Team:NU Kazakhstan/HP/Gold Integrated

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Introduction to Human Practices

Our project created new challenges in synthetic biology and integrated fields for us. Our primary goal was to generate an idea, (i) which can shape the newly built scientific body of our university, (ii) which can solve major ecological problems in Kazakhstan and (iii) inspire Kazakhstani population to be more careful and considerate of ecology and science. We believe that ecology is what makes our development healthy and science is what makes our directions clear. In this passage we combined Integrated Human Practices for Gold and Human Practices for Silver Medal.

In the Integrated Human Practices’ part you can find our journey from the baby steps of brainstorming till the final stages of the project. In this journey, we had two meetings with company representatives of mining plants, scientists, experts, and just regular people who care about the ecological situation in Kazakhstan. During these meetings, which was far from Astana for 1500 kilometers (~1000 miles), we gained the whole picture of chromate reduction and waste neutralisation in Kazakhstan, which inspired us to select the target organism in bioremediation of waste waters, to construct the safety forms, and run the project.

The Human Practices part for the Silver medal combines everything we did what we thought to be right to be vocal. We do not believe that ecology of steppes is the only modern problem we face but the ecology of minds in our university too. We think that popularization of science is one of the most important parts what we aimed to accentuate on. We tried to bomb the minds from every single corner available and opened research & internship opportunities for the research starters. We were constantly facing barriers in advocating for minorities in science like women and queers. However, we think that our hard work gave another breath for the new scientists to fulfil their true potential. We could open an internship opportunity for talented high school students and university students, also we supported Women in Tech program to inspire mid school girls. We could advocate for science and ecology problems through media (TV and science-pop journals) and participated in seminars in high schools. In addition, we opened series of seminars on Human Rights in Science and invited the most popular human rights activists and scientists to deliver their speech.

As a result, (i) we could build the body of a lab where researchers can work with algae modification, (ii) to create a developed idea to solve the problem with pollution in Kazakhstan, and (iii) to create a consensus around chromate pollution and science popularisation. We are thankful to everyone who contributed to Human Practices.

Idea Development

The process of idea generation took more than 2 months. Our primary goal was to design something meaningful that would help to solve existing problem in Kazakhstan. We researched through the available sources and decided that we should focus on environmental problems. Kazakhstan has the second largest chromium deposits in the world. Majority of chromium producing plants are located in the West of Kazakhstan: JSC "Aktobe Chromium Compouds Plant", JSC "Aksu Ferroalloys Plant", LLC "Sunrise Chromium", JSC "Ferroalloy", TNS "Kazchromium", Aktobe Paint Factory, and "Sunrise" Chromium Ores Processing Plant. It is not surprising that huge amounts of chromium-containing wastes are produced every year, which is considered as carcinogenic and mutagenic. Before coming up with a solution we evaluated existing methods of chromium utilization in the world.

Each of the existing methods has its own disadvantage such as high price, pH dependence, low selectivity for chromium, formation of precipitate or requirement for the additional reagents. Considering aforementioned obstacles, bioremediation is a perfect candidate to solve the problem.

First Meeting in Aktobe

In order to make sure that our project will be of current interest and relevant to real world problems we organized meeting in Aktobe city, where chromium has been mined and processed for more than 60 years. We invited people from different areas including professors, representatives from industry and governmental officials. Three of our students gave a presentation regarding the chromium pollution, risks for population and bioremediation as a potential solution.

Bioremediation is still on its ground level in Kazakhstan, and we described main principles of bioremediation, advantages over conventional methods in simple and understandable form to our audience. Public was actively engaged in the presentation and we got lots of questions and feedback.

This meeting was important for project development in several ways:

Governmental officials together with industry representatives confirmed the importance of problem of chromium pollution in the region. Based on their feedback we decided to proceed and develop our idea to the full extent.
We learnt about the methods of chromium utilization used nowadays in Kazakhstan. Turned out, that industry uses methods of chemical precipitation using iron oxide to treat wastewaters. However, this method requires large enough amount of reagents to reduce chromium to the appropriate level for further discharge. Moreover, it creates additional problem of sludge formation and aggregation of precipitate containing metals. This can have detrimental long-term consequences for the environment.
One of the important findings during the meeting was that besides wastewaters containing chromates there is a river Ilek which is also highly polluted. Taking this point into consideration, we decided to expand our project to bioremediation of both water from plants and natural waters.

The feedback that we got from this meeting gave us a clear picture about the challenges of chromium utilization in real conditions and led us to a conclusion that Kazakhstan will benefit from new alternative method of bioremediation which promises to be cost-effective, environmentally friendly and effective at the same time. This meeting reinforced us to develop project in bioremediation of both natural and waste waters.

Design of the project

After extensive research on chromium pollution, we found that some of the methods including chemical precipitation are based on the principle of reduction of more soluble hexavalent chromium to the less soluble trivalent. Trivalent form possesses 1000 folds less toxicity compared to Cr(VI) form. Therefore, we decided to create an organism that would perform three major functions: uptake hexavalent chromium, reduce and store it. There were number of aspects that we considered in our design:

Careful selection of model organism. We chose Chlamydomonas reinhardtii because
- It is non-pathogenic organism. Safety comes first.
- Environmentally friendly organism producing oxygen. It doesn’t produce secondary wastes.
- It can survive a wider range of temperatures compared to other model organisms. This especially important for application in Kazakhstan which has extreme continental climate.
- C. reinhardtii is a photosynthetic organism, which can survive in presence of light, salts and CO2. This is a strong advantage over bacteria, which require source of carbon.
- C.reinhardtii can be effectively immobilized on the surface of loofah sponges. This makes our idea easier to implement in real life in bioreactor.
- C.reinhardtii has natural ability to upregulate sulfate transporters which at the same time are channels for chromium under the conditions of sulfur starvation. This ability can be exploited in order to increase uptake of chromium from water.
Selection of Parts
- We chose “Chromate reductase” from Escherichia coli (strain K12)
- Chromodulin serves an important function inside the cell. It prevents pumping out of reduced trivalent chromium into environment. Even though trivalent form is less toxic, but still it can have negative effect on living creatures. This way we ensure that all the reduced chromium is preserved inside the cell.
- SuperNova protein is safety system that helps to prevent GMOs spreading into environment. SuperNova is activated under the 585 nm wavelength contained in daylight.
Bioreactor design
- Loofah sponge is an excellent support for C. reinhardtii that allows effective immobilization.
- Bioreactor will have screen which blocks irradiation from 500-600 nm. In case if C.reinhardtii leaves bioreactor daylight (containing 585 nm) will activate safety system (SuperNova). This protein produces excessive amounts of ROS which kills the cells.

Presentations for university faculty

We had several meetings with our faculty which influenced our project. Our supervisor Dr. Abdulla Mahboob who had an extensive experience working with C.reinhardtii guided us throughout the project. We had meetings with experts in the field of algae in our university Veronika Dashkova and Dmitriy Malashenkov. They gave us some valuable advices on how to set up algae lab. Moreover, most of the biology faculty, including head of biology department Dr. Ivan Vorobyev, evaluated our project for relevance, sustainability and safety. In the very beginning of the design process, Dr. Ivan Vorobyev suggested us to focus on unicellular microalgae instead of macroalgae.

Second meeting with KazChromium

When our primary working model was designed, we organized official meeting with another Aktobe regional ferroalloy factory - KazChromium. After this meeting we introduced major changes in our bioreactor design. Results are the following:

Some part of the wastes produced by KazChromium is in the solid form and needs to be dissolved before the use of our method.
The most important feedback and advice that our team learned is that contamination of natural water was mainly caused through flow of underground water. Because of this discovery our working model was completely modified. Before the meeting we designed bioreactor in such a way that the source of light was Sun during the day. Since it is no longer possible to use the daylight for underground water, we decided to introduce artificial irradiation. Microalgae immobilized on the loofah will be growing in a closed tank under the lamp which gives only red and blue light. There is also a second section which has irradiation of full spectrum including 585 nm. This way, those cells which left the first section of the tank will activate SuperNova and will be killed.

Safety and Governmental regulations

One of the important issues that need to be addressed before starting any project is to check out laws and regulations regarding the use of GMOs in your country and then ensure that the project is safe. According to the Code of the Republic of Kazakhstan on administrative offences (Chapter 40, Article 282): “For genetically modified organisms intended for deliberate release into the environment, users should provide the authorities in the field of environmental protection and the state authority of the sanitary-epidemiological service with detailed information on their characteristics”. In order to get permission for future real life application of our project we took into consideration the following criteria:

We carefully searched for the chassis which will be safe to work with and at the same time be convenient for the purposes of the project. We chose a Chlamydomonas reinhardtii which is a non-pathogenic and environmentally friendly organism.
Parts that we use in the project possess no harm to humans and environment. Central part ChrR is a gene responsible for the reduction of toxic Cr(VI) to insoluble and less toxic Cr(III). Chromodulin tightly binds up to 4 equivalents of Cr(III) to keep all the reduced chromium inside the cells.
To prevent uncontrollable release of GMOs into the environment we designed the bioreactor and introduced a safety system. Firstly, our microalgae will be immobilized on loofah sponges. Secondly, we are introducing membrane-bound photosensitizing protein SuperNova, modified from previously registered cytosolic form.