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BIO111: Microbiology > Lab Practical 2 > Flashcards

Lab Practical 2 Flashcards

1
Q

FTM

A

fluid thioglycollate media

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2
Q

what does thioglycollate do

A

reacts with oxygen to remove it to create an anaerobic environment

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3
Q

What does red indicate in thyioglycollate media?

A

Resazurin is added to FTM, if it turns red/pink it indicates the presence of oxygen

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4
Q

Brewer’s agar

A

a type of agar with methylene blue added.
Blue color indicates specimen has been in oxygenated environment. If placed in gaspak jar, it should turn clear if in anaerobic environment

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5
Q

GasPak jar

A

creates an anaerobic environment - packets are put inside that get rid of oxygen

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6
Q

Obligate Aerobes

A
  • Have SOD and catalase
  • must have oxygen for metabolism (aerobic respiration) and growth
  • Most fungi and protists, some bacteria (Bacillus, Pseudomonas)
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7
Q

Facultative Anaerobes

A
  • Usually have SOD and catalase
  • “flexible” – can survive with or without oxygen
  • Grow best with oxygen (aerobic respiration)
  • E. coli, Staphylococcus and Saccharomyces (yeast)
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8
Q

Microaerophiles

A
  • Only small amounts of SOD and catalase
  • Need small concentrations of oxygen (2-10%) for aerobic respiration
  • Large amounts of oxygen are inhibitory
  • Found in mucous linings of hollow organs
  • Helicobacter pylori
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9
Q

Obligate Anaerobes

A
  • Usually lack both SOD and catalase
  • Cannot grow if oxygen is present
  • No aerobic respiration
  • deep mud, lakes, oceans, inside animal bodies
  • Clostridium
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10
Q

Aerotolerant Anaerobes

A
  • May have SOD, but not catalase
  • Indifferent to oxygen
  • Do not use oxygen – obligate fermenters
  • Streptococcus pyogenes
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11
Q

catalase

A

degrates hydrogen peroxide into oxygen and water

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12
Q

superoxide dismutase

A

converts superoxide ion to oxygen and hydrogen peroxide

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13
Q

peroxidase

A

breaks down hydrogen peroxide to water with the help of NAD+

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14
Q

anaerobic respiration

A

inorganic compounds such as sulfate or nitrates replace oxygen as the terminal electron acceptor

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15
Q

results of growth patterns of FTM

A

Aerobic - grows only on top
Microaerophilic - towards top but not exposed to oxygen
Facultative - throughout media
Anaerobic - in the bottom of media

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16
Q

*Add lab 13

A

*study lab 13

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17
Q

Psychrophiles

A
  • cold loving, - 5C to 15C

- Grow in polar and glacial regions

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18
Q

Psychrotrophs

A
  • “cold feeding,” 20C to 30C
  • do not cause infections in humans
  • responsible for spoiling of refrigerated and frozen food
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19
Q

Mesophiles

A
  • middle-loving, 25C to 45C

- optimum temperature for human pathogens is around 37C

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20
Q

Thermophiles

A
  • heat loving, 45oC to 70oC

- Found in natural hot springs, compost

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21
Q

Hyperthermophiles

A
  • extreme heat loving, 70oC +

- usually Archaea, hydrothermal vents

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22
Q

Bacteria usually live in what kind of solution

A

hypotonic

cell wall gives protection from high osmotic pressure

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23
Q

what will bacteria do to maintain their environment

A
  • Bacteria will maintain hypotonic environment outside cell

- Pump in K+ or produce extra amino acids

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24
Q

Non-tolerant

A

needs hypotonic environment

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25
Q

Halotolerant

A
  • up to 10% NaCl
  • can tolerate moderate concentrations of salt
  • Staphylococcus on skin
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26
Q

Halophiles

A
  • (obligate halophiles)
  • require a high level of NaCl
  • marine microbes, extreme halophiles are Archaea living in salt lakes
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27
Q

Osmophiles

A

grow in high sugar concentrations

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28
Q

Temperature Requirement Classifications (5)

A 
1. Psychrophiles (cold-loving) 
Psychrotroph (Cold-feeding)
2. Mesophiles (middle-loving) 
3. Thermophiles (heat-loving) 
4. Hyperthermophiles
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29
Q

Classification using solute concentration (4)

A
  1. Halotolerant
  2. Halophilis (obligate halophile)
  3. Losmophiles
  4. Non-tolerant
30
Q

Oxidation & Fermentation Reactions (4)

A
  1. Sugar fermentation (Ferment Manitol, Glucose, Lactose)
  2. MR-VP (Methyl Red - Vogues Proskauer) Test
  3. Catalase Production Experiment
  4. Citrate Utilization Experiment
31
Q

Sugar fermentation (Ferment Manitol, Glucose, Lactose)

A
  • Durham tube (upside down tube w/in a tube)
  • Indicates the presence of gas that was produces as a result of fermentation (+/-)
  • Each tube contains a phenol red pH indicator
  • *Tube begins red:
  • NO color change= NO fermentation occurred
  • Color changed to yellow= fermentation occurred
32
Q

MR-VP (Methyl Red - Vogues Proskauer) Test

A
  • ->Results:
  • two tubes (original & new w/ 1 ml of broth) & 2 solutions to add.
  • Original tube: add methyl red indicator: Will turn red if pH dropped below 5 (red= mixed acid fermentation occurred)
  • Tube w/ 1 ml: add VP solution A (15) & VP solution B (5): If turns PINK/RED = 2,3 butanedial and ethanol fermentation (instead of acid fermentation) occurred.
  • *always a + MR/- VP or -MR/+ VP result**
33
Q

Catalase Production Experiment

A
  • Uses nutrient agar plate

- ->Results: Drop hydrogen peroxide on plate; if bubbling occurs = catalase positive

34
Q

Citrate Utilization Experiment

A
  • Uses Simmons agar slant (starts dark green) - double inoculate; stab with needle, smear slant
  • ->Positive= Change to blue
  • ->Negative= remains green
35
Q

Hydrolysis

A

-refers to bacteria using enzymes to do catabolic chemical reactions.

36
Q

Hydrolysis Experiments

A
  1. Starch Hydrolysis
  2. Urea Hydrolysis
  3. Tryptophan Hydrolysis
37
Q

Starch Hydrolysis

A

-experiment determines if bacteria have enzyme called amylase that breaks down starch to sugar.

38
Q

Starch Hydrolysis Results

A

Results–> Flood starch agar plate with Grams Iodine (IKI), IKI turns starch to a blue or dark color.
*Positive= “zone of clearing” (lighter coloration) around the colonies

39
Q

Urea Hydrolysis

A

-experiment determines if bacteria have enzyme called urease that breaks down urea into ammonia.

40
Q

Urea Hydrolysis Results

A

Results–> Tubes contain phenol red pH indicators (nothing added to tube)

  • Positive= broth turns red/purple (indicates presence of urea)
  • Negative= NO color change
41
Q

Tryptophan Hydrolysis

A

-experiment determines if bacteria have enzyme called tryptophanase that break down the amino acid tryptophan into indole and pyruvate.

42
Q

Tryptophan Hydrolysis Results

A

Results–> Add “indole reagent” to tryptophan broth

*Positive= Indole is present (and tryptophan broken down), RED layer will form at top of tube.

43
Q

Kligler’s Iron Agar

A

-experiment determines if the bacteria are capable of conducting acidic fermentation with glucose and lactose and if they produce hydrogen sulfide gas from amino acid cysteine.

44
Q

Kligler’s Iron Agar Results

A

Results–> Possible color changes in tube:

  • Bottom YELLOW, top REDDISH= glucose used during fermentation
  • Entire tube YELLOW= glucose and lactose used during fermentation
  • Tube turns BLACK= Amino acid cysteine into pyruvic acid, ammonia, and hydrogen sulfide
45
Q

Diagnostic key

A

-used to identify unknown bacteria.

46
Q

Dichotomous key

A
  • the key always has two answers for each question asked, determines next question or test.
47
Q

thermal death time

A

-time required to destroy a population of bacteria at a specific temperature

48
Q

thermal death point

A

-temperature required to destroy a population of bacteria in 10 minutes

49
Q

the area around the disk (antibiotic or cleanser) with no bacteria

A

zone of inhibition

- measured in mm

50
Q

Oxygen Requirement Classifications (5)

A
  1. Obligate aerobe
  2. Facultate anaerobe
  3. Obligate anaerobes
  4. Microphile
  5. Aerotolerant (Aerotolerant Anaerobe, Obligate Fermenter)
51
Q

0

A

no colony growth

52
Q

+

A

1-10 colonies

53
Q

++

A

11-100 colonies

54
Q

+++

A

101 - uncountable

55
Q

why is UV light lethal bacteria

A

It causes mutations, such as thymine dimers

56
Q

Dilution Method Experiment

A
  • toothpicks soaked in bacteria where then soaked in a disinfectant for varying lengths of time and then rinsed and placed in nutrient broth.
57
Q

filter paper disk method

A

a plate is inoculated and filter paper disks soaked in disinfectant are placed on the plate and incubated. The disinfectant with the greatest zone of inhibition is the most effective.

58
Q

When performing plate counts what dilution is easiest to count

A

1:10,000,000

59
Q

how to calculate cells/mL

A

(total # of cells counted in six med squares) x dilution factor x 1000
divided by 0.024

60
Q

How to count squares

A

counts 6 squares, ignore cells touching upper and left borders of a box, but count the cells found on lower or right borders

61
Q

Only count plates with how many colonies

A

30 - 300
If the cfus are >300, overcrowding on the plate could have inhibited some cells from growing. If cfus are <30, could involve a sampling error.

62
Q

name of device used to count squares

A

Naubauer counting chamber

63
Q

Types of direct counts

A

viable

non-viable

64
Q

viable count

A

standard count, living organisms

65
Q

non-viable

A

living & dead cells

66
Q

How are population counts conducted?

A

A sample can be diluted and cells in sample can be counted with a microscope and Petroff-Hauser chamber.

67
Q

Why are population counts important?

A

To determine the number of bacteria in a sample.

Ex: A diagnosis of bladder infection depends on a certain threshold level of bacteria present in a urine sample.

68
Q

What is a serial dilution?

A

Start with culture.
Add 1 mL of sample to 100 mL of sterile water = 1:100
Remove 1 mL of 1:100 sample and add to 100 mL of sterile water = 1:1000
Remove 1 mL of 1:1000 sample and add to 100 mL of sterile water = 1:10,000
Remove 1 mL of 1:10,000 sample and add to 100 mL of sterile water = 1:1,000,000

69
Q

Why are serial dilutions necessary?

A
  • Bacteria cells are diluted to an end point where a single cell divides giving rise to the visible colony on a plate.
  • Multiply the number of colonies by the dilution factor to determine the number of bacteria in the original sample.
70
Q

Advantages/disadvantages of SPC

A

Advantages: SPC determines only viable cells

Disadvantages: Specific conditions and media are used and these factors may exclude certain bacteria.

BIO111: Microbiology (12 decks)

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