| Gram Negative Unknown | Biology 3444-006 | | Lena Wallace | 11/7/2011 |
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Abstract:
The purpose of this lab was to identify an unknown bacteria culture using differential tests. The identification of the unknown culture was accomplished by identifying the bacteria based on its specific metabolic characteristics and morphology. It is suggested that culture 11 is a sample of Enterobacter aerogenes.
Introduction:
This experiment was centered on metabolic and biochemical testing procedures. The rationale of performing these tests was to distinguish six different microbes from one another and to compare how their metabolic and biochemical processes differ from species to species to determine the unknown sample.
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If the broth turns yellow, it means that acid was produced and reported as A. If the organism can break down the amino acids be de-amination and ammonia is produced, this will raise the pH level turning it pink. This alkaline result was reported as K.
The Methyl Red test is a differential test for bacterial respiration used to differentiate strains of coliform bacteria capable of performing mixed acid fermentation that will lower the pH despite the phosphate buffer (http://faculty.deanza.fhda.edu). Mixed acid fermentation is confirmed by using methyl red as an indicator. It is red ant pH 4.4 and below, yellow at pH 6.2 and above, and orange in between. Red is a positive result reported as (+), yellow is a negative result reported as (-), and orange is negative or inconclusive.
The Voges-Proskauer test to detect organisms that are able to ferment glucose, but convert the products to acetoin and 2,3-butanediol. This is deduced by the addition of Reagent A and Reagent B, and the observation of the color change thereafter. Reagent A is a solution of -naphthol and alcohol. Reagent A catalyzes the conversion of acetoin to diacetyl. Diacetyl thens react with guanidine-containing compounds from the peptone to form a red color in the presence of -naphthol. Reagent B is a solution of potassium hydroxide and water. It
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 purpose of this lab was to identify two unknown bacteria from a mixed culture. The reason for identification of unknown bacteria was to help students recognize different bacteria through different biochemical tests and characteristics. This is important in the medical field because identification of unknown bacteria can help treat a patient by knowing the contributing source of a disease. Also knowledge of different bacteria helped others make antibiotics used today. This lab was completed by using the methods learned thus far in identification of bacteria.
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
The purpose of this study project was to carefully isolate and identify two unknown bacteria from a mixed culture. The ability to properly evaluate biochemical test results is also necessary for the identification to be successful. The goal was to apply all of the methods and techniques that have been learned in the microbiology laboratory course for the proper identification of unknown bacteria. A certain amount of bacteria that were used throughout the course were possible bacteria that could be found in a mixed culture. The bacteria that were identified in the mixed culture were Staphylococcus Aureus and Kocuria Rhizophila.
I identified Citrobacter freundii, the gram negative rod, by running a series of tests. I began with the Phenol Red Lactose tests, which tests if the organism contains various enzymes that determine if it can ferment lactose. The broth turned yellow after it was incubated, indicating that the lactose was fermented to acid, and there was also gas present in the Durham tube. Since the Phenol Red Lactose Test was positive, I then ran the Phenol Red Sucrose test, which tests if the bacteria contain different enzymes that determine if sucrose can be fermented. After incubation, the broth was yellow, indicating that sugar was fermented to acid, and there was also gas present in the Durham tube. Next, I ran the Sulfide Production, Indole Formation, Motility test, but I was only testing for Hydrogen Sulfide Production to differentiate between the organisms Citrobacter freundii and Enterobacter aerogenes. This test detects if the organisms can metabolize sulfur into hydrogen sulfide, which is revealed by the formation of ferrous sulfide that causes blackening around the growth. The test also reveals if the organism can break tryptophan into indole or migrate away from initial stab area. After incubation, the agar slant was completely black, indicating that the organism produces hydrogen sulfide and is motile proving that it was Citrobacter
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
|MSA Agar |For organisms that are |Isolates for mannitol fermentation |Yellow color change in |Staphylococcus aureus |
An unknown bacterium was handed out by the lab instructor. The methods that have been learned so far in identifying bacteria were applied to this unknown. Procedures were followed as stated in the lab manual and biochemical test handouts. The first procedure that was done was a gram stain followed by a streak of the unknown on a TSA plate in order to determine the gram reaction and observe the colony morphology. After that, specific biochemical tests were performed for gram positive, since unknown number five was determined to be gram positive rod. The other tests were performed in this order: Mannitol Salt (MSA) streak, Blood Agar streak, Catalase test, Nitrate Reduction test, and Phenyl
There are many reasons for knowing the identity of microorganisms. The reasons range from knowing the causative agent of a disease in a patient, so as to know how it can be treated, to knowing the correct microorganism to be used for making certain foods or antibiotics. This study was done by applying all of the methods that have been learned so far in the microbiology laboratory class for the identification of unknown bacteria. The identification process can be completed with a series of deferential stains and biochemical tests. Creating a dichotomous key helps to limit the amount of biochemical tests done on an unknown organism and by observation
To perform this test, a tube of broth rich with glucose is acquired. In this tube is phenol red, a pH indicator. Initially, the tube appeared pink in color, indicating a normal pH level. Next, a sample of unknown #44 is introduced into this medium using the aseptic technique, and this is allowed to sit for several days. If the organism is able to ferment glucose, the pH in the medium would decrease and cause the phenol red to exhibit a yellow color. In addition to the straw color, gas can also be produced and trapped inside the Durham tube placed in the medium. This production of acid and gas is a direct result of the fermentation of glucose, as seen with unknown
The main idea of this experiment was to correctly identify the unknown bacteria, #3. Identification of unknown bacteria yields multiple benefits in many different areas in the research of microorganisms. In this experiment I performed many different test dealing with things such as the presence of enzymes, fermentation abilities and different chemical reactions. Observations made from the tests were then compared to a gram negative unknown chart in order to identify the bacteria. Based off of my results and the chart, I concluded the bacteria #3 was the bacteria Escherichia coli. E. coli is most commonly found in the intestines of warm blooded organisms. Most E. coli strands are non pathogenic however, there are strands
The purpose of this study was to identify the unknown bacterium using biochemical tests and various methods that had been learned from previous the microbiology laboratory class. Identifying the unknown bacterium was determined by separating and differentiating possible
2. Introduction: Each student was given unknown bacteria and was instructed to perform a variety of experimental tests that would help to identify their bacteria. During the process of identification, the unknown bacteria was added to many different testing medias using aseptic technique. They are as follows: lactose fermentation on eosin methylene blue (EMB), TSI (Triple Sugar Iron agar), Phenol red sucrose, the SIM test, H2S by SIM, IMViC (indole, motility, voges-proskauer, and citrate), Urease (urea broth), PDase (Phenylalanine Deaminase), Lysine Decarboxylase, and Ornithine Decarboxylase. Colonial morphology on EMB was used to
Each mixed culture that was tested had one gram positive and one gram negative bacterial species. The possible species of bacteria that could have been isolated from the mixtures included the following: Bacillus subtilis, Corynebacterium glutamicum, Escherichia coli, Staphylococcus aureus, Enterococcus faecalis, Enterobacter aerogenes, Salmonella enterica, and Pseudomonas aeruginosa. The identities of the unknown species were determined through comparing the experimental data against data acquired from earlier experimentation.