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BACTERIOLOGY - LAB > Staining > Flashcards

Staining Flashcards

1
Q

PREPARATION OF SPECIMENS
FOR LIGHT MICROSCOPY

A

BACTERIAL STAINING

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2
Q

PURPOSE OF STAINING

A
  1. Observe and appreciate the
    appearance of microorganism
  2. Differentiate one microorganism or group
    of microorganism from another
  3. Identification of microorganisms and their
    special structures
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3
Q

cationic dyes with positively charged (pentavalent nitrogen) that adhere to the negatively charged molecules

A

Basic Dyes

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4
Q

Example of BASIC DYES

A

Crystal Violet, Methylene Blue
Malachite Green and Safranin

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5
Q

anionic dyes with negatively charged groups
(carboxyl and phenolic) that bind to positively
charged cell structures

A

Acidic dyes

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6
Q

Example of ACIDIC DYES

A

Eosin, Acid Fuchsin and Nigrosin

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7
Q

(TRUE OR FALSE)
Bacteria are slightly negatively charged at pH 7

A

TRUE

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8
Q

kills the microorganisms and fixes them to the slide

A

FIXING

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9
Q

preserves various parts of microbes in their natural
state with only minimal distortion

A

FIXING

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10
Q

Example of FIXING

A

a. Heat-fixed
b. Methanol Fixation

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11
Q

95% Methanol for I minute

A

Methanol Fixation

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12
Q

preserves morphology of host
cells, bacteria

A

Methanol Fixation

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13
Q

especially useful for examining
bloody specimen material

A

Methanol Fixation

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14
Q

Three kinds of staining techniques:

A
  1. SIMPLE
  2. DIFFERENTIAL
  3. SPECIAL
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15
Q

single stain is used

A

SIMPLE STAINS

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16
Q

highlight the entire microorganism so that
cellular shapes and basic structures are visible

A

SIMPLE STAINS

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17
Q

stain is applied to the fixed smear for a certain
length of time and then washed off, dried and
examined

A

SIMPLE STAINS

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18
Q

Example of SIMPLE STAINS

A

Methylene Blue
Carbolfuchsin
Crystal Violet
Safranin

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19
Q

react differently with different kinds of
bacteria and thus can be used to
distinguish them

A

DIFFERENTIAL STAINS

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20
Q

Example of DIFFERENTIAL STAINS

A

Gram Stain
Acid-Fast Stain

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21
Q

classifies bacteria into two large group

A

Gram-positive
Gram-negative

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22
Q

retain the dye and remain
purple

A

Gram-positive cells

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23
Q

do not retain the dye; they are
colorless until counterstained with a red dye

A

Gram-negative cells

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24
Q

chemically bond the alkaline dye to the
bacterial cell wall

A

MORDANT

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25
Q

chemical added to the solution to intensify the
stain

A

MORDANT

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26
Q

increase the affinity of a stain for a biological
specimen

A

MORDANT

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27
Q

coat a structure to make it thicker and easier to
see after it is stained with a dye

A

MORDANT

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28
Q

Example of MORDANT

A

GRAM’S IODINE

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29
Q

Valuable information for the treatment of disease

A

Gram-positive bacteria -> tend to be killed easily by PENICILLINS and CEPHALOSPORINS

Gram-negative bacteria -> generally more resistant because the antibiotics cannot penetrate the lipopolysaccharide layer

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30
Q

tend to be killed easily by
PENICILLINS and CEPHALOSPORINS

A

Gram-positive bacteria

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31
Q

generally more resistant
because the antibiotics cannot penetrate
the lipopolysaccharide layer

A

Gram-negative bacteria

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32
Q

contain a highly cross-linked layer of peptidoglycan
that retains the primary dye-Crystal Violet-following the
application of the mordant-iodine (I)

A

Gram-Positive Organisms

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33
Q

iodine and crystal violet form a complex within the peptidoglycan

A

Gram-Positive Organisms

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34
Q

when decolorized—CV-I complex remains within the cell

A

Gram-Positive Organisms

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35
Q

Appearance of GRAM-POSITIVE ORGANISMS

A

Dark Purple to Deep Blue

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36
Q

do not contain a thick cross-linked layer of
peptidoglycan

A

Gram-Negative Organisms

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37
Q

CV-I complexes are not trapped within
the peptidoglycan

A

Gram-Negative Organisms

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38
Q

decolorizer dehydrates the outer cellular
membrane, leaving holes in the membrane
and effectively washing or removing the
CV-I complex from the cells

A

Gram-Negative Organisms

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39
Q

Secondary stain in GRAM-NEGATIVE ORGANISMS

A

Safranin

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40
Q

Appearance of GRAM-NAGATIVE ORGANISMS

A

Pink to Deep Magenta

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41
Q

Report the Gram stain organism’s cellular
shape, morphology, and Gram reaction

A

Indirect Smear

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42
Q

QUALITY CONTROL
Gram-positive:
Gram-negative:

A

Gram-positive: Staphylococcus aureus
Gram-negative: Escherichia coli

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43
Q

General Rules of Gram Staining!!!

A
  1. All COCCI are GRAM-POSITIVE except for
    Neisseria, Veilonella and Branhamella
    (Moraxella)
  2. All BACILLI are GRAM-NEGATIVE except for
    Arcanobacterium, Actinomyctes, Bacillus,
    Clostridium, Corynebacterium, Erysipelothrix,
    Eubactrium, Gordonia, Kuthria, Lactobacilli,
    Listeria, Mycobacteria, Nocardia,
    Propionibacterium and Tsukamurella
  3. All spirochetes are GRAM-NEGATIVE
44
Q

All COCCI are GRAM-POSITIVE except for
_________________

A

Neisseria, Veilonella and Branhamella
(Moraxella)

45
Q

All BACILLI are GRAM-NEGATIVE except for
________

A

Arcanobacterium, Actinomyctes, Bacillus,
Clostridium, Corynebacterium, Erysipelothrix,
Eubactrium, Gordonia, Kuthria, Lactobacilli,
Listeria, Mycobacteria, Nocardia,
Propionibacterium and Tsukamurella

46
Q

(TRUE OR FALSE)
All spirochetes are GRAM-POSITIVE

A

FALSE
All spirochetes are GRAM-NEGATIVE

47
Q

Reasons Why Gram-Positive Bacteria Becomes
Gram-Negative

A
  1. Removal of MgRNA by precipitation with bile salts
  2. Autolysis, aging and temperature of incubation result
    to loss of gram-positivity
    ***Antibiotic-treated bacterial cell have atypical
    staining reaction
  3. Acidic solution of Gram’s Iodine
  4. Technical error Overdecolorization
48
Q

Reasons Why Gram-Negative Bacteria
Becomes Gram-Positive

A
  1. Incomplete decolorization
  2. Thick smear
49
Q

binds strongly only to bacteria that have a
waxy material in their cell walls

A

ACID-FAST STAIN

50
Q

Principle of ACID-FAST STAIN

A

Acid-fast organism contain MYCOLIC ACID in
their outer membrane, making the cells waxy and
resistant to staining with aqueous based stains
such as the Gram stain

51
Q

Acid-fast organism contain in ______
their outer membrane, making the cells waxy and
resistant to staining with aqueous based stains
such as the Gram stain

A

MYCOLIC ACID

52
Q

Example of ACID-FAST ORGANISM

A

Mycobacteria spp, Nocardia

53
Q

primary stain in ACID-FAST STAIN

A

Carbolfuchsin

54
Q

allow the stain to penetrate into the waxy
surface of acid-fast microorganisms

A

Heat or Tergitol

55
Q

-Ethanol and Hydrochloric Acid
-removed excess stain

A

3% Acid Alcohol

56
Q

secondary stain in ACID-FAST STAIN

A

Methylene Blue or Malachite Green

57
Q

Expected Results

Acid-Fast Organisms:
Nonacid-Fast Oganisms:
***Background material
should stain _____

A

Acid-Fast Organisms: PINK
Nonacid-Fast Oganisms: DARK BLUE
***Background material
should stain BLUE to BLUE-GREEN

58
Q

Ways to Facilitate Acid-Fast Staining

A
  1. Use of heating or steaming process for 5-7
    minutes to temporarily remove the mycolic acid,
    while the smear is flooded with stain
  2. Increasing the concentration of dye and phenol
    in the staining reagent
  3. Prolonged contact of the specimen with the
    primary stain
  4. Addition of a wetting agent like TERGITOL
59
Q

METHODS OF ACID-FAST STAINING

A

Ziehl-Neelsen
Kinyoun’s Method
Pappenheim Method
Baumgarten Method
Auramine-Rhodamine Method

60
Q

Hot Staining Method

A

Ziehl-Neelsen

61
Q

Cold Staining Method

A

Kinyoun’s Method

62
Q

Differentiate M. smegmatis from M.
tuberculosis

A

Pappenheim Method

63
Q

(Pappenheim Method)
decolorized by the mixture
of rosolic acid and alcohol
coloring it BLUE

A

M. smegmatis

64
Q

(Pappenheim Method)
not decolorized and
remains RED

A

M. tuberculosis

65
Q

Differentiate M. tuberculosis from M.
leprae

A

Baumgarten Method

66
Q

(Baumgarten Method)
does not readily take
up the stain and
appears BLUE

A

M. tuberculosis

67
Q

(Baumgarten Method)
easily stained by dilute
alcoholic fuchsin
coloring it RED

A

M. leprae

68
Q

selective for the cell wall of AFB

A

Auramine-Rhodamine Method

69
Q

Mycolic acid renders the bacterial cell resistant to
decolorize-_______

A

ACID-FAST

70
Q

is affected by colonial age, medium for
growth and UV light

A

Acid-Fastness

71
Q

ideal for concentrated smears
partially acid-fast bacilli—Nocardia spp

A

Ziehl-Neelsen method

72
Q

Acid-alcohol is composed of __________

A

Hydrochloric Acid and Ethanol

73
Q

used to color and isolate specific
parts of microorganisms

A

SPECIAL STAINS

74
Q

endospores and flagella, and reveal
the presence of capsules

A

SPECIAL STAINS

75
Q

SPECIAL STAINS

A

CELL WALL STAIN
INDIRECT/NEGATIVE STAINING
NEGATIVE STAINING FOR CAPSULES
CAPSULAR STAINING
ENDOSPORE (SPORE) STAINING
FLAGELLAR STAINING

76
Q

Dyar Method

A

CELL WALL STAIN

77
Q

colorless bacteria against a colored
background

A

INDIRECT/NEGATIVE STAINING

78
Q

excellent technique for studying bacterial
vacuoles and viral morphology

A

INDIRECT/NEGATIVE STAINING

79
Q

Demonstrating the presence of a capsule means of determining the organism’s virulence

A

NEGATIVE STAINING FOR CAPSULES

80
Q

NEGATIVE STAINING FOR CAPSULES APPEARANCE

A

Bacteria as light colored bodies against a
dark background

81
Q

(TRUE OR FALSE)
Cell surface repels acidic stain as a result of
bacterial cells being negatively charged

A

TRUE

82
Q

Example of INDIRECT/NEGATIVE STAINING

A

INDIA INK OR NIGROSIN DYE

83
Q

CAPSULAR STAINING

A

Hiss’s Copper Sulfate Method
Gin’s Method
Anthony’s Method
Welch’s Method
Muir’s Method
Tyler’s Method
Wadsworth’s Method
MacNeal
Lawson

84
Q

capsules appear faint blue
halos around dark blue to purple cells

A

Hiss’s Copper Sulfate Method

85
Q

capsules unstained with margin delineated by
ink; bacteria will be stained

A

Gin’s Method

86
Q

capsule is unstained against a purple
background; cells are deeply stained

A

Anthony’s Method

87
Q

capsules stains a pale violet

A

Welch’s Method

88
Q

cells are stained red and the capsule blue

A

Muir’s Method

89
Q

cannot be stained by ordinary methods
because the dyes do not penetrate the wall of
the endospore

A

ENDOSPORE (SPORE) STAINING

90
Q

most commonly used endospore stain

A

Schaeffer-Fulton Endospore Stain

91
Q

primary stain in ENDOSPORE STAINING

A

MALACHITE GREE

92
Q

helps the stain penetrate the endospore wall

A

Heat steam for about 5 minutes

93
Q

-counterstain
-portions of the cell other than endospores

A

SAFRANIN

94
Q

Appearance in SAFRANIN

A

Endospores appear GREEN
within Red or Pink cells

95
Q

SPORE STAIN METHODS

A

Dorner’s Method
Wirtz and Conklin
Acetic Acid Method

96
Q

spores stain red; bacterial cells almost
colorless against a dark gray background

A

Dorner’s Method

97
Q

spores are green; bodies of bacteria
are red

A

Wirtz and Conklin

98
Q

tedious and delicate staining procedure

A

FLAGELLAR STAINING

99
Q

uses a mordant and the stain CARBOLFUCHSIN
to build up the diameters of the flagella until
they become visible under the light microscope

A

FLAGELLAR STAINING

100
Q

FLAGELLAR STAINING uses a mordant and the stain _____
to build up the diameters of the flagella until
they become visible under the light microscope

A

CARBOLFUCHSIN

101
Q

(FLAGELLAR STAINING)
Treating the cells with an unstable colloidal suspension of _____ cause a heavy precipitate to form on the cell walls and flagella

A

TANNIC ACID SALTS

102
Q

(FLAGELLAR STAINING)
Diameter of flagella is increased to such an extent that subsequent staining with ______ makes the flagella visible in the light microscope

A

BASIC FUCHSIN

103
Q

FLAGELLAR STAIN

A

Leifson Method
Gray’s Method
Fischer and Conn
Casares-Gil’s
Loefflers
Van Ermengen’s

104
Q

bacterial bodies blue; flagella red

A

Leifson Method

105
Q

Mordant -> ______ : swells, coats and forms
precipitate with the flagella

A

TANNIC ACID

BACTERIOLOGY - LAB (6 decks)

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