Microbiology Task 1

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Jan 9, 2024

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Rabeca Plummer Clinical Microbiology Laboratory C453 2/25/2021 Task 1

Introduction In Task 1, we delve into the difference between gram-positive and gram-negative microorganisms. Gram- positive microorganisms react to the staining method invented by Hans Christian Gram and can retain the violet color of the stain in the process. Gram-negative microorganisms are defined as the opposite, they do not retain the violet color and instead take on the red counterstain. Gram-positive cells have as many as 40 layers of peptidoglycan in their cell wall. They have a thicker cell wall (20-80nm across) containing 60-90% peptidoglycan. The cell walls contain very little protein (with the notable exception of streptococci). Yeast cells contain no peptidoglycan but have thick walls and are considered gram-positive, thus a connection to thick walls and gram-positivity has been concluded. Another notable connection to wall thickness is that in aging, cells can become gram-negative or gram- variable as their wall retains damage and can become leaky. Gram-negative cells have thinner and more complex walls than their counterparts. Only 10-20% of the walls are made of peptidoglycan. The wall also covered in an outer membrane which leaves a very narrow periplasmic space. Gram-negative bacteria cannot hold the violet-iodine dye because of both their thin walls and the large quantities of lipoproteins and lipopolysaccharides in their walls. Materials  Glass slides  Pipettes  Sample  Flame  Crystal Violet Dye  Gram’s Iodine  Water  Safranin  Paper towels  Sterile gloves  PPE in compliance with lab standards Method 1. Place sample on the sterile slide using aseptic method 2. Dry the sample with application of heat through open flame on the bottom of the slide 3. Flood the sample on the slide with crystal violet for 1 minute 4. Rinse sample with water 5. Flood the sample on the slide with Gram’s Iodine for 1 minute 6. Rinse sample with water 7. Decolorize sample briefly with alcohol 8. Rinse sample with water 9. Counterstain sample with Safranin for 30-60 seconds 10. Rinse sample with water 11. Blot sample dry 12. Document findings

This method (Unknown, Wiley Engage ) is the basic gram staining method used for most common circumstances. There are four main important steps to the process: Crystal violet, Gram’s iodine, decolorize, and counterstaining. Flooding the sample with crystal violet dyes the bacteria purple which makes it more visible. Adding Gram’s iodine to the sample reacts with the crystal violet and bons to the cell wall, creating a distinguishing characteristic to the different cells. Decolorizing allows the Gram- negative cells to be cleanse of any lingering dye that may compromise the results. Counterstaining the sample with Safranin allows the gram-negative bacteria to take on a red color and differentiates them from the gram-positive violet cells. Results Slide one contains a sample of Staphylococcus Epidermidis (unknown, staphylococcus_epidermidis ). The samples shows spherical cocci cells dyed violet and clumping together. Results indicate that the sample is Gram-positive. Slide two contains a sample of Escherichia Coli (unknown, escherichia_coli ). The sample shows bacilli rods that have been dyed a light red/pink color. The cells are very separate and not clumping together. Results indicate the sample is Gram-negative. Discussion Gram negative bacteria contains an outer membrane that prevents it from absorbing the violet color of the Gram stain. It also prevents certain antibiotics from being able to infiltrate their cell wall and destroy them. The proper identification of Gram-negativity leads to the appropriate antibiotic prescription. Resources Unknown. (n.d.). Wiley Engage: Log in to the site. Retrieved February 25, 2021, from https://wiley.engagelms.com/learn/mod/book/view.php?id=25818 Unknown. (n.d.). Staphylococcus_epidermidis. Retrieved February 25, 2021, from https://web5.wgu.edu/aap/content/staphylococcus_epidermidis.jpg Unknown. (n.d.). Escherichia_coli. Retrieved February 25, 2021, from https://web5.wgu.edu/aap/content/escherichia_coli.jpg

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