Zane Stiles and Justin Beiriger
Mr. Krotec
AP Biology
January 6, 2013
I.
II.
Animal Behavior Lab
Introduction
This experiment is being done to study pillbugs and observe their responses to
environmental variables, as well as to investigate mating behavior in fruit flies.
Understanding what environmental conditions can affect the behavior of these
animals is important to understanding human’s effect on the environment. Knowing
that certain pollutants or other conditions affect these animals’ behaviors will allow
ecologists to understand human impact on the environment or where to predict the
location of these animals.
Armadillidium vulgare, also known as pillbugs, are terrestrial isopods. Terrestrial
isopods are land dwelling crustaceans. Related to lobsters, crabs, and shrimp,
terrestrial isopods breathe with gills. When threatened, pill bugs can curl up into a
tight ball for protection. These pillbugs use their external antennae to observe their
environment.
Drosophila melanogaster, of the fruit fly, have been extensively studied. They
live throughout the world and feed on the fungi of rotting fruit. Its genome has been
sequenced and its behavior has been extensively studied. A choice chamber, which
will be used in this experiment, gives fruit flies two choices, in this case, either bottle
cap of the choice chamber, made of the top of two bottles, is filled with a different
variable. Fruit flies are attracted to substances that offer food or an environment in
which to lay eggs.
Ethology is the study of animal behavior, and includes the movement of animals.
Many animals express orientation behaviors, which place the animal in its most
favorable environment. Taxis is a directed response to a stimulus, either toward or
away from a stimulus. Some common variables which cause taxis are light, heat,
moisture, sound, or chemicals. Kinesis is a non-directional response to a stimulus. For
example, an increase in heat might make reptiles move more quickly, but not toward
any particular location.
The purpose of this investigation is to determine the effect of hydrochloric acid,
hydrogen peroxide, and light intensity on the movement behavior of Armadillidium
vulgare and Drosophila melanogaster. The null hypothesis is that the variables will
not significantly affect the movement of these species.
Materials and Procedure
Materials:
• Fruit fly cultures (approximately 30–40 fruit flies per lab group)
• Choice chambers constructed from two matching plastic water bottles and caps
per lab group
• Extra caps to fit water bottles
• Cotton balls with tape to secure the cotton balls
• 2% hydrogen peroxide solution
• Safety goggles or glasses
• Timers
• Clear plastic packing tape
• Pipette for hydrogen peroxide
• Masking tape
• 2 petri dishes
• 10 pillbugs
• Filter paper
• Tweezers
Pillbug Procedure:
1. Prepare a choice chamber by attaching two petri dishes with a passageway cut
between the two. The choice chamber consists of two large, plastic petri dishes
taped together with an opening cut between them. Cut the opening with scissors
and use tape to hold the dishes together.
2. Line one chamber with a piece of filter paper with 2 mL of hydrogen peroxide
and line the other with a piece of filter paper with 2mL of water.
3. Use a soft brush to transfer 10 pillbugs from the stock culture into the choice
chamber. Place 5 pillbugs in each side of the choice chamber. Cover the
chambers.
4. Count how many pillbugs are on each side of the choice chamber every 30
seconds for 10 minutes and then record your data. Continue to record even if they
all move to one side or stop moving.
5. Return your pillbugs to the stock culture.
6. Clean out the chambers, and line both chambers with a dry piece of filter paper.
Cover one side with a dark sheet so no light can get through. Position a light
above the other side.
7. Repeat steps 3-5.
Fruit Fly Procedure:
8. Prepare a choice chamber by attaching two water bottles after cutting off the
bottom, so that the bottom of the two water bottles are together and no flies can
escape. Place cotton balls in the bottle caps on either side of the bottle.
9. Add 2 mL of hydrogen peroxide to the cotton ball on the left side, and add 2mL of
water to the cotton ball on the right side.
10. Release 20 flies from the culture into the choice chamber
11. Count how many flies are on each side of the choice chamber every 30 seconds
for 10 minutes and then record your data. Continue to record even if they all move
to one side or stop moving.
12. Return your flies to the stock culture.
Results
Pillbugs:
Time (seconds)
0
30
60
90
120
150
180
210
240
270
300
330
360
390
420
450
480
510
540
570
600
Pillbugs on side with
water
5
5
6
7
8
9
9
9
10
10
10
10
10
10
10
10
10
10
10
10
10
Pillbugs on side with
hydrogen peroxide
5
5
4
3
2
1
1
1
0
0
0
0
0
0
0
0
0
0
0
0
0
Number of Pillbugs on Water Side vs. Time
12
Number of Pillbugs on Water Side
III.
10
8
6
4
2
0
0
100
200
300
400
Time Elapsed (seconds)
500
600
700
Time (seconds)
0
30
60
90
120
150
180
210
240
270
300
330
360
390
420
450
480
510
540
570
600
Pillbugs on dark side
5
5
5
5
5
6
6
6
7
7
7
7
7
8
8
8
8
9
9
10
10
Pillbugs on light side
5
5
5
5
5
4
4
4
3
3
3
3
3
2
2
2
2
0
0
0
0
Number of Pillugs on Dark Side vs. Time
Number of Pillbugs on Dark Side
12
10
8
6
4
2
0
0
100
200
300
400
Time Elapsed (minutes)
500
600
700
Chi Square:
(Note: 5 on either side was the expected result, and the final time trial was used as the
observed result)
Water vs. Hydrogen Peroxide:
X2 =(10-5)2 + (0-5)2
5
5
2
X = 25 + 25
5
5
2
X = 10 P<0.01<0.05
Expected
Water
5
Observed
10
Expected
Light
5
Observed
10
Dark vs. Light:
2
2
X =(10-5) + (0-5)
5
5
2
X = 25 + 25
5
5
2
X = 10 P<0.01<0.05
Fruit Flies:
Time (seconds)
0
30
60
90
120
150
180
210
240
270
300
330
360
390
420
450
480
510
540
2
Flies on side with
hydrogen peroxide
13
11
12
12
13
10
9
6
7
6
6
4
4
2
5
2
3
1
3
H2O2
5
0
Dark
5
0
Flies on side without
hydrogen peroxide
7
9
8
8
7
10
11
14
13
14
14
16
16
18
15
18
17
19
17
570
600
3
3
17
17
Number of Fruit Flies on Hydrogen Peroxide
Side
Number of Fruit Flies on Hydrogen Peroxide
Side vs. Time
14
12
10
8
6
4
2
0
0
100
200
300
400
500
600
700
Time Elapsed (seconds)
Chi Square:
(Note: 10 on either side was the expected result, and the final time trial was used as the
observed result)
Water vs. Hydrogen Peroxide:
2
2
2
X =(17-10) + (3-5)
5
5
2
X = 49 + 4
5
5
2
X = 10.6 P<0.01<0.05
IV.
Water
Expected
10
Observed
17
H2O2
10
3
Conclusion
At an alpha of 0.05, hydrogen peroxide had a significant effect on pillbug location since
the P-value < 0.05. Similarly, the hydrogen peroxide seemed to have a significant effect on
fruit fly behavior. This result makes since because hydrogen peroxide is sometimes used as a
substitute for bleach or other cleaning agents, designed to kill bacteria and other living cells.
Thus, it is natural that these organisms would shy away from it.
Secondly, the availability of light had a significant effect on pillbug behavior, since the Pvalue < 0.05. The bugs heavily favored the dark side, which is natural considering their
environment. In the wild, pillbugs live underground or above ground but under rocks or
other objects that could provide complete shade. The pillbugs, therefore, would be more
comfortable shielded from the light than they would in the light, where they would feel like
they were above ground.
Taxis is an organism’s directional response to a stimulus or stimulus gradient. This term
can be used to describe the pillbugs’ and the fruit flies’ behavior in response to the in vitro
stimulus. The pillbugs shy away from light because of an inherent biological indicator that
they have evolved to suit their dark lifestyle. Similarly the organisms are able to sense
hydrogen peroxide and relay messages to the brain that it is bad for the organism. These
signals result in the creature’s behavior, which is largely impacted by its environment.
As with every experiment, this one had limitations and possible opportunities for future
studies. First, some limitations include the evaporation of hydrogen peroxide or water prior
to the organisms’ exposure, the pillbugs becoming reluctant to move (tired), and the light
being too intense so that it stuns the pillbugs. Some possible extensions include varying
intensities and colors of light, testing different peroxides, testing different organisms,
perhaps even mammals, and inducing a predator-prey feeling in vitro to observe the fight or
flight response.