3.6.1 Antimicrobial Assays

The disc diffusion (Kirby-Bauer) method involves placing a compound-impregnated filter paper disc onto a lawn of test organism on Mueller-Hinton agar and...

The disc diffusion (Kirby-Bauer) method involves placing a compound-impregnated filter paper disc onto a lawn of test organism on Mueller-Hinton agar and measuring the resulting zone of inhibition in millimetres, following CLSI or EUCAST reference guidelines. The minimum inhibitory concentration (MIC) is determined by 96-well broth microdilution using doubling dilutions of the test compound, defined as the lowest concentration at which no visible bacterial growth occurs, following the CLSI M07 reference method. The minimum bactericidal concentration (MBC) is subsequently determined by sub-culturing wells at and above the MIC onto fresh agar, with the MBC defined as the concentration achieving 99.9% kill; an MBC-to-MIC ratio below 4 is generally interpreted as indicating bactericidal (rather than merely bacteriostatic) activity. Reference organisms conventionally used include Staphylococcus aureus (ATCC 25923), Escherichia coli (ATCC 25922), and Candida albicans (ATCC 10231), with methicillin-resistant S. aureus (MRSA) strains included where clinical relevance to resistant infection is being assessed.