Integrated Human Practices
To us, Human Practices was all about finding a place in the world for our project and adapting it where it is needed, so that it can fit perfectly in its niche and truly be an improvement for the global environment. This happened gradually over the course of our project and shaped our final outcome.
First, we discussed our ideals amongst ourselves: Since the very beginning we wanted to make a project that could tackle an aspect of a global problem: environmental contamination. Our goal was to create something that would solve a real issue and in a way where it was accessible to those who truly need it. Biosensors were ideal for this as they allow for simple, inexpensive detection of contamination - in ways where even the most advanced technological system is unable to. When Professor Cyril Zipfel - an expert in plant immunity - became our principal investigator, our goal became clear to us: Make a biosensor based on the plant immune system, specifically plant pathogen recognition receptors (PRRs) and do it in a way where anyone can use it: with a simple, binary “present or absent”, colorimetric output. That way we can visualize bacterial contamination in water in a cheap way and without the need for additional equipment making it usable by anyone, anywhere. This of course would benefit people who need it most, those in developing nations and remote locations, where access to safe water is rare. Additionally working with PRRs as biosensors is a novel approach, opening the doors for many new future developments.
As is often the case with scientific projects, where we initially aimed was not where we ended up. And that is a good thing, as our new goal was redefined by experts, whose understanding and knowledge we integrated into our new goal. At first we discovered the issues with our original idea and were lost in the cloud for a while, but by listening to the needs of experts in the field, we adapted our project to a new goal: A water quality sensor that could be used by professionals to determine changes in water quality and track issues in systems, due to its rapid response and low cost per sample. We also redoubled our efforts of groundwork, as the importance of finding new specificities in microbial sensing was highlighted to us. Overall we emerged from the cloud with a system that now had more of a place in the world than our original ever would have. To this end we adapted our goals, procedure and system on every level.
Of course no human practice would be complete without reaching out to the broader community, both so they could learn from us and so that we can learn from them. To read more about this please visit our science communication page.
For our experts we needed to cover 3 essential topics:
- Water safety & developing nations, to gain perspective on what is necessary and lacking in water safety in general and in the context of affected countries.
- Microbial Sensing, as it is essential to understand techniques currently available, to be able to improve on them.
- Biosensors, to better understand the advantages and limitations of systems like ours, and to identify safety risks with our product.
Initial goal: Layman sensing system, with binary present or absent output that is visible to the naked eye.
Layman sensing is difficult. GMOs bring many troubles with regulations and disposal.
Total cell count is useful for detecting post-disinfection regrowth. Chlorination is a simple and harmless water treatment, applicable independently of sensing.
Due to many issues with layman sensing and open niches in the expert sector, we have decided to focus on expert sensing applications.
Total cell counts can be useful for quality measurements. Luminescence is a more sensitive system than colorimetric.
To fulfill our new goal of quantification, we changed our colorimetric output to a luminescence output, to increase sensitivity & quantification at the cost of needing a machine for readouts.
Systems don't need to be perfect, they need to lead to decisions.
Total cell count is a quick and simple indicator of water quality.
Total cell count monitoring, to measure changes across many samples.
Our approach is not suited for developing nations, as distribution systems are limited and general sensing is not as important as specific sensing.
Prof. Jan Roelof van der Meer - University of Lausanne
Our first contact in Human practices was Prof. van der Meer, an expert in Biosensing and someone who has great experience as he has worked on an arsenic bioreporter in the past. He gave us many valuable inputs on our original concept, which at the time was still very idealistic. We discussed the usage & issues of bioreporters in general and specifically their usage in a layman context. Our key take-aways were:
- A yes/no colorimetric output is not that useful, there is too much background of non harmful bacteria.
- An everyman's sensor is complicated to market.
- Proper disposal of GMOs is crucial, but almost impossible to guarantee in a layman context.
- Speed is far less important than accuracy, when it comes to topics where safety is at stake.
EAWAG - Swiss Federal Institute of Aquatic Science and Technology
One of our main contacts, with which we had multiple meetings was the EAWAG. The research at EAWAG focuses heavily on water safety, pollution and obviously microbial detection in water. They were a very relevant contact to us due to their experience with microbial water contamination and their interests in water sensing projects like ours. They also provided valuable input from the perspective of low income countries, as they have many projects that focus on the development of water safety in low income countries. Initially we had planned a partnership where they provide us with contaminated samples, which would enable us to generate data for our sensor on real world samples, to determine sensitivity & specificity of a potential prototype.
Dr. Eva Reynaert - EAWAG
Dr. Reynaert is working on the AUTARKY project, a handwashing station that recycles water directly using a membrane-biofilm system. This was interesting to us, as one of the uses of this system is recycling water in locations with no access to pipeable water in low income countries. We discussed water safety in a non potable, developing nation context and how she tests her system for microbial pollution & what is currently left desirable about microbial sensing. Our key take-aways were:
- Disinfection (Chlorine) is a simple, effective water treatment method with few drawbacks - as such it is done often even independently of water quality sensing.
- Total bacterial cell count is a useful measure to monitor post disinfection bacterial levels and regrowth.
- Changes in total bacterial cell count and removal values are valuable, to compare before and after disinfection steps.
- A threshold or quantification is absolutely necessary.
- Viral detection is currently a far more challenging but equally vital aspect of microbial sensing.
New goal: Expert sensing
At this point, we had received 2 important feedbacks: Layman sensing is difficult to achieve with our current project and serves limited purpose. Expert sensing on the other hand, has a niche for systems like ours, but quantification is necessary for this use. As such we changed our goal to be more oriented towards laboratory personnel, with a focus on making our project quantifiable.
Dr. Alessandro Franchini & Dr. Anselme Fournier – PROMEGA
Promega as a corporation is one of our sponsors and they produce a water sensing kit, Water-Glo™ Microbial Water Testing Kit, with an output similar to ours: Total microbial load. With them we discussed the use of a general sensor vs a specific sensor and what to gain from general sensing. We discussed the specifics of their system and what to expect from our system - and how we could improve sensitivity. Our key take-aways were:
- Luminescence is more suited as a quantifiable output than colorimetric, it has higher sensitivity and thus allows for a lower limit of detection.
- Total bacterial load still serves a purpose, to monitor water quality.
- Our system will have a higher limit of detection and will be less sensitive than comparable systems such as Water-Glo, but will be cheaper and simpler to operate.
- We were advised to focus on general water quality, not water safety & drinking water as those are far more difficult to determine with a system like ours.
- A future workaround to GMO issues could be a cell free membrane capsule system.
System change: Luminescence output
After our discussions with Promega, we decided to forgo the use of a colorimetric output in favour of using a luminescence output to increase our sensitivity and lower our limit of detection. Still, our sensitivity will be lower than other systems, but should be sufficient for observing changes in water quality.
Prof. Eberhard Morgenroth - EAWAG
Prof. Eberhard is an expert in biological water treatment and with him, we discussed specific application potentials of a system like ours. We talked about how single tests do not need to be perfect, they need to lead to decisions and how we would have to adapt our project to lead to meaningful decisions being made. Our key take-aways were:
- Independent, single event test kits are important to make decisions, even if the results are imperfect.
- Results need to lead to decisions.
- Specificity or quantification is absolutely necessary.
- Dangerous species are far more important but less common, thus a threshold based approach is not useful
Dr. Frederik Hammes - EAWAG
As the leader of the Drinking Water Microbiology group at EAWAG, Dr. Hammes was a very important contact for us, as someone who could give us a perspective from inside the lab on microbial detection. In our discussion we learned where the total bacterial cell count could be applied and, just as importantly, where it could not be applied. We discussed alternative water sensing systems and compared ours. We also talked about Legionella & viruses and what is missing in their detection. Key Points:
- Total cell count is an indicator of water quality, not safety. It is an excellent, quick measure for changes in water stability across distribution networks.
- Living cell counts are very important, especially for after disinfection. Proteins (i.e. enzymes) might be too stable to be indicative of live organisms.
- Legionella cannot be related to total bacterial cell count, but protozoans may be an indicator for Legionella biofilms.
- Detecting protozoans is currently very difficult. Even a general protozoan sensor would be extremely valuable, as very few species are inconsequential.
- A layman sensor would have been interesting, but specificity is necessary in a developing nation context.
Proposed implementation: Total cell count monitoring
With our newfound understanding of the importance of the total bacterial cell count, we propose a new implementation of our system: To monitor changes in water quality. This is possible because the cost, simplicity and speed of our system allow for many samples to be taken and quantified.
Dr. Susan Mercado - Special Envoy of the President for Global Health Initiatives, Philippines
We were very lucky to be able to get a meeting with Dr. Mercado, as she is not only an appointed health official, but has a long and impressive history with the WHO and has worked on the ground in developing countries. She was able to give us an all important perspective from outside the first world & laboratory experts, which is essential to consider for a project that seeks to help developing nations. We discussed how our project could be applicable - or more specifically how it couldn't. We discovered many oversights in our goal & system that make sense in a context as we know it, but are far more complex in the reality of a developing country. Key Points:
- Chlorination/Disinfection is the key and often only response to bacterial contamination in water.
- Faecal indicators are enough to chlorinate the water and sufficient tests for that already exist.
- Without alternative solutions other than chlorination, additional information from a test is not that helpful, if it is not very specific to harmful bacteria.
- Many countries do not reach WHO standards or have running water, so quality comparisons are not feasible.
- A system directly connected to an information network would be more applicable.
In summary, we learned a lot about how our perception of what was necessary for a layman system is not what we could offer. But instead of giving up or getting lost in the cloud, we adapted. We learned what was missing in microbial sensing in an expert context. From that we decided to focus our current iteration of the project on expert sensing, and adapted the system to match. Our goal shifted and so did our questions in Human Practices. Our system itself also adapted on a molecular level, to be more in line with the requirements of expert sensing, mainly quantification.
However the current implementation of our project is only part of our goal - the other aspect is laying the groundwork for future usage of PRRs as biosensors.
During our Human Practices research, we also learned that there is a great lack of specificity in the field of cheap sensing, especially when it comes to elusive microbes such as viruses . This is an example of how PRR based biosensing can go beyond our current implementation, as PRRs that recognize viruses are currently being researched. Additionally future development may lead to engineered PRRs  that can be designed with improved or expanded ligand affinity.
The modular design of our system enables an easy integration of newly discovered PRRs. For this reason we believe PRRs have great potential for future use in Biosensing and it was important to lay the groundwork for their advancement, both in the lab by proving their expression and providing the parts, but also on an outreach and education level with our presentations and the PRR guidebook.
To learn more about how we finally implemented our project and what its future looks like in detail, please read our proposed implementation or description page.
Our project uses exclusively risk group 1 components, with our two chassis (Saccharomyces cerevisiae and Chlamydomonas reinhardtii) already existing as free living, harmless organisms in the environment. Our integrated parts come from two very common, non-dangerous plants and are not involved in anything besides sensing. So on a molecular level our project is very safe, as it contains no harmful components.
Still, considering safety is an important aspect of a project like this, especially since it was initially considered to be an application for laymen. Since our implementation involves a bioreporter, a GMO, proper disposal and escape into the environment are our primary safety concerns when it comes to the use of our project.
Indeed, we found the safety risk of handing a system like ours to laymen, especially considering current regulations, too large. Since we could not provide a completely bulletproof solution to proper disposal, we rely on proper disposal by the user, which cannot be guaranteed in a layman context. Thus we moved our project into the hands of experts, who with proper instruction should have no difficulty properly disposing of our bioreporter and the contaminated water sample, by disinfection or autoclaving.
 Gouveia Bianca C., Calil Iara P., Machado João Paulo B., Santos Anésia A., Fontes Elizabeth P. B., Immune Receptors and Co-receptors in Antiviral Innate Immunity in Plants, Frontiers in Microbiology, Vol. 7, 2017, doi: 10.3389/fmicb.2016.02139
 Albert, Markus et al. “Arabidopsis thaliana pattern recognition receptors for bacterial elongation factor Tu and flagellin can be combined to form functional chimeric receptors.” The Journal of biological chemistry vol. 285,25 (2010): 19035-42. doi:10.1074/jbc.M110.124800
 Freddy Boutrot and Cyril Zipfel, Function, Discovery, and Exploitation of Plant Pattern Recognition Receptors for Broad-Spectrum Disease Resistance, Annual Review of Phytopathology 2017 55:1, 257-286, doi:10.1146/annurev-phyto-080614-120106
EAWAG header image: by Andri Bryner, 16. October 2012, shared under CC BY-SA 3.0