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MT6322 MYCOVIRO > E3 LAB: BASIC TECHNIQUES ON FUNGAL ISOLATION > Flashcards

E3 LAB: BASIC TECHNIQUES ON FUNGAL ISOLATION Flashcards

1
Q

Media and Reagents

A
  • Potato Dextrose Agar (PDA)
  • Agar
  • Lactophenol cotton blue
  • Sterile distilled water
  • Plated media (PDA and Water Agar)
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2
Q

Equipment and Materials

A
  • Autoclave
  • Dissecting microscope
  • Compound microscope
  • Sterile Petri dish
  • Sterile Test Tubes (Loeffler’s)
  • Sterile slides and coverslips
  • Alcohol lamp
  • Scalpel, blade 15A
  • Agar block
  • Filter paper
  • Glass pasteur pipette
  • Erlenmeyer flask
  • Micropipettor
  • Bent glass rod
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3
Q

Guide template for Agar block

A

5 mm x 5 mm

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4
Q

Specimen

A
  • Soil
  • fungal sample in PDA (Aspergillus niger ATCC 16404)
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5
Q

Method 1

A

Agar Blocking

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6
Q

Agar Blocking

Scalpel needle must cut down the medium touching the?

A

bottom part of plate

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7
Q

Agar Blocking

Lift the agar block and transfer it on a slide, placed inside a?

A

petri dish

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8
Q

Agar Blocking may be used to?

A

purify fungi in a mixture of microbial elements

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9
Q

Agar Blocking

Small block is usually cut from the?

A

outermost part of fungal growth

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10
Q

T/F: Agar block may also be used in slide-agar culture technique

A

True

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11
Q

Other term for slide culture technique

A

Modified Riddell Technique

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12
Q

Slide Culture Technique

Inoculate small fragment of sample to?

A

four edges of the block

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13
Q

Slide Culture Technique

Incubate at what temp, where, and for how many days?

A

room temperature (22-25C), locker for 10 days

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14
Q

Slide Culture Technique

Once sporulation has developed, transfer cover slip on slide with a?

A

drop of Lactopheral cotton blue

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15
Q

Spore Touch Isolation Technique

A

Pasteur pipet tip elongation

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16
Q

Pasteur pipet tip elongation

Place forcep at?

A

tip of pipet

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17
Q

Pasteur pipet tip elongation

Heat what part of the pipette?

A

middle part of thin portion of pipette

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18
Q

Pasteur pipet tip elongation

Once it softens, you must?

A

pull apart the end

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19
Q

Pasteur pipet tip elongation

When the tip cools, make sure it is?

A

straight

HOMOKOJIC

20
Q

Spore isolation technique allows?

A

purification of fungal isolate in a given medium with mixed microbial growth

21
Q

Spore Isolation Technique

This can be examined microscopically in a dissecting microscope.

A

Aerial spores

22
Q

Soil sprinkling technique

Scoop soil sample and sprinkle using?

A

long handle square head spoon

23
Q

Soil sprinkling technique

Incubate at?

A

room temperature

24
Q

Soil sprinkling technique

Observe growth after?

A

1 week

25
Q

Brush paint technique

Preparation of soil suspension

A
  • 1g of soil in 50mL beaker
  • 3mL distilled water
  • mix using a stirring rod
26
Q

Brush paint technique

Dip what paint brush in suspension?

A

paint brush #6

27
Q

Brush paint technique

Spread ________ layer on plated media

A

3-single layer

28
Q

This decreases number of microorganisms and allow distinct microbial growth.

A

Serial dilution

29
Q

Serial dilution favors what organisms?

A

rapidly growing

30
Q

Examples of rapidly growing organisms

A

Fungi Imperfecti
- Penicillium
- Aspergillus sp.

31
Q

Serial dilution

Preparation of soil suspesion

A
  • 1g of soil
  • 100mL of distilled water
  • shake for 15 minutes
32
Q

Serial dilution

Soil suspension will give a?

A

10-2 dilution

33
Q

Serial dilution

Set up what tubes?

A

Loeffler’s test tubes

34
Q

Serial dilution

Add how many mL of distilled water?

A

9.0mL

35
Q

Serial dilution

Transfer how many mL of the soil supernatant fluid of last dilution, and where?

A

0.1mL, center of agar medium

36
Q

Serial dilution

Incubate at what temperature for how many days?

A

37C for 3-5 days

37
Q

Serial dilution

Further dilution allows isolation of?

A

Actinomycetes

38
Q

Aerial spore isolation

A

Cellophane Technique

39
Q

Cellophane Technique

First step

A

Place drop of Lactophenol Cotton Blue on a slide

40
Q

Cellophane Technique

Extend a?

A

2 inch adhesive tape

41
Q

Cellophane Technique

Touch adhesive side onto the?

A

upper surface of old fungal culture

42
Q

Cellophane Technique

What are expected to stick to the tape?

A

Aerial spores and hyphal elements

43
Q

Cellophane Technique

To secure tape in the slide, you must flip?

A

both ends of the tape

44
Q

For keratinophiles

A

Hair bait technique

45
Q

Hair bait technique

_____ a sterile petri dish with soil specimens.

A

Half-fill

46
Q

Hair bait technique

Moisten soil specimen with?

A

15mL of sterile distilled water

47
Q

Hair bait technique

Place short tufts of what on soil surface?

A

autoclaved hair samples

MT6322 MYCOVIRO (10 decks)

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