Biofilm Formation Studies in Microtiter Plate Format
Although Candida biofilms have been clearly identified as playing an increasingly important role in human disease, their biology and the reason for their poor susceptibility to antifungal agents remain largely unknown. Over recent years, various models ha
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1. Introduction A biofilm is defined as a community of microorganisms attached to a surface: it contains an exopolymer matrix and exhibits distinctive phenotypic properties (1). Biofilm cell physiology clearly differs from that of planktonic cells and is notably characterized by increased antibiotic resistance (2). Although a few studies have been published on biofilm formation in Coccidioides immitis and Cryptococcus neoformans, most of the fungal biofilm work has focused on Candida. The Candida species (and C. albicans and C. glabrata in particular) are major nosocomial pathogens and are responsible for various types of local and (often life-threatening) systemic infections. Medical implants (such as catheters, heart valve prostheses, and joint replacements) are all susceptible to colonization by microorganisms with the ability to attach to the device and
Alexandra C. Brand and Donna M. MacCallum (eds.), Host-Fungus Interactions: Methods and Protocols, Methods in Molecular Biology, vol. 845, DOI 10.1007/978-1-61779-539-8_25, © Springer Science+Business Media, LLC 2012
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form a biofilm on its surface. Candida cells in biofilms display phenotypic traits that differ dramatically from those of their planktonic counterparts. Resistance to therapeutic drugs is remarkably high and is associated with increased expression of certain drug efflux pumps. Biofilms can thus constitute reservoirs for subsequent reinfestation (3). Furthermore, mucosal infections by Candida (thrush, vaginitis) are probably associated with the formation of biofilms that may limit the efficiency of currently available antifungal treatments. Despite clear evidence of the clinical relevance of such structures, little is known about the molecular mechanisms responsible for biofilm formation by C. albicans and other yeasts. Over the last few years, a number of different models have been developed in order to better characterize Candida biofilms, including (1) growth of adherent populations on small portions of catheters, acrylic strips, or silicone squares, (2) growth on cellulose acetate using a perfused biofilm fermenter, and (3) growth on plastic slides using a microfermenter and (4) microtiter plates (4–6). Lastly, different animal models of catheter-associated C. albicans biofilm infection have been set up (7–9). These models have mostly been used to study the structure of Candida biofilms and the resistance to various antifungals. Together, it has been shown that Candida biofilms on catheter materials (1) feature yeast cells in their lower layers and mycelia in their upper layers, (2) contain polysaccharide matrix polymers, and (3) are significantly more resistant than planktonic cells to fluconazole and (to a lesser extent) amphotericin B. Furthermore, in vitro models have enabled characterization of three phases in C. albicans biofilm development. The first step is cell adherence to the surface, which is followed by an intermediate phase during which microcolonies are formed and produce an extracellular matrix. Finally
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