Staphylococcus epidermidis and Escherichia coli Using Various

Growth Inhibition of
Staphylococcus epidermidis and
Escherichia coli Using Various
Anti-microbial Agents.
Domonique Jackson, Janecia Wilson, Railene
Echevarria, Kijhauna Stevens
Introduction
S. epidermidis –
• Gram positive
• Found on the skin
E. coli
• Found in the intestines, bad
when found in the blood
• Gram negative
Antiseptic – used on living things
Detergent – used on fomites
Antibiotics – products of living
things (mold)
• Investigations:
1. Listerine on S. epidermidis
and E. coli
2. Environmental Microbes
1.
What agents can inhibit
the growth of
S.epidermidis and E. coli?
2. What Antibiotics work on
yourself?
Purpose
1. To determine if Listerine will inhibit the
growth of S. epidermidis and E. coli
2. To analyze if bacterial growth on fomites
3. To determine which cleaning agents work
better on bacteria
4. To determine the effectiveness of antibiotics
on human natural flora
Materials & Methods
Part 1:
1.
2.
3.
4.
5.
6.
7.
8.
9.
10.
11.
Add 5 mL of Tryptic Soy Broth (TSB) into test tube marked control.
Add 5 mL of Listerine into another test tube.
Add 5 mL of stock of Staph into each of the two test tubes
After 1 minute, add 100 μL from the test tube containing Listerine and Staph into a flask
containing 100 mL of TSB marked 1 minute.
After 5 minutes, add 100 μL from the test tube containing Listerine and Staph into another
flask containing 100 mL of TSB marked 5 minutes.
Place 100 μL from control test tube into the flask containing 100 mL of TSB marked control.
Set the spectrophotometer to 550 nm and place the TSB blank in the machine to set to 0
absorbance.
Take 100 μL from control flask and place in a cuvette
Place into spectrophotometer and take absorbance reading
Repeat steps 7-9 for the 1 minute and 5 minute flasks
After recording the absorbance of all three flasks, place them in an incubator at 37°C for 48
hours total taking periodic measurements in the same fashion as described above.
Repeat steps 1-11 for E. coli.
12.
Part 2:
1.
Dip a sterile cotton swab in water and swab a large area of the surface being tested (i.e.
elevator button).
2.
Then, wipe cotton swab all over a Tryptic Soy Agar (TSA) plate and cover.
3.
Repeat for each designated place, then incubate the plates at 37 degrees Celsius for 24
hours.
Materials & Methods
Part 3:
1. Swab each plate with a sterile cotton swab (as described in Part 4) containing stock
solution of Staph or E-coli
2. Draw a line down the middle of plates
3. Get 8 disks and dip one in a corresponding cleaning agent
4. Place two disks, containing the cleaning agents, on each plate according to their
number designation
5. Place all the plates in the incubator at 37°C or 98.6°F for 24 hours
6. Measure the diameter of the zones of inhibition after 24 hours
Part 4:
1. Swab the area behind your ear using a cotton swab. Incubate on a TSA plate at 37
degrees Celsius. Recolonize after 24 hrs the bacteria onto 2 plates onto two
TSA plates using the previous TSA plate.
2. Place the 6 disks containing 6 different antibiotics onto the plates (3 on each)
3. Incubate the plates for 24 hrs at 37 degrees Celsius
4. Then, the zones of inhibition were measured and compared against a table of data
to determine resistivity, and intermediate effect, or sensitivity to each of the
antibiotics
1.8
1.6
1.6
1.4
1.4
1.2
1.2
1
Control
1 Minute
5 Minute
0.8
0.6
0.4
A bsorb en ts (55 0n m )
A b s o rb e n ts (5 5 0 n m )
1.8
1
Control
0.8
0.4
0.2
0
0
-0.2
1000
2000
Time (Minutes)
3000
5 Minute
0.6
0.2
0
1 Minute
0
1000
2000
3000
-0.2
Time (Minutes)
Figure 1: Inhibition of Growth using Listerine on Staph
(left graph) and E. coli (right graph)
50
40
Staphepidermidis
E.coli
30
20
10
0
Ly
s
C ol
h
lo
ro
x
H
T
y
i
d
ro L lex
g ist
e
e
R n P rin
u
e
e
b
ro
b
in
x
id
g
e
A
l
D co
a
w ho
D
ia n S l
l
C oa
o
m p
p
le
te
Zones Of inhibition (mm)
60
Cleaning Products
Figure 2: Inhibition of Growth using
Different Cleaning Products
Figure 3: Zone of Inhibition of Dawn
(7) and Dial Complete Antibacterial
Soap (8)
Figure 4: Environmental Microbes
Figure 5: Environmental Microbes
Table 1: Zones Of Inhibition
Resistant
Intermediate
Sensitive
Ampicillin
CD, LB, JW, KS,
DJ, RE <28 mm
-
>29 mm
Bacitracin
CD, LB, JW, KS,
DJ, RE <8 mm
9-12 mm
>13 mm
Chloramphenicol
<17 mm
RE 18-20 mm
CD, LB, JW, KS,
DJ >21 mm
Neomycin
RE <12 mm
JW 13-16 mm
CD, LB, KS, DJ
>17mm
Penicillin
CD, LB, JW, KS,
DJ, RE <28 mm
-
>29 mm
Tetracycline
CD, JW <18 mm
DJ, RE 19-22 mm
LB, KS >23 mm
Conclusions
• Listerine may have had an effect on slowing the
growth of E. coli and S. epidermidis
• Anti-microbial agents differed in effectiveness
based on gram positive versus gram negative
• Be aware of what products used on which types of
bacteria
• Bacteria can be beneficial depending on location
• All were resistant to ampicillin and penicillin
because they were same family
Conclusions
• Sources of Error:
• Contamination
• Pipetting incorrectly
• Further Experiments:
• Different volumes
• Different cleaning agents, i.e. Natural cleaning
products