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MLS 223/L MIDTERMS > LAB S19 > Flashcards

LAB S19 Flashcards

1
Q

Specimens to be obtained for culture depend on

A

the nature of the

staphylococcal infection

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2
Q

G+ cocci, 0.5-1.5 micrometer, grape-like clusters

A

staphylococcus species

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3
Q

staphylococcal species’ specimens are obtained from

A
  • Pus from wound and other pyogenic infections
  • Blood from septicemia
  • CSF from meningitis
  • Sputum from respiratory infections
  • Urine from UTI
  • Nasal swab from suspected carriers
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4
Q

direct smears of staphylococcus species from pus or CSF usually reveals

A

G+ cocci arranged as

  • single cells
  • pairs
  • tetrads
  • short chains
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5
Q

direct smears of staphylococcus species from pus or CSF usually reveals

A

G+ cocci arranged as

  • single cells
  • pairs
  • tetrads
  • short chains
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6
Q

The direct smears of staphylococcus species from pus or CSF should be examined for the presence of

A

inflammatory cells
-these cells along with background tissue debris and proteinaceous
material generally stain pink to red in properly gram stained specimens

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7
Q

Staphylococcus species grow well on routine laboratory media such as

A
  • Blood Agar Medium (BAM)

- Mannitol Salt Agar (MSA)

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8
Q

This an enriched medium that supports the growth of staphylococci and
permits observation of the pattern of hemolysis of blood

A

Blood Agar Medium (BAM)

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9
Q

A culture media that is both a selective and differential medium for
staphylococci

A

Mannitol Salt Agar (MSA)

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10
Q

Approximate formula / Liter of Mannitol Salt Agar (MSA)

A
  • Pancreatic digest of Casein - 5g
  • Peptic digest of animal tissue - 5g
  • Beef extract - 1g
  • Sodium chloride - 75g
  • D-mannitol - 10g
  • Phenol Red - 25mg
  • Agar - 15g
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11
Q

Uninoculated MSA appears

A

light to medium rose red in color

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12
Q

The ______________ concentration of MSA inhibits the growth of
other organisms (except for enterococci), and selectively allows
the growth of halotolerant staphylococci

A

high salt (7.5% NaCl)

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13
Q

Mannitol fermentation results in acids is indicated by

A

a change in the phenol red indicator to yellow

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14
Q

this aids in the differentiation of staphylococcal species

A

Mannitol fermentation

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15
Q

Interpretation of colonial morphologies of staphylococcus species is usually performed after

A

24 to 48 hours of incubation at 35-37 degrees celsius

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16
Q

size of staphylococcal colonies on Blood Agar Medium (BAM)

A

medium-large (1-2 mm in diameter)

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17
Q

other characteristics of staphylococcal species on BAM

A
  • color: off-white or gray
  • surface: smooth
  • margin: entire
  • elevation: slightly raised, low convex
  • opacity: opaque
  • consistency: butyrous (butterlike)
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18
Q

size of S. aureus colonies on BAM

A

usually large (4.6 mm in diameter)

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19
Q

some strains in S. aureus colonies on BAM produce _________ pigments

A

golden yellow (lipochrome)

  • hence they appear creamy yellow or yellow-orange
  • aureus = meaning “golden”
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20
Q

other S. aureus strains may have _____ or ______ zone of B-hemolysis around the colonies

A

distinct; hazy

-this hemolytic property may become apparent only after prolonged incubation

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21
Q

characteristics of S. epidermidis on BAM

A
  • size: small to medium
  • color: gray-white colonies
  • opacity: opaque
  • most colonies are NONHEMOLYTIC
  • Slime-producing strains are extremely sticky and adhere to the agar surface
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22
Q

characteristics of S. saprophyticus colonies on BAM

A
  • size: large
  • color: usually white but can be yellow or orange
  • surface: smooth; very glossy
  • margin: entire
  • elevation: convex
  • opacity: opaque
  • consistency: butyrous (butterlike)
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23
Q

characteristics of Mannitol-fermenting staphylococci (S. aureus)

A

color: yellow colonies (accompanied by yellow discoloration of the medium) > typical for S. aureus

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24
Q

S. saprophyticus may also ferment mannitol and thus resemble S. epidermidis.

A

False. S. saprophyticus may also ferment mannitol and thus resemble S. aureus on MSA medium

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25
Q

S. epidermidis is also a mannitol fermenter.

A

False. S. epidermidis is non-mannitol fermenter.

-Hence, y S. epidermidis and several other species produce SMALL RED colonies with NO COLOR CHANGE to the medium.

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26
Q

Identification Tests are done in what order

A

-Catalase Test
-Coagulase Test
>Slide Coagulase Test
>Tube Coagulase Test
-Novobiocin Test

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27
Q

When performed with bacterial isolates that are
G+ cocci, this test distinguishes staphylococci
form streptococci

A

Catalase Test
because
-Staphylococci = strongly catalase (+)
-streptococci = catalase (-)

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28
Q

The principle of this identification test is based on the ability of the bacteria to produce the catalase enzyme that decomposes hydrogen peroxide into water and oxygen

A

Catalase Test

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29
Q

The evolution of oxygen in Catalase Test leads to

A

effervescence (rapid bubble formation)

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30
Q

what culture slant is unsuitable for Catalase Test

A

Blood Agar Slant

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31
Q

This test is done on catalase-positive, gram-positive cocci, placing them in the genus Staphylococcus

A

Coagulase Test

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32
Q

the single most reliable characteristic for identifying S.

aureus

A

Coagulase Test

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33
Q

The principle of this identification test relies on the coagulase enzyme produced by S. aureus that binds plasma fibrinogen and activates a cascade of reactions causing plasma to clot

A

Coagulase Test

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34
Q

Two types of Coagulase Test

A
  • Slide coagulase test

- Tube coagulase test

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35
Q

The medium for both slide coagulase and tube coagulase

procedures

A

rabbit plasma with EDTA

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36
Q

Rabbit plasma has high amounts of

A

Coagulase-reacting Factor (CRF)

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37
Q

Preferred anticoagulant of rabbit plasma

A

EDTA

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38
Q

Why is human plasma not recommended as medium for slide coagulase and tube coagulase procedures?

A

because it contains variable
amounts of CRF and may contain anti-staphylococcal
antibodies

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39
Q

Why is citrated plasma not used in coagulase test

A

because some bacteria (enterococci) are able to utilize citrate which could yield to false positive results if they are mistaken for staphylococci
NOTE: ALWAYS PERFORM A CATALASE TEST FIRST

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40
Q

This is a screening test for S. aureus

A

Slide Coagulase Test

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41
Q

Strains that are negative with the

slide coagulase test must be

A

confirmed with a tube coagulase test

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42
Q

This identification test is based on the principle that the abundant presence of bound coagulase or “clumping factor” on the surface
of the cell wall of most strains of S. aureus.

A

Slide coagulase test

43
Q

This factor reacts directly with fibrinogen in plasma and converts it into fibrin, causing rapid cell agglutination

A

bound coagulase or clumping factor

44
Q

reaction between bound coagulase (clumping factor) and fibrinogen in plasma results in

A

formation of fibrin which leads to rapid cell agglutination

45
Q

This indicates a positive result in Slide Coagulase Test

A

Formation of white precipitates or clumping

within 10-15 seconds of mixing the plasma with the bacterial suspension.

46
Q

characteristic of the negative control in slide coagulase test

A

saline suspension remains smooth and milky

47
Q

what are some human coagulase-negative species that produce clumping factor and may be slide coagulase-positive

A

S. lugdunensis and S. schleiferi subsp. schleiferi

48
Q

slide coagulase test can be performed with growth from

A

blood agar or other nonselective medium

49
Q

slide coagulase test should not be performed with growth from

A

media having high salt content (mannitol salts agar)

-because high salt causes some strains of S. aureus to AUTOAGGLUTINATE

50
Q

This is a confirmatory test for S. aureus

A

Tube coagulase test

51
Q

this identification test’s principle is based on the detection of free coagulase which is secreted extracellularly by S. aureus. The free coagulase reacts with a substance in the plasma called coagulase-reacting factor (CRF), which in turn reacts with fibrinogen in plasma to form fibrin (clot formation)

A

tube coagulase test

52
Q

positive result in tube coagulase test

A

clotting of the plasma (because of the formation of fibrin clot)

53
Q

what animal isolates may be tube-coagulase positive

A
  • S. intermedius
  • S. hyicus
  • S. delphini
  • S. schleiferi subsp. coagulans
  • strains of S. schleiferi subsp. schleiferi
54
Q

What to do when you are observing for any degree of clot formation (partial or complete) after incubation

A

-gently tilt the tube
-DO NOT AGITATE as small clot formed may be dislodged and will not
reform anymore

55
Q

What to do to tube coagulase tests that are negative after 4 hours of incubation at 35 degrees celsius

A
  • should be held at room temperature

- read again after 18 to 24 hours

56
Q

why does negative tube coagulase tests need to be read again after 18 - 24 hours (at room temp)

A

because some strains will produce fibrinolysin (staphylokinase) on prolonged incubation at 35°C, causing dissolution of the clot during the incubation period

57
Q

one of the methods to distinguish between coagulase-negative
staphylococci, S. epidermidis, S. saprophyticus

A

Novobiocin Test

58
Q

this agent is involved in several

infectious processes

A

S. epidermidis

59
Q

this agent has a recognized clinical significance in urinary tract infections

A

S. saprophyticus

60
Q

in novobiocin test, a zone of inhibition of <12mm indicates that the agent is

A

novobiocin resistant

-presumptive for S. saprophyticus

61
Q

in novobiocin test, a zone of inhibition of >16mm indicates that the agent is

A

novobiocin-susceptible

-coagulase-negative staphylococci (CoNS), S. aureus

62
Q

Novobiocin test is performed as a

A

disk susceptibility test using a novobiocin disk (NB,5 µg)

63
Q

other than S. saprophyticus, some human staphylococcal species resistant to novobiocin

A
  • S. cohnii subspecies,
  • S. hominis subsp. novobiosepticus
  • S. xylosus
  • some S. pseudolugdunensis
64
Q

Additional culture media for isolation of Staphylococcus species

A
  • Columbia CNA Agar
  • Phenylethyl Alcohol (PEA) Agar
  • Vogel-Johnson (VJ) Agar
  • Chapman Stone Agar
  • Baird-Parker Agar
65
Q

This blood supports the growth of fastidious bacteria and allows
detection of hemolytic reactions in Columbia CNA Agar

A

Sheep blood

66
Q

what causes Columbia CNA Agar to be selective for G+ bacteria by inhibiting G-bacteria

A

because it contains the antimicrobial agents: COLISTIN and NALIDIXIC ACID

67
Q

sheep blood in phenylethyl alcohol supports the growth of fastidious bacteria particularly

A

G+ cocci.

68
Q

The medium should not be used for determination of hemolytic
reactions since atypical reactions may be observed

A

phenylethyl alcohol (PEA) agar

69
Q

Phenylethly Alcohol (PEA) Agar inhibits

A

G- bacteria

70
Q

selectivity of Vogel-Johnson (VJ) Agar is achieved by

A

-potassium tellurite
-lithium
-chloride
-glycine
which inhibit non-staphylococcal organisms, both G+ and G- bacteria

71
Q

Tellurite is reduced to metallic tellurium by S. aureus producing
colonies that are

A

black or gray-black in color

72
Q

in vogel-johnson (VJ) Agar, mannitol is degraded

into acid which is indicated by _____________ which changes the color of medium to yellow

A

phenol red

73
Q

the selectivity of Chapman Stone Agar is due to

A

relatively high salt content

74
Q

this serves as a susbtrate for gelatinase activity in chapman stone agar

A

gelatin

75
Q

this allows detection of gelatin hydrolysis in chapman agar which is characterized by clear zone around the colonies

A

ammonium sulfate

76
Q

chapman stone agar is differential due to

A

mannitol fermentation

77
Q

how to determine mannitol on chapman stone agar

A

add few drops of bromcresol purple to areas on the medium

-yellow color indicates mannitol fermentation

78
Q

in chapman stone agar, what identifies S. aureus

A

yellow colonies surrounded by clear halo

79
Q

in chapman stone agar, what identifies S. epidermidis

A

white nonpigmented colonies with or without clear zone

80
Q

in baird-parker agar, these have inhibitory action for organisms other than staphylococci

A
  • tellurite
  • lithium chloride
  • glycine
81
Q

in baird-parker agar, this is incorporated to stimulate the growth of S. aureus without destroying the selectivity

A

Sodium pyruvate

82
Q

reduction of tellurite to _______ produces a black discoloration

A

tellurium

83
Q

Addition of egg yolk is used to demonstrate ____________ producing clear zones around the colonies.

A
  • proteolysis by lecithinase

- lipolysis by lipase

84
Q

in baird-parker agar, these are black, shiny, convex colonies 1-5 mm in diameter with a narrow white edge

A

S. aureus

85
Q

in baird-parker agar, these are black, shiny, irregular shaped colonies with the development of opaque zone around the colonies after 24 hrs

A

S. epidermidis

86
Q

S. aureus produces ________ and ___________ .

A

DNase; thermostable nuclease having endo- and exonucleolytic activities

87
Q

what are the additional identification tests for staphylococcus aureus

A
  • deoxyribonuclease (DNase) test

- Thermostable endonuclease test

88
Q

this identification test is based the detection of bacterial deoxyribonuclease

A

deoxyribonuclease test

89
Q

this is an extracellular enzyme that depolymerizes (breaks down) DNA into
subunits composed of nucleotides.

A

deoxyribonuclease

90
Q

method of inoculation in deoxyribonuclease test could either be

A
  • streak inoculation

- spot inoculation

91
Q

a DNase test agar plate (containing deoxy-ribonucleic acid) must be incubated at _______ for ___________

A

35C for 24-48 hrs

92
Q

after the incubation of a DNase agar plate flood the plate with _________ which will precipitate DNA

A

1N hydrochloric acid (HCl)

93
Q

what is the indication of a positive result in DNase test

A

A clear area surrounding the
growth after addition of 1N
HCL

94
Q

what is the indication of a negative result in DNase test

A

no clearing
and a cloudy precipitate
around the colonies

95
Q

what is the alternative method for DNase test without the use of 1N HCl

A

usage of agar plates with dye

  • methyl green
  • toluidine blue
96
Q

What causes the formation of color green medium when methyl green is used

A

Methyl green forms a complex with intact polymerized DNA to form green
color of the medium

97
Q

what is the indication of a positive result in a DNase test using methyl green

A

DNAse activity depolymerizes the DNA, breaking
down the DNA-methyl green complex, which results in the formation of
colorless zones around the colonies .

98
Q

what is the indication of a positive result in a DNase test using toluidine blue

A

DNAse activity depolymerizes the DNA, breaking
down the DNA-toluidine blue complex, and the toluidine blue takes
on its metachromatic color, forming pink to RED ZONES around the
bacterial growth.

99
Q

what is the indication of a negative result in a DNase test using toluidine blue

A

A negative test is indicated when the medium remains

blue because the intact DNA-toluidine blue complex, the toluidine blue has a normal blue color

100
Q

this test also use the same DNase test agar

A

thermostable endonuclease test

101
Q

procedures involved in thermostable endonuclease test

A

-3-mm holes are cut into the agar with a sterile cork borer
-the wells are filled with a 24-hour broth culture
>from the test organism that has been boiled in a water bath (15 mins)
- The plate is incubated overnight at 35°C.

102
Q

what staphylococcal specie is thermostable endonuclease-positive

A

S. aureus

103
Q

what is the indication of a positive result in thermostable endonuclease test (S. aureus)

A

will show a pink zone surrounding the

well containing the boiled suspension

104
Q

what is the indication of a negative result in thermostable endonuclease test

A

when the blue color in the

medium remains

MLS 223/L MIDTERMS (5 decks)

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