Comparison of Four Methods for the in vitro Susceptibility Testing of Dermatophytes

Abstract

Objectives: Infections caused by dermatophytes affect a high percentage of the population. Antifungal susceptibility testing (AST) can offer useful information about the susceptibility profiles of the pathogens as well as the concomitant documentation of the appropriate treatment. However, the slow growth rate of these fungi and their poor sporulation are factors that can delay and affect the performance of the AST. The proposed methods by the CLSI or the EUCAST are both laborious for the everyday routine. There are alternative applications which propose the use of an inoculum, consisting of a conidia-mycelium mixture or even plain mycelia, as well as the use of resazurin in order to facilitate the reading. The aim of this study was to compare these approaches to the EUCAST method and evaluate their performance. Methods: Three alternative methods were compared to the EUCAST proposed methodology for conidia forming molds. The last was defined as the reference method. The methods under evaluation were (a) a fragmented mycelia method, (b) the EUCAST method with the addition of resazurin sodium salt solution and (c) the fragmented mycelia method with the addition of resazurin sodium salt solution. Twenty-two isolates (8 Trichophyton interdigitale, 8 T. rubrum, and 6 Microsporum canis) were tested against the antifungal agents of griseofulvin, terbinafine, fluconazole, and itraconazole. Results: The essential agreement between the methods was calculated in percentages and a statistical analysis of the results was performed. Data evaluation revealed sufficient overall agreement of the methods with the addition of resazurin to the initial "uncolored" methods (98.9 and 97.5% for the EUCAST and the fragmented mycelia methods, respectively). The fragmented mycelia method exhibited a relatively sufficient overall agreement in comparison to the EUCAST method (90%) and not a satisfactory correlation, probably as a result of various issues of standardization. Conclusion: The EUCAST method was found to be the more reliable one, whereas the addition of resazurin sodium salt solution facilitates the reading and provides a reliable and objective evaluation. The fragmented mycelia method could serve as an alternative that should be applied only in cases of poor or no sporulating dermatophytes.

Predicting a therapeutic cut‐off serum level of itraconazole in recalcitrant tinea corporis and cruris—A prospective trial

“…However, no such standards are available for dermatophytoses. As most of the recent worldwide susceptibility data of ITZ vis a vis dermatophytes suggests that this class of fungi are largely sensitive to ITZ, 1,2,7,19 it is probable that levels achieved in the serum and as a corollary in the stratum corneum are predictive of treatment outcomes. Thus, it is plausible that clinical failures with ITZ may represent inadequate serum levels, and thus, there is a need to study cut-off serum levels which can predict optimum dosing, and lead to better and more predictable treatment outcomes with ITZ in tinea corporis/cruris.…”

Comparison of the Sensititre YeastOne® and CLSI M38‐A2 microdilution methods in determining the activity of nine antifungal agents against dermatophytes

“…A change in colour from blue (oxidised state) to pink (reduced) indicated fungal growth. Some studies have evidenced that use of Alamar Blue does not affect the MIC values, but makes it more evident to read for dermatophyte 31 . Discrepancies in the MICs or MECs between the two methods indicated that the concentration of Alamar Blue should be explored for each antifungal agent and each different species of dermatophyte.…”

Myxobacteria From Vietnam: Isolation, Phylogenetic Analysis and Bioprospection

“…Add 50 µL test microorganism inocula; and 30 µL resazurin at 0.015 mg/mL as an indicator for cell viability; plates were incubated at 35 o C for 16-48 hours [29]. The MIC endpoints were observed visually when viable cells changed their colour from purple to pink [30].…”