Modeling of Fungal Biofilms Using a Rat Central Vein Catheter
Candida frequently grows as a biofilm, or an adherent community of cells protected from both the host immune system and antimicrobial therapies. Biofilms represent the predominant mode of growth for many clinical infections, including those associated wit
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1. Introduction The use of biofilm models has fueled investigation of this unique manner of growth and has been vital for characterizing associated phenotypic properties and gene expression patterns (1–3). Because Candida infection of commonly placed medical devices, such as a dentures, venous catheters, or urinary catheters, involves biofilm growth, models closely mimicking infection at these clinical sites are of interest (4). To date, the in vivo model most commonly used for Candida biofilm study is the venous catheter model described here (5–8). This model emulates one of the most common clinical biofilm infections and mimics environmental host conditions at this site, including anatomical location, flow conditions, and exposure to host cells, serum proteins, and immune factors. The catheter is secured in the jugular vein without disruption of blood flow and then tunneled subcutaneously and positioned in a wire casing for protection. Inoculation of the catheter occurs 24 h after catheter placement to allow for a conditioning period of host protein
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_40, © Springer Science+Business Media, LLC 2012
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deposition on the catheter surface (4, 9, 10). Throughout the experiment, the model uses the clinically relevant anticoagulant, heparin, although other anticoagulants can be utilized. The rat venous catheter model can be used as a tool to answer a variety of scientific questions and has identified C. albicans biofilms with altered morphology, adhesion, matrix production, and drug susceptibility (8, 11, 12). Confocal microscopy and scanning electron microscopy can successfully illustrate intact biofilm architecture, including the fungal cell morphology, presence of extracellular matrix, and the incorporation of host cells (5). Adjusting the duration of biofilm formation from 6 to 72 h can capture the time course of this process from cell adhesion to development of a multicellular community with both yeast and hyphal fungal cell morphologies, host cell components, extracellular matrix, and open areas or channels. Sonication effectively removes cells for microbiological enumeration or cellular analyses, such as gene expression profiling or cell biology studies (13). Microbiological counts can be used to quantify the viable biofilm mass and are a simple method of measuring the impact of a luminal drug therapy or comparing the difference in viable burden among several genetic strains. In addition, organs and blood from distant sites can be collected for measurement of viable burden and assessment of biofilm dispersion or dissemination of disease. Although vascular catheters may be infected by hematogenous seeding from a distant vascular site, the model has primarily been utilized for study following intraluminal infection. The latter results in more reproducible cell number and biofilm cell mass among experiments.
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