Transcript PHillip Ferguson PJAS Slides

Rainwater Toxicity Effects
on
Microbial Life.
Phillip Ferguson
February 2, 2008
Central Catholic High School
Water Pollution
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Contributed to by neighborhood people
(fertilizers, unclean gutters as a source of debris,
other trash items and residue to water sources).
Affects water dwelling organisms (polluting of
the water leads to harming the fish population,
which leads to harming of sea-faring birds.
Surface runoff pollutes lakes, streams, and
rivers (Silt from logging operations, farmland
feces, chemical waste from large industries).
Types of Pollutants
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Organic Pollutants – Bacteria found in animal
feces, tree and brush debris from logging
camps, processed food from landfills.
Inorganic Pollutants – Silt runoff from
construction sites, chemical waste from
industrial products, heavy metals from acid
mine drainage.
Water Cycle
Affects of Farmland Feces
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As rainwater runs through farmland, it may
pick up E coli bacteria. The more E coli the
rainwater absorbs, the more that gets into
water supplies. As the amount of E coli in
water increases, the more of a chance there
is that the bacteria gets into tap water and in
turn infects people.
First Flush
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System designed to collect the first wave of
rainwater from rooftop gutters.
Intercepts harmful debris and stops it from
getting to water sources and polluting water
supplies.
Yeast
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Growth form of
eukaryotic
microorganisms
classified in the
kingdom Fungi.
Has been widely used
in genetics and cell
biology, largely
because the cell cycle
in a yeast cell is very
similar to the cell cycle
in humans.
Purpose and Hypothesis
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To investigate the effects of local rainwater on
the survivorship of yeast colonies. This could
help gauge the toxicity or other inhibitory effects
of rainwater.
Hypothesis – The survivorship of yeast colonies
will be significantly affected by all the tested
concentrations of rainwater.
Null Hypothesis – The survivorship of yeast
colonies will not be significantly affected by the
tested concentrations of rainwater.
Materials
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YEPD agar plates (1% yeast
extract, 2% peptone, 2%
glucose (dextrose), 1.5%
agar)
YEPD Media (1% yeast
extract, 2% peptone, 2%
glucose (dextrose)
Klett spectrophotometer
Sterile pipette tips
Micropipettes
Vortex
Incubator
Sidearm flask
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Spreading platform,
spreader bar, ethanol
Sample rainwater (stock)
0.22 micron syringe filters +
10 mL syringe
Distilled water
Ethanol
Bunsen burner
Incubator
Saccharomyces cerevisiae
(Yeast)
Timer or clock
700 mL rainwater sample
from Fox Chapel PA
Procedure
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S.c. Yeast was grown overnight in sterile YEPD Media.
A sample of the overnight culture was added to fresh media in a
sterile sidearm flask.
The culture was placed in a shaking water bath (300 C) until a density
of 50 Klett spectrophotometer units was reached. This represents a
cell density of approximately 107 cells/ml.
The cell culture was diluted in sterile dilution fluid to a concentration
of approximately 103 cells/ml.
The rainwater samples were sterile filtered using 0.22 micron syringe
filters.
Concentrations of stock (rainwater), yeast, and distilled water were
prepared in 10 test tubes using 5 different varieties of concentrations
(two test tubes per concentration were arranged in rows with the
corresponding concentration vertically aligned with its twin).
After concentrations are prepared, wait 15 minutes.
Procedure (con.)
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After 15 mins,0.1 mL aliquots were spread onto YEPD
auger plates from each concentration in the first row of
tubes to 8 auger plates each.
After 45 mins, 0.1 mL aliquots were spread onto YEPD
auger plates from each concentration in the second
row of tubes to 8 auger plates each.
The plates were incubated at 300 C for 48 hours.
The resulting colonies were counted. Each colony is
assumed to have arisen from one cell.
Concentrations for Procedure
Tubes
1, 2
Stock
(mL)
9.9
5,6
7,8
9,10
5
[~100%] [50%]
0
4.9
2.5
[25%]
7.4
1
[10%]
8.9
0
[0%]
9.9
Yeast
(mL)
0.1
0.1
0.1
0.1
0.1
Total
(mL)
10
10
10
10
10
Distilled
Water
(mL)
3, 4
Rainwater Effects on Yeast Survivorship
(15 min exposure)
p = 1.55202E-13
p = 1.55202E-13
100
100% RW
50% RW
60
25% RW
P < .05
20
P < .05
40
P < .05
80
P < .05
Number of Colonies
120
0
1
Concentration of Rainwater
10% RW
0% RW
Rainwater Effects on Yeast Survivorship
(45 mins)
P = 9.24553E-10
100
P < .05
80
20
P > .05
40
P > .05
60
P < .05
Number of Colonies
120
0
1
Concentration of Rainwater
100% RW
50% RW
25% RW
10% RW
0% RW
Conclusion
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After conducting both an Anova and
Dunnett’s Test for the data, it was concluded
that the 15 minute set of data was
significantly affected by the stock (rainwater),
and the null hypothesis was rejected.
Surprisingly, the yeast colonies were all
positively affected by the concentrations of
stock.
Conclusion (con.)
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After 45 minutes of exposure it was
discovered that both the 100% and 10%
concentrations significantly affected the yeast
colony population. The 100% stock
negatively influenced the yeast colonies,
while the 10% stock positively influenced
yeast colony population.
Limitations and Flaws
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Just one test subject (only yeast) as opposed
to more (yeast and bacteria).
Difficult to get the exact same time of
exposure for each sample because of lag
time between transfer of stock between auger
plates and test tubes.
Extensions
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Use two kinds of test subjects (yeast and
bacteria).
Take sample rainwater from different areas.
Expose the test subjects to the stock for
different amounts of time (elongate exposure
to ensure encompassing results).
Fuse the rainwater into the auger plates to
more evenly distribute the rainwater.
References
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http://en.wikipedia.org/wiki/Rainwater
http://en.wikipedia.org/wiki/Yeast
http://en.wikipedia.org/wiki/Water_pollution
http://www.waterencyclopedia.com/EnGe/Fresh-Water-Nautral-Composition-of.html.