B-189. Investigating Interactions between GI Tract Signaling Molecules on Escherichia coli O157:H7 Chemotaxis Using a Novel Microfluidic Chemotaxis Model

D. Englert, F. Senocak, J. Kim, T. K. Wood, A. Jayaraman;
Texas A&M Univ., College Station, TX.

The GI tract is abundant in a range of eukaryotic (e.g., norepinephrine, dopamine) and prokaryotic (e.g., AI-2, AI-3, and indole) signaling molecules. The high local concentration of these molecules often leads to them spilling over into the intestinal lumen, where can be encountered by pathogens. We have recently demonstrated the importance of these signals in E. coli O157:H7 (EHEC) infections by showing their effect on EHEC chemotaxis, motility, gene expression, and adherence to epithelial cells (Infect. Immun. 75: 4597, 2007; Appl. Environ. Microbiol. 73: 4100, 2007). However, not all signals have the same effect; for example, indole counters the effect of norepinephrine on these EHEC phenotypes. Importantly, EHEC is also likely to encounter concentration gradients of different signals in the lumen, as the signals are not produced uniformly throughout the GI tract. We hypothesize that the gradients of signaling molecules encountered by EHEC governs the initial migration by EHEC that leads to infection. A novel microfluidic chemotaxis model (1050 um wide x 20 um tall x 8000 um long) was developed for generating precise and stable concentration gradients of signaling molecules and used to investigate the effect of signaling molecule gradients, both individually and in combination, on EHEC chemotaxis. Laminar flow-based diffusive mixing was used to generate concentration gradients of different strengths, and the chemotactic response of GFP-expressing EHEC to the gradients was determined using confocal microscopy. EHEC was attracted to 0 to 100 uM gradients of AI-2 and norepinephrine, but repelled from indole (0 to 500 uM). EHEC also migrated towards a 0 to 500 uM gradient of isatin, which is a derivative of indole. Interestingly, EHEC was also attracted towards a combined 0 to 500 uM gradient of indole and isatin. Similarly, concentration-dependent interactions were also observed as EHEC migrated towards 0 to 100 uM, but away from 0 to 500 uM norpeinephrine. This data strongly support the hypothesis that concentration-dependent interaction between signals is an important determinant of EHEC colonization.