Characterisation of mixed virus infections in Ribes species in Switzerland

Various virus disease-like symptoms are frequently observed in Ribes sp. in Switzerland but the aetiology remains poorly documented, although a number of viruses infecting Ribes sp. were described elsewhere. Therefore, symptomatic and apparently healthy plants from diverse origins were analysed by electron microscopy (EM), immunoprecipitation electron microscopy (IPEM), Western blot and (RT-)PCR. By EM, at least four different particle types, often in combination, were observed. (1) Bacilliform particles were typical for the Badnavirus genus with dimensions of 145 x 28 nm. This virus was identified by PCR as the Gooseberry vein banding associated virus (GVBaV). (2) Filamentous particles were mainly observed on black currants with downward rolling of leaves with interveinal reddening during summer and fall. We tentatively named this unknown virus Blackcurrant leafroll-associated virus 1 (BCLRaV-1). In phylogenetic analysis of HSP70h nucleotide sequences, BCLRaV-1 felt in the Closterovirus genus. In Western blot analysis, one dominant protein with an estimated molecular weight of about 28 kDa was detectable. The virus was shown to be different from the Raspberry mottle closterovirus (RMoV) by IPEM and RT-PCR. (3) RTPCR and sequencing of products also clearly demonstrated the presence in our Ribes samples of Rubus chlorotic mottle virus (RuCMV), a Sobemovirus recently described in Scotland. This finding correlates with the presence of the 30 nm diameter particles observed by EM. (4) A further structure with isometrical particles of 60 nm could not yet be attributed to a particular genus. Altogether, our data suggest the presence of multiple virus infections i

Highly analysable, reusable, and realisable architectural designs with XCD

Connector-Centric Design (XcD) is a new approach to specifying software architectures. XcD views complex connectors as highly significant in architectural designs, as it is the complex connectors that non-functional quality properties in systems can emanate from. So, XcD promotes in designs a clean separation of connectors (interaction behaviours) from components (functional behaviours). Designers can then specify connectors in detail explicitly thus easing the analysis of system designs for quality properties. Furthermore, XcD separates control behaviour from connectors as control strategies. Architectural designs in XcD thus become highly modular with re-usable components, connectors, and control strategies (representing design solutions for quality properties). The end result is the eased architectural experimentation with different design solutions by re-using components/connectors and formal analysis of these solutions to find out the optimal ones

Identification of classical swine fever virus protein E2 as a target for cytotoxic T cells by using mRNA-transfected antigen-presenting cells

Vaccination of pigs against Classical swine fever virus (CSFV) by using live-virus vaccines induces early protection before detectable humoral immune responses. Immunological analyses indicate that this is associated with T-cell activation, underlining the importance of targeting cytotoxic T-lymphocyte (CTL) responses for vaccine improvement. Antigen-presenting cells (APCs) transfected with mRNA encoding structural protein E2 or non-structural viral proteins NS3¿NS4A were used to identify viral genes encoding CTL epitopes. Monocyte-derived dendritic cells (DCs) and fibrocytes served as the APCs. In vitro translation of the mRNA and microscopic analysis of transfected cells demonstrated that E2 and NS3¿NS4A could be identified. APCs transfected with either of the mRNA molecules restimulated CSFV-specific T cells to produce gamma interferon and specific cytotoxic activity against CSFV-infected target cells. The presence of CTL epitopes on E2 was confirmed by using d/d-haplotype MAX cells expressing E2 constitutively as target cells in d/d-haplotype CTL assays. A potent CTL activity against E2 was detected early (1¿3 weeks) after CSFV challenge. This work corroborates the existence of CTL epitopes within the non-structural protein domain NS3¿NS4A of CSFV. Furthermore, epitopes on the E2 protein can also now be classified as targets for CTLs, having important implications for vaccine design, especially subunit vaccines. As for the use of mRNA-transfected APCs, this represents a simple and efficient method to identify viral genes encoding CTL epitopes in outbred population

Lenvatinib in Advanced Radioiodine-Refractory Thyroid Cancer - A Retrospective Analysis of the Swiss Lenvatinib Named Patient Program.

javax.xml.bind.JAXBElement@5a6991ed Differentiated thyroid cancer (DTC) accounts for approximately 95% of thyroid carcinomas. In the metastatic RAI-refractory disease, chemotherapy has very limited efficacy and is associated with substantial toxicity. With increasing knowledge of the molecular pathogenesis of DTC, novel targeted therapies have been developed. Lenvatinib is a tyrosine kinase inhibitor (TKI) with promising clinical activity based on the randomized phase III SELECT trial. In Switzerland, a Named Patient Program (NPP) was installed to bridge the time gap to Swissmedic approval. Here, we report the results from the Swiss Lenvatinib NPP including patients with metastatic RAI-refractory DTC. javax.xml.bind.JAXBElement@7407c55a Main inclusion criteria for the Swiss NPP were RAI-refractory DTC, documented disease progression, Eastern Cooperative Oncology Group (ECOG) performance status 0-3. The number of previous therapies was not limited. The Swiss Lenvatinib NPP was initiated in June 2014 and was closed in October 2015 with the approval of the drug. javax.xml.bind.JAXBElement@1c5cb2cc Between June 2014 and October 2015, 13 patients with a median age of 72 years have been enrolled. Most patients (69%) had at least one prior systemic therapy, mainly sorafenib. 31% of patients showed a PR and 31% SD. Median progression free survival was 7.2 months and the median overall survival was 22.7 months. Dose reduction due to adverse events was necessary in 7 patients (53%). At the time of analysis 6 patients (47%) were still on treatment with a median time on treatment of 9.98 months. javax.xml.bind.JAXBElement@713fc2d4 Our results show that lenvatinib has reasonable clinical activity in unselected patients with RAI-refractory thyroid cancer with nearly two-third of patients showing clinical benefit. The toxicity profile of lenvatinib is manageable

Borrelia burgdorferi EbfC defines a newly-identified, widespread family of bacterial DNA-binding proteins

The Lyme disease spirochete, Borrelia burgdorferi, encodes a novel type of DNA-binding protein named EbfC. Orthologs of EbfC are encoded by a wide range of bacterial species, so characterization of the borrelial protein has implications that span the eubacterial kingdom. The present work defines the DNA sequence required for high-affinity binding by EbfC to be the 4 bp broken palindrome GTnAC, where ‘n’ can be any nucleotide. Two high-affinity EbfC-binding sites are located immediately 5′ of B. burgdorferi erp transcriptional promoters, and binding of EbfC was found to alter the conformation of erp promoter DNA. Consensus EbfC-binding sites are abundantly distributed throughout the B. burgdorferi genome, occurring approximately once every 1 kb. These and other features of EbfC suggest that this small protein and its orthologs may represent a distinctive type of bacterial nucleoid-associated protein. EbfC was shown to bind DNA as a homodimer, and site-directed mutagenesis studies indicated that EbfC and its orthologs appear to bind DNA via a novel α-helical ‘tweezer’-like structure

Activation and modulation of antiviral and apoptotic genes in pigs infected with classical swine fever viruses of high, moderate or low virulence

The immune response to CSFV and the strategies of this virus to evade and suppress the pigs’ immune system are still poorly understood. Therefore, we investigated the transcriptional response in the tonsils, median retropharyngeal lymph node (MRLN), and spleen of pigs infected with CSFV strains of similar origin with high, moderate, and low virulence. Using a porcine spleen/intestinal cDNA microarray, expression levels in RNA pools prepared from infected tissue at 3 dpi (three pigs per virus strain) were compared to levels in pools prepared from uninfected homologue tissues (nine pigs). A total of 44 genes were found to be differentially expressed. The genes were functionally clustered in six groups: innate and adaptive immune response, interferon-regulated genes, apoptosis, ubiquitin-mediated proteolysis, oxidative phosphorylation and cytoskeleton. Significant up-regulation of three IFN-γ-induced genes in the MRLNs of pigs infected with the low virulence strain was the only clear qualitative difference in gene expression observed between the strains with high, moderate and low virulence. Real-time PCR analysis of four response genes in all individual samples largely confirmed the microarray data at 3 dpi. Additional PCR analysis of infected tonsil, MRLN, and spleen samples collected at 7 and 10 dpi indicated that the strong induction of expression of the antiviral response genes chemokine CXCL10 and 2′–5′ oligoadenylate synthetase 2, and of the TNF-related apoptosis-inducing ligand (TRAIL) gene at 3 dpi, decreased to lower levels at 7 and 10 dpi. For the highly and moderately virulent strains, this decrease in antiviral and apoptotic gene expression coincided with higher levels of virus in these immune tissues

Development of a diagnostic protocol for dizziness in elderly patients in general practice: a Delphi procedure

<p>Abstract</p> <p>Background</p> <p>Dizziness in general practice is very common, especially in elderly patients. The empirical evidence for diagnostic tests in the evaluation of dizziness is scarce. Aim of our study was to determine which set of diagnostic tests should be part of a diagnostic protocol for evaluating dizziness in elderly patients in general practice.</p> <p>Methods</p> <p>We conducted a Delphi procedure with a panel of 16 national and international experts of all relevant medical specialities in the field of dizziness. A selection of 36 diagnostic tests, based on a systematic review and practice guidelines, was presented to the panel. Each test was described extensively, and data on test characteristics and methodological quality (assessed with the Quality Assessment of Diagnostic Accuracy Studies, QUADAS) were presented. The threshold for in- or exclusion of a diagnostic test was set at an agreement of 70%.</p> <p>Results</p> <p>During three rounds 21 diagnostic tests were selected, concerning patient history (4 items), physical examination (11 items), and additional tests (6 items). Five tests were excluded, although they are recommended by existing practice guidelines on dizziness. Two tests were included, although several practice guidelines question their diagnostic value. Two more tests were included that have never been recommended by practice guidelines on dizziness.</p> <p>Conclusion</p> <p>In this study we successfully combined empirical evidence with expert opinion for the development of a set of diagnostic tests for evaluating dizziness in elderly patients. This comprehensive set of tests will be evaluated in a cross-sectional diagnostic study.</p