Soil Contamination Effects on
Microbial Life
Christian Dresser- Pittsburgh Central Catholic
High School, Grade 10
Common Inorganic Soil
Contaminants
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Lead - poisonous, paint can gradually come off buildings and
contaminant soils
Mercury – poisonous, generated from cement production, coal
fired plants etc.
Acetone – used in fuel for cars and trains, dissipates slowly in soil
Arsenic – poisonous, used in integrated circuits
Barium – poisonous, used in welding railroad tracks together
Benzene – produced from coal, found in exhaust
Coal - carried via near by trains, combustible
Gypsum – used in Ordinary Portland Cement (OPC), drywall, and
plaster, composed of calcium sulfate
Soils
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Soils are among the most precious though least
appreciated resources for human kind.
Broken rock particles that have been altered by chemical and
environmental conditions, weathering, and erosion
Altered by interactions between the lithosphere, hydrosphere,
atmosphere, and biosphere
The world is facing a crisis of soil integrity:
erosion
salinization
compaction
Pollution
Soils
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Soils near industrial, urban, or transportation
centers are considered at high risk for soil pollution.
Organic – bacteria, viruses, molds etc.
Inorganic – lead, mercury, acetone, arsenic,
barium, benzene, coal, and gypsum
Lead
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Poisonous: can cause blood and brain disorders
Highly malleable and ductile
8 million tones produced annually
Used in car batteries, organ pipes, and stained glass
windows
Lead paint gradually sheds off buildings and contaminants
soils
Mercury
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Used in thermometers, barometers, and other scientific
apparatuses
Poisonous: can cause brain damage, can be absorbed and
inhaled through the skin and mucous membranes
Generated from cement production
Acetone
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Component of paints and varnishes
Used to dissolve many plastics
Solvent: can depress the central nervous system
Dissipates slowly in soil
Used in fuel for cars and trains
Arsenic
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Poisonous metalloid with many allotropic forms
Used in pesticides, herbicides, insecticides, and various
alloys
Used in integrated circuits
Exposure to high level can cause multi-system organ failure
Coal
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Carried/delivered via near by trains
Largest source of fuel for the generation of
electricity
World consumption use is 6.2 billion tons annually.
Used to produce syngas: a mixture of carbon
monoxide (CO) and hydrogen gas (H2)
Combustible; sedimentary rock
Barium
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Used in rat poison, making bricks, and glass
making
Extremely poisonous, effects the nervous
system
Used in welding rail tracks together
Benzene
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Component of cigarette smoke and found in
exhaust
Produced from coal
Used to make polymers and plastic
Serious health effects, including multi organ cancer and even death
Experimental Soil Samples
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Industrial Soil – located near trains and a cement
factory; possibly exposed to the inorganic
contaminants such as lead, mercury, and gypsum
Non-Industrial Soil – not located near trains nor a
cement factory; most likely not exposed to inorganic
contaminants
Control Soil – store-bought potting soil, not exposed
to any inorganic contaminants
Cement
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Made by heating limestone with small quantities of
other materials (such as clay) to 1450° C.
The resulting hard substance, called clinker, is
ground with a small amount of gypsum into a
powder to make Ordinary Portland Cement (OPC)
Basic ingredient of concrete, mortar, and grout
Soil
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Broken rock particles that have been altered by chemical
and environmental conditions, weathering, and erosion
Altered by interactions between the lithosphere,
hydrosphere, atmosphere, and biosphere
Potting soil contains peat moss, composted bark, sand, and
perlite.
It also contains vermicompost for water retention.
Gypsum
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Composed of calcium sulfate
CaSO4 · 2H2O
Occurs as twinned crystals and transparent
cleavable masses called selenite
Deposited in lakes, oceans, and hot springs
Used in drywall, plaster, soil conditioner, fertilizer,
blackboard chalk, and OPC
E. Coli
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Escherichia coli (E. coli) – very common, found in
intestinal tract of most mammals
There are many strains of E. coli, most are nonpathogenic.
Pathogenic strains can cause illness and death in
humans.
Frequently studied in biology – ubiquitous, simple
structure, easily manipulated in the laboratory
Purpose
The purpose of this investigation was to
determine the effects of soil
components/contaminants on the survivorship of
E.coli.
Null Hypothesis
The addition of soil to E.coli suspensions will not
significantly affect survivorship.
Materials
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5.00 grams of sterilized soil from an industrial site (railroad tracks and
cement factory)
5.00 grams of sterilized soil from a non-industrial site
5.00 grams of sterilized control soil (store bought)
Sterile Dilution Fluid (SDF) (10mM KH2PO4, 10mM K2HPO4, 1mM MgSO4,
0.1mM CaCl2, 100mM NaCl)
3 tubes for the 3 different soil extracts ( 30.0mL SDF and 5.00 g soil for
each tube)
14 tubes for the low and high concentrations of the extracts (Low- 8.9mL
SDF, 1.0mL extract; High – 9.9mL extract)
E.coli bacteria (.1 mL in each of the 14 tubes (107 cells/mL) )
56 LB Agar plates ( 8 for each concentration)
Sterile Pipettes
LB media (yeast extract (0.5 %), NaCl (1%), tryptone (1.5%))
Bunsen burner
Ethanol
Autoclave
Klett Spectrophotometer
Procedure
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1. E. Coli was grown overnight in sterile media.
2. A sample of the overnight culture was added to fresh
media in a sterile sidearm flask.
3. The culture was placed in a shaking water bath until a
density of 50 Klett spectrophotometer units was reached.
This represents a cell density of approximately 108 cells/mL.
4. The culture was diluted in SDF to a concentration of
approximately 103 cells/mL.
5. Soils from 3 different sites were sterilized in an autoclave
for 45 minutes.
6. 5.00 g of each sterilized type of soil were weighed.
7. 5.00 g of each type of soil were added to 30.0mL of SDF,
creating a soil extract.
Procedure - Concentrations
Low
Concentration
Industrial
NonIndustrial
Control Soil
SDF
Extract
E.coli
8.9mL
1.0mL
0.1mL
8.9mL
1.0mL
0.1mL
8.9mL
1.0mL
0.1mL
High
Concentration
Industrial
Non-Industrial
Control Soil
SDF
Extract
E.coli
0.0mL
9.9mL
0.1mL
0.0mL
9.9mL
0.1mL
0.0mL
9.9mL
0.1mL
Procedure Cont’d
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11. 0.1mL of the E. Coli cell culture was added to each
test tube, yielding a final volume of 10.0mL and a cell
density of approximately 107 cells /mL.
12. The solution in each tube was mixed by vortexing
and allowed to sit at room temperature for 15 minutes.
13. After vortexing to evenly suspended cells, 0.1mL
aliquots were removed from the tubes and spread on
LB-Agar plates.
14. The plates were incubated at 37 degrees Celsius
overnight.
15. The resulted colonies were counted. Each colony is
assumed to have arisen from 1 cell.
Data
Ave. Small
Ave. Large
Ave. Total
Non –
Industrial Low
24
31
55
Non –
Industrial High
11
9
20
Industrial Low
15
21
8
16
16
13
32
37
22
Control Dirt
High
22
14
36
Control SDF
16
14
29
Industrial High
Control Dirt
Low
P-value = 0.000184, p<.05 = Significant
p<.05
60
50
40
p<.05
p<.05
p< .05
30
p<.05
p<.05
p<.05
20
10
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Colonies Counted
Total Average of Colonies Counted
Soils and Concentrations
Dunnet Test Results
Variables
t value
1.2227
Interpretation
Control Dirt Low
a = .05; n = 6.5; a = .01
1.0417
Not Significant
Not Significant
Industrial High
a = .05; n = 7; a =.01
1.3344
Not Significant
Not Significant
Industrial Low
a = .05; n = 8; a = .01
.23666
Not Significant
Not Significant
Non-Industrial High
a = .05; n = 7.5; a = .01
1.3504
Not Significant
Not Significant
Non-Industrial Low
a = .05; n = 7.5; a = .01
3.9529
Significant
Not Significant
Control Dirt High
a = .05; n = 8; a = .01
Not Significant
Not Significant
Conclusion
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In my hypothesis, I stated that the E. coli grown
in the simple SDF fluid would have the greatest
survival rate.
Through the experiment, this was proven
wrong.
The E. Coli grown in a low concentration of the
non-industrial soil had the greatest survival rate,
while the E. Coli grown in the simple SDF
solution (control) had the fifth highest survival
rate.
Conclusion
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Through the experiment, there was a Rejection of
the Null hypothesis. The soil contaminants did have
effects on the survivorship of E.Coli.
In comparison with the control SDF, through a
Dunnet’s test , the survivorship of the E.Coli in the
Non-Industrial Low was significant to the
survivorship of the E.Coli in the control SDF.
This was the only concentration that was significant,
which was proven in the Dunnet’s test.
Future Implications
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More soils from different locations could be tested.
The soils could be tested on the survivorship of yeast.
More LB agar plates could be used for each type of soil.
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Different concentrations of the soil extracts could be tested.
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References
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Dr. John Wilson: biostatistician University of Pittsburgh
http://en.wikipedia.org/wiki/Lead
http://en.wikipedia.org/wiki/Mercury
http://en.wikipedia.org/wiki/Acetone
http://en.wikipedia.org/wiki/Arsenic
http://en.wikipedia.org/wiki/Barium
http://en.wikipedia.org/wiki/Benzene
http://en.wikipedia.org/wiki/Coal
http://en.wikipedia.org/wiki/Cement
http://en.wikipedia.org/wiki/Gypsum
http://en.wikipedia.org/wiki/Soil
http://en.wikipedia.org/wiki/E. Coli
http://www.epa.gov/ebtpages/pollsoilcontaminants.html
http://www.ecy.wa.gov/programs/hwtr/demodebris/pages2/dwinsoil.html
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http://www.egr.msu.edu/~envirotools/cgi-bin/soil.php3
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