ENGINEERING E. COLI TO SENSE AND KILL VIBRIO CHOLERAE, HUMAN PATHOGEN CAUSING THE CHOLERA DISEASE
Chueh Loo Poh1, Maciej Holowko2, Prem Jayaraman2
1National University of Singapore, Singapore;
2Nanyang Technological University, Singapore
The cholera disease, caused by the Vibrio cholerae bacteria, is still claiming thousands of human life’s every year in developing countries, particularly in Asia and Africa. However, being recognised by WHO as a major threat to public healthcare it still lacks a therapy that would efficiently address the cause of the disease. Likewise, cholera preventive methods, including vaccines, are also characterized by low efficacy.
To address the problem, we are engineering a probiotic bacterium, Escherichia coli Nissle strain, which would be able to sense and kill V. cholerae in our human gastrointestinal tract. A key genetic module in such an engineered bacterium is the sensor which would be able to sense V. cholerae and produce a measurable output upon detecting V. cholerae. To this end, we have repurposed and rewired the native CAI-1 based quorum sensing pathway from V. cholerae in E. coli. We designed and constructed a synthetic genetic sensor to detect CAI-1, the quorum sensing molecules produced by V. cholerae. Results show that the sensor was able to produce detectable changes in the presence and absence of V. cholerae supernatant. As this quorum sensing pathway is activated in absence of V. cholerae and repressed in its presence, we have also developed and layered a CRISPRi based inverter to invert and amplify the signal. This enables the sensor to be activated in the presence of V. cholerae. This sensor would then be used to drive the expression of antimicrobial payload upon V. cholerae detection.