UNKNOWN LAB REPORT
Report by TABITHA MATTHEWS
MICROBIOLOGY
FALL 2012
INTRODUCTION
The discipline of microbiology has many applications in science and medicine. This unknown
laboratory study was an exercise in identifying two bacterium using varied scientific techniques
in a lab. The practical application of this study includes determining unknown bacteria in/on
patients, surfaces, and foods using varied laboratory techniques.
MATERIALS AND METHODS
An unknown bacteria tube, number 102, was distributed randomly by the course instructor. Methods
outlined and practiced in Lab Manual for General Microbiology were used in the
determination of unknown bacteria from tube 102 as well as an alternate Gram-negative
bacterium. Procedures outlined in the lab manual were followed.
The first procedure completed was the growth of two isolation streaks on nutrient agar plates
from mixed culture 102. Two different cultures grew and were visibly observed, isolated, and
grown on separate nutrient agar plates labeled cultures “A” and “B”. Culture “B” grew pure on a
nutrient agar plate, and culture “A” was re-plated to isolate a more pure culture. A Gram stain was
then performed on the two different cultures and it was determined that culture “A” was a Gram-positive cocci and culture “B” was a Gram-negative rod.
The following tests were performed on Gram-positive culture “A” and are documented on
Results Flow Chart 1:
1. Nitrate
2. Mannitol Salt Agar
3. Urea
4. Catalase
Note: The results from the above test were inconclusive, reflected in the chart.
The following tests were performed on Gram-negative culture “B” and are documented on
Results Flow Chart 2:
1. Simmon’s Citrate
2. Urea
3. Mannitol Salt Agar
4. Nitrate
6. Methyl Red
7. Fermentation Tubes for glucose, galactose, and maltose
Note: The results from the above test were inconclusive, reflected in the chart.
After completing the above tests for both cultures the identity of the unknown bacteria was still
unknown. After repeating Gram stains for both cultures it was determined that both cultures
were Gram positive. Based on the tests it was not determined what the identity of Culture “A”
was. Culture “A” was discarded. Culture “B” was then determined to be a Gram positive
bacillus based on Gram staining and was an uncontaminated pure culture.
The following tests were performed on Gram positive culture “B” and are documented on
Results Flow Chart 3:
1.Methyl Red
2. Oxidase
3. Glycerol Fermentation
4. Maltose Fermentation
An alternate Gram negative was then randomly assigned as Alternate # 3 and methods and tests
from the lab manual were used to determine its identity. Tests were completed on the Gram
negative pure culture following its growth on a nutrient agar plate and a Gram stain to confirm it
was Gram negative.
The following tests were performed on Gram negative alternate culture and are documented on
Results Flow Chart 4:
1. MRVP
2. Urea
3. Deoxycholate
**Flowcharts in RESULTS was removed due to formatting reasons.
DISCUSSION/CONCLUSION
After a Gram negative pure culture was assigned because of a contaminated mixed culture, a
careful order of tests was followed that led to Proteus vulgaris as the unknown from the alternate
bacterium. A positive methyl red test indicated acid production during glucose fermentation and
narrowed the possible bacterium down to three with a Vogues-Proskauer test following. A
negative VP result indicated the bacterium did not produce acetyl methyl carbinol and could only
be Escherichia coli or P. vulgaris. A urea test, testing for the activated enzyme urease was
positive, ruling out E. coli. It was concluded that P. vulgaris was the unknown. To confirm, the
bacterium was plated on a desoxycholate plate as a differential test for P. vulgaris. Confirming
the results were correct, P. vulgaris grew in clear colonies, fermenting xylose but not producing
acid.
P. vulgaris part of the Enterobacteriaceae family. It is commonly found in fecal and
decomposing matter, soil, and water. P. vulgaris is commonly associated with urinary,
respiratory, skin, and eye infections. Although P. vulgaris is an enteric bacteria, is does not
grow well in the intestinal tract, thriving in the urinary tract instead. Many urinary tract
infections are caused by P. vulgaris. Antimicrobials that are most effective in inhibiting P.
vulgaris are ampicillin and aminoglycosides.
REFERENCES
Mosby’s Medical Dictionary, 8th edition. (2009) from http://medical-dictionary.thefreedictionary.com/Proteus
+vulgaris,
