Bacteriophages evolve enhanced persistence to a mucosal surface.

The majority of viruses within the gut are obligate bacterial viruses known as bacteriophages (phages). Their bacteriotropism underscores the study of phage ecology in the gut, where they modulate and coevolve with gut bacterial communities. Traditionally, these ecological and evolutionary questions were investigated empirically via in vitro experimental evolution and, more recently, in vivo models were adopted to account for physiologically relevant conditions of the gut. Here, we probed beyond conventional phage-bacteria coevolution to investigate potential tripartite evolutionary interactions between phages, their bacterial hosts, and the mammalian gut mucosa. To capture the role of the mammalian gut, we recapitulated a life-like gut mucosal layer using in vitro lab-on-a-chip devices (to wit, the gut-on-a-chip) and showed that the mucosal environment supports stable phage-bacteria coexistence. Next, we experimentally coevolved lytic phage populations within the gut-on-a-chip devices alongside their bacterial hosts. We found that while phages adapt to the mucosal environment via de novo mutations, genetic recombination was the key evolutionary force in driving mutational fitness. A single mutation in the phage capsid protein Hoc-known to facilitate phage adherence to mucus-caused altered phage binding to fucosylated mucin glycans. We demonstrated that the altered glycan-binding phenotype provided the evolved mutant phage a competitive fitness advantage over its ancestral wild-type phage in the gut-on-a-chip mucosal environment. Collectively, our findings revealed that phages-in addition to their evolutionary relationship with bacteria-are able to evolve in response to a mammalian-derived mucosal environment.

Tracheal branching in ants is area-decreasing, violating a central assumption of network transport models.

The structure of tubular transport networks is thought to underlie much of biological regularity, from individuals to ecosystems. A core assumption of transport network models is either area-preserving or area-increasing branching, such that the summed cross-sectional area of all child branches is equal to or greater than the cross-sectional area of their respective parent branch. For insects, the most diverse group of animals, the assumption of area-preserving branching of tracheae is, however, based on measurements of a single individual and an assumption of gas exchange by diffusion. Here we show that ants exhibit neither area-preserving nor area-increasing branching in their abdominal tracheal systems. We find for 20 species of ants that the sum of child tracheal cross-sectional areas is typically less than that of the parent branch (area-decreasing). The radius, rather than the area, of the parent branch is conserved across the sum of child branches. Interpretation of the tracheal system as one optimized for the release of carbon dioxide, while readily catering to oxygen demand, explains the branching pattern. Our results, together with widespread demonstration that gas exchange in insects includes, and is often dominated by, convection, indicate that for generality, network transport models must include consideration of systems with different architectures.

Capacitive Sensing for Monitoring of Microfluidic Protocols Using Nanoliter Dispensing and Acoustic Mixing.

The development of protocols for bio/chemical reaction requires alternate dispensing and mixing steps. While most microfluidic systems use the opening of additional parts of the channel to allow the ingress of fixed volumes of fluid, this requires knowledge of the protocol before the design of the chip. Our approach of using a microfluidic valve to regulate the flow into an initially empty cavity allows for on-chip protocol development and refinement. Mixing is provided by way of surface acoustic wave excitation; this high-frequency vibration causes steady-state streaming flows. We show that capacitive sensing can be used to measure fluid levels, even if multiple fluid types are used, such that nanoliter dispensing accuracy is achieved. Also, the capacitive readout can be used to establish mixing quality and to monitor temperature fluctuations. These capabilities allow for protocols to be conducted without optical assessment and thus will allow for multiplexing, such that reactions could be conducted, simultaneously, in multiple chambers across a chip.

Paper-Based Acoustofluidics for Separating Particles and Cells.

Paper is emerging as a versatile platform for automated fluid handling with a broad range of applications in medical diagnostics and analytical chemistry. However, selectively controlling analyte transport in paper to achieve concentration or selection has been a challenge for functional analysis. Here, by combining paper-based microfluidics with acoustics, we present a rapid and powerful method to size dependently control movement of microparticles and cells in paper using surface acoustic waves (SAW). We demonstrate the unique capability of the paper-based SAW approach to trap and concentrate microparticles in paper and release them when required, achieving collection efficiency of over 98%. Given the correlation between collection efficiency, size, and applied power, the paper-based SAW approach is applied to isolate a mixture of microparticles (1.1, 3.2, and 5 µm in diameter) into different regions and also to trap and concentrate human prostate cancer PC3 cells at a predetermined site. This paper-based SAW approach provides opportunities to develop powerful and low-cost selection and analysis tools, capable of processing complex multicomponent samples, with potential applications in medical diagnostics.

Microfluidic Valves for Selective on-Chip Droplet Splitting at Multiple Sites.

We describe a microfluidic system for control of droplet division at two locations using a T-junction and expansion channel which are placed one after another. Droplets generated at a standard T-junction are introduced into the droplet division section of the microchannel. In the first set of experiments, the droplet division section consists of two consecutive identical T-junctions branching from the main channel. With this geometry, we were able to produce daughter droplets only at the first junction while there was no droplet division at the second junction. Resistive network analysis is used to redesign the microchannel geometry with an expansion channel in place of the second junction, to have the same quantity of flow entering in both the junctions. We observed five different regimes of droplet breakup, namely, (1) no droplet breakup in both junctions, (2) droplet breakup in the first junction, (3) droplet breakup in both junctions with higher daughter droplet volume in the first junction, (4) daughter droplet volume higher in the second junction, and (5) intermittent droplet breakup in both the junctions. Under specific flow conditions, droplet interaction with both the junctions is similar. We then showed design requirements for location of microvalves, simulated by deformation of the main channel wall and by experiments to break the droplet.

Correction: Comparison of bulk and microfluidic methods to monitor the phase behaviour of nanoparticles during digestion of lipid-based drug formulations using in situ X-ray scattering.

Correction for 'Comparison of bulk and microfluidic methods to monitor the phase behaviour of nanoparticles during digestion of lipid-based drug formulations using in situ X-ray scattering' by Ben J. Boyd et al., Soft Matter, 2019, 15, 9565-9578.

Communication Aspects of Visible Light Positioning (VLP) Systems Using a Quadrature Angular Diversity Aperture (QADA) Receiver.

Visible light positioning (VLP) is a promising indoor localization system in which light emitting diode (LED) luminaires are used as positioning beacons. Data communication is an essential aspect of any VLP system, as each luminaire must transmit information about its own location to the receiver. The quadrature angular diversity aperture (QADA) is a new receiver designed specifically for VLP systems using angle-of-arrival estimation. Previous QADA research has focused only on positioning and assumed error-free communication. In this paper, we investigate, via simulations and experiment, the actual communication characteristics of a VLP system that uses a QADA receiver. We calculate the signal-to-noise ratio and bit-error-rates for a range of scenarios and demonstrate the impact of the dimensions of the receiver. We show that reliable communication is assured in typical operating scenarios, proving that communication will not be a limiting factor when using QADA in VLP systems.

Rapid Characterization of Multiple-Contact Miscibility: Toward a Slim-Tube on a Chip.

Carbon dioxide enhanced oil recovery (CO2-EOR) has been widely used to improve production from mature oil fields around the world. To be effective, the injected gas and reservoir oil must develop miscibility, which generally requires prolonged contact between the two phases while in relative motion. Thus, identifying whether miscibility is possible is crucial for determining the feasibility of such EOR projects. The current industry-standard method of characterization, the slim-tube, requires weeks of analysis, while alternative methods are unable to infer all routes to miscibility, producing significant overestimates in required pressures. Microfluidic devices have the potential to simplify and speed up the analysis by offering high levels of fluid control and excellent visualization. Recently, high-pressure microfluidic devices etched into glass and exploiting crude oil's natural fluorescence have been successfully demonstrated. Here we focus on designing a microfluidic channel for identifying the development of miscibility. We prove its accuracy for a known ternary fluid system that mimics the true oil-gas system and can be manipulated at room temperature and pressure. Our chip consists of a single channel with several inline pocket structures. The chip is initially flooded with one phase before a second phase is injected via a flow-rate-controlled pump. The first phase is then rapidly displaced in the primary channel, but small samples are retained within the pockets. Over time, these trapped droplets can be observed as they interact with the continuously flowing second phase. When the fluid concentrations meet the conditions for development of miscibility, a dramatic and visually observable change in behavior occurs, allowing for characterization within 2 h.

Coalescence of Surfactant-Stabilized Adjacent Droplets Using Surface Acoustic Waves.

A novel, on-demand microfluidic droplet merging mechanism is presented in this paper. We demonstrate that a narrow beam surface acoustic wave, targeted at the oil buffer, causes nearby surfactant-stabilized droplets to coalesce. The lack of direct exposure of the droplet to the excitation stimulus makes this method ideal for sensitive samples as harm will not occur. This powerful technique works on a straight channel with no special design, is not affected by surfactant concentration and droplet volume hence promises seamless integration into existing microfluidic systems. It offers high-throughput, biologically safe, on-demand droplet merging for applications ranging from fast reaction kinetics to microfluidic high throughput screening. We thoroughly characterize the physical mechanism triggering droplet-droplet coalescence and observe a cutoff distance from the center of the acoustic beam to the droplet-droplet interface after which the merging mechanism does not work anymore. We establish that the most likely mechanism for merging is acoustic streaming induced droplet deformation.

Comparison of bulk and microfluidic methods to monitor the phase behaviour of nanoparticles during digestion of lipid-based drug formulations using in situ X-ray scattering.

The performance of orally administered lipid-based drug formulations is crucially dependent on digestion, and understanding the colloidal structures formed during digestion is necessary for rational formulation design. Previous studies using the established bulk pH-stat approach (Hong et al. 2015), coupled to synchrotron small angle X-ray scattering (SAXS), have begun to shed light on this subject. Such studies of digestion using in situ SAXS measurements are complex and have limitations regarding the resolution of intermediate structures. Using a microfluidic device, the digestion of lipid systems may be monitored with far better control over the mixing of the components and the application of enzyme, thereby elucidating a finer understanding of the structural progression of these lipid systems. This work compares a simple T-junction microcapillary device and a custom-built microfluidic chip featuring hydrodynamic flow focusing, with an equivalent experiment with the full scale pH-stat approach. Both microfluidic devices were found to be suitable for in situ SAXS measurements in tracking the kinetics with improved time and signal sensitivity compared to other microfluidic devices studying similar lipid-based systems, and producing more consistent and controllable structural transformations. Particle sizing of the nanoparticles produced in the microfluidic devices were more consistent than the pH-stat approach.

Acoustic fields and microfluidic patterning around embedded micro-structures subject to surface acoustic waves.

Recent research has shown that interactions between acoustic waves and microfluidic channels can generate microscale interference patterns with the application of a traveling surface acoustic wave (SAW), effectively creating standing wave patterns with a traveling wave. Forces arising from this interference can be utilized for precise manipulation of micron-sized particles and biological cells. The patterns that have been produced with this method, however, have been limited to straight lines and grids from flat channel walls, and where the spacing resulting from this interference has not previously been comprehensively explored. In this work we examine the interaction between both straight and curved channel interfaces with a SAW to derive geometrically deduced analytical models. These models predict the acoustic force-field periodicity near a channel interface as a function of its orientation to an underlying SAW, and are validated with experimental and simulation results. Notably, the spacing is larger for flat walls than for curved ones and is dependent on the ratio of sound speeds in the substrate and fluid. Generating these force-field gradients with only travelling waves has wide applications in acoustofluidic systems, where channel interfaces can potentially support a range of patterning, concentration, focusing and separation activities by creating locally defined acoustic forces.

Indoor Visible Light Positioning: Overcoming the Practical Limitations of the Quadrant Angular Diversity Aperture Receiver (QADA) by Using the Two-Stage QADA-Plus Receiver.

Visible light positioning (VLP), using LED luminaires as beacons, is a promising solution to the growing demand for accurate indoor positioning. In this paper, we introduce a two-stage receiver that has been specifically designed for VLP. This receiver exploits the advantages of two different VLP receiver types: photodiodes and imaging sensors. In this new receiver design a quadrant angular diversity aperture (QADA) receiver is combined with an off-the-shelf camera to form a robust new receiver called QADA-plus. Results are presented for QADA that show the impact of noise and luminaire geometry on angle of arrival estimation accuracy and positioning accuracy. Detailed discussions highlight other potential sources of error for the QADA receiver and explain how the two-stage QADA-plus can overcome these issues.

Delivery of femtolitre droplets using surface acoustic wave based atomisation for cryo-EM grid preparation.

Cryo-Electron Microscopy (cryo-EM) has become an invaluable tool for structural biology. Over the past decade, the advent of direct electron detectors and automated data acquisition has established cryo-EM as a central method in structural biology. However, challenges remain in the reliable and efficient preparation of samples in a manner which is compatible with high time resolution. The delivery of sample onto the grid is recognized as a critical step in the workflow as it is a source of variability and loss of material due to the blotting which is usually required. Here, we present a method for sample delivery and plunge freezing based on the use of Surface Acoustic Waves to deploy 6-8 µm droplets to the EM grid. This method minimises the sample dead volume and ensures vitrification within 52.6 ms from the moment the sample leaves the microfluidics chip. We demonstrate a working protocol to minimize the atomised volume and apply it to plunge freeze three different samples and provide proof that no damage occurs due to the interaction between the sample and the acoustic waves.

High angular resolution visible light positioning using a quadrant photodiode angular diversity aperture receiver (QADA).

The increasing use of white LEDs for indoor illumination provides a significant opportunity for Visible Light Positioning (VLP). The challenge is to design a small, unobtrusive sensor that can be incorporated into mobile devices to provide accurate measurements for triangulation. We present experimental results for a novel angle of arrival (AOA) detector that has been designed for use in a VLP system. The detector is composed of a transparent aperture in an opaque screen that is located above a quadrant photodiode (PD), separated by a known vertical distance. Light passing through the aperture from an LED casts a light spot onto the quadrant PD. The position of this spot, coupled with knowledge of the height of the aperture above the quadrant PD, provides sufficient information to determine both the incident and polar angles of the light. Experiments, using a prototype detector, show that detector is capable of accurate estimation of AOA. The root mean square errors (rMSE) were less than 0.11° for all the measured positions on the test bed, with 90% of positions having an rMSE of less than 0.07°.

Self-Aligned Acoustofluidic Particle Focusing and Patterning in Microfluidic Channels from Channel-Based Acoustic Waveguides.

Acoustic fields have been widely used for manipulation of particles and cells within microfluidic systems. In this Letter, we explore a novel acoustofluidic phenomenon for particle patterning and focusing, where a periodic acoustic pressure field is produced parallel to internal channel boundaries with the imposition of either a traveling or standing surface acoustic wave (SAW). This effect results from the propagation and intersection of edge waves from the channel walls according to the Huygens-Fresnel principle and classical wave fronts from the substrate-fluid interface. We demonstrate versatile control over this effect to produce both one- and two-dimensional acoustic patterning from one-dimensional SAW fields and its utility for continuous particle focusing. Uniquely, this channel-guided acoustic focusing permits the generation of robust acoustic fields without channel resonance conditions and particle focusing positions that are difficult or impossible to produce otherwise.

Huygens-Fresnel Acoustic Interference and the Development of Robust Time-Averaged Patterns from Traveling Surface Acoustic Waves.

Periodic pattern generation using time-averaged acoustic forces conventionally requires the intersection of counterpropagating wave fields, where suspended micro-objects in a microfluidic system collect along force potential minimizing nodal or antinodal lines. Whereas this effect typically requires either multiple transducer elements or whole channel resonance, we report the generation of scalable periodic patterning positions without either of these conditions. A single propagating surface acoustic wave interacts with the proximal channel wall to produce a knife-edge effect according to the Huygens-Fresnel principle, where these cylindrically propagating waves interfere with classical wave fronts emanating from the substrate. We simulate these conditions and describe a model that accurately predicts the lateral spacing of these positions in a robust and novel approach to acoustic patterning.

Droplet Bouncing and Breakup during Impact on a Microgrooved Surface.

We experimentally investigate the impact dynamics of a microliter water droplet on a hydrophobic microgrooved surface. The surface is fabricated using photolithography, and high-speed visualization is employed to record the time-varying droplet shapes in the transverse and longitudinal directions. The effect of the pitch of the grooved surface and Weber number on the droplet dynamics and impact outcome are studied. At low pitch and Weber number, the maximum droplet spreading is found to be greater in the longitudinal direction than the transverse direction to the grooves. The preferential spreading inversely scales with the pitch at a given Weber number. In this case, the outcome is no bouncing (NB); however, this changes at larger pitch or Weber number. Under these conditions, the following outcomes are obtained as a function of the pitch and Weber number: droplet completely bounces off the surface (CB), bouncing occurs with droplet breakup (BDB), or no bouncing because of a Cassie to Wenzel wetting transition (NBW). In BDB and NBW, the liquid partially or completely penetrates the grooves beneath the droplet as a result of the wetting transition. The former results in droplet breakup alongside bouncing, while the latter suppresses the bouncing. These outcomes are demarcated on the Weber number-dimensionless pitch plane, and the proposed regime map suggests the existence of a critical Weber number or pitch for the transition from one regime to the other. CB and BDB are quantified by plotting the coefficient of restitution of the bouncing droplet and the volume of the daughter droplet left on the surface, respectively. The critical Weber number needed for the transition from CB to BDB is estimated using an existing mathematical model and is compared with the measurements. The comparison is good and provides insights into the mechanism of liquid penetration into the grooves. The present results on microgrooved surfaces are compared with published results on micropillared surfaces in order to assess the water-repelling properties of the two surfaces.

Droplet Manipulation Using Acoustic Streaming Induced by a Vibrating Membrane.

We present a simple method for on-demand manipulation of aqueous droplets in oil. With numerical simulations and experiments, we show that a vibrating membrane can produce acoustic streaming. By making use of this vortical flow, we manage to repulse the droplets away from the membrane edges. Then, by simply aligning the membrane at 45° to the flow, the droplets can be forced to follow the membrane's boundaries, thus steering them across streamlines and even between different oil types. We also characterize the repulsion and steering effect with various excitation voltages at different water and oil flow rates. The maximum steering frequency we have achieved is 165 Hz. The system is extremely robust and reliable: the same membrane can be reused after many days and with different oils and/or surfactants at the same operating frequency.

Shear Assisted Electrochemical Exfoliation of Graphite to Graphene.

The exfoliation characteristics of graphite as a function of applied anodic potential (1-10 V) in combination with shear field (400-74 400 s(-1)) have been studied in a custom-designed microfluidic reactor. Systematic investigation by atomic force microscopy (AFM) indicates that at higher potentials thicker and more fragmented graphene sheets are obtained, while at potentials as low as 1 V, pronounced exfoliation is triggered by the influence of shear. The shear-assisted electrochemical exfoliation process yields large (∼10 µm) graphene flakes with a high proportion of single, bilayer, and trilayer graphene and small ID/IG ratio (0.21-0.32) with only a small contribution from carbon-oxygen species as demonstrated by X-ray photoelectron spectroscopy measurements. This method comprises intercalation of sulfate ions followed by exfoliation using shear induced by a flowing electrolyte. Our findings on the crucial role of hydrodynamics in accentuating the exfoliation efficiency suggest a safer, greener, and more automated method for production of high quality graphene from graphite.