Macomb Mathematics Science Technology Center

The Effect of Ultraviolet Light on the Growth of

Escherichia Coli

Dean Lawrence and Robert Pedder

9B
Mr. Estapa, Mr. Acre, Mrs. Gravel
28 May 2014

Lawrence-Pedder 1
Table of Contents
Introduction

Problem Statement

Experimental Design

Data and Observations

Data Analysis and Interpretation

11

Conclusion

17

Works Cited

21

Lawrence-Pedder 2
Introduction
Around the world, there are nearly 210 million cases of Escherichia coli poisoning, and
380,000 deaths caused by it. Those deaths have been mostly in children (NIH). Over the
decades people have been trying to find ways to kill E. coli and get rid of it from our lives. One
of the ways people have been testing to help kill E. coli is by exposing it to ultraviolet light. The
ultraviolet light damages and destroys the E. coli cells DNA so it cannot reproduce. That is why
radiating E. coli with ultraviolet light is one of the very effective ways people have been testing.
Escherichia coli is most commonly found in our food, specifically beef and other meats.
Since E. coli is in the intestines to break down food, it will sometimes work its way into the
bloodstream and muscle of the human body. This happening can be a serious problem. This does
not only attack the human body but also can affect cows and animals slaughtered for food.
Therefore, sometimes, when one of the animals killed for food has this problem of E. coli in the
meat and bloodstream, the E. coli will grow rapidly within it (USA.gov). If the infected mean is
eaten, the same thing can happen to humans. The E. coli will enter the bloodstream and a serious
disease will ensue: an E. coli infection.
The type of light that was used for this experiment was a UV-C light. It has already been
shown that UV-C light is more effective in killing bacteria than the two other types of UV light:
UV-A and UV-B. This type of light is not naturally found on Earth as it is completely stopped by
our atmosphere (Basslrl). This means that many bacteria (including E. coli) are less capable of
protecting from it. The wavelength of UV-C light is 100-280 nm. UV-A light has a wavelength
of 400-315 nm, and UV-B has a wavelength of 315-280 nm (National Toxicology Program).
Since it has a shorter wavelength, it has a higher amount of energy, which gives it the ability to

Lawrence-Pedder 3
destroy the bacterias DNA more effectively. Another term commonly given to this kind of light
is germicidal, meaning that it can kill germs (Zeman).
In this experiment the E. coli was exposed to ultraviolet light at different distances for
varied amounts of time. It was first exposed the E. coli in the agar to ultraviolet light treatment.
The E. coli was then grown in an incubator at 37 degrees C, for 24 hours. E. coli also grows best
at this temperature. Each Petri dish was exposed to the light only once. The information
collected from this experiment indicates the amount of E. coli colonies in each Petri dish after
the one day of growth.
The whole reason the experiment is being conducted is to find the most efficient way to
sterilize objects using ultraviolet light. This could be useful for companies that would need to
sterilize a product, so that they would be able to sterilize it as quickly as possible to save money.
Another way that it would be useful would be for the average person to easily clean and sterilize
food or other objects in their house. When a person disinfects food or objects, they would most
likely want to get it done as well as possible, in the shortest amount of time.
UV light is not the only way of sterilization though. Another commonly used method is
to expose the object to large amounts of heat. This method was not taken into account when
thinking about what would be most effective in killing E. coli. Exposing the E. coli to large
amounts of heat could possibly be a better method than using ultraviolet light, but only the light
method was considered in this experiment.
The first variable in this experiment was the amount of time the E. coli was exposed to
the UV light. As the amount of time was increased, the amount of E. coli colonies grown should
decrease, as there is a larger amount of light rays entering the cell. The more light that enters the
cell, the more that the DNA inside the nucleus will deteriorate. Even though the DNA is

Lawrence-Pedder 4
destroyed within the cell, the cell will usually not die. However, with the DNA being destroyed,
the cell will no longer be able to reproduce.
The second variable used was the distance between the light bulb and the E. coli. As the
distance was decreased, the amount of E. coli colonies should decrease. This would happen
because the light hitting the E. coli would be more direct. Again, causing more light to be
absorbed by the cell. This in turn, would cause less bacterial cells to be able to reproduce.
Therefore, halting the production of E. coli.

Lawrence-Pedder 5
Problem Statement
Problem:
The effect of ultraviolet light on Escherichia coli.
Hypothesis:
If the E. coli is exposed to ultraviolet light, then the ultraviolet light will keep the E. coli
from reproducing.
Data Measured:
The independent variables are the amount of time (seconds) that the E. coli are exposed
to the ultraviolet light, and the distance (cm) of which the E. coli is from the ultraviolet light
source. The dependent variable is the growth rate of the E. coli after exposed to the light. The
distance will have a high of 5.08 cm, a low of 15.24 cm and a standard of 10.16 cm. The low for
the time will be 30 seconds, high will be 90 seconds, and the standard will be 60 seconds. The
E. coli was exposed to the ultraviolet light for the periods of time noted. Fifteen trials were
performed for each group; in the end sixty-five trials total were done.

Lawrence-Pedder 6
Experimental Design
Materials:
65 Petri dishes
(1) 75 watt 120 volt UV light bulb
agar
(1) desk lamp

37 degree C Incubator
Escherichia coli
11 ml sterile water
transfer loop

Procedures:
SETUP:
1. Pour 1 liter of water into a 1 liter flask.
2. Place flask on hotplate. Turn hot plate on high.
3. Add a stirring magnet to the 1 liter of water.
4. Slowing add 23 grams of agar powder to the flask. Be careful not to get it on the sides.
5. Allow the solution to turn from a cloudy liquid to a clear like apple juice (the
temperature will be near 100 degrees Celsius)
6. Remove from the heat and allow it to cool.
7. Pour the agar into a petri dish just enough to cover the bottom of dish
8. Fill a test tube with 10 ml of water.
9. Use the transfer loop to move E. coli from the starter dish to the water filled test tube.
10. Fill another test tube with 1 ml of water.
11. Use the transfer loop to transfer from the 10 ml test tube to the 1 ml test tube.
12. Use the transfer loop again to transfer E. coli from the 1 ml test tube to the agar filled
petri dish.
TESTING:
13. Place bulb into lamp.
14. Place the newly prepared open petri dish containing the E. coli under the lamp at the
set distance.

Lawrence-Pedder 7
15. Set a timer for the specified amount of time.
16. Turn on the UV light and start the timer at the same time.
17. Turn the UV light off at the same time as the timer goes off.
18. Close the petri dish and incubate at 37 degrees C for 24 hours.
19. Count the number of bacterial colonies in the petri dish under a microscope.
20. Record the number of colonies.

Lawrence-Pedder 8
Data and Observations
Data:
Table 1
Design of Experimental Values
Amount of Time (Seconds)
Standard
30
60

+
90

Distance From Bulb (Centimeters)

Standard
+
15.24
10.16
5.08

Table 1 shows the high, low, and standard variables for the amount of time and distance
from the bulb. The standard levels for the variables were determined by finding a value equally
in between the low and the high amounts. The two variables were the amount of time (measured
in seconds), and the distance from the bulb (measured in centimeters). The low for the time was
30 seconds, the high was 90 seconds, and the standard was 60 seconds. For the distance, the low
was 15.24 centimeters, the high was 5.08 centimeters, and the standard distance was 10.16
centimeters.

Lawrence-Pedder 9
Table 2
Collected Data
DOE

Number of Colonies
(Amount of time, Distance from bulb)
(+,+)

1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
Average

(+,-)
20.00
16.00
16.00
20.00
26.00
22.00
24.00
22.00
27.00
18.00
18.00
20.00
19.00
26.00
20.00
20.93

(-,+)
38.00
27.00
21.00
33.00
29.00
30.00
34.00
40.00
39.00
33.00
34.00
34.00
38.00
30.00
30.00
32.66

(-,-)
40.00
30.00
32.00
35.00
34.00
30.00
36.00
37.00
32.00
38.00
30.00
31.00
34.00
36.00
33.00
33.86

Standard
52.00
45.00
47.00
40.00
40.00
50.00
40.00
40.00
39.00
40.00
40.00
47.00
47.00
50.00
40.00
43.80

31.00
32.00
28.00
35.00
31.00
30.00

31.16

Table 2 shows how many DOEs were completed, and the exact order that the trials were
executed.

Lawrence-Pedder 10
Observations:
Table 3
Observations
Date

Observation

3/11/14

Found that the way we put the bacteria in the petri dish would make the
colonies more difficult to count, which may make the data slightly off.

3/13/14

The bacteria grew large amounts on the perimeter of a few dishes.

3/18/14

The agar was not covering a few spots in two of the dishes.

3/28/14

Accidentally radiated a dish for 15 seconds too long.

4/4/14

Observed that the (-,+) samples seemed to grow in the same range as the
(+,-) samples.

Table 3 shows the different observations of mistakes and data made throughout the span
of data collection. Most of the observations were of mistakes made during the preparation and
treatment of the bacteria. One of the observations was about the range of bacterial colonies. The
dates that the observations were made were March 11, March 13, March 18, March 28, and April
4.

Lawrence-Pedder 11
Data Analysis and Interpretation
Table 4
Averages

Average

(+,+)

(+,-)

(-,+)

(-,-)

20.93

32.66

33.86

43.80

Grand Average

32.8167

Table 4 shows the averages number of colonies for each set of tests. (-,-) ended up
having the highest number of colonies, while (+,+) had the lowest. (+,-) and (-,+) were relatively
close to each other. The first variable was the amount of time that the Escherichia coli was
exposed to the ultraviolet light. The second variable was the distance of the E. coli from the UV
light. The experiment was conducted to determine the effectiveness of time and distance when
E. coli is exposed to a UV light.
38.84

26.8

Table 5
Effect of Time
Time
+
33.87
20.93
43.80
32.67
Avg. 38.83 Avg. 38.235

Figure 1. Effect of Time

Lawrence-Pedder 12
Figure 1. and Table 5 show the effect of time on the growth of E. coli colonies. As the
amount of time increased, the number of E. coli colonies decreased. When the amount of time
the E. coli was exposed to the light was increased from 30 seconds to 90 seconds, the amount of
colonies decreased by 12.035.

38.24

27.4

Table 6
Effect of Distance
Distance (centimeters)
32.67
43.80
Avg. 38.24

+
20.93
33.86
Avg. 27.40

Figure 2. Effect of Distance

Table 6 and Figure 2. show the effect of distance on the amount of E. coli colonies
grown. As the distance decreases, the number of E. coli colonies shrinks on average by 10.835.

Lawrence-Pedder 13
Figure 2. shows that as the distance is decreased (light gets closer to Petri dish), the amount of
colonies decreases.
Table 7
Interaction Effect Data
Distance
(-)
Time

Solid Segment
Dotted Segment

(+)
(-)

(+)
32.67
43.80

20.93
33.86

Figure 3. Interaction Effect of Time and Distance

Table 7 shows the average number of E. coli colonies for each set of experiments. The
averages are all the same as what was shown in Table 4. Figure 3. is a graph showing the dotted
and solid line segments. The dotted segment represents the low amount for time and has a slope
of -4.97. The solid segment represents the high amount for time, and has a slope of -5.86. The
interaction effect of these two factors is -0.9. This is found by subtracting the dashed (low)
segment slope from the slope of the solid (high) segment.

Lawrence-Pedder 14

0 0

Figure 4. Dot Plot of Effects

Figure 4. shows the effects of Time (T), Distance (D), and the interaction of the two
(TD). To identify significant factors in an experiment a dot plot of effects can be drawn. To tell
whether a factor is significant compare it to the range of standards doubled. The vertical line
represents the range of standards doubled, which in this experiment is the absolute value of 14. If
the effects are inside the dotted lines they are not significant. If any effects are outside of the
dotted lines then that factor is significant. In this case no factors were significant.
Table 8
Data of Standards
Standard Number of Colonies Grown
30
28
31
32
35
31
Table 8 signifies the number of colonies that were counted using the factors, Time (60
seconds) and Distance (10.16 centimeters). The range was 7 colonies.

Lawrence-Pedder 15

Figure 5. Standards Plot

Figure 5. shows the number of colonies that grew using the standard values of each of the
factors. The range of standards was 7 colonies. Doubling that range would yield 14 colonies of
E. coli.
Y = 32.8167 0.9(TD) + noise
Figure 6. Parsimonious Prediction Equation
Figure 6 represents the Parsimonious Prediction Equation which consists of only the
significant factors unlike the prediction equation. In this case only the interaction effect of Time
and Distance were significant.

Lawrence-Pedder 16

Y = 32.8167 0.9 (0.5)(0.5) + noise

Y = 32.59
Figure 7. Parsimonious Prediction
Figure 7 shows that if 0.5 was put into the Parsimonious Prediction Equation then a
prediction of 33.04 E. coli colonies would grow. So, if this experiment was done again using 75
seconds and 12.62 centimeters, which is halfway between the standard and the high values, a
total of 32.59 colonies would be expected to grow on average.

Lawrence-Pedder 17

Conclusion
The hypothesis that as the time was extended and the petri dish is closer to the light, more
E. coli would die, was accepted. The hypothesis was accepted because the set of trials with the
shortest distance and highest amount of time, had the lowest average number of colonies. It was
also accepted because the set of trials with the longest distance and shortened amount of time,
had the highest average number of colonies. This effect was expected because of data from
previous experiments that have shown the same general results.
The (+,+) trials resulted with an average of 43.80 colonies. The (+,-), (-,+), and (-,-) trials
had 32.66, 20.93, and 33.86 colonies respectively. The (-,-) trials had the lowest average number
of colonies. Along with the (+,+) trials, the (-,-) shows that as the distance decreases, and the
time increases, less colonies would grow. Which in turn, supports the researchers hypothesis.
The data also shows that the time variable was more effective in killing the E. coli than the
distance variable. But the two variables were very close to each other in terms of their ability to
kill E. coli. Their effects were just over 1 E. coli colony.
The reason that E. coli is killed from UV light is because of ultraviolet lights short
wavelength. The shorter wavelength means that ultraviolet light has more energy. With more
energy, the light is able to penetrate the nucleus of the cell and destroy the DNA within it. The
amount of time impacts the fatality rate of the E. coli because it allows the concentration of light
to be longer. Which in turn, destroys more of the cells DNA. This gives it a higher chance of
killing the E. coli. The distance causes more cells to die, since as the light gets further away, the
area that the UV light hits is more spread out. Each time the distance is doubled, the area that the

Lawrence-Pedder 18
light hits is multiplied by four. Therefore, less light will actually hit the E. coli when the distance
is elongated. The distance from the light and the amount of time exposed to it, work well
together to kill the E. coli for this experiment. This experiment held true compared to many
other experiments. The mortality rate of E. coli radiated at 30 second intervals was nearly 40
percent. When E. coli was radiated for 1 minute the mortality rate was almost 90 percent (Beck).
Of course, there were multiple flaws that could have affected the number of colonies that
were counted. One such problem could have been how each petri dish was seeded with the E.
coli. For this experiment, the E. coli was planted on the petri dish by creating an X mark on the
agar with the transfer loop. Instead, the whole 1 ml of water that was in the test tube should have
been poured directly on the agar. This problem made it difficult to count the colonies reliably,
and also made it so that less E. coli made it onto the petri dish in the first place. There could
have been more reliable numbers if more E. coli made it onto the petri dish in the first place.
Larger differences could have been seen by using larger numbers. Another potential problem
might be that the sterile bag that contained the petri dishes had been open for a few days before
the petri dishes could be used. This could have caused unwanted bacteria to find its way onto the
dish. It is not confirmed if this caused any problems with the data, but it is a possibility. One
other flaw is that the test tubes that were used were being used for other experiments over the
same days as this one. It is possible that the test tubes were not completely sterilized and some
contaminants from other experiments made their way into this experiment. This may not be a
possibility though, as the test tubes were sterilized with boiling water between experiments. One
other small issue could have been that using our method, it wasnt possible to give each petri
dish the exact same amount of time. For one or two of the trials, we went a few seconds over the

Lawrence-Pedder 19
amount of time for that trial. These few seconds could have caused fewer colonies to grow than
there should have been.
At this point in time, it isnt possible to prove if any of these errors actually impacted the
experiment or not. It is also possible that none of these had any impact at all on the experiment
at all. With one exception, none of these could have been controlled in this experiment. The
exception being the seeding of the E. coli on the Petri dish. That problem was a design issue,
while the rest were natural problems that could not be easily avoided.
There are multiple things that would be changed if this experiment were to be rerun. One
of the things that would be changed would be how the agar was seeded with the E. coli. Another
thing that could be changed is the high, low, and standard values for the two variables. There
could be a larger range between each value in order to make the number of colonies more
different from each other.
This experiment is useful outside of the academic environment in many different ways.
At a pharmaceutical plant instruments and machine parts need to be sterilized of E. coli and other
bacteria. In a hospital or doctors office, surgical equipment has to be sterilized before being used
in/on a patient. This should not be interpreted the wrong way though. As much as it is a good
thing that the use of ultraviolet light can kill bacteria it cannot be used on the human body to
disinfect or heal a wound. This would result in terrible tragedy for the human body as it would
try kill too many cells that are vital to the function of the body. And although this can happen
there are defenses in the body to try and protect the body from ultraviolet light such as the
pigment in the skin (Chaffin)
Altogether this was a very interesting, informative, and important experiment. With the
information in this experiment we can make some of the food we eat healthier. We can also make

Lawrence-Pedder 20
the world we live in safer, by discovering new ways to sterilize instruments within the work
force such as in hospitals and pharmaceutical plants. The next step in this type of experiment
would be to test the effects of ultraviolet light on different types of bacteria such as Coccus,
Spirillum, and Vibrio Cholerae.

Lawrence-Pedder 21

Works Cited
Basslrl, Eby. "Effects of UV Irradiation on Microbial Numbers and Population." University Of
Pennsylvania. University Of Pennsylvania, n.d. Web. 4 Feb. 2014.
<http://www.sas.upenn.edu/LabManuals/biol123/Table_of_Contents_files/19-UVEnum.pdf>.
Beck, Alyssa E. "What Are the Effects of Ultraviolet Light on Bacteria Mortality?" University Of
Southern California. University Of Southern California, 2 Apr. 2004. Web. 4 Feb. 2014.
<http://www.usc.edu/CSSF/History/2004/Projects/J1303.pdf>.
"E. Coli." Overview. NIH. Web. 21 May 2014.
<http://www.niaid.nih.gov/topics/ecoli/understanding/pages/overview.aspx>.
"How Does E. Coli Get into Meat? - USA.gov Blog - United States Government Blog." USA.gov
Blog. N.p., n.d. Web. 14 May 2014.
<http://blog.usa.gov/post/6150460032/how-does-e-coli-get-into-meat>
"National Toxicology Program." NTP RSS News. U.S. Department of Health and Human
Services, n.d. Web. 21 May 2014.
<http://ntp.niehs.nih.gov/?objectid=BD4CD88D-F1F6-975E-792094AC1CE4B062>.
Zeman, Gary. "Ultraviolet Radiation." Health Physics Society. Health Physics Society, 27 Aug.
2011. Web. 02 Feb. 2014. <http://hps.org/hpspublications/articles/uv.html>.