobial Activity via the Exploitation of Toxin-antitoxin Systems and Conjugation

Mary Girard (2012 Goldschmidt Female Graduate Student Scholarship Recipient)

Mary’s mentor is Dr. Christophe Herman in the Departments of Molecular Virology & Microbiology and Molecular & Human Genetics, Baylor College of Medicine, Houston, TX

With the rise in antibiotic resistant bacterial infections, we are in need of new antimicrobial methods.  One potential alternative is to exploit the natural bacterial toxin-antitoxin (TA) systems. TA systems are  bacterial operons which encode a stable toxin and an unstable corresponding  antitoxin. Toxin-antitoxin gene modules are always co-expressed to alleviate toxicity as the toxin is detrimental and will kill or inhibit growth of the bacterium when the antitoxin is depleted. The role of TA systems in bacterial physiology remains unclear; one possible role for TA systems is maintenance and proper segregation of large, low-copy plasmids. If errors occur in plasmid segregation during division, the daughter cell which lacks a TA-encoding plasmid will succumb to the toxin. Accordingly, several TA systems have beenfound on conjugative plasmids, which are transferred through a sex pilus from the donor to the recipient in a process known as conjugation.

This project aims to investigate a novel antimicrobial approach that exploits conjugation and TA systems to engineer a bacterium that delivers toxins by conjugation. This strategy relies on uncoupling toxin/antitoxin gene expression by placing the toxin gene on the conjugative plasmid and the antitoxin gene on the chromosome. Upon transfer of the plasmid, only the toxins will be expressed, killing the recipient bacterium. We aim to determine whether conjugative transfer of a plasmid encoding multiple toxins will kill recipient bacteria and how this toxic effect can be targeted to specific bacterial pathogens. We are initially testing this antimicrobial strategy by performing mating assays between the engineered strain carrying a single toxin and non-pathogenic laboratory strains of E. coli, scoring for cell viability.  Preliminary data shows that this method effectively kills recipient E. coli in both planktonic and biofilm  states. Next, we will be testing different combinations of toxins to determine the effect of multiple toxins on cell viability. In the future, mating and viability assays will also be performed with a panel of bacterial pathogens. Ultimately, this work may lead to the development of this system for application in biotherapeutics or biocontainment.