Ευαισθησίες των υφομυκήτων σε νεότερα αντιμυκητικά φάρμακα: κλινική σημασία

Abstract

The continuing increase of systemic and invasive mycoses due to filamentous fungi (moulds) such as species of Aspergillus, zygomycetes of the order Mucorales (Rhizopus, Mucor, θιπ.) and rare hyalohyphomycetes such as Fusarium and Scedosporium, and their difficult management because of their resistance in most antifungals, needed the development and standardization of a method to determine susceptibility to these drugs. The present dissertation aimed to the development of an antifungal susceptibility testing method which would be reliable and reproducible, in order to be used as a standard method. This method should be applicable to all filamentous fungi,The strains used for this study were isolates from cases with systemic mycoses with known antifungal susceptibility and outcome, as well as from a prospective registry study. The experiments of the first phase of the study aimed to verify the optimal conditions for the preparation of the inoculum of conidia to be tested for antifungal susceptibility with the broth microdilution method. The main conclusions of this phase were that the haemocytometer counting of conidia was a more reliable method of inoculum preparation than the spectrophotometric adjustment, and that Tween 20, a nonionic surfactant, should be used as a dispersing agent for the optimal suspension of hydrophobic conidia. The ideal inoculum size was 1.0 × 106 -5.0 × 106 CFU/ml. In the second phase, this new method was evaluated in three independent laboratories and its reliability and reproducibility were confirmed, with interlaboratory agreement of 89.2% and a narrow 95% CI (2.20 -2.65). In the third phase of the study the method was used for the susceptibility testing of strains from a prospective registry study of rare invasive mycoses and performed very well. The results of this study have been adopted by the EUCAST and constitute the basis of the guidelines for the determination of the MICs of antifungals against conidia forming moulds (EUCAST DEFINITIVE DOCUMENT E.DEF 9.1).