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Bio 211 > LAB Test 2 > Flashcards

LAB Test 2 Flashcards

Ex. 9-16

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1
Q

Form for scientific notation

A

a × 10b

1 < a < 10

b gives the order of magnitude

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2
Q

What are plate counts used for?

A

Determining water, milk, food quality

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3
Q

103

A

kilo

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4
Q

100

A

Base unit

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5
Q

10-3

A

milli

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6
Q

10-6

A

micro

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7
Q

10-9

A

nano

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8
Q

Why are serial dilutions used?

A

Method used to efficiently calculate the number of microbes in a sample.

Dilutions will thin out the lawn until we can see single colonies, which we can physically count

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9
Q

Diluent

A

a fluid used to dilute the concentrated sample

(e.g. water)

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10
Q

Aliquot

A

a smaller volume withdrawn from a total sample volume

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11
Q

Dilution factor

A

Fraction by which your original sample concentration is diluted

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12
Q

Equation for dilution factor

A

DF = Vf / Vi

Vf = aliquot volume + diluent volume

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13
Q

Equation for dilution

A

Dilution A = (total dilution)(original concentration)

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14
Q

Equation for final dilution

A

Final dilution = product of total dilutions at every step in the series

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15
Q

Final sets of the tubes have the highest ______ and the lowest ______.

A

dilution; concentration

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16
Q

P1000

A

200-1000µL

0 | 9 | 0 (=900µL)

Larger blue tips

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17
Q

P200

A

20-200µL

1 | 0 | 0 (=100µL)

Smaller yellow tips

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18
Q

Using micropipettors

A
  1. Depress plunger to FIRST STOP befor you place it into the sample
  2. Slowly return the plunger to the up position to suck up the liquid
  3. Release liquid by depressing plunger all the way to the SECOND STOP
  4. Change tips after every​ transfer
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19
Q

Spread Plating

A
  1. Dip the spreader into alcohol; let excess alcohol drip back into the jar before flaming (to sterilize)
  2. Wait until the plate spreader is cool
  3. Apply liquid culture to plate and spread
    1. Take only 100 µL from the tube to spread on the plate
      1. This will result in a 1/10 dilution
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20
Q

When are plates considered countable?

A

30-300 colonies

TNTC (too numerous to count): >300 colonies

TFTC (too few to count): < 30 colonies

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21
Q

Calculating the number of bacteria in the original sample

A

O.T. Concentration = Number of CFUs / (Total Dilution)(Volume Plated)

O.T. Concentration = (Number of CFUs)(Total Dilution Factor) / Volume Plated

units: CFU/mL

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22
Q

Growth Characteristic Tests

A
  1. Osmotic pressure
  2. O2 requirements
  3. pH
  4. Temperature
  5. ​Pigment production
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23
Q

Strict aerobe

A

connot exist without 02

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24
Q

Strict anaerobe

A

cannot live in the presence of O2

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25
Q

Facultative anaerobe

A

can grow with or without O2, but they grow better with O2 present

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26
Q

Aerotolerant anaerobe

A

cannot use O2 for growth, but can tolerate it

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27
Q

Microaerophile

A

Grow best in high CO2 / low O2 environment

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28
Q

Thioglycolate medium

A
  • used in culturing anaerobes
  • sodium thioglycolate binds free O2
  • Creates an O2 gradient (most O2 at the top of the tube and progressively less O2 towards the bottom)
    • agar reduces the rate of diffusion of O2 into and out of medium
  • Contains resazurin, an indicator dye that turns pink in the presence of O2 and is colorless in absence of O2
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29
Q

Examples of strict (obligate) aerobes

A
  • Bacillus subtilis*
  • Mycobacterium tuberculosis*
  • Mycobacterium leprae*
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30
Q

Examples of strict (obligate) anaerobes

A
  • Clostridium perfringens*
  • Clostridium botulinum*
  • Clostridium tetani*
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31
Q

Examples of Facultative Anaerobes

A
  • Staphylococcus spp.*
  • Corynebacterium spp.*
  • Bacillus anthracis*
  • Klebsiella spp.*
32
Q

Examples of Aerotolerant Anaerobes

A

Clostridium sporogenes

33
Q

Anaerobic jar

A
  • used to incubate regular media in an anaerobic environment
  • Uses catalysts to combine O2 with H2 to form H2O
  • packet - gives off CO2 after about 3 hours to create CO2 atmosphere in the absence of O2
  • Contains methylene blue indicator strip
    • Blue= O2 present
    • White = O2 absent
34
Q

What does temperature affect in bacteria?

A

function of enzymes and proteins essential for growth and survival

35
Q

Maximum growth temperature

A

highest temperature in which species will grow (will usually have minimal growth rate)

36
Q

Minimum growth temperature

A

lowest temperature in which species will grow (will usually have minimal growth rate)

37
Q

Optimum growth temperature

A

Temperature at which species have the highest growth rate

38
Q

Mesophile

A

Grows best between 25-40º C

(e.g. human gut flora)

39
Q

Psychrophile

A

Grows between 0-5ºC

(e.g. bacteria in the deep sea, Arctic regions; food spoilage microbes)

40
Q

Thermophile

A

Grows between 45-65ºC

(e.g. bacteria in hot springs; hyperthermophiles can survive near-boiling conditions like deep sea vents)

41
Q

What controls pigment production?

A

enzyme activity

pigment is the product of biochemical pathways involving enzymes

some bacteria make pigments in certain conditions (usually temperature)

42
Q

Diffusible pigment

A

a pigment that can diffuse into agar from the colonies (soluble in water)

43
Q

Non-diffusible pigment

A

a pigment that is confined to the colony

44
Q

Pseudomonas aeruginosa

A
  • opportunistic and nosocomial pathogen
  • produces two types of soluble pigments
    • fluorescent green
    • blue
  • Hallmark of infection: “blue pus”
45
Q

Osmosis

A

a movement of water across a semi-permeable membrane from a lower solute concentration to a higher solute concentration

46
Q

Osmotic pressure

A

pressure that must be applied to the solution to equalize the flow of water across the semi-permeable membrane in both directions

47
Q

Hypertonic environment

A

solute concentration outside the cell is higher

water flows out of the cell

48
Q

Plasmolysis

A

cytoplasm shrinks; cell collapses

result of hypertonic environment

49
Q

Hypotonic environment

A

Solute concentration outsie the cell is lower

water flows into the cell

50
Q

Cell lysis

A

cytoplasm swells; cell explodes

result of hypotonic environment

51
Q

Isotonic environment

A

solute concentrations across the membrane are equal

no water flow in either direction

no osmotic pressure

52
Q

What will happen if there is improper pH?

A

“fold” the proteins differently, impairing their function

53
Q

What pH do most bacteria grow at?

A

neutral pH (6.5-7.5)

some grow at acidic pH (low) or alkaline pH (high)

54
Q

Acidophiles

A

prefer acidic conditions (low pH)

EX: Thiobacillus ferrooxidans lives in coal mines and can survive at pH 1

55
Q

Alkaline bacteria

A

prefer alkaline conditions (high pH)

EX: Vibrio cholerae can live up to pH 9

56
Q

Endospores

A
  • encapsulate genetic material (DNA) and essential cell components
  • Enable bacteria to survive unfavorable environmental conditions
    • starvation
    • Extreme heat/cold
    • Dessication
    • Toxic chemicals/antibiotics/radiation
  • Once favorable conditions return, the endospores can germinate to form vegetative cells
57
Q

Endospore staining

A

Primary stain: Malachite green (10-15 min)

Counterstain: Safranin (30 sec)

Vegetative cells: pink

Endospores: blue

58
Q

Transient bacteria

A

contaminants; may be present for a finite time

may be pathogenic

59
Q

Resident bacteria

A

permanent residents of normal human flora

normally, not pathogenic

60
Q

Ignác Semmelweis

A

introduced concept of hand-washing

suggested washing with chlorinated water

61
Q

Common skin microbiota

A
  • Corynebacterium diphtheriae*
  • Staphylococcus aureus*
  • Micrococcus luteus*
  • Staphylococcus epidermidis*
  • Pityrosporum ovale*
62
Q

Isolation by dilution

A

dilute the culture enough that single cells give rise to single colonies on a plate (all cells in a single colony are clones)

Pour plate, Spread plate, Streak plate

63
Q

Colony morphology

A
  • color
    • white, cream, tan, brown, black, purple, yellow, red, etc
  • size
    • pinpoint, small, medium, large
  • shape
    • raised, flat, convex, fried egg, dimpled (caved in), growth in agar
  • margin
    • smooth, irregular, filamentous, rhizoid, wavy/swarming, entire, filamentous, spreading
  • consistency
    • shiny, glistening, dry, powdery, wrinkled, rough, dull/matte, mucoid
64
Q

T-Streak Plate

A
  1. Pick up culture
  2. Inoculate in Zone 1
  3. Flame loop
  4. Streak through Zone 1 into Zone 2
  5. Flame loop
  6. Streak through Zone 2 into Zone 3
65
Q

Cotton Applicator Stick

A
  • Sterile polyester-tipped swab
  • Swab the surface of interest and place it in the provided case
  • Case = glass ampule which contains general purpose medium
    • survives for up to 72 hours
66
Q

Why is quality control used?

A

to ensure that:

  • procedures and results can be evaluated for accuracy
  • there are controls provided for comparison
  • the dyes/reagents are in good condition

ALWAYS use when identifying an unknown organism with Gram staining

67
Q

Gram staining

A
  • Differentiates two large groups of bacteria based on their cell wall compositon
    • Gram negative (Gm-) [pink] and Gram positive (Gm+) [purple]
      • helps to determine appropriate antibiotic treatments

Primary stain: crystal violet (1 min)

Mordant: Gram’s iodine (1 min) -complexes crystal violet to peptidoglycan layer

Decolorizing agent: ethanol (10-20 sec) - dissolves lipids in Gm-cells enhancing decoloization and dehydrates Gm+ cells closing off pores

Counter-stain: safranin (1 min)

68
Q

Bacterial cell wall

A
  • composed of peptidoglycan
    • polymer of amino acids (“peptido-“) and sugars (“-glycan”)
    • layered on top of the plasma membrane
  • Thickness of the layer of peptidoglycan comprising the cell wall determines Gram character
    • Thick = Gm+
    • Thin = Gm-
69
Q

What is the charge of the exterior of the cell?

A

anionic charge

70
Q

What type of dye is crystal violet?

A

cationic dye

71
Q

What will happen if you forget to add mordant?

A

Gm+ may appear pink

72
Q

What will happen if you decolorize too long?

A

Gm+ may appear pink

73
Q

Acid-Fast Stain

A

Used to identify Mycobacterium spp. on the basis of their mycolic acid layers

Primary dye: Kinyon’s TB carbolfuchsin (KF) - chemically binds to mycolic acid in cell wall

Decolorizer: Acid alcohol - washes away KF if not bound to mycolic acid

Counterstain: Methylene blue

Interpretation: dark red/pink rods (positive); blue (negative)

74
Q

Glycocalyx

A

slimy layer surrouding cells made of polysaccharides (“sugar coat”) or sometimes of proteinaceous material

protects cell from immune system

75
Q

Virulence factor

A

any bacterial adaptation that makes them more pathogenic

76
Q
A
  1. Capsule
  2. Nuclear matter
  3. Cytoplasm
  4. Cell membrane
  5. Cell wall
  6. Pili
  7. Flagellum
77
Q

Capsule staining

A

The capsule repels dyes so it cannot be stained directly

Negative Staining

  • Background is stained
  • Capsule is visualized as a halo around the cell
  • Indirect visualization

Bio 211 (5 decks)

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