Test for unknown:
Day 1: prepare a new stock culture on TSA, Streak for isolation.
Do Gram stain and determine if you have a rod or cocci. And if it is gram – or Gram +
Do the gram stain as many times as necessary to gain confidence in your result.
If you have a gram + cocci perform the catalase test and isolation streak culture to
Mannitol/salt agar, and to blood agar.
If you have a gram – rod (any G - non cocci) prepare an isolation streak culture on MacConkey
media to test for lactose fermentation, and to verify Gram negative. Also culture isolation
streak on HE media to test for H2S gas production and further verification of lactose
fermentation.
Day 2: Reculture on TSA, Streak for isolation to verify maintenance of pure culture.
Set up a perform the following tests:
SIM – Test for motility, H2S production, and indole production.
Simmons Citrate Test – Test to see if the organism can use citrate as its sole source of
carbon
Oxidase Test – Test to see if oxidase in used to convert the final cytochrome oxygen to H2O.
(See me for oxidase test paper)
Urease Test – Test to see if the organism is has the enzyme to split urea into ammonia and
CO2
(NH2)2CO + H2O → CO2 + 2 NH3
MR-VP Test Day 3: Read all tests and prepare table:
Gram
+/-
Shape
Rod/cocci/
vibro/Spiral
/spirochete
Motility
+/-
Depth
in SIM
Top/
bottom
Indole
H2S
Oxidase
+/-
+/-
+/-
MR
+/-
VP
+/-
Oxidase
Catalase
Mannitol
Salt
Growth
Mannitol
Salt
Ferment
+/-
+/-
+/-
+/-
Prepare your report:
Describe your gram result and microscopic appearance.
Using the terminology in the Unknown ID document describe your dish culture results for each plate.
List each test performed, give the reason or purpose for the test, and describe the results.
Make your best estimate from the following organism species based on research and your tests:
Enterobacter aergenes
Enterobacter cloacae
Escherichia coli
Klebsiella pneumoniae
Proteus mirabilis
Pseudomonas aeruginosa
Salmonella typhimurium
Staphylococcus aureus
Staphylococcus epidermidis
Shigella flexneri
Proteus vulgaris
Proteus mirabilis
Shigella boydii
Streptococcus pyogenes
Bacillus megatarium
Streptococcus aglactiae
Streptococcus pyogenes
Explain why you chose the organism. I.e., how does your test profile fit your research for the
organism selected?