Stable Expression of Gal/GalNAc Lectin of Entamoeba Histolytica in Transgenic Chloroplasts and Immunogenicity in Mice Towards Vaccine Development for Amoebiasis

Chloroplast genetic engineering offers several advantages, including high levels of transgene expression, transgene containment via maternal inheritance and multigene engineering in a single transformation event. Entamoeba histolytica infects 50 million people, causing about 100 000 deaths annually, but there is no approved vaccine against this pathogen. LecA, a potential target for blocking amoebiasis, was expressed for the first time in transgenic plants. Stable transgene integration into chloroplast genomes and homoplasmy were confirmed by polymerase chain reaction and Southern blot analyses. LecA expression was evaluated by Western blots and quantified by enzyme-linked immunosorbent assay (up to 6.3% of total soluble protein or 2.3 mg LecA/g leaf tissue). Subcutaneous immunization of mice with crude extract of transgenic leaves resulted in higher immunoglobulin G titres (up to 1 : 10 000) than in previous reports. An average yield of 24 mg of LecA per plant should produce 29 million doses of vaccine antigen per acre of transgenic plants. Such high levels of expression and immunogenicity should facilitate the development of a less expensive amoebiasis vaccine

Chloroplast-Derived Vaccine Antigens And Biopharmaceuticals: Expression, Folding, Assembly And Functionality

Chloroplast genetic engineering offers several advantages, including high levels of transgene expression, transgene containment via maternal inheritance, and multi-gene expression in a single transformation event. Oral delivery is facilitated by hyperexpression of vaccine antigens against cholera, tetanus, anthrax, plague, or canine parvovirus (4%-31% of total soluble protein, TSP) in transgenic chloroplasts (leaves) or non-green plastids (carrots, tomato) as well as the availability of antibiotic free selectable markers or the ability to excise selectable marker genes. Hyperexpression of several therapeutic proteins, including human serum albumin (11.1% TSP), somatotropin (7% TSP), interferon-alpha (19% TSP), interferon-gamma (6% TSP), and antimicrobial peptide (21.5% TSP), facilitates efficient and economic purification. Also, the presence of chap-erones and enzymes in chloroplasts facilitates assembly of complex multisubunit proteins and correct folding of human blood proteins with proper disulfide bonds. Functionality of chloroplast-derived vaccine antigens and therapeutic proteins has been demonstrated by several assays, including the macrophage lysis assay, GM1-ganglioside binding assay, protection of HeLA cells or human lung carcinoma cells against encephalomyocarditis virus, systemic immune response, protection against pathogen challenge, and growth or inhibition of cell cultures. Purification of human proinsulin has been achieved using novel purification strategies (inverse temperature transition property) that do not require expensive column chromatography techniques. Thus, transgenic chloroplasts are ideal bio-reactors for production of functional human and animal therapeutic proteins in an environmentally friendly manner. © 2009 Springer-Verlag Berlin Heidelberg

Vigilance, arousal, and acetylcholine: Optimal control of attention in a simple detection task

Paying attention to particular aspects of the world or being more vigilant in general can be interpreted as forms of 'internal' action. Such arousal-related choices come with the benefit of increasing the quality and situational appropriateness of information acquisition and processing, but incur potentially expensive energetic and opportunity costs. One implementational route for these choices is widespread ascending neuromodulation, including by acetylcholine (ACh). The key computational question that elective attention poses for sensory processing is when it is worthwhile paying these costs, and this includes consideration of whether sufficient information has yet been collected to justify the higher signal-to-noise ratio afforded by greater attention and, particularly if a change in attentional state is more expensive than its maintenance, when states of heightened attention ought to persist. We offer a partially observable Markov decision-process treatment of optional attention in a detection task, and use it to provide a qualitative model of the results of studies using modern techniques to measure and manipulate ACh in rodents performing a similar task

Fast ACh signals and the optimal control of attention in a detection task

Understanding how the brain represents, updates, and accommodates uncertainty is a key challenge for computational neuroscience. Neuromodulators such as acetylcholine (ACh) have been suggested as playing important roles over multiple timescales, regulating excitability and plasticity to mediate various effects of uncertainty on inference and learning. While these influences are consistent with ACh’s long-standing association with general functions of attention and arousal, recent studies using novel techniques to measure and manipulate this system with increasing exactitude have revealed intriguing patterns of activity at fast timescales. Notably, Sarter and colleagues used ACh amperometry (Howe et al., 2013, J.Neurosci., 33(20):8742-8752) and optogenetics (Gritton et al., 2016, PNAS, 113(8):E1089-E1097) as rodents performed a challenging signal detection task; they reported effects such as a serial dependency over multiple trials as to whether ACh is released, and increased false alarm rates when optogenetic stimulation is applied during non-signal trials. Inspired by their task and findings, we construct an abstract detection task, and consider how attentional state might be optimally controlled over each trial, assuming that more focused attention improves sensory information but incurs costs. We show similarities between the resulting attentional dynamics and task performance in the model and experimental results

Alan Frieze ∗

An n-lift of a digraph K, is a digraph with vertex set V (K) × [n] and for each directed edge (i,j) ∈ E(K) there is a perfect matching between fibers {i} × [n] and {j} × [n], with edges directed from fiber i to fiber j. If these matchings are chosen independently and uniformly at random then we say that we have a random n-lift. We show that if h is sufficiently large then a random n-lift of the complete digraph �Kh is hamiltonian whp.

Influence of Epoxy/Nanosilica on Mechanical Performance of Hemp/Kevlar Fiber Reinforced Hybrid Composite with an Ultrasonic Frequency

Ultrasonic vibration was employed in blending the nanosilica into epoxy resin to manufacture hemp/kevlar/nanosilica-based epoxy composites, with an ultrasonic occurrence of 20 kHz and a 900 W capacity of power. An ultrasonic probe was utilized to ensure the consistent dispersion of the nanoparticles in the epoxy. The mechanical characteristics of hemp/kevlar fiber reinforced with epoxy/nanosilica in a mat form have been studied. Hand layup procedures were used to create these composites, including varying weight % of nanosilica and variable fiber stacking sequencing. The different weight % are 3, 6, and 9, and the stacking sequences are B, C, and D. The effectiveness of ultrasonic irradiation on mechanical characteristics was investigated and related. The inclusion of 6 wt.% of SiO2 to the B type resulted in a 25% rise in tension and a 37% in bending. The addition of 6 wt.% silica to the C-type hybridization nanocomposite results in a 34% rise in tension and a 38% rise in bending. Extreme tension behavior is attained at 6 wt.% SiO2 with epoxy with the B type piling order, and extreme bending behavior is obtained at 6 wt.% SiO2 with the C type piling order. A B-type model composite with a 6-wt.% SiO2 addition performed better in hygroscopic than A, C, and D type model composites. An SEM is utilized to observe the microstructure of shattered materials