Susceptibility testing of terbinafine alone and in combination with amphotericin B, itraconazole, or voriconazole against conidia and hyphae of dematiaceous molds.

Studies have demonstrated excellent in vivo efficacy of terbinafine combined with other antifungal agents against dematiaceous molds; however, there is a lack of in vitro studies. Most studies evaluated conidia inocula, but susceptibility testing of hyphae could mimic the fungal status in infected tissues and might reflect the therapeutic potential of the agent. We investigated the in vitro susceptibility of terbinafine alone and in combination with amphotericin B, itraconazole, or voriconazole against conidia by microdilution and dynamic measurement of hyphae growth of dematiaceous molds. The MIC values for hyphae were, until 3 dilutions, below the MIC obtained for conidia. The results indicated 100% synergistic interactions between terbinafine and azoles or amphotericin B in all tests, but lower MICs for hyphae. In conclusion, our findings allow us to say that the hyphal form of tested dematiaceous molds showed high susceptibility to all antifungal agents evaluated, alone and in combination with terbinafine.

In vitro evaluation of the type of interaction obtained by the combination of terbinafine and itraconazole, voriconazole, or amphotericin B against dematiaceous molds.

In vitro associations using the checkerboard microdilution method indicated lower MIC ranges and MIC median values for each drug (terbinafine, itraconazole, voriconazole, and amphotericin B) in association than those obtained for each single drug. Fractional inhibitory concentration index (FIC) results showed 100% synergism in the association of terbinafine with voriconazole, 96.5% in the association of terbinafine with amphotericin B, and 75.9% in the association of terbinafine with itraconazole. Drug combinations may be useful for treatment of dematiaceous mold infections as an alternative treatment to enhance the effectiveness of each drug.

Standardization of hyphal growth inhibition rate as a means of evaluating Microsporum spp. in vitro susceptibility to terbinafine, griseofulvin, and ciclopiroxolamine.

Reference methods for antifungal susceptibility tests recommend the use of conidia as inoculum. However, some isolates produce few conidia, while the invasive form of filamentous fungi in general is hyphae making susceptibility tests infeaseble. These facts suggest that other than conidia broth dilution method is required for susceptibility tests. The aim of this study was to clarify if the hyphal growth inhibition rate could be used as a method of determining the antifungal susceptibility of genus Microsporum. For this reason, a method which traces hyphal tips automatically and measures their growth rate was standardized for Microsporum spp. Control growth curves and test growth curves obtained by real-time observation of the hyphae groups responses to different concentrations of terbinafine, griseofulvin, and ciclopiroxolamine were used to compare with minimum inhibitory concentrations (MICs) obtained by conidia broth microdilution method. A visible reduction in the growth inhibition rate was observed when hyphal activity was evaluated using the third or fourth serial two-fold dilution below the MIC determined by broth microdilution for terbinafine and ciclopiroxolamine. For griseofulvin, this reduction occurred after the fifth dilution below the MIC. This study highlights the importance of the inoculum type used to determine the in vitro susceptibility of Microsporum strains. We conclude that measurement of hyphal growth inhibition, despite being time consuming, could be a suitable method for evaluating antifungal susceptibility, particularly for fungi as Microsporum spp. that produce a small (or not at all) number of conidia.

Preanalytical conditions for broth microdilution antifungal susceptibility of Microsporum spp.

Dermatophytoses caused by the genus Microsporum require a long-duration therapy compared to infections caused by other genera. Treatment of these cutaneous infections includes topical and systemic antifungal agents. Tinea capitis and tinea unguium caused by M. canis and M. gypseum are the most difficult-to-treat dermatophytoses. There are few specific studies about corresponding antifungal susceptibility in vitro. Recently, the Clinical and Laboratory Standards Institute proposed the M38A document as standard to determine the minimum inhibitory concentrations (MICs) of several antifungal agents against conidium-forming filamentous fungi; however, dermatophytes were not included in this document. This study aimed to contribute to continuing investigations concerning the optimal antifungal susceptibility testing conditions of Microsporum spp. to terbinafine, ciclopiroxolamine and griseofulvin. The results pointed out potato dextrose agar as the best culture medium for inducing conidia sporulation, inoculum density amounting to 1 x 10(3) conidia ml(-1), containing only microconidia, with an incubation time of 7 days at 28 degrees C and 100% growth inhibition serving as an endpoint. The minimum fungicidal concentration values were in accordance with the MICs values, showing a fungicidal activity of these drugs towards the tested strains. According to our results, in general, terbinafine was more active than griseofulvin and ciclopiroxolamine.