INTRODUCTION
The purpose of this investigation was to identify an unknown bacterium. “At any time there are millions of bacteria living around, on, or inside us” (The Plague). Bacterium can’t be identified by merely looking at it. Many bacteria have the similar appearances in growth. “In most cases, detection is based on the reaction of an enzyme with a certain substrate” (Sigma-Aldrich). Identification is usually based on the results of the bacterium’s cells metabolic capacities.
Some bacteria are harmless and are part of our normal flora. While other bacteria are capable of causing diseases or death. In order to determine what type of bacteria is present, a set of biochemical test must be performed. With biochemical testing, the ability to identify the bacterium responsible for causing illnesses or diseases. Once this is determined, other tests can be performed to discover what treatments would work best in ridding the illness causing bacteria.
During the investigation, seven test were used to determine the unknown bacterium as Salmonella typhimurium. “Salmonella typhimurium is a genus of gram-negative, non-lactose fermenting, medium-sized, rod-shaped, bacteria, members of the family Enterobacteriaceae, most species having flagella and pili” (medical-dictionary). This bacterium is known for causing food poisoning. This illnesses usually last 4-7 days. Treatments for the illnesses caused by this bacterium are dependent upon the severity of the symptoms, the age of the patient, and the comorbidities.
MATERIALS AND METHODS
During this investigation to identify the unknown bacterium, seven different biochemical tests were performed. All biochemical tests were performed according to the Eastfield Microbiology Lab Manual. As with...
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REFERENCES
Eastfield College Microbiology Lab. (2013). [Comparison Chart of Biochemical Test Results]. Unpublished raw data.
Eastfield College Microbiology Laboratory Manual, 1st edition, Oliver, T. D. (Book Must Be Purchased New from Eastfield Bookstore and Cannot Be Sold Back to Bookstore at the End of the Semester), Kendall Hunt Publishing, 2013, Dubuque, IA. ISBN 9781465223784.
Microbiology with Diseases by Body System (Hardcover) & MasteringMicrobiology, 3rd edition, Bauman, R. W., Benjamin- Cummings Publishing Company, 2012, San Francisco, CA. ISBN 9780321716378. OR
Cefrey, Holly. The Plague. New York: Rosen, 2001. Print. http://www.sigmaaldrich.com/analytical-chromatography/microbiology/identification-tests/reagents.html 2013
“Welcome to Microbugz.” Welcome to Microbugz. Linda Beaver and Mary Rutter, web. 2000
The isolate possesses some enzymes required for hydrolytic reactions. Hydrolytic enzymes found to be secreted from the bacterium, are -amylase, casein, and PYRase. In the starch hydrolysis and casein tests, there was a zone of clearing around the bacterium, which was indicative of the secreted enzymes necessary to break down starch and casein. In the PYR test, the presence of PYRase was detected by a color change to red on the PYR disc after the addition of the PYR reagent (p-dimethylaminocinnamaldehyde). Hydrolytic enzymes for which the EI tested negative were urease, gelatinase, and DNAse. In the Urea Hydrolysis test, it was observed that the urea broth did not have a color change, indicating that there was no urease secreted to break down urea in the broth. Similarly, there was no gelatinase present to break down gelatin in the Gelatin Hydrolysis test, so the nutrient gelatin remained solid. It was concluded that the EI does not possess DNase because there was no clearing zone around the bacteria, indicating that DNA had not been
After 48 hours of incubation the agar plates were viewed. Individual colonies were tested for successful isolation by gram staining and then viewing the stained bacteria under a microscope. Isolation was successful. One colony of each unknown bacteria was transferred to an agar slant for growth. The agar slants were stored at room temperature over the weekend so that they would not grow too much.
The purpose of this study is to identify an unknown bacterium from a mixed culture, by conducting different biochemical tests. Bacteria are an integral part of our ecosystem. They can be found anywhere and identifying them becomes crucial to understanding their characteristics and their effects on other living things, especially humans. Biochemical testing helps us identify the microorganism present with great accuracy. The tests used in this experiment are rudimentary but are fundamental starting points for tests used in medical labs and helps students attain a better understanding of how tests are conducted in a real lab setting. The first step in this process is to use gram-staining technique to narrow down the unknown bacteria into one of the two big domains; gram-negative and gram-positive. Once the gram type is identified, biochemical tests are conducted to narrow down the specific bacterial species. These biochemical tests are process of elimination that relies on the bacteria’s ability to breakdown certain kinds of food sources, their respiratory abilities and other biochemical conditions found in nature.
The purpose of this laboratory is to learn about cultural, morphological, and biochemical characteristics that are used in identifying bacterial isolates. Besides identifying the unknown culture, students also gain an understanding of the process of identification and the techniques and theory behind the process. Experiments such as gram stain, negative stain, endospore and other important tests in identifying unknown bacteria are performed. Various chemical tests were done and the results were carefully determined to identify the unknown bacteria. First session of lab started of by the selection of an unknown bacterium then inoculations of 2 tryptic soy gar (TSA) slants, 1 nutrient broth (TSB), 1 nutrient gelatin deep, 1 motility
Stout, M.A, et al. "Microbiology Lab Notebook". Lab handbook. University of Texas. Arlington. 2014. Print.
The eighteenth exercise of the laboratory manual titled Unknown Identification and Bergey’s Manual is an experiment to identify an unknown bacterium. In this exercise, a student must randomly choose a numbered bacterium available to the class. The keys in Appendix H, located on the last pages of the book, are the major helpful tools in this exercise because it provides completed steps of tests that needs to be performed in order to distinguish certain bacteria. This means that in this exercise, various types of tests and techniques must be performed to identify the chosen unknown bacterium. The unknown bacterium that I selected was number thirty-nine in which I discovered as the Bacillus megaterium after conducting several tests.
I was given unknown organism #14, in order to find out what organism I had, I had to perform several different biochemical tests to identify it. Starting with the Gram stain test, which is performed to differentiate Gram-positive and Gram-negative cells. After staining, when observed through the microscope Gram-positive cells are a purple color with thick peptidoglycan cell walls. Gram-negative cells are a pinkish/red color with thinner cell walls. (handout G. s.) My organism was observed to be pinkish rod shaped meaning it is Gram-negative bacteria.
The objective of this lab was to identify unknown bacteria culture by using various differential tests. There are many reasons for knowing the identity of microorganisms including to find the correct antibiotic to treat infections the bacteria may have caused. All the methods and techniques used to identify unknown bacterium #79 was learned in the microbiology laboratory.
The unknown bacterium that was handed out by the professor labeled “E19” was an irregular and raised shaped bacteria with a smooth texture and it had a white creamy color. The slant growth pattern was filiform and there was a turbid growth in the broth. After all the tests were complete and the results were compared the unknown bacterium was defined as Shigella sonnei. The results that narrowed it down the most were the gram stain, the lactose fermentation test, the citrate utilization test and the indole test. The results for each of the tests performed are listed in Table 1.1 below.
Identifying microorganisms can provide information on diagnosing diseases and discovering the most beneficial treatment possible. The purpose of this assignment was to identify an unknown microorganism using biochemical tests and various methods that were learned in my microbiology laboratory classes. In this paper, I will discuss the processes of how I came to identify my unknown microorganism.
Slack, John M. and I. S. Snyder. Bacteria and Human Disease. Chicago: Year Book Medical Publishers, Inc., 1978.
...nvironmental Microbiology. New York: A John Wiley & Sons, Inc; 1992. pp. 125?156. Accessed December 2, 2013.
To further determine the species of the unknown bacteria, an API 20E was used. API 20E system utilized a plastic strip with 20 separate compartments with each compartment consisting of cupule or a depression and a small tube containing a specific dehydrated medium (1). The ONPG tube consisted of an ingredient that functioned as an internal indicator. The ADH, LDC, ODC and URE tubes contain phenol red as the indicator. The CIT, GLU, MAN, INO, SOR, RHA, SAC, MEL, AMY and ARA tubes contain bromthymol blue as an indicator. The GEL tube contains charcoal and the H2S tube contains iron salts as indicators. The TDA, IND and VP tubes contain no indicator. All the tubes contain buffers and all the tubes with the exception of the CIT and URE contain
Leboffe, M. J., & Pierce, B. E. (2010). Microbiology: Laboratory Theory and Application, Third Edition 3rd Edition (3rd Ed.). Morton Publishing
This may sound like the beginning of a horror movie, but it isn’t. It is actually a very basic description of a very broad subject: microbiology.