WHAT DOES OUR TEAM CARE ABOUT?
We wanted our project to reflect our values. During the ideation process it became clear that all our team members are concerned about environmental issues the world is nowadays facing. As Finland is a land of thousands of lakes and located by the struggling Baltic Sea, we settled on protecting water resources. After deciding on this general topic, all our team started an exhaustive research on environmental problems involving waters in Finland, the Baltic Sea and the European Union in general.
During the ideation we familiarized ourselves with many interesting iGEM projects. The ones we found particularly inspiring were Sydney 2016 and Westminster UK 2019 teams. The team from Sydney aimed to develop a biosensor in form of a sticker that could indicate the ripeness of avocado. We all thought a biosensor could be an interesting idea for use of synthetic biology tools to tackle many issues. On the other hand, the Westminster team attempted to generate electricity from the process of plastic degradation. They were planning to take advantage of the Mtr pathway of Shewanella. It gave us the idea we could use this pathway to create an electrical output in our biosensor. Even though we were not able to test it experimentally, we believe this pathway to have a potential in environmental biosensors. More information about the Mtr pathway in our project can be found here.
MICROPOLLUTANTS AND MACROLIDES: WHY A BIOSENSOR?
During our research we realized that in recent years there has been a growing concern about presence of pharmaceuticals in water. These compounds are very problematic due to their persistence and the fact that they remain biologically active . Even in relatively low doses they may have a negative impact on the environment. In addition to that, the consequences of interactions of many different pharmaceuticals are unknown and may have a synergistic negative effect . For all these reasons, it is extremely important that people start realizing how dangerous pharmaceuticals can be for the environment and there has to be a community effort to try to remove the compounds from waters.
A particularly problematic example are macrolide antibiotics, which in addition to above mentioned issues also carry another risk: development of antibiotic microbial resistance . In fact, in a status report about pharmaceuticals in the aquatic environment of the Baltic Sea region , it is explicitly mentioned that macrolide antibiotics, concretely erythromycin, clarithromycin and azithromycin, are in a “watch list” of priority substances that are of significant risk for the aquatic environment. More about the environmental impact of pharmaceuticals and macrolides can be found in our Incentive section.
Although these compounds are in the “watch list”, currently there are no regulations regarding the monitoring or removal of these compounds in the European Union. Nonetheless, several experts of the wastewater treatment industries that we contacted (see our Integrated Human Practices page) confirmed us that it is expected that there will be new legislation and more strict regulations regarding the compounds in the “watch list” in the near future. This will create a need for a cheap and reliable way of monitoring their levels in wastewater. However, these substances are present in rather low concentrations, which makes their monitoring challenging .
Current detection methods usually include LC-MS/MS or UHPLC-MS/MS which are quite costly, time-consuming and require qualified personnel . A biosensor could solve this issue, since they are cost-effective, easy to use and can be made portable due to relatively small sizes . Such a sensor could be used, among others, in wastewater treatment plants to optimize the removal process of pharmaceuticals.
Our iGEM project introduces SINISENS – engineered bacteria for detection and quantification of macrolide antibiotics. Biosensors for environmental use have been research for over a decade, yet there are hardly any that have reached beyond academia. We wanted to design a product with a potential for commercial use. To achieve this, we focused not only on designing a genetic circuit, but also on optimization, calibration, costs and ease of use.
BIOSENSOR FUNCTION: DETECTION
A sample of wastewater would be introduced to our biosensor. When there are no macrolides present, the bacteria would not give an output signal. In presence of macrolides, green fluorescence would be produced, which could be correlated with the amount of the compound (Fig. 1). More about our proposed implementation can be found here.
To improve the detection limits of the biosensor, we also modelled mutations in the MphR transcription factor to increase its binding affinity to macrolides. See our modelling page.
BIOSENSOR IMPROVEMENT: CONCENTRATION
Since concentrations of macrolides in wastewater are rather low , it is important to have a sensor with a very sensitive range. For that reason, we also focused on finding an optimal method for concentrating macrolides in cells using tools within, as well as outside of synthetic biology. We researched, among many others, potential applications of microfluidics technology and modifying bacterial outer membrane channels (Fig. 2). More about our concentration research can be found here.
As everyone this year, our work has been affected by COVID-19. In addition to general uncertainty of the situation and cancellation of iGEM-related events and meet-ups, our access to the laboratory was delayed over two weeks. Moreover, the number of people allowed in the building was limited during the whole summer. For that reason we also were not allowed to use certain lab spaces and equipment. Luckily, we were able to utilize quite a lot of dry-lab in our project. The downside was that unfortunately all the work had to be done from home, which made having smooth communication between team members more difficult. However, we understand we are relatively lucky in that regard, since many iGEM teams this year did not have an opportunity to access lab at all.
1. "Science for Environment Policy": European Commission DG Environment News Alert Service, edited by SCU, The University of the West of England, Bristol.
2. Arnold, K. E., Boxall, A. B. A., Brown, A. R., Cuthbert, R. J., Gaw, S., Hutchinson, T. H., et al. (2013). Assessing the exposure risk and impacts of pharmaceuticals in the environment on individuals and ecosystems. Biology Letters (2005), 9(4), 20130492. doi:10.1098/rsbl.2013.0492
3. Schafhauser, B. H., Kristofco, L. A., de Oliveira, Cíntia Mara Ribas, & Brooks, B. W. (2018). Global review and analysis of erythromycin in the environment: Occurrence, bioaccumulation and antibiotic resistance hazards. Environmental Pollution, 238, 440-451. doi:10.1016/j.envpol.2018.03.052
4. Agency for Healthcare Research and Quality. (2018). 2017 national healthcare quality and disparities report (Report No. 18-0033-EF). U.S. Department of Health and Human Services. https://www.ahrq.gov/research/findings/nhqrdr/nhqdr17/index.html