Orientia tsutsugamushi secretes two ankyrin repeat-containing effectors via a type 1 secretion system to inhibit host NF-κB function

Scrub typhus is a potentially fatal infection that threatens one billion persons in the Asia-Pacific region and is caused by the obligate intracellular bacterium, Orientia tsutsugamushi. How this organism facilitates its intracellular survival and pathogenesis is poorly understood. Intracellular bacterial pathogens utilize the Type 1 (T1SS) or Type 4 secretion system (T4SS) to translocate ankyrin repeat-containing proteins (Anks) into the host cell to modulate host cell processes. The O. tsutsugamushi genome encodes one of the largest known bacterial Ank libraries as well as Type 1 and Type 4 secretion systems (T1SS and T4SS), which are expressed during infection. In silico analyses of the Anks’ C-termini revealed that they possess characteristics of T1SS secretion signals. Escherichia coli expressing a functional T1SS was able to secrete chimeric hemolysin proteins bearing the C-termini of 19 of 20 O. tsutsugamushi Anks. In addition to infecting endothelial cells, O. tsutsugamushi infects professional phagocytes. To better understand why these innate immune cells are unable to eliminate O. tsutsugamushi, we addressed the activity of host NF-κB proinflammatory transcription factor. Screening of O. tsutsugamushi infected cells at an MOI of 1 revealed inhibition of NF-κB nuclear accumulation as early as 8 hours in HeLa and bone-marrow derived macrophage cells. When stimulating infected cells with TNF-α, IκBα degradation still occurs, however NF-κB dependent gene transcription remains downregulated. Immunofluorescence microscopic analysis of TNF-α treated cells ectopically expressing all O. tsutsugamushi Anks revealed that two nuclear trafficking Anks, Ank1 and Ank6, result in a significant decrease in NF-κB nuclear accumulation. Additionally, these Anks also significantly inhibited NF-κB dependent gene transcription. Co-immunoprecipitation experiments revealed that both Anks interact with importin-β1, exportin-1, and the p65 NF-κB subunit. Treating cells with importazole significantly reduces the nuclear accumulation of Ank1 and Ank6. Finally, treating infected cells or cells ectopically expressing Ank1 or Ank6 with leptomycin B resulted in restoration of NF-κB nuclear accumulation. With these data, we propose that O. tsutsugamushi secretes Ank1 and Ank6 to initially interact with importin-β1, which permits their nuclear entry where they then interact with NF-κB and subsequently exportin-1 to prevent NF-κB nuclear accumulation

Identification and characterization of point defects in aluminum nitride and zinc oxide crystals

Electron paramagnetic resonance (EPR) and electron-nuclear double resonance (ENDOR) studies have been performed on single crystals of aluminum nitride (AlN) and zinc oxide (ZnO), two wide-band-gap semiconductors having the wurtzite crystal structure. These studies were used to characterize point defects in each material. In the first study in AlN, new EPR and ENDOR spectra were acquired from a deep donor. Although observed in as-grown crystals, exposure to x rays significantly increased the concentration of this center. ENDOR identified a strong hyperfine interaction with one aluminum neighbor along the c axis and weaker equivalent hyperfine interactions with three additional aluminum neighbors in the basal plane. These aluminum interactions indicate that the responsible center was located at a nitrogen site. The observed paramagnetic defect is either an oxygen substituting for nitrogen or a nitrogen vacancy. An analysis of the hyperfine data suggests that substitutional oxygen is the most likely candidate.;The second point defect studied in AlN was silicon substituting for aluminum. Silicon is a shallow donor in AlN, and its neutral charge state is paramagnetic. Two samples containing silicon were studied. Only one of the samples was intentionally doped with silicon. The silicon-related EPR signals from these two samples had different behaviors. The signal from the doped sample had behavior similar to that described in previous studies where the silicon was explained as a DX center. The undoped sample had behavior that was inconsistent with a DX center.;In ZnO, EPR was used to monitor oxygen vacancies and zinc vacancies in a ZnO crystal irradiated near room temperature with 1.5 MeV electrons. Out-of-phase detection at 30 K greatly enhanced the EPR signals from these vacancies. Following the electron irradiation, but before illumination, Fe3+ ions and nonaxial singly ionized zinc vacancies were observed. Illumination with 325 nm laser light at low temperature eliminated the Fe3+ signal while producing spectra from singly ionized oxygen vacancies, neutral zinc vacancies, and axial singly ionized zinc vacancies. This light also produced EPR spectra from zinc vacancies having an OH- ion at an adjacent oxygen site. The low temperature response of the irradiated crystal to illumination wavelengths between 350 and 750 nm is described. Wavelengths shorter than 600 nm converted Fe3+ ions to Fe2+ ions and converted neutral oxygen vacancies to singly ionized oxygen vacancies. Neutral zinc vacancies were formed by wavelengths shorter than 500 nm as electrons were removed from isolated singly ionized zinc vacancies. Warming above 120 K in the dark reversed the effect of the illuminations. These wavelength-dependence results suggest that the ground state of the neutral oxygen vacancy is deep, approximately 1.3 eV above the valence band, and that the ground state of the singly ionized zinc vacancy is also deep, about 0.9 eV above the valence band.;The hyperfine associated with the isolated nitrogen acceptor in ZnO was studied with EPR. The sample used in this study was grown by the seeded chemical vapor transport method, with N2 added to the gas stream to serve as the doping source. This study further characterized the hyperfine interactions with the nitrogen nucleus (I = 1) and the nearest-neighbor zinc nuclei (I = 5/2). Angular dependence data were obtained from EPR and were analyzed by complete diagonalizations of the full spin Hamiltonian. Nuclear electric quadrupole effects were included in the nitrogen hyperfine analysis, thus yielding a value for the nuclear quadrupole and more accurate g values and nitrogen hyperfine parameters

Changed Patterns of Genomic Variation Following Recent Domestication: Selection Sweeps in Farmed Atlantic Salmon

The introduction of wild Atlantic salmon into captivity, and their subsequent artificial selection for production traits, has caused phenotypic differences between domesticated fish and their wild counterparts. Identification of regions of the genome underling these changes offers the promise of characterizing the early biological consequences of domestication. In the current study, we sequenced a population of farmed European Atlantic salmon and compared the observed patterns of SNP variation to those found in conspecific wild populations. This identified 139 genomic regions that contained significantly elevated SNP homozygosity in farmed fish when compared to their wild counterparts. The most extreme was adjacent to versican, a gene involved in control of neural crest cell migration. To control for false positive signals, a second and independent dataset of farmed and wild European Atlantic salmon was assessed using the same methodology. A total of 81 outlier regions detected in the first dataset showed significantly reduced homozygosity within the second one, strongly suggesting the genomic regions identified are enriched for true selection sweeps. Examination of the associated genes identified a number previously characterized as targets of selection in other domestic species and that have roles in development, behavior and olfactory system. These include arcvf, sema6, errb4, id2-like, and 6n1-like genes. Finally, we searched for evidence of parallel sweeps using a farmed population of North American origin. This failed to detect a convincing overlap to the putative sweeps present in European populations, suggesting the factors that drive patterns of variation under domestication and early artificial selection were largely independent. This is the first analysis on domestication of aquaculture species exploiting whole-genome sequence data and resulted in the identification of sweeps common to multiple independent populations of farmed European Atlantic salmon

Assessing Ecological and Economic Effects of Derelict Fishing Gear: a Guiding Framework

Developing standardized protocols to assess the ecological and socio-economic effects of marine debris – especially, derelict fishing gear – is critical for the protection of natural resources and for evaluating policies and programs designed to reduce and remove debris. This document outlines a Derelict Fishing Gear Assessment Framework to guide the development and implementation of derelict gear assessment, management and mitigation. The framework draws from techniques and protocols developed to assess derelict crab traps effects in the Chesapeake Bay and on past derelict gear assessments either conducted by or known to the framework authors. However, this framework is generalized and intended to be used by any stakeholder with a need to assess the status of derelict fishing gear and its economic and ecological effects on living resources, habitats, ecosystems, and local economies. It provides a generalized pathway and processes for assessing the effects of derelict fishing gear, and is flexible and scalable so that users of the framework can make informed decisions when data are limited, and can tailor it to satisfy their specific assessment goals and objectives if a full scale assessment is not required. The framework recommends best practices for each of five key elements

Adjuvant chemotherapy with or without bevacizumab in patients with resected non-small-cell lung cancer (E1505): an open-label, multicentre, randomised, phase 3 trial.

BackgroundAdjuvant chemotherapy for resected early-stage non-small-cell lung cancer (NSCLC) provides a modest survival benefit. Bevacizumab, a monoclonal antibody directed against VEGF, improves outcomes when added to platinum-based chemotherapy in advanced-stage non-squamous NSCLC. We aimed to evaluate the addition of bevacizumab to adjuvant chemotherapy in early-stage resected NSCLC.MethodsWe did an open-label, randomised, phase 3 trial of adult patients (aged ≥18 years) with an Eastern Cooperative Oncology Group performance status of 0 or 1 and who had completely resected stage IB (≥4 cm) to IIIA (defined by the American Joint Committee on Cancer 6th edition) NSCLC. We enrolled patients from across the US National Clinical Trials Network, including patients from the Eastern Cooperative Oncology Group-American College of Radiology Imaging Network (ECOG-ACRIN) affiliates in Europe and from the Canadian Cancer Trials Group, within 6-12 weeks of surgery. The chemotherapy regimen for each patient was selected before randomisation and administered intravenously; it consisted of four 21-day cycles of cisplatin (75 mg/m2 on day 1 in all regimens) in combination with investigator's choice of vinorelbine (30 mg/m2 on days 1 and 8), docetaxel (75 mg/m2 on day 1), gemcitabine (1200 mg/m2 on days 1 and 8), or pemetrexed (500 mg/m2 on day 1). Patients in the bevacizumab group received bevacizumab 15 mg/kg intravenously every 21 days starting with cycle 1 of chemotherapy and continuing for 1 year. We randomly allocated patients (1:1) to group A (chemotherapy alone) or group B (chemotherapy plus bevacizumab), centrally, using permuted blocks sizes and stratified by chemotherapy regimen, stage of disease, histology, and sex. No one was masked to treatment assignment, except the Data Safety and Monitoring Committee. The primary endpoint was overall survival, analysed by intention to treat. This trial is registered with ClinicalTrials.gov, number NCT00324805.FindingsBetween June 1, 2007, and Sept 20, 2013, 1501 patients were enrolled and randomly assigned to the two treatment groups: 749 to group A (chemotherapy alone) and 752 to group B (chemotherapy plus bevacizumab). 383 (26%) of 1458 patients (with complete staging information) had stage IB, 636 (44%) had stage II, and 439 (30%) had stage IIIA disease (stage of disease data were missing for 43 patients). Squamous cell histology was reported for 422 (28%) of 1501 patients. All four cisplatin-based chemotherapy regimens were used: 377 (25%) patients received vinorelbine, 343 (23%) received docetaxel, 283 (19%) received gemcitabine, and 497 (33%) received pemetrexed. At a median follow-up of 50·3 months (IQR 32·9-68·0), the estimated median overall survival in group A has not been reached, and in group B was 85·8 months (95% CI 74·9 to not reached); hazard ratio (group B vs group A) 0·99 (95% CI 0·82-1·19; p=0·90). Grade 3-5 toxicities of note (all attributions) that were reported more frequently in group B (the bevacizumab group) than in group A (chemotherapy alone) were overall worst grade (ie, all grade 3-5 toxicities; 496 [67%] of 738 in group A vs 610 [83%] of 735 in group B), hypertension (60 [8%] vs 219 [30%]), and neutropenia (241 [33%] vs 275 [37%]). The number of deaths on treatment did not differ between the groups (15 deaths in group A vs 19 in group B). Of these deaths, three in group A and ten in group B were considered at least possibly related to treatment.InterpretationAddition of bevacizumab to adjuvant chemotherapy did not improve overall survival for patients with surgically resected early-stage NSCLC. Bevacizumab does not have a role in this setting and should not be considered as an adjuvant therapy for patients with resected early-stage NSCLC.FundingNational Cancer Institute of the National Institutes of Health

SARS-CoV-2 spreads through cell-to-cell transmission

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a highly transmissible coronavirus responsible for the global COVID-19 pandemic. Herein, we provide evidence that SARS-CoV-2 spreads through cell-cell contact in cultures, mediated by the spike glycoprotein. SARS-CoV-2 spike is more efficient in facilitating cell-to-cell transmission than is SARS-CoV spike, which reflects, in part, their differential cell-cell fusion activity. Interestingly, treatment of cocultured cells with endosomal entry inhibitors impairs cell-to-cell transmission, implicating endosomal membrane fusion as an underlying mechanism. Compared with cell-free infection, cell-to-cell transmission of SARS-CoV-2 is refractory to inhibition by neutralizing antibody or convalescent sera of COVID-19 patients. While angiotensin-converting enzyme 2 enhances cell-to-cell transmission, we find that it is not absolutely required. Notably, despite differences in cell-free infectivity, the authentic variants of concern (VOCs) B.1.1.7 (alpha) and B.1.351 (beta) have similar cell-to-cell transmission capability. Moreover, B.1.351 is more resistant to neutralization by vaccinee sera in cell-free infection, whereas B.1.1.7 is more resistant to inhibition by vaccinee sera in cell-to-cell transmission. Overall, our study reveals critical features of SARS-CoV-2 spike-mediated cell-to-cell transmission, with important implications for a better understanding of SARS-CoV-2 spread and pathogenesis

Expedition 306 summary

The overall aim of the North Atlantic paleoceanography study of Integrated Ocean Drilling Program Expedition 306 is to place late Neogene–Quaternary climate proxies in the North Atlantic into a chronology based on a combination of geomagnetic paleointensity, stable isotope, and detrital layer stratigraphies, and in so doing generate integrated North Atlantic millennial-scale stratigraphies for the last few million years. To reach this aim, complete sedimentary sections were drilled by multiple advanced piston coring directly south of the central Atlantic “ice-rafted debris belt” and on the southern Gardar Drift. In addition to the North Atlantic paleoceanography study, a borehole observatory was successfully installed in a new ~180 m deep hole close to Ocean Drilling Program Site 642, consisting of a circulation obviation retrofit kit to seal the borehole from the overlying ocean, a thermistor string, and a data logger to document and monitor bottom water temperature variations through time