virtual lab week 3

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North Carolina Central University *

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242

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Biology

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Apr 3, 2024

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BIOS242, Week 3-OL Name: Lab 3: The Gram Stain: Identify and differentiate bacteria Learning Outcomes  Describe the structure of the Gram-positive and Gram-negative bacteria.  Identify theoretical and technical aspects of the Gram staining procedure.  Identify the most commonly made mistakes in Gram staining.  Analyze the results of a Gram staining experiment using a light microscope. About the Gram Stain: Identify and differentiate bacteria Virtual Simulation Lab Did you know that there are approximately 5 million-trillion-trillion bacteria in the world? Most of them are harmless, but some can induce disease in an affected host. In this simulation, you will help doctors identify bacteria in a cerebrospinal fluid sample from a patient suspected of suffering from bacterial meningitis. Explore the bacterial cell wall Compare and contrast the cell wall of Gram-positive and Gram-negative bacteria by building your very own bacterial 3D models on the hologram table. Enter the exploration pod to observe in an immersive animation how the four reagents of the Gram stain interact with structural components of the cell wall to color the bacteria. Perform the gram stain When the patient’s fluid sample arrives at the laboratory, equip yourself with protective gear to prepare a bacterial smear and heat fix it to a glass slide. You are now ready to perform the Gram stain in a safe virtual environment. Made a mistake? No worries, hit the big red button on the workbench to repeat the staining procedure until it becomes second nature. Interpret your findings using a microscope In the end, you will use a light microscope to interpret the results of your Gram stain. View the microscopic image on the computer screen, and apply immersion oil to increase magnification 1000x! Will you be able to identify the presence of any bacteria in the patient´s cerebrospinal fluid? Questions: 1. Purpose: Please describe in complete sentences and in your own words, the purpose of this experiment. The purpose of this experiment was to perform a Gram stain using the provided sample of cerebrospinal fluid to detect the presence of bacteria. The aim was to determine whether the patient might be suffering from bacterial meningitis based on the results of the Gram stain. 2. Complete the following table by predicting colors of bacteria with- and without cell wall as they are processed through the steps of Gram staining. 1

BIOS242, Week 3-OL Name: Steps of Gram Staining Bacteria containing thick cell wall Bacteria containing thin cell wall (LPS) Crystal violet treatment Purple Purple Iodine Purple Purple Decolorization Purple Pink Safranin Purple Pink 3. A fellow student showed you a gram stained slide where cells containing thick cell walls were stained pink. What would you tell her about the staining procedure? Why? If cells with thick cells walls stained pink in the Gram stain, it indicates they are likely Gram- negative bacteria. This occurs because Gram-negative bacteria have thinner peptidoglycan layers, allowing them to retain the pink safranin counterstain after decolorization, unlike Gram-positive bacteria. 4. A fellow student showed you a gram stained slide where cells containing LPS were stained purple. What would you tell her about the staining procedure? Why? If cells with LPS stained purple in the Gram stain, it suggests incomplete decolorization. Gram-negative bacteria usually lose the crystal violet-iodine color and appear pink, but if they remain purple, it indicates decolorization was not effective, likely due to the outer membrane’s resistance. Adjustments in the staining procedure may be needed for accurate results. 5. Reflection: Write 5 sentences on what you learned from this simulation. What did you like and what was something that you would prefer not be a part of this simulation? From the simulation, I learned about the differences between Gram-positive and Gram-negative bacteria and how they affect staining. Understanding the basics and practical aspects of staining was important. Learning about common mistakes showed the need for careful technique. Looking at stained samples under a microscope helped me see the differences between the bacteria. I liked the hands-on aspect of the simulation, but I was wished there was more help for fixing mistakes. 2

BIOS242, Week 3-OL Name: Grading Rubric: Activity Deliverable Points Lab Report and Questions Complete lab report and answer questions  Purpose (1 point)  Questions (9 points)  Reflection (5 points) 15 All Lab Deliverables Complete ALL lab work and lab report 15 3

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