Unknown Identification Lab Report: Bacterial Culture

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UNKNOWN IDENTIFICATION LABORATORY REPORT
Introduction
The Gram Stain Test: This test utilizes differences in bacterial cell wall types to distinguish between two groups of bacteria namely: Gram positive and Gram negative bacteria. The Gram positive bacteria have their cell wall made up of a thick layer of 90 percent peptidoglycan chains and teichoic acid cross-linkages. On the other hand, the Gram negative bacteria have a thin layer of cell wall, mostly comprising 10 percent peptidoglycan chains and high lipid content. This forms the basis for the color differences observed in Gram Stain test. In the Gram positive bacteria, a blue to purple color is retained for a positive test, while the Gram negative bacteria will give a red to pink color. The Gram Stain test can also be used to detect bacterial morphology. For example, once a culture is stained and observed under a microscope with the help of the oil immersion lens, different sizes and shapes of bacteria can be seen. These sizes range from 0.5 - 10 microns. Bacterial shapes include: spherical (or coccus); helical (spirilum); curved (or comma-shaped) and rod-shaped (or bacillus).
The Sugar Fermentation test: This test is used to distinguish different bacteria based on their abilities to ferment certain sugars and use them for their growth and respiration. Different bacteria require different sugars as substrates. The main sugars utilized by different bacteria include: glucose, lactose, and sucrose. The sugar under test is usually incorporated into a phenol red broth. Bacteria that ferment the specific sugar will cause the broth to change from red to yellow colour due to a fall in pH. For those bacteria that do not ferment the sugar in question, the broth remains red.
The Catalase test: Microorganisms that live in well-oxygenated niches produce the enzyme catalase in response to toxic oxygen radicals and metabolites, especially hydrogen peroxide. Therefore, the test can be used to distinguish between aerobic and anaerobic bacteria. Still, the test can be used to differentiate the anaerobic bacteria, especially aerotolerant Clostridium strains (Catalase-negative) from Bacillus species which are Catalase-positive. Catalase-positive bacteria will give effervescence (or bubble formation) immediately hydrogen peroxide is added to the bacterial culture. Catalase-negative bacteria do not give any effervescence.
The Urease test: This test is based on urea, the main by-product of the amino acid decarboxylation process. The test determines the ability of bacteria to hydrolyze urea into ammonia and carbon dioxide gas. For the Urease-positive bacteria, it is an indication that they possess the enzyme urease responsible for urea hydrolysis. Usually, Urease-positive bacteria will cause a color change in phenol red broth from light orange to pink because of the alkaline nature of the ammonia formed. On the other hand, Urease-negative bacteria do not give any color change and the phenol red broth remains light orange.
Methyl Red (MR) test: This is a test used to determine whether bacteria undergo mixed acids fermentation, especially when glucose is supplied to a bacterial culture. Different types and proportions of fermentation products will be produced by anaerobic glucose fermentation. The different types as well as proportions of the products are the basis of identifying various genera of the Enterobacteriaceae family. After the addition of methyl red into a test sample, Methyl red-positive bacteria will turn the medium red. The change in color means that the bacteria are capable of producing a lot of acids (lactic, acetic, and succinic acids) on fermentation of glucose. On the contrary, Methyl red-negative bacteria will give no color change in the medium, and the medium retains its yellow color. This shows that less acids are produced (this accounts for the higher pH).
The Voges-Proskauer test: This test is used to distinguish between acetoin-producing and non-acetoin producing bacteria based on utilization of pyruvate, an important substrate in glucose fermentation. Certain groups of bacteria utilize a unique pathway in pyruvate metabolism resulting in the production of acetyl methyl carbinol (or acetoin). Acetoin is known to be a neutral end product in these groups of bacteria. Positive tests are confirmed by a color change when 5% alpha naphthol and 40% potassium hydroxide are added to Methyl red/Voges-Proskauer broth. A red color develops after 15 minutes of incubation. The color change is due to the formation of diacetyl, the oxidation end product of acetoin. However, this test should not be read after one hour because negative Voges-Proskauer bacterial cultures ...