In vitro susceptibility tests are performed on microorganisms suspected of causing disease, particularly if the organism is thought to belong to a species that may exhibit resistance to frequently used antimicrobial agents. The tests are also important in resistance surveillance, epidemiological studies of susceptibility and in comparisons of new and existing agents.
Dilution procedures are used to determine the minimum inhibitory concentrations (MICs) of antimicrobial agents and are the reference method for antimicrobial susceptibility testing. MIC methods are used in resistance surveillance, comparative testing of new agents, to establish the susceptibility of organisms that give equivocal results in routine tests, for tests on organisms where routine tests may be unreliable and when a quantitative result is required for clinical management. In dilution tests, microorganisms are tested for their ability to produce visible growth on a series of agar plates (agar dilution) or in broth (broth dilution) containing serial dilutions of the antimicrobial agent.
The lowest concentration of an antimicrobial agent (in mg/l) that, under defined in vitro conditions, prevents the appearance of visible growth of a microorganism within a defined period of time is known as the MIC. The MIC is a guide for the clinician to the susceptibility of the organism to the antimicrobial agent and aids treatment decisions. Careful control and standardisation is required for intra- and inter-laboratory reproducibility, as results may be significantly influenced by the method used. It is generally accepted that broth MIC tests are reproducible to within one doubling dilution of the real end point (i.e. one well or tube in a doubling dilution series).
Broth dilution is a technique in which containers holding identical volumes of broth with antimicrobial agent solutions in incrementally (usually geometrically) increasing concentrations are inoculated with a known number of microorganisms.
Broth microdilution denotes the performance of the broth dilution test in microdilution trays.
The method described in this part of ISO 20776 is intended for the testing of pure cultures of aerobic bacteria that are easily grown by overnight incubation on agar and grow well in Mueller-Hinton broth, which may be supplemented. The broth microdilution method described in this part of ISO 20776 is essentially the same as those used in many countries, including France, Germany, Sweden, the United Kingdom, and the United States. The method is also essentially the same as the broth microdilution method published by the European Committee on Antimicrobial Susceptibility Testing (EUCAST). All these methods are based on those described by Ericsson and Sherris.
This part of BS EN ISO 20776 describes one reference method, broth microdilution, for determination of MICs. The MIC reflects the activity of the drug under the described test conditions, and can be interpreted for clinical management purposes by taking into account other factors, such as drug pharmacology or bacterial resistance mechanisms. This allows categorization of bacteria as “susceptible” (S), “intermediate” (I), or “resistant” (R). In addition, MIC distributions can be used to define wild type or non-wild type bacterial populations. Although clinical interpretation of the MIC value is beyond the scope of this part of ISO 20776, modifications of the basic method are required for certain antimicrobial agent - bacteria combinations to facilitate clinical interpretation. These modifications are included in a separate table. It is advisable to compare other susceptibility testing methods (e.g. routine methods or diagnostic test devices) with this reference.