Multi-tiered, high through-put screen for compounds effective against bacterial biofilm compounds effective for inhibiting and eradicating bacterial biofilm
Inventors
Hassett, Daniel J. • Lamkin, Thomas J. • Panmanee, Warunya • Taylor, Deborah E. • Shea, Chloe J. A.
Assignees
University of Cincinnati • United States Department of the Air Force
Publication Number
US-10202631-B2
Publication Date
2019-02-12
Expiration Date
2034-03-14
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Abstract
A high through-put screening method for identifying agents effective for inhibiting biofilm formation and/or killing established biofilm are disclosed. The method includes three tiers, and each tier includes three specific biological process assays. The tier levels are a primary screen, a confirmation screen, and a dose-response screen, and the biological process assays include as says for total bacterial growth, bacterial metabolic activity, and biofilm formation.
Core Innovation
The invention provides a multi-tiered high through-put screening (HTS) method for identifying agents effective in inhibiting bacterial biofilm formation and killing established bacterial biofilms. The method comprises conducting three tiers: a primary screen, a confirmation screen, and a dose-response screen, with each tier including three specific biological process assays selected to reflect different mechanisms—total bacterial growth, bacterial metabolic activity, and biofilm formation assays.
The background describes the problem of bacterial biofilms, which are complex communities of bacteria protected by a matrix that confer high resistance to antibiotics and biocides, making biofilms difficult to eradicate. These biofilms form on diverse surfaces in medical, industrial, and environmental settings, causing infections, material corrosion, and safety hazards. Current methods targeting early biofilm formation stages using toxic antimicrobials have downstream environmental problems. There remains a significant need for safe, non-toxic agents that effectively inhibit biofilm formation and/or kill established biofilms across problematic bacterial species such as Pseudomonas aeruginosa, Acinetobacter baumannii, and Staphylococcus epidermidis.
The invention solves this problem by developing an HTS platform that screens compounds through sequential tiers, each with assays measuring bacterial growth, metabolic activity, and biofilm formation. Compounds passing primary and confirmation screens at specific inhibition thresholds advance to dose-response screening, generating dose-response curves to identify putative hits. These hits are confirmed and validated using confocal laser scanning microscopy (CLSM) to assess biofilm inhibition and killing, with further toxicity and minimum inhibitory concentration (MIC) testing performed.
Claims Coverage
The patent includes two independent claims focusing on methods of controlling bacterial biofilms on substrates by applying compositions comprising specified compounds.
Method for controlling bacterial biofilms on substrates
Applying a composition consisting of one or more compounds selected from a specific set of chemical compounds outlined in Table 1 to a substrate to control bacterial biofilms.
Method comprising specific compound application
Applying a composition comprising the compound 5-chloro-2-methyl-3-oxo-1,2-thiazole-4-carbonitrile to control bacterial biofilms on a substrate.
The claims cover methods of controlling bacterial biofilms by applying compositions containing one or more defined compounds from Table 1, emphasizing specified chemical entities effective against biofilms on substrates.
Stated Advantages
The multi-tiered high through-put screening method increases the reliability and robustness of identifying compounds effective against biofilms by applying confirmation and dose-response tiers.
The identified compounds show efficacy in inhibiting biofilm formation and killing established biofilms across multiple problematic bacterial species, including P. aeruginosa, A. baumannii, and S. epidermidis.
Broad spectrum agents identified demonstrate low cytotoxicity in human cell lines, suggesting safety for potential applications.
The method allows identification of compounds effective in aerobic and anaerobic growth conditions, addressing biofilm environments with oxygen deprivation.
Documented Applications
Controlling bacterial biofilms on substrates such as medical devices, hospital surfaces, nursery appliances, toy surfaces, fabrics, and wound dressing materials that contact human skin vulnerable to bacterial infection.
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