Staples STEM Journal: Issue No. 3.5 by Staples STEM Journal

JUNE 2017

STAPLES

ISSUE NO. 3.5

STEM JOURNAL Student Research

DROUGHT RESISTANCE

BIOFUEL PRODUCTION STAPLES HIGH SCHOOL

NOCTURNAL MIGRATION OF BIRDS

Copyright © 2017 Staples High School STEM Journal P UBLISHED BY S TAPLES H IGH S CHOOL STEM J OURNAL Licensed under the Creative Commons Attribution-NonCommercial 3.0 Unported License (the “License”). You may not use this file except in compliance with the License. You may obtain a copy of the License at http://creativecommons.org/licenses/by-nc/3.0. Unless required by applicable law or agreed to in writing, software distributed under the License is distributed on an “AS IS ” BASIS , WITHOUT WARRANTIES OR CONDITIONS OF ANY KIND, either express or implied. See the License for the specific language governing permissions and limitations under the License. Printed for June 2017.

Contents

Letter from the Editor . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . A LICE S ARDARIAN ‘17

Drought Resistance . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . J ULIANA H OPPER ‘18

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Production of Methanol From Carbon Dioxide . . . . . . . . . . . . . . . . . . . . Z ACHARY D E B RINO ‘19 AND A NIRUDDHA M URALI ‘19

Nocturnal Migration of Birds . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . J ORY G. T ELTSER ‘19

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Biofuel Production . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . JASON K ATZ ‘19

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Antibiotics in Wastewater . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . J OSHUA Z HANG ‘19

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Journal Staff . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

1. Letter from the Editor

Dear Reader, Here we are again, one last issue for the year and one last letter from me, as Editor-in-Chief. I am thrilled to present this special edition, featuring student research proposals. They are quite impressive and certainly a testament to our great and high quality public education. It is bitter sweet saying goodbye, but I am confident that our new executive board will take the helms and continue the growth we began a mere two years ago. The STEM fields are vital to our advancement, so I encourage all students, who may excel and value the arts, to also participate in the STEM Journal. Ms. Thompson, you have been an inspiration and a phenomenal support; thank you for your faith and trust in me and our contributing students, and for your support throughout the entire process. To all those who contributed, graduating members, and underclassmen who will continue to transform and improve this publication, thank you for making our STEM Journal a success. It was simply a pleasure to work with and to learn from all of you. I would also like to extend an immense gratitude to the PTA for their financial support. You are truly visionary and we appreciate your services for our school. We are very fortunate to have a caring and supportive community of educators and parents. On behalf of Staples STEM Journal team members, I thank you all for having our best interest in mind. As a senior, I leave this great school with so much hope and with pride, and look forward to reading many more upcoming, exciting STEM Journal issues. Stay passionate, stay curious, and stay informed! Truly Yours,

Alice Sardarian â&#x20AC;&#x2DC;17, Founder & Editor-in-Chief

2. Drought Resistance

The Effect of EsMCsu1, from Eutrema Salsugineum, on Transformed Arabidopsis Related to Drought Resistance By Juliana Hopper ‘18

Abstract Due to the growing drought problem in California, it would be advantageous to transform crops to be more drought resistant. This would aid in maintaining enough water to support California’s large agricultural community. Arabidopsis, a model organism, can be transformed with the Eutrema Salsugineum gene EsMCsu1. This will allow it to go through more ABA synthesis, which closes the stomata to keep water that would be transpired through the leaves inside the plant, thus conserving water for future use. Transforming the arabidopsis can be achieved through a simplified agrobacterium transfer process (Weeks et. al. 2008). When the arabidopsis plants, transformed and wild type, reach 10% bloom, the water use efficiency (WUE) will be measured through the percent change in plant mass. Since arabidopsis is a model organism, the success of this transformation would be beneficial for farmers and water conservation efforts in the future because it could lead to all crops having the ability to use water more efficiently.

Introduction California, one of the United States’s top agricultural producers, has been faced with a monumental drought crisis for the past four years. Data shows that California is experiencing extreme drought conditions with 44.84% of the state categorized as exceptional drought, while at least 97.33% of California is categorized as moderate drought conditions (Figure 1) (Heim, 2015). Farmers are struggling to obtain enough water for their crops due to Governor Brown’s mandate to minimize water usage by 25% (CA.gov, 2015). As a result, many farmers are reducing their number of crops since there is not enough water to sustain them. Arabidopsis, from the Brassicaceae family, serves as the model organism for genetic transformation research because it has a small genome and short growth cycle. The genome size of the

Chapter 2. Drought Resistance arabidopsis is 125 Mb (Bennett, 2003), which is small compared to the size of the genome of most cereal crops, such as rice, that has a genome of 389 Mb. The growth cycle for arabidopsis from seed germination to the first seed pod shattering is completed in 48 days (Swarbreck et al., 2008), which is faster than the growth cycle of other cereal crops. These two variables cause the transformation of arabidopsis to be a faster and more efficient process compared with the transformation of other plants. The extremophile eutrema salsugineum has a high tolerance against abiotic stresses, including drought resistance. The organism also shares a lineage and similar morphology with the model organism arabidopsis, but it also has certain protein-coding genes that respond to abiotic stress, which are not inherited from the arabidopsis genome (Kazachkova et al., 2013). One of these genes, EsMCsu1, is the master regulator for the synthesis of abscisic acid (ABA), which regulates the transpiration of eutrema salsugineum through the management of the osmotic pressure of the paired stomata guard cells. When the cells remain turgid, the stomata are open. When ABA binds to the membrane of the guard cells, it causes several cell receptors to activate interconnecting pathways that raise the pH of the cytosol and releases stored Ca2 + from the vacuoles into the cytosol. These changes in the cell result in a loss of anions and K+ in the cell, which reduces the osmotic pressure of the stoma, thus causing it to close. This allows the eutrema salsugineum to retain more water, as less is being lost through transpiration. In an experiment to improve alfalfa’s tolerance to drought conditions, the tested alfalfa were transformed by an agrobacterium method to express the gene EsMCsu1. The results demonstrated that the ABA content of the transgenic alfalfa lines was 25.9% higher for TG2, and 39.5% higher for TG7 compared to the WT alfalfa in response to drought (Figure 2). The transgenic plants with higher ABA contents also demonstrated increased drought resistance because after eight and sixteen days of drought treatment, the leaf water potential and relative water content were significantly lower in the WT plants compared to both TG2 and TG7 (Zhou et.al, 2015) (figure 3). In a similar way, arabidopsis can be transformed through an agrobacterium method to express the eutrema salsugineum gene, EsMCsu1, in order to increase ABA synthesis and conserve the water lost through transpiration. The transformation of arabidopsis to be more drought resistant with the gene EsMCsu1 will demonstrate that this gene is a viable option to transform agriculturally important crops across the board to be more drought resistant. Based on the success of previous research conducted with this gene, transforming arabidopsis to overexpress the gene EsMCsu1 will result in enhanced drought resistance through increased ABA synthesis.

Methods The total RNA will be extracted from eutrema salsugineum by the Japanese reagent TRIzol, then will be reverse transcribed into cDNA to act as a template for PCR analysis, where the gene EsMCsu1 will be identified and separated. Next, the gene will be cut and inserted into the plasmid pBI121(Figure 4). The agrobacterium strain, GV301, will need to be suspended in 100 mL Luriabertani (LB) liquid medium (pH 7.0) containing 50 g mL–1 kanamycin, 50 g mL–1 gentamicin, and 25 g mL–1 rifampicin overnight at 28°C under constant rotation at 200 rpm. After suspension, the agrobacterium will go through electroporation in order to transfer the vector into the bacterium (Wu et al., 2015). Via the simplified process of transformation outlined by Weeks et. al. 2008, 80 arabidopsis seedlings will be sterilized when vortexed in a solution of ethanol and bleach. After the seeds are sterilized they will be germinated in Murashige and Skoog with 1% sucrose for 2 days in the dark at 24°C. Prior to transformation the seeds, will be cold treated at 4°C for 16 hours. The prepared seedlings will be placed in a 50mL centrifuge tube along with 15 mL of the suspended agrobacterium that was prepared with the cloning vector in the previous section, 20mL of LB liquid medium that is inoculated with antibiotics and agrobacterium, and 2.4g of white quartz sand. The

contents of the centrifuge tube will be stirred on the highest setting on the vortex genie II for 30 minutes at room temperature. The control seeds will use the same procedure except the suspended agrobacterium solution will be replaced with distilled water. The transformed and WT seeds will first be cultivated on a Murashige and Skoog (MS) medium for 14 days with a 16 hour photoperiod at 23Â°C. The healthiest cultivated plants, 20 transgenic and 20 WT, will be planted in a soil mixture of vermiculite, perlite, and peat moss (v/v, 1:1:1) in a greenhouse set at 23Â°C, with 16 hours of artificial light (26Â°C) per day. The plants will also be in well watered conditions for the first 3 weeks. Prepare the same artificial soil mixture as above, in individual plastic cylindrical pots that are 8 cm diameter and 10 cm deep (Bao et al., 2015). Measure the mass of each pot with the soil in it, and then transplant the 20 transgenic arabidopsis plants and 20 WT arabidopsis plants. All arabidopsis plants will start testing under well watered conditions for 1 week. A pilot test will be performed to determine the quantity and the frequency of the watering that meets the criteria of well watered conditions. Every day during data collection, each plant will be weighed and the percent change in mass will be calculated. Next, withhold water from the plants, and continue to weigh the plants and calculate the percent change in mass. After all the data is collected a T-test will be performed, with a p value of 0.05, to determine if the transgenic arabidopsis is significantly more tolerant to drought conditions compared to the WT arabidopsis.

Implications If the transformation of the EsMCsu1 gene in the arabidopsis is successful, there could be economic, environmental, and agricultural benefits in the alfalfa industry. California is the largest agricultural state in the U.S because 43 million acres out of the 100 million acre state are used for agriculture (Thompson, 2009). If this transformation can be utilized across the board for crops that require large amounts of water, such as almonds and produce, farmers will still be able to grow a diverse amount of crops under drought conditions, which will keep our agricultural economy stable. Farmers will also save money on watering drought resistant crops since the cost for an irrigation system in large agricultural farms is $70 per acre of water (Schaible & Aillery, 2013). Drought resistant crops also benefit the environment because they use water more efficiently through the expression of EsMCsu1, which increases ABA synthesis in order to close the stomata and limit the amount of evapotranspiration that occurs. The heightened efficiency of the transgenic crops will allow Californians to use the water they were previously using for farming for other circumstances, which will lower the risk of California running out of water while drought conditions persist. Drought tolerant crops can benefit the agricultural community by offering farmers, who live in more arid climates, the opportunity to grow certain species that were once out of their reach. Current farmers can also benefit by introducing transgenic crops into their farms because they will be able to yield the same amount of crops in drought conditions as they would in optimal growing conditions.

Chapter 2. Drought Resistance

Figure 2.1: Map of drought intensities in California measured in fall of 2015. The scale ranges from abnormally dry to exceptional drought (Heim, 2015).

Figure 2.2: The ABA content of the WT, TG2, and TG7 lines measured every 8 days for 16 days during drought treatment (Zhou et.al, 2015).

Figure 2.3: The LWP and RWC concentrations in WT, TG2, and TG7 alfalfa plants after 0, 8, and 16 days of drought treatment (Zhou et.al, 2015).

Figure 2.4: The Vector pBI121 1302 and the restriction enzymes and where they will be used (pBI121, 2017).

Chapter 2. Drought Resistance

References Bao, A., Du, B., Touil, L., Kang, P., Wang, Q., & Wang, S. (2015). Co-expression of tonoplast Cation/H+ antiporter and H+-pyrophosphatase from xerophyte Zygophyllum xanthoxylum improves alfalfa plant growth under salinity, drought and field conditions. Plant Biotechnology Journal, 1-12. Doi:10.1111 Bennett, M. D., Leitch, I. J., Price, H. J., & Johnston, J. S. (2003). Comparisons with Caenorhabditis ( 100 Mb) and Drosophila ( 175 Mb) using flow cytometry show genome size in Arabidopsis to be 157 Mb and thus 25% larger than the Arabidopsis genome initiative estimate of 125 Mb. Annals of botany, 91(5), 547-557. Governor Brown Issues Executive Order to Bolster Stateâ&#x20AC;&#x2122;s Drought Response. (2015, November 13). Retrieved November 15, 2015, from http://ca.gov/drought/topstory/top-story-50.html Heim, R. (2015, November 10). U.S. Drought Monitor California. Retrieved November 15, 2015, from http://droughtmonitor.unl.edu/Home/StateDroughtMonitor.aspx?CA Kazachkova, Y., Batushansky, A., Cisneros, A., Tel-Zur, N., Fait, A., & Barak, S. (2013). Growth platform-dependent and-independent phenotypic and metabolic responses of Arabidopsis and its halophytic relative, Eutrema salsugineum, to salt stress. Plant physiology, 162(3), 15831598. pBI121 [Map; PDF]. (2017, February 15). Schaible, G., & Aillery, M. (2013, September 2). Western Irrigated Agriculture: Production Value, Water Use, Costs, and Technology Vary by Farm Size. Retrieved January 4, 2016. Swarbreck, D., Wilks, C., Lamesch, P., Berardini, T. Z., Garcia-Hernandez, M., Foerster, H., ... & Radenbaugh, A. (2008). The Arabidopsis Information Resource (TAIR): gene structure and function annotation. Nucleic acids research, 36(suppl 1), D1009-D1014. Thompson, E., Jr. (2009, July). Agricultural Land Loss & Conservation. Weeks, J. T., Ye, J., & Rommens, C. M. (2008). Development of an in planta method for transformation of alfalfa (Medicago sativa). Transgenic Research, 17(4), 587-597. Retrieved December 15, 2015. Wu, G., Feng, R., Wang, S., Bao, A., Wei, L., & Yuan, H. (2015). Co-expression of xerophyte Zygophyllum xanthoxylum ZxNHX and ZxVP1-1 confers enhanced salinity tolerance in chimeric sugar beet (Beta vulgaris L.). Frontiers in Plant Science. Retrieved January 7, 2016, from NCBI. Zhou, C., Ma, Z. Y., Zhu, L., Guo, J. S., Zhu, J., & Wang, J. F. (2015). Overexpression of EsMcsu1 from the halophytic plant Eutrema salsugineum promotes abscisic acid biosynthesis and increases drought resistance in alfalfa (Medicago sativa L.). Genetics and molecular research: GMR, 14(4), 17204.

3. Production of Methanol From Carbon Dioxide

The Effect of Different Catalysts on the Production of Methanol From Carbon Dioxide By Zachary DeBrino ‘19 and Aniruddha Murali ‘19

Abstract Two major issues facing our planet today are climate change and the decrease in the supply of energy for production of electricity. The increase in global temperature caused by high carbon dioxide levels is hurting the environment. There are currently no defined solutions that efficiently return a large supply of energy without harming the environment, but converting carbon dioxide to fuel appears to be a possible solution. In this experiment, carbon dioxide will be directly converted into methanol (CO2 + 3H2 → CH3 OH + H2 O). Six catalysts (silicon, cobalt, copper, iron, nickel, and zinc) will be tested to see which catalyst yields the highest rate of reaction. Hydrogen gas will be used in combination with carbon dioxide to produce methanol and water. Distillation will be used to separate the methanol from the water at the end of each trial. By converting carbon dioxide into methanol, the amount of carbon dioxide in the atmosphere will decrease, and there will be an efficient way to produce energy without harming the environment.

Introduction The temperature of the globe has been rising over time. The presence of active gasses in the atmosphere, like carbon dioxide, has increased the global surface temperature (Mitchell, 2010). It also leads to more extreme weather conditions, more ice melting, sea levels rising, and acidification of the oceans (Bradford, 2014). Its effects are profound: plants and animals are both migrating away from the equator, warmer temperatures are expanding the range of many disease-causing pathogens, farmers are experiencing crop failures and livestock shortages, there is a loss of food security, and the number of cases of disease are on the rise (Bradford, 2014). One of the biggest issues with climate change is the rising amount greenhouse gases in the atmosphere. Carbon dioxide makes up

Chapter 3. Production of Methanol From Carbon Dioxide the majority of these greenhouse gases at 81% (Figure 3). The amount of carbon dioxide in the atmosphere has been increasing exponentially since 1958 and is currently at 406.35 ppm (Figure 1). One of the main things that is causing this rise is the increased use of fossil fuels, including coal, oil, and natural gas, all of which produce high carbon emissions and have led to an increase in air and water pollution in the environment. Another thing causing this issue is an increasing human population, so there is an increased demand for energy. Some solutions to these problems include solar panels, wind turbines, hydrogen fuel cells, and nuclear energy (Wirkus, 2016). Some professional experiments have looked at these issues and decided to tackle them with an alternative solution: converting carbon dioxide into fuel. By converting carbon dioxide to fuel, the amount of greenhouse gases in the atmosphere would substantially decrease, and because carbon dioxide is constantly in the air, it has the potential to produce lots of fuel which can be used to generate electricity without being highly dependent on a certain condition (ex. weather). Carbon dioxide has a very high free Gibbs energy compared to most molecules, which means that it is difficult to convert carbon dioxide into a different molecule (Jiang, Xiao, Kuznetsov, Edwards, 2010). Research has shown that catalysts are needed to be able to convert carbon dioxide into fuel (Jiang, Xiao, Kuznetsov, Edwards, 2010). This experiment focuses on the impact of different catalysts on the rate of reaction and addresses the following: “What catalyst can help yield the most amount of methanol when combining carbon dioxide and hydrogen gas?”

Methods This experiment uses different catalysts to compare their ability to convert carbon dioxide to methanol. Hydrogen gas will react with carbon dioxide to produce methanol and water. In a 4L container, set up two pieces of lead as the anode and cathode. Pour 3L of water into the container. Pour sodium bicarbonate into the water to act as an electrolyte solution. Set up one bottle over the anode, which is the site of oxidation, to collect oxygen. Place another bottle over the cathode, which is the site of reduction, where hydrogen is collected. Connect alligator clips to the anode and cathode and then the other side to a 9V battery. Collect the gases. Oxidation Reaction: 2H2 O → O2 + 4H+ + 4eReduction Reaction: 4H2 0 + 4e- → 2H2 + 4OHTo produce the methanol, place the hydrogen gas into a 250mL beaker. Add carbon dioxide into the beaker. Now take 0.045 moles of the powder catalyst that is being used for that trial and pour it into the beaker. Then, heat this beaker to 473K and set to a pressure of 0.5 atm. From this reaction water and methanol should be produced. The equation of this reaction is as follows: CO2 + 3H2 → CH3 OH + H2 O The methanol will now be separated from the water by the process of distillation. Once distillation has been completed, the mass of the methanol produced will be measured. Three trials will be performed for each catalyst to improve the validity of the experiment and to make comparisons more accurate. The catalyst used in the reaction that produces the most methanol is the best catalyst. The results will be compared with an ANOVA test to determine the difference in the amount of methanol produced. If the P value of the test is less than 0.05, that will show that the data is statistically different.

Expected Results The six catalysts used in this experiment are cobalt, copper, iron, zinc, nickel, and silicon. The first five metals are transition metals, while silicon is a metalloid. The transition metals are very

similar to each other. They have similar charges in their ions, and they have similar structures and properties. Silicon differs from these metals because it is a metalloid and has 4 valence electrons. However, because the metal powders are catalysts, this will impact the rate of the reaction, but will not be directly involved in the reaction. This means that the catalysts will most likely not be chemically affected, so the working ability of the catalysts will not impact the amount of methanol produced. Another observation is that all catalysts used in this experiment have nearly the same electronegativity. Electronegativity of elements does not have to be involved in direct chemical reactions; it is just an atomâ&#x20AC;&#x2122;s ability to pull electrons closer to itself. The null hypothesis states that the type of element in the catalyst will not impact the amount of methanol produced.

Discussion The data gained from the experiment will show which catalyst yields the most amount of methanol. If the results are not significantly different, then the different types of catalysts do not affect the amount of methanol produced. If the hypothesis is not supported, it will be able to be determined what properties of an element or a compound make a good catalyst for the conversion of carbon dioxide into methanol (ex. conductivity, electronegativity, etc.). A possible next step for this experiment would be to test what would be the optimal amount of catalyst to be used to maximize methanol production. Another next step would be to see if the use of an amine in this process would help speed up the reaction thus producing more methanol. Based on the properties of the most optimal catalyst in this experiment, a next step could be to test other catalysts with similar properties and see how they would effect this experiment.

Figure 3.1: The carbon dioxide concentration at the Mauna Loa Observatory from 1958 through January 31, 2017 (Mauna Loa Observatory 2017).

Chapter 3. Production of Methanol From Carbon Dioxide

Figure 3.2: The setup of distillation for the separation of methanol from water.

Figure 3.3: The amount of each cause of greenhouse gas emissions in the U.S. in 2014 (United States Environmental Protection Agency).

References A new leaf: Scientists turn carbon dioxide back into fuel. (2016, July 29). Retrieved from https://www.anl.gov/articles/ new-leaf-scientists-turn-carbon-dioxide-back-fuel Jiang, Xiao, Kuznetsov, & Edwards. (n.d.). Turning carbon dioxide into fuel. Philosophical Transactions of the Royal Society. Mauna Loa Observatory. (2016). The Keeling Curve [Chart]. Retrieved from https://scripps.ucsd. edu/programs/keelingcurve/ Mitchell, J. F. B. (1989), The “Greenhouse” effect and climate change, Rev. Geophys., 27(1), 115–139, http://dx.doi.org/10.1029/RG027i001p00115. Northon, K. (2015, November 9). NASA Holds Media Briefing on Carbon’s Role in Earth’s Future Climate [Map]. Retrieved from https://www.nasa.gov/press-release/nasa-holds-mediabriefing-on-carbon-s-role-in-earth-s-future-climate Sun, W., & Qian, C. (2016). Heterogeneous reduction of carbon dioxide by hydride-terminated silicon nanocrystals. Nature Communications. http://dx.doi.org/10.1038/ncomms12553 U.S. Greenhouse Gas Emissions in 2014 [Image]. (2014). Retrieved from https://www.epa.gov/ ghgemissions/overview-greenhouse-gases Wirkus, H. (Presenter). (2016). Alternative Fuel Sources. Lecture presented at Staples High School, Westport, CT.

4. Nocturnal Migration of Birds

Proposed Study on the Effect of Weather Patterns on the Nocturnal Migration of Birds By Jory G. Teltser â&#x20AC;&#x2DC;19

Abstract The goal of this experiment is to determine the weather conditions in which birds move in large numbers in coastal Southwestern Connecticut during nocturnal migration. It is predicted that a period of sustained strong southwesterly winds followed by a cold front in the spring will produce the highest concentration of movement. In the fall, the same conditions but with a period of sustained strong northwesterly winds followed by a warm front will also produce the largest movements of birds. One study tracked the bird flights in multiple coastal locations in the Northeast. The researchers did so in an effort to understand how topography, winds, and the size of the migration movement affect morning flight (defined as the visible migration carried over from nocturnal movements in the very early morning) (Van Doren et al., 2014). There has also been review done on the significance of research in the field (Farnsworth, 2005). The hope is that the knowledge of which weather conditions produce large movements of birds can help predict when such movements will occur. More accurate measurements of these windows can help in the preservation and protection of birds in the Northeast, as well as across the country, specifically those threatened by extinction.

Introduction While there are species of birds that migrate during the day (diurnal migration), most species migrate at night. Many bird species that migrate at night give flight calls (Figure 1), however the reason for this is uncertain. Nocturnal vocalizations have been documented from at least 232 of 749 bird species that breed in the United States and Canada, spanning at least 51 of 82 families and 18 of 22 orders (La, 2012). During nights of favorable winds, birds will move in large numbers, increasing the frequency of flight calls heard. One study tracked the bird movements in different

Chapter 4. Nocturnal Migration of Birds locations in the Northeast in an effort to understand how topography, winds, and the size of the migration movement affect morning flight (defined as the visible migration carried over from nocturnal movements in the very early morning) (Van Doren et al., 2014). In addition to this, there has also been a review done on the importance of research in this field as well as further information regarding how weather and topographical features affect migration. Although migration occurs across the country, evidence suggests that topographic features such as mountains, hills, and coastlines concentrate birds. These features also appear to concentrate flight calls as a result of migration concentrations. In one study, it was found that changes in wind conditions produced an increase in the number of flight calls that were recorded along the coastline of Texas. Southwesterly winds forced birds migrating inland in the direction of the coast, and in order to avoid veering over the Gulf of Mexico, the birds concentrated on the coast before moving north along it. During this study, Evans reported that when the cloud ceiling is at a low elevation, the variations of the terrain cause a disruption of flight paths of migrating passerines, and cause them to concentrate in areas of lower altitudes (Evans and Mellinger, 1999). Flight calls may be important for comparative study in species placement in taxonomic order, benefiting in the resolution of cryptic species (Farnsworth, 2005). The knowledge of which weather conditions produce large movements of birds can likely help predict when such movements will occur. More accurate measurements of these timeframes can, in turn, help in the conservation of birds in the Northeast, specifically those threatened by extinction. The data gathered by this study may be used to allow for the temporary shutdown of hazardous wind turbines and other migration barriers in more precisely-defined areas during such movements to cut down on bird fatalities. Behind habitat loss and feral cats, window strikes pose as the leading cause of bird deaths, with an estimated 599 million birds killed annually in the United States alone (Figure 2). Another study found that artificial lights interfere with a birdâ&#x20AC;&#x2122;s ability to direct themselves. Nocturnal bird fatalities happen in places where an illuminated obstacle, such as a skyscraper, lighthouse, or offshore oil platform, are present and reach a height where birds are flying (Figure 3). It was observed that when the lights of an offshore oil platform were switched on, the number of birds on and around the platform quickly grew. However, when the lights were switched off, the birds rapidly dispersed, indicating that the birds were attracted to the lights. Around the world, hundreds of millions of birds are affected by this disruption each year, many of which do not survive. Changing the color of the lights could very well result in a decrease of the number of birds affected. The challenge of this however consists of developing bird-friendly lighting that is visible to the human eye, but does not attract and disorient nocturnally migrating birds (Poot et al., 2008).

Methods In this experiment, the independent variable will be the wind direction during a given time of the year. The dependent variable will be the number of flight calls of birds. This study will focus on tracking the nocturnal migration of birds via the use of an acoustic recording setup, which will be used to record flight calls during nocturnal migration (Evans and Rosenberg, 2000) (Evans, 2000). For each night data is collected, the microphone will record flight calls throughout the night, and data from the radar (via the National Weather Service) and local meteorological data will be taken down. After each night of recording, the flight calls will be analysed and categorised (via eBird). Data will be collected throughout the Spring (late April - late May) and the Fall (August - early November). The use of doppler radar to track bird migration has also been employed to aid in conservation implications (Gauthreaux, Jr. and Belser, 2003). In another study, bird movement data were collected in several different locations in the Northeast, in relation to wind patterns and direction (Van Doren et al., 2014). The latter experiment will be similar to this one in that a recording setup will be used to count migrant species, as well as their flight calls, and compare the data to weather patterns. The time of year, amount of data collected, and duration of the study are

all factors that will differ from previous experiments.

Expected Results It is predicted that a period of sustained strong southwesterly winds following by a cold front in the spring will produce the highest concentration of movement. In the fall, the same conditions but with a period of sustained strong northwesterly winds following by a warm front will also produce the largest movements of birds.

Discussion The knowledge of what weather conditions produce large movements of birds can help predict when such movements will occur. More accurate measurements of these time frames can aid in the conservation of birds in the Northeast, particularly those threatened by extinction, such as Cerulean Warbler, Golden-winged Warbler, and those threatened particularly in the Northeast United States, such as the Upland Sandpiper. The data gathered by this study may be used to possibly allow for the temporary shutdown of hazardous wind turbines, controlled uses of disorienting and unnecessary lights in highly populated areas, and other migration barriers in more precisely-defined areas during such movements to cut down on bird fatalities. These recommendations could lead to a significant leap forward in wildlife conservation. Next steps in research could be applying to genealogic study in bird morphology and evolutionary trends.

Figure 4.1: In areas where flocks of birds commonly fly just above the forest canopy, wire strikes can be mitigated by placing the lines just below the treetops. The horizontal, dotted line indicates minimum ground clearance of rth conductors, and lowering the line while maintaining this clearly requires more towers and shorter spans. (Biological Services Program, 1978).

Chapter 4. Nocturnal Migration of Birds

Figure 4.2: Examples of passerine flight calls (from Evans and O’Brien 2002): (A) Bobolink (Dolichonyx oryzivorus), (B) Indigo Bunting (Passerine cyanea), (C) Blackpoll Warbler (Dendroica striata), and (D) Swainson’s Thrush (Catharus ustulatus). Note that the axes of these spectrograms have identical scales, which facilitates comparing the different species’ calls. (B) and (C) depict modulated calls, whereas (A) and (D) depict flight calls of parts of flight calls with pure tones. (Farnsworth, 2005)

Figure 4.3: This chart show the leading causes of bird fatalities in the United States and Canada (The State of the Birds, 2014).

References Evans, W. R., & Rosenberg, K. V. (2000). Acoustic monitoring of night-migrating birds: A progress report. Strategies for Bird Conservation: The Partners in Flight Planning Process; Proceedings of the 3rd Partners in Flight Workshop; 1995 October 1-5; Cape May, NJ, 151-159. Retrieved December 5, 2016, from http://www.fs.fed.us/rm/pubs/rmrs_p016/rmrs_p016_151_159.pdf Evans, W. R. and D. K. Mellinger. 1999. Monitoring grassland birds in nocturnal migration. Pages 219â&#x20AC;&#x201C;229 in Ecology and Conservation of Grassland Birds of the Western Hemisphere (P. D. Vickery and J. R. Herkert, Eds.). Studies in Avian Biology, no. 19. Retrieved April 27, 2017. Evans, W. R. (2000, May). Applications of acoustic bird monitoring for the wind power industry. In Proceedings of the National Avian-Wind Power Planning Meeting III, San Diego, California, May 2000. Retrieved December 5, 2016. Farnsworth, A. (2005). Flight calls and their value for future ornithological studies and conservation research. The Auk, 122(3), 733-746. Retrieved December 5, 2016, from http://www.aoucospubs. org/doi/full/10.1642/0004-8038(2005)122[0733:FCATVF]2.0.CO;2 Gauthreaux, S. A., Jr., & Belser, C. G. (2003). Radar ornithology and biological conservation. The Auk, 266-277. Retrieved December 5, 2016, from http://www.aoucospubs.org/doi/pdf/10.1642/00048038(2003)120[0266:ROABC]2.0.CO;2 Impacts of transmission lines on birds in flight. (1978). Biological Services Program, 43-43. Retrieved January 14, 2017, from https://pubs.usgs.gov/fwsobs/1978/0048/report.pdf La, V. T. (2012). Diurnal and nocturnal birds vocalize at night: a review. The Condor, 114(2), 245-257. Retrieved December 5, 2016, from http://dx.doi.org/10.1525/cond.2012.100193 Poot, H., Ens, B. J., Vries, H. D., Donners, M. A., Wernand, M. R., & Marquenie, J. M. (2008). Green light for nocturnally migrating birds. Ecology and Society, 13(2). doi:10.5751/es-02720130247 The state of the birds 2014. (2014). The North American Bird Conservation Initiative U.S. Committee. Retrieved December 3, 2016, from http://www.stateofthebirds.org/2014/2014%20SotB_ FINAL_low-res.pdf Van Doren, B. M., Sheldon, D., Geevarghese, J., Hochachka, W. M., & Farnsworth, A. (2014). Autumn morning flights of migrant songbirds in the northeastern United States are linked to nocturnal migration and winds aloft. The Auk, 132(2015), 105-118. Retrieved December 5, 2016, from http://www.aoucospubs.org/doi/pdf/10.1642/AUK-13-260.1?code=coop-site

5. Antibiotics in Wastewater

The Effect of MN250 on the Removal of Sulfonamide Antibiotics in Wastewater By Joshua Zhang â&#x20AC;&#x2DC;19

Abstract Sulfonamide are antibiotics used to treat burns, urinary tract infections, and malaria. It is also used in animal feed. This excess use leads to the presence of these antibiotics in runoff and bodies of water. There are many current processes of antibiotics removal, but hypercrosslinked resins provide more advantages than traditional processes. They exhibit high adsorption capacity for polar and nonpolar compounds, easy regeneration, and low cost. MN250 was determined in a previous experiment to be the most effective of hypercrosslinked adsorbents in removing sulfamethazine in distilled water. Another previous experiment determined the effectiveness of MN250 in removing sulfamethazine in humic acid, differing pH, and different ion concentrated solutions. This experiment will study the effectiveness of MN250 in removing four other sulfonamide antibiotics in the same differing conditions: pH, concentrations of KCl, and simulated groundwater.

Introduction Antibiotics are one of the greatest inventions of the modern world, but they pose the greatest threats. The more they are produced and consumed, the more they pollute the Earth. Many have tried to solve this, but the â&#x20AC;&#x153;solutionâ&#x20AC;? which had made the most progress is in question. A current problem of antibiotics is their runoff and excessive contact with bacteria and viruses, as they can grow resistant to that antibiotic. These antibiotics become ineffective when used to treat those specific bacteria or viruses in the human body, as they are resistant to that antibiotic. One popular family of antibiotics is the sulfonamide family, which is a group of antibiotics that contain the sulfanilamide molecular structure (Image 1). They are currently used to treat urinary tract infections, prevent infection from burns, treat malaria, and produce animal feed. They are designed to competitively inhibit the conversion of p-aminobenzoic acid, PABA, by inhibiting the

Chapter 5. Antibiotics in Wastewater biosynthetic pathway of folate (Overview: Sulfonamides). Although sulfonamide concentrations are low in the environment, they are continuously being released. There is very little that is known about the impact of sulfonamides on the environment and its respective organisms, so much of analytical and environmental chemistry is focused on this issue (Stolte & Stepnowski 2014). Hypercrosslinked synthetic adsorbents are a relatively new method to remove organic contaminants from air and liquid systems (Tsyurupa & Davankov 2006). They are typically 100% crosslinked with methylene bridges between phenyl rings of long polystyrene chains (Tsyurupa e.t. 2007). Some advantages of hypercrosslinked resins are their high adsorption capacity for both polar and nonpolar organic compounds, their easy regeneration, their high mechanical strength to withstand water forces, and their low cost (Tsyurupa, M. P., & Davankov, V. A., 2006). One experiment explored the use of bamboo biochar to remove fluoroquinolone. It investigated the effectiveness of Bamboo Biochar on the removal of enrofloxacin and ofloxacin. A conclusion was made that the maximum adsorption capacity is 45.88 ± 0.90 mg·g1. The effects of differing pH were found to be low while higher ion strength led to a decrease in the adsorption of Fluoroquinolone antibiotics (Wang e.t. 2015). Another experiment determined the most effective commercially available adsorbent of Purolite in removing sulfamethazine from distilled water. The most effective adsorbent was MN250, a hypercrosslinked adsorbent produced by Purolite. MN250 displayed a high adsorption capacity. This experiment was used as a basis for a second experiment (Grimmett, M. E., 2012) An additional experiment focused on the effects of humic acids, ion concentrations, and pH concentrations on the absorbance of sulfamethazine onto the adsorbent MN250. This experiment simulated environmental conditions to determine how well MN250 would perform in real environmental conditions. The results proved to be statistically significant and supported the fact that MN250 was a very effective adsorbent for sulfamethazine. Therefore, the adsorbent MN250 will be as effective in removing other sulfonamide antibiotics as it was with sulfamethazine (Grimmett, M. E., 2015).

Methods Sulfamethazine, sulfathiazole, sulfamethoxazole, sulfamerazine, and sulfadimethoxine concentrations will approximately 30 mg L-1 for all batch studies and will be prepared by dissolving 450 mg of the respective antibiotic into 18 mL of acetone and adding it to 15 L of deionized water. The rate of adsorption will be determined by adding 100-mL of stock sulfamethazine solution to nine flasks each containing 25 mg of MN250 and then periodically measuring the antibiotic concentration over 240 hours to establish the time to reach equilibrium (defined as a <5% change in concentration over 24 hours) by using the ELISA test procedure. A control flask containing respective antibiotic solutions without MN250 will be matched to each batch adsorption series. Various salts (5 mM NaCl, 1 mM Na2 SO4 , 0.8 mM CaCl2 ·2H2 O, and 3 mM NaHCO3 ) will be added to the solutions to mimic groundwater. The pH of the solutions will be adjusted to 5, 7, and 9 by the addition of 0.2 M NaOH and/or HCl. Another set of solutions of dissolved humic acid will be made by adding 1 g of humic acid to 500 mL of deionized water. The pH will be kept constant at 7 by the addition of 0.2 M NaOH and/or HCl. Potassium chloride will added to a third set of solutions to create 0.005, 0.05, and 0.5 M KCl solutions, and the pH will adjusted to 7 with 0.2 M NaOH (Grimmett, M. E., 2015). A graph of the results of each set of solutions will be created using the Langmuir and Ho’s Pseudo-Second Order Model in order to calculate the constant adsorption rate and maximum adsorption capacity of MN250 for sulfathiazole, sulfamethoxazole, sulfamerazine, and sulfadimethoxine (Figure 1 & 2). Each set of data will be recorded (Figure 3). A second part of the experiment will be carried out to determine if the functional group of an antibiotic or the backbone influences its adsorption properties. Different antibiotics with the sulfanilamide functional group and a 4-aminobenzene ring backbone. Concentrations will be

approximately 30 mg L-1 for all batch studies and will be prepared by dissolving 450 mg of the respective antibiotic into 18 mL of acetone and adding it to 15 L of deionized water. The same process will be used to periodically calculate the concentration of each antibiotic in the solutions. A graph of the results of each set of solutions will be created using the Langmuir and Hoâ&#x20AC;&#x2122;s Pseudo-Second Order Model in order to calculate the constant adsorption rate and maximum adsorption capacity of MN250.

Predicted Results The constant adsorption rate and maximum adsorption capacity for MN250 will prove to be at least as large in removing sulfathiazole, sulfamethoxazole, sulfamerazine, and sulfadimethoxine from wastewater as it is removing sulfamethazine. The functional group will have a much greater influence on the adsorption capacity and constant adsorption rate of MN250 than the backbone of the antibiotic.

Discussion Results from this experiment can be used to further confirm the effectiveness of MN250 in removing the sulfonamide family of antibiotics. These results can be used to design another experiment to look at differing methods or the effectiveness of MN250 in removing emerging antibiotics in the Great Lakes.

Figure 5.1: Adsorption isotherms of sulfamethazine by Purolite MN250 at 25Â°C (Qe), fit to the Langmuir model. Ce is the aqueous sulfamethazine concentration at equilibrium (mg L1) (Grimmett, M.E., 2015).

Chapter 5. Antibiotics in Wastewater

Figure 5.2: Sulfamethazine adsorption by Purolite MN250 over time (Qt) at pH 5, 7, and 9 and 25Â°C in the presence of common groundwater ions, fit to Hoâ&#x20AC;&#x2122;s modeling equations, neutral, and alkaline environments. The time to reach equilibrium was 144, 120, and 144 h at pH 5, 7, and 9, respectively (Grimmett, M.E., 2015).

Figure 5.3: Data table that I will use to record the concentrations of antibiotics at certain time periods. I will use these concentrations to calculate the rate of adsorption or maximum adsorption capacity of MN250 in that specific environmental condition.

Chapter 5. Antibiotics in Wastewater

References Bialk-Bielinska, A., Stolte, S., Kumirska, J., & Stepnowski, P. (2014). Analysis and Fate Assessment of Sulphonamides in the Environment. Retrieved from International Proceedings of Chemical, Biological and Environmental Engineering website: http://www.ipcbee.com/vol69/023ICEST2014-A3001.pdf Grimmett, M. E., (2012, December 6). Removal of Sulfamethazine by Hypercrosslinked Adsorbents in Aquatic Systems. Grimmett, M. E., (2015, July 10). Adsorption of Sulfamethazine from Environmentally Relevant Aqueous Matrices onto Hypercrosslinked Adsorbent MN250. Overview: Sulfonamides. (2016, December 6). Retrieved from LiverTox website: https://livertox. nlm.nih.gov/Sulfonamides.htm Tsyurupa, M. P., & Davankov, V. A. (2006, January 4). Porous structure of hypercrosslinked polystyrene: State-of-the-art mini-review. Retrieved from https://www.researchgate.net/profile/ Vadim_Davankov/publication/222385510_Porous_structure_of_hypercrosslinked_polystyrene_Stateof-the-art_mini-review/links/5421a62c0cf26120b79ea538.pdf Valderrama, C., Cortina, J.L., Farran, A., Gamisans, X., & Lao, C. (2007, March 23). Kinetics of sorption of polyaromatic hydrocarbons onto granular activated carbon and Macronet hyper-crosslinked polymers. Retrieved from http://fulltext.study/download/612310.pdf Wang, Y., Lu, J., Wu, J., Liu, Q., Zhang, H., & Jin, S. (2015, September 22). Adsorptive Removal of Fluoroquinolone Antibiotics Using Bamboo Biochar. Retrieved from http://www.mdpi.com/20711050/7/9/12947/htm

6. Biofuel Production

The Effect of Algal and Cyanobacterial Species on Biofuel Production By Jason Katz â&#x20AC;&#x2DC;19

Abstract The fuels that humans are extremely reliant on in their daily lives are rapidly depleting. Conventional crops are currently being used but it only contradicts efforts to impede a potential global food crisis. The central question in the experiment is related to determining which species of algae and cyanobacteria would prove to be the best producers of oils for the synthesis of biofuels. It is hypothesized that the organism that can create the greatest amount of lipids will produce the greatest quantity of oils. Between Oedogonium, Spirulina, and Spirogyra, Botryococcus braunii, Scenedesmus, Nannochloropsis, Synechocystis, Synechococcus sp., and Microcystis aeruginosa, it is predicted that Oedogonium will produce the greatest quantity of oils. This algae has the greatest lipid productivity, its biomass before oil extraction is greater than the other organisms, and it was recorded to have the highest oil extraction in the experiment. (Figure 2). The materials required for this experiment are minimal and solely consist of a photobioreactor/s, a light source, a source of carbon dioxide/food, and samples of the various algae and cyanobacteria being tested. An ANOVA test will be used to analyze the data, and the results should display that the amount of oil produced from each organism is significantly different. This experiment can be further expanded by testing a greater variety of species of algae and cyanobacteria for oil extraction in future experiments and to see if other characteristics of these organisms such as dry mass are as vital for high oil extraction, and if the same results can be concluded.

Introduction/Literature Review The availability of fuels that humans are extensively dependent on in their daily lives is rapidly declining, and soon enough, they will be gone. Alternative fuel sources such as corn and soybeans are currently being used but, it only contradicts efforts to impede the global food crisis that might arise in the future. In order to solve these crises, the questions about whether or not algae and

Chapter 6. Biofuel Production cyanobacteria can be alternatively used for biofuel production and which species are the most efficient in doing so, needs to be resolved. Corn and soybeans are presently being used for biofuel production, but it is an considerable incongruity against the prevention of an upcoming food contingency. â&#x20AC;&#x153;Corn functions as feed for farm animals such as cows and chicken, which humans are both heavily reliant onâ&#x20AC;? (Hirsch 2008). Using corn as a source for a process other than food production would severely limit the number of cattle and chicken that can be used for milk and egg harvesting, but it would also decrease the annual production of beef, poultry, and pork. Not only would this negatively affect the current human population, but it would not help to create more arable land/use it more efficiently to grow more crops for future generations. Each individual species of algae and cyanobacteria differs from one another in their physical and chemical characteristics. Having a high lipid productivity is an essential characteristic of an algae or a cyanobacteria for biodiesel production, but growth rate is an extremely advantageous trait for biofuel generation as well (Griffiths et al 2009). It has been known that organisms with higher rates of biomass and oil production can generate more oil than most conventional crops (Griffith et al 2009). This was a critical characteristic that helped to decide the species of algae and cyanobacteria that would be used in the experiment. The central question behind this experiment is: which species of algae and cyanobacteria can produce the greatest amount of oils for biofuel production? The hypothesis stated that Oedogonium (algae) will have the greatest quantity of oils produced, as its lipid component is greater than the other species chosen. If one of the selected specimens is observed to produce greater amounts of oil for biofuel production than what other researchers claim conventional crops can produce, then the goal of finding an alternative source for biofuel synthesis will be achieved.

Materials and Methods The independent variable in this experiment is the species of algae and cyanobacteria being tested, and the species used were Oedogonium, Spirulina, Spirogyra, Botryococcus braunii, Scenedesmus, Nannochloropsis, Synechocystis, Synechococcus sp., and Microcystis aeruginosa. The dependent variable is quantity of oil extracted from the algae and cyanobacteria that can be used for synthesis of biofuels. The only materials needed for this experiment are a photobioreactor similar to those in Figure 1, Figure 3, and Figure 4, a light source, a source of carbon dioxide, and the multiple species of algae and cyanobacteria being tested. The statistical test used to analyze the data would be an ANOVA test, and the resulting conclusion will be that the amount of oil produced between each organism is statistically different.

Predicted Results The most competent algae in this experiment should be Oedogonium, due to its high lipid productivity and fast growth rate in comparison to the other algae. In terms of cyanobacteria, Synechococcus sp. should be the most productive. One factor that is often linked with oil production is biomass production, and Oedogonium has 3.8 grams of biomass, which is greater than the other species (Hossain et al 2008)(Figure 2). Both of these characteristics are critical in determining which organism will have the greatest oil production, which is why Oedogonium and Synechococcus sp. will be more likely to produce more.

Discussion/ Implications The data collected will reveal that Oedogonium is the most effective for biofuel production. Its high lipid productivity and growth rate will create more oils that can be extracted. The algae

or cyanobacteria that will end up with the greatest amount of extracted oils is the one that should be used as an alternative biofuel source as opposed to conventional crops. The results from the ANOVA test will affirm the prediction that the amount of oils extracted from each organism are significantly different from each other and that one of the specimen truly is the most effective in this procedure. To further expand the idea behind this experiment, a greater variety of algae and cyanobacteria/other photosynthetic organisms can be tested for oil extraction in future experiments and other characteristics that are a vital for biofuel generation could be tested for.

Figure 6.1: This image displays the structure and processes that occur in a photobioreactor that is very similar to the one used in this experiment.

Figure 6.2: The results from an experiment completed that compared Spirogyra to Oedogonium and their dry weights, extracted oils, and biomasses.

Chapter 6. Biofuel Production

Figure 6.3: An example of a real life photobioreactor is displayed in this image and the unique structure can be observed.

Figure 6.4: Unlike photobioreactors, this is an example of the Airlift system that can be used as an alternative. The structure of this growing container is more tank-like than the usual photobioreactor (Spirulina Systems).

References Airlift System. Digital image. Spirulina Systems. Spirulina Systems, n.d. Web. 11 Jan. 2017. Anderson, M. (2015, July). Algae as Biofuel. Griffiths, M. J., & Harrison, S. (2009, January). Lipid productivity as a key characteristic for choosing algal species for biodiesel production. Hirsch, J. (2008, March 2). Corn is King â&#x20AC;&#x201C; And therefore a growing problem. Los Angeles Times. Retrieved from http://articles.latimes.com/2008/mar/02/business/fi-corn2 Hossain, S., Salleh, A., & Boyce, A. N. (2008). Biodiesel Fuel Production from Algae as Renewable Energy. Retrieved from American Journal of Biochemistry and Biotechnology database. Laboratory Photobioreactor. Digital image. IMG. N.p., n.d. Web. 19 Jan. 2017. <http://img.cdn2. vietnamnet.vn/Images/english/2013/01/27/13/20130127133139-8.jpg>.

7. Journal Staff

Faculty Advisor Ms. Karen Thompson

Editor-in-Chief Alice Sardarian ‘17

Assistant Editor Alyssa Hyman ‘18

Writers Aniruddha Murali ‘19 Jason Katz ‘19 Jory Teltser ‘19 Joshua Zhang ‘19 Juliana Hopper ‘18 Zachary DeBrino ‘19 Special thanks to PTA Wrecker Grants for sponsoring this issue. To contribute to our next issue, kindly email ah49508@students.westport.k12.ct.us