Introduction 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). Rebekah Worley February 21, 2012 Mitchell Section 4 Biol 311 Staining and Identifying Unknown Bacteria Introduction: The microbiology lab up to this point has been used to teach the students how to stain and identify bacteria. There are several types of staining through which the bacteria can be identified based on the color and shape. The staining methods used in the lab are Gram Staining, Capsule Staining, Endospore Staining, and Acid Fast staining.
One of the most significant method of staining is the Gram Staining, as it is highly dependent (McCarthy, 25). In the specific experiment that was done, Gram Staining was used and the bacteria that was found was purple and round (cocci) shaped. Through this the bacteria was identified as Staphylococcus epidermis. Material and Methods: The first step to identifying the bacteria was to heat fix it to the slide. The materials used were a slide, water, a Bunsen Burner, bibulous paper and clothes pin. The unknown bacteria was in a vial in solid form.
The steps on page 19 and 20 of the Customized Biol 311 General Microbiology Laboratory Manual were followed to heat fix the bacteria. After this gram staining was used to identify the unknown bacteria. The materials used for gram staining include the slide the was heat fixed, bibulous paper, crystal violet, distilled water, Gram’s iodine, 95% ethyl alcohol, safranin, oil and a microscope. The steps on page 26 of the Customized Biol 311 General Microbiology Laboratory Manual were used to stain the bacteria. Several changes were made in the procedure.
The crystal violet was on the slide for 1 minute rather than 20 seconds. The decolorizing step was used with alcohol for 10 seconds rather than 20 seconds. The only other change was that the safranin was on the slide for 1 minute instead of the recommended 20 seconds. The slide was put under the microscope at 1000x magnification using oil immersion. Results: When looking under the microscope the bacteria was found to… Material & Methods The tests performed on the unknown bacteria cultures were all used to determine the identity of the bacteria.
Each of the tests performed provided some key information about the bacteria in question and how it functions. Not all of the tests were performed on every culture, however, as some of the tests were used only for gram (+) or gram (–) bacteria, while others were even more specific and used only for cocci bacteria. The tests performed and what constitutes a positive and negative test are as follows. Gram staining was used to determine whether the cells were gram positive or gram negative based on the color they appear at the end.
The Gram stain utilizes four basic steps: apply a primary stain (crystal violet), fix it with Gram’s iodine (which fixes the primary dye inside the cell), decolorize with 95% ethyl alcohol to wash out the crystal violet-iodine (CVI) complex, and apply a counterstain, Safarin. Gram positive cells have a thick peptidoglycan layer that is external and has a higher degree of cross-thinking that traps the CVI complex better than gram negative cells. Therefore, they are less susceptible to decolorization by alcohol making them appear purple in color. Introduction
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. There are many reasons for identifying an unknown bacterium.
The reasons range from medical purposes, such as determining if the unknown could cause ailments in living things or knowing what microorganisms are needed to make antibiotics to other purposes such as knowing the exact microorganism has to be used to make certain foods. This experiment was done by applying methods in order to identify an unknown bacterium. 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 Red Broth test for lactose and sucrose fermentation. After performing a gram stain on unknown number five, number five was determined to be gram positive rod.
On the TSA plate, number five had the following morphology: very large, raised, opaque cream color colonies that covered the entire plate and had beta-hemolysis. All the results from the biochemical tests performed are listed in Table 1 and are also shown in the preliminary (Figure 1) and final (Figure 2) flow charts that are included at the end of the lab report. Table 1: Results from Biochemical Tests Test Results Interpretations Lab12-Medical Microbiology- Part1– Differential Media/Biochemical Tests, Sp2012 (Set all of your margins to ? ”)
Purpose: The purpose of this lab is to help you become a little familiar with some of the tests that can be typically performed in a clinical or research lab facility. These tests may help in determining a particular pathogen’s growth needs. There are several sections to this lab. Find each section and complete the “Preparing for Class” sections. Preparing for class – Day 1 Read in your lab manual about the following agar mediums: Blood Agar (pg 168), EMB Agar (pg 170), Mannitol Salt Agar (MSA)(pg 172) ), MacConkey Agar (pg 174), and PEA Agar (pg 176) to answer the following: 1.
What does the blood agar select for? Blood agar allows distinction among bacteria based on their ability to lyse red blood cells (hemolytic activity). 2. What color is the blood agar? Blood red color. 3. What are the 3 types of blood agar results and how can you recognize them? Beta hemolysis, which is the complete lysis of red blood cells and hemoglobin. This results in complete clearing of the blood around the colonies. Alpha hemolysis refers to the partial lysis of red blood cells and hemoglobin. This results in a greenish-grey discoloration of the blood around the colonies.
No hemolysis, sometimes called gamma hemolysis results in no change in the medium. 3. What color is the EMB agar? Dark blue colonies with green metallic sheen or pink. 4. What does the EMB agar select for? Gram-negative bacteria. 5. What bacteria can easily be differentiated on EMB agar? Gram-positive. How is it recognized? It contains the dyes eosin and methylene blue, which inhibit the growth of gram-positive bacteria. 6. What color is the Mannitol Salt Agar (MSA)? Yellow color change in surrounding media. 7. What does the MSA agar select for? It contains 7. 5% sodium chloride, which selects for organisms that are halotolerant.
