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Microbiology > Lab 6: Bacterial Enumeration > Flashcards

Lab 6: Bacterial Enumeration Flashcards

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Biology 101 flashcards
1
Q

What is the primary stain of gram stains?

A

Crystal violet

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2
Q

What is the primary stain step of acid fast stains?

A

Carbol fuchsin flood smear (paper towel)

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3
Q

What is the primary stain step of endospore staining?

A

Malachite green flood smear (paper towel)

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4
Q

What is the mordant step of a gram stain?

A

Iodine

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5
Q

What is the mordant step of an acid fast stain?

A

Heat carbol fuchsin to 90-95 degrees C (steaming) for 3.5 minutes, then rinse with water

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6
Q

What is the mordant step of an endospore stain?

A

Heat malachite green to 90-95 degrees C (steaming), rinse with water

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7
Q

What is the decolorization step of a gram stain?

A

95% alcohol

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8
Q

What is the decolorization step of an acid fast stain?

A

Add alcohol dropwise for 20-30 seconds, rinse with water

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9
Q

What is the decolorization step of an endospore stain?

A

There isn’t one

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10
Q

What is the counterstain step of a gram stain?

A

Safranin

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11
Q

What is the counterstain step of an acid fast stain?

A

Methylene blue for 1 minute

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12
Q

What is the counterstain step of an endospore stain?

A

Safranin for 1 minute, rinse with water

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13
Q

What are the steps of an endospore?

A

1) Primary stain: Malachite green flood smear (paper towel)
2) Mordant: Heat at 90-95C (steaming), keeping paper towel wet, for 3.5 minutes. Rinse with water.
3) Counterstain: Safranin for 1 minute, rinse with water

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14
Q

What are the steps to an acid fast stain?

A

1) Primary stain: Carbol fuschin (paper towel) flood smear
2) Mordant: Heat to 90-95C for 3.5 minutes, rinse with water
3) Decolorization: Add alcohol dropwise 20-30 seconds, rinse with water
4) Counterstain: Methylene blue for 1 minute

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15
Q

What are the acid fast colors?

A

Acid fast positive = purple/ red
Acid fast negative = blue

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16
Q

How do you make bacteria make spores?

A

Leave them in an incubator for a week or more

17
Q

Would you ever acid-fast stain a species that gram-stains well?

A

No

18
Q

How do you calculate stepwise dilution factor? Give an example

A

1 divided by the volume of dilution, then convert to a decimal.
-Ex: 1: 1ml + 99ml equals 1:100, then 1/100 = 0.01, which = 10^-2

19
Q

1) How do you calculate total dilution factor? Give an example
2) What is the typical initial total dilution factor of the culture?

A

1) Stepwise dilution factor + previous total dilution factor
-Ex: 10^-2 x 10^-4 = 10^-6 (adding not multiplying)
2) Typically 10^0 for the culture

20
Q

In the 1890s, we learned the causative agent of what two things? What was the causative agent? Why wasn’t this very beneficial at first?

A

1) Typhoid fever and cholera
2) Causative agent was mammalian feces
3) Monitoring those agents was time consuming

21
Q

What technique was created to monitor for potentially contaminated water?

A

Monitoring fecal contamination with an indicator species called coliforms

22
Q

What do coliforms do?

A

1) Indicate fecal contamination
2) Lactose fermentation (usually within 24 hours)

23
Q

Define lactose fermentation

A

The production of acid + gas

24
Q

Historically, the issue of enumerating bacteria arose due to what?

A

Public health concerns for clean, uncontaminated water, which led to attempts to monitor water quality

25
Q

How is water quality counted?

A

Using MPN tests with phenol red lactose fermentation

26
Q

What are other ways to count bacteria? (3)

A

1) Serial dilutions
2) Viable plate counts
3) Hemocytometer (graph thing)

27
Q

When does phenol red turn yellow?

A

At pH greater than or equal to 6

28
Q

What does an MPN test do, and what does MPN stand for?

A

Tests water quality; stands for ‘most probable number’

29
Q

What are the 3 parts of an MPN test?

A

1) Presumptive: data generated
2) Confirmed: data verified
3) Complete: published and acted upon (recommendations, policy, etc)

30
Q

What is the formula to calculate the number of bacteria in the original culture (cells/mL)?

A

Number of colonies divided by total dilution factor

31
Q

True or false: The number of colonies on a plate is the minimum number of cells originally put on that plate

A

True

32
Q

The statistically valid range of colony counts depends on what?

A

The size of the plate

33
Q

What range is considered a reliable colony count?

A

30-300

34
Q

How

A

Microbiology (42 decks)

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