1. The Effects of Pesticide on Microbial Life
Jose Esquivel
11th Grade
Central Catholic High School

2. Pesticide
Primary factors affecting the effectiveness of pesticides include:
Type of pesticide
Degradation
Low temperatures

3. Non Target Effects Cause cancer in humans
Negative interaction with vertebrate immune systems
Passed on through food chains
Removal of food sources

5. Pesticide Use
DEET is the most common active ingredient in insect repellents across the market
toxic for species of freshwater zooplankton

6. Pesticide (OFF Deep Woods)
25% DEET, 75% other ingredients (not revealed)
Stored at room temperature
Provides 8 Hours of protection

7. Yeast Single celled fungi
Most commonly studied organism- similar biochemistry and genes as other Eukaryotes
Saccharomyces cerevisiae (Yeast)

8. Purpose
To examine the effects of OFF DEEP WOODS on Yeast cell survivorship

9. Hypothesis
Null Hypothesis: The pesticide will not have a significant effect on Yeast cell survivorship
Alternative Hypothesis: The pesticide will significantly reduce Yeast cell survivorship

10. Materials YEPD agar plates (YEPD media + 1.5% agar)
YEPD media (1% yeast extract, 2% peptone, 2% glucose)
Sterile Dilution Fluid [SDF] (100mM KH2PO4, 100mM K2HPO4, 10mM MgSO4, 1mM NaCI)
Sterile pipette tips
Micropippettes
Vortex
Incubator
Sidearm flask
Spreading turntable
Spreader bar
Ethyl alcohol
Sterile capped test tubes with Sterile distilled water
Saccharomyces cerevisiae (Yeast)
Pesticide

11. Procedure 1. Pesticide was obtained from local Home Depot
2. The pesticide was sprayed into a 50 mL to achieve a fluid form of the pesticide
3. The pesticide was sterile filtered
4. Yeast cells were grown overnight in sterile YEPD media
5. The cultures were placed in an incubator until a density of 50 Klett spectrophotometer was reached. This is a density of approximately 10^7 cells/mL
6. The cultures were then diluted in Sterile Dilution Fluid to a concentration of 10^5 cells/mL

12. Procedure (Continued)
7. The different concentrations were made: 0%
0.1% 1%
10%
Yeast
0.1 mL
Dilution Fluid
8.9 mL
Pesticide Solution
0 mL
0.01 mL
1 mL
Sterile Water
.99 mL
0.9 mL
Final Volume
10mL
10 mL

13. Procedure (Continued)
8. The tubes were allowed to incubate at room temperature for 15 minutes
9. After vortexing to evenly suspend cells, 0.1 mL aliquots were removed from the tubes and spread on the YEPD agar plates
10. The Yeast cells incubated for 48 hours at 30C
11. The resulting colonies were counted, each colony is assumed to have arisen from one cell

14. Pesticide Effect on Yeast Cells
848
P Value- less than 10^-6
Colonies
521.14
334.29
131.43 1%
10% 0%
0.1%

15. Dunnett’s Test Table T crit.=3.01 Alpha=0.05
Yeast T Value Interpretation
0.1% Significant
1% Significant
10% Significant

16. Yeast Analysis Anova revealed significant variation between the means
The Dunnett's test revealed significant negative effects on yeast survivorship at all concentrations

17. Limitations and Extensions
Synchronization of spreading plates more precisely
Limited exposure times
Only tested liquid pulse
Only survivorship was measured- not growth
Extensions
Increase trial size to 20 or 30 for more accurate results
Vary exposure times to the water
Conduct test on different micro organisms
Agar infusion for prolonged exposure
Synergistic effects?

18. References http://www.gmasw.com/chemlist.pdf
woods.aspx#off-deep-woods-insect-repellent-v

19. Yeast ANOVA