The Effects of Hydrochloric Acid and UV Radiation on E. Coli David Foody Central Catholic High School Grade 9

Escherichia coli (E. coli) Large and diverse group of gram (-) bacteria Free living, symbionts, or pathogens Most strains are not pathogenic Serve as a common prokaryotic cell model Common mammalian intestinal symbiont Environmental prokaryotic model

Electromagnetic Spectrum Diagram showing the range of all types of frequencies of electromagnetic radiation The types of radiation are radio waves, microwaves, infrared, visible, ultraviolet, x-rays, and gamma rays

Ultraviolet Radiation Ultraviolet (UV) rays are light rays that have shorter wavelengths than visible light Range from 150nm – 300nm They are naturally given off by the sun, but most are absorbed by the ozone layer The wavelength used in this experiment is 254nm light

pH Measures how acidic or basic a substance is Scale from 0-14 pH Scale Neutralization Measures how acidic or basic a substance is Scale from 0-14 pH < 7 acidic, pH > 7 basic, pH = 7 neutral The mixture of an acid and a base. Mixture of acid and base commonly yields water

Hydrochloric Acid Forms an organic salt when it reacts with an organic base Strong mineral acid pH level of 3 Highly corrosive Often used to restore metals

Purpose Test the survivorship of E. Coli when exposed to Ultraviolet Radiation and Hydrochloric Acid. Determine if radiation from the sun affects the refinements of ores

Question Will differing amounts of Hydrochloric Acid and ultraviolet radiation have a significant effect on E. Coli?

Hypothesis Null Hypothesis: The Hydrochloric Acid and ultraviolet Radiation will not have a significant effect on the resulting amount of E. Coli. Alternative Hypothesis: As the concentration of Hydrochloric Acid and exposure to ultraviolet radiation increases, the survivorship of the E. Coli will decrease.

Materials LB Agar Plates Spreader Bars Ethanol Matches Hydrochloric Acid (HCl) Water Escherichia Coli UV Lamp (254 nm) Incubator Test Tubes Stopwatch Vortex Machine PH Strips Sterile Dilution Fluid

Procedure Bacteria (E. Coli) was grown overnight in LB Media A Sample of the Overnight Culture was added to fresh media in a Sterile Sidearm Flask The Culture was placed in an incubator until a density of 50 Klett spectrophotometer units was reached. This represents a cell density of approximately 108 cells/mL. The culture was diluted in sterile dilution fluid to a concentration of approximately 108 cells/mL. Various concentrations of Hydrochloric Acid were created in test tubes containing SDF, resulting in 9.9 mL per tube

Concentrations pH of 0 pH of 5 pH of 3.5 mL of E. Coli 0.1 mL mL of Water 8.9 mL mL of Acid 0 mL 0.55 mL mL of SDF 1 mL 0.9 mL 0.45 mL Total Volume 10 mL K

Procedure (Continued) 0.1 ml of E. Coli was then added to the test tubes, yielding a final volume of 10 mL. The solutions were mixed by vortexing and allowed to sit at room temperature for 10 minutes. After vortexing to evenly suspend cells, 0.1 ml was removed from the tubes and spread on LB plates. Plates were then placed under a UV lamp for specific times (3 seconds and 6 seconds) The plates were incubated at 37°C for 24 hours The resulting colonies were counted. Each colony is assumed to have arisen from one cell.

E. Coli Colonies with No Radiation Results

Individual Stat Analysis T-Value/P-Value T-Crit Significant/Not Significant pH 5 vs control T = 3.35 3.06 Significant pH 3 vs control T = 6.94 pH 5 vs pH 3 P = 0.000000002

E. Coli with 3 Second Radiation Results

Individual Stat Analysis T-Value/P-Value T-Crit Significant/Not Significant pH 5 vs control T = 3.61 3.06 Significant pH 3 vs control T = 5.92 pH 5 vs pH 3 P = 0.00000000009

E. Coli with 6 Second Radiation Results

Individual Stat Analysis T-Value/P-Value T-Crit Significant/Not Significant pH 5 vs control T = 4.92 3.02 Significant pH 3 vs control T = 7.04 pH 5 vs pH 3 P = 0.00004

Conclusion Null hypothesis: The Hydrochloric Acid and ultraviolet Radiation will not have a significant effect on the resulting amount of E. Coli. Null Rejected All colonies exposed to Ultraviolet Radiation and Hydrochloric Acid had a significant effect. The longer the E. Coli was exposed to UV Radiation and the lower the pH level of Acid, the lower the E. Coli survivorship

Future Experimentation and Limitations More pH levels and times for UV Radiation could be used Use different chemicals Use other sources of radiation Some sunlight was shining into the room while spreading the plates Only 1 species tested Only survivorship was tested

Sources Website Title: Centers for Disease Control and Prevention Article Title: E.coli (Escherichia coli) Publisher: Centers for Disease Control and Prevention Electronically Published: January 25, 2018 https://www.cdc.gov/ecoli/index.html Website Title: Healthline Article Title: E. Coli Infection: Causes, Symptoms, and Prevention Publisher: Healthline Media https://www.healthline.com/health/e-coli-infection Website Title: National Center for Biotechnology Information. PubChem Compound Database Article Title: hydrochloric acid Publisher: U.S. National Library of Medicine https://pubchem.ncbi.nlm.nih.gov/compound/hydrochloric_acid Website Title: HYDROCHLORIC ACID, SOLUTION | CAMEO Chemicals | NOAA Article Title: Search Chemicals Author: NOAA Office of Response and Restoration, US GOV https://cameochemicals.noaa.gov/chemical/3598 Website Title: American Cancer Society Article Title: What Is Ultraviolet (UV) Radiation? https://www.cancer.org/cancer/skin- cancer/prevention-and-early-detection/what-is-uv-radiation.html Website Title: LiveScience Article Title: What Is Ultraviolet Light? Publisher: Purch Electronically Published: September 15, 2017 Author: Jim Lucas https://www.livescience.com/50326-what-is-ultraviolet-light.html