title: A Rapid and Quantitative Flow Cytometry Method for the Analysis of Membrane Disruptive Antimicrobial Activity.
abstract: We describe a microbial flow cytometry method that quantifies within 3 hours antimicrobial peptide (AMP) activity, termed Minimum Membrane Disruptive Concentration (MDC). Increasing peptide concentration positively correlates with the extent of bacterial membrane disruption and the calculated MDC is equivalent to its MBC. The activity of AMPs representing three different membranolytic modes of action could be determined for a range of Gram positive and negative bacteria, including the ESKAPE pathogens, E. coli and MRSA. By using the MDC50 concentration of the parent AMP, the method provides high-throughput, quantitative screening of AMP analogues. A unique feature of the MDC assay is that it directly measures peptide/bacteria interactions and lysed cell numbers rather than bacteria survival as with MIC and MBC assays. With the threat of multi-drug resistant bacteria, this high-throughput MDC assay has the potential to aid in the development of novel antimicrobials that target bacteria with improved efficacy.
authors.affiliation: Oral Health Cooperative Research Centre, Melbourne Dental School, Bio21 Institute, The University of Melbourne, Melbourne, Australia.
authors.forename: Neil M
authors.initials: NM
authors.lastname: O'Brien-Simpson
authors.affiliation: Oral Health Cooperative Research Centre, Melbourne Dental School, Bio21 Institute, The University of Melbourne, Melbourne, Australia.
authors.forename: Namfon
authors.initials: N
authors.lastname: Pantarat
authors.affiliation: Oral Health Cooperative Research Centre, Melbourne Dental School, Bio21 Institute, The University of Melbourne, Melbourne, Australia.
authors.forename: Troy J
authors.initials: TJ
authors.lastname: Attard
authors.affiliation: Oral Health Cooperative Research Centre, Melbourne Dental School, Bio21 Institute, The University of Melbourne, Melbourne, Australia.
authors.forename: Katrina A
authors.initials: KA
authors.lastname: Walsh
authors.affiliation: Oral Health Cooperative Research Centre, Melbourne Dental School, Bio21 Institute, The University of Melbourne, Melbourne, Australia.
authors.forename: Eric C
authors.initials: EC
authors.lastname: Reynolds
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology; D023181:Antimicrobial Cationic Peptides / Q000494:pharmacology*; D002462:Cell Membrane / Q000187:drug effects*; D004926:Escherichia coli / Q000187:drug effects; D005434:Flow Cytometry / Q000379:methods*; D006090:Gram-Negative Bacteria / Q000187:drug effects*; D006094:Gram-Positive Bacteria / Q000187:drug effects*; D057166:High-Throughput Screening Assays / Q000379:methods; D055624:Methicillin-Resistant Staphylococcus aureus / Q000187:drug effects; D008826:Microbial Sensitivity Tests / Q000379:methods
chemical_list: D000900:Anti-Bacterial Agents; D023181:Antimicrobial Cationic Peptides