A closer look at mammalian antiviral condensates.

Several biomolecular condensates assemble in mammalian cells in response to viral infection. The most studied of these are stress granules (SGs), which have been proposed to promote antiviral innate immune signaling pathways, including the RLR-MAVS, the protein kinase R (PKR), and the OAS-RNase L pathways. However, recent studies have demonstrated that SGs either negatively regulate or do not impact antiviral signaling. Instead, the SG-nucleating protein, G3BP1, may function to perturb viral RNA biology by condensing viral RNA into viral-aggregated RNA condensates, thus explaining why viruses often antagonize G3BP1 or hijack its RNA condensing function. However, a recently identified condensate, termed double-stranded RNA-induced foci, promotes the activation of the PKR and OAS-RNase L antiviral pathways. In addition, SG-like condensates known as an RNase L-induced bodies (RLBs) have been observed during many viral infections, including SARS-CoV-2 and several flaviviruses. RLBs may function in promoting decay of cellular and viral RNA, as well as promoting ribosome-associated signaling pathways. Herein, we review these recent advances in the field of antiviral biomolecular condensates, and we provide perspective on the role of canonical SGs and G3BP1 during the antiviral response.

Broadening of the Divertor Heat Flux Profile in High Confinement Tokamak Fusion Plasmas with Edge Pedestals Limited by Turbulence in DIII-D.

Multimachine empirical scaling predicts an extremely narrow heat exhaust layer in future high magnetic field tokamaks, producing high power densities that require mitigation. In the experiments presented, the width of this exhaust layer is nearly doubled using actuators to increase turbulent transport in the plasma edge. This is achieved in low collisionality, high confinement edge pedestals with their gradients limited by turbulent transport instead of large-scale, coherent instabilities. The exhaust heat flux profile width and divertor leg diffusive spreading both double as a high frequency band of turbulent fluctuations propagating in the electron diamagnetic direction doubles in amplitude. The results are quantitatively reproduced in electromagnetic XGC particle-in-cell simulations which show the heat flux carried by electrons emerges to broaden the heat flux profile, directly supported by Langmuir probe measurements.

Mechanisms and consequences of mRNA destabilization during viral infections.

During viral infection there is dynamic interplay between the virus and the host to regulate gene expression. In many cases, the host induces the expression of antiviral genes to combat infection, while the virus uses "host shut-off" systems to better compete for cellular resources and to limit the induction of the host antiviral response. Viral mechanisms for host shut-off involve targeting translation, altering host RNA processing, and/or inducing the degradation of host mRNAs. In this review, we discuss the diverse mechanisms viruses use to degrade host mRNAs. In addition, the widespread degradation of host mRNAs can have common consequences including the accumulation of RNA binding proteins in the nucleus, which leads to altered RNA processing, mRNA export, and changes to transcription.

An Engineered Monovalent Anti-TNF-α Antibody with pH-Sensitive Binding Abrogates Immunogenicity in Mice following a Single Intravenous Dose.

Therapeutic Abs directed toward TNF-α display significant immunogenicity in humans, frequently leading to lower serum concentrations of the Ab that are associated with lower treatment efficacy. The enhanced incidence of immunogenicity observed with this class of therapeutics may be mediated by the expression of TNF-α as a homotrimer, both as a soluble serum protein and as a membrane-associated protein (mTNF-α) on the surface of dendritic cells. The TNF-α homotrimer enables the formation of polyvalent Ab-TNF-α immune complexes (ICs) that enhance binding to FcR and neonatal FcR. Polyvalent ICs and Ab bound to mTNF-α on the surface of dendritic cells can internalize, traffic to the lysosomes, and be processed for presentation by MHC molecules. To diminish immunogenicity caused by trafficking of ICs and mTNF-α to the lysosomes, we engineered a monovalent format of adalimumab with pH-sensitive binding to TNF-α. The engineered variant, termed AF-M2637, did not cross-link TNF-α trimers and consequently formed small, nonprecipitating ICs only. AF-M2637 bound TNF-α with high affinity at pH 7.4 (EC50 = 1.1 nM) and displayed a significantly faster dissociation rate than adalimumab at pH 6.0. No immune response to AF-M2637 was detected in mice following a single i.v. dose. In contrast, rapid immunization was detected following the injection of a single i.v. dose of adalimumab, monovalent adalimumab, or the bivalent form of the pH-sensitive variant. These data suggest that ICs and mTNF-α both contribute to the immunogenicity of adalimumab in mice and provide a general strategy for engineering less immunogenic therapeutic TNF-α Abs.

Profile of and risk factors for early unplanned readmissions in patients with acute necrotizing pancreatitis.

INTRODUCTION: Acute necrotizing pancreatitis (ANP) complicates up to 15% of acute pancreatitis cases. ANP has historically been associated with a significant risk for readmission, but there are currently no studies exploring factors that associate with risk for unplanned, early (<30-day) readmissions in this patient population. METHODS: We performed a retrospective review of all consecutive patients presenting to hospitals in the Indiana University (IU) Health system with pancreatic necrosis between December 2016 and June 2020. Patients younger than 18 years of age, without confirmed pancreatic necrosis and those that suffered in-hospital mortality were excluded. Logistic regression was performed to identify potential predictors of early readmission in this group of patients. RESULTS: One hundred and sixty-two patients met study criteria. 27.7% of the cohort was readmitted within 30-days of index discharge. The median time to readmission was 10 days (IQR 5-17 days). The most frequent reason for readmission was abdominal pain (75.6%), followed by nausea and vomiting in (35.6%). Discharge to home was associated with 93% lower odds of readmission. We found no additional clinical factors that predicted early readmission. CONCLUSION: Patients with ANP have a significant risk for early (<30 days) readmission. Direct discharge to home, rather than short or long-term rehabilitation facilities, is associated with lower odds of early readmission. Analysis was otherwise negative for independent, clinical predictors of early unplanned readmissions in ANP.

An epidemiological investigation of COVID-19 outbreaks in a group of care homes in Wales, UK: a retrospective cohort study.

BACKGROUND: This study describes the epidemiology of COVID-19 outbreaks in four care homes in terms of spread, severity, presentation and interventions. METHODS: Participants were 100 residents and 102 staff from four co-located care homes in Wales. Data were collected from the homes and Public Health Wales, including demographics, presentations, test status and results, hospital admissions and deaths. Genomic sequencing of confirmed case samples was completed, where possible. Epi-curves, crude attack rates, a Kaplan-Meier survival curve and adjusted hazard ratios were calculated using R. RESULTS: About 14 confirmed and 43 possible resident cases, 23 confirmed and 47 possible staff cases occurred. Crude attack rates of possible and confirmed cases were 57% (residents) and 69% (staff). Genomic sequencing for 10 confirmed case PCR samples identified at least 5 different UK lineages of COVID-19.42 (42%) residents died, 23 (55%) with COVID-19 or suspected COVID-19 recorded on the death certificate. The hazard ratio for death amongst resident possible and confirmed cases compared to null cases, adjusting for age and sex, was 13.26 (95% CI 5.61-31.34). CONCLUSIONS: There were extensive outbreaks of COVID-19 in these homes with high crude attack rates and deaths. Universal testing and early isolation of residents are recommended.

Prenatal infection with Mycoplasma pulmonis in rats exaggerates the angiotensin II pressor response in adult offspring.

Exposure to different stressors in utero is linked to adult diseases such as obesity and hypertension. In this study, the impact of prenatal infection (PNI) on adult body weight and cardiovascular function was evaluated using a naturally occurring rodent pathogen, Mycoplasma pulmonis (MP). Pregnant Sprague-Dawley rats were infected with MP on gestationalday 14 and gave birth naturally. Adult PNI offspring weighed more than controls, but resting mean arterial pressure (MAP) was unchanged. Subcutaneous injection of angiotensin II (10 µg/kg) elicited a rise in MAP that was greater in both male and female PNI offspring compared with controls (P < 0.03). The accompanying reflex bradycardia was similar to the controls, suggesting that PNI induced baroreflex dysfunction. Subcutaneous nicotine administration, a potent cardiorespiratory stimulus, also elicited a transient rise in MAP that was generally greater in the PNI group, but the change in MAP from baseline was only significant in the PNI females compared with controls (P < 0.03). Elevated body weight and cardiovascular reactivity in the PNI offspring was associated with an increase in the ratio of hypothalamic corticotrophin-releasing hormone receptors type 1 to type 2 gene expression in both sexes compared with controls. These findings support previous studies demonstrating that PNI induces alterations in cardiovascular function and body weight. Yet, unlike previous studies utilizing other models of PNI (e.g., endotoxin), MP PNI did not induce resting hypertension. Thus, our study provides a foundation for future studies evaluating the cardiovascular risks of offspring exposed to microbial challenges in utero.

First Evidence of Local E×B Drift in the Divertor Influencing the Structure and Stability of Confined Plasma near the Edge of Fusion Devices.

The structure of the edge plasma in a magnetic confinement system has a strong impact on the overall plasma performance. We uncover for the first time a magnetic-field-direction dependent density shelf, i.e., local flattening of the density radial profile near the magnetic separatrix, in high confinement plasmas with low edge collisionality in the DIII-D tokamak. The density shelf is correlated with a doubly peaked density profile near the divertor target plate, which tends to occur for operation with the ion B×∇B drift direction away from the X-point, as currently employed for DIII-D advanced tokamak scenarios. This double-peaked divertor plasma profile is connected via the E×B drifts, arising from a strong radial electric field induced by the radial electron temperature gradient near the divertor target. The drifts lead to the reversal of the poloidal flow above the divertor target, resulting in the formation of the density shelf. The edge density shelf can be further enhanced at higher heating power, preventing large, periodic bursts of the plasma, i.e., edge-localized modes, in the edge region, consistent with ideal magnetohydrodynamics calculations.

The 1980s: D-AP5, LTP and a Decade of NMDA Receptor Discoveries.

In the 1960s and 70s, biochemical and pharmacological evidence was pointing toward glutamate as a synaptic transmitter at a number of distinct receptor classes, known as NMDA and non-NMDA receptors. The field, however, lacked a potent and highly selective antagonist to block these putative postsynaptic receptors. So, the discoveries in the early 1980s of D-AP5 as a selective NMDA receptor antagonist and of its ability to block synaptic events and plasticity were a major breakthrough leading to an explosion of knowledge about this receptor subtype. During the next 10 years, the role of NMDA receptors was established in synaptic transmission, long-term potentiation, learning and memory, epilepsy, pain, among others. Hints at pharmacological heterogeneity among NMDA receptors were followed by the cloning of separate subunits. The purpose of this review is to recognize the important contributions made in the 1980s by Graham L. Collingridge and other key scientists to the advances in our understanding of the functions of NMDA receptors throughout the central nervous system.

Brachionus leydigii (Monogononta: Ploima) reported from the western basin of Lake Erie.

Several species of non-indigenous planktonic invertebrates have historically been introduced to the Laurentian Great Lakes. Previous introductions of non-indigenous planktonic invertebrates to the Great Lakes have been crustacean zooplankton, specifically Cladocera and Copepoda. This report documents the first known occurrence of Brachionus leydigii var. tridentatus (Zernov, 1901) in Lake Erie and possibly the first detection of a non-indigenous rotifer species in the Laurentian Great Lakes. The specimen was collected from a U.S. EPA monitoring station in the western basin of Lake Erie on April 4, 2016.

Differences in dehydration tolerance among populations of a gametophyte-only fern.

PREMISE OF THE STUDY: For many plant species, historical climatic conditions may have left lasting imprints that are detectable in contemporary populations. Additionally, if these historical conditions also prevented gene flow among populations, these populations may be differentiated with respect to one another and their contemporary environmental conditions. For the fern, Vittaria appalachiana, one theory is that historical conditions during the Pleistocene largely shaped both the distribution and lack of sporophyte production. Our goals-based on this theory-were to examine physiological differences among and within populations spanning the species' geographic range, and the contribution of historical climatic conditions to this differentiation. METHODS: We exposed explants from five populations to four drying treatments and examined differences in physiological response. Additionally, we examined the role of historical and current climatic conditions in driving the observed population differentiation. KEY RESULTS: Populations differ in their ability to tolerate varying levels of dehydration, displaying a pattern of countergradient selection. Exposure to historical and contemporary climatic conditions, specifically variation in temperature and precipitation regimes, resulted in population divergence observed among contemporary populations. CONCLUSIONS: Historical conditions have shaped not only the distribution of V. appalachiana, but also its current physiological limitations. Results from this study support the hypothesis that climatic conditions during the Pleistocene are responsible for the distribution of this species, and may be responsible for the observed differences in dehydration tolerance. Additionally, dehydration tolerance may be the driving factor for previously reported patterns of countergradient selection in this species.

CMV in the gut: a critical review of CMV detection in the immunocompetent host with colitis.

As scientific techniques for the detection of cytomegalovirus (CMV) improve, we are able to detect small amounts of CMV in the mucosal wall. As clinicians, we are unsure how to interpret the results of this novel test. There is controversy in the literature as to the significance of the detection of CMV in the gut. Whilst the importance of CMV and reactivation of the virus is clear in those patients such as allograft recipients with established immune compromise, the role is less clear in patients with less damaged immune systems. We explore whether the detection of CMV in such cases influences outcome and how it should be optimally managed. We discuss the optimal management of such cases, according to current guidelines, with a review of the literature.

Branched isomeric 1,2,3-triazolium-based ionic liquids: new insight into structure-property relationships.

A series of four isomeric 1,2,3-triazolium-based ionic liquids (ILs) with vary degree of branching were synthesized and characterized to investigate the effect of ion branching on thermal and physical properties of the resulting IL. It was found that increased branching led to a higher ionicity and higher viscosity. The thermal properties were also altered significantly and spectral changes in the near edge X-ray absorption fine structure (NEXAFS) spectra show that branching affects intermolecular interaction. While the ionicity and viscosity varying linearly with branching, the MDSC and NEXAFS measurements show that the cation shape has a stronger influence on the melting temperature and absorptive properties than the number of branched alkyl substituents.

Evaluation of a membrane filtration method for the rapid enumeration of confirmed Clostridium perfringens from water.

UNLABELLED: A modification of the UK reference and ISO 14189 TSCA medium for the enumeration of Clostridium perfringens from water coupled with a membrane filter transfer technique for testing for production of acid phosphatase was evaluated. The new tryptose cycloserine agar (TCA) medium, which lacks sodium metabisulphite but contains sodium pyruvate to improve recovery, allows the isolation and confirmation of Cl. perfringens within 18-24 h of sample processing. Data from a multilaboratory study analysed according to ISO 17994 showed that TCA was equivalent to TSCA for the enumeration of Cl. perfringens. The identification of acid phosphatase-negative isolates revealed a false-negative rate for the TCA method of 0.8%. The TCA membrane filter transfer procedure provides confirmed Cl. perfringens counts in half the time of the TSCA method and is simple to undertake. SIGNIFICANCE AND IMPACT OF THE STUDY: The testing of drinking water for Clostridium perfringens is a regulatory parameter in Europe and the UK. Current UK and ISO methods employ membrane filtration (MF) and TSCA medium followed by subculture and confirmation of isolates by testing for acid phosphatase. This takes 48 h. We present here the results of a multilaboratory evaluation of a MF method that features a simplified isolation medium (TCA) and a membrane transfer procedure for the acid phosphatase test resulting in confirmed results being available in 18-24 h. This development significantly reduces the time to confirmed results for Cl. perfringens from water samples.

RNase L-induced bodies sequester subgenomic flavivirus RNAs and re-establish host RNA decay.

Subgenomic flavivirus RNAs (sfRNAs) are structured RNA elements encoded in the 3'-UTR of flaviviruses that promote viral infection by inhibiting cellular RNA decay machinery. Herein, we analyze the production of sfRNAs using single-molecule RNA fluorescence in situ hybridization (smRNA-FISH) and super-resolution microscopy during West Nile virus, Zika virus, or Dengue virus serotype 2 infection. We show that sfRNAs are initially localized diffusely in the cytosol or in processing bodies (P-bodies). However, upon activation of the host antiviral endoribonuclease, Ribonuclease L (RNase L), nearly all sfRNAs re-localize to antiviral biological condensates known as RNase L-induced bodies (RLBs). RLB-mediated sequestration of sfRNAs reduces sfRNA association with RNA decay machinery in P-bodies, which coincides with increased viral RNA decay. These findings establish a role of RLBs in promoting viral RNA decay, demonstrating the complex host-pathogen interactions at the level of RNA decay and biological condensation.

G3BP1-dependent condensation of translationally inactive viral RNAs antagonizes infection.

G3BP1 is an RNA binding protein that condenses untranslating messenger RNAs into stress granules (SGs). G3BP1 is inactivated by multiple viruses and is thought to antagonize viral replication by SG-enhanced antiviral signaling. Here, we show that neither G3BP1 nor SGs generally alter the activation of innate immune pathways. Instead, we show that the RNAs encoded by West Nile virus, Zika virus, and severe acute respiratory syndrome coronavirus 2 are prone to G3BP1-dependent RNA condensation, which is enhanced by limiting translation initiation and correlates with the disruption of viral replication organelles and viral RNA replication. We show that these viruses counteract condensation of their RNA genomes by inhibiting the RNA condensing function of G3BP proteins, hijacking the RNA decondensing activity of eIF4A, and/or maintaining efficient translation. These findings argue that RNA condensation can function as an intrinsic antiviral mechanism, which explains why many viruses inactivate G3BP proteins and suggests that SGs may have arisen as a vestige of this antiviral mechanism.

A comparison of Ki-67 and mitotic count as prognostic markers for metastatic pancreatic and midgut neuroendocrine neoplasms.

BACKGROUND: The aim of this study was to compare mitotic count (MC) and Ki-67 proliferation index as prognostic markers in pancreatic and midgut neuroendocrine neoplasms (NENs). METHODS: Two hundred eighty-five patients with metastatic NENs were recruited. Concordance between histological grade according to either Ki-67 or MC as defined by the European Neuroendocrine Tumour Society guidelines was assessed and the prognostic significance of Ki-67 or MC were evaluated. RESULTS: There was a discrepancy of 44 and 38% in grade assignment when using Ki-67 or MC in pancreatic and midgut NENs, respectively. In multivariate analysis, grade using Ki-67, but not MC, was a significant prognostic factor in determining overall survival (hazard ratios: midgut G2 2.34, G3 15.1, pancreas G2 2.08, G3 11.3). The prognostic value of Ki-67 was improved using a modified classification (hazard ratios: midgut G2 3.02, for G3 22.1, pancreas G2 5.97, G3 33.8). CONCLUSION: There is a lack of concordance between Ki-67 and MC in assigning tumour grade. Grade according to Ki-67 was a better prognostic marker than MC for metastatic pancreatic and midgut NENs. We suggest that Ki-67 alone should be used for grading pancreatic and midgut NENs and that the current threshold for classifying G1/G2 tumours should be revised from 2 to 5%.

Comparative study of the osteoblastic activity of two implant systems (Endopore versus Entegra) utilizing single photon emission computed tomography (SPECT): experimental study in pigs model.

Implantology has been an important component of dental management for over forty years, and during that period, many configurations of implant materials and methods have been developed. As empirical and clinical research yield new implant materials, there has been need to test and compare these materials to provide the most cost-effective and efficient implants. Evaluation of efficiency of implants has relied heavily on histological and radiological methods, but these one-dimensional measurement methods fail to evaluate the osteoblastic activity and osseointegration properties of putative implants. In this report, we describe the use of a quantitative single photon emission computed tomography (SPECT) as a tool for comparing the osseointegrating capabilities of two types of implants.

Single-cell analysis of RNase L-mediated mRNA decay.

Ribonuclease L (RNase L) is a mammalian endoribonuclease that initiates the mass degradation of cellular mRNAs in response to double-stranded RNA or viral infection. The kinetic rate of mRNA decay upon RNase L activation has been elusive because RNase L is heterogeneously activated with respect to time in individual cells. Herein, we describe a method using immunofluorescence combined with single-molecule fluorescence in situ hybridization (smFISH) to determine single-cell mRNA decay rates upon RNase L activation. Using these approaches, we deduce that the rate of mRNA decay upon RNase L activation is extremely rapid, whereby the half-life of stable mRNAs such as GAPDH mRNA is reduced to ∼15 minutes in individual cells. This allows for RNase L to degrade nearly every mRNA in a cell in less than 1 hour, which is much faster than the decay rate that would be derived using bulk measurement techniques for mRNA levels, such as qRT-PCR. These single-cell approaches can generally be employed to resolve mRNA decay kinetics in additional contexts.