Project Purpose:

When first deciding on our project, we knew we wanted to focus on a topic that would have a meangiful impact on society, such as the importance of clean water, which is present in every aspect of our lives. Issues of contaminated water in places like Flint, Michigan are affecting communities of color more drastically than others and environmental concerns regarding water safety are both major problems that we wanted to help address. In our own community, recent water source changes have also raised questions about the water treatment process and the overall cleanliness of local drinking water. This ongoing nature of this issue is why our team decided to work on a project that could help test the quality of water to check for improvements. From here, we looked into the current industry methods for testing water after which we kept two audiences in mind: water testing facilities and researchers working with biosensors. Our two goals were to make a more efficient water testing method compared to current industry methods and also to develop a system more compatible for biosensor testing.

Biosensor Compatability:

Biosensor compatibility was a big issue we wanted to address with our iGEM project. Biosensors are biological systems that are used for detecting foreign materials by converting signals to measurable responses. Because they are very sensitive to the environments they are in, researchers have to work carefully when testing the parameters they are best suited for. Some of these parameters include, but are not limited to, pH, salinity and temperature. Some potential environments where these engineered biological systems can be deployed are listed below. It is clear that the performance efficiency of these biosensors will be affected by the different parameters of different aqautic environemnts:

We see this in a study from the Journal of Physical Chemistry Letters where scientists at the Ecole Polytechnique Federale De Lausanne (EPFL), Switzerland, “engineered stable optical nanotube biosensors” after testing them in liquid environments of various ionic concentrations (New selective biosensor made using synthetic biology). By analyzing the intensity of signals from the nanotube, they were able to determine the optimal salt concentrations for their biosensors compatibility. The issue arises from having to do this testing without a uniform, efficient testing system. This study focuses on just one parameter: salinity. With biosensors being sensitive to other liquid parameters as well, the need for a more efficient system is evident.

A study conducted by researchers at Ege University looked more into the optimization of liquid parameters for biosensor usage. Specifically, they looked into pH, salinity and temperature. They found that for their biosensors, a temperature of 35 C yielded the best results (as seen in Figure 1). As for pH, biosensor responses at pH levels above 7 were 50% less than that at a pH of 7 (Figure 2). As such, these optimal conditions were noted for future experiments.

From tested parameters like pH and salinity to potentially more parameters in the future, it's clear that a centralized device to test for biosensor compatibility would be a breakthrough. It is because of this that our team decided to work on a project that would be easier, cheaper and ultimately more efficient in the area of biosensor testing and compatibility. More on this can be found in the Integrated Human Practices section.

Water Testing:

To better understand current water testing methods (especially in the wastewater treatment industry), our iGEM team met with program manager and process engineer Caitlin Hunt from the Massachusetts Water Resources Authority. During our meeting, we learned about the current testing processes used by Deer Island Wastewater Treatment Facility as well as the challenges they face. The main problems addressed were the long turnaround times for testing as well as the inability to test for certain elements like Copper during field tests. Caitlin mentioned that for their in lab testing, tests usually take around one to three weeks to get results. These in lab testing facilities also require very large equipment and lots of lab personnel (50-100) for results. And lastly, their testing tubes accumulate biofilms that constantly break down their sensors. We also learned that their facility doesn’t currently have a way to test for copper concentrations at onsite testing centers. We looked into copper detection biosensors and learned how to improve our project. According to the World Health Organization’, copper release is more efficient in lower pH concentrations. In “ New biosensor for detection of copper ions in water based on immobilized genetically modified yeast cells”, researchers tested this by experimenting with their new copper detection biosensor in different pH concentrations. They found the opposite relationship between pH and copper detection biosensors activity as shown in figure 3.

While the biosensor's full range of activity isn’t completely tested and understood, it was helpful to see potential ways to solve Deer Island's inability to test for Copper. More on this in the Integrated Human Practices section.

Integrated Human Practices:

Multifluid Input:

Because our team was lucky enough to meet with researchers and industry workers in the water testing and biosensor fields, we were able to learn and adapt our project to more applicable needs. Our meeting with Bioengineers was on August 16, about 12 weeks into our project. One of the biggest changes we made from our meeting was the incorporation of a multifluid input design. Figure 1 shows the original product design - a design that didn’t take into account the sensitivity of biosensors to various liquid parameters. After our meeting with researchers, we knew we had to make our system more compatible and applicable for those working with a wide variety of biosensors. The difficult part was that most of our project design was completed at this point. But knowing we could make our project more universal, we decided to add sensors and liquid reservoirs to sense and regulate key components like pH and salinity. We also made our system flexible in that the content of the liquid reservoirs as well as the sensors being used, could be swapped out depending on the parameters the user wants to regulate. This new design provides a standardized system for biosensor testing, one that isn’t limited to a few liquid variables. This helped us in reaching the true purpose of our project - to make a system meaningful and compatible with labs from all around the world. Below the figures are our meeting questions and notes from our meeting with bioengineers. We learned a lot from this meeting and documented it all for future use.

Wastewater Treatment Facilities:

One of the main flaws that Deep Island mentioned was the buildup of biofilm in their machinery that requires extensive cleaning, further adding to the turnaround time for results. From learning about these issues, we improved our project by adding a three-stage filter process that would remove the need for daily maintenance/cleaning. It was this along with the reduced cost and results turnaround time of our system that makes it directly applicable for Deer Island Testing facilities. Another big improvement we made from our meeting was meeting the compatibility needs for copper biosensors for the successful testing for Copper. From talking to the Deer Island representative, we learned about their inability to test for metal elements like Copper for onsite testing facilities. We did further research and found the pH values required for current industry biosensors (genetically modified yeast cell- based copper detection biosensors). From this found pH of 8.1, we calibrated our system for the variation of pH from a base solution with pH of 7. In the figure below, you can find results from this calibration. To insure successful application, we did multiple runs of getting our base solution of pH 7 to the optimal value of 8.1 This makes our system directly deployable to Deer Islands onsite testing fields and makes their testing capabilities expand. The rest of our meeting notes and questions are also documented below. They were vital to the improvement of our project in various ways.

Engineering Biosensor Compatibility:

We also wanted to connect Deer islands needs to the work conducted by other iGEM teams in past competitions. After looking through the registry, we discovered that a potential solution to aiding in the Copper sensing compatibility issues lied right in the heart of the competition. We found that the promoter BBa_I760005 in the registry would be the perfect in aiding in Deer Island's issues. After some conversations with our Bioengineer colaborators, from this part, it would be possible to pair this promoter with another fluorescent protein in order to engineer the perfect biological system. Very simply, in the presence of Copper, the Copper would bind to the promoter and allow transcription of the fluorescent protein, thus producing a fluorescent output. This output could be read as a signal which can then be further processed to determine the presence of Copper in a given aqautic environment, or in this case, one of the many testing sites located at Deer Island. As seen by its description in the registry, the level of Copper is able to influence the strength of the fluorescence, proven via flow cytometry. This could work perfectly as a solution to Deer Island's need to test for Copper by telling them both the presence and level of Copper in a sample.

References:

Demirkol, Dilek Odaci. “Dependence of the Biosensor Responses on Ph of the Buffer Solution.” Research Gate, https://www.researchgate.net/figure/Dependence-of-the-biosensor-responses-on-pH-of-the-buffer-solution-Measurements-were_fig4_360607555.
Smith, M. (2021, October 16). More Lead-Tainted Water in Michigan Draws Attention to Nation’s Aging Pipes. New York Times.
Vopalenska, Irene. “New Biosensor for Detection of Copper Ions in Water Based on Immobilized Genetically Modified Yeast Cells.” Biosensors and Bioelectronics, 15 Oct. 2015.
Vopalenska, Irene. “New Biosensor for Detection of Copper Ions in Water Based on Immobilized Genetically Modified Yeast Cells.” Biosensors and Bioelectronics, 15 Oct. 2015.
Zakharov, Gennady. “IGEM Registry of Standard Biological Parts.” IGEM, 20 Aug. 2007.