Question 1

For clinical decision-making carbapenemase production should be detected using:
A)MALDI-TOF mass spectrometry.
B)Modified Hodge test or Carba NP test.
C)molecular methods for specific carbapenemases.
D)routine AST methods using current CLSI interpretive criteria.

Accepted Answer

Current CLSI interpretive criteria are sufficient to detect carbapenem resistance to the carbapenem agents in clinical use at this time.For epidemiologic or research studies or if interpretive guidelines from 2010 or prior are used then Modified Hodge test or Carba NP tests,MALDI-TOF,or molecular methods can be used to confirm the presence of carbapenemase activity or specific carbapenemase genes.

Question 2

Which antimicrobial susceptibility testing method uses a series of plates with different antimicrobial concentrations inoculated with multiple bacterial isolates per plate?
A)Agar dilution
B)Automated broth dilution
C)Disk Diffusion
D)E-test

Accepted Answer

In the agar dilution method,the antimicrobial agent is contained within the agar medium.Each Mueller-Hinton agar plates with a specific concentration of antimicrobial agent representing series of twofold dilutions of antimicrobial to be tested are poured.Each plate is inoculated with 1× 10⁴ CFU/mL of bacterial isolates from multiple patients as well as a spot for positive and negative controls.After overnight incubation,the MIC is determined by identifying the plate containing the lowest concentration of antimicrobial where no growth is observed for each patient's bacterial isolate.

Question 3

A sputum culture from a cystic fibrosis patient grew Stenotrophomonas maltophila.A broth microdilution panel was inoculated.Antimicrobials tested included: ceftazidime,imipenem,ticarcillin-clavulanate,levofloxacin,and trimethoprim-sulfamethoxazole.After 20-hour incubation at 35°C in ambient air,no growth was seen in any well of the microtiter plate.Which of the following statements is true?
A)The isolate is susceptible to all antimicrobials tested.
B)The microdilution plate was incubated under incorrect conditions.
C)The test results are invalid and retesting would be recommended.
D)A Cefinase test for β-lactamase activity should be done as an adjunct to MIC results.

Accepted Answer

Since there was no growth in the positive (i.e. ,growth control well)the test is invalid and would indicate that the organism did not grow or was not inoculated.The test is invalid and must be repeated.Furthermore,this organism is known to be highly resistant to many antimicrobials and have intrinsic resistance to imipenem.

Question 4

A newly defined interpretive category termed "Susceptible-Dose Dependent" (SDD)implies susceptibility of an isolate is dependent on dosing regimen.Which of the following statements is correct?
A)SDD is included within the susceptible category definition.
B)SDD is assigned to therapeutic agents that are approved under investigational use only.
C)The infecting organism may be effectively treated by using higher doses or more frequent doses or both of the drugs.
D)Drugs in this category have narrow therapeutic indices and their use at nonconventional ways must be monitored closely.

Accepted Answer

"Susceptible-Dose Dependent" (SDD)is included within the susceptible category definition.SDD implies susceptibility of an isolate is dependent on dosing regimen and by altering dosing (higher or more frequent doses or both)results in higher drug exposure than the dose used to establish the susceptible breakpoint for AST guidelines.SDD is assigned when doses well above those used to calculate the susceptible breakpoint are approved and used clinically,and where sufficient data to justify the designation exist and have been reviewed.

Question 5

Antimicrobial susceptibility testing should be performed when
A)a single bacterial species grows from any body site.
B)an isolate has a predictable susceptibility profile to therapeutic drugs of choice.
C)reliable standardized methods for testing a clinically significant isolate exist.
D)administering intravenous or oral antibiotics to treat an infection.

Accepted Answer

AST should be done on clinically significant isolates only.The decision as to what defines clinical significant may require consultation between microbiology staff,medical director,supervisor,infectious disease specialist,and attending physician in charge of the patient's care.Also,for isolates that are significant but there are not standardized methods for testing,either information from the literature should be used to guide treatment or testing can be done with a disclaimer that results must be viewed with caution.

Question 6

A D-test is set up on a S.aureus isolate.There was no zone around the Erythromycin disk and the zone of inhibition around the clindamycin disk adjacent to the erythromycin disk was interpreted as susceptible.Thus,the resistance mechanism for the erythromycin is due to:
A)altered target.
B)increased efflux.
C)decreased porin channels.
D)degradative enzymes.

Accepted Answer

This describes a negative D zone test.Thus,the erythromycin-R is due to msrA gene,which results in increased efflux of erythromycin from the bacterial cell and does not induce clindamycin (macrolide-lincosamide-streptogramin B)resistance mediated by products of the erm genes (i.e. ,erthryomycin ribosomal methylase)which alters the MLS antibiotic target.

Question 7

Antimicrobial susceptibility testing should be performed when:
A)a single bacterial species grows from any body site.
B)an isolate has a predictable susceptibility profile to therapeutic drugs of choice.
C)reliable standardized methods for testing a clinically significant isolate exist.
D)administering intravenous or oral antibiotics to treat an infection.

Accepted Answer

AST should be done on clinically significant isolates only.The decision as to what defines clinical significant may require consultation between microbiology staff,medical director,supervisor,infectious disease specialist,and attending physician in charge of the patient's care.Also,for isolates that are significant but there are not standardized methods for testing,either information from the literature should be used to guide treatment or testing can be done with a disclaimer that results must be viewed with caution.Note: Reporting susceptibility results for organisms with questionable clinical significance may be incorrectly interpreted by the clinician.

Question 8

Advantages of automated susceptibility tests (e.g. ,Vitek 2 and Microscan Walkaway)include but are not limited to:
A)automated preparation of inoculum.
B)resistance detection by molecular methods (e.g. ,real-time PCR).
C)advanced (e.g. ,expert)software that recognizes and reports resistance patterns.
D)testing of most aerobic and commonly isolated anaerobic bacterial species.

Accepted Answer

Sophisticated software that incorporates algorithms and review of resistance patterns that can be programmed and customized to each clinical microbiology laboratory is a distinct advantage of automated systems that helps in review of microbiology reports and detection of errors or emergence of resistant isolates or abnormal results before release to physicians.This allows troubleshooting,retesting,and confirmation of results,and notification of infection control and pharmacy personnel as needed.

Question 9

A staphylococcal isolate that gives a negative result using the Cefinase disk and tests resistant to penicillin by disk diffusion means that:
A)the isolate may contain the mecA gene.
B)the bacteria produce the b-lactamase enzyme.
C)the isolate is also resistant to ampicillin,carbenicillin,ticarcillin,and imipenem.
D)the isolate is also resistant to oxacillin,cefoxitin,and ticarcillin-clavulanate.

Accepted Answer

The negative Cefinase test indicates that the bacterium does not produce the b-lactamase enzyme;the organism be expressing its resistance to Penicillin due to the presence of the mecA gene product,an altered Penicillin-Binding Protein (i.e. ,PBP2a).

Question 10

Data review process for AST profiles:
A)requires susceptibility testing results only.
B)is the responsibility of a single individual in the laboratory.
C)can be done on "as needed" basis,no less frequently than weekly.
D)allows for troubleshooting of unusual profiles and results to be held until resolved.

Accepted Answer

Data review process for AST profiles should be done daily and identification and susceptibility test results should be read early in the day so as not to delay workflow and results reporting.Technologists need to read identification and susceptibility reports simultaneously to judge if the AST profile is appropriate or not.If not,then troubleshooting should begin and results not released at this point in time.Workflow should allow time for corrective action for errors found during data review so corrected,or substantiated,results can be provided to physicians as soon as possible.Supervisor or medical director performs second tier review to take ultimate responsibility for the accuracy of results,track and monitor the efficiency of the first tier,provide constructive and educational feedback to the technologists,and provide guidance for resolution of the unusual profiles.Computer-based review process that searches all reports for predefined unusual profiles to enhance the efficiency and accuracy of the second level of review.The review process must be flexible and continuously updated and the list of unusual profiles should be reviewed and updated frequently.

Question 11

What action should be taken for any isolate found to be nonsusceptible (NS)to a particular antimicrobial agent?
A)No additional action is needed,as these results can occur with any organism.
B)Organism's identification and susceptibility test results should be confirmed.
C)The patient report should indicate the isolate is resistant to the antimicrobial.
D)Patient's physician should be contacted and advised that effectiveness of this antimicrobial against the isolate cannot be judged by in vitro testing.

Accepted Answer

Used for isolates for which only susceptible interpretive criterion has been established because the absence or rare occurrences of resistant strains;antimicrobials agents with MICs above or zone diameters are below the susceptible breakpoint should be reported as NS.Note: Confirmation of the organism's identification and susceptibility test results should be performed.

Question 12

Standardization of antimicrobial susceptibility testing optimizes bacterial growth so inhibition of growth can be attributed to the antimicrobial agents tested and results are accurate,reproducible,and consistent regardless of the microbiology laboratory performing the test.Which of the following incubation conditions are used for common (i.e. ,nonfastidious)bacterial isolates using manual techniques?
A)16 to 20 hours,ambient temperature,5% to 7% CO₂
B)16 to 20 hours,35°C,ambient air
C)20 to 24 hours,ambient temperature,5% to 7% CO₂
D)20 to 24 hours,35°C,ambient air

Accepted Answer

Incubation conditions for antimicrobial susceptibility testing (AST)of rapidly growing organisms frequently isolated from clinical samples and known pathogens of human infections include duration of 16 to 20 hours,approximate body temperature of 35°C,and room air.Some more fastidious organisms may require longer incubation times and CO₂ enriched atmosphere,as well supplements added to media used for testing.

Question 13

MIC endpoints can be reported results obtained by the following conventional AST methods EXCEPT: A)agar dilution. B)automated broth dilution. C)disk Diffusion. D)E-test.

Accepted Answer

Quantitative MIC results (in ug/mL)are available by agar dilution,microdilution,macrodilution,and E-test methods.However,only qualitative results are obtained from disk diffusion methods.Zones of inhibition measured in mm are correlated with interpretive categories that are reported as susceptible,intermediate,or resistant.These categories are used to predict whether the bacterial isolate will likely respond (susceptible)or will not respond (resistant)to common dose of the antimicrobial tested.

Question 14

Assaying bacterial isolates for resistance genes by molecular methods:
A)can detect unknown resistance genes not yet defined.
B)cannot detect phenotypic resistance due to a combination of mechanisms.
C)means the resistance gene is expressed if found to be present.
D)is currently part of the routine clinical microbiology workflow for all isolates.

Accepted Answer

Molecular methods for resistance genes can only detect those resistance genes that have been characterized and identified,it does not detect novel resistance genes.Also,just because a resistance gene is present in the bacteria it does not mean that it is expressed.Moreover,if resistance is manifested due to a number of different mechanisms (e.g. ,decreased uptake and altered antimicrobial target)and not a single mechanism associated with a single gene,that resistance would be missed.

Question 15

S.pneumoniae was isolated from a spinal fluid sample from an inpatient.An oxacillin disk screening test was performed and the zone of inhibition measured 19 mm.Which action is most appropriate?
A)Call patient's physician and report the isolate as oxacillin-resistant;no further work-up required.
B)Repeat testing to confirm the identification of the isolate and qualitative susceptibility profile.
C)Report the isolate as penicillin-resistant;inform infection control and patient's physician.
D)Do E-test and report quantitative MIC result and interpretive category using meningitis MIC criteria.

Accepted Answer

An oxacillin screening test for penicillin-susceptibility can be done on S.pneumoniae isolates.For zone sizes less than 20 isolates may upon testing by MIC methods found to be susceptible,intermediate,or resistant to penicillin.In this case,the MIC quantitative result (in ug/mL)be reported along with interpretive category specific for meningitis isolates.Nonmeningitis criteria should not be reported.Penicillin resistance is not that unusual and meningitis caused by S.pneumoniae is not thought to be contagious and thus,the other actions in answers a-c are incorrect or not warranted.

Question 16

Quality control results for disk diffusion AST using appropriate ATCC strains of bacteria showed zone size diameters decreasing over time for a few antimicrobials.Which of the following statements is a possible explanation? A)Antimicrobial disks were stored in a frost-free freezer. B)Concentration of the bacterial inoculum was 5× 10⁵ CFU/mL. C)Depth of Mueller-Hinton agar plates measured 3 mm. D)Plates were incubated less than 16 hours in ambient air.

Accepted Answer

Decreasing zone sizes would indicate that the bacteria are becoming resistant to the antimicrobials tested.However,it can be that antimicrobial disks could lose their potency,which is likely if they are stored in a frost-free freezer that goes through cycles of freezing and thawing.Answers b-d would all be reasons why zone sizes would be increased in size.Accurate and reproducible AST results depend on standardized conditions for inoculum size,depth of agar,and duration of incubation.

Question 17

AST was performed using Mueller-Hinton agar with 5% sheep blood,35°C in 5% CO₂ from 24 hours according to standard protocols,what is the most likely identity of the organism being tested?
A)S.aureus
B)Streptococcus pneumoniae
C)Neisseria gonorrhoeae
D)Moraxella catarrhalis

Accepted Answer

Standardized conditions for AST by disk diffusion of Streptococcus spp.require M-H media supplemented with 5% sheep blood and enriched atmosphere for 20 to 24 hours.S.aureus does not require specialized media (although 2% NaCl can be incorporate to aid in the detection of oxacillin-R).Routine susceptibility testing is not usually done on N.gonorrhoeae or M.catarrhalis.If AST is done on N.gonorhoeae,then growth supplements are added and incubated in 5% CO₂.

Quiz 11: Laboratory Methods and Strategies for Antimicrobial Susceptibility Testing

For clinical decision-making carbapenemase production should be detected using:

Which antimicrobial susceptibility testing method uses a series of plates with different antimicrobial concentrations inoculated with multiple bacterial isolates per plate?

A newly defined interpretive category termed "Susceptible-Dose Dependent" (SDD) implies susceptibility of an isolate is dependent on dosing regimen.Which of the following statements is correct?

Antimicrobial susceptibility testing should be performed when

A D-test is set up on a S.aureus isolate.There was no zone around the Erythromycin disk and the zone of inhibition around the clindamycin disk adjacent to the erythromycin disk was interpreted as susceptible.Thus,the resistance mechanism for the erythromycin is due to:

Antimicrobial susceptibility testing should be performed when:

Advantages of automated susceptibility tests (e.g. ,Vitek 2 and Microscan Walkaway) include but are not limited to:

A staphylococcal isolate that gives a negative result using the Cefinase disk and tests resistant to penicillin by disk diffusion means that:

Data review process for AST profiles:

What action should be taken for any isolate found to be nonsusceptible (NS) to a particular antimicrobial agent?

MIC endpoints can be reported results obtained by the following conventional AST methods EXCEPT:

Assaying bacterial isolates for resistance genes by molecular methods:

S.pneumoniae was isolated from a spinal fluid sample from an inpatient.An oxacillin disk screening test was performed and the zone of inhibition measured 19 mm.Which action is most appropriate?

Quality control results for disk diffusion AST using appropriate ATCC strains of bacteria showed zone size diameters decreasing over time for a few antimicrobials.Which of the following statements is a possible explanation?

AST was performed using Mueller-Hinton agar with 5% sheep blood,35°C in 5% CO₂ from 24 hours according to standard protocols,what is the most likely identity of the organism being tested?