Bacillus Badius free essay sample

Dr. Robert Suran May 1 2013 Microbiology Abstract Over a three week time span I was able to successfully identify an unknown microorganism by combining the results of several tests. The first test was the Gram stain test which showed the microorganism was Gram positive and rod shaped. Using an Unknown Identification Flowchart, I proceeded to inoculate a Starch plate. In week 2, I analyzed the Starch plate and was able to determine that the results for Starch Hydrolysis were negative due to the dark color. Next, I inoculated Simmons Citrate Agar slants. By week 3 I was able to analyze the Citrate Agar, which tested negative for citrate utilization. With each successive test, the list of possible identities was being lessened until I finally came to the conclusion that only one logical microorganism remained; Bacillus badius. Introduction In this experiment I am attempting to identify an unknown microorganism. It is important to identify an unknown microorganism for many reasons, such as, if doctors need to treat an infection they need to first identify the bacteria that is causing the infection in order to know how to kill it. We will write a custom essay sample on Bacillus Badius or any similar topic specifically for you Do Not WasteYour Time HIRE WRITER Only 13.90 / page Unknown bacteria may also have clinical uses. For example, Penicillin, which is sometimes used to treat ear infections, is actually isolated from a fungus (Types of antibiotics). Microorganisms are classified based on many characteristics and different microorganisms may share several of these characteristics. Thus, making the possibility of identifying the microorganism, using a single test, none. Over a three week time span I was able to successfully identify the organism Bacillus badius by combining the results of several tests. For majority of the procedures I utilized the septic technique which consists of sanitary measures, such as, washing your hands. I used the Gram Stain test to determine the organisms’ morphology. I also, used the streak plate method for the Starch Hydrolysis Test. Materials and Methods The majority of the procedures for this experiment were performed as detailed for Lab 12, 13, and 14: Identification of an Unknown Microorganism as described in the s pring 2013 BIO 230 Laboratory Manual. Several procedures contributed to successfully determining the identification of the organism. With each successive result, the list of possible organisms lessened. This was done over a three week time span. The first procedure was the Gram stain. Gram staining is a simple test that has four steps: stain with crystal violet, treat with iodine, decolorize with ethanol, counter stain with safranin, and the results will either be positive (dark purple) or negative (pink). In order to do the procedure I heat fixed the unknown bacteria to a slide and positioned the slide in a staining tray. Next, I flooded the slide with crystal violet, letting it stand for 30 seconds and then rinsed it with water. Then, I flooded the smear with Gram’s iodine, letting that sit for 1 minute, before I rinsed it with water. Crystal violet is a basic dye that has a positive charge which attracts the negative charge of Gram’s iodine. The iodine is added and forms a complex with the crystal violet, thus improving the cells retention of the stain. Next, I flooded the slide with ethanol and I let it stand for 15-30 seconds, following with a water rinse. Ethanol dehydrates the peptidoglycan layer which shrinks it; making it impossible for the large crystal violet-iodine complex to penetrate through. Thus, making it trapped in the Gram positive bacteria. On the other hand, the thinner walls make it easier for the crystal violet-iodine complex to be washed out, resulting in the loss of color. Finally, the counter stain, safranin (red dye), is added to the sample by flooding it and letting it stand for 1 minute, staining it red. Finally, I rinsed both sides of the slide with water and blotted it dry with bibulous paper. Because the safranin is lighter than the crystal violet, it does not disturb the dark purple coloration in Gram positive cells. The decolorized Gram negative cells are stained pink. With the results from the Gram stain I was able to follow the â€Å"Unknown Identification Flowchart† to the next step, which was to prepare for the Starch Hydrolysis Test by inoculating a starch plate. Starch is a polysaccharide made up of glucose molecules. Some bacteria have an enzyme called amylase which breaks starch down into glucose subunits. The Starch Hydrolysis test is used to determine the production of amylase. Iodine, which is the mordant used in Gram staining, is used in this test to detect the presence of starch. In order to do the Starch Hydrolysis Test I first inoculated a starch plate by using aseptic technique. I streaked the middle of the starch plate with the bacteria into a single line. Then, I let it incubate overnight at 37 °C. In week 2, after I added several drops of Gram’s iodine to the starch plate it was ready to be analyzed for starch hydrolysis. The iodine complexes with starch to form a blue-black color in the culture. Clear halos surrounding colonies is the result of their ability to digest the starch due to the presence of amylase. With the results of the Starch Hydrolysis Test I preceded to the final test which is the Citrate Test. Using aseptic technique, I streaked the organism onto the surface of the Simmons’ citrate slant and let it incubate overnight at 37 °C. The Citrate Test determines if the organism is able to utilize citrate as its carbon and energy source. The medium contained citrate, ammonium salts, and bromothymol blue (pH indicator). Bromothymol blue is a deep blue color with a pH gt;7. 6. The production of alkaline waste products of citrate metabolism causes the medium to become royal blue which indicates a positive result. If the organism is unable to eat citrate, there will be no alkaline products and the agar will remain green. Results From the Gram stain I was able to determine that the organism was Gram positive and rod shaped. The Starch Hydrolysis test showed no color change to blue-black and no halo appeared. Therefore, the organism was negative for starch utilization. By observing the Simmons’ Citrate slant, which was a green color, I determined the organism tested negative for citrate utilization. With the results of these tests I was able to conclude that the only logical microorganism remaining was Bacillus badius. |Test Done |Description of |Results | | |results | | |Gram Stain |Bacilli rod |+ | |Starch Hydrolysis Dark color |- | | |No halo | | |Citrate |Green |- | | |(no color change) | | Discussion Bacillus badius is a Gram positive and rod shaped bacteria organism which is not able to hydrolyze starch, or utilize citrate as its carbon and energy source. The bacteria bacillus badius forms distinct colonies with rhizoid outgrowths. It has been isolated from dust, feces, marine sources, foods and antacids (Online Textbook of Bacteriology). Reference: Scientific Paper Examples of Student Papers. (n. d. ). George Mason University Classweb. Retrieved April 28, 2013, from http://classweb. gmu. edu/biologyresources/writingguide/Students. htm Types of antibiotics. (n. d. ). Retrieved  April  28, 2013, from http://www. biotopics. co. uk/g11/antibiotic_types. html Devenny, S (2013) Laboratory Manual for Microbiology, Delaware County Community College. Bacillus and related endospore-forming bacteria. Online Textbook of Bacteriology. Retrieved April 27, 2013, from http://textbookofbacteriology. net/Bacillus_6. html