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Microbial Ecology (BIOL 355) > Lecture 7: Sampling and Enumeration > Flashcards

Lecture 7: Sampling and Enumeration Flashcards

1
Q

What are the components of a quality assurance plan?

A
  1. Sampling
    - Number, type, location, timing
    - Replication
  2. Methods
    - Specific techniques, etc.
  3. Storage
    - Preservation (refrigeration, freezing, fixation)
    - Duration issues
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2
Q

What are the challenges in sampling?

A
  • Replication: can be difficult to ensure it’s the same
  • Little to no baseline data
  • Controls: need to find something that doesn’t have the thing you’re researching
  • Little to no data in acceptable ranges
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3
Q

What is the function of transects?

A

You can just sample along the transects instead of the entire sample area
- Ex. a lake, sampling along even x amount of meters

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4
Q

What are some issues to consider in soil/sediment sample collection?

A
  • Contamination: hard to decontaminate such a large drill
  • Compaction: a portion of sediment might clog your piston
  • Water content: sometimes too much water
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5
Q

How are soil/sediment samples collected?

A
  • Coring: piston corer (can get nice stratification)
  • Van Veen grab
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6
Q

Why is aquatic sampling easier than soil/sediment sampling?

A
  • More homogeneous
  • Easier to get samples, even remotely
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7
Q

What are some difficulties in aquatic sampling?

A
  1. May have to filter/concentrate large volumes of water
  2. Viruses require care
    - High volumes
    - Adsorption, elution, reconcentration
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8
Q

What are some bottles and nets used in aquatic sampling? When would you use each one?

A
  • Plankton net tows: to isolate microbes of different sizes
  • Niskin bottle:
  • CTD rosette: measure physical characteristics of water
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9
Q

Compare and contrast crossflow filtration and direct flow filtration.

A
  1. Crossflow (aka tangential flow)
    - High permeate rate
  2. Direct flow (aka dead-end)
    - Low permeate rate
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10
Q

What is eDNA sampling?

A
  • Way to monitor different species in an environment
  • Looking at all the DNA in a sample
  • Snapshot of total community diversity
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11
Q

How is air sampling conducted?

A

Impaction on agar

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12
Q

What is enumeration?

A

Determining how many microbes there are in a sample

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13
Q

What are the difficulties in the environment in enumeration?

A
  • Small size of microbes - hard to see to count
  • Hard to determine viability of samples
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14
Q

What is the Great Plate Anomaly?

A

In certain environments, <1% of microorganisms are cultivable (VBNC)

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15
Q

List viable count methods (aka standard plate count).

A
  • Plate counts and most probable number
  • Selective growth medium
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16
Q

What does growth (as in on a plate) represent?

A

Presence and growth potential
- NOT the environmental potential
- Does not give total count like direct counting

17
Q

Explain serial dilution.

A
18
Q

How do you get the most probably number (MPN)?

A
  • Serial dilution of a liquid sample
  • Growth in successive dilutions allows estimation of numbers in original smaple
19
Q

How is turbidity measured?

A

Optical density
- Measure how much light passes through the sample
- Use the same media as your blank

20
Q

What tools are used to take direct counts?

A
  1. Microscopic measures
    - Counting chamber
    - Gridded filters
  2. Epifluorescence microscopy
    - Fluoresce when you hit them (natural autofluorescence, acridine orange, DAPI)
21
Q

What are fluorochromes?

A

Molecules that emit light in response to irradiation

22
Q

What is stokes shift?

A

Emission is always of longer wavelength than excitation

23
Q

What is photobleaching?

A

Fading of FCs

24
Q

What are the advantages and drawback of direct counts?

A
  1. Advantages
    - Higher numbers (10-100x more than viable counts)
    - Not environment-specific
    - Correlation with biomass
  2. Drawbacks
    - Hard to tell live from dead
    - Debris interference
    - Inability to take any further analysis (typically fix cells with formaldehyde)
    - Laborious
25
Q

What are automated direct counts?

A

Digital image analysis software to count cells
- Must be a “clean”, clear, bright sample
- Expensive

26
Q

What is flow cytometry? When would you use it?

A
  • Analyzes the size, shape, and properties of cells by passing them through a tube single-file and then shooting a laser beam at them
  • Soil sample, algal blooms, seawater samples for counting phytoplankton
27
Q

What is cryopreservation protocol?

A
28
Q

Which habitats are easiest to get a representative sample?

A. Soil
B. Seawater
C. Deep subsurface
D. Lake sediment

A

A. Soil
B. Seawater
C. Deep subsurface
D. Lake sediment

Microbial Ecology (BIOL 355) (17 decks)

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