User:Sarah Yacoub/Notebook/2/1/16

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This lab experiment was focused on microbiology and being able to identify and characterize bacteria. In order to be able to characterize bacteria, we had to identify the bacteria based on tests of motility, gram stain, colony morphology and sequencing of the 16s ribosomal subunit gene.

Hypothesis: If the Hay Infusion Culture is developing a more intense smell and more opaque water at the top of the jar, then this indicates that the soil and other solid components of the transect are slowly, but completely sinking to the bottom of the jar, making the water at the top appear clearer. The smell is also more intense because over time, the bacteria in the hay infusion culture begins to age, making the smell more intense.

Archaea species require a very harsh environment in order to be able to grow and develop and such environments do not exist in the agar plates we were using in this lab. Archaea tend to exist in extremely hot temperatures and in acid filled waters. Such environments could not have been replicated in the agar plates.

Methods and Materials: - A transect of an area on campus was collected - Transect was placed into a jar - Once it was inspected and observations were recorded, the transect was left in the jar for a week -After the week was over, drawings of the niches were created to document what state they were in after a week - Using a dichotomous key and a microscope, the kinds of bacteria present in the transect were determined -The size, color and shape of each bacteria observed was recorded - 8 agar plates were created, 4 with tetracycline spread across the surface and 4 without - A sample of the bacteria was then spread across the agar plates -The plates were left to incubate at room temperature for another week - Then different tests were done on the contents of the transect -The bacteria was left in agar plates for another week, 4 agar plates containing tetracycline and 4 plates without the tetracycline - First motility was tested - Then a gram stain test was performed - Lastly a PCR was created

Observations (before transect was tested): - The smell of the content of the transect is more intense and pungent - The water at the top of the jar is clearer and more opaque than it was the previous week.

Table 1: Dilution Agar Type # of Colonies on Plate 10^2 nutrient 1000+ 10^4 nutrient 150 10^6 nutrient 7 10^8 nutrient 1 10^2 nutrient + tet 86 10^4 nutrient + tet 2 10^6 nutrient + tet 0 10^8 nutrient + tet 0

As can be seen in Table 1, the presence and lack of presence of the antibiotic made a difference in the outcome of bacteria growth. The agar plates that contained bacteria overall had a tendency to have more bacterial growth than the plates with the antibiotic. In 2 cases, there were agar plates with antibiotic that did not grow any bacteria at all. This is an indicator that the bacteria cannot survive in the conditions that the antibiotic creates, as it is too harsh, and ends up killing the bacteria. The tetracycline in the antibiotic works to kill off the bacteria at early stages, before it is able to replicate. However, there are some species of bacteria that can survive in environments that contain tetracycline, as can be seen by the agar plates of 10^2 and 10^4 which grew 86 and 2 colonies respectively, despite the presence of the antibiotic. This is because there are kinds of bacteria that are resistant or insensitive to the antibiotic present.

One of the first kinds of tetracycline resistant bacterias is shigella dysenteriae. along with other bacterium in their phylum. It has also been discovered that 433 members of the enterobacteriaceae family are also resistant to tetracycline. The reason behind the resistance is the genetic makeup of the specific type of bacteria, which explains why there were some colonies able to thrive on two of the agar plates with tetracycline.

Table 2: Colony Label Plate Type Colony Description Cell Description "Gram + or Gram -" 10^4 -tet multiple, green clusters motility, circular + 10^6 -tet multiple, green clusters motility, circular + 10^2 +tet irregularly shaped, green no motility - 10^2 +tet irregularly shaped, green no motility -

Works Cited: Chopra, Ian, and Marilyn Roberts. "Tetracycline Antibiotics: Mode of Action, Applications, Molecular Biology, and Epidemiology of Bacterial Resistance." Microbiology and Molecular Biology Reviews. American Society for Microbiology, June 2001. Web. 01 Feb. 2