Engineering bacterial thiosulfate and tetrathionate sensors for detecting gut inflammation.

Laura C Ortiz‐Velez,Kristina N‐M Daeffler,Jeffrey J Tabor,Christopher O Bibb,Noah F Shroyer,Ravi U Sheth,Robert A Britton,Jeffrey D Galley

doi:10.15252/msb.20167416

Laura C Ortiz‐Velez, Kristina N‐M Daeffler + Show 6 more

Open Access

https://doi.org/10.15252/msb.20167416

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Abstract

There is a groundswell of interest in using genetically engineered sensor bacteria to study gut microbiota pathways, and diagnose or treat associated diseases. Here, we computationally identify the first biological thiosulfate sensor and an improved tetrathionate sensor, both two‐component systems from marine Shewanella species, and validate them in laboratory Escherichia coli. Then, we port these sensors into a gut‐adapted probiotic E. coli strain, and develop a method based upon oral gavage and flow cytometry of colon and fecal samples to demonstrate that colon inflammation (colitis) activates the thiosulfate sensor in mice harboring native gut microbiota. Our thiosulfate sensor may have applications in bacterial diagnostics or therapeutics. Finally, our approach can be replicated for a wide range of bacterial sensors and should thus enable a new class of minimally invasive studies of gut microbiota pathways.

Highlights

The mammalian colon plays important roles in metabolism (Tremaroli & Backhed, 2012), and immune (Hooper et al, 2012) and brain function (Mayer et al, 2014)
We hypothesized that a bioinformatic search for a two-component system (TCS) containing a sensor kinase (SK) with a PhnD-like sensor domain located near a thiosulfate reductase might reveal an uncharacterized thiosulfate sensor
Mice treated with dextran sodium sulfate (DSS) had elevated inflammation relative to healthy controls and similar histologic scores as the DSS-treated mice given the thiosulfate sensor (ThsSR) strain (Appendix Fig S26B). These results suggest that either our engineered S. baltica TtrSR construct does not function in vivo, or tetrathionate concentration in the lumen of DSS-treated mice, where our sensor bacteria likely reside in this 6-h protocol, is below the ~1 lM limit of detection (Fig 4F)

Summary

Introduction

The mammalian colon (gut) plays important roles in metabolism (Tremaroli & Backhed, 2012), and immune (Hooper et al, 2012) and brain function (Mayer et al, 2014). Gut processes are orchestrated by metabolic and signaling interactions between host cells and the dense and diverse community of resident bacteria (the microbiota). The mice in this study, and the three previous studies, were either raised in germfree conditions or pre-treated with antibiotics to clear the native microbiota prior to administration of the engineered sensor strains This sweeping perturbation has major effects on gut physiology, making these methods poorly suited to the analysis of gut pathways. In light of these studies, two major current challenges are to (i) engineer bacterial strains that sense other metabolites produced in the gut, and (ii) develop methods to assay reporter gene expression from those strains in animals with an intact microbiota

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Results

Conclusion