Multi-pathogens sequence containing plasmids as positive controls for universal detection of potential agents of bioterrorism

BACKGROUND: The limited circulation of many of the agents that are likely to be used in a bioterrorism attack precludes the ready availability of positive controls. This means that only specialized laboratories can screen for the presence of these agents by nucleic amplification assays. Calibrated controls are also necessary for quantitative measurements. Primers and probes to be used in both conventional and real-time PCR assays were designed for the detection of agents likely to be used by a bioterrorist. Three plasmids, each of which contains 4 to 6 specific sequences from agents on the CDC Category A and B list (excluding RNA viruses) were constructed. Two plasmids incorporate the sequences of Category A and B agents, respectively. The third plasmid incorporates sequences from Variola major and organisms that cause rash-like illnesses that may be clinically confused with smallpox. An "exogenic sequence", introducing a NotI restriction site was incorporated in the native sequences of the bioterrorism agents inserted in plasmids. The designed molecular system for detection of bioterrorism agents was tested on each of these agents (except Monkeypox virus, Smallpox virus and 2 Burkholderia species for which no native DNA was available) and a collection of 50 isolates of C. burnetii using constructed plasmids as positive controls. RESULTS: Designed primers and probes allowed molecular detection, in either single or multiplex assays, of agent-specific targets with analytical sensitivities of between 1 and 100 DNA copies. The plasmids could be used as positive controls. False-positive results due to contamination by the positive control were easily detected by sequencing and eliminated by digestion with NotI. CONCLUSION: Plasmid A and B can be used as positive controls in molecular assays for the detection of bioterrorism agents in clinical specimens or environmental samples. Plasmid C can be used as a positive control in differentiation of vesicular rashes. It is also possible to avoid or to ensure immediate detection of false positive results due to contamination by positive controls using these plasmids. These plasmids and the corresponding primers and probes are immediately available for all clinical microbiology laboratories provided they have molecular amplification equipment

Point of Care Strategy for Rapid Diagnosis of Novel A/H1N1 Influenza Virus

Within months of the emergence of the novel A/H1N1 pandemic influenza virus (nA/H1N1v), systematic screening for the surveillance of the pandemic was abandoned in France and in some other countries. At the end of June 2009, we implemented, for the public hospitals of Marseille, a Point Of Care (POC) strategy for rapid diagnosis of the novel A/H1N1 influenza virus, in order to maintain local surveillance and to evaluate locally the kinetics of the pandemic.Two POC laboratories, located in strategic places, were organized to receive and test samples 24 h/24. POC strategy consisted of receiving and processing naso-pharyngeal specimens in preparation for the rapid influenza diagnostic test (RIDT) and real-time RT-PCR assay (rtRT-PCR). This strategy had the theoretical capacity of processing up to 36 samples per 24 h. When the flow of samples was too high, the rtRT-PCR test was abandoned in the POC laboratories and transferred to the core virology laboratory. Confirmatory diagnosis was performed in the core virology laboratory twice a day using two distinct rtRT-PCR techniques that detect either influenza A virus or nA/N1N1v. Over a period of three months, 1974 samples were received in the POC laboratories, of which 111 were positive for nA/H1N1v. Specificity and sensitivity of RIDT were 100%, and 57.7% respectively. Positive results obtained using RIDT were transmitted to clinical practitioners in less than 2 hours. POC processed rtRT-PCR results were available within 7 hours, and rtRT-PCR confirmation within 24 hours.The POC strategy is of benefit, in all cases (with or without rtRT-PCR assay), because it provides continuous reception/processing of samples and reduction of the time to provide consolidated results to the clinical practitioners. We believe that implementation of the POC strategy for the largest number of suspect cases may improve the quality of patient care and our knowledge of the epidemiology of the pandemic

Online Monitoring of the Osiris Reactor with the Nucifer Neutrino Detector

Originally designed as a new nuclear reactor monitoring device, the Nucifer detector has successfully detected its first neutrinos. We provide the second shortest baseline measurement of the reactor neutrino flux. The detection of electron antineutrinos emitted in the decay chains of the fission products, combined with reactor core simulations, provides an new tool to assess both the thermal power and the fissile content of the whole nuclear core and could be used by the Inter- national Agency for Atomic Energy (IAEA) to enhance the Safeguards of civil nuclear reactors. Deployed at only 7.2m away from the compact Osiris research reactor core (70MW) operating at the Saclay research centre of the French Alternative Energies and Atomic Energy Commission (CEA), the experiment also exhibits a well-suited configuration to search for a new short baseline oscillation. We report the first results of the Nucifer experiment, describing the performances of the 0.85m3 detector remotely operating at a shallow depth equivalent to 12m of water and under intense background radiation conditions. Based on 145 (106) days of data with reactor ON (OFF), leading to the detection of an estimated 40760 electron antineutrinos, the mean number of detected antineutrinos is 281 +- 7(stat) +- 18(syst) electron antineutrinos/day, in agreement with the prediction 277(23) electron antineutrinos/day. Due the the large background no conclusive results on the existence of light sterile neutrinos could be derived, however. As a first societal application we quantify how antineutrinos could be used for the Plutonium Management and Disposition Agreement.Comment: 22 pages, 16 figures - Version

Exploring CEvNS with NUCLEUS at the Chooz Nuclear Power Plant

Coherent elastic neutrino-nucleus scattering (CE$\nu$NS) offers a unique way to study neutrino properties and to search for new physics beyond the Standard Model. Nuclear reactors are promising sources to explore this process at low energies since they deliver large fluxes of (anti-)neutrinos with typical energies of a few MeV. In this paper, a new-generation experiment to study CE$\nu$NS is described. The NUCLEUS experiment will use cryogenic detectors which feature an unprecedentedly low energy threshold and a time response fast enough to be operated in above-ground conditions. Both sensitivity to low-energy nuclear recoils and a high event rate tolerance are stringent requirements to measure CE$\nu$NS of reactor antineutrinos. A new experimental site, denoted the Very-Near-Site (VNS) at the Chooz nuclear power plant in France is described. The VNS is located between the two 4.25 GW$_{\mathrm{th}}$ reactor cores and matches the requirements of NUCLEUS. First results of on-site measurements of neutron and muon backgrounds, the expected dominant background contributions, are given. In this paper a preliminary experimental setup with dedicated active and passive background reduction techniques is presented. Furthermore, the feasibility to operate the NUCLEUS detectors in coincidence with an active muon-veto at shallow overburden is studied. The paper concludes with a sensitivity study pointing out the promising physics potential of NUCLEUS at the Chooz nuclear power plant

CeLAND: search for a 4th light neutrino state with a 3 PBq 144Ce-144Pr electron antineutrino generator in KamLAND

The reactor neutrino and gallium anomalies can be tested with a 3-4 PBq (75-100 kCi scale) 144Ce-144Pr antineutrino beta-source deployed at the center or next to a large low-background liquid scintillator detector. The antineutrino generator will be produced by the Russian reprocessing plant PA Mayak as early as 2014, transported to Japan, and deployed in the Kamioka Liquid Scintillator Anti-Neutrino Detector (KamLAND) as early as 2015. KamLAND's 13 m diameter target volume provides a suitable environment to measure the energy and position dependence of the detected neutrino flux. A characteristic oscillation pattern would be visible for a baseline of about 10 m or less, providing a very clean signal of neutrino disappearance into a yet-unknown, sterile neutrino state. This will provide a comprehensive test of the electron dissaperance neutrino anomalies and could lead to the discovery of a 4th neutrino state for Delta_m^2 > 0.1 eV^2 and sin^2(2theta) > 0.05.Comment: 67 pages, 50 figures. Th. Lasserre thanks the European Research Council for support under the Starting Grant StG-30718

White paper: CeLAND - Investigation of the reactor antineutrino anomaly with an intense 144Ce-144Pr antineutrino source in KamLAND

We propose to test for short baseline neutrino oscillations, implied by the recent reevaluation of the reactor antineutrino flux and by anomalous results from the gallium solar neutrino detectors. The test will consist of producing a 75 kCi 144Ce - 144Pr antineutrino source to be deployed in the Kamioka Liquid Scintillator Anti-Neutrino Detector (KamLAND). KamLAND's 13m diameter target volume provides a suitable environment to measure energy and position dependence of the detected neutrino flux. A characteristic oscillation pattern would be visible for a baseline of about 10 m or less, providing a very clean signal of neutrino disappearance into a yet-unknown, "sterile" state. Such a measurement will be free of any reactor-related uncertainties. After 1.5 years of data taking the Reactor Antineutrino Anomaly parameter space will be tested at > 95% C.L.Comment: White paper prepared for Snowmass-2013; slightly different author lis

Essential versus accessory aspects of cell death: recommendations of the NCCD 2015

Cells exposed to extreme physicochemical or mechanical stimuli die in an uncontrollable manner, as a result of their immediate structural breakdown. Such an unavoidable variant of cellular demise is generally referred to as ‘accidental cell death’ (ACD). In most settings, however, cell death is initiated by a genetically encoded apparatus, correlating with the fact that its course can be altered by pharmacologic or genetic interventions. ‘Regulated cell death’ (RCD) can occur as part of physiologic programs or can be activated once adaptive responses to perturbations of the extracellular or intracellular microenvironment fail. The biochemical phenomena that accompany RCD may be harnessed to classify it into a few subtypes, which often (but not always) exhibit stereotyped morphologic features. Nonetheless, efficiently inhibiting the processes that are commonly thought to cause RCD, such as the activation of executioner caspases in the course of apoptosis, does not exert true cytoprotective effects in the mammalian system, but simply alters the kinetics of cellular demise as it shifts its morphologic and biochemical correlates. Conversely, bona fide cytoprotection can be achieved by inhibiting the transduction of lethal signals in the early phases of the process, when adaptive responses are still operational. Thus, the mechanisms that truly execute RCD may be less understood, less inhibitable and perhaps more homogeneous than previously thought. Here, the Nomenclature Committee on Cell Death formulates a set of recommendations to help scientists and researchers to discriminate between essential and accessory aspects of cell death

Defining Life: The Virus Viewpoint

Are viruses alive? Until very recently, answering this question was often negative and viruses were not considered in discussions on the origin and definition of life. This situation is rapidly changing, following several discoveries that have modified our vision of viruses. It has been recognized that viruses have played (and still play) a major innovative role in the evolution of cellular organisms. New definitions of viruses have been proposed and their position in the universal tree of life is actively discussed. Viruses are no more confused with their virions, but can be viewed as complex living entities that transform the infected cell into a novel organism—the virus—producing virions. I suggest here to define life (an historical process) as the mode of existence of ribosome encoding organisms (cells) and capsid encoding organisms (viruses) and their ancestors. I propose to define an organism as an ensemble of integrated organs (molecular or cellular) producing individuals evolving through natural selection. The origin of life on our planet would correspond to the establishment of the first organism corresponding to this definition