Optimal growth conditions for the determination of the antifungal susceptibility of three species of dermatophytes with the use of a microdilution method

As a prerequisite to standardization of dermatophyte susceptibility testing , conditions that support optimal growth of different dermatophyte species must be established. Eighteen isolates of Trichophyton? spp. (T rubrum, T m entagrophytes, T tonsurans) were grown in 4 different media: RPMI 1640 with L-glutamine, without sodium bicarbonate and buffered at pH = 7.0; antibiot ic medium #3 (Penassay); yeast nitrogen base with 0.5% dextrose buffered at pH = 7.0; and Sabouraud dextrose broth. Incubation for 6 days at 35 degree s C produced the following results: RPMI and Sabouraud dextrose supported e qually sufficient growth for all strains tested; Penassay supported growth of only 33% of the isolates tested, and buffered yeast nitrogen base did no t support growth of any isolates. RPMI was selected as the optimal medium, and organisms were tested at both 30 degrees C and 35 degrees C with a stan dardized inoculum density of 10(3) conidia/ml, No temperature differences w ere noted in the amount of growth of the dermatophytes tested. With RPMI at an incubation temperature of 35 degrees C, 3 inoculum sizes (10(3), 10(4), and 10(5) conidia/mL) were tested against 4 antifungal agents: griseofulvi n, itraconazole, terbinafine, and fluconazole. Inoculum size did not affect minimum inhibitory concentration (MIC) results for itraconazole or terbina fine, but a larger inoculum produced a slightly higher MIC for griseofulvin and a noticeably higher MIC for fluconazole. Our data support the use of R PMI 1640, 35 degrees C, and 4 days as an incubation temperature and time, r espectively, and an inoculum of 10(3) conidia/ml as optimal conditions for the determination of the antifungal susceptibility of dermatophytes.