3 tests to identify Escherichia coli

A positive result changes to a dark color and a negative test did not. This result shows that my bacteria do not ferment the sugars glucose, lactose and sucrose. Bacteria that can ferment these three sugars have the ability to produce hydrogen sulfide gas. The Gelatin test was to determine whether or not the organism has the enzyme gelatinase that hydrolyzes gelatin. A gelatin broth was obtained and was stabbed with the unknown bacteria.

Several lab experiments were performed to determine the species of an unknown culture. During staining the bacteria was determined to be gram negative after a differential gram stain was performed. The bacteria stained purple. When visualizing the bacteria, the bacteria were rod shaped and largely individuals instead of in groups. It was determined using Thioglycollate agar deep the unknown culture was inoculated using a stabbing technique, and it was determined that the bacteria is facultative.

Basic Microbiol. 41 57–64. 8. Haq, I., Ashraf, H., Qadeer, M.A., Iqbal, J. (2005) A source of alpha amylase production by Bacillus licheniformis, Bioresour.

The Catalase Test indicated a positive result. This showed that the bacterium contained the enzyme catalase which converts hydrogen peroxide into water and gaseous oxygen. The positive control used for this test was Micrococcus and the negative control for this test was Entercoccus. The Sulfur Reduction, Indole Production, and Motility (SIM) Test results indicated that the bacterium lacked the enzymes cysteine reductase or thiosulfate reductase and thus was unable to reduce sulfur. The positive control used for sulfur reduction was Proteus mirabilis and the negative control used was Staphylococcus epidermidis.

In this experiment, a series of biochemical test and API 20 E test are carried out to identify the unknown bacterial species provided. MacConkey agar, a selective and differential medium which is designed to isolate and differentiate the gram negative enteric bacteria. Bile salts and crystal violet inhibit the growth of gram positive organisms. Lactose provides a source of fermentable carbohydrate, allowing for differentiation of lactose fermenting bacteria from lactose non-fermenting gram negative bacteria. It uses for differentiate gram negative bacteria between phenotypes with mutations that confer varying abilities to ferment particular sugar.

Based on the different tests that were conducted in lab during the semester it was determined that the blood agar, MSA, and catalase test are used for gram positive bacteria while Macconkey, EMB, TSI, and citrate tests are used for gram negative bacteria. The results of the gram stain test were cocci and purple. This indicated that the unknown bacteria were gram positive. The gram stain test eliminated Escherichia coli, Klebsiella pneumonia, Salmonella enterica, and Yersinia enterocolitica as choices because these bacteria are gram negative. Next a Blood Agar plate was used because in order to do a MSA or a Catalase test there needs to be a colony of the bacteria.

The bacterium to be identified was sample B from wounds. Materials and Methods Prior to the performance of biochemical tests, a Gram stain was performed in order to label the unknown bacterium as either Gram positive or negative. Crystal violet and iodine are the first stains used, and cause the cell wall to possess a purple color. Ethanol is then used to clean off the purple dye. In Gram positive bacteria, the ethanol is unable to successfully wash out the purple color because of the thick cell walls that these bacteria possess.

Works Cited Angumeenal AR, Venkappayya D. 2013. An overview of citric acid production. LWT Food and Science Technol. 50:367-370. Berovic M, Legisa M. 2007.

High performance liquid chromatography-mass spectrometry (HPLC/MS). Organic Chemistry Unit-Bristol Research Centre  LAST, R.L., JONES, A.D. & SHACHAR-HILL, Y. 2007. Towards the plant metabolome and beyond. Nature Reviews Molecular Cell Biology.

The EMB plate was used to isolate the gram negative rod because the EMB plate is selective for gram negative rods, inhibiting the growth of gram positive organisms. This will effectively isolate the gram negative bacteria, thus creating a pure colony. Furthermore, the EMB plate differentiates for a gram negative rod that has the enzyme to break down lactose or sucrose, releasing an acidic waste product. If the organism appears to be dark purple to black, then it means that the microbe can ferment lactose and sucrose. The purple colonies on the EMB plate reveal that the gram negative bacteria is able to ferment lactose and sucrose.