Biology subjects

Stewart, E. J.

Publications and source records attributed to Stewart, E. J..

2 recordsLinked to original sources

bioRxiv2026

In situ three-dimensional mapping of oxygen gradients in Staphylococcus epidermidis biofilms using a solution-based, ratiometric imaging platform

Staphylococcus epidermidis biofilm oxygen gradients are spatially mapped during biofilm development and after vancomycin treatment using a solution-based ratiometric imaging platform. By integrating the oxygen-sensitive, tris(2,2-bipyridyl)dichlororuthenium(II) hexahydrate with oxygen-insensitive, water-soluble CdSe/ZnS quantum dots, we achieve in situ microscale resolution of dissolved oxygen (DO) concentrations within biofilms. The oxygen-sensing platform is calibrated within alginate hydrogels to mimic probe confinement within biofilms and subsequently validated in biofilms using chemical oxygen depletion (sodium sulfite) and thermal inactivation (60{degrees}C). We demonstrate that the probes do not significantly alter planktonic bacterial growth or biofilm development. Using confocal laser scanning microscopy and quantitative image analysis, 3D microscale oxygen maps of biofilms are visualized and evaluated. During S. epidermidis biofilm development from 12 to 24 hours, average biofilm DO concentrations decrease from 2.62{+/-}0.22 mg/L to 2.02{+/-}0.47 mg/L, corresponding with increased S. epidermidis biofilm biomass and bacterial metabolic activity. While treatment of S. epidermidis biofilms with vancomycin at the minimum inhibitory concentration (MIC) (2 {micro}g/mL) results in negligible DO decreases and biofilm biomass and metabolic activity comparable to untreated biofilms, higher vancomycin concentrations (20, 200 {micro}g/mL) lead to increases in biofilm DO, higher dead-cell biovolumes, and decreased metabolic activity. This work establishes a microscale, solution-based ratiometric platform for quantifying the interplay between biofilm oxygen gradients, structure, and metabolic activity, providing a framework for understanding biofilm resilience during antimicrobial treatment.

microbiology↗

bioRxiv2026

Biophysical properties and phenotypes of cell clusters detached from Staphylococcus epidermidis biofilms after matrix-targeted disruption

Bacterial cells detached from Staphylococcus epidermidis biofilms are found to release predominantly as small oblate clusters ([~]1.9 {micro}m) in both untreated biofilms and biofilms treated with matrix-targeted disruptors. Quantitative image analysis common to colloidal science was applied to quantitatively evaluate the physical properties of 9,147 bacterial clusters detached from S. epidermidis biofilms with and without targeted disruption of individual matrix components (polysaccharides, proteins, extracellular DNA) or solubilization of the extracellular polymeric substances (EPS). Concentrations of S. epidermidis biofilm-detached cells are highest after matrix-targeted disruption of polysaccharides. K-means clustering, an unsupervised machine learning technique, was used to reveal that S. epidermidis biofilm-detached cells are released in five distinct phenotypes: small oblate, mid-sized oblate, large oblate, small spherical, and mid-sized prolate clusters. S. epidermidis biofilm detached cell clusters are predominantly oblate across three size groups (79.5%), with the small oblate phenotype representing 60.1% of cell clusters that have 3.1 {+/-} 1.2 cells per cluster, Euclidean diameters of 1.9 {+/-} 0.4 {micro}m, anisotropy indices of 0.98 {+/-} 0.05, and asphericities of -1.75 {+/-} 0.31 on average. The proportion of S. epidermidis cell clusters within each biofilm-detached cell phenotype differs between matrix-targeted disruptors. There are also variations in the abundance of S. epidermidis biofilm detached cells after matrix-targeted disruption between growth conditions and strains. Evaluating the physical properties of biofilm-detached cells after matrix-targeted disruption is critical to understanding their translocation in fluid flow and susceptibility to the host immune response as well as in evaluating matrix-targeted disruption for biofilm control.

microbiology↗