Measuring Drug Susceptibility

Minimal Inhibitory Concentration (MIC)

Minimum Inhibitory Concentration is the lowest drug concentration that prevents visible microorganism growth after overnight incubation.

Learning Objectives

Analyze data to interpret minimal inhibitory concentration values

Key Takeaways

Key Points

  • Minimum inhibitory concentration (MIC) can be determined by culturing microorganisms in liquid media or on plates of solid growth medium.
  • A lower MIC value indicates that less drug is required for inhibiting growth of the organism; therefore, drugs with lower MIC scores are more effective antimicrobial agents.
  • By identifying appropriate drugs and their effective concentrations, MIC scores aid in improving outcomes for patients and preventing evolution of drug-resistant microbial strains.

Key Terms

  • culture: The process of growing a bacterial or other biological entity in an artificial medium.
  • minimum inhibitory concentration: This is the lowest concentration of an antimicrobial drug that prevents visible growth of a microorganism after overnight incubation with media.
  • bacteriostatic: A drug that prevents bacterial growth and reproduction but does not necessarily kill them. When it is removed from the environment the bacteria start growing again.

Definition and Measurement

In microbiology, minimum inhibitory concentration (MIC) is the lowest concentration of an antimicrobial (like an antifungal, antibiotic or bacteriostatic) drug that will inhibit the visible growth of a microorganism after overnight incubation. MICs can be determined on plates of solid growth medium (called agar, shown in the “Kirby-Bauer Disk Susceptibility Test” atom) or broth dilution methods (in liquid growth media, shown in ) after a pure culture is isolated. For example, to identify the MIC via broth dilution, identical doses of bacteria are cultured in wells of liquid media containing progressively lower concentrations of the drug. The minimum inhibitory concentration of the antibiotic is between the concentrations of the last well in which no bacteria grew and the next lower dose, which allowed bacterial growth. There are also several commercial methods available to experimentally measure MIC values.

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Microbroth Dilution Method: To identify the lowest concentration required for a given antibiotic to inhibit bacterial growth, an identical amount of bacteria is introduced into wells of liquid media containing progressively lower concentrations of the drug. (Here, the dilution series of the drug is set up from left to right: for example, well E1 might contain 100 units of drug; E2, 50 units; E3, 25 units; E4, 12.5 units; etc.). Because bacterial growth made the media in well E5 cloudy and the media in well E4 is indistinguishable from clear media, this indicates that the minimum inhibitory concentration is between the drug concentrations in wells E4 and E5. (Image courtesy of Microrao, Dept. of Microbiology, JJMMC, Davangere).

Significance and Applications

An MIC is generally regarded as the most basic laboratory measurement of the activity of an antimicrobial agent against an organism. Because a lower MIC value indicates that less of the drug is required in order to inhibit growth of the organism, drugs with lower MIC scores are more effective antimicrobial agents. Currently, there are a few web-based, freely accessible MIC databases. MIC scores are important in diagnostic laboratories to confirm resistance of microorganisms to an antimicrobial agent and also to monitor the activity of new antimicrobial agents. Clinicians use MIC scores to choose which antibiotics to administer to patients with specific infections and to identify an effective dose of antibiotic. This is important because populations of bacteria exposed to an insufficient concentration of a particular drug or to a broad-spectrum antibiotic (one designed to inhibit many strains of bacteria) can evolve resistance to these drugs. Therefore, MIC scores aid in improving outcomes for patients and preventing evolution of drug-resistant microbial strains.

Kirby-Bauer Disk Susceptibility Test

Kirby-Bauer testing measures sensitivity of bacteria to antibiotics by culturing bacteria on solid growth media surrounding sources of drug.

Learning Objectives

Review the procedure for the Kirby-Bauer antibiotic test

Key Takeaways

Key Points

  • KB tests are performed under standard conditions, so the minimum inhibitory concentration for a given antibiotic can be calculated by comparing the observed zone of inhibition ‘s size to known values.
  • Clinicians use KB test results to choose antibiotics effective against the specific bacteria causing a patient’s infection. Using specifically-targeted antibiotics helps decrease the frequency of drug-resistant bacteria evolving.

Key Terms

  • Kirby-Bauer antibiotic testing: This is a method to determine the sensitivity of microorganisms to specific antimicrobial drugs; greater drug efficacy yields larger microbe-free zones surrounding drug-containing disks after overnight growth on solid media.
  • zone of inhibition: This is an area of media where bacteria are unable to grow, due to presence of a drug that impedes their growth.
  • minimum inhibitory concentration: This is the lowest concentration of an antimicrobial drug that prevents visible growth of a microorganism after overnight incubation with media.

Kirby-Bauer antibiotic testing (also called KB testing or disk diffusion antibiotic sensitivity testing) uses antibiotic-containing wafers or disks to test whether particular bacteria are susceptible to specific antibiotics. First, a pure culture of bacteria is isolated from the patient. Then, a known quantity of bacteria are grown overnight on agar ( solid growth media) plates in the presence of a thin wafer that contains a known amount of a relevant antibiotic. If the bacteria are susceptible to the particular antibiotic from a wafer, an area of clear media where bacteria are not able to grow surrounds the wafer, which is known as the zone of inhibition. A larger zone of inhibition around an antibiotic-containing disk indicates that the bacteria are more sensitive to the antibiotic in the disk.

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Kirby-Bauer test: In Kirby–Bauer testing, discs containing antibiotics are placed on agar where bacteria are growing, and the antibiotics diffuse out into the agar. If an antibiotic stops the bacteria from growing, one can see circular areas around the wafers where bacteria have not grown.

KB tests are performed under standardized conditions and standard-sized zones of inhibition have been established for each antibiotic. KB test results are usually reported as sensitive, intermediate, or resistant, based on the size of the zone of inhibition. If the observed zone of inhibition is greater than or equal to the size of the standard zone, the microorganism is considered to be sensitive to the antibiotic. Conversely, if the observed zone of inhibition is smaller than the standard size, the microorganism is considered to be resistant. The size of a zone of inhibition in a KB test is inversely related to the minimum inhibitory concentration (MIC), which is the amount of antibiotic required to prevent bacterial growth in an overnight culture. The MIC (in µg/ml) can be calculated from known standard-curve (linear regression) graphs based on the diameter of the observed inhibition zone diameter (in millimeters).

Clinicians can use KB test results to choose appropriate antibiotics to combat a particular infection in a patient. Administering antibiotics that specifically target the particular bacteria that are causing the infection can avoid using broad-spectrum antibiotics, which target many types of bacteria. Thus, clinical application of KB testing results can decrease the frequency with which antibiotic-resistant bacteria evolve.