Synergy, Indifference, or Antagonism? In Vitro Susceptibility of Fusarium and Aspergillus spp Isolated From Keratitis in South India Against Combinations of Natamycin, Voriconazole, and Anidulafungin

Abstract

Purpose: : A limited number of antimicrobial agents are available for the treatment of fungal keratitis. The non-ocular infectious disease literature suggests susceptibility testing of fungal isolates provides clinically relevant information. Nevertheless, physicians often combine drugs empirically rather than based on laboratory or clinical evidence. A popular combination used by U.S. ophthalmologists is natamycin and voriconazole. Here, we investigate whether combination pairs of natamycin, voriconazole, and anidulafungin interact synergistically in vitro against Fusarium spp and Aspergillus spp isolated from keratitis in South India.

Methods: : Fifteen isolates of Fusarium spp and 17 isolates of Aspergillus spp were randomly selected from culture-positive fungal keratitis treated at Aravind Eye Hospital from 11/2007 - 5/2008 that were enrolled in the Mycotic Ulcer Treatment Trial Pilot Study. Each isolate was tested for susceptibility to natamycin (N), voriconazole (V), and anidulafungin (A) alone and in combination: [N+V], [N+A], and [V+A]. Synergy was measured using the checkerboard microdilution method at 24, 48, and 72 hours, and quantified using the Fractional Inhibitory Concentration Index (FICI). Synergy was defined as FICI ≤ 0.5, indifference as FICI > 0.5 and ≤ 4, and antagonism as FICI > 4.

Results: : Of the 96 checkerboard tests performed on 32 isolates, synergy was reliably demonstrated in eight plates representing five unique isolates: 3/15 Fusarium spp, 2/4 Aspergillus fumigatus, and 0/11 Aspergillus flavus isolates. All five isolates showed synergy to [N+V]. Two isolates showed synergy to both [N+V] and [N+A]. One isolate showed synergy to all three combinations. The remaining 88 plates showed indifference or were contaminated. None of the plates showed antagonism.

Conclusions: : Synergy was demonstrated in five of the 32 filamentous fungal isolates. The most effective combination was natamycin and voriconazole. These results suggest that there may be additional benefit to treating fungal ulcers with combination therapy. Future studies are necessary to assess how in vitro susceptibilities correlate with in vivo clinical results.

Keywords: keratitis • fungal disease • cornea: basic science