What is media (medium)
Used to grow any microbe. Exactly was in it, how much of component in it.
Differentiate between complex media and defined media
Complex media- that is, media for which the exact chemical composition varies slightly from batch to batch. Defined media- a medium whose exact chemical composition is known.
What is agar? How is it used?
Agar- an extract from marine red algae, has some unique properties that make it useful in culture media. It liquefies at 100 degrees Celsius and remains in a liquid state until cooled to 40 degrees Celsius. Its used to make liquid media solid
Differentiate between nutrient broth and nutrient agar.
Nutrient broth- is a commonly used liquid complex medium. Nutrient agar- when agar is added, is becomes a solid medium
What is a colony?
A population of cells that arises from a single bacterial cell.
What are petri dishes
Containing solid media provide a large surface area for examination of colonies
What does turbid mean? Is turbidity associated with liquid or broth cultures
Turbid- or cloudy, as a result of bacterial growth. Associated with broth cultures.
What is an autoclave? What time and temperature is used for sterilizing by autoclaving?
Autoclave- most common method of sterilizing culture media that are heat stable by using steam under pressure. 121 degrees Celsius for 15 minutes
Be able to name places where microbes are found.
Microbes are found everywhere: water we drink, the air we breathe, and the earth on which we walk.
Be able to recognize and describe what pellicle, flocculent and sediment means as it relates to a broth culture.
Pellicle- membrane across the surface of the broth, flocculent- clumps of microbial cell, sediment- microbial cells have settled on the bottom of the tube
Be able to look at a colony and describe it based on the chart on page 26
Whole colony appearance: circular, irregular, biconvex, filamentous, and rhizoid. Margin: entire, undulate, lobate, filamentous, and curled. Elevation: flat raised, convex, and umbonate.
Why are agar slants used? Why are petri dishes used? Why are agar deeps used
Agar slants- test tubes containing solid culture media that were left at an angle while the agar solidified. Provide a solid growth surface and are easier to store and transport. Petri dishes- containing solid media provide a large surface area for examin
What is aseptic technique? Why is it important to flame your loop or needle?
Aseptic technique- used in microbiology to exclude contaminants. To sterilize the loop before and after use to kill any unwanted bacteria.
When do you use an inoculating needle? When do you use an inoculating loop?
Needle- deep cultures. Loop- slants and broth cultures.
Why is simple staining used?
Staining procedures that use only one stain. Can be used to determine cell morphology, size, and arrangement.
What is a smear?
To stain bacteria, thin film of bacterial cells must be placed on a slide
How do you prepare a smear? Differentiate between preparing a smear from a broth culture and solid culture
Spreading a small amount of a bacterial broth culture on a clean slide and allowing it to dry. From solid medium: place one or two loopfuls of water on the slide, transfer a very small amount of the culture with a sterile loop and mix with the water on th
What is the purpose of fixing a smear?
To kill the bacteria or help cell stick to the slide for staining, also preserves microbes with minimal shrinkage or distortion when stained.
What are the two methods of fixing a smear? Which method did you use in class?
Heat fix and chemically fix are the two methods. We used heat fix in class.
What is a chromophore?
Ion that is colored.
What is a direct stain? What is a negative stain?
-Direct stain- simple stain that stains the bacteria. Negative stain- simple stain that stains the background but leaves the bacteria unstained
Basic stains produce positive stains, Why?
If the chromophore is a positive ion like the methylene blue in the equation shown.
Acidic stains produce negative stains, Why?
Negative ion.
Which dye was used for the simple staining procedure?
Methylene blue
Know how you prepared a simple stain in lab.
How did you know that a microbe grew in the broth culture?
Be able to identify shape and arrangement of a bacteria that has been simple stained under the microscope.
...
What is the purpose of the gram stain?
A differential stain that allows you to classify bacteria as either gram-positive or gram-negative
Know the steps of the gram stain and the reagents used and the effect each will have on a gram positive or gram negative cell
Gram-stain reagents: crystal violet for 30 sec(gram + purple and gram - purple), grams iodine 10 sec(gram+ purple and gram - purple), ethanol for 20 sec(gram + purple and gram -clear), safranin.for 30 sec( gram + purple and gram - pinkish red)
Know the gram staining procedure.
Wash off each reagents with water and once finished blot it dry.
Be able to identify a gram-positive or gram-negative microbe under the microscope.
Gram positive: staphylococcus epidermidis and bacillus subtilis. Gram negative: E.coli.
What is the purpose of the acid fast stain?
A differential stain.
What diseases can be diagnosed using the acid fast stain?
Mycobacterium tuberculosis and leprosy.
Know the steps of the acid fast stain and reagents used.
Reagents used: kinyouns carbolfuchsin for 5 minutes, acid alcohol for 1 minute, methylene blue for 1 minute
How do you determine if a cell is acid fast or non acid fast?
Acid fast- red, non acid-fast- purple or blue
Which organisms were used in the acid fast stain?
Mycobacterium non chromogencium(acid-fast) and bacillus subilitis or megaterium(non acid-fast)
What are endospores?
Are formed by several genera in the orders bacillales and clostridiales. Called resting bodies because they do not metabolize and are resistant to heating, various chemicals, and many harsh environmental conditions.
Why do some microbe produce endospores?
...
Study Figure 1, page 59. Know the different types of endospores based on location in the microbial cell
Free endospores after the cell has disintegrated, subterminal endospores, central swollen endospores, central endospores, terminal swollen endospores
Know the procedure for the endospore stain.
Absorbent paper over smear, cover paper with malachite green, steam the slide for 7 minutes, wash the smear with water, cover the smear with safranin for 30 seconds, and wash the smear with water and blot it dry
Differentiate between selective, differential and enrichment media.
Selective- contain chemicals that prevent the growth of unwanted bacteria without inhibiting the growth of the desired organism. Enrichment- usually liquid media contain chemicals that enhance the growth of desired bacteria. Differential- media contain va
Be able to look at experiments and be able to recognize selective versus differential media
...
Why are dyes used in media?
Products of bacterial metabolism can react with these dyes to produce a color change in the medium or colonies. Some dyes inhibit the growth of some bacteria.
What makes mannitol salt agar both differential and selective? What makes eosin methylene blue media selective and differential?
Mannitol salt agar- 7.5% Nacl makes it selective and manitol makes it differential. EMB- methylene blue makes it selective so inhibits gram positive from growing and eosin makes it differential.
In this experiment what was the purpose of the nutrient agar
Used to grow everything
Which organisms did you work with in this experiment?
E.coli, microccus luteus, pseudomonas aerguinosa, staphylococcus epidermidis,
What was the purpose of the EMB media? What was the purpose of the mannitol salt media?
Mannitol salth agar- used to grow gram positive and EMB- used to grow gram negative.
Define metabolism.
...
Define catabolism and anabolism
Catabolism- chemical reaction that release energy from the decomposition of complex organic molecules Anabollism
- Differentiate between exoenzymes and endoenzymes.
...
What is the purpose of the OF glucose test?
A nutrient semisolid agar deep containing a high concentration of carbohydrate and a low concentration of peptone.
What type of media is used for OF glucose test? How do you determine if the microbe has used glucose or peptone?
Indicated by a dark blue color due to ammonia production
What do the color changes in the OF glucose media indicate
OF medium contains the indicator bromthymol blue, which turns yellow in the presence of acids, indicating catabolism of the carbohydrate.
What are the components of OF glucose media?
Glucose-acid-yellow-ph
Why can OF glucose media be used to detect motility? What is the purpose of bromthymol blue? What was the color of the media before it was inoculated? How do you determine if the microbe is an oxidizer or fermenter? What was the purpose of adding the mine
...
What enzyme breaks down starch?
Amylase
Is amylase a type of exoenzyme or endoenzyme
Alpha (endoenzymes)-within cell, beta(exoenzeme)-outside cell; two types of amylase
Be able to identify a positive or negative starch hydrolysis test.
Clearing around the bacteria indicates that the starch was hydrolyzed
What is the purpose of the fermentation tube?
Is used to detect acid and gas production from carbohydrates
What are the components of the fermentation tube?
Peptone, acid base indicator (phenol red turns yellow), invert tube (trap gas), and sugar present (dactose, sucrose, and glucose)
What is the purpose of each component of the fermentation tube?
...
Which sugars were used in the fermentation test
Glucose, lactose, sucrose
How did you determine if acid was produced?
If the test was positive
How did you determine gas production?
If there was a bubble in the invert tube
What is media (medium)
Used to grow any microbe. Exactly was in it, how much of component in it.
Differentiate between complex media and defined media
Complex media- that is, media for which the exact chemical composition varies slightly from batch to batch. Defined media- a medium whose exact chemical composition is known.
What is agar? How is it used?
Agar- an extract from marine red algae, has some unique properties that make it useful in culture media. It liquefies at 100 degrees Celsius and remains in a liquid state until cooled to 40 degrees Celsius. Its used to make liquid media solid
Differentiate between nutrient broth and nutrient agar.
Nutrient broth- is a commonly used liquid complex medium. Nutrient agar- when agar is added, is becomes a solid medium
What is a colony?
A population of cells that arises from a single bacterial cell.
What are petri dishes
Containing solid media provide a large surface area for examination of colonies
What does turbid mean? Is turbidity associated with liquid or broth cultures
Turbid- or cloudy, as a result of bacterial growth. Associated with broth cultures.
What is an autoclave? What time and temperature is used for sterilizing by autoclaving?
Autoclave- most common method of sterilizing culture media that are heat stable by using steam under pressure. 121 degrees Celsius for 15 minutes
Be able to name places where microbes are found.
Microbes are found everywhere: water we drink, the air we breathe, and the earth on which we walk.
Be able to recognize and describe what pellicle, flocculent and sediment means as it relates to a broth culture.
Pellicle- membrane across the surface of the broth, flocculent- clumps of microbial cell, sediment- microbial cells have settled on the bottom of the tube
Be able to look at a colony and describe it based on the chart on page 26
Whole colony appearance: circular, irregular, biconvex, filamentous, and rhizoid. Margin: entire, undulate, lobate, filamentous, and curled. Elevation: flat raised, convex, and umbonate.
Why are agar slants used? Why are petri dishes used? Why are agar deeps used
Agar slants- test tubes containing solid culture media that were left at an angle while the agar solidified. Provide a solid growth surface and are easier to store and transport. Petri dishes- containing solid media provide a large surface area for examin
What is aseptic technique? Why is it important to flame your loop or needle?
Aseptic technique- used in microbiology to exclude contaminants. To sterilize the loop before and after use to kill any unwanted bacteria.
When do you use an inoculating needle? When do you use an inoculating loop?
Needle- deep cultures. Loop- slants and broth cultures.
Why is simple staining used?
Staining procedures that use only one stain. Can be used to determine cell morphology, size, and arrangement.
What is a smear?
To stain bacteria, thin film of bacterial cells must be placed on a slide
How do you prepare a smear? Differentiate between preparing a smear from a broth culture and solid culture
Spreading a small amount of a bacterial broth culture on a clean slide and allowing it to dry. From solid medium: place one or two loopfuls of water on the slide, transfer a very small amount of the culture with a sterile loop and mix with the water on th
What is the purpose of fixing a smear?
To kill the bacteria or help cell stick to the slide for staining, also preserves microbes with minimal shrinkage or distortion when stained.
What are the two methods of fixing a smear? Which method did you use in class?
Heat fix and chemically fix are the two methods. We used heat fix in class.
What is a chromophore?
Ion that is colored.
What is a direct stain? What is a negative stain?
-Direct stain- simple stain that stains the bacteria. Negative stain- simple stain that stains the background but leaves the bacteria unstained
Basic stains produce positive stains, Why?
If the chromophore is a positive ion like the methylene blue in the equation shown.
Acidic stains produce negative stains, Why?
Negative ion.
Which dye was used for the simple staining procedure?
Methylene blue
Know how you prepared a simple stain in lab.
How did you know that a microbe grew in the broth culture?
Be able to identify shape and arrangement of a bacteria that has been simple stained under the microscope.
...
What is the purpose of the gram stain?
A differential stain that allows you to classify bacteria as either gram-positive or gram-negative
Know the steps of the gram stain and the reagents used and the effect each will have on a gram positive or gram negative cell
Gram-stain reagents: crystal violet for 30 sec(gram + purple and gram - purple), grams iodine 10 sec(gram+ purple and gram - purple), ethanol for 20 sec(gram + purple and gram -clear), safranin.for 30 sec( gram + purple and gram - pinkish red)
Know the gram staining procedure.
Wash off each reagents with water and once finished blot it dry.
Be able to identify a gram-positive or gram-negative microbe under the microscope.
Gram positive: staphylococcus epidermidis and bacillus subtilis. Gram negative: E.coli.
What is the purpose of the acid fast stain?
A differential stain.
What diseases can be diagnosed using the acid fast stain?
Mycobacterium tuberculosis and leprosy.
Know the steps of the acid fast stain and reagents used.
Reagents used: kinyouns carbolfuchsin for 5 minutes, acid alcohol for 1 minute, methylene blue for 1 minute
How do you determine if a cell is acid fast or non acid fast?
Acid fast- red, non acid-fast- purple or blue
Which organisms were used in the acid fast stain?
Mycobacterium non chromogencium(acid-fast) and bacillus subilitis or megaterium(non acid-fast)
What are endospores?
Are formed by several genera in the orders bacillales and clostridiales. Called resting bodies because they do not metabolize and are resistant to heating, various chemicals, and many harsh environmental conditions.
Why do some microbe produce endospores?
...
Study Figure 1, page 59. Know the different types of endospores based on location in the microbial cell
Free endospores after the cell has disintegrated, subterminal endospores, central swollen endospores, central endospores, terminal swollen endospores
Know the procedure for the endospore stain.
Absorbent paper over smear, cover paper with malachite green, steam the slide for 7 minutes, wash the smear with water, cover the smear with safranin for 30 seconds, and wash the smear with water and blot it dry
Differentiate between selective, differential and enrichment media.
Selective- contain chemicals that prevent the growth of unwanted bacteria without inhibiting the growth of the desired organism. Enrichment- usually liquid media contain chemicals that enhance the growth of desired bacteria. Differential- media contain va
Be able to look at experiments and be able to recognize selective versus differential media
...
Why are dyes used in media?
Products of bacterial metabolism can react with these dyes to produce a color change in the medium or colonies. Some dyes inhibit the growth of some bacteria.
What makes mannitol salt agar both differential and selective? What makes eosin methylene blue media selective and differential?
Mannitol salt agar- 7.5% Nacl makes it selective and manitol makes it differential. EMB- methylene blue makes it selective so inhibits gram positive from growing and eosin makes it differential.
In this experiment what was the purpose of the nutrient agar
Used to grow everything
Which organisms did you work with in this experiment?
E.coli, microccus luteus, pseudomonas aerguinosa, staphylococcus epidermidis,
What was the purpose of the EMB media? What was the purpose of the mannitol salt media?
Mannitol salth agar- used to grow gram positive and EMB- used to grow gram negative.
Define metabolism.
...
Define catabolism and anabolism
Catabolism- chemical reaction that release energy from the decomposition of complex organic molecules Anabollism
- Differentiate between exoenzymes and endoenzymes.
...
What is the purpose of the OF glucose test?
A nutrient semisolid agar deep containing a high concentration of carbohydrate and a low concentration of peptone.
What type of media is used for OF glucose test? How do you determine if the microbe has used glucose or peptone?
Indicated by a dark blue color due to ammonia production
What do the color changes in the OF glucose media indicate
OF medium contains the indicator bromthymol blue, which turns yellow in the presence of acids, indicating catabolism of the carbohydrate.
What are the components of OF glucose media?
Glucose-acid-yellow-ph
Why can OF glucose media be used to detect motility? What is the purpose of bromthymol blue? What was the color of the media before it was inoculated? How do you determine if the microbe is an oxidizer or fermenter? What was the purpose of adding the mine
...
What enzyme breaks down starch?
Amylase
Is amylase a type of exoenzyme or endoenzyme
Alpha (endoenzymes)-within cell, beta(exoenzeme)-outside cell; two types of amylase
Be able to identify a positive or negative starch hydrolysis test.
Clearing around the bacteria indicates that the starch was hydrolyzed
What is the purpose of the fermentation tube?
Is used to detect acid and gas production from carbohydrates
What are the components of the fermentation tube?
Peptone, acid base indicator (phenol red turns yellow), invert tube (trap gas), and sugar present (dactose, sucrose, and glucose)
What is the purpose of each component of the fermentation tube?
...
Which sugars were used in the fermentation test
Glucose, lactose, sucrose
How did you determine if acid was produced?
If the test was positive
How did you determine gas production?
If there was a bubble in the invert tube