Introduction:
Although the majority of microorganisms are either harmless or beneficial, certain species are
capable of causing disease. The human body is home to approximately 90 trillion microbial cells some of
which are opportunistic pathogens. Opportunistic pathogens are organisms that do not normally cause
infections in healthy individuals, only when they enter into the body were they are not normally found
or if a patient is immune-compromised do they cause infection. Modern clinical diagnosis of bacterial
infections usually involves the use of the Gram stain and observation of cellular morphology, colony
morphology, and biochemical testing. The Gram stain is the first step in identifying an unknown
microorganism, based on the Gram stain reaction the proper battery of biochemical tests are used for
further diagnosis. Cellular and colony morphology allows microbiologists to narrow the search when
identifying the disease causing organism. Biochemical tests are tests that differentiate between bacterial
species based on metabolic by-products. Biochemical testing is however based on percentages therefore
they must be used in combination with the Gram stain and colony morphology in order for the correct
identification of the disease causing bacteria.
In this case study a patient presented to the hospital with a severe skin wound infection on the
upper forearm. The wound area showed swelling localized to the site of infection with redness
spreading from the wound. Applying pressure to the wound resulted in pain and oozing pus. The
infection site was swabbed and Gram stained, additional swabs were taken for diagnostic testing. Based
on the signs and symptoms and the initial Gram stain showing Gram-positive coccus arranged in clusters
indicates the skin wound infection is most likely caused by an organism from the genus Staphylococcus.
The following biochemical tests that will be discussed will determine if the causative agent is
Staphylococcus aureus or Staphylococcus epidermidis which will allow for proper treatment.
Materials and Methods:
Sugar fermentation tubes
A pure culture of the suspected pathogen was used for all biochemical tests. Sugar fermentation
tubes include glucose, lactose, sucrose, and mannitol these are biochemical tests which test for an
organisms ability to ferment sugar. A positive test is indicated by a color change from red to yellow
indicating the production of an acid by-product. The pH must drop below 6.8 in order for a change to
occur. Weak positives may present if the pH is close to 6.8, indicated by a color change from red to
yellow-orange. To test for sugar fermentation capabilities one loopful of organism was aseptically
inoculated into each of the four sugar tubes. Sugar tubes were then incubated for 24 hours at 37
degrees Celsius.
Test for Motility
Motility agar was used to test for the presence of flagella. A positive test is indicated by growth
and media color change to violet/red. A spreading pattern into surrounding media may also be
observed. Motility agar was incubated for 5 days at 37 degrees Celsius.
Test for oxygen usage
A fluid thioglycolate tube was used to test for this organisms capability to use oxygen during cell
respiration. The upper portion of the agar medium contains the greatest amount of free oxygen
because the gas may easily diffuse into the medium from the atmosphere. The large center zone
contains a decreasing amount of oxygen because the gas has more difficulty reaching that portion of the
medium. If turbid growth is only seen at the top of the media, this indicates an aerobic organism. If
growth is observed throughout the entire media this indicates a facultative anaerobe. Fluid thioglycolate
tubes were inoculated with one loopful of bacteria directly below the surface of the media, the loop was
not agitated in the media only dipped as to not introduce oxygen into the lower portion of the media.
The tube was then incubated at 37 degrees Celsius for 24 hours.
Continue describing the tests you performed using the above format!!!
Results (put in same order as you did for the materials and methods)
Sugar fermentation
Following incubation of the sugar fermentation tubes the results show that this organism
fermented glucose, lactose, and sucrose indicated by the media changing to a yellow color. Gas was not
recovered in the gluscose broth durham tube indicating a slow fermentation rate. The mannitol
fermentation tube was weakly positive showing a yellow/orange color.
Test for motility
Following incubation of the motility agar results show this organism is non-motile indicated by
growth only along the stab line from the inoculating needle. No swarming affect was observed.
Continue describing your results using the above format!!!!
Discussion (must compare results to Bergey’s manual for diagnosis!)
Things to include:
What was your organism? General description of your unknown: recap some of the
major findings that helped you determine your unknown. Discussion of erroneous
results. Provide a statement like I have below
Analysis of biochemical testing indicates results consistent with Staphylococcus aureus. As
mentioned biochemical testing is based on percentages 90% of the results reported are consistent with
the characteristics described in Bergey’s Manual for Systematic Bacteriology. Staphylococcus aureus is a
Gram positive coccus shaped bacterium that is usually characterized as catalase positive. The catalase
test is one of the key tests used to differentiate Gram positive organisms such as Staphylococcus and
Streptococcus organisms. According to Bergey’s Manual S. aureus is classified as a Mannitol fermenter.
The strain causing the skin wound infection described earlier only weakly fermented Mannitol. Longer
incubation may have resulted in a strong positive result. The key test to conclude the causative agent for
the skin wound infection described earlier was the growth and fermentation of Mannitol on Mannitol
Salt Agar. The fermentation of Mannitol confirmed the organism was Staphylococcus aureus not
Staphylococcus epidermidis.
Name some notable diseases caused by this organism
Discussion of virulence factors
Propose Treatment
Conclusion
The human body is composed of approximately 100 trillion cells, only 10 percent of those cells
are actually human cells the remaining 90 percent of cells are microorganisms which are mainly
harmless or beneficial. When given the opportunity some species of normal microbiota are capable of
causing disease. One such organism is Staphylococcus aureus, S. aureus is a Gram-positive coccus
arranged in clumps. This organism is considered to be normal microbiota and equally known to cause
diseases such as skin wound infections, Toxic shock syndrome, and food borne intoxication. Often
infections caused by S. aureus are easily treated with antibiotics however, resistance to antibiotics such
as methicillin is becoming more prevalent. Humans are constantly exposed to potential disease causing
microbes including some found on the human body itself. To avoid a severe skin wound infection caused
by S. aureus, such as the one described in this study, proper wound care should be administered
promptly upon becoming injured.