Environmental Microbiology > Week two: Techniques > Flashcards

Week two: Techniques Flashcards

(58 cards)

Study These Flashcards
1
Q

What is the one of the main issues when sampling microbes?

A

They respond rapidly to a change in the environment (air contamination of anaerobic samples, temp/pressure changes) making ex situ analysis difficult.

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
2
Q

What is the best sample storage method for a direct measure of cellular components?

A

Storing at -20C to -70C, often kills the cells but prevents the decomposition of components (e.g. RNA)

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
3
Q

What is the best sample storage method for a cell count?

A

Sample fixation using formaldehyde or alcohol as this prevents ruptures of cells, even though it kills them

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
4
Q

What is the best sample storage method for a cultivation or activity measurement?

A

Must be conducted ASAP after storage at 4C, microbial communities may still have changes

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
5
Q

What are some of the equipment used for soil analysis?

A

Trowel (shallow)
Auger (depth)
Corer (water bodies)
Cylinder for extraction
Geoprobe for core soils (very deep)

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
6
Q

How do Johnson-ZoBell water samplers work?

A

Creates a vacuum to suck in water samples

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
7
Q

What are Niskin bottles?

A

Used for greater depth (40m) water sampling, lids open until desired depth reached, then triggered closed, airtight seals to avoid contamination before analysis conducted on board.

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
8
Q

What is a CTD sampler?

A

In situ water analysis, measures conductivity, temperature and depth, can attach chlorophyll fluorescence, pH, oxygen or turbidity measures. Measured through the column. Commonly set up as a series of bottles for deployment.

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
9
Q

What are the steps for analysis of microbes in water samples?

A

Concentration (usually), centrifugation or filtration. May require large volumes of water. 0.1um or 0.22um pore size filters used.

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
10
Q

How are microbes associated in sediment typically living?

A

Symbiotically within a complex 3D structure containing: quartz, organic matter, clay, water, microcolonies of bacteria

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
11
Q

How are microbial sediments analysed?

A

Complex 3D structure destroyed then varies

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
12
Q

What are some tests for geochemical indicators?

A

pH probe, REDOX probe, ion chromatography, ICP-AES, ICP-MS, GC, total inorganic/organic carbon

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
13
Q

How does ion chromatography work?

A

Filtered, unacidified samples undergo ion exchange with volatile fatty acids and carboxylates. Typical detection is limited to 50ppb.

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
14
Q

What are some common major inorganic anions that ion chromatography can identify?

A

F-, Cl-, Br-, BrO3-, NO3-, NO2-, SO42-, PO43-, I- and IO3-

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
15
Q

What are some selected metalloid oxyanions that ion chromatography can identify?

A

Selenite, selenate and arsenate

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
16
Q

What does ICP-AES stand for?

A

Inductively coupled plasma- atomic emission spectroscopy

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
17
Q

What does ICP-MS stand for?

A

Inductively coupled plasma- mass spectrometry

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
18
Q

How do ICP-AES and ICP-MS compare?

A

AES-
50ppb-500ppm
Typical 1% dissolved solid tolerance

MS-
1-1000ppb
<0.1% dissolved solid tolerance
Isobaric interferences make measurement difficult in some matrices

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
19
Q

What do ICP-AES and ICP-MS do?

A

Detect trace chemicals in soil, water and sediment samples to detect pollutants, such as heavy metals, and chemicals that can act as geochemical indicators

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
20
Q

What are some mineral characterisation/ identification methods?

A

Electron microscopy, x-ray photoelectron spectroscopy (surface level), x-ray diffraction (crystalline materials), x-ray absorption spectroscopy (changes in elements), x-ray fluorescence, IR spectroscopy (biomolecules), microbe examination, nanoSIMS and more

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
21
Q

What are some measurement methods of microbial numbers?

A

Direct counts, biomass estimation and culturing.

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
22
Q

What are the pros of direct count procedures?

A

Highest estimate of cells, good for large morphologically distinguishable cells (protozoa, fungi and algae)

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
23
Q

What are the cons of direct count procedures?

A

Counted individually with microscope (tedious), hard to distinguish the dead, background debris, little info about present organism

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
24
Q

How does a counting chamber stage micrometre work?

Study These Flashcards
A

Count cells in one chamber grid then assume that as an average for all the covered grids

25
How are florescent and epifluorescent dyes used in cell counts?
Acridine Orange Direct Counts (AODC) is a general nucleic acid stain, appears green or red, making count easier
26
How accurate are direct counts?
AODC gives counts 1-2 orders of magnitude higher than viable bacterial counts (by culturing)
27
How is propidium iodide used for cytological activity measurements?
Stains DNA fluorescent red, but only penetrates dead cells
28
How does flow cytometry work?
Sample mixed with flowing liquid, passed through an oscillating needle to produce droplets, droplets pass optical detector and measures light scatter-size, fluorescence and specific stains can be used for more information (direct count method)
29
What are some of the cons of flow cytometry?
Particulate material can clog the needle and clumps of cells can underestimate numbers
30
What are the pros of cultivation techniques?
Target specific organisms, gives indication of activity, gives model organisms for lab study
31
What are the cons of cultivation techniques?
Will underestimate diversity
32
How are cultivation techniques carried out?
Enumeration and isolation using selective media
33
What are the two main methods for viable count procedures (not direct counts)?
Plate counts and MPN (most probable number) counts (in liquid cultures)
34
How are plate counts conducted?
Dilution if necessary, spread evenly on agar, incubate and count colonies. Selective media for particular organisms and diagnostic media to identify organisms
35
How are MPN counts conducted?
Serially diluted, inoculated in appropriate medium and incubated, statistical analysis based on number of positive scores in replicate sub samples at each dilution. Needs clear positive reaction.
36
What are the cons of MPN counts?
Laborious, poor for total counts, often grow poorly on solid media
37
What are the pros of MPN counts?
Selectively imposed growth medium and good for lithotrophic organisms with low growth yields
38
How are direct chemical measurements used to asses biological activity?
Quantifying changes in concentration of biologically transformed chemical species (added manually or already in sample)
39
What biochemical processes can be measured with direct chemical measurements?
Nitrification/ denitrification- ammonium, nitrate, nitrite accumulation/loss Sulphate reduction- sulphate depletion (anoxic) Metal reduction Methane oxidation- methane depletion Methanogenesis- methane accumulation (anoxic)
40
What are the cons of direct chemical measurements?
Often limited sensitivity, measure potentials rather than in situ rates
41
How are radiotracers used to assess biological activity?
Uptake of radiolabelled compounds measured (typically C14). Degradation of C14 compounds (photosynthesis) Sulphate reduction with S35
42
How are specific metabolic inhibitors used?
Added to sample to measure effects on the processes, accumulation of electron donors can determine pathways of terminal carbon oxidation, and identify mechanism of biotransformation.
43
What are some metabolic inhibitors for sulfate reduction?
Sodium molybdate and fluoracetate
44
What is an inhibitor of methanogenesis?
Bromoethane sulfonic acid
45
How can isotopes be used to identify if a mineral is biogenic or abiogenic?
Organisms prefer lighter isotopes, isotopic ration compared to standard will be the same or lower for biogenic minerals. Abiogenic minerals will have a heavier isotopic ratio than the standard.
46
What percentage of microbes can be isolated in the lab?
<10%
47
What is the ideal molecular marker?
16S rRNA gene as it is highly conserved
48
What does PCR stand for?
Polymerase Chain Reaction
49
What happens during PCR?
Amplification of rRNA gene (any eDNA really)
50
What are the requirements for PCR?
Thermostable DNA polymerase, oligodeoxynucleotide triphosphates, Mg2+ ions, template DNA, primers (base pairs) and a buffer
51
What are the cons of PCR?
Very sensitive, potential bias through amplification of some templates and contamination, takes a few hours, becomes reagent-limited
52
How are PCR products analysed and purified?
Molecular cloning, prior to typing and sequencing. Gel electrophoresis, identify against database, high throughout sequencing approaches to negate purification/ separation needs
53
What does FISH stand for and what is it used for?
Fluorescence in situ hybridisation, identifies rRNA genes within a cell culture
54
What happens during FISH?
Cells dehydrated with ethanol, target ribosomes, fixed cells attach to microscope slides, immersed in hybridisation buffer with oligonucleotide probes labelled with fluorescent dye to recognise sequence, cell washed with buffer to remove unwanted probe, viewed with epifluorescence microscopy, cell fluoresces at appropriate wavelength
55
What are some water sampling methods for extraction?
Johnson-ZoBell water sampler and Niskin bottles
56
What are some in situ water sampling methods?
CTD sampler
57
What are the three components of environmental systems?
Microbial cell, aqueous environment and mineral surfaces
58
What are some methods of analysing microbial activities?
Direct chemical measurements, radiotracers and specific metabolic inhibitors
Environmental Microbiology
flashcards
Decks in class (10)
# Cards
Brainscape helps you reach your goals faster, through stronger study habits.
© 2026 Bold Learning Solutions. Terms and Conditions