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

Aspergillus fumigatus undergoes morphological transition throughout its growth and development. These changes have direct implications for the effectiveness of antifungal treatment. Here we report the in vitro antifungal activity of voriconazole, amphotericin B and caspofungin against three specific phases of multicellular development of A. fumigatus. A. fumigatus conidia were propagated for 8, 12 and 24 h prior to antifungal challenge. The resultant activity of the three agents tested was determined using an XTT reduction assay to assess both endpoint and time-kill susceptibility profiles. Endpoint susceptibility testing demonstrated a time-dependent decrease in efficacy for all three antifungal agents as the complexity of the A. fumigatus hyphal structure developed. Overall, amphotericin B exhibited the best spectrum of activity at each phase of growth, but was comparable to voriconazole against germinated conidial growth (8 h). Later, both voriconazole and caspofungin were ineffective against complex mycelial structures (12 and 24 h). Time-kill studies demonstrated that amphotericin B was significantly more efficacious at reducing A. fumigatus metabolism than both voriconazole and caspofungin for all three growth phases examined, most notably after 1 h of drug exposure (P < 0.001). 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