Iridescent biofilms of Cellulophaga lytica are tunable platforms for scalable, ordered materials.

Nature offers many examples of materials which exhibit exceptional properties due to hierarchical assembly of their constituents. In well-studied multi-cellular systems, such as the morpho butterfly, a visible indication of having ordered submicron features is given by the display of structural color. Detailed investigations of nature's designs have yielded mechanistic insights and led to the development of biomimetic materials at laboratory scales. However, the manufacturing of hierarchical assemblies at industrial scales remains difficult. Biomanufacturing aims to leverage the autonomy of biological systems to produce materials at lower cost and with fewer carbon emissions. Earlier reports documented that some bacteria, particularly those with gliding motility, self-assemble into biofilms with polycrystalline structures and exhibit glittery, iridescent colors. The current study demonstrates the potential of using one of these bacteria, Cellulophaga lytica, as a platform for the large scale biomanufacturing of ordered materials. Specific approaches for controlling C. lytica biofilm optical, spatial and temporal properties are reported. Complementary microscopy-based studies reveal that biofilm color variations are attributed to changes in morphology induced by cellular responses to the local environment. Incorporation of C. lytica biofilms into materials is also demonstrated, thereby facilitating their handling and downstream processing, as would be needed during manufacturing processes. Finally, the utility of C. lytica as a self-printing, photonic ink is established by this study. In summary, autonomous surface assembly of C. lytica under ambient conditions and across multiple length scales circumvent challenges that currently hinder production of ordered materials in industrial settings.

In situ Linkage of Fungal and Bacterial Proliferation to Microbiologically Influenced Corrosion in B20 Biodiesel Storage Tanks.

Renewable fuels hold great promise for the future yet their susceptibility to biodegradation and subsequent corrosion represents a challenge that needs to be directly assessed. Biodiesel is a renewable fuel that is widely used as a substitute or extender for petroleum diesel and is composed of a mixture of fatty acid methyl esters derived from plant or animal fats. Biodiesel can be blended up to 20% v/v with ultra-low sulfur diesel (i.e., B20) and used interchangeably with diesel engines and infrastructure. The addition of biodiesel, however, has been linked to increased susceptibility to biodegradation. Microorganisms proliferating via degradation of biodiesel blends have been linked to microbiologically influenced corrosion in the laboratory, but not measured directly in storage tanks (i.e., in situ). To measure in situ microbial proliferation, fuel degradation and microbially influenced corrosion, we conducted a yearlong study of B20 storage tanks in operation at two locations, identified the microorganisms associated with fuel fouling, and measured in situ corrosion. The bacterial populations were more diverse than the fungal populations, and largely unique to each location. The bacterial populations included members of the Acetobacteraceae, Clostridiaceae, and Proteobacteria. The abundant Eukaryotes at both locations consisted of the same taxa, including a filamentous fungus within the family Trichocomaceae, not yet widely recognized as a contaminant of petroleum fuels, and the Saccharomycetaceae family of yeasts. Increases in the absolute and relative abundances of the Trichocomaceae were correlated with significant, visible fouling and pitting corrosion. This study identified the relationship between fouling of B20 with increased rates of corrosion and the microorganisms responsible, largely at the bottom of the sampled storage tanks. To our knowledge this is the first in situ study of this scale incorporating community and corrosion measurements in an active biodiesel storage environment.

Melanin Produced by the Fast-Growing Marine Bacterium Vibrio natriegens through Heterologous Biosynthesis: Characterization and Application.

Melanin is a pigment produced by organisms throughout all domains of life. Due to its unique physicochemical properties, biocompatibility, and biostability, there has been an increasing interest in the use of melanin for broad applications. In the vast majority of studies, melanin has been either chemically synthesized or isolated from animals, which has restricted its use to small-scale applications. Using bacteria as biocatalysts is a promising and economical alternative for the large-scale production of biomaterials. In this study, we engineered the marine bacterium Vibrio natriegens, one of the fastest-growing organisms, to synthesize melanin by expressing a heterologous tyrosinase gene and demonstrated that melanin production was much faster than in previously reported heterologous systems. The melanin of V. natriegens was characterized as a polymer derived from dihydroxyindole-2-carboxylic acid (DHICA) and, similarly to synthetic melanin, exhibited several characteristic and useful features. Electron microscopy analysis demonstrated that melanin produced from V. natriegens formed nanoparticles that were assembled as "melanin ghost" structures, and the photoprotective properties of these particles were validated by their protection of cells from UV irradiation. Using a novel electrochemical reverse engineering method, we observed that melanization conferred redox activity to V. natriegens Moreover, melanized bacteria were able to quickly adsorb the organic compound trinitrotoluene (TNT). Overall, the genetic tractability, rapid division time, and ease of culture provide a set of attractive properties that compare favorably to current E. coli production strains and warrant the further development of this chassis as a microbial factory for natural product biosynthesis.IMPORTANCE Melanins are macromolecules that are ubiquitous in nature and impart a large variety of biological functions, including structure, coloration, radiation resistance, free radical scavenging, and thermoregulation. Currently, in the majority of investigations, melanins are either chemically synthesized or extracted from animals, which presents significant challenges for large-scale production. Bacteria have been used as biocatalysts to synthesize a variety of biomaterials due to their fast growth and amenability to genetic engineering using synthetic biology tools. In this study, we engineered the extremely fast-growing bacterium V. natriegens to synthesize melanin nanoparticles by expressing a heterologous tyrosinase gene with inducible promoters. Characterization of the melanin produced from V. natriegens-produced tyrosinase revealed that it exhibited physical and chemical properties similar to those of natural and chemically synthesized melanins, including nanoparticle structure, protection against UV damage, and adsorption of toxic compounds. We anticipate that producing and controlling melanin structures at the nanoscale in this bacterial system with synthetic biology tools will enable the design and rapid production of novel biomaterials for multiple applications.

Hidden Gratings in Holographic Liquid Crystal Polymer-Dispersed Liquid Crystal Films.

Dynamic diffraction gratings that are hidden in the field-off state are fabricated utilizing a room-temperature photocurable liquid crystal (LC) monomer and nematic LC (NLC) using holographic photopolymerization techniques. These holographic LC polymer-dispersed LCs (HLCPDLCs) are hidden because of the refractive index matching between the LC polymer and the NLC regions in the as-formed state (no E-field applied). Application of a moderate E-field (5 V/µm) generates a refractive index mismatch because of the NLC reorientation (along the E-field) generating high-diffraction efficiency transmission gratings. These dynamic gratings are characterized by morphological, optical, and electrooptical techniques. They exhibit a morphology made of oriented LC polymer regions (containing residual NLC) alternating with a two-phase region of an NLC and LC polymer. Unlike classic holographic polymer-dispersed LC gratings formed with a nonmesogenic monomer, there is index matching between the as-formed alternating regions of the grating. These HLCPDLCs exhibit broad band and high diffraction efficiency (≈90%) at the Bragg angle, are transparent to white light across the visible range because of the refractive index matching, and exhibit fast response times (1 ms). The ability of HLCPDLCs not to consume electrical power in the off state opens new possibilities for the realization of energy-efficient switchable photonic devices.

Electrical Control of Shape in Voxelated Liquid Crystalline Polymer Nanocomposites.

Liquid crystal elastomers (LCEs) exhibit anisotropic mechanical, thermal, and optical properties. The director orientation within an LCE can be spatially localized into voxels [three-dimensional (3-D) volume elements] via photoalignment surfaces. Here, we prepare nanocomposites in which both the orientation of the LCE and single-walled carbon nanotube (SWNT) are locally and arbitrarily oriented in discrete voxels. The addition of SWNTs increases the stiffness of the LCE in the orientation direction, yielding a material with a 5:1 directional modulus contrast. The inclusion of SWNT modifies the thermomechanical response and, most notably, is shown to enable distinctive electromechanical deformation of the nanocomposite. Specifically, the incorporation of SWNTs sensitizes the LCE to a dc field, enabling uniaxial electrostriction along the orientation direction. We demonstrate that localized orientation of the LCE and SWNT allows complex 3-D shape transformations to be electrically triggered. Initial experiments indicate that the SWNT-polymer interfaces play a crucial role in enabling the electrostriction reported herein.

The impact of culture medium on the development and physiology of biofilms of Pseudomonas fluorescens formed on polyurethane paint.

Microbial biofilms cause the deterioration of polymeric coatings such as polyurethanes (PUs). In many cases, microbes have been shown to use the PU as a nutrient source. The interaction between biofilms and nutritive substrata is complex, since both the medium and the substratum can provide nutrients that affect biofilm formation and biodeterioration. Historically, studies of PU biodeterioration have monitored the planktonic cells in the medium surrounding the material, not the biofilm. This study monitored planktonic and biofilm cell counts, and biofilm morphology, in long-term growth experiments conducted with Pseudomonas fluorescens under different nutrient conditions. Nutrients affected planktonic and biofilm cell numbers differently, and neither was representative of the system as a whole. Microscopic examination of the biofilm revealed the presence of intracellular storage granules in biofilms grown in M9 but not yeast extract salts medium. These granules are indicative of nutrient limitation and/or entry into stationary phase, which may impact the biodegradative capability of the biofilm.

Layer-in-layer hierarchical nanostructures fabricated by combining holographic polymerization and block copolymer self-assembly.

We report the combination of top-down and bottom-up nanomanufacturing techniques to fabricate active, hierarchically structured volume reflection gratings. Holographic polymerization (H-P) formed lamellar structures of approximately 200 nm in thickness, confining a block copolymer (BCP) to approximately 100 nm domains. Subsequently, the BCP self-assembles into nanolayers with a period of approximately 21 nm. We envisage that this approach opens a gateway to fabricating hierarchical nanostructures at different length scales.

Polymer crystallization/melting induced thermal switching in a series of holographically patterned Bragg reflectors.

Holographic photopolymerization (H-P) is a simple, fast and attractive means to fabricate one-, two- and three-dimensional complex structures. Liquid crystals, nanoparticles and silicate nano-plates have been patterned into submicron periodical structures. In this article, we report fabrication of a one-dimensional reflection grating structure by patterning a semicrystalline polymer, polyethylene glycol (PEG), in Norland resin (thiol-ene based UV curable resin) matrix using the H-P technique. Sharp notches observed in the reflection grating of this Norland/PEG system indicate a finite Δ present in the system due to spatial segregation of the PEG and Norland resin. The notch position red shifts upon heating and the diffraction efficiency (ratio between diffraction and incident light intensity, DE) increases from ∼20% to 60% for the Norland 65/PEG 4600 grating. This dynamic behavior of the reflection grating is also fully reversible. The unique thermal switching behavior is attributed to the melting/formation of PEG crystals during heating/cooling. By employing different molecular weight PEGs which have different melting temperatures, a series of switching temperatures have been achieved. Since PEG can be easily coupled with a variety of functional groups, this research might shed light on fabricating multifunctional Bragg gratings using the H-P technique.