Developing a Crystal Violet Assay to Quantify Biofilm Production Capabilities of Staphylococcus aureus
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Date
2016-05
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The Ohio State University
Abstract
Staphylococcus aureus is an opportunistic pathogen responsible for significant morbidity and mortality in clinical settings in both humans and animals. This emerging pathogen is among the top three nosocomial pathogens in human and veterinary hospitals due to its ability to survive in these environments for long periods. This survival is likely the result of S. aureus’s production of biofilm: a protective matrix of bacterially secreted proteins that allow colonies to attach to environmental surfaces. Preventing and controlling this pathogen, specifically within small animal veterinary hospitals, becomes critical for two reasons. One, the presence of this pathogen increases the risk of animals developing a hospital-acquired infection. Two, this pathogen poses an occupational risk to veterinary hospital staff. Therefore, it is important to know the biofilm production potential and characteristics of S. aureus isolates in these animal facilities. This information can be used to more effectively prevent or control a uniquely natured biofilm-producing S. aureus. To quantify biofilm production potential of S. aureus isolates the Crystal Violet (CV) assay is commonly used. This assay is preferred due to its simplicity, reliability, and quick throughput. With this assay isolates can be categorized as high, moderate, or non-biofilm producers. To adapt this assay, we used two high producer bacterial control strains NE95 and NE1241, a moderate producer wild type JE2 (WT), and a low producer NE1193 to validate the CV assay. Assay optimization included changes in the quantity of overday culture growth, overnight incubation time, aspiration quantities, washing technique, and microplate reader absorbance wavelength. Through assay optimization a sensitive and specific CV assay was developed for the quantification and characterization of S. aureus biofilm production.
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Keywords
Crystal Violet Assay, Biofilm, Staphylococcus aureus, Veterinary Medicine