1.1 Minimum inhibitory concentrations

The minimum inhibitory concentration (MIC) is the lowest concentration of an antibiotic agent that inhibits the visible growth of a micro-organism. MICs are determined by incubating the micro-organism with the antibiotic agent at a range of different concentrations.

The classic test is broth dilution: this is done by doing doubling dilutions of the antibiotic agents to give a range of concentrations, each one twice as concentrated as the one before. Each tube in the dilution series is then inoculated with a fixed amount of bacterial inoculum. The tubes are incubated, and after 18–24 hours the dilution series is examined for bacterial growth which appears as turbidity, or cloudiness (Figure 1). A variation is to use a microdilution method: again, the doubling dilutions are used and bacterial inoculum added, but this time the test is performed using much smaller volumes, usually in a microtitre plate. The turbidity can be read visually, or using a special plate reader. Broth dilution or microdilution methods are considered the Gold Standard for AST; however, broth dilution is rarely used in routine diagnostic laboratories. The microdilution method is often used in automated systems, which you will learn about later in the module.

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Figure 1 Determining the MIC using a manual broth dilution method. Tubes containing increasing concentrations of antibiotic (from right to left in the figure) and a fixed amount of inoculum are examined for turbidity following incubation.
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A photo of seven test tubes with increasing concentrations of antibiotic: 64 µg/ml, 32 µg/ml, 16 µg/ml, 8 µg/ml, 4 µg/ml, 2 µg/ml and 1 µg/ml.

Figure 1 Determining the MIC using a manual broth dilution method. Tubes containing increasing concentrations of antibiotic (from right ...

  • What is the MIC for the antibiotic agent shown in Figure 1?

  • In Figure 1, visible bacterial growth (turbidity) can be observed in the three tubes containing 4, 2 and 1 μg/ml of antibiotic. The remaining four tubes appear to be clear. At these antibiotic concentrations, visible bacterial growth is inhibited. The lowest concentration where there is no visible bacterial growth is 8 μg/ml; therefore, the MIC is 8 μg/ml.

It is important to note that the MIC of an antibiotic is the concentration at which this antibiotic is bacteriostatic – it stops bacterial growth. However, the test does not show whether there are any viable bacteria still in the suspension. The concentration of antibiotic that kills bacteria is known as the minimum bactericidal concentration (MBC). The MBC will often be a higher concentration of antibiotic than the MIC. You would need to subculture the clear tubes to see if any live bacteria were still present in order to find the MBC.

MICs are used to evaluate the antibiotic efficacy of compounds: the lower the MIC, the less antibiotic is required to inhibit the growth of an organism, so the more susceptible it is. However, the value of a MIC is influenced by the method used to determine it, so MICs are only meaningful if they are compared to other values that were obtained using the same standardised method.

  • Can MICs be directly compared between antibiotics?

  • No: the MIC value for one antibiotic cannot be compared to the MIC value of another antibiotic. A lower figure does not necessarily mean the antibiotic will be more effective in practice. This is affected by a range of other factors, such as the achievable tissue concentration, half-life in the blood or whether the antibiotic goes on killing bacteria after the levels in the body have fallen.

MICs are not very useful clinically on their own to predict treatment success or failure. It is therefore necessary to set breakpoints using the MIC. If the MIC of an isolate is below the breakpoint, the organism is considered susceptible. Breakpoints are discussed in the following section.

1 The theory underpinning AST

1.2 Interpretive criteria