Essential oils (EOs) are widely investigated for their antimicrobial activities, but comparing MIC data across studies is challenging due to the lack of a harmonized reference assay. This article presents a standardized protocol for determining the Minimum Inhibitory Concentrations (MICs) of EOs against Staphylococcus aureus ATCC 29213, Enterococcus faecalis ATCC 29211, Escherichia coli ATCC 25922, Pseudomonas aeruginosa ATCC 27853 and Klebsiella pneumoniae ATCC 700603 by using a broth microdilution technique. The protocol was developed and validated using fourteen pharmacopoeia- or ISO-listed essential oils. These EOs were tested against five ATCC reference strains and 250 clinical isolates. The method incorporates ISO/EUCAST aligned inoculum preparation, twofold serial dilutions in 96-well plates and visual MIC endpoint determination together with optimized procedures. By following this standardized workflow, MIC values for EOs can be reproducibly and accurately determined, enabling reliable comparison across laboratories and supporting future translational applications.
Suggested by Dr. Maura Di Vito