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We investigate the effect of micrometer-scale surface wrinkling on the attachment and proliferation of model bacteria (Staphylococcus aureus, Pseudomonas aeruginosa, and Escherichia coli K12) and fungi (Candida albicans). Specifically, sinusoidal (1D), checkerboard (C), and herringbone (H) patterns were fabricated by mechanical wrinkling of plasma-oxidized polydimethylsiloxane (PDMS) bilayers and contrasted with flat (F) surfaces. Microbial deformation and orientation were found to correlate with the aspect ratio and commensurably with surface pattern dimensions and local pattern order. Significantly, the proliferation of P. aeruginosa could be described by a linear scaling between bacterial area coverage and available surface area, defined as a fraction of the line integral along each profile with negative curvature. However, in the early stages of proliferation (up to 6 h examined), that C and H patterns disrupt the spatial arrangement of bacteria, impeding proliferation for several hours and reducing it (by ∼50%) thereafter. Our findings suggest a simple framework to rationalize the impact of micrometer-scale topography on microbial action and demonstrate that multiaxial patterning order provides an effective strategy to delay and frustrate the early stages of bacterial proliferation.

A quartz-crystal-microbalance with dissipation (QCM-D) can measure molecular mass adsorption as well as register adhesion of colloidal particles. However, analysis of the QCM-D output to quantitatively analyze adhesion of (bio) colloids to obtain viscoelastic bond properties is still a subject of debate. Here, we analyze the QCM-D output to analyze the bond between two hydrophilic streptococcal strains 91 nm long and without fibrillar surface appendages and micron-sized hydrophobic polystyrene particles on QCM-D crystal surfaces with different hydrophobicities, using the Kelvin-Voigt model and the Maxwell model. A Poisson distribution was implemented in order to determine the possible virtues of including polydispersity when fitting model parameters to the data. The quality of the fits did not indicate whether the Kelvin-Voigt or the Maxwell model is preferable and only polydispersity in spring-constants improved the fit for polystyrene particles. The Kelvin-Voigt and Maxwell models both yielded higher spring-constants for the bald streptococcus than for the fibrillated one. In both models, the drag coefficients increased for the bald streptococcus with the ratio of electron-donating over electronaccepting parameters of the crystal surface, while for the fibrillated strain the drag coefficient was similar on all crystal surfaces. Combined with the propensity of fibrillated streptococci to bind to the sensor crystal as a coupled-resonator above the crystal surface, this suggests that the drag experienced by resonator-coupled, hydrophilic particles is more influenced by the viscosity of the bulk water than by interfacial water adjacent to the crystal surface. Hydrophilic particles that lack a surface tether are mass-coupled just above the crystal surface and accordingly probe the drag due to the thin layer of interfacial water that is differently structured on hydrophobic and hydrophilic surfaces. Hydrophobic particles without a surface tether are also mass-coupled, but their drag coefficient decreases when the ratio of electron-donating over electron-accepting parameters increases, suggesting that hydrophobic particles experience less drag due to the structured water adjacent to the surface.

The quartz-crystal-microbalance-with-dissipation (QCM-D) has become a powerful tool for studying the bond viscoelasticity of biotic and abiotic colloidal particles adhering to substratum surfaces. A window equipped QCM-D allows high-throughput analysis of the average bond viscoelasticity, measuring over 106 particles simultaneously in one single experiment. Other techniques require laborious analyses of individual particles. In this protocol, the quantitative derivation of the spring-constant and drag-coefficient of the bond between adhering colloidal particles and substratum surfaces using QCM-D is explained for bacteria and silica particles, using the particle-mass derived for validation. Bond viscoelasticity is calculated using a coupled resonator model, paying special attention to the protocol for mathematical fitting needed to obtain reliable quantitative output. Knowledge of the viscoelasticity of the bond between colloidal particles and substratum surfaces facilitates development of new strategies to detach adhering particles from or retain them on a surface. (C) 2016 Elsevier B.V. All rights reserved.

Yeastolate or yeast extract, which are hydrolysates produced by autolysis of yeast, are often employed as a raw material in the media used for industrial mammalian cell culture. The source and quality of yeastolate can significantly affect cell growth and production; however, analysis of these complex biologically derived materials is not straightforward. The best current method, liquid chromatographymass spectrometry (LC-MS), is time-consuming and requires extensive expertise. This study describes the use of surface-enhanced Raman scattering (SERS) and fluorescence excitationemission matrix (EEM) spectroscopy coupled with robust principal component analysis (ROBPCA) for the rapid and facile characterization and discrimination of yeast extracts in aqueous solution. SERS using silver colloids generates time-dependent signals, where adenine is the strongest contributor, and the spectra are stable and reproducible (< ~3%) at 180 minutes after mixing. Combining this spectral behavior with chemometric methods enables SERS to be used for discriminating between different yeastolate sources, for assessing lot-to-lot variability, and potentially to monitor storage-induced compositional changes. Fluorescence EEM combined with multiway ROBPCA also provides a rapid and inexpensive method for discrimination of yeastolate, yielding very similar results in terms of sample discrimination to that obtained by SERS. However, the EEM data does not provide the same level of chemical information as provided by the SERS. Thus the combination of the two methodologies has the potential to be extremely useful in biopharmaceutical manufacturing for the rapid characterization and screening of biogenic hydrolysates from animal or plant sources. Irish Industrial Development Authority peer-reviewed

Amino acid transport in right-side-out membrane vesicles of Acinetobacter johnsonii 210A was studied. L-Alanine, L-lysine, and L-proline were actively transported when a proton motive force of -76 mV was generated by the oxidation of glucose via the membrane-bound glucose dehydrogenase. Kinetic analysis of amino acid uptake at concentrations of up to 80 microM revealed the presence of a single transport system for each of these amino acids with a Kt of less than 4 microM. The mode of energy coupling to solute uptake was analyzed by imposition of artificial ion diffusion gradients. The uptake of alanine and lysine was driven by a membrane potential and a transmembrane pH gradient. In contrast, the uptake of proline was driven by a membrane potential and a transmembrane chemical gradient of sodium ions. The mechanistic stoichiometry for the solute and the coupling ion was close to unity for all three amino acids. The Na+ dependence of the proline carrier was studied in greater detail. Membrane potential-driven uptake of proline was stimulated by Na+, with a half-maximal Na+ concentration of 26 microM. At Na+ concentrations above 250 microM, proline uptake was strongly inhibited. Generation of a sodium motive force and maintenance of a low internal Na+ concentration are most likely mediated by a sodium/proton antiporter, the presence of which was suggested by the Na(+)-dependent alkalinization of the intravesicular pH in inside-out membrane vesicles. The results show that both H+ and Na+ can function as coupling ions in amino acid transport in Acinetobacter spp.