MICR 290 Exam Questions with Correct Answers| Latest Update Guaranteed Success
What is the disk diffusion test? One of the most popular methods for measuring antibiotic
activity is the disk diffusion test, also known as the Kirby-Bauer test. This test uses small pieces
of filter paper that are impregnated with an antibiotic. An agar plate is coated with a very thin
layer of the indicator strain of bacteria (i.e. the bacteria that is being tested for antibiotic
susceptibility), the antibiotic disks are placed on top, and the plate is incubated.
- If the indicator bacteria is susceptible to the antibiotic, there will be a clear zone of inhibition
around the disk.
- If the bacteria is fully resistant to the antibiotic, the area around the disk will be cloudy with
bacterial growth.
The zone diameter, generally measured in millimetres, can be related to the susceptibility of
the indicator bacteria to the antibiotic.
As you can appreciate from the provided images, a major advantage of the disk diffusion test is
that multiple antibiotics can be tested on the same agar plate. Disks containing different
antibiotics are placed on different parts of the agar surface. The diameter of each zone of
clearing can be measured, allowing the antibiotic susceptibility of the indicator strain to a broad
panel of antibiotics to be quickly determined.
What are the steps of a disk diffusion test? 1. Indicator Bacteria Preparation:
First, a culture of the indicator bacteria is grown in liquid media.
2. Agar Plate Preparation:
Once the culture is grown, a sterile swab is used to spread the culture over the surface of an
agar plate. It is important to swab the whole surface of the agar plate, so that the indicator
bacteria will grow everywhere on the agar (i.e. so that the plate will become confluent).
3. Disk Placement:
,After the agar plate has been swabbed and allowed to dry, antibiotic-impregnated disks are
applied to the surface of the agar plate (employing aseptic technique!). Then, the plate is
incubated under suitable conditions to allow the indicator bacteria to grow.
4. Zone of Clearing Measurement:
Once the indicator bacteria has grown, any zones of clearing that appear can be measured.
What is the minimum inhibitory concentration (MIC)? The MIC is inversely related to the
susceptibility of this strain to the antibiotic.
If the bacterial strain becomes resistant to the antibiotic, the MIC of this antibiotic for the
resistant strain will be greater.
Although disk diffusion tests give a quantitative measure of the antibiotic susceptibility of an
indicator bacteria (i.e. a zone diameter), this value by itself is not very meaningful. It is only
useful when compared to other results from disk diffusion tests. A more generally useful way of
measuring the antibiotic susceptibility of a bacterial strain is by determining a value known as a
minimum inhibitory concentration (MIC). The MIC is the smallest concentration of antibiotic
that fully inhibits the growth of a particular bacterial strain.
An MIC is particular to the combination of a specific antibiotic and a specific strain of bacteria.
What is the Epsilometer test (E-test)? Several antibiotic susceptibility tests can be used to
determine MIC values. One of the easiest methods is the Epsilometer test (E test), which is
conceptually very similar to the disk diffusion tests.
In an E test, an antibiotic-impregnated strip is used to inhibit the growth of an indicator strain
on an agar plate. While Kirby-Bauer disks contain a uniform amount of antibiotic throughout
the disk, E test strips contain a gradient of antibiotic. One end of the E test strip contains a large
amount of antibiotic, and the other end contains a small amount of antibiotic.
As in a disk diffusion test, the surface of an agar plate is first swabbed with a very thin layer of
the culture of the indicator bacterial strain. Then, an E test strip is applied to the surface of the
agar plate, and the plate is incubated to allow the indicator strain to grow.
,If the indicator strain is susceptible to the antibiotic, there will be a zone of clearing around the
E test strip. However, this zone will not be uniform around the entire strip. The zone will be
largest around the end of the strip that contains the greatest amount of antibiotic, and will
decrease in size along the strip as the local antibiotic concentration decreases.
The end of the E test strip with the lowest amount of antibiotic typically does not kill the
indicator bacteria, and there is no zone of clearing at this end. As a result, the zone of clearing
will intersect the strip somewhere in the middle. This offers a very easy way to determine the
MIC for the antibiotic against the indicator bacteria.
What is a broth micro dilution test? One of the most commonly used methods for
determining the MIC of an antibiotic is the broth microdilution test. In this test, liquid growth
media is added to the wells of a 96-well microplate. Then, different amounts of an antibiotic are
added to the wells of the plate. The different wells span a wide range of antibiotic
concentrations (e.g. 64 μg/mL, 32 μg/mL, 16 μg/mL, 8 μg/mL, 4 μg/mL, and so on). Finally, the
wells are all inoculated with a small volume of a liquid culture that contains the indicator
bacterial strain. Once all of these components have been added, the microplate is incubated
under conditions that promote the growth of the indicator bacteria. Much like in E tests, at low
antibiotic concentrations, the growth of the indicator bacteria may not be impacted by the
antibiotic, and the cultures in these wells become cloudy. At high antibiotic concentrations, the
bacterial cells are killed and/or their growth is inhibited, and the liquid growth media in these
wells remains clear. The lowest antibiotic concentration that fully inhibits bacterial growth is
the MIC. Compared to disk diffusion tests and E tests, broth microdilution tests are relatively
labour intensive. This test requires the addition of growth media, antibiotic, and indicator
bacteria culture to all of the wells in a microplate. In addition, the different dilutions of the
antibiotic must be prepared. Overall, this test requires a lot of pipetting, and is relatively slow
and time-consuming. For this reason, disk diffusion tests and E tests are more commonly used
in clinical microbiology labs. However, broth microdilution tests are commonly carried out in
research labs, allowing new antibiotics to be characterized against a variety of different
bacterial species. Often, it is important to test the susceptibility of a bacterial strain to a whole
What are antibiotic susceptibility tests? If a clinical lab suspects a bacterial isolate is
responsible for an infection, they will test how susceptible this isolate is to a panel of
antibiotics. These assays will provide zone diameters or MICs. Antibiotic susceptibility tests
must be performed with a high level of uniformity so that the results of these tests can be
compared. There are very strict guidelines that define how exactly these tests should be
performed. If these guidelines are followed, the results from antibiotic susceptibility tests can
, be compared within a lab, and also between different labs. This standardization can help guide
clinical decision making, allowing for the identification of the optimal antibiotic treatment for a
particular infection. In the rest of this section, you will learn more about the guidelines for
antibiotic susceptibility tests that are applied in many labs around the world, including in
Canada.
What are the antibiotic susceptibility testing guidlines? In Canada, many labs follow the
guidelines from the Clinical and Laboratory Standards Institute (CLSI) for antibiotic susceptibility
testing. These guidelines define how exactly these tests should be carried out. For example, for
a disk diffusion test, the CLSI guidelines define what kind of agar should be used, how much
agar should be used, how much bacteria should be swabbed on the plate, how much antibiotic
should be in the disks, and so on.
The CLSI also publishes antibiotic clinical breakpoints, which relate the zone diameter (from a
disk diffusion test) or an MIC (from a broth microdilution test) to the susceptibility or resistance
of a bacterial strain to an antibiotic.
Clinical breakpoints are commonly used by clinical microbiology labs to decide on the best
antibiotic to be used for patient therapy. Clinical labs identify the bacterial species responsible
for an infection, and test its susceptibility to a panel of antibiotics. These results are compared
to the published clinical breakpoints for zone diameters or MICs, revealing which antibiotics the
bacterial isolate is susceptible to. The lab can then recommend which antibiotic is most likely to
successfully treat the infection.
Clinical breakpoints quantitatively define the susceptibility of a bacterial species to an
antibiotic. Depending on the zone diameter or the MIC, bacteria may be categorized as
resistant (R), intermediate (I), or susceptible (S) to an antibiotic.
a) Resistant (R)
If a bacterial strain is categorized as R for an antibiotic, this means that it is resistant to that
antibiotic, and treatment is unlikely to be successful.
b) Intermediate (I)
What is the disk diffusion test? One of the most popular methods for measuring antibiotic
activity is the disk diffusion test, also known as the Kirby-Bauer test. This test uses small pieces
of filter paper that are impregnated with an antibiotic. An agar plate is coated with a very thin
layer of the indicator strain of bacteria (i.e. the bacteria that is being tested for antibiotic
susceptibility), the antibiotic disks are placed on top, and the plate is incubated.
- If the indicator bacteria is susceptible to the antibiotic, there will be a clear zone of inhibition
around the disk.
- If the bacteria is fully resistant to the antibiotic, the area around the disk will be cloudy with
bacterial growth.
The zone diameter, generally measured in millimetres, can be related to the susceptibility of
the indicator bacteria to the antibiotic.
As you can appreciate from the provided images, a major advantage of the disk diffusion test is
that multiple antibiotics can be tested on the same agar plate. Disks containing different
antibiotics are placed on different parts of the agar surface. The diameter of each zone of
clearing can be measured, allowing the antibiotic susceptibility of the indicator strain to a broad
panel of antibiotics to be quickly determined.
What are the steps of a disk diffusion test? 1. Indicator Bacteria Preparation:
First, a culture of the indicator bacteria is grown in liquid media.
2. Agar Plate Preparation:
Once the culture is grown, a sterile swab is used to spread the culture over the surface of an
agar plate. It is important to swab the whole surface of the agar plate, so that the indicator
bacteria will grow everywhere on the agar (i.e. so that the plate will become confluent).
3. Disk Placement:
,After the agar plate has been swabbed and allowed to dry, antibiotic-impregnated disks are
applied to the surface of the agar plate (employing aseptic technique!). Then, the plate is
incubated under suitable conditions to allow the indicator bacteria to grow.
4. Zone of Clearing Measurement:
Once the indicator bacteria has grown, any zones of clearing that appear can be measured.
What is the minimum inhibitory concentration (MIC)? The MIC is inversely related to the
susceptibility of this strain to the antibiotic.
If the bacterial strain becomes resistant to the antibiotic, the MIC of this antibiotic for the
resistant strain will be greater.
Although disk diffusion tests give a quantitative measure of the antibiotic susceptibility of an
indicator bacteria (i.e. a zone diameter), this value by itself is not very meaningful. It is only
useful when compared to other results from disk diffusion tests. A more generally useful way of
measuring the antibiotic susceptibility of a bacterial strain is by determining a value known as a
minimum inhibitory concentration (MIC). The MIC is the smallest concentration of antibiotic
that fully inhibits the growth of a particular bacterial strain.
An MIC is particular to the combination of a specific antibiotic and a specific strain of bacteria.
What is the Epsilometer test (E-test)? Several antibiotic susceptibility tests can be used to
determine MIC values. One of the easiest methods is the Epsilometer test (E test), which is
conceptually very similar to the disk diffusion tests.
In an E test, an antibiotic-impregnated strip is used to inhibit the growth of an indicator strain
on an agar plate. While Kirby-Bauer disks contain a uniform amount of antibiotic throughout
the disk, E test strips contain a gradient of antibiotic. One end of the E test strip contains a large
amount of antibiotic, and the other end contains a small amount of antibiotic.
As in a disk diffusion test, the surface of an agar plate is first swabbed with a very thin layer of
the culture of the indicator bacterial strain. Then, an E test strip is applied to the surface of the
agar plate, and the plate is incubated to allow the indicator strain to grow.
,If the indicator strain is susceptible to the antibiotic, there will be a zone of clearing around the
E test strip. However, this zone will not be uniform around the entire strip. The zone will be
largest around the end of the strip that contains the greatest amount of antibiotic, and will
decrease in size along the strip as the local antibiotic concentration decreases.
The end of the E test strip with the lowest amount of antibiotic typically does not kill the
indicator bacteria, and there is no zone of clearing at this end. As a result, the zone of clearing
will intersect the strip somewhere in the middle. This offers a very easy way to determine the
MIC for the antibiotic against the indicator bacteria.
What is a broth micro dilution test? One of the most commonly used methods for
determining the MIC of an antibiotic is the broth microdilution test. In this test, liquid growth
media is added to the wells of a 96-well microplate. Then, different amounts of an antibiotic are
added to the wells of the plate. The different wells span a wide range of antibiotic
concentrations (e.g. 64 μg/mL, 32 μg/mL, 16 μg/mL, 8 μg/mL, 4 μg/mL, and so on). Finally, the
wells are all inoculated with a small volume of a liquid culture that contains the indicator
bacterial strain. Once all of these components have been added, the microplate is incubated
under conditions that promote the growth of the indicator bacteria. Much like in E tests, at low
antibiotic concentrations, the growth of the indicator bacteria may not be impacted by the
antibiotic, and the cultures in these wells become cloudy. At high antibiotic concentrations, the
bacterial cells are killed and/or their growth is inhibited, and the liquid growth media in these
wells remains clear. The lowest antibiotic concentration that fully inhibits bacterial growth is
the MIC. Compared to disk diffusion tests and E tests, broth microdilution tests are relatively
labour intensive. This test requires the addition of growth media, antibiotic, and indicator
bacteria culture to all of the wells in a microplate. In addition, the different dilutions of the
antibiotic must be prepared. Overall, this test requires a lot of pipetting, and is relatively slow
and time-consuming. For this reason, disk diffusion tests and E tests are more commonly used
in clinical microbiology labs. However, broth microdilution tests are commonly carried out in
research labs, allowing new antibiotics to be characterized against a variety of different
bacterial species. Often, it is important to test the susceptibility of a bacterial strain to a whole
What are antibiotic susceptibility tests? If a clinical lab suspects a bacterial isolate is
responsible for an infection, they will test how susceptible this isolate is to a panel of
antibiotics. These assays will provide zone diameters or MICs. Antibiotic susceptibility tests
must be performed with a high level of uniformity so that the results of these tests can be
compared. There are very strict guidelines that define how exactly these tests should be
performed. If these guidelines are followed, the results from antibiotic susceptibility tests can
, be compared within a lab, and also between different labs. This standardization can help guide
clinical decision making, allowing for the identification of the optimal antibiotic treatment for a
particular infection. In the rest of this section, you will learn more about the guidelines for
antibiotic susceptibility tests that are applied in many labs around the world, including in
Canada.
What are the antibiotic susceptibility testing guidlines? In Canada, many labs follow the
guidelines from the Clinical and Laboratory Standards Institute (CLSI) for antibiotic susceptibility
testing. These guidelines define how exactly these tests should be carried out. For example, for
a disk diffusion test, the CLSI guidelines define what kind of agar should be used, how much
agar should be used, how much bacteria should be swabbed on the plate, how much antibiotic
should be in the disks, and so on.
The CLSI also publishes antibiotic clinical breakpoints, which relate the zone diameter (from a
disk diffusion test) or an MIC (from a broth microdilution test) to the susceptibility or resistance
of a bacterial strain to an antibiotic.
Clinical breakpoints are commonly used by clinical microbiology labs to decide on the best
antibiotic to be used for patient therapy. Clinical labs identify the bacterial species responsible
for an infection, and test its susceptibility to a panel of antibiotics. These results are compared
to the published clinical breakpoints for zone diameters or MICs, revealing which antibiotics the
bacterial isolate is susceptible to. The lab can then recommend which antibiotic is most likely to
successfully treat the infection.
Clinical breakpoints quantitatively define the susceptibility of a bacterial species to an
antibiotic. Depending on the zone diameter or the MIC, bacteria may be categorized as
resistant (R), intermediate (I), or susceptible (S) to an antibiotic.
a) Resistant (R)
If a bacterial strain is categorized as R for an antibiotic, this means that it is resistant to that
antibiotic, and treatment is unlikely to be successful.
b) Intermediate (I)
