Module 2 Case Study - Up Against a Cell Wall

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Utah Valley University *

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2065

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Biology

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Dec 6, 2023

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docx

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Case Study: Up Against a [Cell] Wall As the most senior member in the diagnostic microbiology lab at American Fork Hospital, you know a thing or two about microorganisms. Many of the other members of the lab come to you with questions and ask for help with laboratory techniques. On a typical day, you process dozens of patient samples and read the results as if it were second nature. One sample is a sputum sample from a young woman who has had a chronic cough since she returned from visiting her grandparents in Peru. You have been instructed to perform the Gram stain on this sample and, if necessary, an Acid-Fast Stain. The Gram stain is a differential stain that allows microbiologist to determine the shape of a bacterium, its morphological arrangement, and differentiate between the two major groups: Gram-positive and Gram- negative. Except for bacteria that lack a cell wall and those that have a waxy coat attached to the outer surface of their cell wall, virtually all bacterial species will stain either Gram+ or Gram- based on the structure of their cell wall. The cell wall is a rigid structure composed of two repeating sugar units (glycans) called N-acetyl muramic acid (NAM) and N-acetylglucosamine (NAG) that are represented as blue rectangles in the figure below. These repeating chains are attached to each other by short chains of amino acids (peptides). For this reason, the cell wall is often referred to as the peptidoglycan layer (peptides and sugars). Gram+ cells have a thick cell wall just outside the cytoplasmic membrane. Gram-negative cells have a thin cell wall generally only 1-2 layers thick sandwiched between their cytoplasmic membrane and an outer membrane. The outer membrane is an asymmetric bilayer unique to Gram- organisms. The cell wall contributes to the shape or morphology of the cell and prevents the cell from rupturing in a hypotonic environment. Bacteria with a thicker peptidoglycan layer (Gram+) will retain a dye used in the
Gram stain called crystal violet. Under the microscope, Gram+ bacteria will have a dark purple appearance (A). In contrast, bacteria with a thin layer of peptidoglycan (Gram-) will not retain the crystal violet dye, so when another dye called safranin is used, the bacteria will have a lighter purple/pink appearance (B). After Gram-staining the sputum sample, you view the bacteria under a microscope to see if it is G- positive or G-negative and you see something surprising: the cells on the slide under the microscope are rod-shaped microbes but there is no stain! You determine that the acid-fast stain is necessary to properly identify this bacterium.
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