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Applied and Environmental Microbiology, July 2007, p. 4592-4601, Vol. 73, No. 14
0099-2240/07/$08.00+0     doi:10.1128/AEM.02506-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Cryptococcus neoformans Biofilm Formation Depends on Surface Support and Carbon Source and Reduces Fungal Cell Susceptibility to Heat, Cold, and UV Light

Department of Microbiology and Immunology, Albert Einstein College of Medicine, Bronx, New York

Received 27 October 2006/ Accepted 8 May 2007

The fungus Cryptococcus neoformans possesses a polysaccharide capsule and can form biofilms on medical devices. We describe the characteristics of C. neoformans biofilm development using a microtiter plate model, microscopic examinations, and a colorimetric 2,3-bis(2-methoxy-4-nitro-5-sulfophenyl)-5-[(phenylamino) carbonyl]-2H-tetrazolium-hydroxide (XTT) reduction assay to observe the metabolic activity of cryptococci within a biofilm. A strong correlation between XTT and CFU assays was demonstrated. Chemical analysis of the exopolymeric material revealed sugar composition consisting predominantly of xylose, mannose, and glucose, indicating the presence of other polysaccharides in addition to glucurunoxylomannan. Biofilm formation was affected by surface support differences, conditioning films on the surface, characteristics of the medium, and properties of the microbial cell. A specific antibody to the capsular polysaccharide of this fungus was used to stain the extracellular polysaccharide matrix of the fungal biofilms using light and confocal microscopy. Additionally, the susceptibility of C. neoformans biofilms and planktonic cells to environmental stress was investigated using XTT reduction and CFU assays. Biofilms were less susceptible to heat, cold, and UV light exposition than their planktonic counterparts. Our findings demonstrate that fungal biofilm formation is dependent on support surface characteristics and that growth in the biofilm state makes fungal cells less susceptible to potential environmental stresses.

Published ahead of print on 18 May 2007.