Podcast
Questions and Answers
What color is Escherichia coli under a gram stain? Why?
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?
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?
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?
What color is Bacillus megaterium under a gram stain? Why?
What color is Escherichia coli under an acid fast stain? Why?
What color is Escherichia coli under an acid fast stain? Why?
What color is Mycobacterium smegmatis in an acid fast stain? Why?
What color is Mycobacterium smegmatis in an acid fast stain? Why?
What color is Staphylococcus epidermidis in an acid fast stain? Why?
What color is Staphylococcus epidermidis in an acid fast stain? Why?
What color is Bacillus megaterium in an acid fast stain? Why?
What color is Bacillus megaterium in an acid fast stain? Why?
Which of the four samples from lab is the only acid fast positive bacteria?
Which of the four samples from lab is the only acid fast positive bacteria?
Which of the four samples from lab is the only gram negative bacteria?
Which of the four samples from lab is the only gram negative bacteria?
What is the primary stain in the gram stain?
What is the primary stain in the gram stain?
What is the mordant in the gram stain?
What is the mordant in the gram stain?
What is the decolorizer in the gram stain?
What is the decolorizer in the gram stain?
What is the secondary stain in the gram stain?
What is the secondary stain in the gram stain?
What is the primary stain in the acid fast stain?
What is the primary stain in the acid fast stain?
What is the decolorizer in the acid fast stain?
What is the decolorizer in the acid fast stain?
What is the counterstain in the acid fast stain?
What is the counterstain in the acid fast stain?
What is the primary stain in the spore stain?
What is the primary stain in the spore stain?
What is the counterstain in the spore stain?
What is the counterstain in the spore stain?
Is there a decolorizer in the spore stain?
Is there a decolorizer in the spore stain?
Which of the four samples from lab is positive for the spore stain?
Which of the four samples from lab is positive for the spore stain?
What are the results of Escherichia coli if you apply a gram stain, acid fast, and spore stain?
What are the results of Escherichia coli if you apply a gram stain, acid fast, and spore stain?
What are the results of Mycobacterium smegmatis if you apply a gram stain, acid fast, and spore stain?
What are the results of Mycobacterium smegmatis if you apply a gram stain, acid fast, and spore stain?
What are the results of Staphylococcus epidermidis if you apply a gram stain, acid fast, and spore stain?
What are the results of Staphylococcus epidermidis if you apply a gram stain, acid fast, and spore stain?
What are the results of Bacillus megaterium if you apply a gram stain, acid fast, and spore stain?
What are the results of Bacillus megaterium if you apply a gram stain, acid fast, and spore stain?
Name 4 gram positive bacteria.
Name 4 gram positive bacteria.
Name 4 gram negative bacteria.
Name 4 gram negative bacteria.
Name 4 acid fast positive bacteria.
Name 4 acid fast positive bacteria.
Name 4 acid fast negative bacteria.
Name 4 acid fast negative bacteria.
Name 4 spore positive (spore forming) bacteria.
Name 4 spore positive (spore forming) bacteria.
Name 4 spore negative (non-spore forming) bacteria.
Name 4 spore negative (non-spore forming) bacteria.
What is the purpose of a negative stain?
What is the purpose of a negative stain?
What is total magnification?
What is total magnification?
What is a petri sample for?
What is a petri sample for?
What is a slant for?
What is a slant for?
What is the dip sample for?
What is the dip sample for?
What is the broth for?
What is the broth for?
Why do you store the petri dish upside down?
Why do you store the petri dish upside down?
What temperature is a petri dish stored at? Why?
What temperature is a petri dish stored at? Why?
What is a basic dye?
What is a basic dye?
What is an acidic dye?
What is an acidic dye?
Why does basic dye attach to bacteria?
Why does basic dye attach to bacteria?
What are chromophores?
What are chromophores?
What is the purpose of a simple stain?
What is the purpose of a simple stain?
What is a simple stain?
What is a simple stain?
What does the condenser do?
What does the condenser do?
What does the diaphragm do?
What does the diaphragm do?
What does coccus mean?
What does coccus mean?
What does bacillus mean?
What does bacillus mean?
What does diplo mean?
What does diplo mean?
What does strepto mean?
What does strepto mean?
What does tetrads mean?
What does tetrads mean?
What does sarcinae mean?
What does sarcinae mean?
What does staphy mean?
What does staphy mean?
What does chemotaxis mean?
What does chemotaxis mean?
What does phototaxis mean?
What does phototaxis mean?
What happens to the gram positive bacteria cell when the decolorizer (alcohol acetone) is added?
What happens to the gram positive bacteria cell when the decolorizer (alcohol acetone) is added?
What happens to the gram negative bacteria cell when the decolorizer (alcohol acetone) is added?
What happens to the gram negative bacteria cell when the decolorizer (alcohol acetone) is added?
What are 5 considerations when using the oil immersion objective?
What are 5 considerations when using the oil immersion objective?
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.
