Microbiology Lab Test Review
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Questions and Answers

What color is Escherichia coli under a gram stain? Why?

Red/pink because it is gram negative. It does not have a thick peptidoglycan cell wall therefore, does not retain the CV-1 stain.

What color is Mycobacterium smegmatis under a gram stain? Why?

Purple because it is gram positive. It does have a thick peptidoglycan cell wall and therefore retains the crystal violet stain.

What color is Staphylococcus epidermidis under a gram stain? Why?

Purple because it is gram positive. It does have a thick peptidoglycan cell wall and therefore retains the crystal violet stain.

What color is Bacillus megaterium under a gram stain? Why?

<p>Purple because it is gram positive. It does have a thick peptidoglycan cell wall and therefore retains the crystal violet stain.</p> Signup and view all the answers

What color is Escherichia coli under an acid fast stain? Why?

<p>Blue because it is acid fast negative. The carbolfuchsin washes out and you see the methylene blue.</p> Signup and view all the answers

What color is Mycobacterium smegmatis in an acid fast stain? Why?

<p>Red because it is acid fast positive. The carbolfuchsin is more soluble in the cell wall lipids therefore, retains the red color.</p> Signup and view all the answers

What color is Staphylococcus epidermidis in an acid fast stain? Why?

<p>Blue because it is acid fast negative. The carbolfuchsin washes out and you see the methylene blue.</p> Signup and view all the answers

What color is Bacillus megaterium in an acid fast stain? Why?

<p>Blue because it is acid fast negative. The carbolfuchsin washes out and you see the methylene blue.</p> Signup and view all the answers

Which of the four samples from lab is the only acid fast positive bacteria?

<p>Mycobacterium smegmatis.</p> Signup and view all the answers

Which of the four samples from lab is the only gram negative bacteria?

<p>Escherichia coli.</p> Signup and view all the answers

What is the primary stain in the gram stain?

<p>Crystal violet.</p> Signup and view all the answers

What is the mordant in the gram stain?

<p>Iodine.</p> Signup and view all the answers

What is the decolorizer in the gram stain?

<p>Alcohol-acetone.</p> Signup and view all the answers

What is the secondary stain in the gram stain?

<p>Safranin.</p> Signup and view all the answers

What is the primary stain in the acid fast stain?

<p>Carbolfuchsin.</p> Signup and view all the answers

What is the decolorizer in the acid fast stain?

<p>Alcohol-hydrochloric acid.</p> Signup and view all the answers

What is the counterstain in the acid fast stain?

<p>Methylene blue.</p> Signup and view all the answers

What is the primary stain in the spore stain?

<p>Malachite green.</p> Signup and view all the answers

What is the counterstain in the spore stain?

<p>Safranin.</p> Signup and view all the answers

Is there a decolorizer in the spore stain?

<p>False</p> Signup and view all the answers

Which of the four samples from lab is positive for the spore stain?

<p>Bacillus megaterium.</p> Signup and view all the answers

What are the results of Escherichia coli if you apply a gram stain, acid fast, and spore stain?

<p>Gram neg = red, acid fast neg = blue, spore neg = red.</p> Signup and view all the answers

What are the results of Mycobacterium smegmatis if you apply a gram stain, acid fast, and spore stain?

<p>Gram pos = purple, acid fast pos = red, spore neg = red.</p> Signup and view all the answers

What are the results of Staphylococcus epidermidis if you apply a gram stain, acid fast, and spore stain?

<p>Gram pos = purple, acid fast neg = blue, spore neg = red.</p> Signup and view all the answers

What are the results of Bacillus megaterium if you apply a gram stain, acid fast, and spore stain?

<p>Gram pos = purple, acid fast neg = blue, spore positive = red and green.</p> Signup and view all the answers

Name 4 gram positive bacteria.

<p>Mycobacterium smegmatis, Mycobacterium tuberculosis, Mycobacterium leprae, Mycobacterium bovis, Staphylococcus epidermidis, Staphylococcus aureus, Bacillus megaterium.</p> Signup and view all the answers

Name 4 gram negative bacteria.

<p>Escherichia coli, Vibrio cholera, Neisseria meningitis, Neisseria gonorrhoeae.</p> Signup and view all the answers

Name 4 acid fast positive bacteria.

<p>Mycobacterium smegmatis, Mycobacterium tuberculosis, Mycobacterium leprae.</p> Signup and view all the answers

Name 4 acid fast negative bacteria.

<p>Escherichia coli, Staphylococcus epidermidis, Bacillus megaterium.</p> Signup and view all the answers

Name 4 spore positive (spore forming) bacteria.

<p>Bacillus anthracis, Bacillus megaterium, Clostridium botulinum.</p> Signup and view all the answers

Name 4 spore negative (non-spore forming) bacteria.

<p>Escherichia coli, Mycobacterium smegmatis, Staphylococcus epidermidis.</p> Signup and view all the answers

What is the purpose of a negative stain?

<p>To see if the bacteria has a capsule.</p> Signup and view all the answers

What is total magnification?

<p>It is the power of magnification of the objective multiplied by the power of magnification of the ocular.</p> Signup and view all the answers

What is a petri sample for?

<p>Isolation.</p> Signup and view all the answers

What is a slant for?

<p>Preservation.</p> Signup and view all the answers

What is the dip sample for?

<p>To see if there is flagella.</p> Signup and view all the answers

What is the broth for?

<p>So the bacteria can grow without any restrictions.</p> Signup and view all the answers

Why do you store the petri dish upside down?

<p>To prevent condensation from forming on the lid.</p> Signup and view all the answers

What temperature is a petri dish stored at? Why?

<p>38°C, close to human body temperature which is the temperature that most bacteria grow the best in.</p> Signup and view all the answers

What is a basic dye?

<p>A dye with + ions, also called cations.</p> Signup and view all the answers

What is an acidic dye?

<p>A dye with negative ions, also known as anions.</p> Signup and view all the answers

Why does basic dye attach to bacteria?

<p>Because the basic dye has a positive charge and the bacteria have a negative charge.</p> Signup and view all the answers

What are chromophores?

<p>The colored ion pigment within stains.</p> Signup and view all the answers

What is the purpose of a simple stain?

<p>To improve the visibility of bacterial cells by increasing the contrast between the cell and the background.</p> Signup and view all the answers

What is a simple stain?

<p>A basic stain that is rinsed with water.</p> Signup and view all the answers

What does the condenser do?

<p>Focuses the light.</p> Signup and view all the answers

What does the diaphragm do?

<p>Controls the amount of light.</p> Signup and view all the answers

What does coccus mean?

<p>Round.</p> Signup and view all the answers

What does bacillus mean?

<p>Rod.</p> Signup and view all the answers

What does diplo mean?

<p>Two.</p> Signup and view all the answers

What does strepto mean?

<p>Chainlike.</p> Signup and view all the answers

What does tetrads mean?

<p>Four.</p> Signup and view all the answers

What does sarcinae mean?

<p>Eight.</p> Signup and view all the answers

What does staphy mean?

<p>Grapelike cluster.</p> Signup and view all the answers

What does chemotaxis mean?

<p>Toward or away from a chemical stimulus.</p> Signup and view all the answers

What does phototaxis mean?

<p>Toward or away from a light stimulus.</p> Signup and view all the answers

What happens to the gram positive bacteria cell when the decolorizer (alcohol acetone) is added?

<p>The peptidoglycan dehydrates and makes it more impermeable to the crystal violet-iodine. The CV-1 does not wash out, appears purple.</p> Signup and view all the answers

What happens to the gram negative bacteria cell when the decolorizer (alcohol acetone) is added?

<p>The alcohol dissolves the outer membrane and leaves small holes in the thin peptidoglycan layer. Makes it clear.</p> Signup and view all the answers

What are 5 considerations when using the oil immersion objective?

<ol> <li>Put the oil on between the 40x and the 100x. 2. One drop of oil. 3. Clean it before and after. 4. The 100x objective must be touching the oil. 5. Focus using the fine focus adjustment knob.</li> </ol> Signup and view all the answers

Study Notes

Gram Staining

  • Escherichia coli: Red/pink (gram-negative); does not retain crystal violet due to a thin peptidoglycan wall.
  • Mycobacterium smegmatis: Purple (gram-positive); retains crystal violet due to a thick peptidoglycan wall.
  • Staphylococcus epidermidis: Purple (gram-positive); thick peptidoglycan retains crystal violet.
  • Bacillus megaterium: Purple (gram-positive); retains crystal violet due to a thick peptidoglycan wall.

Acid-Fast Staining

  • Escherichia coli: Blue (acid-fast negative); carbolfuchsin washes out, leaving methylene blue visible.
  • Mycobacterium smegmatis: Red (acid-fast positive); retains red from carbolfuchsin due to lipid-enriched cell wall.
  • Staphylococcus epidermidis: Blue (acid-fast negative); similar to E. coli.
  • Bacillus megaterium: Blue (acid-fast negative); similar to E. coli.

Unique Bacterial Characteristics

  • Only acid-fast positive bacteria in lab: Mycobacterium smegmatis.
  • Only gram-negative bacteria in lab: Escherichia coli.

Staining Components

  • Primary stain in gram stain: Crystal violet.
  • Mordant in gram stain: Iodine.
  • Decolorizer in gram stain: Alcohol-acetone.
  • Secondary stain in gram stain: Safranin.
  • Primary stain in acid-fast stain: Carbolfuchsin.
  • Decolorizer in acid-fast stain: Alcohol-hydrochloric acid.
  • Counterstain in acid-fast stain: Methylene blue.
  • Primary stain in spore stain: Malachite green.
  • Counterstain in spore stain: Safranin.
  • Decolorizer in spore stain: None; only a water rinse.

Staining Results for Specific Bacteria

  • Escherichia coli: Gram negative (red), acid-fast negative (blue), spore negative (red).
  • Mycobacterium smegmatis: Gram positive (purple), acid-fast positive (red), spore negative (red).
  • Staphylococcus epidermidis: Gram positive (purple), acid-fast negative (blue), spore negative (red).
  • Bacillus megaterium: Gram positive (purple), acid-fast negative (blue), spore positive (red and green).

Bacterial Classification

  • Gram Positive Bacteria: Mycobacterium smegmatis, Staphylococcus epidermidis, Bacillus megaterium, Streptococcus species.
  • Gram Negative Bacteria: Escherichia coli, Neisseria species, Vibrio cholera.
  • Acid-Fast Positive Bacteria: Mycobacterium species, Nocardia asteroides.
  • Acid-Fast Negative Bacteria: Escherichia coli, Staphylococcus species, Bacillus megaterium.
  • Spore-Forming Bacteria: Bacillus anthracis, Clostridium species.
  • Non-Spore Forming Bacteria: Escherichia coli, Mycobacterium smegmatis.

Staining Techniques and Purposes

  • Negative stain: Used to see if bacteria have a capsule.
  • Petri dish: Isolation of bacteria.
  • Slant: Preservation of cultures.
  • Dip sample: Examination for flagella presence.
  • Broth culture: Unrestricted growth of bacteria.

Storage and Maintenance

  • Petri dishes stored upside down to prevent condensation.
  • Incubation temperature for petri dishes: 38°C, optimal for most bacteria.

Staining Characteristics

  • Basic dye: Positive ions (cations), attract negatively charged bacteria.
  • Acidic dye: Negative ions (anions).
  • Chromophores: Colored ions in stains.

Microscope Adjustments

  • Condenser: Focuses light.
  • Diaphragm: Controls light amount.

Bacterial Morphology Terms

  • Coccus: Round shape.
  • Bacillus: Rod shape.
  • Diplo: Pairs.
  • Strepto: Chains.
  • Tetrads: Groups of four.
  • Sarcinae: Groups of eight.
  • Staphy: Grapelike clusters.

Bacterial Movement Responses

  • Chemotaxis: Movement toward or away from chemicals.
  • Phototaxis: Movement toward or away from light.

Gram Staining Mechanism

  • Gram-positive bacteria retain crystal violet after decolorization due to dehydrated peptidoglycan.
  • Gram-negative bacteria lose crystal violet due to dissolved outer membrane and porous peptidoglycan layer.

Oil Immersion Microscope Usage

  • Apply oil between 40x and 100x magnification.
  • Use one drop of oil; clean before and after use.
  • Ensure 100x objective touches oil.
  • Use fine focus adjustments for clarity.
  • Oil maximizes numerical aperture and resolution; avoid using cloudy oil.

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Description

Test your knowledge of microbiology concepts with this lab review focused on Gram staining. Learn the differences between Gram-positive and Gram-negative bacteria, including Escherichia coli and Mycobacterium smegmatis. Use these flashcards to prepare for your microbiology exams effectively.

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