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We present high-resolution measurements of the electron beam energy spread at the SwissFEL injector as a function of the electron bunch charge, the optics, and the longitudinal dispersion of the lattice. The measured values are in general an order of magnitude higher than what is predicted by standard simulation codes. The measured dependences indicate that the energy spread blowup is caused primarily by intrabeam scattering and microbunching instabilities, effects not covered in the conventional modeling of radio-frequency injectors. We present further a numerical model that qualitatively reproduces the experimental data and an approach to mitigate the energy spread deterioration. The work underlines the importance of considering the energy spread in the optimization and design of high-brightness electron beam sources and the need to develop new models to adequately understand and simulate the observed physics effects., Physical Review Accelerators and Beams, 25 (10), ISSN:2469-9888

SARS CoV-2 antibody assays measure antibodies against the viral nucleoprotein (NP) or spike protein. The study examined if testing of antibodies against both antigens increases the diagnostic sensitivity. Sera (N=98) from infected individuals were tested with ELISAs based on the NP, receptor-binding domain (RBD), or both proteins. The AUROCs were 0.958 (NP), 0.991 (RBD), and 0.992 (NP/RBD). The RBD- and NP/RBD-based ELISAs showed better performance than the NP-based assay. Simultaneous testing for antibodies against NP and RBD increased the number of true and false positives. If maximum diagnostic sensitivity is required, the NP/RBD-based ELISA is preferable. Otherwise, the RBD-based ELISA is sufficient. Supplementary Information The online version contains supplementary material available at 10.1007/s10096-021-04284-5.

‘Staggering disease’ is a neurological disorder considered a threat to European domestic cats (Felis catus) for almost five decades. However, its aetiology has remained obscure. Rustrela virus (RusV), a relative of rubella virus, has recently been shown to be associated with encephalitis in a broad range of mammalian hosts. Here, we report the detection of RusV RNA and antigen by metagenomic sequencing, RT-qPCR, in-situ hybridization and immunohistochemistry in brain tissues of 28 out of 29 cats with non-suppurative meningoencephalomyelitis and ‘staggering disease’-like neurological disorder from Sweden, Austria, and Germany, but not in non-affected control cats. Screening of possible reservoir hosts in Sweden revealed RusV infection in wood mice (Apodemus sylvaticus). Our work strongly indicates RusV as the long-sought cause of feline ‘staggering disease’. Given its broad host spectrum and considerable geographic range, RusV may be the aetiological agent of neuropathologies in further mammals, possibly even including humans.

We present the first lasing results of SwissFEL, a hard X-ray free-electron laser (FEL) that recently came into operation at the Paul Scherrer Institute in Switzerland. SwissFEL is a very stable, compact and cost-effective X-ray FEL facility driven by a low-energy and ultra-low-emittance electron beam travelling through short-period undulators. It delivers stable hard X-ray FEL radiation at 1-A wavelength with pulse energies of more than 500 μJ, pulse durations of ~30 fs (root mean square) and spectral bandwidth below the per-mil level. Using special configurations, we have produced pulses shorter than 1 fs and, in a different set-up, broadband radiation with an unprecedented bandwidth of ~2%. The extremely small emittance demonstrated at SwissFEL paves the way for even more compact and affordable hard X-ray FELs, potentially boosting the number of facilities worldwide and thereby expanding the population of the scientific community that has access to X-ray FEL radiation. The first lasing results at SwissFEL, an X-ray free-electron laser, are presented, highlighting the facility’s unique capabilities. A general comparison to other major facilities is also provided.

Orthohantaviruses are re-emerging rodent-borne pathogens distributed all over the world. Here, we report the isolation of a Puumala orthohantavirus (PUUV) strain from bank voles caught in a highly endemic region around the city Osnabrück, north-west Germany. Coding and non-coding sequences of all three segments (S, M, and L) were determined from original lung tissue, after isolation and after additional passaging in VeroE6 cells and a bank vole-derived kidney cell line. Different single amino acid substitutions were observed in the RNA-dependent RNA polymerase (RdRP) of the two stable PUUV isolates. The PUUV strain from VeroE6 cells showed a lower titer when propagated on bank vole cells compared to VeroE6 cells. Additionally, glycoprotein precursor (GPC)-derived virus-like particles of a German PUUV sequence allowed the generation of monoclonal antibodies that allowed the reliable detection of the isolated PUUV strain in the immunofluorescence assay. In conclusion, this is the first isolation of a PUUV strain from Central Europe and the generation of glycoprotein-specific monoclonal antibodies for this PUUV isolate. The obtained virus isolate and GPC-specific antibodies are instrumental tools for future reservoir host studies. Electronic supplementary material The online version of this article (10.1007/s11262-020-01755-3) contains supplementary material, which is available to authorized users.

Erratum in Correction to Lancet Microbe 2021; published online Nov 16. https://doi.org/10.1016/S2666-5247(21)00211-1. [No authors listed] Lancet Microbe. 2022 Jan;3(1):e8. doi: 10.1016/S2666-5247(21)00325-6. Epub 2021 Nov 26. PMID: 34870252 Free PMC article.; International audience; Biobanking infrastructures, which are crucial for responding early to new viral outbreaks, share pathogen genetic resources in an affordable, safe, and impartial manner and can provide expertise to address access and benefit-sharing issues. The European Virus Archive has had a crucial role in the global response to the COVID-19 pandemic by distributing EU-subsidised (free of charge) viral resources to users worldwide, providing non-monetary benefit sharing, implementing access and benefit-sharing compliance, and raising access and benefit-sharing awareness among members and users. All currently available SARS-CoV-2 material in the European Virus Archive catalogue, including variants of concern, are not access and benefit-sharing cases per se, but multilateral benefit-sharing has nevertheless occurred. We propose and discuss how a multilateral system enabling access and benefit-sharing from pathogen genetic resources, based on the European Virus Archive operational model, could help bridge the discrepancies between the current bilateral legal framework for pathogen genetic resources and actual pandemic response practices.

The Accelerator on a Chip International Program (ACHIP) is an international collaboration, funded by the Gordon and Betty Moore Foundation, with the goal of demonstrating that laser-driven accelerator can be integrated on a chip to fully build an accelerator based on dielectric structures. PSI will provide access to the high brightness electron beam of SwissFEL to test structures, approaches and methods towards achieving the final goal of the project. In this contribution, we will describe the two interaction chambers installed on SwissFEL to perform the proof-of-principle experiments. In particular, we will present the positioning system for the samples, the magnets needed to focus the beam to sub-micrometer dimensions and the diagnostics to measure beam properties at the interaction point.

Background Newcastle disease is a devastating disease in poultry caused by virulent Newcastle disease virus (NDV), a paramyxovirus endemic in many regions of the world despite intensive vaccination. Phylogenetic analyses reveal ongoing evolution of the predominant circulating genotype 2.VII, and the relevance of potential antigenic drift is under discussion. To investigate variation within neutralization-sensitive epitopes within the protein responsible for receptor binding, i.e. the Hemagglutinin-Neuraminidase (HN) spike protein, we were interested in establishing genotype-specific monoclonal antibodies (MAbs). Methods An HN-enriched fraction of a gradient-purified NDV genotype 2.VII was prepared and successfully employed to induce antibodies in BalbC mice that recognize conformationally intact sites reactive by haemagglutination inhibition (HI). For subsequent screening of mouse hybridoma cultures, an NDV-ELISA was established that utilizes Concanavalin A (ConA-ELISA) coupled glycoproteins proven to present conformation-dependent epitopes. Results Six out of nine selected MAbs were able to block receptor binding as demonstrated by HI activity. One MAb recognized an epitope only present in the homologue virus, while four other MAbs showed weak reactivity to selected other genotypes. On the other hand, one broadly cross-reacting MAb reacted with all genotypes tested and resembled the reactivity profile of genotype-specific polyclonal antibody preparations that point to minor antigenic differences between tested NDV genotpyes. Conclusions These results point to the concurrent presence of variable and conserved epitopes within the HN molecule of NDV. The described protocol should help to generate MAbs against a variety of NDV strains and to enable in depth analysis of the antigenic profiles of different genotypes.

Background: Some recipients of ChAdOx1 nCoV-19 COVID-19 Vaccine AstraZeneca develop antibody-mediated vaccine-induced thrombotic thrombocytopenia (VITT), associated with cerebral venous and other unusual thrombosis resembling autoimmune heparin-induced thrombocytopenia. A prothrombotic predisposition is also observed in Covid-19. We explored whether antibodies against the SARS-CoV-2 spike protein induced by Covid-19 cross-react with platelet factor 4 (PF4/CXLC4), the protein targeted in both VITT and autoimmune heparin-induced thrombocytopenia.Methods: Immunogenic epitopes of PF4 and SARS-CoV-2 spike protein were compared via prediction tools and 3D modelling software (IMED, SIM, MacMYPOL). Sera from 222 PCR-confirmed Covid-19 patients from five European centers were tested by PF4/heparin ELISA, heparin-dependent and PF4-dependent platelet activation assays. Immunogenic reactivity of purified anti-PF4 and anti-PF4/heparin antibodies from patients with VITT were tested against recombinant SARS-CoV-2 spike protein. Results: Three motifs within the spike protein sequence share a potential immunogenic epitope with PF4. Nineteen of 222 (8.6%) Covid-19 patient sera tested positive in the IgG-specific PF4/heparin ELISA, none of which showed platelet activation in the heparin-dependent activation assay, including 10 (4.5%) of the 222 Covid-19 patients who developed thromboembolic complications. Purified anti-PF4 and anti-PF4/heparin antibodies from two VITT patients did not show cross-reactivity to recombinant SARS-CoV-2 spike protein. Conclusions: The antibody responses to PF4 in SARS-CoV-2 infection and after vaccination with COVID-19 Vaccine AstraZeneca differ. Antibodies against SARS-CoV-2 spike protein do not cross-react with PF4 or PF4/heparin complexes through molecular mimicry. These findings make it very unlikely that the intended vaccine-induced immune response against SARS-CoV-2 spike protein would itself induce VITT.

This work describes a method for the generation of one and two-color ultra-short X-ray free-electron-laser pulses, based on tailoring the Coulomb interaction between the electrons of a highly compressed beam in the nonlinear regime.

The energy spread of the electron beam is a critical parameter in x-ray free-electron lasers (XFELs) and needs to be optimized for best performance. The uncorrelated energy spread of the electrons can be a few keV or less in XFEL injectors, thus very challenging to measure. The standard method to characterize the electron beam energy spread, consisting in streaking the beam with a transverse deflector and measuring the time-resolved beam size of the electrons in a dispersive location for a single electron beam energy, has a typical resolution of several keV. To overcome this limitation we introduce a novel method to measure the beam size at a dispersive location for different beam energies so that it is possible to disentangle the beam size contributions related to the energy spread, the intrinsic beam size and the monitor resolution. As a consequence, the energy spread can be characterized with a much higher precision and resolution than in the standard approach. We also suggest to perform measurements for different deflection amplitudes so that the energy spread induced by the transverse deflector can be subtracted properly. The scheme does not require any additional hardware and thus can be readily applied in any standard XFEL facility. Numerical simulations and experimental results at SwissFEL confirm the validity of our method. Our calculations show that the approach can be used to significantly overcome the resolution of the standard approach and measure energy spreads well below 1 keV. As an example we present energy spreads of few keV measured at the SwissFEL injector., Physical Review Accelerators and Beams, 23 (9), ISSN:2469-9888

Ebola virus (EBOV) causes severe outbreaks of viral hemorrhagic fever in humans. While virus-host interactions are promising targets for antivirals, there is only limited knowledge regarding the interactions of EBOV with cellular host factors. Recently, we performed a genome-wide siRNA screen that identified the nuclear RNA export factor 1 (NXF1) as an important host factor for the EBOV life cycle. NXF1 is a major component of the nuclear mRNA export pathway that is usurped by many viruses whose life cycles include nuclear stages. However, the role of NXF1 in the life cycle of EBOV, a virus replicating in cytoplasmic inclusion bodies, remains unknown. In order to better understand the role of NXF1 in the EBOV life cycle, we performed a combination of co-immunoprecipitation and double immunofluorescence assays to characterize the interactions of NXF1 with viral proteins and RNAs. Additionally, using siRNA-mediated knockdown of NXF1 together with functional assays, we analyzed the role of NXF1 in individual aspects of the virus life cycle. With this approach we identified the EBOV nucleoprotein (NP) as a viral interaction partner of NXF1. Further studies revealed that NP interacts with the RNA-binding domain of NXF1 and competes with RNA for this interaction. Co-localization studies showed that RNA binding-deficient, but not wildtype NXF1, accumulates in NP-derived inclusion bodies, and knockdown experiments demonstrated that NXF1 is necessary for viral protein expression, but not for viral RNA synthesis. Finally, our results showed that NXF1 interacts with viral mRNAs, but not with viral genomic RNAs. Based on these results we suggest a model whereby NXF1 is recruited into inclusion bodies to promote the export of viral mRNA:NXF1 complexes from these sites. This would represent a novel function for NXF1 in the life cycle of cytoplasmically replicating viruses, and may provide a basis for new therapeutic approaches against EBOV, and possibly other emerging viruses.

We have produced hard x-ray free-electron laser (FEL) radiation with unprecedented large bandwidth tunable up to 2%. The experiments have been carried out at SwissFEL, the x-ray FEL facility at the Paul Scherrer Institute in Switzerland. The bandwidth is enhanced by maximizing the energy chirp of the electron beam, which is accomplished by optimizing the compression setup. We demonstrate continuous tunability of the bandwidth with a simple method only requiring a quadrupole magnet. The generation of such broadband FEL pulses will improve the efficiency of many techniques such as x-ray crystallography and spectroscopy, opening the door to significant progress in photon science. It has already been demonstrated that the broadband pulses of SwissFEL are beneficial to enhance the performance of crystallography, and further SwissFEL users plan to exploit this large bandwidth radiation to improve the efficiency of their measurement techniques.

The transverse emittance of the electron beam is a fundamental parameter in linac-based x-ray free-electron lasers (FELs). We present results of emittance measurements carried out at SwissFEL, a compact x-ray FEL facility at the Paul Scherrer Institute in Switzerland, including a description of the novel high-resolution measurement techniques and the optimization procedure. We obtained slice emittance values at the undulator entrance down to 200 nm for an electron beam with a charge of 200 pC and an rms duration of 30--40 fs. Furthermore, we achieved slice emittances as low as 100 nm for 10 pC beams with few fs duration. These values set new standards for electron linear accelerators. The quality, verification, and control of our electron beams allowed us to generate high-power FEL radiation for a wavelength as short as 0.1 nm using an electron beam with an energy of only 6 GeV. The emittance values demonstrated at SwissFEL would allow producing hard x-ray FEL pulses with even lower-energy beams, thus paving the way for even more compact and cost-effective FEL facilities.

Objectives To determine the seroprevalence of dengue in central and eastern Sudan and the breadth of neutralising antibody responses. Methods Blood was drawn from 483 patients with fever who visited outpatient clinics in Port Sudan, Red Sea state, in three towns in Kassala state and in El Obeid, North Kordofan, in December 2012 and January 2013. Sera were tested for dengue virus IgG and IgM by ELISA (Panbio) and sera without serologic evidence of acute infection (IgM negative) were used for the analysis of the seroprevalence. DENV neutralisation tests were performed to determine the specificity of the ELISA and to examine the degree of cross-neutralisation of multiple DENV serotypes. Results Sixty-seven per cent (302 of 448) of the sera were dengue virus IgG-positive. The seroprevalence in Port Sudan was 89% (106 of 119 sera), in Kassala 61% (128 of 209) and in North Kordofan 56.7% (68 of 120). Thirty-one of 32 ELISA-positive sera neutralised dengue viruses indicating that the ELISA was highly specific. The majority of the sera broadly neutralised all four dengue virus serotypes indicating multiple infections. Conclusions The majority of the population in central and eastern Sudan has been infected with dengue viruses, many people repeatedly. The high seroprevalence underscores the need for extended dengue surveillance in Sudan, broad disease awareness in medical institutions and in the population and diagnostic capacity building for severe dengue infections.

Schmallenberg virus (SBV) induces fetal malformation, abortions and stillbirth in ruminants. While the non-structural protein NSs is a major virulence factor, the biological function of NSm, the second non-structural protein which consists of three hydrophobic transmembrane (I, III, V) and two non-hydrophobic regions (II, IV), is still unknown. Here, a series of NSm mutants displaying deletions of nearly the entire NSm or of the non-hydrophobic domains was generated and the intracellular distribution of NSm was assessed. SBV-NSm is dispensable for the generation of infectious virus and mutants lacking domains II - V showed growth properties similar to the wild-type virus. In addition, a comparable intracellular distribution of SBV-NSm was observed in mammalian cells infected with domain II mutants or wild-type virus. In both cases, NSm co-localized with the glycoprotein Gc in the Golgi compartment. However, domain IV-deletion mutants showed an altered distribution pattern and no co-localization of NSm and Gc.

A simple method to significantly enhance the coherence of X-ray free-electron laser (FEL) pulses by combining self-seeding and the harmonic generation mechanism is presented. This work is a fundamental step towards obtaining fully coherent FEL pulses, solving in this way a critical problem in the FEL field and its multidisciplinary scientific applications. In particular, the coherence improvement will be extremely beneficial for many photon science applications requiring a small bandwidth such as resonant inelastic X-ray scattering, therefore opening new areas of research and playing a crucial role in the future of all scientific users of the FEL facilities., X-ray free-electron lasers (FELs) are cutting-edge scientific tools able to generate transversely coherent radiation with very high power and ultra-short pulse durations. The self-seeding mechanism has been proven to increase the longitudinal coherence of the FEL radiation but its efficiency could be significantly improved, especially for soft X-rays. This paper proposes the enhancement of the performance of self-seeding by combining it with the harmonic generation mechanism. In particular, by starting the process with a subharmonic of the wavelength of interest, the coherence of the produced radiation is improved, the undulator beamline becomes more compact, and the monochromator realization is simplified. Numerical simulations for SwissFEL are presented showing that the method can be employed, within a given space, to increase the spectral brightness by one order of magnitude or more with respect to standard self-seeding. This coherence enhancement will be fundamental for many photon science applications and techniques such as resonant inelastic X-ray scattering.

We demonstrate the operation of a compact x-ray free-electron laser (FEL) using the optical klystron mechanism. This effect speeds up the FEL process using the dispersion of magnetic chicanes placed between the undulator modules of the FEL beamline. The demonstration was performed at the soft x-ray beamline of SwissFEL, called Athos, which is, as far as we are aware, the only x-ray FEL to date with magnetic chicanes between every two undulator modules. In our measurements, we show that, compared to standard operation without chicanes, the required undulator length to achieve FEL saturation is reduced between 15% and 30% for radiation wavelengths between 1 and 2 nm. Fully exploiting the optical klystron effect represents an important step toward more compact FEL designs, rendering this key technology more affordable and hence accessible to a larger science community.

A transverse tilt of the electron beam is normally not wanted in X-ray free-electron-laser (XFEL) facilities, since in a typical undulator beamline with quadrupole magnets only the central part of the beam with negligibly small betatron oscillations will produce significant XFEL radiation. In this work we present special configurations for which a transverse tilt can actually be used to improve the XFEL performance. In particular, we show with numerical simulations how tilted electron beams can be employed to generate two-color XFEL pulses, ultra-large-bandwidth XFEL radiation, and high-power and short duration XFEL radiation. We present and assess different mechanisms to generate the transverse tilt, namely a transverse deflecting structure, transverse wakefields, and introducing dispersion to an energy-chirped beam.

The Accelerator on a Chip International Program (ACHIP), funded by the Gordon and Betty Moore Foundation for a 5 year period, pursues basic research and development for a super-compact accelerator on a chip, where the accelerating structure is a dielectric microstructure excited by femtosecond laser pulses. The Paul Scherrer Institute (PSI) will contribute to this by providing the international collaboration access to the high-brightness electron beams in SwissFEL, where we plan to do a proof-of-principle demonstration of the acceleration of a highly relativistic beam. In this contribution we present the conceptual layout of the experiment, in which we will focus the beam down to sub-micrometer beam sizes. Start-to-end simulation results of the tracked electron beam, and first calculations of the accelerating field of the microstructure will be shown.

We present a new and simple method to generate two-color free-electron laser (FEL) pulses. In our scheme an electron bunch is quadratically tilted in the transverse direction and exhibits an energy chirp. The head and tail of the bunch are aligned to the undulator beamline and will lase, while the middle part of the bunch undergoes orbit oscillations and thus is unable to amplify the FEL radiation. Since the beam is energy chirped, the central wavelengths of the SASE amplification at the two aligned parts of the bunch are different. The beam tilt is introduced by a variation of sextupole strength in a bunch compressor section. We present simulation results of this new method for the hard X-ray FEL beamline Aramis at SwissFEL. This includes the general performance and tunability of the scheme, the requirements on the energy chirp and tilt generation, and adverse effects associated with the beam tilt.

Background Dengue IgG testing is being recommended before dengue vaccination. Presently, the diagnostic method of choice is the dengue IgG ELISA. Objective Determine the test performance and concordance of two commercial dengue IgG ELISA kits. Study design A diagnostic study to examine the sensitivity, specificity, accuracy and concordance of the Panbio Dengue Indirect IgG ELISA kit and the NovaLisa Dengue IgG ELISA kit. Sera (483) were from dengue-endemic regions in Sudan. Test performance characteristics were determined when tests were performed as indicated in the test kits and when the Panbio calibrator sample was used for both tests. Results The sensitivity of the Panbio and the NovaLisa ELISA was 91.1% and 99.0% and the specificity was 79.4% and 50.9%. The Panbio test was slightly more accurate (87.5% compared with 84.0%). Quantitative measurement readings of the tests correlated. The calibrator samples gave different cutoff values. Replacing the NovaLisa cutoff sample with the Panbio calibrator sample raised the accuracy of the NovaLisa assay to 88% and increased the concordance of the tests from 82.8 to 93%. Conclusions The study shows that the two dengue IgG ELISAs differed clearly in sensitivity and specificity and gave discordant results for 17.2% of the sera. For the most part the discrepancy depended on the calibrator sample. The findings indicate that an optimized dengue IgG calibrator standard can enhance accuracy and concordance of commercial dengue ELISAs. An optimized standard calibrator would make dengue IgG seroprevalence testing more reliable.

The SwissFEL soft X-ray free-electron laser (FEL) beamline Athos will be ready for user operation in 2021. Its design includes a novel layout of alternating magnetic chicanes and short undulator segments. Together with the APPLE X architecture of undulators, the Athos branch can be operated in different modes producing FEL beams with unique characteristics ranging from attosecond pulse length to high-power modes. Further space has been reserved for upgrades including modulators and an external seeding laser for better timing control. All of these schemes rely on state-of-the-art technologies described in this overview. The optical transport line distributing the FEL beam to the experimental stations was designed with the whole range of beam parameters in mind. Currently two experimental stations, one for condensed matter and quantum materials research and a second one for atomic, molecular and optical physics, chemical sciences and ultrafast single-particle imaging, are being laid out such that they can profit from the unique soft X-ray pulses produced in the Athos branch in an optimal way., Journal of Synchrotron Radiation, 26 (4), ISSN:0909-0495, ISSN:1600-5775

A novel, flexible yet simple method to generate gigawatt X-ray free-electron-laser radiation with unprecedented spectral bandwidth above the 10% level is presented. Such broadband radiation will improve substantially the efficiency of techniques like X-ray crystallography and spectroscopy, paving the way for outstanding progress in fields like biology and material science., A new and simple method to generate X-ray free-electron-laser radiation with unprecedented spectral bandwidth above the 10% level is presented. The broad bandwidth is achieved by sending a transversely tilted beam through a transverse-gradient undulator. The extent of the bandwidth can easily be controlled by variation of the beam tilt or the undulator gradient. Numerical simulations confirm the validity and feasibility of this method.

During herpesvirus replication, newly synthesized nucleocapsids exit the nucleus by a vesicle-mediated transport, which requires the nuclear egress complex (NEC), composed of the conserved viral proteins designated as pUL31 and pUL34 in the alphaherpesviruses pseudorabies virus (PrV) and herpes simplex viruses. Oligomerization of the heterodimeric NEC at the inner nuclear membrane (INM) results in membrane bending and budding of virus particles into the perinuclear space. The INM-derived primary envelope then fuses with the outer nuclear membrane to release nucleocapsids into the cytoplasm. The two NEC components are necessary and sufficient for induction of vesicle budding and scission as shown after co-expression in eukaryotic cells or in synthetic membranes. However, where and when the NEC is formed, how membrane curvature is mediated and how it is regulated, remains unclear. While monospecific antisera raised against the different components of the PrV NEC aided in the characterization and intracellular localization of the individual proteins, no NEC specific tools have been described yet for any herpesvirus. To gain more insight into vesicle budding and scission, we aimed at generating NEC specific monoclonal antibodies (mAbs). To this end, mice were immunized with bacterially expressed soluble PrV NEC, which was previously used for structure determination. Besides pUL31- and pUL34-specific mAbs, we also identified mAbs, which reacted only in the presence of both proteins indicating specificity for the complex. Confocal microscopy with those NEC-specific mAbs revealed small puncta (approx. 0.064 μm2) along the nuclear rim in PrV wild type infected cells. In contrast, ca. 5-fold larger speckles (approx. 0.35 μm2) were detectable in cells infected with a PrV mutant lacking the viral protein kinase pUS3, which is known to accumulate primary enveloped virions in the PNS within large invaginations of the INM, or in cells co-expressing pUL31 and pUL34. Kinetic experiments showed that while the individual proteins were detectable already between 2–4 hours after infection, the NEC-specific mAbs produced significant staining only after 4–6 hours in accordance with timing of nuclear egress. Taken together, the data indicate that these mAbs specifically label the PrV NEC.

We have installed an interaction chamber in the electron beam line of SwissFEL. Electrons with a particle energy of 3 GeV are focused to a few-micrometer beam size. Samples placed in this beam can be aligned to the electron beam with a hexapod. The goal of this installation is to demonstrate laser-driven acceleration inside dielectric structures. We present here the layout of this interaction chamber as well as first measurements with the electron beam.

Introduction: The complete inactivation of infectious tissues of large animal carcasses is one of the most challenging tasks in high-containment facilities. Steam sterilization is a method frequently in use to achieve biological inactivation of liquid and solid waste. Objective: This study aims to highlight parameters most effective in creating reproducible cycles for steam sterilization of pig and calf carcasses. Methods: Two pigs or 1 calf were sterilized by running a liquid cycle (n = 3) at 121°C for at least 120 minutes in a pass-through autoclave. To assess the physical and biological parameters, temperature data loggers and biological indicators (BIs) with spores of Geobacillus stearothermophilus (ATCC 7953) were placed at defined positions within animal carcasses. After completion of each cycle, data loggers were analyzed and BIs were incubated for 7 days at 60°C. Results: Initial testing with an undissected pig carcass resulted in suboptimal temperatures at the tissue level with growth on 1 BI. After modifications of the used stainless-steel boxes and by placing the reference probe of the autoclave in the animal carcass, reproducible cycles could be created. A complete inactivation of BIs and a temperature profile of >121°C for at least 20 minutes could be achieved in almost all probed tissues. Conclusion: Only minor modifications in carcass preparation and the used sterilization equipment resulted in effective and reproducible cycles to inactivate large animal carcasses by using a steam autoclave.

A broadband energy-chirped hard X-ray pulse has been demonstrated at the SwissFEL (free electron laser) with up to 4% bandwidth. We consider the characteristic parameters for analyzing the time dependence of stationary protein diffraction with energy-chirped pulses. Depending on crystal mosaic spread, convergence, and recordable resolution, individual reflections are expected to spend at least &asymp, 50 attoseconds and up to &asymp, 8 femtoseconds in reflecting condition. Using parameters for a chirped XFEL pulse obtained from simulations of 4% bandwidth conditions, ray-tracing simulations have been carried out to demonstrate the temporal streaking across individual reflections and resolution ranges for protein crystal diffraction. Simulations performed at a higher chirp (10%) emphasize the importance of chirp magnitude that would allow increased observation statistics for the temporal separation of individual reflections for merging and structure determination. Finally, we consider the fundamental limitation for obtaining time-dependent observations using chirped pulse diffraction. We consider the maximum theoretical time resolution achievable to be on the order of 50&ndash, 200 as from the instantaneous bandwidth of the chirped SASE pulse. We then assess the ability to propagate ultrafast optical pulses for pump-probe cross-correlation under characteristic conditions of material dispersion, in this regard, the limiting factors for time resolution scale with crystal thickness. Crystals that are below a few microns in size will be necessary for subfemtosecond time resolution.

A simple and compact scheme that enhances the brightness of self-amplified spontaneous-emission (SASE) free-electron lasers is presented. The method combines the high-brightness SASE scheme and the optical klystron concept to increase the temporal coherence of the produced radiation and to reduce the required length of the undulator beamline at the same time. The scheme is very simple and only requires compact chicanes between the modules of the undulator beamline. Simulations show that, in comparison with SASE, the brightness can be improved by up to a factor of ten and the required length to achieve saturation can be reduced by 20% or more.

Rift Valley fever virus is a mosquito-borne virus which is associated with acute hemorrhagic fever leading to large outbreaks among ruminants and humans in Africa and the Arabian Peninsula. RVFV circulates between mosquitoes, ruminants, camels and humans, which requires divergent amplification and maintenance strategies that have not been fully explored on the cellular and molecular level. We therefore assessed monoclonal antibodies for their applicability to monitor the expression pattern and kinetics of viral proteins in different RVFV infected cell species. Sequences of RVFV vaccine strain MP-12 were used in a bacterial expression system to produce recombinant non-structural proteins directed to NSs and NSm. After immunization of balb/c mice a set of monoclonal antibodies were generated and extensively characterized. The kinetics of RVFV proteins in vertebrate (Vero76) and mosquito-derived (C6/36) cells were evaluated with monoclonal antibodies against the nucleocapsid protein (NP) and the glycoproteins (Gn and Gc) as well as with the newly generated NSs and NSm derived monoclonal antibodies. Significant differences of viral protein distribution and accumulation in vertebrate compared to mosquito-derived cells could be demonstrated. Differences were observed for the nonstructural NSm and most intriguingly for the NSs protein indicating significant divergency of replication strategies of RVFV in Vero 76 cells and C6/36 cells. The described monoclonal antibodies are therefore powerful tools to elucidate the discrepancies of virus replication and interaction within the mammalian host compared to the mosquito vector.

A new large-scale facility is currently in the design phase at the Paul Scherrer Institute. SwissFEL is foreseen to accelerate electrons up to an energy of 7 GeV with a pulsed time structure. Unavoidable interactions of the primary beam with beam line components create secondary radiation. The shielding of the accelerator vault has to reduce the radiation level for accessible areas in compliance with legal constraints. In addition, the position and layout of cable ducts and infrastructure shafts through the shielding of the accelerator tunnel have to be investigated from a radiation protection point of view. In this work, the dose rate distribution inside the accelerator vault has been evaluated for expected beam loss scenarios (normal and diagnostic mode of operation) based on Monte Carlo calculations. By means of the multi-particle transport code FLUKA, a simplified model has been defined and the results have been parameterized. From these, a dose rate evaluation has been extracted for different positions inside the accelerator tunnel. The results can be used to investigate the leakage of radiation through cable ducts and infrastructure shafts for different design layouts or modes of operation.

The present paper describes a simple approach to obtain three-dimensional (3D) cartilage constructs using human normal dermal fibroblasts (hNDFs) cultured in a self-assembling peptide nanofibre scaffold. During the first days of culture, the 3D constructs underwent morphological changes consisting of a substantial contraction process that ended in a small compact structure. During this process the system became sensitive to induction with standard chondrogenic medium, evidenced by the expression of specific markers of mature cartilage. First, it was detected that the samples become highly stained with toluidine blue dye over time (40-50 days), indicating that the system produced significantly high amounts of glycosaminoglycans. By quantitative PCR, it was confirmed that the system significantly upregulated the expression of the proteoglycan aggrecan, a good indicator of cartilage commitment. Moreover, collagen type II was upregulated at protein level, confirming that the system differentiated to a chondrocyte-like construct. Additionally, during the first days of culture in control medium analysed hNDFs proliferation capacity in this 3D system was analysed. This platform could be used in the future to obtain an autologous source of cells from a simple patient skin biopsy, which could be easily translated into a low-cost and effective regenerative therapy.

The design of a photoinjector, in particular that of the electron source, is of central importance for free electron laser (FEL) machines where a high beam brightness is required. In comparison to standard designs, an rf traveling-wave photocathode gun can provide a more rigid beam with a higher brightness and a shorter pulse. This is illustrated by applying a specific optimization procedure to the SwissFEL photoinjector, for which a brightness improvement up to a factor 3 could be achieved together with a double gun output energy compared to the reference setup foreseeing a state-of-the-art S-band rf standing-wave gun. The higher brightness is mainly given by a (at least) double peak current at the exit of the gun which brings benefits for both the beam dynamics in the linac and the efficiency of the FEL process. The gun design foresees an innovative coaxial rf coupling at both ends of the structure which allows a solenoid with integrated bucking coil to be placed around the cathode in order to provide the necessary focusing right after emission.

Several beam manipulation methods have been studied and experimentally tested to generate two-color photon beams in free electron laser facilities to accommodate the user requests. We propose to use the interaction of the beam with an oscillating longitudinal wakefield source to obtain a suitable electron beam structure. The bunch generates two subpulses with different energies and delayed in time passing through a magnetic chicane after its longitudinal phase space has been modulated by the wakefield source. According to this approach the power of the emitted radiation is not degraded compared to the monochromatic beam, and the setup in the machine is quite simple because the bunch is manipulated only in the high energy section, where it is more rigid. We present the design applied to SwissFEL. We identified the parameters and the corresponding range of tunability of the time and energy separation among the two subbunches.

The advent of x-ray free electron lasers has made possible the study of matter at the characteristic space and time scales of atomic and molecular phenomena using intense coherent x-ray pulses. This article describes the physical principles and the theoretical models governing the interaction of charged particles, electromagnetic waves, and external magnetic fields that comprise the x-ray free-electron lasers. It also includes a discussion of existing facilities and avenues for increasing the peak power and improving the control of spectral and coherence properties to allow the exploration of an even larger range of phenomena.\hskip-0.22em

An optimization of the undulator layout of X-ray free-electron-laser (FEL) facilities based on placing small chicanes between the undulator modules is presented. The installation of magnetic chicanes offers the following benefits with respect to state-of-the-art FEL facilities: reduction of the required undulator length to achieve FEL saturation, improvement of the longitudinal coherence of the FEL pulses, and the ability to produce shorter FEL pulses with higher power levels. Numerical simulations performed for the soft X-ray beamline of the SwissFEL facility show that optimizing the advantages of the layout requires shorter undulator modules than the standard ones. This proposal allows a very compact undulator beamline that produces fully coherent FEL pulses and it makes possible new kinds of experiments that require very short and high-power FEL pulses.

With the successful operation of free-electron lasers (FELs) as user facilities there has been a growing demand for experiments with two photon pulses with variable photon energy and time separation. A configuration of an undulator with variable-gap control and a delaying chicane in the middle of the beamline is proposed. An injected electron beam with a transverse tilt will only yield FEL radiation for the parts which are close to the undulator axis. This allows, after re-aligning and delaying the electron beam, a different part of the bunch to be used to produce a second FEL pulse. This method offers independent control in photon energy and delay. For the parameters of the soft X-ray beamline Athos at the SwissFEL facility the photon energy tuning range is a factor of five with an adjustable delay between the two pulses from −50 to 950 fs.

PHASE is a software tool for physical optics simulation based on the stationary phase approximation method. The code is under continuous development since about 20 years and has been used for instance for fundamental studies and ray tracing of various beamlines at the Swiss Light Source. Along with the planning for SwissFEL a new hard X ray free electron laser under construction, new features have been added to permit practical performance predictions including diffraction effects which emerge with the fully coherent source. We present the application of the package on the example of the ARAMIS 1 beamline at SwissFEL. The X ray pulse calculated with GENESIS and given as an electrical field distribution has been propagated through the beamline to the sample position.We demonstrate the new features of PHASE like the treatment of measured figure errors, apertures and coatings of the mirrors and the application of Fourier optics propagators for free space propagation

Next-generation X-ray sources, based on the X-ray Free Electron Laser (XFEL) concept, will provide highly coherent, ultrashort pulses of soft and hard X-rays with peak intensity many orders of magnitude above that of a synchrotron. These pulses will allow studies of femtosecond dynamics at nanometer resolution and with chemical selectivity. They will produce coherent-diffraction images of organic and inorganic nanostructures without the deleterious effects of radiation damage.

A central, yet unresolved issue in the pathogenesis of HIV disease is the mechanism of antibody perturbation. In this study, HIV-specific memory B-cells were quantified in groups of infected subjects and compared with memory responses to other antigens and antibody titers. HIV-specific memory B-cell responses were vigorous in individuals with CD4(+) T-cell counts350/microl and weak or undetectable in subjects with CD4(+) T-cell numbers200/microl. Memory B-cell loss was permanent, because antiretroviral therapy failed to restore HIV-specific memory responses while influenza- and tetanus toxoid-specific memory B-cells remained unaffected or recovered. Antibody titers to Gag strongly correlated with memory B-cell frequencies. In contrast, Env-specific antibodies were maintained in advanced disease despite low or undetectable levels of memory B-cells. These results provide a potential mechanism by which destruction of HIV-specific CD4(+) T-cells affects the humoral immune response against HIV and compromises the ability to maintain an effective antibody response.

Protein tagging with a peptide is a commonly used technique to facilitate protein detection and to carry out protein purification. Flexibility with respect to the peptide tag is essential since no single tag suites all purposes. This report describes the usage of two short peptides from the SARS-associated coronavirus nucleocapsid (SARS-N) protein as protein tags. Plasmids for the generation of tagged proteins were generated by ligating synthetic oligonucleotides for the peptide-coding regions downstream of the protein coding sequence. The data show recognition of prokaryotically expressed HIV-1 Gag/p24 fusion protein by Western blot and efficient affinity purification using monoclonal antibodies against the tags. The SARS peptide antibody system described presents an alternative tagging opportunity in the growing field of protein science.

There is a large interest in the production of high power, ultra-short, 1 fs or less, coherent X-ray pulses, for atomic physics and other applications. However, the present design of X-ray SASE FELs leads to an X-ray pulse about 100 times longer. The time structure of the X-ray pulse is determined by several factors, mainly the electron bunch length and the FEL cooperation length. Until now all FELs have been working in the regime where the electron bunch length is much larger than the cooperation length. In the case of an X-ray SASE FEL this means that the X-ray pulse consists of a series of spikes, a few hundred in the case of the LCLS, and the bunch length is of the order of 200 fs. Several methods to reduce the bunch length to the 10 - 1 fs region have been proposed. These methods are based on electron bunch manipulation to cut the lasing part of the bunch to a fraction of the total length, thus reducing the X-ray pulse length. We are considering here a different method, using ultra-short, very low charge electron bunches, with a length of the order or shorter than the FEL cooperation length. In this case the X-ray pulse length after amplification in the undulator is a few times the electron bunch length. Our simulations show that in the case of LCLS and SPARX, operating with a 1 pC electron beam, we obtain X-ray pulses as short as 300 as, transform limited in the SPARX case, and almost transform limited in the LCLS case. Compared to other modes of producing short X-ray bunches this one offers the advantage of very small background signal, since all electrons are used to lase, and the charge is very small.

The capability for simulation of the dynamics of a free-electron laser including the higher-order harmonics in linear undulators exists in several existing codes as MEDUSA [H.P. Freund, S.G. Biedron, and S.V. Milton, IEEE J. Quantum Electron. 27 (2000) 243; H.P. Freund, Phys. Rev. ST-AB 8 (2005) 110701] and PERSEO [L. Giannessi, Overview of Perseo, a system for simulating FEL dynamics in Mathcad, 〈 http://www.jacow.org 〉, in: Proceedings of FEL 2006 Conference, BESSY, Berlin, Germany, 2006, p. 91], and has been recently implemented in GENESIS 1.3 [See 〈 http://www.perseo.enea.it 〉]. MEDUSA and GENESIS also include the dynamics of even harmonics induced by the coupling through the betatron motion. In addition MEDUSA, which is based on a non-wiggler averaged model, is capable of simulating the generation of even harmonics in the transversally cold beam regime, i.e. when the even harmonic coupling arises from non-linear effects associated with longitudinal particle dynamics and not to a finite beam emittance. In this paper a comparison between the predictions of the codes in different conditions is given.

Free-Electron Lasers as high-brilliance radiation sources, rely on a high quality of the electron beam driving the FEL process. The amount of energy, transferred from the electrons to the radiation field, and thus the efficiency of the FEL depends on the provided beam parameters. The presentation discusses the impact of various beam parameter and how current designs of FEL injector try to accomplish the demands on the beam quality for reaching saturation.

A recent breakthrough in laser-plasma accelerators, based upon ultrashort high-intensity lasers, demonstrated the generation of quasi-monoenergetic GeV-electrons. With future Petawatt lasers ultra-high beam currents of ~100 kA in ~10 fs can be expected, allowing for drastic reduction in the undulator length of free-electron-lasers (FELs). We present a discussion of the key aspects of a table-top FEL design, including energy loss and chirps induced by space-charge and wakefields. These effects become important for an optimized table-top FEL operation. A first proof-of-principle VUV case is considered as well as a table-top X-ray-FEL which may open a brilliant light source also for new ways in clinical diagnostics., Comment: 6 pages, 4 figures; accepted for publication in Appl. Phys. B

X-ray free-electron lasers (XFELs) are cutting-edge research tools that produce almost fully coherent radiation with high power and short-pulse length with applications in multiple science fields. There is a strong demand to achieve even shorter pulses and higher radiation powers than the ones obtained at state-of-the-art XFEL facilities. In this context we propose a novel method to generate terawatt-attosecond XFEL pulses, where an XFEL pulse is pushed through several short good-beam regions of the electron bunch. In addition to the elements of conventional XFEL facilities, the method uses only a multiple-slotted foil and small electron delays between undulator sections. Our scheme is thus simple, compact, and easy to implement both in already operating as well as future XFEL projects. We present numerical simulations that confirm the feasibility and validity of our proposal.