DISCUSSION
The purpose of this study was to isolate, characterize, and identify an unknown species of bacteria collected from soil in Flagstaff, Arizona. The environmental isolate (EI) was found to be non-motile, this limits the bacteria from spreading across an area without outside forces. The EI had a positive reaction to the catalase test this indicates that the bacteria can convert harmful hydrogen peroxide into water and free oxygen (Shand and Fitchett 2017). It was also discovered that the EI was a strict aerobe which is significant because it cannot live without oxygen. This limits the area the bacteria can survive in. It was discovered that the EI was predominantly arranged in clusters and had the ability to produce a biofilm. This
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This was narrowed down to Table 17.1 because the EI is a strict aerobe. Table 17.1 was viewed and it contained only six genera. Within the table there were twelve characteristics the genera were tested for, but only four of those tests were done on the EI. Those four tests were the motility test, the catalase test production of acid from carbohydrates, and cell arrangement (Table 2). The catalase test was positive for all genera, but the motility test was only positive for Marinococcus and Planococcus (Table 2). This difference helped to make the decision that the EI was not one of these two genera. Deinococcus was ruled out based on its cells predominantly arranging in pairs or tetrads (Table 2). There were only two genera that seemed close to the cell arrangement of the EI. The genera were Micrococcus which predominantly shows clusters or tetrads and Salinicoccus which forms pairs, tetrads, and clumps (Table 2). The EI is mostly arranged in clusters or clumps (Table 2). These results left Salinicoccus and Micrococcus as the two genera that the majority of tests in Bergey’s Manual of Determinative Bacteriology match with. Miccrococcus was chosen from the remaining two genera because Salinicoccus prefers moderately saline environment making it much less likely to be found in Arizona mountain soil (Ventosa …show more content…
Micrococcus is nonmotile, catalase positive, and does not form endospores (Busse 2015). The EI shares all of the previous characteristics with Micrococcus (Table 2). The EI and Micrococcus form the same cell arrangements and Micrococcus can be found in soil, where the EI was taken from (Sims et al. 1986). Micrococcus can often produce acid from carbohydrates, but not all species produce acid from the same carbohydrates (Holt et al. 2000). The EI did not produce any acid with the four carbohydrates tested but this genus is still valid because the EI could be one of the species that does not produce acid for these carbohydrates. Micrococcus can also withstand harsh habitats even without an endospore, making it well suited for environments like Flagstaff Arizona where the EI was collected (Greenblatt et al.
Data from Table 1. confirms the theory that as the concentration of glucose increases so will the absorbance of the solution when examined with the glucose oxidase/horseradish peroxidase assay. Glucose within the context of this assay is determined by the amount of ferricyanide, determined by absornace, which is produced in a one to one ratio.1 Furthermore when examining the glucose standards, a linear calibration curve was able to be produced (shown as Figure 1). Noted the R2 value of the y = 1.808x - 0.0125 trend line is 0.9958, which is statistically considered linear. From this calibration curve the absorbance values of unknowns samples can be compared, and the correlated glucose concentration can then be approximated.
Data table 1 Well plate Contents Glucose concentration A 3 drops 5% sucrose + 3 drops distilled water Negative B 3 drops milk+3 drops distilled water Negative C 3 drops 5% sucrose +3 drops lactase Negative D 3 drops milk +3 drops lactase 15+ E 3 drops 20% glucose +3 drops distilled water 110 ++ Questions B. In this exercise, five reactions were performed. Of those reactions, two were negative controls and one was a positive control.
In the lab, Inhibiting the Action of Catechol Oxidase we had to investigate what type of enzyme inhibition occurs when an inhibitor is added. Catechol oxidase is an enzyme in plants that creates benzoquinone.Benzoquinone is a substance that is toxic to bacteria. It is brown and is the reason fruit turns brown. Now, there are two types of inhibitors, the competitive inhibitor and non-competitive inhibitor. For an enzyme reaction to occur a substrate has to bind or fit into the active site of the enzyme. In competitive inhibition there is a substrate and an inhibitor present, both compete to bind to the active site. If the competitive inhibitor binds to the active site it stops the reaction. A noncompetitive inhibitor binds to another region
Table 6 shows the results of the biochemical tests. The isolate can obtain its energy by means of aerobic respiration but not fermentation. In the Oxidation-Fermentation test, a yellow color change was produced only under both aerobic conditions, indicating that the EI can oxidize glucose to produce acidic products. In addition to glucose, the EI can also utilize lactose and sucrose, and this deduction is based on the fact that the color of the test medium broth changed to yellow in all three Phenol Red Broth tests. These results are further supported by the results of the Triple Sugar Iron Agar test. Although the EI does perform fermentation of these three carbohydrates, it appears that this bacterium cannot perform mixed acid fermentation nor 2,3-butanediol fermentation due to the lack of color change in Methyl Red and Vogues-Proskauer
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 affects of pH, temperature, and salt concentration on the enzyme lactase were all expected to have an effect on enzymatic activity, compared to an untreated 25oC control. The reactions incubated at 37oC were hypothesized to increase the enzymatic activity, because it is normal human body temperature. This hypothesis was supported by the results. The reaction incubated to 60oC was expected to decrease the enzymatic activity, because it is much higher than normal body temperature, however this hypothesis was not supported. When incubated to 0oC, the reaction rate was hypothesized to decrease, and according to the results the hypothesis was supported. Both in low and high pH, the reaction rate was hypothesized to decrease, which was also supported by the results. Lastly, the reaction rate was hypothesized to decrease in a higher salt concentration, which was also supported by the results.
Bacteria play a large role in our health, the environment, and most aspects of life. They can be used in beneficial ways, such as decomposing wastes, enhancing fertilizer for crops, and breaking down of substances that our bodies cannot. However, many bacteria can also be very harmful by causing disease. Understanding how to identify bacteria has numerous applications and is incredibly important for anyone planning to enter the medical field or begin a career in research. Having the background knowledge of identifying an unknown bacteria may one day aid healthcare professionals diagnose their patient with a particular bacterial infection or help researchers determine various clinical, agricultural, and numerous other uses for bacteria.
Each test that was used in the lab for the unknown bacteria had been performed on many different bacteria and shown that each test has different results depending on the bacteria given. The first test, the Gram stain, confirmed that the unknown bacterium was a gram negative bacilli. After performing the remainder of the tests and comparing them to the twelve negative bacteria that it could be out of it was basically a process of elimination. Basically looking at all the results and seeing which tests separated positive verses negative results the most. After reviewing all of the tests the first test that stuck out besides the gram stain was the lactose fermentation, followed by the citrate utilization test and then by the indole test. The lactose fermentation test eliminated seven of the 12 bacteria. From the five bacteria left the citrate utilization test eliminated who more of the bacteria, and last the indole test eliminated two of the three bacteria left leaving only one bacterium left. After comparing the results to the results of the 12 tests and separating which tests were positive and negative for each it was obvious that the bacteria had to be Shigella
I identified the genus and species of an unknown bacterial culture, #16, and I applied the following knowledge of morphologic, cultural and metabolic characteristics of the unknown microorganism according to the laboratory manual as well as my class notes and power point print outs. I was given an incubated agar slant labeled #16 and a rack of different tests to either examine or perform myself; the tests are as follows: Gram Stain; Nutrient Gelatin Test; Carbohydrate Fermentation; Dextrose, Lactose and Sucrose; IMVIC tests; Citrate, Indole, Mythel-Red and Vogues Proskauer test; as well as a Urease and TSI Test.
In this experiment the enzyme peroxidase and the substrate hydrogen peroxide were not mixed initially, instead they were both placed in separate tubes and were incubated at a specific temperature, to prevent hydrogen peroxide from undergoing any reaction with peroxidase until they both acquire the required temperature.
7. Madigan, Michael T., John M. Martinko, and Jack Parker. Brock Biology of Microorganisms, Ninth Edition. New Jersey: Prentice Hall, 2000.
The Effect of pH on the Activity of Catalase Planning Experimental Work Secondary Resources Catalase is a type of enzyme found in different types of foods such as potatoes, apples and livers. It speeds up the disintegration of hydrogen peroxide into water because of the molecule of hydrogen peroxide (H2O2) but it remains unchanged at the end of the reaction.
According to the graph on amylase activity at various enzyme concentration (graph 1), the increase of enzyme dilution results in a slower decrease of amylose percentage. Looking at the graph, the amylose percentage decreases at a fast rate with the undiluted enzyme. However, the enzyme dilution with a concentration of 1:3 decreased at a slow rate over time. Additionally, the higher the enzyme dilution, the higher the amylose percentage. For example, in the graph it can be seen that the enzyme dilution with a 1:9 concentration increased over time. However, there is a drastic increase after four minutes, but this is most likely a result of the error that was encountered during the experiment. The undiluted enzyme and the enzyme dilution had a low amylose percentage because there was high enzyme activity. Also, there was an increase in amylose percentage with the enzyme dilution with a 1: 9 concentrations because there was low enzyme activity.
Dr. Patricia Stock’s particular area of study does not include the biochemical investigation of their composition to find their chemical usefulness or the cure for cancer or anything of that nature. Her aim is simply to research and study the mutuality between the bacteria and their nematode hosts in order to better understand their evolutionary biology and pathogenesis.
Madigan, M., & Martinko, J. (2006). Brock biology of microorganisms. (11 ed.). New York, NY.