Engineering novel beneficial therapeutic microorganisms for Vibrio cholerae therapy using synthetic biology

• Main Author: Holowko, Maciej Bartosz
• Other Authors: Poh Chueh Loo
• Format: Theses and Dissertations
• Language: English
• Published: 2017
• Subjects: DRNTU::Engineering::Chemical engineering::Biotechnology
• Online Access: http://hdl.handle.net/10356/72578
• Institution: Nanyang Technological University

Cholera disease is still a major global threat according to WHO. This notion is reinforced by the fact that the antibiotic resistance of Vibrio cholerae, the causative agent of the disease, is increasing in the epidemic strains. In year 2016 alone numerous occurrences of cholera outbreaks were reported, e.g. in Haiti and Sudan. The currently available methods of detection, treatment and prevention of cholera lack the required characteristics. Detection is slow or costly, treatment is mostly aimed at the symptoms and prevention in forms of vaccines is ineffective. To mitigate some of these problems, the goal of this research project is to engineer novel beneficial probiotic microbes to fight V. cholerae infections. To this end, a set of synthetic genetic circuits were created using the tools of synthetic biology. These circuits aim was to create a genetically modified Escherichia coli able to sense and kill V. cholerae. First, a whole cell E. coli based biosensor for V. cholerae detection has been designed, constructed and characterized. Second, this sensor circuit was redesigned to create sense and kill mechanism. Artilysin, a recently developed antimicrobial agent that can be directly expressed by cells was shown to be effective against V. cholerae cells. This work is first both to show a working V. cholerae biosensor in E. coli and an E. coli strain that is able to destroy V. cholerae upon its detection. Main advantage of this approach is that it should be possible not only to mitigate the cholera symptoms, but also to eliminate the cause of the disease, which was not possible before. This study is also a showcase of design principles for synthetic biology with examples of novel circuits with new, previously unknown applications and combinations. Finally, this integrated system forms a basis for a further development of viable detection and therapy option for V. cholerae infection.