Characterization of fusarium keratitis outbreak isolates: contribution of biofilms to antimicrobial resistance and pathogenesis.
OBJECTIVE
Fusarium is a major cause of microbial keratitis, and its ability to form biofilms was suggested as a contributing factor in recent outbreaks. We investigated the ability of outbreak Fusarium isolates (F. solani species complex [FSSC] and F. oxysporum species complex [FOSC]) to form biofilms in vitro and in vivo, and evaluated their antifungal susceptibilities.


METHODS
Biofilm formation was assessed using our in vitro contact lens model and in vivo murine model. Biofilm architecture was assessed using confocal laser scanning microscopy (CLSM). Susceptibility against amphotericin B (AmB), voriconazole (VCZ), and natamycin (NAT) was determined using the CLSI-M38-A2 method and XTT metabolic assay.


RESULTS
FSSC strains formed more biofilms than FOSC, in a strain- and clade-dependent manner. CLSM analyses revealed that "high biofilm forming" (HBF) strains had denser and thicker biofilms than "low biofilm forming" (LBF) strains of both species (thickness 51 vs. 41 μm for FSSC and 61 vs. 45 μm for FOSC strains, P < 0.05 for both comparisons). Fusarium biofilms exhibited species-dependent antifungal susceptibilities (e.g., FSSC biofilms AmB minimal inhibitory concentrations [MIC] ≥16 μg/mL, while NAT or VCZ MICs were 2-8 μg/mL). FSSC-infected mice had severe corneal opacification independent of biofilm thickness, while FOSC infection resulted in moderate corneal opacification. Corneal fungal burden of mice infected with HBF strains was higher than those of the LBF strains. In contrast, the reference ATCC isolate was unable to cause infection.


CONCLUSIONS
The ability to form biofilms is a key pathogenicity determinant of Fusarium, irrespective of the thickness of these biofilms. Further studies are warranted to explore this association in greater detail.
Center for Medical Mycology, Department of Dermatology, Case Western Reserve University, Cleveland, Ohio 44106, USA. Pranab.Mukherjee@case.edu
Pranab K
PK
Mukherjee
Jyotsna
J
Chandra
Changping
C
Yu
Yan
Y
Sun
Eric
E
Pearlman
Mahmoud A
MA
Ghannoum
D000666:Amphotericin B / Q000494:pharmacology; D000818:Animals; D000935:Antifungal Agents / Q000494:pharmacology; D018441:Biofilms / Q000254:growth & development*; D003320:Corneal Ulcer / Q000453:epidemiology / Q000382:microbiology*; D004195:Disease Models, Animal; D004196:Disease Outbreaks; D025141:Drug Resistance, Fungal; D019573:Epithelium, Corneal / Q000382:microbiology; D015821:Eye Infections, Fungal / Q000453:epidemiology / Q000382:microbiology*; D060585:Fusariosis / Q000453:epidemiology / Q000382:microbiology*; D005670:Fusarium / Q000187:drug effects / Q000502:physiology*; D051379:Mice; D008810:Mice, Inbred C57BL; D008826:Microbial Sensitivity Tests; D018613:Microscopy, Confocal; D010866:Natamycin / Q000494:pharmacology; D011743:Pyrimidines / Q000494:pharmacology; D014230:Triazoles / Q000494:pharmacology; D014774:Virulence; D065819:Voriconazole
D000935:Antifungal Agents; D011743:Pyrimidines; D014230:Triazoles; D000666:Amphotericin B; D010866:Natamycin; D065819:Voriconazole