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Microbiology Unit 1 > Unit 1 lab Exam > Flashcards

Unit 1 lab Exam Flashcards

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1
Q

What’s the purpose of an autoclave?

A

sterilizing

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2
Q

How do you prepare a smear?

A

FROM A SLANT

  1. aseptically transfer (w/ loop) a drop of water to slide
  2. transfer a small amount of bacteria to water (on slide) & spread to a thin film
  3. air dry a thin layer of specimen
  4. heat fix the slide (before staining)
FROM A BROTH
no need to add water
roll tube between hands to mix bacteria
transfer a loop full of broth to slide
air dry
heat fix
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3
Q

Why must slide be air dried before heat-fixing?

A

If slide is heat fixed when wet, the water surrounding the bacterial cells will boil, causing the cells to lyse (burst).

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4
Q

Purpose of heat-fixing?

A
  1. Kills bacteria

2. Adheres them to the slide so no need for a coverslip.

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5
Q

simple staining procedure

A

After heat-fixing:

  1. cover smear with crystal violet
  2. gently rinse with water bottle
  3. blot with bibulous paper
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6
Q

Basic dyes

A

crystal violet
safranin
methylene blue

surface of bacteria with neutral pH attract basic dyes

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7
Q

Acidic dyes

A

Congo red
Nigrosine

stain the background & leave microbe transparent

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8
Q

Gram stain: what does it do?

A

G(+) have thick cell wall so stain adheres to it; G(-) has thin cell wall so are decolorized when rinsed with acetone alcohol.

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9
Q

Gram stain - procedure

A
  1. Primary stain - crystal violet
  2. Rinse w/distilled water
  3. Mordant - iodine
  4. Decolorize - acetone alcohol
  5. Counterstain - safranin
  6. Rinse w/distilled water
  7. Blot w/bibulous paper
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10
Q

Acid Fast Stain (Kinyoun method) - purpose

A

distinguishes acid-fast Mycobacterium from other bacteria due to a waxy substance in its cell wall.

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11
Q

Acid Fast stain process

A
  1. Primary stain - carbol fuchsin
  2. Rinse - distilled water
  3. Decolorize - acid alcohol
  4. Rinse - distilled water
  5. Counterstain - Kinyoun brilliant green
  6. Rinse - distilled water
  7. Blot w.bibulous paper
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12
Q

Endospore stain - purpose

A

Endospores have a tough outer covering made of the protein keratin that resists staining. Heat (steaming) is used to drive the stain inside the endospore.

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13
Q

Endospore stain process

A
  1. Cover smear w/small piece of bibulous paper; apply malachite green to paper
  2. Steam slide
  3. Decolorize - distilled water
    4 Counterstain - safranin
  4. Rinse - distilled water
  5. Blot w/bibulous paper
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14
Q

Negative stain (capsule stain) - purpose

A

Used to identify bacteria with a thick glycocalyx capsule. Both G(+) and G(-) bacteria can form capsules. The negative stain (congo red) stains the background & the positive stain (crystal violet) stains the cell body. The capsule is left unstained.

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15
Q

Negative stain (capsule stain) process

A
  1. Add one drop of Congo Red to slide
  2. Aseptically transfer bacteria to stain, swirl loop around to mix the two
  3. Place edge of “spreader slide” in dye & push dye across slide, making a thin smear
  4. Dispose of spreader slide in bleach
  5. Allow smear to completely air-dry (do not heat fix - can cause cells to shrink, creating a possible “halo effect” which looks like a capsule, but isn’t)
  6. Add drop of Crystal Violet
  7. Rinse - distilled water
  8. Air dry
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16
Q

Which species needs Acid Fast stain?

A

Mycobacterium smegmatis - G(+) - cell walls are thick and waxy

17
Q

Which Acid Fast species are pathogenic?

A

Mycobacterium leprae - leprosy

Mycobacterium tuberculosis - tuberculosis

18
Q

Bacillus subtilis

A

G(+) - purple/blue
bacilli
endospore former

19
Q

Staphylococcus aureus

A

G(+) - purple/blue

cocci

20
Q

Escherichia coli

A

G(-) - pink/red
bacilli
pathogenic - food poisoning

21
Q

Streptococcus pyogenes

A

G(+) - purple/blue

cocci

22
Q

Pathogenic endospore formers

A 
Bacillus cereus - food poisoning
Bacillus anthracis - anthrax
Clostridium tetani - tetanus
Clostridium botulinum - botulism
Clostridium difficile - diarrhea
Clostridium perfringens - gas gangrene
23
Q

Capsule formers

A
  • use negative stain

- Klebsiella pneumoniae

24
Q

Mycobacterium smegmatis

A

G(+)
bacilli
use acid fast stain (b/c it has a waxy cell wall)

25
Q

Klebsiella pneumoniae

A

G(-)
bacilli
capsule former (use negative stain)

26
Q

Proteus mirabilis

A

G(-)
bacilli
causes UTIs

27
Q

Pathogenic bacteria

A 
Proteus mirabilis - UTIs
Mycobacterium leprae - leprosy
Mycobacterium tuberculosis - tuberculosisBacillus cereus - food poisoning
Bacillus anthracis - anthrax
Clostridium tetani - tetanus
Clostridium botulinum - botulism
Clostridium difficile - diarrhea
Clostridium perfringens - gas gangrene
28
Q

Pseudomonas aeruginosa

A

G(-)
bacilli
pathogenic - can cause pneumonia, UTI

29
Q

Conditions of an autoclave

A

121 degrees
15-20 minutes
15 psi

Microbiology Unit 1 (9 decks)

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