Robust biofilm assay for quantification and high throughput screening applications.

Autor: Rajamani S; US Army Medical Research Institute of Infectious Diseases, 1425 Porter Street, Ft. Detrick, Frederick, MD 21702, USA; General Dynamics Information Technology, Frederick, MD, USA. Electronic address: sathish.rajamani.ctr@mail.mil., Sandy R; US Army Medical Research Institute of Infectious Diseases, 1425 Porter Street, Ft. Detrick, Frederick, MD 21702, USA., Kota K; US Army Medical Research Institute of Infectious Diseases, 1425 Porter Street, Ft. Detrick, Frederick, MD 21702, USA., Lundh L; US Army Medical Research Institute of Infectious Diseases, 1425 Porter Street, Ft. Detrick, Frederick, MD 21702, USA., Gomba G; US Army Medical Research Institute of Infectious Diseases, 1425 Porter Street, Ft. Detrick, Frederick, MD 21702, USA., Recabo K; US Army Medical Research Institute of Infectious Diseases, 1425 Porter Street, Ft. Detrick, Frederick, MD 21702, USA., Duplantier A; US Army Medical Research Institute of Infectious Diseases, 1425 Porter Street, Ft. Detrick, Frederick, MD 21702, USA; Cherokee Nation Assurance, Frederick, MD, USA., Panchal RG; US Army Medical Research Institute of Infectious Diseases, 1425 Porter Street, Ft. Detrick, Frederick, MD 21702, USA. Electronic address: rekha.g.panchal.civ@mail.mil.
Jazyk: angličtina
Zdroj: Journal of microbiological methods [J Microbiol Methods] 2019 Apr; Vol. 159, pp. 179-185. Date of Electronic Publication: 2019 Feb 28.
DOI: 10.1016/j.mimet.2019.02.018
Abstrakt: Bacterial biofilms are populations of bacteria within a self-produced adherent extracellular matrix that are notoriously resistant to treatment. Existing methods for biofilm quantification are often limited in their dynamic range of detection (signal-to-background), throughput, and require modifications to the protocol depending on the bacterial species. To address these limitations, a broad utility, high-throughput (HTP) method was required. Using a fluorescent dye, FM1-43, we stained the biofilm, followed by solvent extraction and quantitation of biofilm employing a fluorescent plate reader. Utilizing eight different bacterial pathogens, we demonstrate that this method is widely applicable for biofilm quantification. Depending on the species, this biofilm assay offered a large dynamic range of 8-146 fold change compared to 2-22 fold for crystal violet staining under similar conditions. In addition to routine biofilm quantification using this new assay, as a proof-of-concept, 1200 compounds were screened against two different bacterial species to identify biofilm inhibitors. In our HTP screens we successfully identified compounds rifabutin and ethavarine as potential biofilm inhibitors of Burkholderia pseudomallei Bp82 and Acinetobacter baumannii biofilm production respectively. This newly validated biofilm assay is robust and can be readily adapted for antibiofilm screening campaigns and can supplant other less sensitive and low throughput methods.
(Copyright © 2019 Elsevier B.V. All rights reserved.)
Databáze: MEDLINE
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