Unknown Lab Report The mixed unknowns were gram stained and looked under the microscope. There were gram-positive cocci and gram-negative bacilli. The unknowns were inoculated in blood agar and Mac-conkey. Three types of colonies were seen in the blood agar. All the three organisms showed different hemolysis, as there were alpha, beta and gamma hemolysis present. In the mac-conkey, there was colorless colony that denotes lactose non-fermenter. Each colonies were then inoculated into different blood agar to do further testing.
Organism A
I. Microscope- Gram positive cocci
II. Blood Agar – Beta hemolysis Complete hemolysis.
III. Catalase test– Positive Presence of bubbles when catalase was added to the slide with the organism in it as it can convert hydrogen peroxide into water and oxygen.
2H2O2---- 2H2O + O2
The organism falls in the genus Staphylococcus or Micrococcus.
IV. Coagulase test, Staphylococcus Latex test – Positive Clumping of the latex reagent seen within 20 seconds. The organism makes coagulase.
These tests suggest that the organism is Staphylococcus aureus. So for confirmatory test, the organism was inoculated in mannitol salt agar and incubated. The result was yellow color colony (mannitol fermenter), which means it is S. aureus.
Organism B
I. Microscope- Gram negative bacilli
II. Blood agar- Gamma hemolysis No hemolysis.
III. Mac. Conkey- Colorless colony The organism does not ferment lactose.
IV. Oxidase test- Negative The organism falls under enterobacteriacea.
V. Indole test- Negative The organism does not metabolize tryptophan so no indole formation. This test narrowed down the organism to salmonella, shigella and Proteus mirabilis, as these organisms are enterobacteriacea, indole negative and lactose non-fermenters. LIA was the next test done.
V. LIA- K/K (purple top/ purple butt) and H2S production The organism ferments glucose, decarboxylates lysine and it produces H2S.
These results suggest that the organism is salmonella. To confirm, the organism was inoculated in Salmonella Shigella (SS) agar. The colony appeared colorless with black center. So, the second organism is salmonella as only salmonella or shigella can grow in that plate. The presence of H2S confirms it is salmonellas because shigella does not produce H2S.
Organism C
I. Microscope- Gram positive cocci
II. Blood Agar- Alpha hemolysis Partial hemolysis. Green color.
III. Catalase test- Negative (No bubbles) The organism is not able to convert hydrogen peroxide into water and oxygen. The genus of the organism is Streptococcus. There are two Streptococcus organisms that show alpha hemolysis. S. pneumonia and S. viridians. To confirm it is one of them, the organism was inoculated with optochin disk, as they are sensitive and resistant respectively.
IV. Optochin disk- Zone of inhibition around the impregnated disk. The organism was sensitive to optochin. Therefore the third organism is Streptococcus pneumonia, as other alpha hemolytic streptococci do not display the zone of inhibition with the presence of optochin.
The bonus organisms 1. Klebsiella 2. Abiotrophia 3. Escherichia coli 4. Micrococcus