Eimeria tenella protein trafficking: differential regulation of secretion versus surface tethering during the life cycle

Eimeria spp. are intracellular parasites that have a major impact on poultry. Effective live vaccines are available and the development of reverse genetic technologies has raised the prospect of using Eimeria spp. as recombinant vectors to express additional immunoprotective antigens. To study the ability of Eimeria to secrete foreign antigens or display them on the surface of the sporozoite, transiently transfected populations of E. tenella expressing the fluorescent protein mCherry, linked to endogenous signal peptide (SP) and glycophosphatidylinositol-anchor (GPI) sequences, were examined. The SP from microneme protein EtMIC2 (SP2) allowed efficient trafficking of mCherry to cytoplasmic vesicles and following the C-terminal addition of a GPI-anchor (from surface antigen EtSAG1) mCherry was expressed on the sporozoite surface. In stable transgenic populations, mCherry fused to SP2 was secreted into the sporocyst cavity of the oocysts and after excystation, secretion was detected in culture supernatants but not into the parasitophorous vacuole after invasion. When the GPI was incorporated, mCherry was observed on the sporozites surface and in the supernatant of invading sporozoites. The proven secretion and surface exposure of mCherry suggests that antigen fusions with SP2 and GPI of EtSAG1 may be promising candidates to examine induction of protective immunity against heterologous pathogens

An Optimal Control Formulation for Inviscid Incompressible Ideal Fluid Flow

In this paper we consider the Hamiltonian formulation of the equations of incompressible ideal fluid flow from the point of view of optimal control theory. The equations are compared to the finite symmetric rigid body equations analyzed earlier by the authors. We discuss various aspects of the Hamiltonian structure of the Euler equations and show in particular that the optimal control approach leads to a standard formulation of the Euler equations -- the so-called impulse equations in their Lagrangian form. We discuss various other aspects of the Euler equations from a pedagogical point of view. We show that the Hamiltonian in the maximum principle is given by the pairing of the Eulerian impulse density with the velocity. We provide a comparative discussion of the flow equations in their Eulerian and Lagrangian form and describe how these forms occur naturally in the context of optimal control. We demonstrate that the extremal equations corresponding to the optimal control problem for the flow have a natural canonical symplectic structure.Comment: 6 pages, no figures. To appear in Proceedings of the 39th IEEEE Conference on Decision and Contro

Viral proteins expressed in the protozoan parasite Eimeria tenella are detected by the chicken immune system

BACKGROUND: Eimeria species are parasitic protozoa that cause coccidiosis, an intestinal disease commonly characterised by malabsorption, diarrhoea and haemorrhage that is particularly important in chickens. Vaccination against chicken coccidiosis is effective using wild-type or attenuated live parasite lines. The development of protocols to express foreign proteins in Eimeria species has opened up the possibility of using Eimeria live vaccines to deliver heterologous antigens and function as multivalent vaccine vectors that could protect chickens against a range of pathogens. RESULTS: In this study, genetic complementation was used to express immunoprotective virus antigens in Eimeria tenella. Infectious bursal disease virus (IBDV) causes Gumboro, an immunosuppressive disease that affects productivity and can interfere with the efficacy of poultry vaccination programmes. Infectious laryngotracheitis virus (ILTV) causes a highly transmissible respiratory disease for which strong cellular immunity and antibody responses are required for effective vaccination. Genes encoding the VP2 protein from a very virulent strain of IBDV (vvVP2) and glycoprotein I from ILTV (gI) were cloned downstream of 5’Et-Actin or 5’Et-TIF promoter regions in plasmids that also contained a mCitrine fluorescent reporter cassette under control of the 5’Et-MIC1 promoter. The plasmids were introduced by nucleofection into E. tenella sporozoites, which were then used to infect chickens. Progeny oocysts were sorted by FACS and passaged several times in vivo until the proportion of fluorescent parasites in each transgenic population reached ~20 % and the number of transgene copies per parasite genome decreased to < 10. All populations were found to transcribe and express the transgene and induced the generation of low titre, transgene-specific antibodies when used to immunise chickens. CONCLUSIONS: E. tenella can express antigens of other poultry pathogens that are successfully recognised by the chicken immune system. Nonetheless, further work has to be done in order to improve the levels of expression for its future use as a multivalent vaccine vector. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s13071-016-1756-2) contains supplementary material, which is available to authorized users

Classroom Demonstrations: Learning Tools Or Entertainment?

We compared student learning from different modes of presenting classroom demonstrations to determine how much students learn from traditionally presented demonstrations, and whether learning can be enhanced by simply changing the mode of presentation to increase student engagement. We find that students who passively observe demonstrations understand the underlying concepts no better than students who do not see the demonstration at all, in agreement with previous studies. Learning is enhanced, however, by increasing student engagement; students who predict the demonstration outcome before seeing it, however, display significantly greater understanding

Optimization Of Detergent-Mediated Reconstitution Of Influenza A M2 Protein Into Proteoliposomes

We report the optimization of detergent-mediated reconstitution of an integral membrane-bound protein, full-length influenza M2 protein, by direct insertion into detergent-saturated liposomes. Detergent-mediated reconstitution is an important method for preparing proteoliposomes for studying membrane proteins, and must be optimized for each combination of protein and membrane constituents used. The purpose of the reconstitution was to prepare samples for site-directed spin-labeling electron paramagnetic resonance (SDSL-EPR) studies. Our goals in optimizing the protocol were to minimize the amount of detergent used, reduce overall proteoliposome preparation time, and confirm the removal of all detergent. The liposomes were comprised of (1-palmitoyl-2-oleyl-sn-glycero-phosphocholine (POPC) and 1-palmitoyl-2-oleyl-sn-glycero-3-[phospho-rac-(1-glycerol)] (POPG), and the detergent octylglucoside (OG) was used for reconstitution. Rigorous physical characterization was applied to optimize each step of the reconstitution process. We used dynamic light scattering (DLS) to determine the amount of OG needed to saturate the preformed liposomes. During detergent removal by absorption with Bio-Beads, we quantified the detergent concentration by means of a colorimetric assay, thereby determining the number of Bio-Bead additions needed to remove all detergent from the final proteoliposomes. We found that the overnight Bio-Bead incubation used in previously published protocols can be omitted, reducing the time needed for reconstitution. We also monitored the size distribution of the proteoliposomes with DLS, confirming that the size distribution remains essentially constant throughout the reconstitution process

Meta-analysis of the Sea Change Consultation Reports

Sea Change is a three-year, EU funded, Horizon 2020 project with 17 partners in nine European countries (Belgium, Denmark, Greece, Sweden, France, Ireland, Portugal, Spain and UK). During April and May 2016, eight of these countries (except France) carried out consultations with education stakeholders (classified as incumbents, regulating agencies and challengers, see p. xii) for the purpose of gaining deeper insights into the barriers to teaching 12-19 year olds about the ocean, and to identify options for overcoming these barriers. The Sea Change partners used the method Collective intelligence (CI) to consult with the education stakeholders. The CI method used facilitation and problem solving to harmonise input from education stakeholders from different backgrounds and perspectives. This report was built on the Collective Intelligence consultation work completed and reported in Sea For Society (D2.4 Global Analysis of the SFS Consultation Process from a Social Sciences Perspective), as reported in Domegan et al., 2014

The New Political Economy of EU State Aid Policy

Despite its importance and singularity, the EU’s state aid policy has attracted less scholarly attention than other elements of EU competition policy. Introducing the themes addressed by the special issue, this article briefly reviews the development of EU policy and highlights why the control of state aid matters. The Commission’s response to the current economic crisis notably in banking and the car industry is a key concern, but the interests of the special issue go far beyond. They include: the role of the European Commission in the development of EU policy, the politics of state aid, and a clash between models of capitalism. The special issue also examines the impact of EU policy. It investigates how EU state aid decisions affect not only industrial policy at the national level (and therefore at the EU level), but the welfare state and territorial relations within federal member states, the external implications of EU action and the strategies pursued by the Commission to limit any potential disadvantage to European firms, and the conflict between the EU’s expanding legal order and national

Stochastic Streams: Sample Complexity vs. Space Complexity

We address the trade-off between the computational resources needed to process a large data set and the number of samples available from the data set. Specifically, we consider the following abstraction: we receive a potentially infinite stream of IID samples from some unknown distribution D, and are tasked with computing some function f(D). If the stream is observed for time t, how much memory, s, is required to estimate f(D)? We refer to t as the sample complexity and s as the space complexity. The main focus of this paper is investigating the trade-offs between the space and sample complexity. We study these trade-offs for several canonical problems studied in the data stream model: estimating the collision probability, i.e., the second moment of a distribution, deciding if a graph is connected, and approximating the dimension of an unknown subspace. Our results are based on techniques for simulating different classical sampling procedures in this model, emulating random walks given a sequence of IID samples, as well as leveraging a characterization between communication bounded protocols and statistical query algorithms

Existence of solutions for a higher order non-local equation appearing in crack dynamics

In this paper, we prove the existence of non-negative solutions for a non-local higher order degenerate parabolic equation arising in the modeling of hydraulic fractures. The equation is similar to the well-known thin film equation, but the Laplace operator is replaced by a Dirichlet-to-Neumann operator, corresponding to the square root of the Laplace operator on a bounded domain with Neumann boundary conditions (which can also be defined using the periodic Hilbert transform). In our study, we have to deal with the usual difficulty associated to higher order equations (e.g. lack of maximum principle). However, there are important differences with, for instance, the thin film equation: First, our equation is nonlocal; Also the natural energy estimate is not as good as in the case of the thin film equation, and does not yields, for instance, boundedness and continuity of the solutions (our case is critical in dimension $1$ in that respect)

Resolving the Azimuthal Ambiguity in Vector Magnetogram Data with the Divergence-Free Condition: Application to Discrete Data

We investigate how the divergence-free property of magnetic fields can be exploited to resolve the azimuthal ambiguity present in solar vector magnetogram data, by using line-of-sight and horizontal heliographic derivative information as approximated from discrete measurements. Using synthetic data we test several methods that each make different assumptions about how the divergence-free property can be used to resolve the ambiguity. We find that the most robust algorithm involves the minimisation of the absolute value of the divergence summed over the entire field of view. Away from disk centre this method requires the sign and magnitude of the line-of-sight derivatives of all three components of the magnetic field vector.Comment: Solar Physics, in press, 20 pages, 11 figure