Effects of Surface Topography and Cellular Biomechanics on Nanopillar-Induced Bactericidal Activity.

Bacteria are mechanically resistant biological structures that can sustain physical stress. Experimental data, however, have shown that high-aspect-ratio nanopillars deform bacterial cells upon contact. If the deformation is sufficiently large, it lyses the bacterial cell wall, ultimately leading to cell death. This has prompted a novel strategy, known as mechano-bactericide technology, to fabricate antibacterial surfaces. Although adhesion forces were originally proposed as the driving force for mechano-bactericidal action, it has been recently shown that external forces, such as capillary forces arising from an air-water interface at bacterial surfaces, produce sufficient loads to rapidly kill bacteria on nanopillars. This discovery highlights the need to theoretically examine how bacteria respond to external loads and to ascertain the key factors. In this study, we developed a finite element model approximating bacteria as elastic shells filled with cytoplasmic fluid brought into contact with an individual nanopillar or nanopillar array. This model elucidates that bacterial killing caused by external forces on nanopillars is influenced by surface topography and cell biomechanical variables, including the density and arrangement of nanopillars, in addition to the cell wall thickness and elastic modulus. Considering that surface topography is an important design parameter, we performed experiments using nanopillar arrays with precisely controlled nanopillar diameters and spacing. Consistent with model predictions, these demonstrate that nanopillars with a larger spacing increase bacterial susceptibility to mechanical puncture. The results provide salient insights into mechano-bactericidal activity and identify key design parameters for implementing this technology.

Gut-on-a-chip models for dissecting the gut microbiology and physiology.

Microfluidic technologies have been extensively investigated in recent years for developing organ-on-a-chip-devices as robust in vitro models aiming to recapitulate organ 3D topography and its physicochemical cues. Among these attempts, an important research front has focused on simulating the physiology of the gut, an organ with a distinct cellular composition featuring a plethora of microbial and human cells that mutually mediate critical body functions. This research has led to innovative approaches to model fluid flow, mechanical forces, and oxygen gradients, which are all important developmental cues of the gut physiological system. A myriad of studies has demonstrated that gut-on-a-chip models reinforce a prolonged coculture of microbiota and human cells with genotypic and phenotypic responses that closely mimic the in vivo data. Accordingly, the excellent organ mimicry offered by gut-on-a-chips has fueled numerous investigations on the clinical and industrial applications of these devices in recent years. In this review, we outline various gut-on-a-chip designs, particularly focusing on different configurations used to coculture the microbiome and various human intestinal cells. We then elaborate on different approaches that have been adopted to model key physiochemical stimuli and explore how these models have been beneficial to understanding gut pathophysiology and testing therapeutic interventions.

Emerging computational paradigms to address the complex role of gut microbial metabolism in cardiovascular diseases.

The human gut microbiota and its associated perturbations are implicated in a variety of cardiovascular diseases (CVDs). There is evidence that the structure and metabolic composition of the gut microbiome and some of its metabolites have mechanistic associations with several CVDs. Nevertheless, there is a need to unravel metabolic behavior and underlying mechanisms of microbiome-host interactions. This need is even more highlighted when considering that microbiome-secreted metabolites contributing to CVDs are the subject of intensive research to develop new prevention and therapeutic techniques. In addition to the application of high-throughput data used in microbiome-related studies, advanced computational tools enable us to integrate omics into different mathematical models, including constraint-based models, dynamic models, agent-based models, and machine learning tools, to build a holistic picture of metabolic pathological mechanisms. In this article, we aim to review and introduce state-of-the-art mathematical models and computational approaches addressing the link between the microbiome and CVDs.

Liquid-infused microstructured bioadhesives halt non-compressible hemorrhage.

Non-compressible hemorrhage is an unmet clinical challenge that accounts for high mortality in trauma. Rapid pressurized blood flows under hemorrhage impair the function and integrity of hemostatic agents and the adhesion of bioadhesive sealants. Here, we report the design and performance of bioinspired microstructured bioadhesives, formed with a macroporous tough xerogel infused with functional liquids. The xerogel can rapidly absorb interfacial fluids such as whole blood and promote blood clotting, while the infused liquids facilitate interfacial bonding, sealing, and antibacterial function. Their synergy enables the bioadhesives to form tough adhesion on ex vivo human and porcine tissues and diverse engineered surfaces without the need for compression, as well as on-demand instant removal and storage stability. We demonstrate a significantly improved hemostatic efficacy and biocompatibility in rats and pigs compared to non-structured counterparts and commercial products. This work opens new avenues for the development of bioadhesives and hemostatic sealants.

Surface Wettability Is a Key Feature in the Mechano-Bactericidal Activity of Nanopillars.

Nanopillar-textured surfaces are of growing interest because of their ability to kill bacteria through physical damage without relying on antimicrobial chemicals. Although research on antibacterial nanopillars has progressed significantly in recent years, the effect of nanopillar hydrophobicity on bactericidal activity remains elusive. In this study, we investigated the mechano-bactericidal efficacy of etched silicon nanopillars against Pseudomonas aeruginosa at nanopillar hydrophobicities from superhydrophilic to superhydrophobic. Assessing cell viability and bacterial morphology in immersed wet conditions, we observed negligible bactericidal activity; however, air/liquid interface displacement during water evaporation established a bactericidal effect that strongly depends on substrate hydrophobicity. Specifically, bactericidal activity was highest on superhydrophilic surfaces but abated with increasing hydrophobicity, diminishing at substrate contact angles larger than 90°. Calculation of the surface tension and Laplace pressure forces during water evaporation for each substrate subsequently highlighted that the total capillary force, as an external driving force responsible for bacterial deformation, is significantly weaker on hydrophobic substrates. These findings suggest that superhydrophilic nanopillared surfaces are a superior choice for mechano-bactericidal activity, whereas superhydrophobic surfaces, although not bactericidal, may have antibiofouling properties through their self-cleaning effect. These findings provide new insights into the design and application of nanopillared surfaces as functional antibacterial materials.

Microfluidic Study of Bacterial Attachment on and Detachment from Zinc Oxide Nanopillars.

Nanopillars can influence how bacterial cells attach to a surface. Herein, we investigated whether self-assembled zinc oxide (ZnO) nanopillars synthesized on glass substrates via the conventional hydrothermal route possess anti-biofouling properties either by reducing the amount of initially attached cells or promoting the detachment of cells from the surface or both. To avoid complications associated with manual intervention methods of assessing bacterial attachment on nanopillar surfaces, we implemented a microfluidic approach. In our study, we synthesized two nanopillar topographies: a low surface density of ZnO nanopillars and a high surface density of ZnO nanopillars. Next, we mounted microfluidic channels to each of these substrates. This microfluidic approach allowed us to gently flow Pseudomonas aeruginosa, Staphylococcus aureus, or Bacillus subtilis cells onto the nanopillars for initial attachment before systematically increasing the flowrate to attempt to detach remaining attached cells without introducing air-liquid interface artefacts during the assay. Generally, initial bacterial attachment was similar across all substrates. However, cells consistently detached more readily from high-surface-density nanopillars compared to low-surface-density nanopillars. Electron microscopy revealed that cells that attached to high-surface-density nanopillars rested atop the nanopillars, fully exposed to microfluidic shear, whereas many cells became trapped in the void space between neighboring low-surface-density nanopillars, shielding these cells from detachment. Our findings indicate that self-assembled ZnO nanopillars can provide anti-biofouling properties under submerged flow but only if synthesized at high surface density.

Ionotronic Tough Adhesives with Intrinsic Multifunctionality.

Ionotronic hydrogels find wide applications in flexible electronics, wearable/implantable devices, soft robotics, and human-machine interfaces. Their performance and practical translation have been bottlenecked by poor adhesiveness, limited mechanical properties, and the lack of biological functions. The remedies are often associated with complex formulations and sophisticated processing. Here, we report a rational design and facile synthesis of ionotronic tough adhesives (i-TAs), which have excellent mechanical, physical, electrical, and biological properties and promise high scalability and translational potential. They consist of an interpenetrating network with high-density amine groups and highly mobile chains, which enable intrinsic adhesiveness, self-healing, ionic stability, cytocompatibility, and antimicrobial functions. The i-TAs in both pristine and swollen states possess high toughness, stretchability, and strong adhesion to diverse substrates such as tissues and elastomers. The superior mechanical performance is achieved simultaneously with high ionic conductivity and stability in electrolyte solutions. We further demonstrate the use of i-TAs as wearable devices, strain sensors, and sensory sealants. This work is expected to open avenues for new ionotronics with novel functions and stimulate the development and translation of ionotronics.

Bioinspired tough gel sheath for robust and versatile surface functionalization.

Sutures pervade surgeries, but their performance is limited by the mechanical mismatch with tissues and the lack of advanced functionality. Existing modification strategies result in either deterioration of suture's bulk properties or a weak coating susceptible to rupture or delamination. Inspired by tendon endotenon sheath, we report a versatile strategy to functionalize fiber-based devices such as sutures. This strategy seamlessly unites surgical sutures, tough gel sheath, and various functional materials. Robust modification is demonstrated with strong interfacial adhesion (>2000 J m-2). The surface stiffness, friction, and drag of the suture when interfacing with tissues can be markedly reduced, without compromising the tensile strength. Versatile functionalization of the suture for infection prevention, wound monitoring, drug delivery, and near-infrared imaging is then presented. This platform technology is applicable to other fiber-based devices and foreseen to affect broad technological areas ranging from wound management to smart textiles.

Highly Absorbent Antibacterial and Biofilm-Disrupting Hydrogels from Cellulose for Wound Dressing Applications.

In this study, a carboxyl-modified cellulosic hydrogel was developed as the base material for wound dressings. ε-poly-l-lysine, a natural polyamide, was then covalently linked to the hydrogel through a bioconjugation reaction, which was confirmed by X-ray photoelectron spectroscopy (XPS) and Fourier transform infrared (FTIR). The antibacterial efficacy of the hydrogel was tested against two model bacteria, Staphylococcus aureus and Pseudomonas aeruginosa, two of the most commonly found bacteria in wound infections. Bacterial viability and biofilm formation after exposure of bacteria to the hydrogels were used as efficacy indicators. Live/Dead assay was used to measure the number of compromised bacteria using a confocal laser scanning microscope. The results show that the antibacterial hydrogel was able to kill approximately 99% of the exposed bacteria after 3 h of exposure. In addition, NIH/3T3 fibroblasts were used to study the biocompatibility of the developed hydrogels. Water-soluble tetrazolium salt (WST)-1 assay was used to measure the metabolic activity of the cells and Live/Dead assay was used to measure the viability of the cells after 24, 48, and 72 h. The developed antibacterial hydrogels are light weight, have a high water-uptake capacity, and show high biocompatibility with the model mammalian cells, which make them a promising candidate to be used for wound dressing applications.

Hydrophilic Mechano-Bactericidal Nanopillars Require External Forces to Rapidly Kill Bacteria.

Nanopillars have been shown to mechanically damage bacteria, suggesting a promising strategy for future antibacterial surfaces. However, the mechanisms underlying this phenomena remain unclear, which ultimately limits translational potential toward real-world applications. Using real-time and end-point analysis techniques, we demonstrate that in contrast to initial expectations, bacteria on multiple hydrophilic "mechano-bactericidal" surfaces remained viable unless exposed to a moving air-liquid interface, which caused considerable cell death. Reasoning that normal forces arising from surface tension may underlie this mechano-bactericidal activity, we developed computational and experimental models to estimate, manipulate, and recreate the impact of these forces. Our experiments together demonstrate that a critical level of external force acting on cells attached to nanopillar surfaces can rapidly deform and rupture bacteria. These studies provide fundamental physical insight into how nanopillar surfaces can serve as effective antibacterial materials and suggest use-conditions under which such nanotechnology approaches may provide practical value.

Anodized Aluminum with Nanoholes Impregnated with Quaternary Ammonium Compounds Can Kill Pathogenic Bacteria within Seconds of Contact.

Bacterial contamination of surfaces results in the spread of pathogens in public spaces such as hospitals and public transport. The development of antibacterial surfaces that rapidly kill bacteria is therefore highly desirable. Here, we investigate the antibacterial efficacy of a novel anodized aluminum surface featuring nanoholes impregnated with quaternary ammonium compounds, referred to as A3S. The antimicrobial activity of A3S was assessed using both Gram-positive and Gram-negative bacteria in a novel assay which simulates pathogen transfer from a contaminated "finger" to a clean finger in a real-world scenario. Enumeration of colony-forming units shows that the number of viable bacteria on the second "finger" contacting A3S is significantly reduced compared to a control surface. Furthermore, bacterial contact with the A3S material results in compromised cell membranes in less than 1 min, and a kill zone assay shows that an exposure time as short as 5 s is sufficient to kill pathogenic bacteria. The rapid antimicrobial action of A3S was particularly evident against Gram-positive bacteria, that account for more than 70% of nosocomial infections. Taken together, these findings demonstrate that A3S is a promising candidate for the fabrication of antibacterial surfaces that can be used in a wide range of clinical and commercial applications to stop the spread of harmful bacteria.

Protocol for biofilm streamer formation in a microfluidic device with micro-pillars.

Several bacterial species possess the ability to attach to surfaces and colonize them in the form of thin films called biofilms. Biofilms that grow in porous media are relevant to several industrial and environmental processes such as wastewater treatment and CO2 sequestration. We used Pseudomonas fluorescens, a Gram-negative aerobic bacterium, to investigate biofilm formation in a microfluidic device that mimics porous media. The microfluidic device consists of an array of micro-posts, which were fabricated using soft-lithography. Subsequently, biofilm formation in these devices with flow was investigated and we demonstrate the formation of filamentous biofilms known as streamers in our device. The detailed protocols for fabrication and assembly of microfluidic device are provided here along with the bacterial culture protocols. Detailed procedures for experimentation with the microfluidic device are also presented along with representative results.

A web of streamers: biofilm formation in a porous microfluidic device.

Using a microfabricated porous media mimic platform, we investigated how fluid flow influences the formation of filamentous structures, known as streamers, between porous media structures. We demonstrate how hydrodynamics govern the formation, morphology and the distribution of these biofilm streamers in the device. Our work establishes that, under favorable hydrodynamic conditions, streamers can often act as precursors to mature microbial structures found in complex geometries, such as those involved in porous media.