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kyleannkelsey
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What is the limit for visible growth?
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10^7 CFU/mL represents what?
Visible growth
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Define the Minimum inhibitory concentration (MIC):
The minimum concentration that inhibits visible growth
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What is the term for the minimal concentration to inhibit visible growth?
Minimum inhibitory concentration (MIC)
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What is the definition for significant bactericidal activity?
Killing of three logs (99.9%) or greater
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What is the term for killing of three logs or greater (i.e. 99.9%)
Significant bacterial activity
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Minimum bactericidal concentration is defined as:
Lowest concentration with significant bactericidal activity
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What is the term for the lowest concentration with significant bactericidal activity?
Minimum bactericidal activity
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Frequency of resistant mutants is:
1 mutant per 10^6 to 10^9 viable bacterial cells
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1 mutant per 10^6 to 10^9 viable bacteria cell represents what statistic?
Frequency of resistant mutants
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What is a breakpoint?
A determining committee, such as the FDA
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Define CLSI:
Clinical and Laboratory Standards Institute
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Define EUCAST:
European Committee on Antimicrobial Susceptibility Testing
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What categories of test results for susceptibility exist?
- Susceptible
- Intermediate
- Resistant
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Give examples of determining committees or Breakpoints:
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The FDA, EUCAST and CLSI all fall under what greater category?
Breakpoints or Determining committees
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How do we detect resistance?
Susceptibility tests
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Good susceptibility is a guarantee of success with a given antimicrobial agent (T/F)?
False
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Susceptibility tests are not adequate (T/F)?
True, we need to optimize detection of resistance
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Therapeutic success depends on what other factors other than susceptibility?
- Penetration of drug at infection site
- Immune status
- Need for surgery
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___________ is dependent on Immune status, penetration of drug at infection site and need for surgery?
Therapeutic success
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Disk diffusion can be described as what type of measure of susceptibility?
Qualitative
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Name a qualitative test for susceptibility:
Disk diffusion
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Potency of drugs can be compared using qualitative means (T/F)?
False
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What was discussed in class as not accomplishable via Qualitative testing?
Comparison of drug potency
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What examples were given in class for Qualitative tests?
- Measure of susceptibility
- Comparison of drugs
- Macrobroth dilution
- Microbroth Dilution
- Agar dilution
- E-test
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E-test, measure of susceptibility, drug comparisons, macrobroth dilution, microbroth dilution and agar test are all what general type test?
Quantitative
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Describe the process of creating a Macrobroth dilution:
- 1-3 mL Volumes of broth
- Dilute drug in 2 fold dilution to get to 0.06 ug/mL of the drug
- Inoculate with 5E5 CFU/mL
- Incubate for 18-24 hours
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What is the inherent Error of the susceptibility test?
- Inherent 2 fold error of MIC test
- i.e. 2ug/ml can be 1-4 ug/ml
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What is the Inoculum effect?
- The inoculum in the MIC test is much lower than in a real infection
- Does not account for pathogens production of antibiotic inactivating enzymes
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What technical difficulties may arise in susceptibility tests?
- Fuzzy or trailing endpoints
- Difficult to interpret
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What limitations are inherent in the MIC test?
- Inherent error = inherent 2 fold error in dilutions
- Inoculum effect = inoculation in MIC test is usually much lower than in vivo/does not account for pathogen production of antibiotic inactivating enzyme
- Technical difficulties= diff. to interpret results
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Macrobroth dilution is what general type of assay?
Quantitative
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What are the advantages of a Macrobroth dilution?
- Quantitative
- Most conservative (gives the highest MIC)
- Gold standard
- Can measure Minimum Bactericidal concentration
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What test is quantitative, conservative (giving the highest MICs), able to measure MBC and considered the Gold standard for susceptibility assays?
Macrobroth Dilution
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What disadvantages are there for Macrobroth Dilution?
- Labor intensive
- Inoculums below 10^6 = missed mutants
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What test is unable to detect inoculums below 1066 and is considered labor intensive?
Macrobroth Dilution
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What is the dilution volume for Microbroth dilution?
0.1mL
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Describe the Microbroth Dilution method:
- 0.1mL volumes
- Dilute 2 fold to reach 0.06ug/mL
- 50,000 CFU/mL inoculum
- Incubate 18-24 hours
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Why is Microbroth considered less accurate than a Macrobroth?
The smaller the sample size the less accurate the results
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Why is a Macrobroth Dilution considered more accurate than a Microborth?
Larger sample size equates to more accurate results
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When accuracy is the most important component of your testing, what susceptibility assay would you choose, Microbroth or Macrobroth?
Macrobroth
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Describe an agar dilution susceptibility test:
Spread plating of agar is performed on plates of varying concentration
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What are the benefits of using an Agar dilution susceptibility test?
- Quantitative
- 32 strains per drug plate
- Easier than macrobroth
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What susceptibility test is Quantitative, easier than macroborth and allows for 32 strains to be plated per plate?
Agar Dilution
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What disadvantages exist for the Agar dilution susceptibility test?
- Does not yield a minimum bactericidal concentration
- MICs are lower than macrobroth
- Lower inoculum
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What test does not yield MBCs, has MICs that are lower than macrobroth and a low inoculum?
Agar Dilution
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An E-test is similar to what other tests?
Agar dilution and disk diffusion
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What susceptibility test is similar to an agar dilution and disk diffusion?
E-test
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Describe the method for an E-test:
- Spread a lawn of bacteria
- Apply E-test strip
- Incubate for 18-24 hours
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What test requires a lawn of bacteria, an antibiotic gradient strip and an incubation period of 18-24 hours?
E-test
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Describe the characteristics of an E-test strip:
Plastic strip coated with antibiotic gradient
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What is the name for a strip coated with an antibiotic gradient?
E-test Strip
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In the E-test what happens to the antibiotic during incubation?
Diffuses into the agar at rate dependent on the concentration
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Describe how you read an E-test:
The point of the E-test strip that aligns with the lowest concentration point where bactericidal activity can be detected= MIC
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What are the advantages of the E-test:
- Quantitative
- 6 drugs can be tested concurrently
- Easy
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What susceptibility test allows for 6 drugs to be tested concurrently, is quantitative and easy to perform?
E-test
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What are the disadvantages of the E-test?
- Does not yield MBCs
- Costly
- Results are difficult to interpret
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What susceptibility test has results that are difficult to interpret, does not produce MBC data and is costly?
E-test
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Describe the method for Disk Diffusion:
- Spread a lawn of bacteria
- Apply antibacterial impregnanted disk
- Incubate 18-24 hours
- What test requires a lawn to be spread, antibacterial impregnated disks to be layed on the lawn and an incubation period of 18-24 hours?
- Disk Diffusion
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How is a disk diffusion test interpreted?
By measuring the zone of inhibition
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What susceptibility test is interpreted by measuring the zone of inhibition?
Disk Diffusion
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What variables control the zone of inhibition in the Disk diffusion test?
- Temperature
- Concentration of Drug disk
- Inoculum size
- Susceptibility of test organism
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Temperature, Conc. Of drug, inoculum size and susceptibility of test organism are all variable factors for what susceptibility test?
Disk Diffusion
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What are the advantages of disk diffusion?
- Fast and easy to perform
- Inexpensive
- Many drugs can be tested concurrently
- Drug interactions may be detected
- New drugs are usually available for this test type
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What is the disadvantage of the disk diffusion assay?
You cannot tell HOW sensitive the organism is from the zone size
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What I easy, fast, inexpensive, able to determine the susceptibility of many drugs concurrently, can detect drug interactions and has new drugs available for testing?
Disk Diffusion
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What are the advantages of automated susceptibility tests?
- Rapid, accurate and cheap
- Reduce workload
- Very specific computer analyzed results
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What are the disadvantages for automated susceptibility tests?
Algorithms may be inaccurate for certain organisms, drugs or inoculum concentrations
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A negative result for a Molecular detection of resistance genes test means what?
Does not mean the organism is susceptible
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What are the shortcomings of Molecular detection of resistance genes?
Genes that are too diverse may not be detected
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What are the positive aspects of Molecular detection of resistance genes?
- Potential for:
- Rapid results
- Accurate results
- Highly specific results
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What issue require resolution prior to routine use of Molecular detection of resistance gene testing?
- Cost
- Convenience
- Preventative measures for over and understating resistance
- Needs methods for detecting new resistance genes
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Phenotypic tests for resistance have a broader scope than Molecular detection of resistance genes test (T/F)?
True
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What is the point of Irreversibility?
The point beyond which treatment will have no impact on an outcome of an infection
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What is the term for the point beyond which most appropriate treatment will have no impact on the outcome?
Point of irreversibility
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What has been well studied in terms of the point of irreversibility?
- Pneumonia (most well studied)
- Sepsis
- Meningitis
- Neutropenia
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Pneumonia, Sepsis, Meningitis and Neutropenia have all been studied (Pneumonia more so than the others) in terms of what characteristic?
Point of irreversibility
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What differences in Rapid vs microdilution susceptibility methods were discovered by the Doern study?
- The mortality rate was lower for those receiving a rapid susceptibility method
- Fewer procedures were used on patients receiving rapid methods
- Faster witch from empiric Rx
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Why is early antibiotic administration more robust?
Attacks the pathophysiological process
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What is the social impact of using Rapid susceptibility methocs?
- Reduction of therapy cost
- 2.4 million Saved/year
- Early isolation of patient=less unnecessary spread
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How can we diminish the development of resistance?
Diminish drug use
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