Gram Negative Lab Report

Topics: Escherichia coli, Gram-negative bacteria, Gram staining Pages: 12 (3004 words) Published: May 13, 2015
Gram Negative Unknown Lab Report

Abstract
The Unknown Gram Negative bacterium inoculated in a Tryptic Soy broth medium was randomly selected from a group of other unknowns. In order to identify this unknown the seven different types of biochemical tests will be conducted on this unknown bacterium to identify it out of 6 possible bacteria; Escherichia coli(E. coli), Enterobacter aerogenes (E.aerogenes), Klebsiella pneumoniae (K.pneumoniae), Proteus mirabilis(P.miranilis), Pseudomonas aeruginosa(P.aeruginosa), and Salmonella typhimurium (S.typhimurium). The biochemical tests utilized were; Triple Sugar Iron Agar (TSIA) test, Sulfur Indole Motility (SIM) test, Methyl red test and Voges-Proskauer (MR-VP) test, Citrate test, Urea test, and Gelatin test. After conducting each of these tests, the unknown bacteria number 24 was concluded to be Proteus mirabilis. Introduction

This probe purpose was to help identify an unknown gram negative bacterium, distributed by our TA instructor. It is relatively important to identify unknown organisms, those that are unidentified can certainly be harmful and have the potential to cause harm to the public. The use for these several test experiments are very important, mainly because it aids in the identification of unknown and potentially harmful organisms, Along with reflecting how well an organism can grow or react in certain environments.

The T-streak procedure for isolation of unknown was conducted before identifying the morphological properties. A T-streak is performed to isolate bacteria over three regions of a Tryptic Soy Agar plate (TSA), into single colonies that are able to be inoculated for biochemical tests. It is in the third quadrant of the TSA plate is where a pure culture can be found and used for the biochemical tests. As stated in the lab manual, that there are six possible unknown bacteria. The T- streak isolation method will make sure there is no contamination in the unknown.

The bacteria being utilized in this experimentation will be unknown gram negative bacterium number 22 that had been inoculated in a Tryptic Soy Broth. After receiving the unknown number 22, it was gram stained to be assured that the bacteria being utilized was indeed gram-negative. The purpose of the gram stain was to be used to identify the morphological characteristics of the unknown. The gram stain is a differential stain that utilizes a primary stain of crystal violet (i.e. a purple dye), iodine, alcohol decolorization, and safranin (i.e. a tinted red dye); used as counterstain. Standard results of a proper gram stain would show a bacterium to be gram positive if purple, and gram negative if pink. The results from the Gram Stain of unknown number 24 showed that unknown number 24 has bacillus shaped cell morphology, holding a dark-pink color confirming it to be a gram negative organism indeed. In both gram-positive and gram-negative the cell walls house peptidoglycan, which is a blend of carbohydrates and amino-acids. Gram negative bacteria cell walls have a thin peptidoglycan layer, in which an ample amount of lipids cross throughout the peptidoglycan and cell membrane. During the gram stain, lipids are prevalent in the gram negative bacteria are usually transparent as a result of the alcohol decolorizing agent. This allows the peptidoglycan of gram negative bacteria to retain the counterstain safranin (Leboffe & Pierce, 105)

Specific biochemical tests where implemented to follow, after the gram stain found unknown number 24 to be gram negative. These tests include a Triple Sugar Iron Agar (TSIA) test that uses a rich media that separates bacteria by the fermentation of Lactose, Sucrose, and Glucose, and have the capacity to reduce sulfur (Leboffe & Pierce 206). The objective of the test is to show if the unknown could exploit the carbohydrates in the medium to ferment into gas or reduce the sulfur. Any bacteria that possess the ability to ferment glucose and...

References: Todar, Kenneth. "Nutrition and growth of Bacteria." Online Textbook of Bacteriology. Web. 30 Mar. 2012. .
Leboffe, M. J. and B.E. Pierce. 2010. Microbiology Laboratory Theory and Application. 3rd ed. Morton Publishing Company
"E. Coli Enteritis." Medline Plus (n.d.): n. pag. 10 Jan. 2011. Web. 28 Mar. 2013.
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