1. Tap Water Intrusion Effects on Algal Populations Anthony DeRenzo Grade 9 Pittsburgh Central Catholic High School

2. Surface Runoff Is part of the water cycle and describes the water that flows over a land surface. It can come from precipitation and flow into natural bodies of water. It can have an effect on the marine life in the various water sources it enters depending on the type of runoff.

3. Types of Surface Runoff Petroleum Pesticides Many organic and inorganic chemicals Silt Fertilizers Tap water

4. Tap Water Water directly drawn from a faucet. Used in every day activities such as plumbing, washing machines, and dishwashers. Contains a wide variety of contaminants. Goes through a purification process to set each contaminant at a harmless standard.

5. Common Components in Tap Water Calcium9.84 mg/L Iron0.0034 mg/L Potassium1.36 mg/L Magnesium1.72 mg/L Manganese0.0002 mg/L Sodium8.76 mg/L Sulfur4.45 mg/L Silicon0.77 mg/L Strontium0.56 mg/L Zinc.0022 mg/L Chlorine0.2 mg/L Fluoride0.8 mg/L

6. Trace Contaminants in Tap Water cryptosporidiumasbestos giardia lambliabarium legionellalead coli forms (including fecal coli form and E. coli) mercury carbofurannitrate and nitrite bromateatrazine chloritebenzene antimonymany more various organic contaminants

7. Eutrophication Eutrophication is the process by which a body of water becomes enriched in dissolved nutrients that stimulate the growth of aquatic plant life. A eutrophic body of water promotes the growth of algae due to increased nutrients which promotes population growth. This reflects the purpose of this experiment because it was tested if tap water or tap water + sodium nitrate would have an effect on the nutrients in the water. This would then affect the growth of algae if there is a negative effect.

8. Sodium Nitrate A colorless, odorless crystalline compound that has chemical properties that resemble potassium nitrate. Formed by a chemical reaction of sodium chloride and nitric acid. Used in the production of explosives, fertilizers, and potassium nitrate. It is also used to preserve meats.

9. Relationship People make use of sodium nitrate by growing plants with fertilizers. People make use of tap water through sewage systems, plumbing, and many other activities. A fraction of these two substances undoubtedly enter natural water sources. There are numerous species of macroscopic and microscopic organisms that could be affected by runoff that contains either tap water or tap water + sodium nitrate.

10. Algae All algae lack organic structures such as roots, leaves, seeds, and flowers. All algae are photosynthetic and usually inhabit damp places or bodies of water. Algae play very important roles in aquatic ecology. There are two major groups of algae; prokaryotic and eukaryotic.

11. Chlamydomonas reinhardtii Single celled green algae that are common in soil and freshwater. Have two flagella; one visible that is used for gliding and swimming, or metaboly and one that is not usually not visible. Typically can grow in the light on a medium of inorganic salts.

12. Euglena sp. There are many different shapes of euglena such as oval, ribbon-shaped, and cylindrical. They also have two flagella; one that is used to swim and one that is not visible. Have a red pigmented eyespot that is part of a light receptive system. They reproduce through binary fission and vary from 20 to 300 nanometers.

13. Purpose Test tap water and a mixture of tap water + sodium nitrate on two types of algae: Chlamydomonas reinhardtii and Euglena sp.

14. Null and Alternative Hypotheses Null- tap water will not significantly affect the population survivorship of Chlamydomonas reinhardtii or Euglena sp. Null- tap water with sodium nitrate will not significantly affect the population survivorship of Chlamydomonas reinhardtii or Euglena sp. Alternative hypothesis- the addition of different concentrations of tap water and tap water with sodium nitrate will significantly (adversely) affect the algal models.

15. Materials science kit educator spectrophotometer orange test tube rack 64 13 X 100 Borosilicate culture tubes 5 ml pipette 100W incandescent light bulb 68 ml of a population of chlamydomonas reinhardtii 68 ml of a population of euglena 16 ml of spring water 8 ml of tap water 1 ml of sodium nitrate.22 micron syringe filter

16. Procedure 1 Tap water was sterile filtered with a 0.22 micron syringe filter. The following ingredients were added to the culture tubes to create a total of 5 ml in each tube: tap water, spring water, sodium nitrate, and the two types of algae (chart on next page). The ingredients were added at different concentrations and were made into 8 separate groups for each type of algae.

17. Procedure 2 Algae 2 ml Spring Water 3 ml2.5 ml2 ml.5 ml2.95 ml 2.45 ml 1.95 ml.45 ml Tap Water 0 ml.5 ml1 ml2.50 ml.5 ml1 ml2.5 ml Sodium Nitrate 0 ml.05 ml [.1%] Total 5 ml

18. Procedure 3 4 equivalent tubes were created for each treatment. 64 tubes were created in all. The 64 tubes were placed in the orange rack and set on a table 1.5 meters below the 100W incandescent light bulb. The light bulb was on the entire time. The tubes were left in the rack for a total of 17 days. The absorbance at 430 nm was recorded every other day. There were 9 recordings total.

22. ANOVA Chlamy. Source of VariationSSdfMSFP-valueF crit Sample428.0843142.694714.059851.7E-053.008787 Columns188.0831 18.53201 0.000243 4.259677 Interaction10.6870633.5623540.3510020.7887723.008787 Within243.57822410.14909 Total870.432231

26. ANOVA Euglena Source of VariationSSdfMSFP-valueF crit Sample45.89373437315.297911462.2474820.1086653.008787 Columns153.78195311 22.59277 7.78E-05 4.259677 Interaction353.21118443117.737061517.297263.39E-063.008787 Within163.360525246.806688542 Total716.247396931

27. Results and Conclusions Chlamydomonas The null hypothesis that tap water will not significantly affect the population survivorship of Chlamydomonas reinhardtii was rejected (p<.05). The null hypothesis that tap water + sodium nitrate will not significantly affect the population survivorship of Chlamydomonas reinhardtii was rejected (p<.05). However, the two factor ANOVA revealed no significant interaction between the effects of tap water and sodium nitrate (no synergistic effects). Euglena The null hypothesis that tap water will not significantly affect the population survivorship of Euglena was rejected (p<.05). The null hypothesis that tap water + sodium nitrate will not significantly affect the population survivorship of Euglena was rejected (p<.05). The two factor ANOVA did reveal significant interaction between the effects of tap water and sodium nitrate. There were synergistic effects.

28. Observed Results Tap water had a separate effect on Chlamydomonas reinhardtii than tap water + sodium nitrate did. The two lines on the third graph are parallel, showing increases and decreases at the same concentrations. This shows that tap water and sodium nitrate were two independent factors. Tap water had a separate effect on Euglena than tap water + sodium nitrate had but there was a different pattern. The lines on the third graph intersect at about 10% and then go in two separate directions. This shows that tap water and sodium nitrate were dependent factors. The effect of tap water depended on the presence of sodium nitrate and visa versa.

29. Limitations and Extensions There were only 4 trials for each test group. This experiment will be furthered by using larger sample sizes. The cultures of algae were not sterile. There will be a longer duration of data collection, which could reveal more long term effects. There will be more concentrations of tap water and sodium nitrate tested. Only spring water was used in testing. This experiment will be expanded by the addition of true aquatic sources. There will be tests with more types of algae.

30. References “About Chlamydomonas.’’ Chlamy.org. 11 Dec. 2007. <http://www.chlamy.org/info.html “Algae.” Tuberose.com. 11 Dec. 2007. <http://www.tuberose.com/Algae.html “Chlamydomonas Reinhardtii.” Wikipedia.org. 11 Dec. 2007. <http://www.wikipedia.org/wiki/Chlamydomonas_reinhardtii Cloem, James E. “Eutrophication.” 12 Jan. 2008. <http://www.eoearth.org/article/Eutrophication Dr. John Wilson, biostatistician, University of Pittsburgh Dr. Stanley States, Water Quality Manager for the Pittsburgh Water Authority “Euglena.” Silicasecchidisk.conncoll.edu. 11 Dec. 2007 <http://silicasecchidisk.conncoll.edu/LucidKeys/Carolina_Key/ html/Euglena_Main.html “Euglena.” Jracademy.com. 12 Jan. 2008. <http://www.jracademy.com/~mlechner/archive1999/euglena.html “Introduction to the Green Algae.” Ucmp.berkley.edu. 12 Jan. 2008.<http://ucmp.berkley.edu/greenalgae.html “Sodium Nitrate.” Encyclopedia.com. 11 Dec. 2007. <http://www.encyclopedia.com/doc/1E1-sodiumni.html