Phase-dependent antifungal activity against Aspergillus fumigatus developing multicellular filamentous biofilms

Abstract

Overall, the data presented demonstrate that treatment of actively growing A. fumigatus cells with antifungal agents is more efficacious than treating mature structures in vitro. Amphotericin B was consistently more effective against each phase and displayed rapid effects, and therefore may be a suitable option for managing patient groups at risk from aspergillosis infections.

In Vitro Interaction between Alginate Lyase and Amphotericin B against Aspergillus fumigatus Biofilm Determined by Different Methods

“…However, Ͼ95% of the reduction in metabolic activity was obtained with Af293 at the highest concentrations (32 g/ml for AMB and 16 g/ml for LAMB), and there was still 10% activity 24 h after challenging CG261 with 32 g/ml of both AMB and LAMB ( Fig. 1 and 2), in agreement with previous studies (6,13,18). Conversely, AlgL did not affect the time-kill curve of A. fumigatus Af293 and CG261 biofilms challenged with 1 U/ml of the compound, while a minimal reduction in metabolic activity over the 24-h period was shown following exposure to 10 U/ml AlgL, with a maximal 26% metabolism reduction displayed after 24 h ( Fig.…”

“…This growth phenotype, which complies with the definition of a biofilm (10), may help A. fumigatus to colonize the host substratum and to resist phagocytic and antimicrobial attacks, mimicking the typical Candida albicans or bacterial biofilm (8,11). Recent observations have consistently shown that all antifungal drugs are significantly less effective when A. fumigatus is grown as a biofilm than when it is grown in the planktonic state (4,7,9,12,13), presumably as a reflection of multiple resistance mechanisms, including the ECM, which would prevent drug diffusion by acting as a physical barrier (14). This could contribute to the overall mortality with invasive aspergillosis, which remains high, despite the use of newer broad-spectrum antifungal agents and diagnostic adjuncts (15).…”

“…To validate the results of the checkerboard microdilution analysis, preformed biofilms of A. fumigatus Af293 and CG261 were challenged with AMB and LAMB (at 0.5ϫ, 1ϫ, and 2ϫ SMICs), alone or combined with defined concentrations of AlgL (1 and 10 U/ml), in order to monitor cell viability (i.e., the OD) versus time using the XTT assay, which measures the aforementioned fungal biomass-associated metabolic activity (31). As it was expected (13,18), the effects of polyenes against A. fumigatus cells in biofilms were rapid, with the Af293 isolate exhibiting after 4 h of drug challenge 30.7% and 39.4% OD reductions at the lowest (8 g/ml) and highest (32 g/ml) concentrations of AMB, respectively. Notably, more pronounced effects were seen with LAMB, with which 38.5% and 59.7% OD reduction rates were achieved after 4 h of drug challenge (8 and 32 g/ml, respectively) for the Af293 isolate (Fig.…”

“…However, high concentrations of AMB (at least 8 g/ml) are needed to inhibit hyphal clumps (6,18) or mature mycelia in vitro (13), although AMB was seen to be significantly more effective against each phase of A. fumigatus biofilm development and to display more rapid effects than other antifungals, such as voriconazole or caspofungin (13). In contrast to AMB deoxycholate, unique efficacy against Candida (C. albicans and C. parapsilosis) biofilms grown on a bioprosthetic model was displayed by lipid-formulated AMB (19), probably due to the higher doses of drug present in these formulations.…”

Microbial glycoside hydrolases as antibiofilm agents with cross-kingdom activity

“…The Pel polysaccharide enhances resistance to several antibiotics including aminoglycosides and colistin (22,24,25). Because biofilm formation by A. fumigatus is associated with increased resistance to a number of antifungal agents (26)(27)(28), we hypothesized that GAG may have an analogous function to Pel in enhancing resistance to antifungal agents. To test this hypothesis, we investigated whether Sph3 h or PelA h could potentiate the activity of commonly used antifungal drugs.…”