Flow Cytometry Antifungal Susceptibility Testing of Aspergillus fumigatus and Comparison of Mode of Action of Voriconazole vis-à-vis Amphotericin B and Itraconazole

Abstract

Aspergillus fumigatus isolates were tested with three antifungals by flow cytometry (FC) and fluorescenceactivated cell sorting. FC results after 4 h correlated well with MICs obtained by the NCCLS M38-A method; voriconazole exhibited fungicidal activity, albeit to a lesser extent than amphotericin B, but to a greater extent than itraconazole.

Comparison of Two Probes for Testing Susceptibilities of Pathogenic Yeasts to Voriconazole, Itraconazole, and Caspofungin by Flow Cytometry

“…This effect only occurred in the resistant strains when challenged with the highest drug concentrations (4 to 16 g/ml); the effect was similar to that observed previously with fluconazole (14) and flucytosine (19,14). We were able to identify itraconazole categorical endpoints after 1 h of incubation with this agent; the overall time in other studies for this incubation period is 4 to 6 h for yeasts (20,2) and 2 to 3 h for A. fumigatus (21).…”

“…The availability of rapid and reliable tools to determine the susceptibility of yeasts, namely, Candida and Cryptococcus spp., to antifungal agents is mandatory. In the last few years, the susceptibility to antifungal agents has been evaluated by several flow cytometry methods (16,17,18,19,20,21,23); these methods can save time and provide additional information regarding mechanisms of action and resistance of the drugs tested. We have already developed cytometric protocols to study amphotericin B, flucytosine, and fluconazole (16,19), but each antifungal class may require a different protocol that takes into consideration its molecular characteristics and mechanisms of action.…”

“…Cytometric methodology has also been evaluated for testing itraconazole with acridine orange, ethidium bromide, and fluorescein diacetate, but these methods require 8 to 24 h to yield results (9). A flow cytometry assay has recently been described to study Aspergillus fumigatus versus voriconazole using PI as the marker of fungicidal activity (21). However, although voriconazole may be fungicidal for Aspergillus spp., it is only fungistatic against yeasts.…”

Human Dendritic Cells followingAspergillus fumigatusInfection Express the CCR7 Receptor and a Differential Pattern of Interleukin-12 (IL-12), IL-23, and IL-27 Cytokines, Which Lead to a Th1 Response

“…Since addition of either of these two antifungal agents did not affect cytokine secretion from LPSstimulated DC, we hypothesized that the action of VCZ and AB on A. fumigatus development might directly influence the signal controlling cytokine production by infected DC. AB is known to bind to existing ergosterol in the fungal membrane, while VCZ blocks ergosterol biosynthesis, affecting A. fumigatus growth in two different stages, as previously suggested by Ramani et al (34). VCZ might allow the early stages of germination (swelling) and the release of intracellular fungal constituents that are able to stimulate cytokine production, whereas killing by AB might be more efficient.…”

Hitting the Caspofungin Salvage Pathway of Human-Pathogenic Fungi with the Novel Lasso Peptide Humidimycin (MDN-0010)

“…Flow cytometry (FC) assays were performed with the C. albicans strain treated with the following compounds (concentration ranges): humidimycin (8 to 0.06 g/ml), CAS (1.0 to 0.004 g/ml), CAS at a sublethal dose (0.03 g/ml), ITZ at a sublethal dose (0.06 g/ml), humidimycin (8 to 0.06 g/ml) plus CAS at a sublethal dose, and humidimycin (8 to 0.06 g/ml) plus ITZ at a sublethal dose. Propidium iodide (PI) was used as a probe to determine cell viability and membrane permeability (18). Cells that were not treated, with and without the probe, and AMB curves were used as controls.…”