20.02.2025

Reference MIC testing of M.tuberculosis released

The EUCAST AMST reference method for M. tuberculosis MIC determination has now been released in the Mycobacteria methods section. A summary is below:

• A 96-well U-bottom-shaped polystyrene microtiter plate with an untreated surface.
• Liquid Middlebrook 7H9 medium (7H9) made from the broth base with a final concentration of 0.2% glycerol (no Tween 80), sterilized and then enriched with 10% Oleic Albumin Dextrose Catalase (OADC) growth supplement.
• A final inoculum of 10⁴-10⁶ CFU/mL of a MTBC isolate.
• A range of log₂ concentration values of the ATTB to be tested, calculated from 1 mg/L and encompassing the targeted MIC value.
• At least two positive growth controls: one with 100% of the inoculum (GC100%) and one with 1% (100-fold dilution, GC1%).
• A negative control: a well with 200 µL of uninoculated broth.
• For water-insoluble agents, the final proportion of the solvent should not exceeding 1% and should be the same concentration in all ATTB-containing and growth controls wells (no ATTB).
• Incubation of the microtiter plate should be at 36°C ± 2 °C in ambient air (no CO₂).
• The MIC value defined as the lowest concentration in mg/L, where no visible growth is observed at the time point when both GC100% and GC1% are showing visible growth.
• Mycobacterium tuberculosis H37Rv ATCC 27294 as the quality control (QC) strain.