M13 bacteriophage production for large-scale applications.

Bacteriophage materials have the potential to revolutionize medicine, energy production and storage, agriculture, solar cells, optics and many other fields. To fulfill these needs, this study examined critical process parameters during phage propagation to increase phage production capability. A representative scale-down system was created in tube spin reactors to allow parallel experimentation with single- and multi-variable analysis. Temperature, harvest time, media composition, feed regime, bacteriophage, and bacteria concentration were analyzed in the scale-down system. Temperature, media composition, and feeding regimens were found to affect phage production more than other factors. Temperature affected bacterial growth and phage production inversely. Multi-variate analysis identified an optimal parameter space which provided a significant improvement over the base line method. This method should be useful in scaled production of bacteriophage for biotechnology.

A systems toxicology approach to elucidate the mechanisms involved in RDX species-specific sensitivity.

Interspecies uncertainty factors in ecological risk assessment provide conservative estimates of risk where limited or no toxicity data is available. We quantitatively examined the validity of interspecies uncertainty factors by comparing the responses of zebrafish (Danio rerio) and fathead minnow (Pimephales promelas) to the energetic compound 1,3,5-trinitroperhydro-1,3,5-triazine (RDX), a known neurotoxicant. Relative toxicity was measured through transcriptional, morphological, and behavioral end points in zebrafish and fathead minnow fry exposed for 96 h to RDX concentrations ranging from 0.9 to 27.7 mg/L. Spinal deformities and lethality occurred at 1.8 and 3.5 mg/L RDX respectively for fathead minnow and at 13.8 and 27.7 mg/L for zebrafish, indicating that zebrafish have an 8-fold greater tolerance for RDX than fathead minnow fry. The number and magnitude of differentially expressed transcripts increased with increasing RDX concentration for both species. Differentially expressed genes were enriched in functions related to neurological disease, oxidative-stress, acute-phase response, vitamin/mineral metabolism and skeletal/muscular disorders. Decreased expression of collagen-coding transcripts were associated with spinal deformity and likely involved in sensitivity to RDX. Our work provides a mechanistic explanation for species-specific sensitivity to RDX where zebrafish responded at lower concentrations with greater numbers of functions related to RDX tolerance than fathead minnow. While the 10-fold interspecies uncertainty factor does provide a reasonable cross-species estimate of toxicity in the present study, the observation that the responses between ZF and FHM are markedly different does initiate a call for concern regarding establishment of broad ecotoxicological conclusions based on model species such as zebrafish.

Effect of Cellulose Nanofibrils and TEMPO-mediated Oxidized Cellulose Nanofibrils on the Physical and Mechanical Properties of Poly(vinylidene fluoride)/Cellulose Nanofibril Composites.

Cellulose nanofibrils (CNFs) are high aspect ratio, natural nanomaterials with high mechanical strength-to-weight ratio and promising reinforcing dopants in polymer nanocomposites. In this study, we used CNFs and oxidized CNFs (TOCNFs), prepared by a 2,2,6,6-tetramethylpiperidine-1-oxyl radical (TEMPO)-mediated oxidation process, as reinforcing agents in poly(vinylidene fluoride) (PVDF). Using high-shear mixing and doctor blade casting, we prepared free-standing composite films loaded with up to 5 wt % cellulose nanofibrils. For our processing conditions, all CNF/PVDF and TOCNF/PVDF films remain in the same crystalline phase as neat PVDF. In the as-prepared composites, the addition of CNFs on average increases crystallinity, whereas TOCNFs reduces it. Further, addition of CNFs and TOCNFs influences properties such as surface wettability, as well as thermal and mechanical behaviors of the composites. When compared to neat PVDF, the thermal stability of the composites is reduced. With regards to bulk mechanical properties, addition of CNFs or TOCNFs, generally reduces the tensile properties of the composites. However, a small increase (~18%) in the tensile modulus was observed for the 1 wt % TOCNF/PVDF composite. Surface mechanical properties, obtained from nanoindentation, show that the composites have enhanced performance. For the 5 wt % CNF/PVDF composite, the reduced modulus and hardness increased by ~52% and ~22%, whereas for the 3 wt % TOCNF/PVDF sample, the increase was ~23% and ~25% respectively.

Trends in Synthetic Biology Applications, Tools, Industry, and Oversight and Their Security Implications.

Recent developments in synthetic biology tools and techniques are driving commercialization of a wide range of products for human health, agriculture, environmental stewardship, and other purposes. This article reviews some of the trends in synthetic biology applications as well as some of the tools enabling these and future advances. These tools and capabilities are being developed in the context of a rapidly changing industry, which may have an impact on the rate and direction of progress. Final products are subject to a regulatory framework that is being challenged by the pace, scale, and novelty of this new era of biotechnology. This article includes discussion of these factors and how they may affect product design and the types of applications that are most likely to be supported and pursued commercially. The final section provides perspective on the security implications of these advances, with a focus on US interests.

Production of tunable nanomaterials using hierarchically assembled bacteriophages.

Large-scale fabrication of precisely defined nanostructures with tunable functions is critical to the exploitation of nanoscience and nanotechnology for production of electronic devices, energy generators, biosensors, and bionanomedicines. Although self-assembly processes have been developed to exploit biological molecules for functional materials, the resulting nanostructures and functions are still very limited, and scalable synthesis is far from being realized. Recently, we have established a bacteriophage-based biomimetic process, called 'self-templating assembly'. We used bacteriophage as a nanofiber model system to exploit its liquid crystalline structure for the creation of diverse hierarchically organized structures. We have also demonstrated that genetic modification of functional peptides of bacteriophage results in structures that can be used as soft and hard tissue-regenerating materials, biosensors, and energy-generating materials. Here, we describe a comprehensive protocol to perform genetic engineering of phage, liter-scale amplification, purification, and self-templating assembly, and suggest approaches for characterizing hierarchical phage nanostructures using optical microscopy, atomic-force microscopy (AFM), and scanning electron microscopy (SEM). We also discuss sources of contamination, common mistakes during the fabrication process, and quality-control measures to ensure reproducible material production. The protocol takes ∼8-10 d to complete.