Biology subjects

Reimhult, E.

Publications and source records attributed to Reimhult, E..

4 recordsLinked to original sources

bioRxiv2025

Cryo-SEM Reveals Native Architecture and Matrix Complexity in P. aeruginosa Biofilms

Pseudomonas aeruginosa (PA) biofilms resist antibiotics and immune clearance through their multicellular community organization. Yet, the native spatial arrangement of cells and the extracellular matrix (ECM) remain poorly understood, as research has largely focused on genetics and regulatory networks rather than physical structure. This has limited our understanding of how physical structural organization connects to biofilm function. Here, we combine high-pressure-freezing cryo-SEM, depth-resolved confocal microscopy, and quantitative spatial analysis to examine 4-day-old mucoid and PAO1 biofilms in a near-native state. In contrast to the dense cellular aggregates often inferred from dehydrated samples, cryo-SEM revealed bacterial cells as individually embedded within a continuous extracellular matrix. Spatial statistics revealed a preferred intercellular spacing of approximately 1 m, broad spacing distributions, and only weak short-range clustering. Depth-resolved confocal analysis confirmed this sparse organization across larger biofilm volumes and revealed vertical stratification, with the highest bacterial volume fraction near the substrate and lower cell volume fraction toward the biofilm surface. Cryo-SEM further showed that mucoid biofilms contained aligned fibrillar structures within the biofilm interior, whereas PAO1 biofilms exhibited a denser, mesh-like matrix. These findings challenge prevailing views of biofilms as densely packed bacterial aggregates and establish a quantitative framework for understanding antimicrobial tolerance, cell-cell interactions, nutrient access, and biofilm mechanics in both clinically and environmentally relevant contexts.

microbiology↗

bioRxiv2025

Beyond the Matrix: Rethinking Antibiotic Tolerance in CF Biofilms Using 3D Models

Chronic lung infections in cystic fibrosis (CF) patients are associated with Pseudomonas aeruginosa biofilms exhibiting high antibiotic tolerance with no clear explanation. We investigate the role of the biofilm matrix in this antibiotic tolerance using 3D biofilm models based on acetylated alginate and DNA, mimicking mucoid biofilms. Printed from these bioinks seeded with P. aeruginosa (PAO1), these models support robust microcolony formation as observed in vivo and enable high-throughput assessment of antibiotic diffusion and efficacy. Surprisingly, antibiotic diffusion is not significantly impeded by acetylation or DNA incorporation. Despite this, bacterial tolerance increases tremendously upon encapsulation in alginate. Acetylation further enhances tolerance, particularly to tobramycin, ciprofloxacin, and colistin. The addition of DNA mitigates this effect in a drug-specific manner. While mucoid biofilms, in contrast to the biofilm models, show significant retardation of antibiotic penetration, they also get saturated with all tested antibiotics within 20 h. This demonstrates that direct interaction with alginate or DNA does not explain the slow diffusion of antibiotics in mucoid P. aeruginosa biofilms. Our findings challenge the view that diffusion limitation or antibiotics binding by biofilm exopolysaccharides dominate biofilm resilience and highlight the need to target matrix-induced bacterial adaptation in the development of antibiofilm therapies.

microbiology↗

bioRxiv2024

The native glycocalyx is an ordered, self-assembled hierarchical micro- and nanoarray lamellar structure conserved in evolution

The native ultrastructure of the glycocalyx remained unknown despite its functional importance in cellular recognition/adhesion and selective filtration. The major components of this universal extracellular coat, mucins, proteoglycans, glyconectins, and hyaluronan, share similar physicochemical properties of high molecular weight, glycan richness, and amply hydrated bottlebrush polymer morphologies with comparable intramolecular anionic charge distribution. The diversity of these glycoconjugate intermolecular binding under physiologically highly hydrated and specific ionic conditions keeps the native glycocalyx structure enabling it to function. Irrespective of the intricacy of the glycocalyx physiological milieu preservation and molecular organization, only a dehydrated non-native state presenting an artefactual unorganized fiber mesh was imaged. Using cryo-SEM after cryo-preservation with minimal sublimation to conserve water, ion distribution, and the native intermolecular interactions, we unveil well-organized lamellae of glycoconjugates self-assembled in hierarchical micro- and nanoarrays for the glycocalyx of human cell and self-assembled glyconectin glycocalyx from an evolutionary most distant sponge despite differences in sequence and composition. Our combined AFM binding strength measurements and cryo-SEM imply that evolutionarily preserved glycocalyx micro- and nano-morphologies are formed by thermodynamically driven self-assembly of glycoconjugates having similar physico-chemical properties. TeaserThe extracellular glycocalyx coat is a self-organizing ultrastructure in human and sponge cells.

cell biology↗

bioRxiv2024

Adhesion of E. coli bacteria is force-modulated due to fimbriae-mediated surface repulsion and multivalent binding irrespective of surface specificity

Escherichia coli bacteria that express type 1 fimbriae migrate along surfaces when pushed by a slow flow but stick more firmly when the flow increases. This and other examples of force-modulated biological binding are often described as due to lectin-glycan catch-bonds. Here we quantitatively track the 3D movements of fimbriated E. coli flowing over surfaces nanopatterned with mannose or hydrophobic binding sites. We reveal that flow-modulated surface adhesion and motion are consequences of bacteria adhering via polydisperse, elastic fimbriae, irrespective of binding affinity and specificity. The fimbria-mediated surface repulsion and the flow forces on tethered bacteria establish an equilibrium bacteria-surface separation. The separation controls the number of potential tethers between the bacterium and the surface. Combined with the individual fimbria affinity, this determines the surface avidity and surface motion. This provides a broadly applicable mechanism by which bacteria acquire adaptive surface avidity, responding super-selectively to different flow environments, concentration, and affinity of available binding sites, essential to explaining how fimbriae govern tropism and surface colonization.

microbiology↗