Antimicrobial Susceptibility Testing Flashcards Preview

Liz's Microbiology Module 2 > Antimicrobial Susceptibility Testing > Flashcards

Flashcards in Antimicrobial Susceptibility Testing Deck (173):
1

What three classes of antimicrobials fall under the category of beta-lactams?

- Pencillins
- Cephalosporins
- Carbapenems

2

What antimicrobial class is penicillin in?

Narrow spectrum penicillins

3

What antimicrobial class is oxacillin in?

Penicillins (beta-lactamase resistant)

4

What antimicrobial class is piperacillin in?

Broad-spectrum penicillin (ureidopenicillins)

5

What antimicrobial class is cefazolin in?

Cephalosporins (1st generation)

6

What antimicrobial class is cefuroxime in?

Cephalosporins (2nd generation)

7

What antimicrobial class is ceftazidime in?

Cephalosporins (3rd generation)

8

What antimicrobial class is cefepime in?

Cephalosporins (4th generation)

9

What antimicrobial class is imipenem in?

Carbapenems (beta-lactams)

10

What antimicrobial class is meropenem in?

Carbapenems (beta-lactams)

11

What antimicrobial class is doripenem in?

Carbapenems (beta-lactams)

12

What antimicrobial class is vancomycin in?

Glycopeptides

13

What antimicrobial class is ciprofloxacin in?

Floroquinolones

14

What antimicrobial class is levofloxacin in?

Fluoroquinolones

15

What antimicrobial class is gentamicin in?

Aminoglycosides

16

What antimicrobial class is tobramycin in?

Aminoglycosides

17

What antimicrobial class is is amikacin in?

Aminoglycosides

18

What antimicrobial class is tetracycline in?

Tetracyclines

19

What antimicrobial class is doxycycline in?

Tetracyclines

20

What antimicrobial class is clindamycin in?

Under its own category

21

What antimicrobial class is is erythromycin in?

Macrolides

22

What antimicrobial class is trimethoprim-sulfamethoxazole (Bacitrim) in?

Antimetabolites

23

Penicillin
- Spectrum of activity
- Mechanism of action

- Against GP and GN (changes depending upon class)
- Inhibits cell wall synthesis

24

Cephalosporins (4th generation)
- Spectrum of activity
- Mechanism of action

- GNRs
- Inhibits peptidoglycan

25

Tetracyclines
- Spectrum of activity
- Mechanism of action

- Broad spectrum: GP, GN, mycoplasma, chlamydiae, rickettsiae
- Inhibits protein synthesis

26

Clindamycin
- Spectrum of activity
- Mechanism of action

- Broad spectrum (aerobic GP + anaerobes)
- Inhibits protein synthesis

27

Fluoroquinolones
- Spectrum of activity
- Mechanism of action

- Broad spectrum: GP, GN
- Inhibits DNA synthesis

28

Cephalosporins (2nd generation)
- Spectrum of activity
- Mechanism of action

- GPC, some GPR
- Inhibits peptidoglycan synthesis

29

Antimetabolites
- Spectrum of activity
- Mechanism of action

- Specific clinical uses...UTI, S. maltophilia
- Inhibits folic acid synthesis

30

Macrolides
- Spectrum of activity
- Mechanism of action

- Broad spectrum
- Inhibits protein synthesis

31

Cephalosporins (3rd generation)
- Spectrum of activity
- Mechanism of action

- GNR, some GPC
- Inhibits peptidoglycan synthesis

32

Aminoglycosides
- Spectrum of activity
- Mechanism of action

- Active against GNRs and S. aureus
- Inhibits protein synthesis

33

Cephalosporins (1st generation)
- Spectrum of activity
- Mechanism of action

- GPC
- Inhibits peptidoglycan synthesis

34

Glycopeptides
- Spectrum of activity
- Mechanism of action

- GP only
- Inhibits peptidoglycan synthesis

35

How many interactions occur b/w a patient, an organism, and an antimicrobial?

Six

36

Substance naturally produced by living organisms such as bacteria and fungi and able, in a dilute solution, to inhibit or kill another microorganism

Antibiotic

37

Chemical substance produced by a microorganism that has the capability of killing of inhibiting the growth of another organism

Antimicrobial agent

38

Range of activity of an antimicrobial agent against certain groups of bacteria

Spectrum of activity

39

Difference b/w intrinsic and acquired resistance

- Intrinsic: all members of the species are resistant
- Acquired: not all members of the species are resistant

40

Examples of intrinsic resistance

Staphylococcus saprophyticus and novobiocin

41

Examples of acquired resistance

Staphylococcus aureus and methicillin/oxacillin

42

Interpret susceptibility/resistance to oxacillin and cefoxitin as MSSA

MSSA is susceptible to oxacillin and cefoxitin

43

Interpret susceptibility/resistance to oxacillin and cefoxitin as MRSA

MRSA is resistant to oxacillin and cefoxitin

44

Three mechanisms used by bacteria in order to exchange genetic material resulting in antimicrobial resistance

- Genes
- Transfer
- Expression

45

Three mechanisms of bacterial resistance

- Impermeability
- Alterations in target molecules
- Enzymatic inactivation

46

Two mechanisms of impermeability

- Altered outer membrane porins
- Altered transport systems

47

Three mechanisms in altering the target molecules

- Methylation of rRNA
- Alterations of ribosomes
- Altered penicillin binding proteins

48

Three mechanisms in enzymatic inactivation

- Beta-lactamases
- Chloramphenicol acetyltransferases
- Aminoglycosides modifying enzymes

49

Three effects of combining antimicrobials

- Autonomous/indifferent
- Antagonistic (a substance that interferes w/ or inhibits the physiological action of another)
- Synergistic (one drug increases the other's effectiveness)

50

What organism is universally susceptible to penicillin?

Streptococcus pyogenes

51

Standardizing susceptibility testing
- Growth medium

Mueller-Hinton Agar or broth
- Depth 3-5mm

52

Standardizing susceptibility testing
- pH

7.2-7.4

53

Standardizing susceptibility testing
- Cation concentration

- Mg2+
- Ca2+
- NaCl

54

Standardizing susceptibility testing
- Incubation conditions

35°C, ambient air, duration varies

55

Standardizing susceptibility testing
- Inoculum density

If not enough organism then not enough enzyme which could lead to a false susceptibility

56

Standardizing susceptibility testing
- Inoculum prep

3-5 colonies made into a McFarland standard

57

In a disk diffusion test (or an E-test), the agar depth is 1mm, what will the result be?

The antibiotic diffuses farther due to less agar → false susceptibility

58

In a disk diffusion test (or an E-test), the agar depth is 6 mm, what will the result be?

The antibiotic cannot diffuse as far → false resistance

59

In a disk diffusion test (or an E-test), a 0.25 McFarland is used, what will the result be?

There is less bacteria in the inoculum that is spread over the plate → Larger zone → false susceptibility

60

In a disk diffusion test (or an E-test), a 2.0 McFarland is used, what will the result be?

There is more bacteria in the inoculum that is spread over the plate → smaller zone → false resistance

61

In a broth dilution, a 0.25 McFarland is used, what will the result be?

There is less bacteria in the inoculum → ↓ MIC

62

In a broth dilution, a 2.0 McFarland is used, what will the result be?

There is more bacteria in the inoculum → ↑ MIC

63

Disk diffusion test
- What is the McFarland standard?

Uses barium sulfate to make a 1.5x10^8 CFU/mL standard which is a 0.5 McFarland

64

Disk diffusion test
- Inoculation of agar

W/in 15 minutes of prep, streak plates in 3 planes so that there is a lawn of confluent growth

65

Disk diffusion test
- Application of disks

W/in 15 minutes of inoculation, NEVER relocate a disk

66

Disk diffusion test
- Incubation

- 16-18 hours (Staph and oxacillin/methicillin for 24 hours)
- 35°C ambient air

67

Disk diffusion test
- Reading results

Read circular zone side in mm;
- If it's resistant, zone size is still 6mm! NEVER 0!

68

Disk diffusion test
- Qualitative or quantitative?
- Interpretation of results

Qualitative → inform MD if its S, I, or R using CLSI charts

69

E-test
- What is the McFarland standard?

Uses barium sulfate to make a 1.5x10^8 CFU/mL standard which is a 0.5 McFarland

70

E-test
- Inoculation of agar

W/in 15 minutes of prep, streak plates in 3 planes so that there is a lawn of confluent growth

71

E-test
- Application of gradient strips

?

72

E-test
- Incubation

- 16-18 hours
- 35°C ambient air

73

E-test
- Reading results

Read elliptical zone side in mcg/mL

74

E-test
- MIC

Where growth intersects strip
- No zone: MIC > highest concentration
- Zone below strip: MIC < lowest concentration

75

E-test
- Qualitative or quantitative?
- Interpretation of results

Quantitative → informing MD of concentration (mcg/mL) and if its S, I, or R using CLSI tables

76

Broth dilution
- Inoculum prep

3.5 colonies and make a McFarland standard

77

Broth dilution
- What is the McFarland standard?

Use barium sulfate to make a 1.5x10^8 CFU/mL standard which is a 0.5 McFarland
- Final concentration: 5x10^5 CFU/mL

78

Broth dilution Macrotube
- Inoculation procedure

??

79

Broth dilution microtiter tray
- Inoculation procedure

- Deliver inoculum, via delivery prongs, into sterility tray with sterility well (neg) and growth control well (pos)
- Make a PURITY PLATE!

80

Broth dilution (Macrotube and microtiter tray)
- Incubation

35°C ambient air, 16-20 hours

81

Broth dilution
- Reading results

MIC is the lowest concentration that inhibits growth
- Needs to be reproducible w/ one well margin of error

82

Broth dilution
- Qualitative or quantitative

Quantitative → informing MD of concentration (mcg/mL) and if it's S, I, or R using CLSI tables

83

What is the significance of a regressive curve?

Linear and inversely proportional
- As zone size ↑, MIC ↓

84

Regressive curve
- X axis
- Y axis

- Y axis: MIC (mcg/mL)
- X axis: Zone size (mm)
- Top left = resistant
- Bottom right = susceptible

85

MIC/MBC procedure

Take clear tubes from MIC and incubate again on a plate. If < 50 colonies can grow on a plate thats considered killed!

86

Minimal bactericidal concentration

Killing capacity of 99.9% of original inoculum

87

MID/MBD procedure

?

88

MID procedure

Patient's serum is serially diluted and a standard inoculum (5x10^5 CFU/ml) of patient organism is added. Incubate and read highest dilution which inhibits growth.

89

MBD procedure

- Highest dilution of patient serum that kills 99.9% of an original inoculum of patients organism
- Must be performed w/ MID
- Results are dilutions 1:8 to 1:32 is adequate

90

SXT is read at what % inhibition?

80%

91

Automated methods: Dade Microscan
- Three methods

- Traditional MIC methods
- Photometric reading (turbidity)
- Fluorometric reading (best method) (degradation of substrates by viable bacteria; detects bacterial inhibition by antibiotics)

92

Automated methods: BD Phoenix
- Three methods

- Gravity based inoculation process
- Redox indicator system
- Data management system

93

Automated methods: Vitek
- Three methods

- Computer-assisted analysis of growth
- Algorithm derived MIC
- Calculated MIC

94

Three methods of beta-lactamase detection

- Chromogenic cephalosporin: cefinase, nitrocefin
- Acidometric
- Iodometric

95

Results of beta-lactamase test

- Positive = organism produces enzyme to degrade penicilin, ampicillin, and amoxicillin
- Negative = does not produce enzyme but it may have some other mechanism of resistance to the same drug

96

What organism requires beta-lactamase testing?

Haemophilus influenzae

97

Which antibiotics which will not be effective against beta-lactamase producers

- Penicillin, ampicillin, amoxicillin
- Cephalosporins
- Carbapenems

98

What is the value of measuring serum levels?

Want to know how much antibiotic is at site of infection so that therapeutic range can be determined

99

Agar dilution
- Prep

Antibiotic is diluted into agar in 2-fold concentrations

100

Agar dilution is used for what type of organisms?

Fastidious organisms (N. gonorrhoeae, N. meningitidis)

101

Agar dilution, steers replicator
- What is the standard inoculum per spot?

10^4 CFU/mL

102

Agar dilution, steers replicator
- Incubation requirements

35°C, ambient air, 18-20 hours

103

What is the reference method for susceptibility testing?

Agar dilution

104

Five mechanisms of action a drug can have

- Inhibit cell wall synthesis
- Inhibit protein synthesis
- Inhibit nucleic acid synthesis
- Antimetabolites
- Alteration of cell membranes

105

What drugs work by inhibiting cell wall synthesis?

- Penicillins
- Cephalosporins
- Glycopeptides (inhibitis peptidoglycan synthesis; precursors of cell wall synthesis)

106

What drugs work by inhibiting protein synthesis?

- Aminoglycosides
- Tetracyclines
- Macrolides
- Clindamycin

107

What drug work by inhibiting the folic acid pathway?

Antimetabolites

108

What drug works by inhibiting DNA synthesis?

Fluoroquinolones

109

What drug alters cell membranes?

Bactracin

110

Mechanism of resistance
- DRSP

- Altered target sight
- PBP-2 pencillin binding protein

111

Mechanism of resistance
- ESBL

- Arise from point mutations of common beta-lactamases
- TEM-1 and SHV-1 genes coding for enzyme

112

Mechanism of resistance
- VRSA/GRSA

- May involve alterations in the cell wall and hyperexpression of PBP
- vanA gene present

113

Mechanism of resistance
- MRSA

- Alteration of target site (classic and modified)
- Hyperproduction of beta-lactamases

114

Mechanism of resistance
- VRE

- Altered peptidoglycan synthesis
- Carried by transposons or plasmids

115

Mechanism of resistance
- ARHI

Plasmid-mediated beta-lactamases (most common)

116

Mechanism of resistance
- CRE

- Carbapenemase (breaks down antibiotic)
- Cephalosporinase combined with porin loss

117

Lab detection of MRSA
- Cefoxitin disk diffusion

Induces mecA gene (best drug to detect MRSA)
- Media: CAMHB with 2% NaCl
- Incubation: 24 Hours, 35°C, ambient air

118

Lab detection of DRSP
- Disk diffusion w/ oxacillin

Disk Diffusion using Oxacillin;
- Media: Mueller-Hinton Agar, 5% SBA;
- Incubation: 20-24 Hrs, 35°C in CO2;

119

Lab detection of ARHI
- Disk diffusion

- Media: HTM agar
- Incubation: 16-18 Hrs, 35°C in CO2

120

Lab detection of VRE
- Disk diffusion

- Media: Mueller-Hinton Agar;
- Incubation: 24 Hrs, 35'C, Ambient Air;

121

Lab detection of VRSA/GRSA

Disk Diffusion and Broth Dilution tests are held for 24 Hrs;
Agar Screen:
- Media: BHI with 6ug/mL Vancomycin,
- Incubation: 24 Hrs, 35'C, ambient air;
D-Test Disk Induction: Erythromycin and Clindamycin disks are placed 15mm apart and flattening is noted; Pos=Resistant to clindamycin, Neg=susceptible

122

Lab detection of ESBL

Confirmatory Test: Use cefotaxime and ceftazidime disks and each disk combined with clavulanic acid; >5mm increase with clav=pos;
Can also use E-Test Strips, Vitek-2, and Microscan

123

Lab detection of CRE

Modified Hodge Test: 1:10 Dilution of 0.5 Mcfarland (E.coli lawn)(because larger zone size with decreased inoculum), streaked with known + and - and Patient.
- Shouldering occurs if enzyme present to break down meropenam = pos result/resistance

124

MRSA
- Infection types

- HA-MRSA
- CA-MRSA

125

What is HA-MRSA?

- Found in ICUs
- S. aureus has acquired several genes that are responsible for its resistance

126

What is CA-MRSA?

- S. aureus has only gene responsible for its resistance

127

VRE
- Infection types

- Nosocomial bacteremia
- Surgical wound infection
- UTI

128

DRSP
- Infection types

- Otitis media
- Sinusitis
- Community acquired pneumonia
- Bacteremia
- Meningitis

129

ARHI
- Infection types

?

130

VISA/VRSA
- Infection types

?

131

ESBL
- Infection types

- E. coli
- Klebseilla spp
- Proteus mirabilis

132

CRE
- Infection types

- Class A: K. pneumoniae
- Class B: P. aeruginosa, A. baumannii
- Class D: A. baumannii

133

Effective treatment or prevention
- MRSA

- Linezolid
- Synercid
- Daptomycin
- Vancomycin to treat

134

Effective treatment or prevention
- ARHI

- Type b vaccine
- Empiric treatment w/ cefotaxime or ceftriaxone

135

Effective treatment or prevention
- DRSP??

??

136

Effective treatment or prevention
- VRE??

??

137

Effective treatment or prevention
- VRSA/VISA??

??

138

Effective treatment or prevention
- ESBL??

??

139

Effective treatment or prevention
- CREs??

??

140

Synercid treats which problematic bacteria?

- MRSA
- VRE

141

Daptomycin treats which problematic bacteria?

- MRSA
- VRE

142

Telithromycin treats which problematic organisms?

Macrolide-resistant pneumococci

143

What test detects the inducible clindamycin resistance mechanism in GP organisms?

D-test

144

Desribe the prinicple behind the D-test

Erythromycin is used to induce the resistance mechanism (ribosome alteration) to clindamycin w/in the GP organism
- If using the D-test, the GP organism should already be resistant to erythromycin

145

Describe the procedure of the D-test

Erythromycin and clindamycin disks are placed 15mm apart and flattening is noted around the clindamycin disk

146

Interpret the results of the D-test

- Pos: GP organism is RESISTANT to clindamycin (CANNOT USE FOR TREATMENT!)
- Neg: GP organism is SUSCEPTIBLE to clindamycin

147

Rationale for identifying VRE to the species level

To confirm if we truly have VRE (E. casseliflavus or E. gallinarium)

148

Resistance mechanism responsible for vancomycin resistance in S. aureus (VRSA)

Alteration of cell wall

149

Resistance mechanism responsible for penicillin resistance in S. pneumoniae

Alteration of target site

150

What antibiotic disk is used to predict penicillin resistance in S. pneumoniae?

Oxacillin

151

What antibiotic disk is used to predict oxacillin resistance in S. aureus?

Cefoxitin

152

What is the purpose of the confirmatory ESBL test?

To detect the presence of extended spectrum beta-lactamase enzymes produced by an isolate

153

Four disks used in the confirmatory ESBL test

- Cephalosporin (CTX)
- Clavulanic acid (CLA)
- Cephalosporin/clavulanic acid (CTX/CLA)
- Ceftazidime/clavulanic acid (CAZ/CLA)

154

What is the purpose of the cefoxitin screen?

To determine if an isolate produces the mecA gene (differentiates MSSA vs. MRSA)

155

What size zone indicates a "positive" (resistant) screen?

≤ 21mm (MRSA)

156

What size zone indicates a "negative" (susceptible) screen?

≥ 22mm (MSSA)

157

CRE/modified Hodge test
- Purpose

To detect the presence of carbapenemases

158

CRE/modified Hodge test
- What organism is streaked all over the plate?

E. coli

159

CRE/modified Hodge test
What disk is placed in the center of the plate?

Meropenem

160

What does "MRSA" stand for?

Methicillin resistant Staphylococcus aureus

161

What does "DRSP" stand for?

Drug resistant Streptococcus pneumoniae

162

What does "ARHI" stand for?

Ampicillin resistant Haemophilus influenzae

163

What does "VRE" stand for?

Vancomycin resistant Enterococcus spp

164

What does VRSA/VISA" stand for?

Vancomycin resistant Staphylococcus aureus

165

What does "ESBL" stand for?

Extended spectrumm beta-lactamases

166

What does "CRE" stand for?

Carbapenem resistant enterics

167

Lab detection of MRSA
- MRSA agar screen

Meuller-Hinton Agar + 4% NaCl + 6 mcg/mL Oxacillin

168

Lab detection of DRSP
- Broth dilution

- Media CAMHB with 2-5% LHB;
- Incubation: 20-24 Hrs at 35°C, ambient air

169

Lab detection of DRSP
- E-test

- Media: Mueller-Hinton agar, 5% SBA;
- Incubation: 20-24 Hours 35°C in CO2

170

Lab detection of ARHI
- Broth dilution

- Media: HTM broth
- Incubation: 20-24 Hours, 35°C, ambient air

171

Lab detection ARHI
- Nitrocefin assay

Chromogenic cephalosporin
- Pos: isolate is resistant to penicillin, ampicillin, and amoxicillin
- Neg: isolate can be resistant by other mechanisms

172

Lab detection of VRE
- Broth dilution

Broth Dilution:
- Media: CAMHB;
- Incubation: 24 Hrs, 35'C, ambient air; Agar

173

Lab detection
- Screening test

- Media: BHI with 6ug/mL vacomycin;
- Incubation: 24 Hrs, 35'C, Ambient air, any growth = resistance;
Confirmation of VRE with gram stain