Considerations

  Applications

We have successfully demonstrated that E. Coli can be engineered to detect the presence of Pseudomonas Aeruginosa autoinducers. Our project opens up a wide range of possibilities for future iGEM projects and even commercial devices. We hope that one day our work will make it easier to detect and combat P. aeruginosa infections in clinical settings.

In the immediate future, there are several ways to expand upon our project. One goal we were not able to accomplish is to match up the two different quorum sensing receptors with two different reporter genes. This would allow for more detailed information about the progression of a Pseudomonas infection. In addition, the two separate signals could be used as part of an AND gate to reduce the risk of false positives. Another area that could be expanded upon is mining the large number of other P. aeruginosa genomic promoters that are autoinducer sensitive. We have sent two autoinducer-activated genomic promoters to the registry, but there are literally hundreds more within the genome. Each promoter likely has slightly different properties and sensitivities, so perhaps there are promoters particularly well suited for a detection system. Characterization and analysis of genomic promoters could allow for increased signal strength or decreased response time.

We propose to utilize our construct as a biosensor in a device which can read fluorescence. The theoretical design of such a device is depicted below in Figure 1.

Figure 1: A prototype design for a device which can detect and quantify the presence of P. aeruginosa. The image on the left is a circuit/schematic view, whereas the one on the right is a side view.

Figure 1 is a schematics of how a prototype device which can detect P. aeruginosa would look. The device will initially radiated light (from a blue LED source) which shines on the well. The well contains the autoinducer-sensing E. coli and sample to be analyzed. An orange optical filter, that blocks the blue light, but passes the GFP GFP fluorescence (which is in the well) is placed between the well and photodiode sensor. Once the GFP fluorescence begins, the photo-diode will convert the light based signal into an electrical one. This electrical signal will be processed by an Arduino microprocessor, which feeds the display with the relevant concentration.

As a proof of concept, we subjected each of our three key component systems (which was also submitted for judging) to an array of blue LEDs with an orange filter. Figure 2 illustrates the validation of our concept and in turn, its practicality.

Figure 2: Proof of concept demonstration of the binary detection biosensor for PAI-1 via las, concentration-responsive detection of PAI-2 via rhl , and concentration-responsive detection of PAI-2 via a novel genomic promoter.

In the long term, this technology offers great potential for a rapid and routine Pseudomonas detection system. Existing commercial methods require a potential sample to be grown overnight, but our system reported measurable GFP transcription within an hour of induction. To do routine monitoring of the hospital environs, thousands of samples would need processing, which is impractical by the current methods. We envision the development of a handheld device that can be carried through a hospital, consisting of two chambers. One chamber would contain our engineered E. Coli, while a potential Pseudomonas sample would be placed in the other. The contents of the chambers would then be mixed and the device could be analyzed for the presence of a reporter. This faster diagnosis method could allow for quicker treatment in patients, and the detection of Psedeumonas in the hospital environment could also dramatically lower the incidence of this common nosocomial infection.