Oxidase Test Lab Report

An unknown bacterium was handed out by Dr. Honer. The appropriate tests were prepared and applied. The first procedure that was done was the gram stain. Under a microscope, if the gram stain is purple, the bacterium is gram positive, if the stain is red, it is gram negative. The next test was the fermentation tests for glucose, sucrose and

The preparation for the experiment started by gathering the solutions of enzyme Peroxidase, substrate hydrogen peroxide, the indicator guaiacol and distilled water. Two small spectrometer tubes and three large test tubes with numbered labels. In addition, one test tube rack, one pipet pump and a box of kimwipes were also gathered. Before the experiment, the spectrometer must be set up to use by flipping the power switch to on. Following, the machine was warmed up for 10 minutes and the filter lever was moved to the left. In addition, I set the wavelength to 500 nm with the wavelength control knob. Before the experiment, I had to create the blank solution by pipetting 0.1 ml of guaiacol, 1.0 ml of turnip extract and 8.9 ml water into tube #1. Following the creation of the blank, a control 2% solution was created.

The first result of importance was the result of the Gram stain. The observations of the unknown bacteria from the slant culture after Gram staining showed that the unknown bacteria were Gram negative bacilli (Image 1). After determining the unknown bacteria was Gram negative, an oxidase test was conducted on a sample from the slant culture. The cotton swap with the sample of bacteria did not change color when the oxidase reagent was applied, thus providing a negative result. With a negative oxidase test, further tests were conducted to determine various characteristics of the unknown bacteria. A MR-VP broth was inoculated with a sample from a slant culture of unknown bacteria. After incubation, the methyl red reagent was added to the broth, and the broth turned red, providing a positive result (Image 2). An EMB agar streak plate was inoculated with a sample from a slant culture of the unknown bacteria, and after incubation, growth was found on the plate, providing a positive result (Image 3). A Citrate agar slant was inoculated, and after incubation, growth was found on the media, providing a positive result (Image 4). A Urea agar slant was inoculated, and after incubation, the agar had changed from a peach color to a bright pink color, providing a positive result (Image 5). Using the flowchart (Figure 1) developed from the Table of Expected Results, the lab partners started at the oxidase test. Given the negative result of the oxidase test, the flowchart is

I began by running the starch test, which tests for the presence of starch hydrolyzing enzymes. After doing a one-line inoculation of the organism, the plate had to be incubated. Once I received an appropriate amount of growth I added the reagent iodine. The iodine turned the plate purple, formed no clear zone, and lifted the organism off of the plate, which revealed that the starch was not degraded and the enzyme was not present. The organism being lifted off the plate is unique to the bacteria Corynebacterium xerosis indicating that it was my gram positive rod. For reassurance, I ran the Phenol Red Glucose test, which tests if the organism contains various enzymes that determine if the bacteria can ferment glucose. After incubation, the broth turned orange, but this did not provide a clear positive or negative result so I ran the Nitrate Broth Reduction test. The Nitrate Broth Reduction test detects if the organism utilizes nitrate. After incubation for forty-eight hours I added Nitrate A and Nitrate B indicators. However, there was no color change indicating that the test was inconclusive. Since the test was inconclusive, I proceeded to the following step, which included adding a small amount of zinc to the broth, and this turned the broth a red color. The red color indicated that

This test is used to detect if the bacteria contains any deoxyribonuclease activity. Because no color change was observed from blue to clear my unknown bacteria displayed a negative result.

In a laboratory setting, it often becomes necessary to identify an unknown organism. In this experiment, researchers classified an unidentified bacterium based on its physical structure, colony morphology, optimal conditions and metabolic properties. A Gram stain using crystal violet, iodine, and safranin and a simple stain using methylene blue characterized the organism’s cell wall. Cultural behavior was classified by inoculating the organism onto nutrient agar and incubating it at 37° C for 48 hours, and observing its behavior, as well as using SIM medium to test for motility. Optimal growth temperature was

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

The purpose of this lab was to identify unknown bacteria cultures using various differential tests, and my unknown bacteria is #17. 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 Gram stain, Catalase, Mannitol Salt Agar (MSA), Blood Agar, Novobiocin, Coagulase, and DNAse (Alachi, 2007).

The enzyme urease breaks urea down into NH3 and CO2. An orange broth containing urea is used for this test and needs to be inoculated with the gram negative bacteria. A pink color in the medium indicates a urease-positive organism, an orange or yellow is negative.

The first step toward identifying this unknown organism was to perform a Gram Stain to differentiate between gram positive and gram negative bacteria. This is an important step because it directs what the next tests will be. My Gram Stain on sample #12 showed that the bacteria was gram negative, however, after receiving the results of the OF glucose, H2S, Citrate, Urease and Motility tests, it was apparent that my Gram Stain was contaminated. I then performed a catalase test which came back negative, so I ordered a Bacitracin disc, Optochin disc and a CAMP test which had to be incubated overnight. After receipt of those test results,

For the Urease test, I incoluated my Urea test tube with my unkown bacteria from a TSA plate using and inoculating loop. The Urea tube was then incubated at 37 degrees Celsius for 8 days to observe for a color change. The Urea tests for the ability of a bacteria grown in urea broth produces urease. This medium contains the pH indicator phenol red. If urease is produced the pH of the media will raise thus causing the phenol red to change from yellow to a pink color.

This experiment was given to us to utilized previous knowledge learned throughout the semester to identify a gram negative unknown bacterium. We had to first learn the difference between a gram negative and a gram positive organism. We started off with doing gram stains to determine whether it was positive or negative. Based on the gram stains, a gram positive stains purple and a gram negative stains pink. A gram positive stains purple because the cell walls is made of a thick peptidoglycan layer and doesn’t

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.

A great deal of cultural tests and physiological procedures were performed on the unknown before the conclusion of P. aeruginosa was made. The first test preformed was Gram’s staining which involves the use of crystal violet, Gram’s Iodine, Ethyl Alcohol, and Safranin. Gram staining is known as a differentiating stain, since it can determine the presence or absence of peptidoglycan in the cell wall. The unknown bacteria were Gram stain negative. After that the motility of the cell was tested using a hanging drop slide. The slide was examined, but the results were inconclusive due to the inability to make a clear distinction between motility and Brownian motion. This test was not

The Indole portion of the test is performed by adding Kovac's reagent to the inoculated SIM medium. The Kovac's reagent reacts with the indole (if indole is present) to produce a pinkish-red or reddish-purple ring around the top of the test tube. If indole isn't present, there will be no color change. The presence of indole indicates that the bacteria produces tryptophanase, an enzyme which breaks down tryptophan into smaller components, one of which being indole.