Genetically modified crop cultivation's reduction of soil's decomposition capacity and consequent implications for water pollution *Young Suk SONG1 1Seoul International School, 15, Seongnam-daero 1518beon-gil, Sujeong-gu, Seongnam-si, Gyeonggi-do 461-830, Republic of Korea

ABSTRACT: The increasingly widespread cultivation of genetically modified crops (abbreviated as GM crops) has led to mounting concern over its environmental impacts. Research in the field of genetic engineering has so far succeeded in identifying the harms that GM crops pose on other organisms in the ecosystem, especially on non-GM crops. However, the literature has not yet fully explored the multifaceted causes of these environmental damages, nor the overreaching impacts that GM crop cultivation has on other ecosystems. Such uncertainty in the discussion has led to a lack of consensus on how restrictive the government should be on GM crop cultivation and whether GM crops will ultimately have a net positive or negative effect on the world. This study serves to bring more clarity to the discussion in two ways: first, by revealing one of the fundamental reasons GM crops harm other organisms in their ecosystem, and second, by establishing an unprecedented correlation between GM crop cultivation and water contamination. Through a series of eight comprehensive experiments involving the comparison of GM and non-GM soybeans, the inoculation of Nutrient Agar and Nutrient Broth culture mediums, and water quality tests, the study proves that GM crop cultivation leads to a significant reduction in the number of soil bacteria, which diminishes the soil’s decomposition capacity and leads to extremely high levels of nitrogen in the soil. The study also demonstrates that when the soil enters groundwater or nearby rivers through soil run-off, the excessive levels of nitrogen in the soil lead to eutrophication and red tides, ultimately causing severe water contamination. By demonstrating how GM crop cultivation poses significant threats to nearby aquatic ecosystems, the study aims to warn policymakers worldwide of these dangers and to encourage scientists to re-engineer GM crop seeds to minimize the negative impacts on other organisms and the aquatic ecosystem. Until then, the safest and most responsible way to deal with the situation would be to curtail the cultivation of GM crops to preserve the quality and purity of the soil and water.

1. INTRODUCTION

Since its appearance in 1983, genetically modified (GM) food has been a topic of wide controversy worldwide. Its purported benefits, according to International Union of Nutritional Sciences, range from increased crop yields and disease resistance to the enhancement of food's nutritional values. [1] Although many of these benefits may be well-substantiated by scientific evidence, other findings directly challenge

*Correspondence to : Young Suk SONG (youngsuk0598@gmail.com)

APEC Youth Scientist Journal Vol. 8, No. 2, August, 2016, pp. 95~112 http://www.sigs.or.kr

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the safety and benefits of GM foods. For instance, it is claim that GM foods pose various harms to the human health in the form of food allergies and toxic substances. One study conducted by the Department of Obstetrics and Gynaecology of University of Sherbrooke Hospital Center revealed that Bt toxins that are present in most GM crops remain as residue not only in humans but also in unborn babies in the womb, which can potentially lead to “allergies, miscarriage, abnormalities, or even cancer”. [2] Another study refuted the nutritional superiority of GM foods by demonstrating that GM soybeans generate lower levels of phytoestrogen compounds, a nutrient believed to help protect against heart disease and cancer. [3]

This clash in view is well-embodied in the widespread controversies concerning the implementation of GM crop biotech laws. Since 2013, for instance, thousands of Ugandan farmers and activists have protested against the adoption of GM crops because they believed that it would not only ruin the methods of traditional agriculture, but also damage people's health. On the other hand, the Uganda National Council of Science and Technology dismissed these people's fears as “devoid of scientific evidence” and proposed a GM food law to the government. Although still controversial, the tide has been turning in favor of the GM food advocates; in July 2015, the Ugandan government's ruling party has agreed to pass the National Biotechnology and Biosafety Bill, which would allow and regulate the production, import, and export of GM foods. This announcement devastated thousands of Ugandan citizens who hoped to keep their nation GMO-free.

Such conflicts can be found in many other countries, including South Korea, Philippines, Kenya, China, France, and the United States. These controversies, though unfortunate, are also quite understandable, because there are an overwhelming number of factors that the government must take into consideration when deciding whether to implement GM biotech policies: political stakes, economic projections, environmental consequences, people's health, public opinion, and so on. The multitude of scientific studies that often contradict each other only muddles the case even further. This study thus aims to provide clarity to the discussion in two ways: first, by revealing one of the fundamental reasons GM crops harm other organisms in their ecosystem, and second, by establishing an unprecedented correlation between GM crop cultivation and water contamination.

The study hypothesizes that the cultivation of GM crops will reduce the soil's capacity to decompose organic matter, which will lead to dangerously high levels of nitrogen content in the soil and ultimately cause water contamination in surrounding aquatic ecosystems. If the hypothesis is to be proven true, it would establish a clear correlation between GM crop cultivation and water pollution, and would provide anti-GM crop activists with a powerful tool in arguing for a stricter regulation or ban on GM crop cultivation.

2. MATERIALS & METHODS

2.1 Comparison of growth between GM and non-GM beans in fertile soil

To begin, 21 GM soybeans were obtained and washed with alcohol. Then, six plastic pots were each filled three-fourths with fertile soil, and a measuring cylinder was used to pour 300 millilitres (mL) of deionized water into each pot to provide sufficient moisture for the soil. Afterwards, a varying number of GM beans—from one to six beans per pot—were planted in the plastic pots. The same process was immediately repeated using non-GM beans, so as to compare the growth of the GM and non-GM bean

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plants over a period of three weeks. After three weeks of cultivation in room temperature (25 ºC) and adequate sunlight, the leaves of the

GM and non-GM bean plants were collected and inserted into separate 1.5 mL microtubes. Next, a micropipette was used to insert 1 mL of 80% ethanol into each of the microtubes. These tubes were then stored in the refrigerator at 4 ºC for one hour, thereby extracting the plants' chlorophyll into the ethanol. Afterwards, 1 mL of this ethanol was inserted into a cuvette, and a UV-spectrophotometer was used at a wavelength of 645 and 663 nanometers (nm) to measure the absorbance rate of GM and non-GM soybean plants, which yielded the amount of chlorophyll a and b.

Figure 1. GM and non-GM beans were obtained and planted in fertile soil to assess the difference in growth between the two types of beans.

2.2 Comparison of growth between cabbages cultivated in GM and non-GM bean’s soil

10 cabbage seeds were planted in each of the 12 plastic pots in which GM and non-GM bean plants were growing. Three weeks later, the leaves of the cabbage plants—both those grown with GM beans and those grown with non-GM beans—were collected and inserted into separate 1.5 mL microtubes. Using a micropipette, 1 mL of 80% ethanol was inserted into each microtube. These microtubes were then stored in the refrigerator at 4ºC for an hour, thereby extracting the plants' chlorophyll into the ethanol. 1 mL of this ethanol was extracted and inserted into a cuvette, and the UV-spectrophotometer was again used at a wavelength of 645 nm and 663 nm to measure the absorbance rate of the cabbage leaves, which yielded the amount of chlorophyll a and b.

2.3 Impact of GM and non-GM beans’ protein extractions on the growth of cabbage

The GM and non-GM soybeans were immersed in deionized water in a 50 mL conical tube. After the beans have thoroughly absorbed the water, a mixer was used to blend them completely. Then, using a spatula, the soybean solution was placed into a 50 mL conical tube. The tube was placed in the centrifuge, which was set at 6,000 rotations per minute. After the solution had been stratified in the centrifuge, the

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supernatant was extracted and inserted into a new 50 mL conical tube. Subsequently, a plastic pot was filled about three-fourths with fertile soil. A measuring cylinder was

then used to add 300 mL of deionized water to the pot. When the soil had been thoroughly hydrated, a tube was inserted into the center of the soil. Using this tube, 500 ㎕ of the supernatant containing the soybean protein were injected into the soil daily. After a week, once the solution had been absorbed into the soil, 0.1 g of the soil was gathered and placed into a 1.5 mL microtube.

Next, the 1.5 mL microtube was moved to the clean bench and a micropipette was used to add 1 mL of sterilized distilled water to the microtube. The vortex mixer was then used to mix the sterilized distilled water and the soil. The microtube was placed into the centrifuge and run at 6,000 rotations per minute for five to ten minutes, separating the soil from the supernatant containing soil bacteria. Then, a micropipette was used to extract 5 ㎕ of the supernatant and inoculate the NA culture. A sterilized spreader was used to rub the supernatant onto the surface of the culture. Subsequently, the inoculated NA culture was placed an incubator at 30 ºC for 12 hours. After 12 hours, the cultivated NA medium was removed from the incubator and the bacteria colony that had been generated was carefully observed.

Then, on the clean bench, 3 mL of NB culture was inserted into a 15 mL conical tube. A micropipette was used to add 5 ㎕ of supernatant to the conical tube, and the solution was cultivated in a stirred incubator at 30 ºC for 12 hours. After cultivation, 1 mL of the NB medium was gathered, and its optical density absorbance was measured using a UV-spectrophotometer in order to assess the amount of soil bacteria.

2.4 Growth and variety comparison of soil bacteria in GM and non-GM beans’ soils

Before starting the experiment of comparing the growth and variety of soil bacteria that exist in GM and non-GM beans' soils, a preparatory step is needed to produce Nutrient Agar (NA) and Nutrient Broth (NB) culture mediums, on which the soil bacteria are going to later be cultured. If you have already prepared the NA and NB mediums, then please proceed to Section 2.4.2.

2.4.1 Preparation of NA and NB culture mediums First, an electronic scale was used to measure out 23 g of Peptone Dickinson Korea Inc.'s Difco™

Nutrient Agar (ingredients: beef extract 3.0 g, Peptone 5.0 g, agar 15.0 g), which was then placed into a conical flask. Take care to avoid spilling the culture powder on the lip of the flask. Next, a measuring cylinder was used to add 1 L of deionized water to the same conical flask. A magnetic bar was also placed inside the flask, carefully so as to avoid damaging the flask. The flask was then sealed using aluminium foil, and a special tape was attached to the flask, indicating exposure to high temperature and high pressure by showing black stripes. Afterwards, the flask was placed above a magnetic stirrer, and the Nutrient Agar powder and the deionized water were mixed by running the magnetic stirrers and causing the magnetic bar within the flask to rotate. The solution was then mixed for 20 minutes, while taking care to avoid excessive frothing. Then, the flask containing the mixed solution was placed into the autoclave and run at 121 ºC and 2 atm for 15 minutes.

The sterilized conical flask was then removed from the autoclave and placed once more onto the magnetic stirrer. The magnetic bar was set into rotation by running the stirrer, allowing the NA culture to cool evenly. Should the solution be left to cool at room temperature, the conical flask, in coming into contact with air, would cool from the outside first. This would lead to the hardening of the NA culture

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medium's outer surface, causing unbalanced composition. Therefore, it is essential to cool the solution using a magnetic stirrer.

As the culture medium in the conical flask cooled down and the speed of the magnetic bar's rotation decreased, 70% alcohol was applied to the outer surface of the flask for sterilization purposes, and the flask was removed to a clean bench. Next, the aluminium foil was removed from the flask and poured into separate petri dishes, filling each dish two-thirds.

Afterwards, the petri dishes were set under the ultraviolet lamp. Avoiding contamination, NA culture medium was left to harden from its liquid state until it reached a semi-solid state. Once the NA completely reached a semi-solid state, the lids of the petri dishes were closed, wrapped in plastic wraps, and stored in the refrigerator at 4 ºC, in order to prevent contamination from micro-organisms. In addition, in order to ensure that any water droplets produced by condensation due to the difference in temperature between the refrigerator and the dish does not come into contact with the culture medium, petri dishes were stored upside down.

Unlike the NA culture medium, Peptone Dickinson Korea Inc.'s Difco™ Nutrient Broth (ingredients: beef extract 3.0 sag, and Peptone 5.0 g) does not contain any agar ingredients, and it is a medium for culturing bacteria in liquid form. Thus, when dealing with nutrient broths, a different set of procedures was followed: 8 g of the NB culture medium powder was measured using an electric scale, and then 1 L of deionized water was added. Following the process of sterilization, the mixture was divided into 50 mL conical tubes and refrigerated at 4 ºC until usage, as shown in Figure 2.

Figure 2. The finished products of NA and NB culture mediums were stored in the refrigerator at 4 ºC until usage.

2.4.2 Comparison of soil bacteria that exist in GM and non-GM beans' soils After the preparation of NA and NB culture mediums, an electronic scale was used to measure and

collect 0.1 g of both the GM and non-GM beans’ soils. These soil collections were placed in 1.5 mL microtubes, which were then moved to a clean bench. On the clean bench, a micropipette was used to insert 1 mL of deionized water in each tube. After closing the lids of the microtubes, a vortex mixer was used to properly and efficiently mix the soil and deionized water. Afterwards, the 1.5 mL microtubes were placed into a centrifuge, which was set at 6000 rotations per minute. The centrifuge was run at this speed for approximately 10 minutes to stratify the solution into soil and supernatant containing soil bacteria. Then, on a clean bench, micropipettes were used to collect the supernatant and move it into new 1.5 mL microtubes. These microtubes were then immersed in 40 ºC deionized water for approximately 10 minutes to activate soil bacteria.

Next, the NA and NB cultures that had been kept in the refrigerator at 4 ºC from Step 2.3 were taken

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out of the refrigerator and left in the room until they reached room temperature; when the cultures had fully reached room temperature, they were moved to the clean bench. A micropipette was then used to inject into the NA culture 5 ㎕ of the supernatant, which contained activated soil bacteria. Then, a sterilized spreader was used to spread the supernatant onto the surface of the NA medium. Afterwards, the NA medium was placed in an incubator and cultivated for 12 hours at a temperature of 30 ºC. After cultivation, the NA culture was removed from the incubator and the colony of soil bacteria was carefully assessed. Then, 3 mL of the NB medium were placed into a 15 mL conical tube on a clean bench. Similar to the aforementioned procedure for the NA medium, a micropipette was used to add to the NB medium 5 ㎕ of the supernatant containing activated soil bacteria. Subsequently, the NB medium was cultivated in a stirred incubator at 30 ºC for 12 hours. After cultivation, 1 mL of the NB medium was gathered and its optical density absorbance was measured using a UV-spectrophotometer in order to assess the amount of soil bacteria.

Figure 3. The soil bacteria of GM and non-GM bean plants were cultivated on the NA and NB culture mediums and assessed using the UV-spectrophotometer.

2.5 Comparison of E. coli proliferation rate in soils of GM and non-GM beans

An electronic scale was used to measure out 3 g of soil in which GM beans were cultivated. The 3 g of soil were then placed on a small petri dish and inoculated with 100 ㎕ NB culture cultivated with E. coli. The soil was then placed in an airtight container. After storing at room temperature (25 ºC) for a week, it was placed onto a clean bench and 0.1 g of the soil was relocated into a 1.5 mL microtube with 1 mL of deionized water. Next, this microtube was placed in the centrifuge at 6000 rotations per minute for five to ten minutes. After the solution had stratified, the NA and NB cultures were inoculated with 5 ㎕ of the supernatant and cultivated in a temperature-regulated stirring incubator at 37ºC for 12 hours. After cultivation, 1 mL of the NB medium was gathered, and its optical density absorbance was measured using a UV-spectrophotometer in order to assess the amount of soil bacteria. Afterwards, the procedure of this experiment was repeated once more using soils of non-GM beans instead of GM beans.

2.6 Impact of GM and non-GM bean cultivation on soil’s decomposition capacity

An electronic scale was used to measure out 3 g each from GM and non-GM bean's soils. The soils were then placed in small petri dishes and inoculated with 9 mL of deionized water. After the soil absorbed the water, it was inoculated with 1% urea solution and stored for one week at room temperature (25 ºC). After a week, sterilized equipment was used to place the soils inoculated with urea in 50 mL conical tubes. 5 mL of sterilized distilled water were added and properly mixed using the vortex mixer. Next, the conical tubes were placed in the centrifuge and run for 10 minutes at 6,000 rotations per minute

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for five to ten minutes. Following stratification of the solutions, 1 mL of each supernatant solution was gathered and placed in new 1.5 mL microtubes. Using an EZ WM test kit, a water quality indicator that measures the concentration of different nitrogen compounds and the water's chemical oxygen demand and dissolved oxygen levels as shown in Figure 4, the changes in the concentration of nitrogen content were carefully assessed.

Ammonium Nitrogen (NH4-N) 0-5 ppm (9 levels: 0/0.1/0.2/0.4/0.8/1.2/2/3/5)

Nitrite Nitrogen (NO2-N) 0-1 ppm (9 levels: 0/0.025/0.05/0.1/0.2/0.4/0.6/0.8/1)

Nitrate Nitrogen (NO3-N) 0-240 ppm (9 levels: 0/0.5/1/5/10/15/30/50/80)

Chemical Oxygen Demand (COD) 0-20 ppm (9 levels: 0/2/3/4/5/8/10/15/20)

Dissolved Oxygen (DO) 0-10 ppm (7 levels: 0/1/2/4/6/8/10)

Figure 4. EZ WM Test Kit's Components and Respective Scales

2.7 Effects of GM and non-GM beans’ soils on the growth of phytoplankton

1 g each of GM and non-GM bean soils was added to its respective 50 mL conical tube. After 10 mL of deionized water were added to each tube, a vortex mixer was used to properly mix both solutions. Then, the tubes were placed in the centrifuge at 6,000 rotations per minute for five to ten minutes, stratifying the deionized water and the supernatant. Subsequently, the supernatant was extracted and placed into new 50 mL tubes. 100 ㎕ of the supernatant containing phytoplankton was placed on a small petri dish, and 5mL of sterilized distilled water was added to the solution. The petri dishes were then stored under controlled lighting in order to grow the phytoplankton. Next, the petri dishes were inoculated with phytoplankton everyday with 100 ㎕ of supernatant from GM and non-GM bean soils for three weeks. After three weeks, 1mL of the phytoplankton culture was extracted and carefully observed. Finally, the optical density absorbance of the soils was measured using a UV-spectrophotometer at 645 nm and 663 nm in order to assess the increase in phytoplankton.

2.8 Effect that introduction of GM beans soil to water has on water quality

1 g each of GM and non-GM bean soils was added to its respective 50 mL conical tube. After 10 mL of deionized water were added to each tube, a vortex mixer was used to properly mix both solutions. Then, the tubes were placed in the centrifuge at 6,000 rotations per minute for five to ten minutes, stratifying the deionized water and the super-natant. Subsequently, the supernatant was extracted and placed into new conical tubes. Then, everyday for three weeks, 100 ㎕ of this supernatant was inoculated to a petri dish containing 5mL of water from Yangjae Stream, an ordinary local stream. Using the EZ WM test kit once again, changes in the water's COD (chemical oxygen demand) level and DO (dissolved oxygen) level were carefully assessed.

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3. RESULTS & DISCUSSION

3.1 Comparison of growth between GM and non-GM beans in fertile soil

Three weeks of observing the growths of GM and non-GM soybeans in fertile soil demonstrated that the GM soybeans were healthier in terms of both growth status and germination rate. As can be seen in Figure 5, it could be observed from the GM bean plants' longer stems, larger leaves, and more developed roots that GM beans generally grew better than non-GM beans.

Figure 5. The growth of non-GM and GM soybeans three weeks after planting in fertile soil

Since one cannot precisely assess the difference in plant growth through the plants' outer appearance alone, the plants' chlorophyll contents were measured using the UV-spectrophotometer to make a more objective comparison. Chlorophyll is known to be an essential element to plant life because it is the most important pigment in the photosynthesis process. [4] It is thus fair to say that the greater the amount of chlorophyll the plant contains, the more healthily it will grow. Results depicted in Figure 6 showed that non-GM beans had lower levels of chlorophyll than did GM beans, thereby proving that GM beans are statistically more likely to see healthier and more active growth in a controlled environment. The fact that GM beans saw healthier growth than non-GM beans despite the identical conditions of cultivation (amount of water applied daily, amount of sunlight, amount of soil, size of pot) verified the widely-

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supported notion that the genetic modification of GM crops does indeed enhance their growth.

Figure 6. Comparison of growth in GM and non-GM beans in fertile soil by comparing their respective

absorbance rates of chlorophyll

3.2 Comparison of growth between cabbages cultivated in GM and non-GM bean’s soil

Three week after planting GM and non-GM soybeans, cabbage seeds were added to both soils. Observation of the cabbage plants' growth for three weeks revealed that the cabbages planted alongside non-GM soybeans experienced a normal and healthy state of growth, with straight stems, green leaves, and long roots, as depicted in Figure 7. On the other hand, cabbages grown alongside GM beans experienced a clear lack of growth, with flimsy stems, yellow leaves, and relatively short roots.

Figure 7. Changes in the growth of cabbage grown in the soil of non-GM and GM soybeans

This qualitative data was corroborated with statistics acquired using the UV-spectrophotometer. Data demonstrated in Figure 8 showed that the non-GM soybeans and the cabbages that were grown alongside

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had an absorbance rate approximately four times higher than those of GM beans and their adjacent cabbages.

These results prove that, although GM beans grow better than non-GM beans in an isolated environment as demonstrated in the previous experiment, the situation is reversed when other plants are introduced to the same ecosystem. In other words, the data suggest the notion that GM beans have a negative impact on the growth and nutrition of nearby organisms and possibly even on the soil that they have been raised in.

Figure 8. A comparison of changes in growth of cabbages grown in soil with normal soybeans and with cabbages grown in soil with GM soybeans by comparing their chlorophyll levels

3.3 Impact of GM and non-GM beans’ protein extractions on the growth of cabbage

One group of cabbages was grown in soil injected with protein extracted from GM beans, while the other group was grown in soil injected with protein extracted from non-GM beans. A week of observation and daily injection of protein revealed that the cabbages whose soil was injected with protein extracted from GM soybeans were withered, with yellow leaves and thin, flimsy stems. On the other hand, the cabbages whose soil was injected with protein extracted from non-GM beans experienced healthy and consistent growth, as depicted in Figure 9.

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Figure 9. Contrasting growths of cabbages whose soil was injected with proteins from non-GM beans (left)

and cabbages whose soil was injected with proteins from GM beans (right) A UV-spectrophotometer was used to compare the absorbance rates of proteins extracted from GM and

non-GM beans. Data depicted in Figure 10 showed that non-GM soybean proteins had an absorbance rate more than four times higher than that of GM soybean proteins.

These results strengthened the notion that GM beans harm their surrounding ecosystem, because the proteins of GM soybeans clearly had a detrimental effect on the growth of cabbages. The data collected in this experiment provided increased validity to the inference that the cultivation of GM crops causes a decline in the quality of soil.

Figure 10. Contrasting absorbance rates of proteins extracted from GM beans (left)

and those extracted from non-GM beans (right)

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3.4 Growth and variety comparison of soil bacteria in GM and non-GM beans’ soils

In the case of non-GM soybeans, regardless of the number of soybeans planted, be it as low as one or as many as six, there was little difference evident in the quantity and the variety of soil bacteria, as depicted in Figure 11. However, in the case of GM beans, the variety and quantity of soil bacteria declined as the number of GM beans planted increased.

Figure 11. Varieties and total quantities of soil bacteria present in the soils of GM and non-GM soybeans,

with the number of beans planted increasing from left to right (from one to six beans)

This qualitative data was again verified using the UV-spectrophotometer. The absorbance rate of GM beans were on an average 0.101 nm, whereas that of non-GM beans were more than five times higher, with an average of 0.548 nm, as depicted in Figure 12. Such data, both qualitative and quantitative, validated the notion that the cultivation of GM beans damages soil bacteria in both their variety and quantity.

Figure 12. Absorbance rates of soil bacteria present in the soils of GM and non-GM soybeans,

with the left six representing GM beans and the right six representing non-GM beans

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3.5 Comparison of E. coli proliferation rate between soils of GM and non-GM beans

After E. coli was introduced to soils of GM and non-GM beans, it was stored in an airtight container for a week and left to cultivate. The degree of E. coli proliferation in the soil of GM beans was significantly larger than that of non-GM beans, as demonstrated in Figure 13.

The data make it clear that the excessive and dangerous proliferation of E. coli in GM beans' soil occurred due to a lack of soil bacteria in the GM beans' soil that would have otherwise stopped the proliferation of E. coli as they did in the non-GM beans' soil. The data show that the damage inflicted upon soil bacteria by the GM crops significantly reduces the soil's capacity to decompose organic matter, which gives rise to dire environmental consequences as will be demonstrated in the following experiments.

Figure 13. Contrasting degrees of E. coli proliferation in soils of GM beans (left six) and non-GM beans (right six)

3.6 Impact of GM and non-GM bean cultivation on soil’s decomposition capacity

When organic matter such as urea is injected into non-GM soil, the soil bacteria play the role of breaking the organic matter down, thus maintaining a non-toxic state. If organic matter is not broken down properly, it can lead to unnaturally high levels of nitrogen content in the soil. Such was the case in Step 4.6, in which an experiment using the EZ WM test kit revealed significantly higher levels of nitrogen in the GM bean's soil. As demonstrated in Figure 14, the concentration of ammonium nitrogen rose from Level 1 (0 ppm) to Level 3 (0.2 ppm); the concentration of nitrite nitrogen rose from Level 2 (0.025 ppm) to Level 8 (0.8 ppm); and the concentration of nitrate nitrogen rose from Level 2 (0.5 ppm) to Level 5 (15 ppm). To put things in perspective, the maximum limit of nitrate nitrogen for human consumption is 10 ppm, and most surface and ground water ideally contains under 3 ppm of nitrate nitrogen (Agency for Toxic Substances and Disease Registry) [5], which is significantly lower than the 15 ppm of nitrate nitrogen observed in the experiment.

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Figure 14. Comparison of decomposition capacities of GM and non-GM soils using the EZ WM test kit. The more contaminated a solution is, the farther on the left it is placed.

Ammonium Nitrogen (NH4-N)

Nitrite Nitrogen (NO2-N)

Nitrate Nitrogen (NO3-N)

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These results thus demonstrate that a substantial decrease in the GM bean soil's ability to break down organic matter leads to high levels of nitrogen. This conclusion leads to a possible inference about the impact of the soil's reduced decomposition capacity: when the soil of GM crops, unable to breakdown organic matter, enters nearby rivers through soil run-off, the unprocessed nitrogen waste will also enter the aquatic ecosystem. This, by definition, is a phenomenon called eutrophication, in which an excess amount of nutrients exist in a body of water, and can lead to the disruption of the aquatic ecosystem and its inhabiting organisms.

3.7 Effects of GM and non-GM beans’ soils on the growth of phytoplankton

The inoculation of water containing phytoplankton with this solution and storing for three weeks was a dramatic increase in the number of phytoplankton, unlike the phytoplankton whose water was inoculated with a soil solution from non-GM beans. This data, as represented in Figure 15, shows the tangible consequence of a reduction of soil's decomposition capacity. A reduction in the soil's ability to decompose organic matter results in an excessive influx of nitrogen component, which causes a sharp rise in phytoplankton numbers as shown.

The sharp rise in phytoplankton numbers has a number of severe environmental consequences. It results in algal blooms, which disrupts the normal functioning of the aquatic ecosystem. An excessive amount of algae can consume all the oxygen in the water and block the marine plants' access to sunlight, causing trouble for the organisms coexisting in the ecosystem. [6]

Figure 15. Comparison of phytoplankton growth in water inoculated with soil solution from non-GM beans and that inoculated with soil solution from GM beans

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3.8 Effect that introduction of GM beans soil to water has on water quality

Tests confirmed that introduction of GM beans soil to water containing phytoplankton led to a large increase in the water's COD level from Level 1 (0 ppm) to Level 9 (20 ppm) and a sharp decrease in the water's DO level from Level 6 (8 ppm) to Level 1 (0 ppm), as depicted in Figure 16.

Figure 16. Comparison of COD and DO levels between liquid soil solutions taken from GM and non-GM beans

The COD test is an effective measure of water quality that indicates the mass of oxygen consumed per liter of solution. [7] Likewise, the DO test is also important in that it shows the amount of oxygen that is present in the water, which has a direct influence on the organisms living within the water. [8] The large increase in the water's COD level and the sharp decrease in the water's DO level show that the water solutions exposed to GM bean soil possess harmful and undesirable conditions for the organisms living in the aquatic ecosystem. These results together confirm the severity of the environmental consequences of GM crop cultivation, by conclusively proving the notion that GM crop cultivation leads to contamination of water.

Normal Soybeans GM Soybeans

COD Level

Normal Soybeans GM Soybeans

DO Level

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Genetically modified crop cultivation's reduction of soil's decomposition capacity and consequent implications for water pollution

4. CONCLUSION Through a series of eight comprehensive experiments, the study proved correct its hypothesis that GM

crop cultivation would not only harm other organisms within the ecosystem but also lead to severe water pollution. A source of error in the study was that the experiments were conducted in a regulated, laboratory environment. Although conditions such as moisturization and amounts of sunlight were adjusted to simulate the outdoor environment as much as possible, it was nevertheless not an accurate reflection of the outdoor environment, as the soybean plants were isolated in separate pots and were therefore not exposed to the full ecosystem. This error can be remedied by increasing the scale of the experimental setting by conducting the experiments directly on the outdoor fields, rather than in the laboratory. Although this enlarged scale may be challenging to pursue due to space and resource constraints, it is the best way one can be sure of the validity of the results.

Ultimately, the results of this study has significant implications for the future of water security. The study proves that mass cultivation of GM crops would inevitably lead to a severe water crisis with dire consequences, as the contaminated groundwater would be used for drinking water and irrigation. Thus, it is crucial that citizens understand the long-term harms that GM crop cultivation can have on the aquatic environment. It is particularly essential to raise awareness about this concern in developing countries, as GM crop cultivation is becoming an increasing trend among many. The citizens must realize that they cannot risk the prospect of contaminating their water supplies, which are already limited and scarce in many developing regions.

So what action can we take to reduce the environmental damages caused by GM crop cultivation? Short-term solutions, such as improving the quality of the GM crops' contaminated soil via soil replacement, can certainly be implemented, but would be insufficient and futile in the long run. Therefore, the ideal situation would be that scientists find a way to re-engineer GM crop seeds to reduce the harms inflicted upon surrounding organisms and the aquatic ecosystem. Until then, however, the safest and most responsible way to deal with the situation is to restrict and curtail the cultivation of GM crops in order to preserve the quality and purity of our environment.

5. REFERENCES

[1] International Union of Nutritional Sciences. Statement on Benefits and Risks of Genetically Modified Foods for Human Health and Nutrition. 2012. Available at: http://www.iuns.org/2012/05/statement-on-benefits-and-risks-of-genetically-modified-foods-for-human-health-and-nutrition/. Accessed November 24, 2015.

[2] Aris A, Leblanc S. (2010). Maternal And Fetal Exposure To Pesticides Associated To Genetically Modified Foods In Eastern Townships Of Quebec, Canada. 1st ed. Quebec: Elsevier; 2010. Available at: https://www.uclm.es/Actividades/repositorio/pdf/doc_3721_4666.pdf. Accessed November 23, 2015.

[3] Bakshi A. (2013). Potential Adverse Health Effects of Genetically Modified Crops. Journal of Toxicology and Environmental Health, Part B. 6(3), 211-226. doi:10.1080/10937400306469.

[4] Higdon J. (2004). Chlorophyll and Chlorophyllin | Linus Pauling Institute | Oregon State University. Linus Pauling Institute. 2004. Available at: http://lpi.oregonstate.edu/mic/dietary-factors/phytochemicals/chlorophyll-chlorophyllin. Accessed December 10, 2015.

[5] Environmental Health and Medicine Education. Nitrate/Nitrite Toxicity 2013: Cover Page. 2014. Available at:

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http://www.atsdr.cdc.gov/csem/csem.asp?csem=28. Accessed January 10, 2016. [6] Harmful Algal Blooms. EPA. Available at: https://www.epa.gov/nutrientpollution/harmful-algal-blooms. Accessed

January 7, 2016. [7] Chemical Oxygen Demand (COD) - Real Tech Inc. Real Tech Inc. 2014. Available at: http://realtechwater.com/

chemical-oxygen-demand/. Accessed January 14, 2016. [8] Kemker, C. (2013). Dissolved Oxygen - Environmental Measurement Systems. Environmental Measurement

Systems. 2013. Available at: http://www.fondriest.com/environmental-measurements/parameters/water-quality/dissolved-oxygen/. Accessed January 14, 2016.

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