A Catalytic Nanoreactor Based on in Vivo Encapsulation of Multiple Enzymes in an Engineered Protein Nanocompartment

Bacterial protein compartments concentrate and sequester enzymes, thereby regulating biochemical reactions. Here, we generated a new functional nanocompartment in Escherichia coli by engineering the MS2 phage capsid protein to encapsulate multiple cargo proteins. Sequestration of multiple proteins in MS2‐based capsids was achieved by SpyTag/SpyCatcher protein fusions that covalently crosslinked with the interior surface of the capsid. Further, the functional two‐enzyme indigo biosynthetic pathway could be targeted to the engineered capsids, leading to a 60 % increase in indigo production in vivo. The enzyme‐loaded particles could be purified in their active form and showed enhanced long‐term stability in vitro (about 95 % activity after seven days) compared with free enzymes (about 5 % activity after seven days). In summary, this engineered in vivo encapsulation system provides a simple and versatile way for generating highly stable multi‐enzyme nanoreactors for in vivo and in vitro applications.     

Vitamin E acetate addition to poly(d,l)lactic acid modifies its mechanical behavior without affecting biocompatibility

Mechanical properties of poly(d,l )lactic acid films enriched with Vitamin E and Vitamin E Acetate (5–40% w/w) were investigated. The addition of both formulations resulted in increased polymer Young's modulus and tensile strength. Human foreskin fibroblasts and murine pre‐osteoblasts were used to assess the biocompatibility of polymers. Pre‐osteoblasts adhesion and proliferation were strongly decreased by Vitamin E, whereas Vitamin E Acetate did not alter cell proliferation. Collagen deposition was lower onto Vitamin E blended polymers than onto native and Vitamin E Acetate blended ones. Fibroblasts adhesion and proliferation were increased by both Vitamin E and Vitamin E Acetate addition. © 2013 Wiley Periodicals, Inc. J. Appl. Polym. Sci., 2014 , 131, 39970.     

In-House Validation and Comparison of Two Wheat (<Emphasis Type="Italic">Triticum aestivum</Emphasis>) Taxon-Specific Real-Time PCR Methods for GMO Quantification Supported by Droplet Digital PCR

Common wheat is one of the most important staple food crops worldwide. However, unlike other important staple crops such as maize or soybean, genetically modified (GM) wheat is not yet present in the global food market. Nonetheless, in the recent past, the adventitious presence of GM glyphosate-tolerant volunteers was reported in open wheat fields in the USA. The European Union Reference Laboratory for GM Food and Feed (EURL-GMFF) was therefore called to develop a strategy to detect such unauthorised GM wheat in wheat samples by using both taxon-specific and screening tests. Two candidate common wheat taxon-specific real-time PCR methods were suggested, one targeting ssII-D gene coding for starch synthase and the other targeting waxy-D1 gene, coding for granule-bound starch synthase. In the present study, the two above-mentioned real-time PCR taxon-specific methods were in-house verified and compared, proposing droplet digital PCR (ddPCR) as a new tool for supporting the application of the European Network of GMO Laboratories (ENGL) established method performance criteria. Preliminary performance data of waxy-D1 and ssII-D methods in ddPCR format are shown too to give a contribution to the bridging process from the consolidated to the emerging quantitative PCR methodology.     

Prevalence of the Campylobacter multi-drug efflux pump (CmeABC) in Campylobacter spp. Isolated from freshly processed Turkeys

The prevalence of the Campylobacter multi-drug efflux pump (CmeABC) was evaluated in Campylobacter isolates recovered from freshly processed turkeys at two Midwestern processing plants. A total of 94 Campylobacter isolates recovered from processed turkeys were examined using polymerase chain reaction (PCR) to determine the presence of the multi-drug efflux pump genes cmeA, cmeB, and cmeC. Results from this study found that 51% of all isolates tested were positive for CmeABC. 46.6% of these positive isolates were from plant A and 55.1% from plant B. Differences were observed in the prevalence of individual genes found among Campylobacter isolates from each plant. Additional analysis found that among the isolates positive for CmeABC, 85.5% were identified as C. jejuni and 14.5% identified as C. coli. There was a relatively high occurrence of the Campylobacter multi-drug efflux pump genes in Campylobacter spp. recovered from processed turkeys, however, the presence of the genes could not be significantly linked to antimicrobial resistance observed in the test strains and suggests that the CmeABC genes are only one factor associated with antimicrobial resistance in Campylobacter spp.     

Cholesterol oxidation in heated lard enriched with two levels of cholesterol

Cholesterol oxidation in lard containing two levels of added cholesterol was monitored using capillary gaschromatography. Loss of cholesterol and formation of cholesterol oxidation products (COPs) were measured. Lard samples with 10 times (Test I) and 2 times (Test II) the amount of cholesterol originally found in each batch of lard were heated at 180°C for 10 hr a day for 240 and 160 hr, respectively. Cholesterol steadily decreased throughout the heating period in both tests. Cholesterol loss followed a first-order reaction rate, with a rate constant (k) of −1.18×10⁻³ h⁻¹ for Test I and −9.45×10⁻³ h⁻¹ for Test II. The COPs accumulated during both heating tests. But the amount of COPs formed did not total the amount of cholesterol lost. During heating, thermal degradation of cholesterol likely occurred, and those products were not detected. During cooling, hydroperoxides formed, which further oxidized into the COPs that were detected. The 7-ketocholesterol and 5α,6α-epoxycholesterol were the predominant COPs formed. The isomeric 7α-and 7β-hydroxycholesterols also accumulated in the heating tests. The 3β,5α,6β-cholestantriol was found in very small amounts and the 25-hydroxycholesterol was not detected. Presented in part at the 80th AOCS Annual Meeting, Cincinnati, OH, in May, 1989.     

Treatability Study on Membrane Concentrate Containing Pesticides Using Advanced Oxidation Processes

This treatability study evaluated the overall effectiveness of advanced oxidation processes (AOPs) to treat membrane concentrates containing the pesticides bromoxynil and trifluralin. The results of study indicate that high levels of pesticide degradation were achieved using ozone (O₃) plus hydrogen peroxide (H₂O₂) for all concentrate matrices. However, the toxicity of the samples during the O₃/H₂O₂ process was higher than that obtained during ultraviolet (UV) light combined with H₂O₂. Low levels of pesticide oxidation were observed in experiments using a mixture of pesticides during all treatment options.     

Relative abundance,age structure,and body size in mudpuppy populations in southwestern Ontario

Little is known of mudpuppy (Necturus maculosus) population structure and ecology; some populations in the Great Lakes are thought to be in decline. Mudpuppies are the obligate hosts for the mudpuppy mussel (Simpsonaias ambigua), a species that is endangered in Canada and in many Great Lakes states. We surveyed mudpuppies from the Sydenham River, the only known Canadian locality of the mudpuppy mussel, in order to generate information on relative density, deformity rates and population age/size structure and used this information to compare them to known mudpuppy populations from Great Lakes sites in the Detroit River, Lake St. Clair and Long Point in Lake Erie. Deformity rates were elevated at some sites in the Sydenham River. The relative density of mudpuppies in the Sydenham River was lower than that of other Great Lakes sites and their age was skewed towards younger individuals. Although at lower densities than at other Great Lakes sites, the mudpuppy population in the Sydenham River appears stable and is showing signs of recruitment which bodes well for the future of the mudpuppy mussel population of the river.     

Evaluation of Cell Migration and Cytokines Expression Changes under the Radiofrequency Electromagnetic Field on Wound Healing In Vitro Model

Wound healing (WH) proceeds through four distinct phases: hemostasis, inflammation, proliferation, and remodeling. Impaired WH may be the consequence of the alteration of one of these phases and represents a significant health and economic burden to millions of individuals. Thus, new therapeutic strategies are the topics of intense research worldwide. Although radiofrequency electromagnetic field (RF-EMF) has many medical applications in rehabilitation, pain associated with musculoskeletal disorders, and degenerative joint disorders, its impact on WH is not fully understood. The process of WH begins just after injury and continues during the inflammatory and proliferative phases. A thorough understanding of the mechanisms by which RF-EMF can improve WH is required before it can be used as a non-invasive, inexpensive, and easily self-applicable therapeutic strategy. Thus, the aim of this study is to explore the therapeutic potential of different exposure setups of RF-EMF to drive faster healing, evaluating the keratinocytes migration, cytokines, and matrix metalloproteinases (MMPs) expression. The results showed that RF-EMF treatment promotes keratinocytes’ migration and regulates the expression of genes involved in healing, such as MMPs, tissue inhibitors of metalloproteinases, and pro/anti-inflammatory cytokines, to improve WH.     

Morphological segmenting and neighborhood pixel-based locality preserving projection on brain fMRI dataset for semantic feature extraction: an affective computing study

Two specific chemical receptive fields of brain, namely the amygdala and the orbital-frontal cortex, are related to valence and arousal in medical experiments. Functional magnetic resonance imaging (fMRI), which is a noninvasive, repeatable, and atomical tool for medical imaging in clinic system, was widely used in affective computing; however, it faces its dataset processing difficulty for dimensional reduction as well as for decreasing the computational complexity. In addition, features extraction from those de-dimensionality datasets is a challenging issue. The current work solved the de-dimensionality issue by using some preprocessing algorithms including clustering, morphological segmenting, and locality preserving projection. In order to keep useful information in fMRI dataset for reduction process, improved neighborhood pixel-based locality preserving projection (NP-LPP) algorithm was addressed and continuously for feature extraction operating using Otsu weighted sum of histogram. Furthermore, a modified covariance power spectral density (MC-PSD) separately in an fMRI Valence–Arousal experiments was measured. The results were analyzed and compared with affective norms English words system. The experiments established that the proposed methods of NP-LPP effectively simplified high complexity of fMRI, and Otsu weighted sum of histogram exhibited superior performance for features extraction compared to the MC-PSD through the calculation root mean standard error. The current proposed method provided a potential application and promising research direction on human semantic retrieval through medical imaging dataset.