Results for
Unknown #21
Gram Stain
Gram +
Other Staining:
None
Morphology
Large, rods
Arrangement
Chains, and sometimes pairs
Cell size
Large
Colony:
Color/Pigmentation
Margin
Elevation
Appearance
Opacity

Unpigmented, beige
Smooth
Flat to Raised
Dull
Opaque
Enzymes:
Catalase
Amylase (Starch Test)

+, Bubbles, 02 produced
+
Fermentation:
Glucose
Lactose
Sucrose

+, A, Red to Yellow
+, V, Slight Red to Yellow
+, A, Red to Yellow
MR-VP Tests:
Methyl Red
Voges-Proskauer
Citrate

+ Red to Yellow
- No color change
+ Green to Blue
H2S Production

-
Kligler’s Iron Agar (glucose and lactose)
+, A/A, Yellow produced
Organism Name:
Bacillus megaterium

Results:
Microorganisms are all around us playing a role in our everyday lives. Knowing the difference between different microorganisms can not only help to identify a disease, but also what’s causing a disease in a patient, and what antibiotic can help treat that disease. Using the tests observed and learned in microbiology, correct identification of an unknown microorganism (Unknown 21) was performed. Verifying the purity of the unknown was crucial in this experiment. Contamination was always a concern throughout the entire process. The aseptic technique was of the utmost importance throughout the discovery of my unknown. With each procedure done carefully, contamination was not an issue. From incinerating the loop to inoculating a plate, it is with meticulous efforts, that each step was performed aseptically. Gram staining played a critical role in the identification of the microorganism. Gram staining is widely used to eliminate thousands of possible organisms because it confirms Gram negative or Gram positive organisms and cell morphology. In this experiment, the goal was to correctly identify one of seven microorganisms, however, in this case, the use of Gram staining eliminated most from the very beginning.
The process of Gram staining includes the use of crystal violet, iodine, decolorizer, and safranin. Depending on whether the microorganism is Gram positive or Gram negative, a purple or pink pigment will appear. Unknown 21 appeared as a purple color, therefore it was possible to rule out all of the Gram negative bacteria including Escherichia coli, Eneterobacter aerogenes, Proteus vulgaris, and Pseudomonas aeruginosa. Upon closer examination with the microscope, the morphology of the unknown was observed. A Gram stain was performed numerous times in the laboratory, it was clear that the unknown was purple, Gram positive, large rods. Having an idea of the unknown organism from the beginning was helpful, however, it was fascinating to perform each experiment to confirm the findings. All of the remaining six microorganisms could have potentially been eliminated and it is with each experiment, more and more confidence was built with identifying the unknown. While performing the Gram stain, amongst the purple bacilli, a clear area in the center of the rods. These clear areas were identified as the bacteria’s endospores. Because endospores are sturdy and impermeable to crystal violet, they appeared as transparent holes in the cells. This was a crucial point in the unknown process to identifying unknown 21 microorgamism, as Bacillus megaterium was the only organism to contain endospores.
Another test that was conducted to determine the unknown is the fermentation test using glucose, lactose, and sucrose. Fermentation tests are used to determine if a bacteria is capable of carrying out various fermentation reactions. If an organism ferments a sugar, an acid is usually produced, and gas can be an end product. The presence of acid is indicated by a color change in the pH indicator, phenol red, from red (alkaline pH values) to yellow (acidic pH values.) The production of gas such as hydrogen and carbon dioxide is revealed by the displacement of medium in the tube. In the experiment, the