Unknown Experiment
Introduction: The purpose of this experiment is to distinguish and indentify an unknown bacterium. There are several tests that can help one eliminate and narrow down the options. The most useful test, and the very first one done, is a gram stain. This test will tell whether the bacterium is gram-positive or gram-negative. After the type of gram stain is identified, the tester has a wide array of differentiating tests at their disposal. Based on the results from these tests, and the numerous others that are available, one can accurately establish the identity of an unknown bacterium.
Materials: 1-Tube containing unknown bacteria, crystal violet, iodine, decolorizing agent, Safranin, Bunsen burner,
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4.) To further differentiate the bacteria from other options, obtain 1 TSA plate and perform a bacitracin susceptibility test. 5.) Inoculate the plate with a lawn coverage streak and place 1 bacitracin antibiotic disk in the center of the plate. 6.) Incubate the plate, right side up, at 37 C for 24-48 hours. 7.) Record the results from the bacitracin test. 8.) Combine the results and information from the previous tests and establish the identity of the unknown bacteria.
Results: Test Result
Gram Stain
Purple, gram-positive cocci in chains
Morphology
Cocci (in chains), lancelet-shaped and slightly curved
Hemolytic Test
Alpha (Partial Clearing, Green)
Bacitracin Susceptibility
+ (Susceptible)
Conclusion and Background Information: The gram stain test was able to identify the bacterium as a gram-positive organism, and the morphology distinguished it as a strain of streptococcus, and these finds helped direct the other tests. Hemolytic tests are very useful in differentiating specific species of a certain family of bacteria. The hemolytic test showed the bacterium to be of alpha clearing, which meant it only partially lysed the red blood cells and left a greenish tint. This identified the bacterium as either strep pneumoniae or faecalis. The bacitracin test, which tests whether the bacterium is susceptible to the antibiotics in the disk, posted a positive
In this experiment, an unknown bacterium was given to each individual student. The main purpose of this lab was to identify the given unknown bacteria going through a series of biochemical tests as one of the gram negative bacteria among six different Gram negative bacteria Escherichia coli, Enterobacter aerogenes, Klebsiella pneumoniae, Proteus mirabilis, Pseudomonas aeruginosa and Salmonella typhimurium. At the very beginning, streaking method; T-streak technique was used to isolate the pure colonies. For the morphological identification of unknown bacteria, Gram Stain Method was done. Biochemical tests that were conducted for the experiment
Record the results of your investigations of each unknown in Table 1 by completing the following steps:
C. Capsules- does not stain, it supplies nutrients and prevents dehydration for the bacteria. It also helps the bacteria stick to surfaces and increases its
The purpose of this lab was to identify two unknown bacteria cultures using various differential tests. The identification of these unknown cultures was accomplished by separating and differentiating possible bacteria based on specific biochemical characteristics. Whether the tests performed identified specific enzymatic reactions or metabolic pathways, each was used in a way to help recognize those specifics and identify the unknown cultures. The differential tests used to identify the unknown cultures were oxidase, catalase, lactose and sucrose fermentation, Kugler/iron agar, nitrate reduction, gelatin hydrolysis, starch hydrolysis, manitol salt, MR-VP, citrate, bile esculin,
The next step of the project included preparing a Gram stain to discover the cell shape, arrangement, and if the bacteria is gram positive or
The goal of this paper is to demonstrate comprehension which will fully explain why the test were chosen and how they were performed and lastly to to identify the unknown species. Material and Methods: A broth tube labeled as number 24 containing two unknown bacteria was given out by the lab instructor. To begin the identification of the unknown bacteria a streak plate was performed on a Tryptic Soy Agar (TSA) plate, using the quadrant method. The TSA streak plate was labeled at #24-1.
This process is done by using several techniques. In the microbiology lab students were given numbered specimens. The number for this report will be specimen number # 24. The Unknown specimen # 24 was to be identified.
The bases of this experiment was to discover the identify of the unknown from three possible specimens: Klebsiella pneumonia, Escherichia coli, and Enterobacter aerogenes. Utilizaing the T streak technique, the bacteria was isolated into pure colonies for further study. The Gram Stain method was used to identity the morhphology of the bacteria such as the shape and whether the bacteria was Gram positive or Gram negative. Biochemical test were also used to help identify the unknown bacteria. The biochemical test used was the Triple Sugar Iron Agar, Sulfur Indole Motility test, Methyl Red test, Voges-Proskauer test, Citrate test, Urease test, and the Gelatin test. After observing the morphology of the bacteria using the Gram Stain method and utilizing all the possible biochemical test, the bacteria was identified to be Enterobacter aerogenes.
Significant and measurable bacterial growth was observed on all four of the R2A plates. The control disc yielded no zone of inhibition, while the garlic, cleaning solution, and cinnamon oil discs all created measurable zones of inhibition. The zones of inhibition for the TSA+G
The proposed identity of the unknown organism is Staphylococcus hyicus subspecies chromogenes though, due to variability of results for the specie strains, the unknown organism may also be Staphylococcus haemolyticus. All test results were consistent with my partner’s and other groups who had the same unknown organism. No test results contradicted the physiological concept of another test.
The interpretation of the results led us to believe the organism was Staphylococcus aureus. Although some of the results did have complications: The Mannitol Salt Agar appeared to show no signs of growth, but we assumed the color transition of the medium signified minuscule growth. Moreover, the gram stain had complications as well; at first, it appeared to be gram-negative cocci, then after another gram stain we determined that the Unknown was gram-positive cocci. After concluding the gram reaction of the bacterium, a series of tests were performed to differentiate between different genera of gram-positive cocci. The catalase test was a definite positive, which implied the bacterium belonged to Group VI in the lab manual.
Description of the Microorganism: Write a paragraph describing your organism. Please be sure to include the type of organism (bacterial, viral, fungal, protozoa, helminth, etc), morphology (shape, arrangement, colony morphology if applicable), description of structure (gram result, type of nucleic acid or virion structure, spore type, etc if applicable) and also the type of microscope and/or stain you would use to view the organism.
The colonies were smooth, translucent, and had a white brownish color. The Gram stain resulted in Gram positive cocci. After the Gram stain was completed, the bacteria were streaked on a Mannitol-Salt Agar plate and a Catalase test was performed. After these test were completed a Phenol Red Dextrose Fermentation tube was inoculated, and a SIM Tube inoculated.
On the first day of testing, a gram stain test, a KOH test, a catalase test, and an oxidase test were completed. The gram stain test indicated that the unknown species is a gram positive rod, and the bacteria showed no strings during the KOH test confirming that the species is gram positive. From these two tests, the unknown was concluded to be in the family Bacillicae. The bacteria remained yellow on the cotton swab even after the oxidase reagent was dropped onto the swab showing that the bacteria is oxidase negative. For the second day of testing, the bacteria were inoculated on an SBA plate, and the analysis of this plate was difficult. The result of the SBA plate was difficult to discern, and it was unclear if the bacteria completed beta hemolysis or alpha hemolysis. Therefore, an additional SBA test needed to be completed, and with this second test it was more obvious that the SBA plate underwent alpha hemolysis. The SBA result showing negative beta hemolysis, along with the negative result from the oxidase test, indicated that Bacillus cereus was not the identity of the unknown. At the same time that the SBA hemolysis test was undergone, the inoculation of an MSA plate at 37 degrees for 24 hours was also completed. There was yellow growth on the MSA plate indicating that it could grow at 7.5% NaCl and ferment mannitol.
The identification of the unknown organism was first tested using a Gram’s staining procedure. This test uses a series of dyes and liquid solutions that give a bacterial smear a color contrast and aid in its identification. This method differentiates bacteria based on the thickness of their peptidoglycan composed cell walls. This Gram’s staining test was initiated by first donning gloves and obtaining a clean microscopic slide which was cleaned with alcohol. The slide was then marked with a culture number, and a circle was made in the center with a marker. A drop of water was placed in the center and the excess was absorbed. A wire loop was