Exercise 5-8 PDF

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Answers to Lab Exercise 5-8 (assigned by Dr. Amy Zimmerman)...

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Carolyn Straub Exercise 5-8 Questions 1. a. The base broth can be used with any of the specific amino acid substrates because what is being tested is the organisms’ response decarboxylase medium, not the base medium. Base broth contains the nutrients necessary for the bacteria to grow, as well as pH indicator (bromocresol purple). When an organism grows in broth, it either ferments the glucose (producing acid and changing the color of the pH indicator to yellow) or does not (in which case the color is unchanged). The response of the organism to the amino acids after this initial stage determines whether the result is ultimately negative (the color remains unchanged) or positive (the color becomes purple). Hence this principle of initial color determination based on glucose fermentation will apply for each decarboxylase medium and there is no need of different broth. b. The base broth is a negative control since the tube contains no amino acid substrates. c. No base broth was used in this experiment, but if it were, it would have been a negative control. It would have allowed the scientist to observe how the organisms grow when no amino acids are available. 2. a. The uninoculated tube is a negative control since there should be no organisms growing in it. b. The uninoculated control was a negative control. Negative controls are important because if any growth is seen in them, the scientist knows it must be due to contamination. Therefore, it cannot be determined whether any growth in the experimental tubes is due to contamination or not. It also gives the scientist an opportunity to observe a negative result for the unknown to be compared to. c. Based on the scientific method, this would not be acceptable. In order to compare across trials, the scientist must keep every variable the same and change only one at a time. If the medium was changed between the experimental and control trials, not only would the medium be different, but the inoculating organisms would be different. This is not acceptable. The mediums must be the same so that the only difference between the two trials is the inoculating organism. 3. It depends on how the organism initially reacts to the base broth. When an organism grows in broth, it either ferments the glucose (producing acid and changing the color of the pH indicator to yellow) or does not ferment glucose (in which case the color is unchanged). The response of the organism to the amino acids after this initial stage determines whether the result is ultimately negative (the color remains unchanged) or

Carolyn Straub Exercise 5-8 positive (the color becomes purple). If the color remains unchanged after the first step (remains original color or yellow), then the result is negative. If the color changes a second time (to purple), then the result is positive. 4. a. If a tube turns purple, you know the result is positive and an early reading can be taken because a purple color will not occur under any other circumstances than a positive result. b. An early reading is not allowable if the tubes still appear to be the original color or yellow because they may be decarboxylase-positive but not have had the time to turn purple yet. 5. a. An overlay of mineral oil is used to seal the organism from external oxygen sources to promote fermentation. Omitting this step would not affect the specificity of the test because the mineral oil is not what makes the test selective. b. Omitting this step would affect the selectivity of the test because the mineral oil helps the fermentation process to run optimally. Without this, the results would not be as pronounced or as clear....