Orthogonal inactivation of influenza and the creation of detergent resistant viral aggregates: towards a novel vaccine strategy

<p>Abstract</p> <p>Background</p> <p>It has been previously shown that enveloped viruses can be inactivated using aryl azides, such as 1-iodo-5-azidonaphthalene (INA), plus UVA irradiation with preservation of surface epitopes in the inactivated virus preparations. Prolonged UVA irradiation in the presence of INA results in ROS-species formation, which in turn results in detergent resistant viral protein fractions.</p> <p>Results</p> <p>Herein, we characterize the applicability of this technique to inactivate influenza. It is shown that influenza virus + INA (100 micromolar) + UVA irradiation for 30 minutes results in a significant (<it>p </it>< 0.05) increase in pelletablehemagglutinin after Triton X-100 treatment followed by ultracentrifugation. Additionally, characterization of the virus suspension by immunogold labeling in cryo-EM, and viral pellet characterization via immunoprecipitation with a neutralizing antibody, shows preservation of neutralization epitopes after this treatment.</p> <p>Conclusion</p> <p>These orthogonally inactivated viral preparations with detergent resistant fractions are being explored as a novel route for safe, effective inactivated vaccines generated from a variety of enveloped viruses.</p

Studies of uncontrolled air traffic patterns, phase 1

The general aviation air traffic flow patterns at uncontrolled airports are investigated and analyzed and traffic pattern concepts are developed to minimize the midair collision hazard in uncontrolled airspace. An analytical approach to evaluate midair collision hazard probability as a function of traffic densities is established which is basically independent of path structure. Two methods of generating space-time interrelationships between terminal area aircraft are presented; one is a deterministic model to generate pseudorandom aircraft tracks, the other is a statistical model in preliminary form. Some hazard measures are presented for selected traffic densities. It is concluded that the probability of encountering a hazard should be minimized independently of any other considerations and that the number of encounters involving visible-avoidable aircraft should be maximized at the expense of encounters in other categories

AGEN/BSEN 112 Final project: Moving and Temperament of Cattle

Cattle movement and weather may affect the body temperature of cows which in turn affects their natural behavior and can influence their metabolism. Cattle take several days to resume their normal eating patterns after being overheated, and that can affect dairy production. This project was assigned to study the effect of temperament (calm vs excitable) and evaporative cooling on the body temperature of moving animals to optimize the environmental conditions around the cattle and consequently, dairy production. The project began with processing of a data set from Dr. Tami Brown-Brandl who was the client/adviser for the group. The data contained the body temperature of six heifer cows recorded every minute for 24 hours. During the recording period the cows were moved around from their pen, and either had water sprayed on them to aid in their cooling, or left dry. The cows were classified by their temperament as calm or excitable, depending on their reaction to the presence of humans. The results from the analysis showed movement of cattle had a consistent effect on their average body temperature. As time went on during the movement process, all cattle saw an increase in body temperature with excitable cows reaching a higher maximum temperature than calm cows. Unaided cooling (dry treatment) was also affected by temperament of the cows. Calm heifer’s temperature dropped 0.1 °C about every 2 hours, while excitable heifer’s temperature dropped 0.1 °C about every 6 hours. Evaporative cooling or wet treatments help the heifers shorten excess periods of elevated body temperature with no notable difference due to temperament. Based on the analysis it is recommended to treat all heifers with a wet treatment after movement in an effort to keep heifers calm and therefore easier to corral and handle

Identification of harmful cyanobacteria in the Sacramento-San Joaquin Delta and Clear Lake, California by DNA barcoding.

Accurate identification of cyanobacteria using traditional morphological taxonomy is challenging due to the magnitude of phenotypic plasticity among natural algal assemblages. In this study, molecular approach was utilized to facilitate the accurate identification of cyanobacteria in the Sacramento-San Joaquin Delta and in Clear Lake in Northern California where recurring blooms have been observed over the past decades. Algal samples were collected from both water bodies in 2011 and the samples containing diverse cyanobacteria as identified by morphological taxonomy were chosen for the molecular analysis. The 16S ribosomal RNA genes (16S rDNA) and the adjacent internal transcribed spacer (ITS) regions were amplified by PCR from the mixed algal samples using cyanobacteria generic primers. The obtained sequences were analyzed by similarity search (BLASTN) and phylogenetic analysis (16S rDNA) to differentiate species sharing significantly similar sequences. A total of 185 plasmid clones were obtained of which 77 were successfully identified to the species level: Aphanizomenon flos-aquae, Dolichospermum lemmermannii (taxonomic synonym: Anabaena lemmermannii), Limnoraphis robusta (taxonomic synonym: Lyngbya hieronymusii f. robusta) and Microcystis aeruginosa. To date, Dolichospermum and Limnoraphis found in Clear Lake have only been identified to the genus lavel by microscopy. During the course of this study, morphological identification and DNA barcoding confirmed A. flos-aquae as the predominant cyanobacterium in the Sacramento-San Joaquin Delta indicating a shift from M. aeruginosa that have dominated the blooms in the past decade. Lastly, the species-specific identification of Limnoraphis robusta in Clear Lake is another significant finding as this cyanobacterium has, thus far, only been reported in Lake Atitlan blooms in Guatemala

Functional Amyloid Formation within Mammalian Tissue

Amyloid is a generally insoluble, fibrous cross-β sheet protein aggregate. The process of amyloidogenesis is associated with a variety of neurodegenerative diseases including Alzheimer, Parkinson, and Huntington disease. We report the discovery of an unprecedented functional mammalian amyloid structure generated by the protein Pmel17. This discovery demonstrates that amyloid is a fundamental nonpathological protein fold utilized by organisms from bacteria to humans. We have found that Pmel17 amyloid templates and accelerates the covalent polymerization of reactive small molecules into melanin—a critically important biopolymer that protects against a broad range of cytotoxic insults including UV and oxidative damage. Pmel17 amyloid also appears to play a role in mitigating the toxicity associated with melanin formation by sequestering and minimizing diffusion of highly reactive, toxic melanin precursors out of the melanosome. Intracellular Pmel17 amyloidogenesis is carefully orchestrated by the secretory pathway, utilizing membrane sequestration and proteolytic steps to protect the cell from amyloid and amyloidogenic intermediates that can be toxic. While functional and pathological amyloid share similar structural features, critical differences in packaging and kinetics of assembly enable the usage of Pmel17 amyloid for normal function. The discovery of native Pmel17 amyloid in mammals provides key insight into the molecular basis of both melanin formation and amyloid pathology, and demonstrates that native amyloid (amyloidin) may be an ancient, evolutionarily conserved protein quaternary structure underpinning diverse pathways contributing to normal cell and tissue physiology

Nanoscale Mechanical Characterisation of Amyloid Fibrils Discovered in a Natural Adhesive

Using the atomic force microscope, we have investigated the nanoscale mechanical response of the attachment adhesive of the terrestrial alga Prasiola linearis (Prasiolales, Chlorophyta). We were able to locate and extend highly ordered mechanical structures directly from the natural adhesive matrix of the living plant. The in vivo mechanical response of the structured biopolymer often displayed the repetitive sawtooth force-extension characteristics of a material exhibiting high mechanical strength at the molecular level. Mechanical and histological evidence leads us to propose a mechanism for mechanical strength in our sample based on amyloid fibrils. These proteinaceous, pleated β-sheet complexes are usually associated with neurodegenerative diseases. However, we now conclude that the amyloid protein quaternary structures detected in our material should be considered as a possible generic mechanism for mechanical strength in natural adhesives

Intrinsic regulation of hemangioma involution by platelet-derived growth factor

Infantile hemangioma is a vascular tumor that exhibits a unique natural cycle of rapid growth followed by involution. Previously, we have shown that hemangiomas arise from CD133+ stem cells that differentiate into endothelial cells when implanted in immunodeficient mice. The same clonally expanded stem cells also produced adipocytes, thus recapitulating the involuting phase of hemangioma. In the present study, we have elucidated the intrinsic mechanisms of adipocyte differentiation using hemangioma-derived stem cells (hemSCs). We found that platelet-derived growth factor (PDGF) is elevated during the proliferating phase and may inhibit adipocyte differentiation. hemSCs expressed high levels of PDGF-B and showed sustained tyrosine phosphorylation of PDGF receptors under basal (unstimulated) conditions. Inhibition of PDGF receptor signaling caused enhanced adipogenesis in hemSCs. Furthermore, exposure of hemSCs to exogenous PDGF-BB reduced the fat content and the expression of adipocyte-specific transcription factors. We also show that these autogenous inhibitory effects are mediated by PDGF receptor-β signaling. In summary, this study identifies PDGF signaling as an intrinsic negative regulator of hemangioma involution and highlights the therapeutic potential of disrupting PDGF signaling for the treatment of hemangiomas

A Transgenic Minipig Model of Huntington\u27s Disease

Background: Some promising treatments for Huntington\u27s disease (HD) may require pre-clinical testing in large animals. Minipig is a suitable species because of its large gyrencephalic brain and long lifespan. Objective: To generate HD transgenic (TgHD) minipigs encoding huntingtin (HTT)1–548 under the control of human HTT promoter. Methods: Transgenesis was achieved by lentiviral infection of porcine embryos. PCR assessment of gene transfer, observations of behavior, and postmortem biochemical and immunohistochemical studies were conducted. Results: One copy of the human HTT transgene encoding 124 glutamines integrated into chromosome 1 q24-q25 and successful germ line transmission occurred through successive generations (F0, F1, F2 and F3 generations). No developmental or gross motor deficits were noted up to 40 months of age. Mutant HTT mRNA and protein fragment were detected in brain and peripheral tissues. No aggregate formation in brain up to 16 months was seen by AGERA and filter retardation or by immunostaining. DARPP32 labeling in WT and TgHD minipig neostriatum was patchy. Analysis of 16 month old sibling pairs showed reduced intensity of DARPP32 immunoreactivity in neostriatal TgHD neurons compared to those of WT. Compared to WT, TgHD boars by one year had reduced fertility and fewer spermatozoa per ejaculate. In vitro analysis revealed a significant decline in the number of WT minipig oocytes penetrated by TgHD spermatozoa. Conclusions: The findings demonstrate successful establishment of a transgenic model of HD in minipig that should be valuable for testing long term safety of HD therapeutics. The emergence of HD-like phenotypes in the TgHD minipigs will require more study

Author Correction: Mosaic nanoparticle display of diverse influenza virus hemagglutinins elicits broad B cell responses.

In the version of this article initially published, the labels (50 Å) above the scale bars in Fig. 1b were incorrect. The correct size is 50 nm. The error has been corrected in the HTML and PDF versions of the article