Hybrid surfaces combining electropolymerization and lithography: fabrication and wetting properties.

In the current work, we developed a novel method to fabricate hybrid surfaces consisting of mixed hydrophilic/superhydrophobic properties. These surfaces specifically consist of a regular array of hydrophilic pillars (displaying a receding contact angle lower than 90°) surrounded by a superhydrophobic thinner layer made via the electropolymerization of a fluorinated monomer. Then, we determined the wetting properties of various forms of this complex surface, i.e., displaying different surface properties, by specifically determining their advancing (θa) and receding (θr) contact angles. Two main parameters were varied: the pillar density (from 21.2% to 6.5% based on using a spacing d between pillars varying from 25 to 45 micrometers) and the polymer charge density (from 0 to 100 mC cm-2). We observed that, for low charge density values, only the ground surface was covered by the hydrophobic polymers; while for higher charge density values, polymerization reached higher levels on the lateral surfaces of the nonconductive cylindrical pillars, eventually up to their top surfaces and covering them for the highest charge densities. This feature gave us an additional parameter that we could use to control the surface wettability. We also found that contact angles (advancing and receding) increased markedly with increasing polymer charge density above a critical value (which was higher for receding angles). And we measured advancing and receding contact angles to, respectively, increase and decrease with increasing pillar density. We interpreted qualitatively these behaviors, the main point being the importance of the impalement (null, partial or total).

Capillary bridge technique to study superhydrophobic surfaces.

We present here the use of the capillary bridge technique to study the wetting properties (advancing and receding contact angles) of transparent, textured and superhydrophobic surfaces over large wetted area. Apparent contact angles on such surfaces are classically measured using a goniometer in combination with video camera side visualization and a drop shape analysis. Recent experiments of Schellenberger et al. [F. Schellenberger, N. Encinas, D. Vollmer and H. J. Butt, Phys. Rev. Lett., 2016, 116(9), 096101] show that this method can significantly underestimate the apparent advancing contact angle. We use for the first time the capillary bridge setup for such textured surfaces, leading to a large (up to several cm2) wetted area, instead of having a reduced contact zone as in the drop case (mm2 or less). (1) We show here how to use the method and its characteristics to explore the wetting properties of superhydrophobic surfaces. We have developed a new analysis method in order to obtain the value of the contact angle for any position of the substrate. (2) We compare with the classical drop side view method, showing that advancing contact angles are systematically higher. (3) We compare to a few existing models, concluding a good agreement for receding values but not for advancing angles, for which models must be refined.

Automated high-content image-based characterization of microorganism behavioral diversity and distribution.

Microorganisms have evolved complex systems to respond to environmental signals. Gradients of particular molecules and elemental ions alter the behavior of microbes and their distribution within their environment. Microdevices coupled with automated image-based methods are now employed to analyze the instantaneous distribution and motion behaviors of microbial species in controlled environments at small temporal scales, mimicking, to some extent, macro conditions. Such technologies have so far been adopted for investigations mainly on individual species. Similar versatile approaches must now be developed for the characterization of multiple and complex interactions between a microbial community and its environment. Here, we provide a comprehensive step-by-step method for the characterization of species-specific behavior in a synthetic mixed microbial suspension in response to an environmental driver. By coupling accessible microfluidic devices with automated image analysis approaches, we evaluated the behavioral response of three morphologically different telluric species (Phytophthora parasitica, Vorticella microstoma, Enterobacter aerogenes) to a potassium gradient driver. Using the TrackMate plug-in algorithm, we performed morphometric and then motion analyses to characterize the response of each microbial species to the driver. Such an approach enabled to confirm the different morphological features of the three species and simultaneously characterize their specific motion in reaction to the driver and their co-interaction dynamics. By increasing the complexity of suspensions, this approach could be integrated in a framework for phenotypic analysis in microbial ecology research, helping to characterize how key drivers influence microbiota assembly at microbiota host-environment interfaces.

Cross-linked polymer microparticles with tunable surface properties by the combination of suspension free radical copolymerization and Click chemistry.

We propose a general, versatile and broad in scope two-steps approach for the elaboration of cross-linked polymer microparticles (µPs) with tunable functionalities and surface properties. Surface-functionalized cross-linked polymer µPs with diameter in the 80 µm range are prepared by the combination of: 1) suspension free radical copolymerization of styrene, propargyl methacrylate and 1,6-hexanediol dimethacrylate, 2) subsequent covalent tethering of a variety of azide-functionalized moieties (i.e. rhodamine B fluorescent dye or poly(ethylene glycol) (PEG) brush precursor) by copper(I)-catalyzed azide-alkyne cycloaddition (CuAAC) and, 3) optional N-alkylation of the 1,2,3-triazole groups followed by anion exchange reaction to afford covalently-tethered 1,2,3-triazolium ionic liquids with iodide or cresol red counter-anions. The resulting µPs are characterized by laser diffraction, differential scanning calorimetry, as well as by optical, confocal fluorescence, scanning electron and atomic force microscopies. Finally, the rheological properties of concentrated suspensions (volume fractions of 0.40 and 0.44) of the different synthesized µPs dispersed in a 1:1 (vol/vol) mixture of polyalkylene glycol and water are studied. The modification of µPs surface properties contributes not only to change the stability of the suspensions against flocculation, but also to significantly modify their rheological behavior at high shear stresses. This represents a clear experimental evidence of the importance of non-hydrodynamic contact forces in the rheology of non-Brownian suspensions (NBSs).

Coordination of two opposite flagella allows high-speed swimming and active turning of individual zoospores.

Phytophthora species cause diseases in a large variety of plants and represent a serious agricultural threat, leading, every year, to multibillion dollar losses. Infection occurs when their biflagellated zoospores move across the soil at their characteristic high speed and reach the roots of a host plant. Despite the relevance of zoospore spreading in the epidemics of plant diseases, individual swimming of zoospores have not been fully investigated. It remains unknown about the characteristics of two opposite beating flagella during translation and turning, and the roles of each flagellum on zoospore swimming. Here, combining experiments and modeling, we show how these two flagella contribute to generate thrust when beating together, and identify the mastigonemes-attached anterior flagellum as the main source of thrust. Furthermore, we find that turning involves a complex active process, in which the posterior flagellum temporarily stops, while the anterior flagellum keeps on beating and changes its gait from sinusoidal waves to power and recovery strokes, similar to Chlamydomonas's breaststroke, to reorient its body to a new direction. Our study is a fundamental step toward a better understanding of the spreading of plant pathogens' motile forms, and shows that the motility pattern of these biflagellated zoospores represents a distinct eukaryotic version of the celebrated 'run-and-tumble' motility class exhibited by peritrichous bacteria.

Microorganisms of the Phytophthora genus are serious agricultural pests. They cause diseases in many crops, including potato, onion, tomato, tobacco, cotton, peppers, and citrus. These diseases cause billions of dollars in losses each year. Learning more about how the tiny creatures disseminate and reach host plants could help scientists develop new ways to prevent such crop damage. The spore cells of Phytophthora, also known as zoospores, have two appendages called flagella on their bodies. A tinsel-shaped flagellum is near the front of the creature and a long smooth filament-like flagellum is near the posterior. Zoospores use their flagella to swim at high speeds through liquid toward potential plant hosts. Their complex swimming patterns change in response to different physical, chemical, and electrical signals in the environment. But exactly how they use their flagella to generate these movements is not clear. Tran et al. reveal new details about zoospore locomotion. In the experiments, Tran et al. recorded the movements of zoospores in a tiny 'swimming pool' of fluid on top of a glass slide and analyzed the movements using statistical and mathematical models. The results uncovered coordinated actions of the flagella when zoospores swim in a straight line and when they turn. The tinsel-like front flagellum provides most of the force that propels the zoospore forward. To do this, it beats with an undulating wave pattern. It shifts the beating to a breast-stroke pattern to change direction. The posterior flagellum provides a smaller forward thrust and temporarily pauses during turns. The study provides new details about zoospore's movements that may help scientists develop new strategies to control these pests. It also offers more information about how flagella coordinate their actions to switch speeds or change directions that may be of interest to other scientists studying organisms that use flagella to move.

Guidance of zoospores by potassium gradient sensing mediates aggregation.

The biflagellate zoospores of some phytopathogenic Phytophthora species spontaneously aggregate within minutes in suspension. We show here that Phytophthora parasitica zoospores can form aggregates in response to a K+ gradient with a particular geometric arrangement. Using time-lapse live imaging in macro- and microfluidic devices, we defined (i) spatio-temporal and concentration-scale changes in the gradient, correlated with (ii) the cell distribution and (iii) the metrics of zoospore motion (velocity, trajectory). In droplets, we found that K+-induced aggregates resulted from a single biphasic temporal sequence involving negative chemotaxis followed by bioconvection over a K+ gradient concentration scale [0-17 mM]. Each K+-sensing cell moved into a region in which potassium concentration is below the threshold range of 1-4 mM, resulting in swarming. Once a critical population density had been achieved, the zoospores formed a plume that migrated downward, with fluid advection in its wake and aggregate formation on the support surface. In the microfluidic device, the density of zoospores escaping potassium was similar to that achieved in droplets. We discuss possible sources of K+ gradients in the natural environment (zoospore population, microbiota, plant roots, soil particles), and implications for the events preceding inoculum formation on host plants.

Contrasted enzymatic cocktails reveal the importance of cellulases and hemicellulases activity ratios for the hydrolysis of cellulose in presence of xylans.

Various enzymatic cocktails were produced from two Trichoderma reesei strains, a cellulase hyperproducer strain and a strain with ß-glucosidase activity overexpression. By using various carbon sources (lactose, glucose, xylose, hemicellulosic hydrolysate) for strains growth, contrasted enzymatic activities were obtained. The enzymatic cocktails presented various levels of efficiency for the hydrolysis of cellulose Avicel into glucose, in presence of xylans, or not. These latter were also hydrolyzed with different extents according to cocktails. The most efficient cocktails (TR1 and TR3) on Avicel were richer in filter paper activity (FPU) and presented a low ratio FPU/ß-glucosidase activity. Cocktails TR2 and TR5 which were produced on the higher amount of hemicellulosic hydrolysate, possess both high xylanase and ß-xylosidase activities, and were the most efficient for xylans hydrolysis. When hydrolysis of Avicel was conducted in presence of xylans, a decrease of glucose release occurred for all cocktails compared to hydrolysis of Avicel alone. Mixing TR1 and TR5 cocktails with two different ratios of proteins (1/1 and 1/4) resulted in a gain of efficiency for glucose release during hydrolysis of Avicel in presence of xylans compared to TR5 alone. Our results demonstrate the importance of combining hemicellulase and cellulase activities to improve the yields of glucose release from Avicel in presence of xylans. In this context, strategies involving enzymes production with carbon sources comprising mixed C5 and C6 sugars or combining different cocktails produced on C5 or on C6 sugars are of interest for processes developed in the context of lignocellulosic biorefinery.

Cellulase activity mapping of Trichoderma reesei cultivated in sugar mixtures under fed-batch conditions.

BACKGROUND: On-site cellulase production using locally available lignocellulosic biomass (LCB) is essential for cost-effective production of 2nd-generation biofuels. Cellulolytic enzymes (cellulases and hemicellulases) must be produced in fed-batch mode in order to obtain high productivity and yield. To date, the impact of the sugar composition of LCB hydrolysates on cellulolytic enzyme secretion has not been thoroughly investigated in industrial conditions. RESULTS: The effect of sugar mixtures (glucose, xylose, inducer) on the secretion of cellulolytic enzymes by a glucose-derepressed and cellulase-hyperproducing mutant strain of Trichoderma reesei (strain CL847) was studied using a small-scale protocol representative of the industrial conditions. Since production of cellulolytic enzymes is inducible by either lactose or cellobiose, two parallel mixture designs were performed separately. No significant difference between inducers was observed on cellulase secretion performance, probably because a common induction mechanism occurred under carbon flux limitation. The characteristics of the enzymatic cocktails did not correlate with productivity, but instead were rather dependent on the substrate composition. Increasing xylose content in the feed had the strongest impact. It decreased by 2-fold cellulase, endoglucanase, and cellobiohydrolase activities and by 4-fold ß-glucosidase activity. In contrast, xylanase activity was increased 6-fold. Accordingly, simultaneous high ß-glucosidase and xylanase activities in the enzymatic cocktails seemed to be incompatible. The variations in enzymatic activity were modelled and validated with four fed-batch cultures performed in bioreactors. The overall enzyme production was maintained at its highest level when substituting up to 75% of the inducer with non-inducing sugars. CONCLUSIONS: The sugar substrate composition strongly influenced the composition of the cellulolytic cocktail secreted by T. reesei in fed-batch mode. Modelling can be used to predict cellulolytic activity based on the sugar composition of the culture-feeding solution, or to fine tune the substrate composition in order to produce a desired enzymatic cocktail.

Contact angle and contact angle hysteresis measurements using the capillary bridge technique.

A new experimental technique is proposed to easily measure both advancing and receding contact angles of a liquid on a solid surface, with unprecedented accuracy. The technique is based on the analysis of the evolution of a capillary bridge formed between a liquid bath and a solid surface (which needs to be spherical) when the distance between the surface and the liquid bath is slowly varied. The feasibility of the technique is demonstrated using a low-energy perfluorinated surface with two different test liquids (water and hexadecane). A detailed description of both experimental procedures and computational modeling are given, allowing one to determine contact angle values. It is shown that the origin of the high accuracy of this technique relies on the fact that the contact angles are automatically averaged over the whole periphery of the contact. This method appears to be particularly adapted to the characterization of surfaces with very low contact angle hysteresis.