WASTEWATER TREATMENT

When wastewater enters a plant, the first step is to remove coarse solids and large materials using a grit screen (figure___). The water can then be processed in three main stages: Primary, Secondary, and sometimes Tertiary Treatment (Pescod 1992). In Primary Treatment, heavy solids are removed by sedimentation, and floating materials (such as oils) can be taken out by skimming. However, dissolved materials and colloids—small, evenly dispersed solids such as nanoparticles—are not removed here (Pescod 1992). Secondary Treatment generally involves the use of aeration tanks, where aerobic microbes help to break down organic materials. This is also known as the activated sludge process (Davis 2005). In a subsequent sedimentation step, the microbes are removed and the effluent is disinfected (often by chlorine or UV) before it is released into the environment. In certain WWTPs, wastewater may go through Tertiary Treatment, an advanced process typically aimed to remove nitrogen and phosphorous, and assumed to produce an effluent free of viruses. However, Tertiary Treatment requires additional infrastructure that is expensive and complex, limiting its global usage (Pescod 1992; Malik 2014).

Biosafety

We have chosen to use a safe and common lab strain of E. coli, K-12, as our chassis (Environmental Protection Agency 1977). In both approaches, our constructs do not express proteins associated with virulence: PR is a membrane protein that commonly exists in marine bacteria, and for biofilm production we were careful to avoid known virulence factors such as alpha hemolysins (Fattahi et al. 2015). Most importantly, biosafety is built into WWTPs. Before treated effluent is released back into the environment, it must go through a final disinfection step, where chlorine, ozone, or UV radiation are used to kill microbes still present in the wastewater (Pescod 1992).

APPLYING PR IN WWTPs

Tap Water Museum

We visited the tap water museum hoping to find out more about how tap water is treated. We learned that water filtration methods vary in different areas of Taiwan, with Taipei’s filtration method being the simplest since the water is relatively clean compared to other regions, such as Kaohsiung, where the city is heavily industrialized. In Taipei, the source of tap water comes from a protected zone upstream of Xindian river. We also learned that they use sedimentation tanks and flocculation to help clump up and remove impurities. Due to the lack of a disinfection step, however, we realized that our project would not be applicable here, since our project depends on the use of E. coli bacteria. (Whole team activity)

Nanoparticle and Wastewater experts

Dr. Eric Lee

Before we started to conduct experiments, we emailed Dr. Eric P. Lee, senior member of technical staff at Maxim Integrated, and TAS alumnus, to ask him some general questions about our approach of our project. We told him about our two approaches, one with E. coli receptors that bind to the capping agents of nanoparticles, the other with biofilm that traps nanoparticles. Dr. Lee suggested that our membrane receptor must be specific to a particular capping agent. He also commented that the biofilm approach was a good idea since we could trap multiple types of nanoparticles regardless of their capping agent. (Interviewed by Emily C.)

Dr. Gwo-Dong Roam

We interviewed Professor Roam of National Central University and former general director of the Environmental Analysis Labs (EAL) of the Taiwan Environmental Protection Agency to learn more about the background and potential threat of nanoparticles. Dr. Roam informed us that the most common nanoparticles used in Taiwan include: TiO2, ZnO, Ag, Au, Fe, Carbon Nanotubes, Fullerenes, Clay, and Graphene. He also told us that the toxicity of a nanoparticle is directly related to its size, but there are currently no regulations or guidelines that specify the toxicity of different types and sizes of nanoparticle. With the increased use of nanoparticles in society, Dr. Roam believes that more attention should be placed on waste management, risk assessment and regulations.

After our first visit to the Dihua WWTP, we learned that the sludge removed from wastewater is either 1) sent to landfills, 2) used as fertilizer, or 3) incinerated. We asked Dr. Roam if sludge containing aggregated nanoparticles would still be harmful to the environment if disposed of using current methods. He said that all of these sludge disposal solutions are still harmful to the environment, but they are still better than letting nanoparticles flow into bodies of water. He advised us to target removal of nanoparticles in the wastewater treatment process before it is discharged. (Interviewed by Candice L. and Justin Y.)

Thomas J. Brown

Thomas J. Brown, the Water Program Specialist of the Pennsylvania Department of Environmental Protection (DEP) occasionally helps with the Boswell Wastewater Treatment Plant. He has also worked with the EPA in Taiwan on wastewater treatments. We interviewed Mr. Brown about our methods to clean nanoparticles in wastewater treatment plants and how to achieve our goal of implementation. With his expertise in the field of wastewater treatment, he provided us some suggestions as to how we could turn our project into reality.

For example, we asked him if there were differences between rural and urban plants that we should take into consideration when thinking about implementing our project. He responded that the biological processes used for treatment remains the same regardless of facility size. This helped us think about and design our final prototype, which can potentially be used in both rural and urban treatment plants.

Nanoparticle Manufacturers and Disposal Services

Apex Nanotek

To learn more about the applications of nanoparticles, we visited a nanotech company that uses silver nanoparticles to make various antimicrobial products. The researcher and manager of Apex Nanotek, Chery Yang, introduced us to their main product, which is antimicrobial nanosilver activated carbon. Pure activated carbon, commonly used to treat sewage and industrial exhaust, is prone to bacterial growth. To overcome this problem, they integrate crystallized nanosilver into the activated carbon for their antimicrobial effects. One of their products is a showerhead, with nanosilver activated carbon filters to kill bacteria when water flows through the showerhead.

We tested the product by comparing SEM images between tap water and filtered water from the showerhead. The showerhead decreased the number of bacteria and larger particles from tap water! However, we also observed the release of nanoparticles from the filter, which will flow into wastewater. (Interviewed by Christine C., Kelly C., Yvonne W., Chansie Y., and Justin Y.)

Chery Yang (third person from the left), the main researcher of Apex Nanotek Corporation

The image on the left shows a tap water sample under the SEM, in which we observed some bacteria (round objects that are approximately 1 μm in diameter). The SEM image on the left shows water that was filtered by the showerhead from Apex nanotek. There is less bacteria as the showerhead uses embedded nanosilver antibacterial filters. (SEM images: Christine C. and Florence L.)

THEPS Environmental Protection Engineering Company (中港環保工程股份有限公司)

We contacted the company that removes our nanoparticle waste because we wanted to know what happens when it leaves our lab. They directed us to National Cheng Kung university who actually treats the waste for them. The university uses chemicals and burning to aggregate nanoparticles. Through literature research, we discovered that burning nanoparticles is the most prevalent way for removal, however it is not 100% effective at removing all types of nanomaterials (Marr et. al. 2013). (Interviewed by Katherine H, Audrey T. and Christine C.)

Public Opinion

Survey Results

We created a survey that helped us identify public knowledge and misconceptions about synthetic biology and nanoparticle usage. Over 240 people completed the survey. (Survey created by Abby H., Christine C. and Emily C.)

Here are some results from our survey:

General Questions

• The majority of people think that gene modification is acceptable if the goal is to save or improve quality of life; however, it is not acceptable for non-medical related reasons, such as changing hair or eye color. In addition, most people do not have a preference between chemical or biological drug synthesis. These results suggest that people are accepting of genetic engineering when it is related to health and medicine.
• Environmentally, people are generally concerned with the wastewater that enters the ocean and the river. This gives weight to our project, because the quality of water is an important concern for the general public.

Project-Specific Questions

• The majority of people have heard of nanoparticles and know that nanoparticles are used in consumer products; however, they do not know why nanoparticles are used.
• Most people believe that the government and nanoparticle manufacturers should share responsibility for the regulation of nanoparticle usage and disposal.

Click to see all survey results:

Bioethics Panel

We hosted a Bioethics Panel, where we invited students and teachers to discuss the moral, social and environmental concerns of our project. To encourage participants to consider the problems from multiple perspectives, we created a role-playing game and assigned different roles to participants. We then asked for their opinions on nanoparticle usage and disposal from the perspective of their assigned role. (Whole team activity)

For instance, one of our questions was:

“Dihua WWTP has no nanoparticle removal plan. Should this be the job of the wastewater plant? Or the nanoparticle manufacturer?”

The following roles were assigned:

• Wastewater plant manager
• Nanoparticle manufacturer
• Citizen
• Fisherman
• Fish

IMPACT

Even though we can’t implement our project in an actual wastewater treatment system, we still wanted to make a difference! We decided on two areas where we could make an immediate impact: 1) Creating an policy brief to highlight current obstacles in effective nanoparticle regulation and propose new policy solutions, and 2) Raising funds for two organizations that promote environmental protection.

Policy Brief -- Nanoparticle Regulation Issues and Case Studies

Our team has conducted extensive research on existing regulatory laws and policies regarding nanoparticles and nanomaterials. We have investigated chemical regulations, including the Restriction, Evaluation, Authorization, and Restriction of Chemicals (REACH), A Toxic Substances Control Act (TCSA), CLP, and the Clean Air Act (CAA). There are significant obstacles to successfully regulating nanoparticles, such as conflicting definitions on nanoparticles that lead to an inability to successfully regulate manufacturers. Research has also been conducted on the hazardous effects of nanoparticles on the human body and environment. We decided to compose a policy brief highlighting the existing challenges in nanoparticle regulation and the lessons learned from previous failure to regulate new chemical substances. The brief was sent out to regulatory agencies, government agencies, and news outlets to raise awareness about the issue. We feel responsible to let others know about the damage nanoparticle waste can do to the environment. (Policy Brief created by Ashley L.)

We sent this policy brief to the Environmental Protection Administration (EPA) minister in Taiwan, and they responded! They read our policy brief and said that they will take it into consideration when they make policy regulations on the use of nanoparticles in the future. They understand that nanotechnology is still developing and definitely needs more attention and regulation. (Correspondence: Christine C.)

We were interviewed by News Lens International about nanoparticle regulation. Many of the questions focused on why we chose to target nanoparticles and how nanoparticles are dealt with in Taiwan. We emphasized that the lack of regulatory legislation prevents agencies from acquiring regulatory power. We also talked about the lack of nanoparticle filtration in wastewater treatment plants. (Interviewed by Ashley L.)

Fundraising and Donation

We held multiple fundraising sales, selling small ice cream dots (resembling nanoparticles!) and Oreo fudge during our lunch periods in school, and making “glitter slime” at our school’s annual spring fair (see Spring Fair in the Outreach section above). (Team activity)

In total, we raised around 500 USD, and donated the money to two organizations:

WaterisLife is an organization that provides clean drinking water, as well as sanitation and hygiene education programs to schools and communities in need. We donated to this organization in hopes that more people will have access to clean water. Visit WaterisLife here.

Taiwan Environmental Protection Union (TEPU) is a local organization founded in 1987 to promote public awareness and participation to prevent pollution and damage to public resources. Visit TEPU here.

OUTREACH

In Outreach, we raised awareness of the beneficial qualities and harmful consequences associated with nanoparticles. We also educated the general public about nanoparticle usage, synthetic biology, and science in general. Lastly, we communicated with other iGEM teams to share ideas and collaborate on experiments.

Education

Kindergarten -- Observing the “invisible”

Our iGEM team hosted over 120 kindergarten students to teach them the power of observation and the basics of science. For example, we taught them how to use microscopes to look at anti-counterfeiting measures on paper money and how to use refraction lenses to see that white light is made up of various colors. (Whole Team activity)

7th Grade Introduction to Synthetic Biology

We introduced iGEM and the basics of synthetic biology to all 200+ students in the seventh grade. They learned how to use micropipettes, as well as how to load and run dyes through an agarose gel. We also gave students different real world problems. Using paper biobrick parts, students put together constructs that would solve the given problems. (Whole Team activity)

Spring Fair -- Spreading Public Awareness of Nanoparticles

At our school’s annual spring fair, we manned a booth where people could create their own glitter slime by mixing polyvinyl alcohol and sodium borate solutions. The slime was meant to simulate the biofilm we use to trap nanoparticles (in this demo, glitter) in wastewater treatment plants. We also showed a few SEM images of bacteria, as well as everyday products that contain nanoparticles such as toothpaste and sunscreen. Everyone who came by our booth was encouraged to take our survey so we could record opinions on bioethics and concerns about nanoparticles. (Whole team activity)

Research Symposium -- Poster and Oral Presentations

At TAS we conduct research symposiums twice a year to showcase the research of students who take a variety of research courses. Before we decided our project topic, we developed 4 different project ideas to present at our first research symposium (poster session). We received feedback from both students and teachers, then decided on our current project. At our second research symposium, we presented on our current project, Nanotrap! (Presenters: Candice L., William C., Chansie Y., Christine C., Yvonne, W., Justin Y., Dylan L., and Catherine Y.)

5th Annual Asia-Pacific iGEM Conference -- NCTU

In preparation for the Giant Jamboree, we attended the 5th annual Asia-Pacific iGEM conference at NCTU to share and receive valuable feedback from other college and high school teams in Taiwan. This event allowed us to consider different aspects of our project using feedback from other teams. (Presenters: William C., Yvonne W., and Justin Y.)

Public Outreach -- A Tour of Taipei

Some members of the iGEM team went to various popular sites in Taipei to pass out fliers and conduct surveys. We visited National Taiwan University, Chiang Kai-Shek Memorial Hall, and Taipei 101. This helped us collect feedback from different age groups and backgrounds. This was a great and fun way to spread awareness of nanoparticle pollution! (Team members: Ashley L., Emily C., Florence L., Candice L., Yvonne W., Justin Y., Avery W., Christine C., Jesse K., and Laurent H.)

Here's a video we made for this event.

Collaborations

NYMU_TAIPEI

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CGU_Taiwan

We first met the CGU_Taiwan team at the end of our presentation for the 5th Annual Asia Pacific iGEM Conference hosted in National Chiao Tung University (NCTU). They were excited that our biofilms were able to trap nanoparticles and wanted to know whether they might trap ink particles as well. We offered to test this for CGU_Taiwan.

CGU_Taiwan also helped us independently test biofilm production using a different dye, crystal violet. Their results verified that overexpression of OmpR234 (BBa_K2229200) produces more biofilm than control (BBa_K342003).

Human Practices Text Written by Christine C., Candice L., Emily C., Justin Y. Edited by advisors Jude Clapper and Teresa Chiang.

REFERENCES

Ahamed, M., Alsalhi, M. S., & Siddiqui, M. (2010). Silver nanoparticle applications and human health. Clinica Chimica Acta,411(23-24), 1841-1848. doi:10.1016/j.cca.2010.08.016

Marr, L. C., & Holder, A. L. (2013). Nanomaterial disposal by incineration. Environmental Science: Processes & Impacts, 15(9), 1652-1664. https://doi.org/10.1039/C3EM00224A