Transcript Pinning Wrestling Mat Microorganisms

Pinning Wrestling Mat
Microorganisms
Alaina Goos and Ryan Sturm
Introduction
Materials & Methods
Biofilms are a community of microorganisms attached to a
solid surface. A wrestling mat is not usually thought of as a
location for a biofilm. The surface of a wrestling mat has
the potential to become home to millions of microbial cells
including bacteria, viruses, fungi, protozoa, and more. The
wrestling mats at Wartburg College are used almost daily by
numerous athletes with nothing on their minds but winning.
But do they ever think about the cleanliness of the mat?
What other organisms are crawling around on those mats?
A simple inspection of the wresting mat may reveal that we
aren’t the only ones wrestling on these mats. A biofilm that
exists on this mat is not entirely killed when cleaned by the
Wartburg staff as we attempt to show in this experiment.
The experiment was started by first going to the wrestling room
and taking three sterile cotton swabs. Each swab was dipped in
buffer solution and streaked across the wrestling mats. The end
was broken off and the cotton was left in a tryptic soy broth
solution with five percent sucrose and five percent glucose. This
was repeated for all three swabs but on different areas of the mat.
A few hours earlier, the mats were cleaned with Gemesol, an
antibacterial cleaner for the mats. The sample was then
incubated at 35 degrees Celsius overnight. Dilutions of 10-2 to
10-9 were performed and spread plates of each dilution were put
into the incubator for 24 hours. An eight well plate was used to
grow the biofilm of bacteria off the mat. 400 micro liters of Tryptic
Soy Broth were added to each well and then 50 micro liters of the
original bacteria sample were then added to each well. The well
plate was also put into the incubator for 24 hours. After 24 hours,
the spread plate with the dilution of 10-6 had the most isolated
colonies and was used to make bacterial suspensions. By visual
judgment, four different colonies were made into the suspensions
by placing a drop of TSB and a loop of bacteria into the TSB.
Each of the four colonies were streaked onto four different agar
plates. EMB, MSA, TSA, PIA were divided into four sections and
each colony was streaked on each plate. Gram stains were then
performed on each of the four bacterial suspensions to discover if
they were gram positive or gram negative. A live dead stain was
performed by first mixing 1ml of 10% NaCl with 3 micro liters of
live/dead stain while in the dark. The liquid in the eight well plate
was drawn off and PBS was added to rinse the plate twice. Once
the PBS was removed the second time, 10 micro liters of
live/dead stain was added in the dark and let stand for 15
minutes. After 15 minutes, the stain was removed and rinsed
twice with PBS. A fluorescent microscope was used to observe
the stained bacteria.
Isolated
Colony
EMB
TSA
PIA
MSA
Gram
Stain
1
-
+
-
-
-
2
+
+
-
-
-
3
-
+
-
-
+
4
+
+
+
+
+
Results
Table 1
Figure 1a. 10x FITC
Figure 2a. 20x FITC
Figure 3a. 40x FITC
Through our initial spread plate on the TSA plate, we were
able to isolate four different types of colonies. Table 1
shows the results of streaking each of these four colonies on
different media. Table 1 also shows the specific gram stain
on each of the colonies. Colony 1 could be a range of
bacteria. Colony 2 shows characteristics of Salmonella.
Colony 3 could also be a range of bacteria. Colony 4 shows
characteristics of Pseudomonas. The live/dead stains are
shown in figs. 1 through 3. Fig. 1a and 1b show a 10x
magnification of the biofilm created on the eight well slide.
Fig. 2 shows a 20x magnification of the bacteria. Fig. 3
shows a 40x magnification.
Conclusion
•Bacteria was found on the mats although the mats were
cleaned a few hours prior to inspection. This could mean:
•The cleaner used to clean the wrestling mats do not
successfully kill all bacteria.
Figure 1b. 10x TRITC
Figure 2b. 20x TRITC
Figure 3b. 40x TRITC
References
Difco Laboratories. 1998. Difco Manual. 11th ed. Sparks, MD Becton
Dickenson & Co.
Harley, John P., Donald A. Klein, and Lansing M. Prescott. 5th ed. New
York, NY: The McGraw-Hill Companies, Inc., 2002. 620-621.
•A bacterial biofilm may be grown in only a couple
hours.
•Several different types of bacteria can survive on the
wrestling mat by surviving the cleaner, or by improper
cleaning
•To prevent and control skin infections of wrestlers, proper
use of disinfectant/cleaner must occur.