Development of Engineered Native Bacteria as a Tool for Functional Manipulation of the Gut Microbiome

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PROJECT SUMMARY/ABSTRACT Most therapies that target microbiome composition do not have a detectable impact on the gut microbiome and are not robust to the interpersonal diversity and plasticity of the community in human hosts. To develop a better mechanistic understanding of the microbe-host relationship and more effective microbiome-mediated therapies, approaches based on functional modulation of the gut microbiome are necessary. However, these approaches have been difficult to develop. Attaining long-term engraftment in the luminal environment has proven to be quite difficult, and even once engraftment has been achieved, a change in physiology or improvement of pathologic phenotype has not yet been demonstrated. There is a critical need for a tool that will allow investigators to “knock-in” functions into the gut microbiome and investigate their effects on the luminal milieu and, ultimately, physiology in conventionally-raised hosts in non-sterile conditions. The investigators propose a novel approach to address this need by using native bacteria as chassis for the introduction of specific functions into the luminal environment. The proposal's innovation is a new strategy that allows the quick and effective “knock-in” of a beneficial function in a sustained manner into conventional hosts. To date they have demonstrated that tractable native bacteria can be engineered to express a beneficial function ex vivo, reintroduced to the host, engraft the entire gut of the host, and deliver an intended beneficial function. These functions can affect host physiology, help determine the effect of specific bacterial functions and potentially alleviate disease. The central hypothesis of this proposal is that long-term colonization and functional change in the gut microbiome of a conventional host can be performed effectively with engineered native bacteria. In the next five years, the investigators will continue the development of this technology and better understand the chassis-host interactions that will aid in the development of live bacterial therapeutics for clinical use. First, the investigators will engineer regulatory systems for the transgene of interest, including a sense and control, protein secretion, and biocontainment circuit. They will test whether these systems function in vivo in hosts that are in a non-sterile environment. In addition, they will assess the natural biocontainment of engineered native bacteria among co-housed hosts. Second, they will determine how the niche for a bacterial chassis affects function delivery and whether multiple functions can be delivered by the same chassis or whether different chassis are necessary for the delivery of multiple functions. Finally, the investigators will determine the role of microbial community in amplifying the effects of a transgene of interest in gnotobiotic mice. The expected outcome of the proposed studies is attainment of fundamental biological knowledge of how the gut microbiome can be functionally manipulated. These studies will have a positive translational impact because they will demonstrate that synthetic biology approaches can lead to the development of curative interventions to some of the most debilitating and costly chronic diseases.