30 Questions
What is the primary purpose of performing antibiotic susceptibility tests?
To determine which antimicrobial agents might be effective in treating infections
Which of the following is NOT a factor that is standardized and controlled in antimicrobial susceptibility testing?
Type of bacteria being tested
What organization publishes guidelines and recommendations for antimicrobial susceptibility testing?
Clinical and Laboratory Standards Institute (CLSI)
What is the name of the system that uses a microtiter plate with a specific concentration of antimicrobial agents?
Alamar System
What is the purpose of controlling the bacterial inoculum size in antimicrobial susceptibility testing?
To ensure accurate results
What type of methods use instruments to measure the growth of bacteria in the presence of antimicrobial agents?
Automated Methods
What is the primary difference between the macrobroth dilution method and the microbroth dilution method?
The volume of broth and inoculum used
What is the purpose of the 'alamar blue' indicator in the Alamar system?
To indicate the absence of bacterial growth
What is the primary step in the agar dilution method after adding the inoculum to the MHA?
Allowing the MHA to solidify
What is the term used to describe the area where bacterial growth is inhibited in the diffusion method?
Zone of inhibition
What is the primary difference between the E-test and the agar dilution method?
The presence of a plastic strip
What is the primary purpose of the Vitek system?
To automate the antimicrobial susceptibility testing
What is the primary purpose of incubating the inoculated surface with disks in disk diffusion testing?
To allow the antimicrobial agent to diffuse into the agar
What is the interpretation of a susceptible result in disk diffusion testing?
The antimicrobial agent is an appropriate choice for treating the infection
According to the CLSI protocol, what is the recommended temperature range for short-term storage of a working supply of disks?
2°C to 8°C
What is the recommended timeframe for using inoculum suspensions after preparation?
15 minutes
What is the term for the selection of antimicrobial agents to be tested against a particular bacterial isolate?
Antimicrobial battery
What does the intermediate category in disk diffusion testing imply?
The antimicrobial agent may be effective in certain circumstances or at high concentrations
What is the purpose of pressing the disks firmly onto the agar surface?
To ensure contact between the disk and the agar
How long should the inoculated plates be incubated in a 35°C ambient air incubator?
16 to 18 hours
What is the main purpose of disk diffusion testing?
To determine the antimicrobial agent's effectiveness against the bacterial isolate
Why are certain antimicrobials excluded from the test battery?
Because the organism is intrinsically resistant to them
What is the recommended storage condition for long-term storage of disks?
-20°C or below in a non-frost-free freezer
What is the purpose of excluding certain antimicrobials from the test battery due to acquired resistance patterns?
To prioritize more potent antimicrobials and improve treatment outcomes
What is not a possible interpretation of an intermediate result in disk diffusion testing?
The antimicrobial agent is completely ineffective against the bacterial isolate
Which of the following is not a category in disk diffusion testing?
Uncertain
What is the purpose of using a sterile cotton swab to inoculate the Mueller-Hinton agar plate?
To swab the surface of the agar plate evenly
Why is ceftazidime included in the test battery for Pseudomonas aeruginosa but not for Staphylococcus aureus?
Because it was specifically developed for use against Pseudomonas aeruginosa
What is the significance of a susceptible result in disk diffusion testing?
The antimicrobial agent is effective against the bacterial isolate and may be used to treat the infection
What is the role of the CLSI in selecting antimicrobial agents for testing?
To publish up-to-date tables of potential antimicrobial agents
Study Notes
Inoculum Preparation
- Inoculum suspensions should be used within 15 minutes of preparation
- Suspensions may be diluted or supplemented with more organisms if they do not match the standard's turbidity initially
Selection of Antimicrobial Agents
- Antimicrobial agents chosen for testing against a particular bacterial isolate are referred to as the antimicrobial battery or panel
- The CLSI publishes up-to-date tables listing potential antimicrobial agents for testing against particular organisms or organism groups
Criteria for Antimicrobial Battery Content and Use
- Antimicrobials to which the organism is intrinsically resistant are excluded from the test battery (e.g., vancomycin vs. Gram-negative bacilli)
- Antimicrobials specifically developed for use against particular organisms but not others (e.g., ceftazidime for Pseudomonas aeruginosa but not Staphylococcus aureus) are only included in the appropriate battery
- Antimicrobials with resistance common to local microbial flora are excluded from routine testing, and only more potent antimicrobials are included in the test battery
Antibiotic Susceptibility Test Laboratory Activity
- Performed on isolated bacteria from clinical specimens to determine effective antimicrobial agents for treating infections
- Only bacteria likely to be contributing to an infection are tested
- Standard conditions for antimicrobial susceptibility testing methods have been established by the CLSI
Standardized and Controlled Components
- Bacterial inoculum size
- Growth medium
- pH
- Cation concentration
- Blood and serum supplements
- Thymidine content
- Incubation atmosphere
- Incubation temperature
- Incubation duration
- Antimicrobial concentrations tested
Methods
Dilution Methods
- Macrodilution
- Drug is diluted with MH broth in several tubes together with the inoculum
- Incubate for 24 hours
- Check for presence of growth
- Plate them by overlaying a MIC media with the broth
- Incubate plates for 24 hours
- Determine the MBC
- Microdilution
- Utilizes microtiter plates
- Utilizes minute volume of broth and inoculum
- Almost the same with macrodilution
- Incubate microtiter plates for 24 hours
- Check for growth
- Agar dilution
- Utilizes a type of tray where antibiotics are added first in the MHA, then just before it solidifies, you add the inoculum using a “replicator”
- Incubate for 24 hours
- Determine MIC by checking for presence or absence of colonies
Diffusion Method
- E-test or Epsilometer test
- Utilizes MHA plus a plastic strip impregnated with antibiotics on one side and the label of concentration on the other side
- Inoculate organism to MHA, then place the strip (antibiotic side, face-down)
- Incubate for 24 hours
- Observe for ZOI (MIC)
Alamar System
- Utilizes a type of tray with many wells and each row has a specific antibiotic, MHB, the inoculum + “alamar blue” indicator
- Initially, all wells will appear blue in color
- Incubate
- Presence of “pink” colored well means it is negative for growth
Automated Methods
- Vitek Systems
- Conventional Testing Methods
- Paper disks containing specific concentrations of antimicrobial agent are placed onto the inoculated surface
- After overnight incubation, the diameters of the zones produced by antimicrobial inhibition of bacterial growth are measured
- Result is interpreted as nonsusceptible, susceptible, intermediate, or resistant to a particular drug according to preset criteria
This quiz is based on a laboratory activity on antimicrobial susceptibility testing, which determines the effectiveness of antimicrobial agents against bacteria isolated from clinical specimens. It covers the standard conditions and methods used in this process. Test your knowledge of this important laboratory procedure!
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