Unknown Sample Experiment
G. Brian Blaga
“Sample C”
Day 1
The sample was received on an aqueous solution and then plated on growth
medium in hopes of observing growth of colonies. Using sterile techniques, the
sample was plated on MacConkey Agar (MAC) and Phenylethyl Alcohol Agar (PEA).
The reasoning for this is to determine if the sample contained Gram + and/or Gram . MacConkey is known to be selective for Gram – while it inhibits Gram +. The
reverse of is true for the Phenylethyl alcohol agar, selective for Gram +, inhibits
Gram -.
Results:
The PEA grew multiple white colonies, indicating the presence of at least one type of
Gram + organism. The MAC agar grew multiple colorless colonies, indicating the
presence of a Gram – organism. Not only is MAC selective for Gram – bacteria, but is
also capable of differentiation lactose fermenting and non-lactose fermenting
depending on the color. Since it was colorless and not red, we determined our Gram
– sample was lactose non-fermenting, helping narrow down the choices.
Day 2
To further narrow down the options for our Gram – species, a colony from the MAC
agar was removed and plated on a Xylose Lysine Deoxychioate agar (XLD). The
reasoning XLD was chosen is because it would be able to differentiate whether the
species was Salmonella, shigella, or pseudomonas aerguinosa depending on the
colors of the colonies.
Colonies of the Gram + unknown were taken from the PEA and administered both
catalase and oxidase tests.
Both species were then inoculated in their own broth to ensure there was a back up
sample.
Results:
Gram +: The sample gave a positive results for the catalase test. The sample then
gave a negative result for the oxidase test.
Gram -: A problem occurred with our Gram – sample as yellow colonies grew on
XLD which actually indicates that the sample is a lactose fermenter and could
possibly be Enterobacteria.
Day 3
The Gram – species was plated on EMB to determine if the sample was (or if we
could eliminate the possibility of it being) E. coli.
The Gram + sample was then streaked onto a blood agar for hemolysis to be noticed.
Results:
The Gram + sample which was plated on the blood agar turned out to be white, with
no changes to the media, indicating that the sample is gamma-hemolysis.
The Gram – species grew as pink colonies on the EMB plate, excluding E. coli.
Day 4
Narrowing down our choices for the Gram + with it being oxidase negative, catalase
positive, and non-hemolytic, we narrowed it down to Staphylococcus aureus and S.
epidermidis. To differentiate between the two, the species was grown on a Mannitol
plate, which is selective for Staphylococcus aureus.
Due to confusion and mixed test results for the Gram – sample, an Enteripluri tube
was inoculated with the sample.
Results:
Colonies ended up growing on the Mannitol plate, indicating that our Gram + sample
was Staphylococcus aureus.
The Enteripluri tube gave negative VP and Indole tests. It also confirmed that the
sample was non lactose fermenting. The results of the Enteripluri tube leads to
believe that a problem occurred during the XLD test. The results of the Enteripluri
tube indicated that the Gram – sample was Klebsiella ozaenae.
Gram +: Staphylococcus aureus
Gram - : Klebsiella ozaenae.