A permeability-increasing drug synergizes with bacterial efux pump inhibitors and restores susceptibility to antibiotics in multi-drug resistant Pseudomonas aeruginosa strains

Resistance to antibiotics poses a major global threat according to the World Health Organization. Restoring the activity of existing drugs is an attractive alternative to address this challenge. One of the most efficient mechanisms of bacterial resistance involves the expression of efflux pump systems capable of expelling antibiotics from the cell. Although there are efflux pump inhibitors (EPIs) available, these molecules are toxic for humans. We hypothesized that permeability-increasing antimicrobial peptides (AMPs) could lower the amount of EPI necessary to sensitize bacteria to antibiotics that are efflux substrates. To test this hypothesis, we measured the ability of polymyxin B nonapeptide (PMBN), to synergize with antibiotics in the presence of EPIs. Assays were performed using planktonic and biofilm-forming cells of Pseudomonas aeruginosa strains overexpressing the MexAB-OprM efflux system. Synergy between PMBN and EPIs boosted azithromycin activity by a factor of 2,133 and sensitized P. aeruginosa to all tested antibiotics. This reduced several orders of magnitude the amount of inhibitor needed for antibiotic sensitization. The selected antibiotic-EPI-PMBN combination caused a 10 million-fold reduction in the viability of biofilm forming cells. We proved that AMPs can synergize with EPIs and that this phenomenon can be exploited to sensitize bacteria to antibiotics

High-Density Targeting of a Viral Multifunctional Nanoplatform to a Pathogenic, Biofilm-Forming Bacterium

SummaryNanomedicine directed at diagnosis and treatment of infections can benefit from innovations that have substantially increased the variety of available multifunctional nanoplatforms. Here, we targeted a spherical, icosahedral viral nanoplatform to a pathogenic, biofilm-forming bacterium, Staphylococcus aureus. Density of binding mediated through specific protein-ligand interactions exceeded the density expected for a planar, hexagonally close-packed array. A multifunctionalized viral protein cage was used to load imaging agents (fluorophore and MRI contrast agent) onto cells. The fluorescence-imaging capability allowed for direct observation of penetration of the nanoplatform into an S. aureus biofilm. These results demonstrate that multifunctional nanoplatforms based on protein cage architectures have significant potential as tools for both diagnosis and targeted treatment of recalcitrant bacterial infections

Rapid Diffusion of Fluorescent Tracers into Staphylococcus epidermidis Biofilms Visualized by Time Lapse Microscopy

The transient diffusion of fluorescent tracers into biofilm cell clusters of Staphylococcus epidermidis was visualized by time lapse confocal scanning laser microscopy. Rhodamine B diffused into the center of cell clusters that were 200 to 600 μm in diameter within a few minutes. The apparent effective diffusion coefficient calculated from these data averaged 3.7 × 10(−7) cm(2) s(−1) or 11% of the value in pure water. Fluorescein diffused into biofilm more rapidly, with a diffusion coefficient that averaged 1.6 × 10(−6) cm(2) s(−1), or 32% of the value in water. This study provides direct, visual confirmation that solutes the size of many antibiotics and biocides can diffuse rapidly into biofilms

Spatial and Temporal Patterns of Biocide Action against Staphylococcus epidermidis Biofilms▿ ‡

The dynamic antimicrobial action of chlorine, a quaternary ammonium compound, glutaraldehyde, and nisin within biofilm cell clusters of Staphylococcus epidermidis was investigated using time-lapse confocal scanning laser microscopy. The technique allowed for the simultaneous imaging of changes in biofilm structure and disruption of cellular membrane integrity through the loss of an unbound fluorophore loaded into bacterial cells prior to antimicrobial challenge. Each of the four antimicrobial agents produced distinct spatial and temporal patterns of fluorescence loss. The antimicrobial action of chlorine was localized around the periphery of biofilm cell clusters. Chlorine was the only antimicrobial agent that caused any biofilm removal. Treatment with the quaternary ammonium compound caused membrane permeabilization that started at the periphery of cell clusters, then migrated steadily inward. A secondary pattern superimposed on the penetration dynamic suggested a subpopulation of less-susceptible cells. These bacteria lost fluorescence much more slowly than the majority of the population. Nisin caused a rapid and uniform loss of green fluorescence from all parts of the biofilm without any removal of biofilm. Glutaraldehyde caused no biofilm removal and also no loss of membrane integrity. Measurements of biocide penetration and action time at the center of cell clusters yielded 46 min for 10 mg liter−1 chlorine, 21 min for 50 mg liter−1 chlorine, 25 min for the quaternary ammonium compound, and 4 min for nisin. These results underscore the distinction between biofilm removal and killing and reinforce the critical role of biocide reactivity in determining the rate of biofilm penetration

Gel-Entrapped Staphylococcus aureus Bacteria as Models of Biofilm Infection Exhibit Growth in Dense Aggregates, Oxygen Limitation, Antibiotic Tolerance, and Heterogeneous Gene Expression

Cathedral, model of the nave; Gaudí’s masterpiece, and, although only partially completed, it is among the most impressive buildings of the 20th century. He took charge of the works at the age of 31 and continued for the rest of his life; it thus summarizes his evolution as an architect (from Modernisme to a mature individualism) as well as the increasing depth of his spiritual conviction, measured in terms of his increasing dedication to the task. His assistants on the project included Jujol, Berenguer, Rubió, the sculptor Lorenzo Matamala i Pinyol (1856-1927), Gaudí’s friend and site supervisor, and Carlos Mani i Roig (1866-1911). The concept of a church dedicated to the Holy Family was widened to become a cathedral for the new metropolitan areas of Barcelona, and del Villar’s neo-Gothic design had been under construction for more than a year when Gaudí took over in 1883. The towers and the church show Gaudí’s structural aesthetic at work in the idiom of the Gothic cathedral, eliminating the need for flying buttresses. The triple portals (1903) of the Nativity façade, represent Faith, Hope and Charity. The four great facade towers are named after the Apostles: from south to north, Barnabas, Simon, Thaddeus and Matthew. Only Barnabas was completed before Gaudí’s death. The project is scheduled to be completed in 2026, the 100th anniversary of Gaudi's death. Source: Grove Art Online; http://www.oxfordartonline.com/ (accessed 7/16/2010

Vesicle Self-Assembly of Amphiphilic Siderophores Produced by Bacterial Isolates from Soap Lake, WA

Soap Lake, located in Washington State, is a meromictic soda lake which was the subject of a prior NSF funded Microbial Observatory. Several organisms inhabiting this lake have been identified as producers of siderophores that are unique in structure. Two isolates found to be of the species Halomonas, SL01 and SL28 were found to produce suites of amphiphilic siderophores consisting of a peptidic head-group, which binds iron appended to fatty acid moieties of various lengths. The ability for siderophores to self-assemble into vesicles was determined for three suites of amphiphilic siderophores of unique structure (two from SL01 and one from SL28). These siderophores resemble the amphiphilic aquachelin siderophores produced by Halomonas aquamarina strain DS40M3, a marine bacterium. Vesicle self-assembly studies were performed by dynamic light scattering (DLS) and epifluorescence microscopy. The addition of ferric iron (FeThe accepted manuscript in pdf format is listed with the files at the bottom of this page. The presentation of the authors' names and (or) special characters in the title of the manuscript may differ slightly between what is listed on this page and what is listed in the pdf file of the accepted manuscript; that in the pdf file of the accepted manuscript is what was submitted by the author

Diffusion of Macromolecules in Model Oral Biofilms▿

The diffusive penetration of fluorescently tagged macromolecular solutes into model oral biofilms was visualized by time-lapse microscopy. All of the solutes tested, including dextrans, proteases, green fluorescent protein, and immunoglobulin G, accessed the interior of cell clusters 100 to 200 μm in diameter within 3 min or less