Extracellular Vesicles and Bacteriophages: New Directions in Environmental Biocolloid Research.

There is a long-standing appreciation among environmental engineers and scientists regarding the importance of biologically derived colloidal particles and their environmental fate. This interest has been recently renewed in considering bacteriophages and extracellular vesicles, which are each poised to offer engineers unique insights into fundamental aspects of environmental microbiology and novel approaches for engineering applications, including advances in wastewater treatment and sustainable agricultural practices. Challenges persist due to our limited understanding of interactions between these nanoscale particles with unique surface properties and their local environments. This review considers these biological particles through the lens of colloid science with attention given to their environmental impact and surface properties. We discuss methods developed for the study of inert (nonbiological) particle-particle interactions and the potential to use these to advance our understanding of the environmental fate and transport of extracellular vesicles and bacteriophages.

Characterizing the Transport and Surface Affinity of Extracellular Vesicles Isolated from Yeast and Bacteria in Well-Characterized Porous Media.

Extracellular vesicles (EVs) are membrane-bounded, nanosized particles, produced and secreted by all biological cell types. EVs are ubiquitous in the environment, operating in various roles including intercellular communication and plant immune modulation. Despite their ubiquity, the role of EV surface chemistry in determining transport has been minimally investigated. Using the zeta (ζ)-potential as a surrogate for surface charge, this work considers the deposition of EVs from the yeast, Saccharomyces cerevisiae, and two bacterial species, Staphylococcus aureus and Pseudomonas fluorescens, in well-characterized porous medium under various background conditions shown to influence the transport of other environmental colloidal particles: ionic strength and humic acid concentration. The affinity of S. cerevisiae EVs for the porous medium (glass beads) appeared to be sensitive to changes in ionic strength, as predicted by colloid stability (Derjaguin, Landau, Verwey, and Overbeek or DLVO) theory, and humic acid concentration, while P. fluorescens EVs deviated from DLVO predictions, suggesting that mechanisms other than charge stabilization may control the deposition of P. fluorescens. Calculations of attachment efficiency from these deposition studies were used to estimate EV transport using a clean-bed filtration model. Based on these calculations, EVs could be transported through such homogeneous porous media up to 15 m.

Investigating and Modeling the Regulation of Extracellular Antibiotic Resistance Gene Bioavailability by Naturally Occurring Nanoparticles.

Extracellular antibiotic resistance genes (eARGs) are widespread in the environment and can genetically transform bacteria. This work examined the role of environmentally relevant nanoparticles (NPs) in regulating eARG bioavailability. eARGs extracted from antibiotic-resistant B. subtilis were incubated with nonresistant recipient B. subtilis cells. In the mixture, particle type (either humic acid coated nanoparticles (HASNPs) or their micron-sized counterpart (HASPs)), DNase I concentration, and eARG type were systematically varied. Transformants were counted on selective media. Particles decreased bacterial growth and eARG bioavailability in systems without nuclease. When DNase I was present (≥5 µg/mL), particles increased transformation via chromosomal (but not plasmid-borne) eARGs. HASNPs increased transformation more than HASPs, indicating that the smaller nanoparticle with greater surface area per volume is more effective in increasing eARG bioavailability. These results were also modeled via particle aggregation theory, which represented eARG-bacteria interactions as transport leading to collision, followed by attachment. Using attachment efficiency as a fitting factor, the model predicted transformant concentrations within 35% of experimental data. These results confirm the ability of NPs to increase eARG bioavailability and suggest that particle aggregation theory may be a simplified and suitable framework to broadly predict eARG uptake.

Exploring the design implications of bacteriophages in mixed suspensions by considering attachment and break-up.

The validity and usefulness of implementing bacteriophages into water treatment systems as agents of targeted bacterial inactivation is yet to be determined. While some concerns are still more purely biological in nature other concerns are still chiefly rooted in design feasibility. This work investigated bacteriophage heteroaggregation, a process whereby phages attach to non-host background particles, to explore different design options for water quality engineers, especially tuning mixing velocity. This was done by adapting batch/mixing assays, originally developed to study inert particle heteroaggregation, to characterize bacteriophage and kaolinite heteroaggregation using modified Smoluchowski parameters under different ionic strength conditions. This work found that regardless of the ionic strength or the tested phage to kaolinite ratios heteroaggregation occurred rapidly and was likely driven by extended DLVO forces. A model of bacteriophage-kaolinite heteroaggregation was generated and showed promising correspondence with observed laboratory data. This model, along with other findings, suggests that should bacteriophages be utilized as agents of host inactivation they ought to be used following particle separation processes to reduce the likelihood of phage scavenging through attachment to particulate matter rather than the targeted bacteria.

Modeling bacteriophage-induced inactivation of Escherichia coli utilizing particle aggregation kinetics.

Targeted inactivation of bacteria using bacteriophages has been proposed in applications ranging from bioengineering and biofuel production to medical treatments. The ability to differentiate between desirable and undesirable organisms, such as in targeting filamentous bacteria in activated sludge, is a potential advantage over conventional disinfectants. Like conventional disinfectants, bacteriophages exhibit non-linear concentration-time (Ct) dynamics in achieving bacterial inactivation. However, there is currently no workable model for predicting these observed non-linear inactivation rates. This work considers an approach to predicting bacteriophage-induced inactivation rates by utilizing classical particle aggregation theory. Bacteriophage-bacteria interactions are represented as a two-step process of transport by Brownian motion, differential settling, and shear, followed by attachment. Modifying classical expressions for particle-particle aggregation to include bacterial growth, death, and bacteriophage reproduction, the model was calibrated and validated using literature data. The calibrated model captures much of the observed non-linearity in inactivation rates and reasonably predicts the final host concentration. This model was shown to be most useful in systems more likely to reflect an industrial setting, where the initial multiplicity of infection, or MOI (the ratio of bacteriophage to host organisms), was 1 or greater. For systems of an initial MOI of less than 1 the model showed increased sensitivity to changes in input parameters and a less pronounced ability to reasonably predict inactivation rates.

Ultrathin graphene oxide-based hollow fiber membranes with brush-like CO2-philic agent for highly efficient CO2 capture.

Among the current CO2 capture technologies, membrane gas separation has many inherent advantages over other conventional techniques. However, fabricating gas separation membranes with both high CO2 permeance and high CO2/N2 selectivity, especially under wet conditions, is a challenge. In this study, sub-20-nm thick, layered graphene oxide (GO)-based hollow fiber membranes with grafted, brush-like CO2-philic agent alternating between GO layers are prepared by a facile coating process for highly efficient CO2/N2 separation under wet conditions. Piperazine, as an effective CO2-philic agent, is introduced as a carrier-brush into the GO nanochannels with chemical bonding. The membrane exhibits excellent separation performance under simulated flue gas conditions with CO2 permeance of 1,020 GPU and CO2/N2 selectivity as high as 680, demonstrating its potential for CO2 capture from flue gas. We expect this GO-based membrane structure combined with the facile coating process to facilitate the development of ultrathin GO-based membranes for CO2 capture.

Investigation of interfacial properties of pure and mixed poloxamers for surfactant-mediated shear protection of mammalian cells.

The Poloxamer family of surfactants are commonly used in the biopharmaceutical industry as cell culture media additives to protect cells from the turbulent environment of sparged bioreactors. Despite the widespread use of poloxamers in cell culture, their performance as cell protectants varies depending on their physical structure, molecular weight, and batch-to-batch composition. In this study, the interfacial properties of Poloxamer 188 (P188), Poloxamer 407 (P407), and a mixture of P188 and P407 were characterized to investigate the mechanism of surfactant-mediated shear protection of mammalian cells. The foam stability and equilibrium surface tension of these surfactant systems correlated with their ability to mitigate physical damage to cells in a turbulent environment. We demonstrate that while P188 can function as highly effective shear protectant, the presence of a surface-active contaminant can greatly hinder its protective characteristics. P407 was found to function as such an interfacially active "impurity," disrupting shear protection when mixed with P188 by preferentially adsorbing to the gas-liquid and membrane-liquid interface. Addition of surface-active impurities altered the interfacial properties of the surfactant system and could be detected using an equilibrium surface tension assay. The mechanism of disruption by P407 was determined to be independent of cell-to-bubble attachment, suggesting that poloxamer adsorption to and subsequent reinforcement of the cell membrane may play a key role in protecting cells in high shear environments. This investigation contributes to our understanding of the mechanism of surfactant-mediated shear protection of cells and demonstrates that a surface tension assay can be utilized as a screening tool to ensure that poloxamer lots are free of surface active impurities.