Dichotomous Key Lab Report

The following tests according to the lab manual were performed: gram stain, fermentation tubes, methyl red, vogues proskauer, sulfur, indole, motility and growing it up on MacConkey agar. The gram stain was performed incorrectly the first time. This is because the decolorizer was not on the bacterium slide for long enough, giving a false outcome.

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.

For many years the identification of microorganisms has been important in the world of medicine. It is essential or correct disease diagnosis in patients and for proper treatment. Knowing the correct identity and characteristics of microorganism is crucial when disease outbreaks occur in populations, also knowing how humans can benefit from microorganisms is important; many can be used in making certain foods or antibiotics.

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

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.

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 I have been learned so far in the microbiology laboratory class for the identification of an unknown bacterium.

3. You test another new unknown bacterial sample, and find the G+C content is identical to one of the samples you have already identified, but the rRNA gene sequence contains one base that is different. What can you conclude: C.

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

A Dichotomous Key was studied to identify bacteria and their relationships. Some of the organisms at the end of the Dichotomous Key had viable characteristics that separate them from different groups, and those that did not students learned how to further classify them. A Dichotomous Key is used to narrow down the search for the unknown organism tested. It is organized by phenotypic characteristics of organisms and conducts a systematic way of identifying the other unknowns. In the lab students were given a tube labeled with a number. Instructions were given to conduct a Gram stain to begin the search followed by the use of a Dichotomous Key and photos as resources to carry out the search. Instructions read to isolate and identify the unknown bacterium with both differential and selective tests to positively identify the given unknown organism. Differential tests used specifically for this unknown microorganism was BEA (Bile Esculin Agar), which interpreted results by the hydrolysis of esculin when the media is blackened around

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).

I saw bacteria growth on both two LB tube which was incubated at 250C and other at 370C. I used one tube for the further test, and the other was incubated at 40C to keep bacteria fresh and growth for other test. On the PEA plate, unknown bacteria did not growth, and on the EMB plate, bacteria had growth very strong. The result confirms one more time the unknown bacteria was gram negative because the PEA plate inhibited DNA synthesis of gram negative so bacteria could not grow on the PEA plate. On the other hand, EMB plate inhibited growth of gram positive so my unknown bacteria grew in this. After I recorded all my result, I continued to set up several test for the next day. Next, I set up PR fermentation test which include 3 broths glucose, sucrose, and lactose. I also set up PAD and Urea

In class, we were given the task of identifying an unknown bacterium broth culture. After receiving number 69, I went through several tests to figure out what bacterium I received. First, I created a slide from my broth by putting a small amount of the unknown broth on to a clean slide and letting it dry for ten minutes. After this, I stained the slide by applying four reagents in order; crystal violet, grams iodine, decolorizer and safranin. From the stained slide, I discovered that this bacterium was gram-negative, which would determine the next couple of tests I would do to identify my unknown bacterium. I began by streaking for confluent growth from my broth culture onto a TSA plate. From the TSA plate, I aseptically transferred a loop

This lab and its procedures are all about finding out the unknown identification of a given bacteria. The lab consists of specific techniques, tests, chemicals, and vocabulary that are necessary for the finding of the bacterial identity. A bacterium is randomly assigned and it is a group effort to find the bacteria name through many of its specialties and characteristics. An example of classifying it would be to determine whether the bacteria is catalase negative or positive, or if the species is gram negative or positive. This lab is of huge significance because of its medical microbiology connections. Scientists Gurtler and Stanisich explained the connections more eloquently. They stated, in their medical article, that, “Medical microbiology

The decolorized Gram negative cells are stained pink. With the results from the Gram stain I was able to follow the “Unknown Identification Flowchart” to the next step, which was to prepare for the Starch Hydrolysis Test by inoculating a starch plate.

Determination of the bacteria being a lactose fermenter or non fermenter is done through the growth on the MacConkey agar. Knowing this allow for the student to perform the necessary tests to determine which lactose fermenter was present in the sample. The Indole test allows for the determination of whether the unknown bacteria is Escherichia coli because this genus and speices is the only lactose fermenter that will produce a positive result here. Moving onto the Methyl red test this indicates glucose fermentation, more specifically microbes that produce mixture of acids as a result of fermentation. The Voges-proskauer tests for glucose fermentation, specifically organisms whose acid is converted to acetoin. The Citrate test differentiates an organism’s ability to use citrate as its only carbon source. The urea broth culture detects the enzyme urease, which allows break down of urea producing acid, which causes a noticeable change in color. The final test is the motility test to determine if the bacteria has the capacity of movement beyond the point of