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Background and Aims: Declines in cardiovascular mortality have stagnated in the USA since 2011. There is growing concern that these patterns reflect worsening cardiovascular health in younger adults. However, little is known about how the burden of acute cardiovascular hospitalizations and mortality has changed in this population. Changes in cardiovascular hospitalizations and mortality among adults aged 25-64 years were evaluated, overall and by community-level income., Methods: Using the National Inpatient Sample, age-standardized annual hospitalization and in-hospital mortality rates for acute myocardial infarction (AMI), heart failure, and ischaemic stroke were determined among adults aged 25-64 years. Quasi-Poisson and quasi-binominal regression models were fitted to compare outcomes between individuals residing in low- and higher-income communities., Results: Between 2008 and 2019, age-standardized hospitalization rates for AMI increased among younger adults from 155.0 (95% confidence interval: 154.6, 155.4) per 100 000 to 160.7 (160.3, 161.1) per 100 000 (absolute change +5.7 [5.0, 6.3], P < .001). Heart failure hospitalizations also increased (165.3 [164.8, 165.7] to 225.3 [224.8, 225.8], absolute change +60.0 (59.3, 60.6), P < .001), as ischaemic stroke hospitalizations (76.3 [76.1, 76.7] to 108.1 [107.8, 108.5], absolute change +31.7 (31.2, 32.2), P < .001). Across all conditions, hospitalizations rates were significantly higher among younger adults residing in low-income compared with higher-income communities, and disparities did not narrow between groups. In-hospital mortality decreased for all conditions over the study period., Conclusions: There was an alarming increase in cardiovascular hospitalizations among younger adults in the USA from 2008 to 2019, and disparities between those residing in low- and higher-income communities did not narrow., (© The Author(s) 2023. Published by Oxford University Press on behalf of the European Society of Cardiology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Cross-presentation of dead cell-associated antigens by conventional dendritic cells type 1 (cDC1s) is critical for CD8 + T cells response against many tumors and viral infections. It is facilitated by DNGR-1 (CLEC9A), an SYK-coupled cDC1 receptor that detects dead cell debris. Here, we report that DNGR-1 engagement leads to rapid activation of CBL and CBL-B E3 ligases to cause K63-linked ubiquitination of SYK and terminate signaling. Genetic deletion of CBL E3 ligases or charge-conserved mutation of target lysines within SYK abolishes SYK ubiquitination and results in enhanced DNGR-1-dependent antigen cross-presentation. We also find that cDC1 deficient in CBL E3 ligases are more efficient at cross-priming CD8 + T cells to dead cell-associated antigens and promoting host resistance to tumors. Our findings reveal a role for CBL-dependent ubiquitination in limiting cross-presentation of dead cell-associated antigens and highlight an axis of negative regulation of cDC1 activity that could be exploited to increase anti-tumor immunity., Competing Interests: Declaration of interests C.R.S. is a founder of Adendra Therapeutics and owns stock options and/or is a paid consultant for Adendra Therapeutics, Bicara Therapeutics, Montis Biosciences, and Bicycle Therapeutics, all unrelated to this work. C.R.S. also has an additional appointment as Visiting Professor in the Faculty of Medicine at Imperial College London and holds honorary professorships at University College London and King’s College London. H.B. received a research grant from Nimbus Therapeutics for unrelated work., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

There is a dearth of research on listening in the context of political conversation. Yet there is theoretical reason to believe that political listening could be an important pathway toward several democratically important outcomes including increased exposure to difference, mutual understanding, and decreased polarization. Unfortunately, listening in political contexts associated with deeply held moral beliefs and strong social identities may be among the most difficult settings for listening to flourish. On the other hand, listening is reciprocal within dyads and so a foothold of listening could, through subsequent social contagion, have potentially widespread effects. This article will review theory and research on political listening, as well as relevant scholarship on listening outside of the political domain., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Conventional dendritic cells type 1 (cDC1) are critical for inducing protective CD8 + T cell responses to tumour and viral antigens. In many instances, cDC1 access those antigens in the form of material internalised from dying tumour or virally-infected cells. How cDC1 extract dead cell-associated antigens and cross-present them in the form of peptides bound to MHC class I molecules to CD8 + T cells remains unclear. Here we review the biology of dendritic cell natural killer group receptor-1 (DNGR-1; also known as CLEC9A), a C-type lectin receptor highly expressed on cDC1 that plays a key role in this process. We highlight recent advances that support a function for DNGR-1 signalling in promoting inducible rupture of phagocytic or endocytic compartments containing dead cell debris, thereby making dead cell-associated antigens accessible to the endogenous MHC class I processing and presentation machinery of cDC1. We further review how DNGR-1 detects dead cells, as well as the functions of the receptor in anti-viral and anti-tumour immunity. Finally, we highlight how the study of DNGR-1 has opened new perspectives into cross-presentation, some of which may have applications in immunotherapy of cancer and vaccination against viral diseases., Competing Interests: Declaration of Competing Interest C.R.S. is a founder of Adendra Therapeutics and owns stock options and/or is a paid consultant for Adendra Therapeutics, Bicara Therapeutics, Montis Biosciences, Bicycle Therapeutics and Sosei Heptares. C.R.S. has an additional appointment as a Visiting Professor in the Faculty of Medicine at Imperial College London and holds honorary professorships at University College London and King’s College London., (Copyright © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.)

The membranes of endosomes, phagosomes and macropinosomes can become damaged by the physical properties of internalized cargo, by active pathogenic invasion or by cellular processes, including endocytic maturation. Loss of membrane integrity is often deleterious and is, therefore, prevented by mitigation and repair mechanisms. However, it can occasionally be beneficial and actively induced by cells. Here, we summarize the mechanisms by which cells, in particular phagocytes, try to prevent membrane damage and how, when this fails, they repair or destroy damaged endocytic organelles. We also detail how one type of phagocyte, the dendritic cell, can deliberately trigger localized damage to endocytic organelles to allow for major histocompatibility complex class I presentation of exogenous antigens and initiation of CD8 + T-cell responses to viruses and tumours. Our review highlights mechanisms for the regulation of endocytic organelle membrane integrity at the intersection of cell biology and immunology that could be co-opted for improving vaccination and intracellular drug delivery.

Spontaneous renal artery dissection is a rare condition with an often non-specific presentation, resulting in a challenging diagnosis for clinicians. This is the case of a 39-year-old man who presented with an acute-onset right flank pain, mild neutrophilia and sterile urine. CT of abdomen and pelvis showed a patchy hypodense area in the right kidney originally thought to represent infection. He was treated as an atypical pyelonephritis with antibiotics and fluids. When his symptoms failed to improve, a diagnosis of renal infarction was considered and CT angiogram of the aorta revealed a spontaneous renal artery dissection. He was managed conservatively with systemic anticoagulation, antihypertensive treatment and analgesia and discharged home with resolution of his symptoms and normal renal function., Competing Interests: Competing interests: None declared., (© BMJ Publishing Group Limited 2021. No commercial re-use. See rights and permissions. Published by BMJ.)

Type 1 conventional dendritic (cDC1) cells are necessary for cross-presentation of many viral and tumor antigens to CD8 + T cells. cDC1 cells can be identified in mice and humans by high expression of DNGR-1 (also known as CLEC9A), a receptor that binds dead-cell debris and facilitates XP of corpse-associated antigens. Here, we show that DNGR-1 is a dedicated XP receptor that signals upon ligand engagement to promote phagosomal rupture. This allows escape of phagosomal contents into the cytosol, where they access the endogenous major histocompatibility complex class I antigen processing pathway. The activity of DNGR-1 maps to its signaling domain, which activates SYK and NADPH oxidase to cause phagosomal damage even when spliced into a heterologous receptor and expressed in heterologous cells. Our data reveal the existence of innate immune receptors that couple ligand binding to endocytic vesicle damage to permit MHC class I antigen presentation of exogenous antigens and to regulate adaptive immunity.

Inflammation triggered by infection or cellular necrosis is initiated by a battery of pattern-recognition receptors, such as Toll-like receptors or IL-1 family receptors. Diverse forms of cell stress, such as ER stress or mitochondrial stress, can also promote inflammatory responses that contribute to the chronic inflammation observed in cancer, obesity, and other conditions. However, the molecular mechanisms of cell-stress-induced inflammation are poorly understood. Here, we show that ER stress initiated NF-κB activation and inflammation through transcriptional upregulation and ligand-independent activation of TRAIL receptors. ER-stress-induced TRAIL receptor activation resulted in caspase-8/FADD/RIPK1-dependent NF-κB activation and inflammatory cytokine production. Silencing or deletion of TRAIL receptors, or their downstream effectors caspase-8, FADD, or RIPK1, suppressed ER-stress-induced inflammation. Furthermore, chemotherapeutic stress-induced inflammatory responses were blunted in DR5/TRAIL-R null animals. We propose that, upon ER stress, TRAIL receptors serve as "stress-associated molecular patterns (SAMPs)" coupling ER stress to NF-κB-dependent inflammation., Competing Interests: Declaration of Interests The authors declare no competing interests., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Damage-associated molecular patterns (DAMPs) are molecules exposed or released by dead cells that trigger or modulate immunity and tissue repair. In vertebrates, the cytoskeletal component F-actin is a DAMP specifically recognised by DNGR-1, an innate immune receptor. Previously we suggested that actin is also a DAMP in Drosophila melanogaster by inducing STAT-dependent genes (Srinivasan et al., 2016). Here, we revise that conclusion and report that α-actinin is far more potent than actin at inducing the same STAT response and can be found in trace amounts in actin preparations. Recombinant expression of actin or α-actinin in bacteria demonstrated that only α-actinin could drive the expression of STAT target genes in Drosophila . The response to injected α-actinin required the same signalling cascade that we had identified in our previous work using actin preparations. Taken together, these data indicate that α-actinin rather than actin drives STAT activation when injected into Drosophila ., Competing Interests: OG, CH, NS, SA, AF, SD, PC, DP, RG, SK, DF, AS, RV, CS, LT, BT, MD, WW, CR No competing interests declared, (© 2018, Gordon et al.)

IL-1 family cytokines act as apical initiators of inflammation in many settings and can promote the production of a battery of inflammatory cytokines, chemokines and other inflammatory mediators in diverse cell types. IL-36α, IL-36β and IL-36γ, which belong to the extended IL-1 family, have been implicated as key initiators of skin inflammation in psoriasis. IL-36γ is highly upregulated in lesional skin from psoriatic individuals, and heritable mutations in the natural IL-36 receptor antagonist result in a severe form of psoriasis. IL-36 family cytokines are initially expressed as inactive precursors that require proteolytic processing for activation. The neutrophil granule-derived protease elastase proteolytically processes and activates IL-36α and IL-36γ, increasing their biological activity ~ 500-fold, and also robustly activates IL-1α and IL-33 through limited proteolytic processing. Consequently, inhibitors of elastase activity may have potential as anti-inflammatory agents through antagonizing the activation of multiple IL-1 family cytokines. Using in silico screening approaches, we have identified small-molecule inhibitors of elastase that can antagonize activation of IL-36γ by the latter protease. The compounds reported herein may have utility as lead compounds for the development of inhibitors of elastase-mediated activation of IL-36 and other IL-1 family cytokines in inflammatory conditions, such as psoriasis.

Neutrophil granule proteases are thought to function as anti-microbial effectors, cooperatively hydrolyzing microorganisms within phagosomes, or upon deployment into the extracellular space. However, evidence also suggests that neutrophil proteases play an important role in the coordination and escalation of inflammatory reactions, but how this is achieved has been obscure. IL-1 family cytokines are important initiators of inflammation and are typically released via necrosis but require proteolytic processing for activation. Here, we show that proteases liberated from activated neutrophils can positively or negatively regulate the activity of six IL-1 family cytokines (IL-1α, IL-1β, IL-33, IL-36α, IL-36β, and IL-36γ) with exquisite sensitivity. In contrast, extracellular neutrophil proteases displayed very poor bactericidal activity, exhibiting 100-fold greater potency toward cytokine processing than bacterial killing. Thus, in addition to their classical role as phagocytes, neutrophils play an important immunoregulatory role through deployment of their granule proteases into the extracellular space to process multiple IL-1 family cytokines., (Copyright © 2018 The Author(s). Published by Elsevier Inc. All rights reserved.)

Sterile inflammation is initiated by molecules released from necrotic cells, called damage-associated molecular patterns (DAMPs). Members of the extended IL-1 cytokine family are important DAMPs, are typically only released through necrosis, and require limited proteolytic processing for activation. The IL-1 family cytokines, IL-36α, IL-36β, and IL-36γ, are expressed as inactive precursors and have been implicated as key initiators of psoriatic-type skin inflammation. We have recently found that IL-36 family cytokines are proteolytically processed and activated by the neutrophil granule-derived proteases, elastase, and cathepsin G. Inhibitors of IL-36 processing may therefore have utility as anti-inflammatory agents through suppressing activation of the latter cytokines. We have identified peptide-based pseudosubstrates for cathepsin G and elastase, based on optimal substrate cleavage motifs, which can antagonize activation of all three IL-36 family cytokines by the latter proteases. Human psoriatic skin plaques displayed elevated IL-36β processing activity that could be antagonized by peptide pseudosubstrates specific for cathepsin G. Thus, antagonists of neutrophil-derived proteases may have therapeutic potential for blocking activation of IL-36 family cytokines in inflammatory conditions such as psoriasis.

Activated neutrophils can undergo a mode of regulated cell death, called NETosis, that results in the extrusion of chromatin into the extracellular space, thereby acting as extracellular traps for microorganisms. Neutrophil-derived extracellular traps (NETs) are comprised of DNA decorated with histones, antimicrobial proteins and neutrophil granule proteases, such as elastase and cathepsin G (Cat G). NET-associated factors are thought to enhance the antimicrobial properties of these structures and localisation of antimicrobial molecules on NETs may serve to increase their local concentration. Because neutrophil-derived proteases have been implicated in the processing and activation of several members of the extended interleukin (IL)-1 family, we wondered whether neutrophil NETs could also serve as platforms for the activation of proinflammatory cytokines. Here, we show that neutrophil NETs potently processed and activated IL-1α as well as IL-36 subfamily cytokines through NET-associated Cat G and elastase. Thus, in addition to their role as antimicrobial traps, NETs can also act as local sites of cytokine processing and activation., (© 2017 Federation of European Biochemical Societies.)

Conjoined twins occur in up to 1 in 50,000 live births with approximately 18% joined in a pygopagus configuration at the buttocks. Twins with this configuration display symptoms and carry surgical risks during separation related to the extent of their connection which can include anorectal, genitourinary, vertebral, and neural structures. Neurophysiologic intraoperative monitoring for these cases has been discussed in the literature with variable utility. The authors present a case of pygopagus twins with fused spinal cords and imperforate anus where the use of neurophysiologic intraoperative monitoring significantly impacted surgical decision-making in division of these critical structures.

TRAIL is a potent inducer of apoptosis and has been studied almost exclusively in this context. However, TRAIL can also induce NFκB-dependent expression of multiple pro-inflammatory cytokines and chemokines. Surprisingly, whereas inhibition of caspase activity blocked TRAIL-induced apoptosis, but not cytokine production, knock down or deletion of caspase-8 suppressed both outcomes, suggesting that caspase-8 participates in TRAIL-induced inflammatory signaling in a scaffold role. Consistent with this, introduction of a catalytically inactive caspase-8 mutant into CASP-8 null cells restored TRAIL-induced cytokine production, but not cell death. Furthermore, affinity precipitation of the native TRAIL receptor complex revealed that pro-caspase-8 was required for recruitment of RIPK1, via FADD, to promote NFκB activation and pro-inflammatory cytokine production downstream. Thus, caspase-8 can serve in two distinct roles in response to TRAIL receptor engagement, as a scaffold for assembly of a Caspase-8-FADD-RIPK1 "FADDosome" complex, leading to NFκB-dependent inflammation, or as a protease that promotes apoptosis., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Recent evidence has strongly implicated IL-36 cytokines as key initiators of inflammation in the skin barrier. IL-36 cytokines belong to the extended IL-1 family and, similar to most members of this family, are expressed as inactive precursors that require proteolytic processing for activation. Because the proteases responsible for activation of members of the IL-36 subfamily have not been reported, we have developed a method for the production of biologically active IL-36 through introduction of a caspase cleavage motif, DEVD, within the N-termini of these cytokines. Here, we show that DEVD-modified IL-36α, IL-36β and IL-36γ cytokines were highly soluble and were readily processed and activated by caspase-3. Caspase-3-processed IL-36 family cytokines exhibited robust biological activity on a range of responsive cell types, including primary keratinocytes. We also generated specific polyclonal antibodies against all three IL-36 family members through immunization with purified recombinant IL-36 cytokines. The modified forms of IL-36 described herein will be useful for production of large quantities of biologically active IL-36 for structure and function studies on these important proinflammatory cytokines.

Recent evidence has strongly implicated the IL-1 family cytokines IL-36α, IL-36β, and IL-36γ as key initiators of skin inflammation. Similar to the other members of the IL-1 family, IL-36 cytokines are expressed as inactive precursors and require proteolytic processing for activation; however, the responsible proteases are unknown. Here, we show that IL-36α, IL-36β, and IL-36γ are activated differentially by the neutrophil granule-derived proteases cathepsin G, elastase, and proteinase-3, increasing their biological activity ~500-fold. Active IL-36 promoted a strong pro-inflammatory signature in primary keratinocytes and was sufficient to perturb skin differentiation in a reconstituted 3D human skin model, producing features resembling psoriasis. Furthermore, skin eluates from psoriasis patients displayed significantly elevated cathepsin G-like activity that was sufficient to activate IL-36β. These data identify neutrophil granule proteases as potent IL-36-activating enzymes, adding to our understanding of how neutrophils escalate inflammatory reactions. Inhibition of neutrophil-derived proteases may therefore have therapeutic benefits in psoriasis., (Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.)

TNF promotes a regulated form of necrosis, called necroptosis, upon inhibition of caspase activity in cells expressing RIPK3. Because necrosis is generally more pro-inflammatory than apoptosis, it is widely presumed that TNF-induced necroptosis may be detrimental in vivo due to excessive inflammation. However, because TNF is intrinsically highly pro-inflammatory, due to its ability to trigger the production of multiple cytokines and chemokines, rapid cell death via necroptosis may blunt rather than enhance TNF-induced inflammation. Here we show that TNF-induced necroptosis potently suppressed the production of multiple TNF-induced pro-inflammatory factors due to RIPK3-dependent cell death. Similarly, necroptosis also suppressed LPS-induced pro-inflammatory cytokine production. Consistent with these observations, supernatants from TNF-stimulated cells were more pro-inflammatory than those from TNF-induced necroptotic cells in vivo. Thus necroptosis attenuates TNF- and LPS-driven inflammation, which may benefit intracellular pathogens that evoke this mode of cell death by suppressing host immune responses.

Unlabelled: Transcranial motor stimulation with noninvasive cortical surface stimulation, using a high-intensity magnetic field referred to as transcranial magnetic stimulation generally, is considered a nonpainful technique. In contrast, transcranial electric stimulation of the motor tracts typically cannot be done in unanesthesized patients. Intraoperative monitoring of motor tract function with transcranial electric stimulation is considered a standard practice in many institutions for patients during surgical procedures in which there is potential risk of motor tract impairment so that the risk of paraplegia or paraparesis can be reduced. Because transcranial electric stimulation cannot be typically done in the outpatient setting, transcranial magnetic stimulation may be able to provide a well-tolerated method for evaluation of the corticospinal motor tracts before surgery., Methods: One hundred fifty-five patients aged 5 to 20 years were evaluated preoperatively with single-stimulation nonrepetitive transcranial magnetic stimulation for preoperative assessment., Results and Conclusions: The presence of responses to transcranial magnetic stimulation reliably predicted the presence of responses to transcranial electric stimulation intraoperatively. No complications occurred during the testing, and findings were correlated to the clinical history and used in the setup of the surgical monitoring.

Apoptosis is a programmed mode of cell death that is accompanied by numerous morphological as well as biochemical changes to the cellular architecture. This results not only in cell death but also in the efficient removal of apoptotic cells by phagocytes. Apoptotic cells display a range of common characteristics that include cell shrinkage, plasma membrane blebbing, cell detachment, nuclear condensation, DNA fragmentation, externalization of phosphatidylserine (PS) and activation of caspases. In contrast, necrotic cell death is characterised by rapid plasma membrane, organelle swelling and plasma membrane rupture with none of the features of apoptosis. Apart from severe physical stresses, necrotic cell death often betrays the activities of viral infection and the activities of bacterial toxins. While necrotic cell death is characterized by the release of endogenous 'danger signals' and subsequent inflammation, apoptosis is largely tolergenic. Therefore, care must be taken when assessing whether cells are dying via apoptosis or necrosis. Here, we highlight a number of assays, utilizing both microscopy and flow cytometry, to determine whether cells have undergone apoptosis or alternative modes of cell death., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Apoptosis is commonly thought to represent an immunologically silent or even anti-inflammatory mode of cell death, resulting in cell clearance in the absence of explicit activation of the immune system. However, here we show that Fas/CD95-induced apoptosis is associated with the production of an array of cytokines and chemokines, including IL-6, IL-8, CXCL1, MCP-1, and GMCSF. Fas-induced production of MCP-1 and IL-8 promoted chemotaxis of phagocytes toward apoptotic cells, suggesting that these factors serve as "find-me" signals in this context. We also show that RIPK1 and IAPs are required for optimal production of cytokines and chemokines in response to Fas receptor stimulation. Consequently, a synthetic IAP antagonist potently suppressed Fas-dependent expression of multiple proinflammatory mediators and inhibited Fas-induced chemotaxis. Thus, in addition to provoking apoptosis, Fas receptor stimulation can trigger the secretion of chemotactic factors and other immunologically active proteins that can influence immune responsiveness toward dying cells., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Members of the caspase family of cysteine proteases coordinate the morphological and biochemical events that typify apoptosis. However, neutralization of caspase activity in mammals fails to block death in response to most proapoptotic stimuli. This is because many cell death triggers provoke mitochondrial dysfunction upstream of caspase activation as a consequence of BAX/BAK channel opening. Although genetic or pharmacological inactivation of caspases fails to block cell death in most instances, it does convert the phenotype from apoptosis to necrosis. This has important implications for how the immune system responds to such cells, as necrotic cells provoke inflammation whereas apoptotic cells typically do not. Here, we propose an alternative perspective on apoptosis-associated caspase function by suggesting that these proteases are activated, not to kill, but to extinguish the proinflammatory properties of dying cells. This perspective unifies the mammalian caspase family as either positive or negative regulators of inflammation., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Much debate surrounds how prosurvival members of the BCL-2 family repress opening of the BAX/BAK channel to block apoptosis; in this issue Llambi et al. (2011) identify two modes of apoptosis inhibition that exhibit surprisingly different behavior upon repeat proapoptotic challenges by BH3-only proteins., (Copyright © 2011 Elsevier Inc. All rights reserved.)

The potential for using plant pathogens and seeds as indicator organisms for assessing sanitization of plant wastes during composting was tested in bench-scale flask and large-scale systems. Plasmodiophora brassicae was unsuitable due to high temperature tolerance in dry to moist composts, and detection of viable inoculum post-composting using bioassay plants not corresponding with that using TaqMan® PCR, possibly due to preservation of nucleic acids at elevated temperatures. Several other plant pathogens (Sclerotinia sclerotiorum, Microdochium nivale, Phytophthora cinnamomi and Phytophthora nicotianae) were unsuitable due their low temperature tolerance. Fusarium oxysporum f.sp. cepae and f.sp. radicis-lycopersici chlamydospores and tomato seeds were suitable indicators due to their moderate temperature tolerance and ease of viability testing post-composting. Abutilon seeds were more tolerant than tomato seeds of compost temperatures ≥52°C but more prone to degradation at lower temperatures and therefore less suitable as indicators. Relationships between compost temperature during exposures of 2-10 days and subsequent viability of the above chlamydospores or seeds enabled the sanitizing effect of composting processes to be predicted within 2-6 days. Plant waste type (woody or vegetable) had a small but significant effect on the relationship for tomato seeds but not for F. oxysporum chlamydospores., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

The rise in international trade of plants and plant products has increased the risk of introduction and spread of plant pathogens and pests. In addition, new risks are arising from the implementation of more environmentally friendly methods of biodegradable waste disposal, such as composting and anaerobic digestion. As these disposal methods do not involve sterilisation, there is good evidence that certain plant pathogens and pests can survive these processes. The temperature/time profile of the disposal process is the most significant and easily defined factor in controlling plant pathogens and pests. In this review, the current evidence for temperature/time effects on plant pathogens and pests is summarised. The advantages and disadvantages of direct and indirect process validation for the verification of composting processes, to determine their efficacy in destroying plant pathogens and pests in biowaste, are discussed. The availability of detection technology and its appropriateness for assessing the survival of quarantine organisms is also reviewed.

Alcohol-induced zinc (Zn) deficiency is one of the mechanisms proposed as a cause of ethanol teratogenicity. Subcutaneous Zn treatment with ethanol in early pregnancy has been shown to prevent birth abnormalities and memory impairments in mice. This study examined whether dietary Zn supplementation throughout pregnancy can prevent cognitive impairments caused by early ethanol exposure. Pregnant C57BL/6J mice were fed either a control (35 microg Zn/g) or Zn-supplemented (200 microg Zn/g) diet throughout pregnancy. On gestational day (GD) 8, mice received two intraperitoneal injections (4h apart) of either saline or 25% ethanol (0.015 mL/g). All offspring were screened for physical and behavioural defects (e.g. growth, visual, exploratory, anxiety, motor deficits). Twenty-four phenotypically-normal offspring were randomly selected from each of the four treatment groups (saline +/- Zn-supplementation, ethanol +/- Zn-supplementation) and tested at 60 d of age using a cross-maze escape task for spatial learning and memory impairments, and an object recognition task. While no differences were observed between treatments for spatial learning, offspring exposed to ethanol demonstrated spatial memory impairments at both 12 and 28 d after learning an escape task, with less correct trials and increased escape latency scores compared with saline-treated mice. Furthermore, these mice also exhibited impairments in object recognition memory. In comparison, ethanol-exposed offspring from dams fed a Zn-supplemented diet throughout pregnancy did not display spatial memory or object recognition deficits, performing at the same level as saline-treated offspring. Therefore, dietary Zn-supplementation during pregnancy prevents spatial and object recognition memory impairments caused by ethanol exposure during early pregnancy.

A multiplex reverse-transcription polymerase chain assay (mRT-PCR) was developed, based on primers designed to distinguish the A and B types of beet necrotic yellow vein virus (BNYVV). RNA was extracted from 72 BNYVV isolates from Asia, Europe and North America, and the type of each isolate determined using an established detection method based on single strand conformation polymorphisms (SSCPs). An area of the 'triple gene block' region on RNA 2 was amplified and sequenced from 16 isolates of the A and B types. These sequences were aligned and two sets of PCR primers were designed to amplify unique areas common to each type. The A type assay produced a single 324 base-pair RT-PCR fragment when positive samples were amplified. The B type assay produced a 178 base-pair product from positive samples. No amplification was observed from healthy Chenopodium quinoa or sugar beet plants and from plants infected by others sugar beet soil-borne viruses. Fragment length differed sufficiently to allow both assays to be run in a single PCR tube. The results obtained using the new multiplex RT-PCR assay were consistent with those from the established SSCP method for all 72 reference samples.

Real-time RT-PCR (TaqMan) assays were developed for the specific detection of Beet necrotic yellow vein virus (BNYVV). The two assays designed were a broad-spectrum one that detected RNA 2 from all types and a second designed to detect types containing RNA 5. The assays were validated against a range of different isolates from Europe and the Far East. These real-time assays were compared to a conventional RT-PCR assay for the detection of RNA 5. Sensitivity comparisons showed that for the detection of RNA 5, TaqMan was 10,000 times more sensitive than the conventional RT-PCR assay. Further improvements were made to the test procedure by using post-ELISA virus release (VR), as an alternative to RNA extraction. This significantly increased the speed of processing samples and reduced the staff input required, allowing the TaqMan assay to be used routinely as part of an annual survey of UK field samples.

A diagnostic test incorporating reverse-transcription polymerase chain reaction (RT-PCR) and nested polymerase chain reaction (nPCR) was developed for the detection of Beet necrotic yellow vein virus (BNYVV). The RT-PCR used the primers designed by (Henry et al., J. Virol. Methods 54 (1995)15) but refinements were made to the protocol including simplification of the extraction method, the use of standard reagents and adoption of a one-step procedure. None of these changes impaired sensitivity or specificity. The RT-PCR could also be used to amplify immunocaptured virus but this was slightly less sensitive than amplification from purified RNA. In nPCR, a second round of amplification was performed using primers, which produce a specific 326 base-pair product. Both RT-PCR and nPCR detected a range of 21 isolates collected from Europe, America and Asia (including A, B and P pathotypes) isolated from either sugar beet or Chenopodium quinoa. Neither assay produced PCR products using total RNA extracted from the roots of healthy sugar beet or beet infected with Beet soil-borne virus. However, the sensitivity of the nPCR was 1000 times greater than the standard RT-PCR. The reliability of the standard RT-PCR and nPCR was demonstrated using a range of cultivars collected from an infected field site. The use of the nPCR assay is recommended for applications where its improved sensitivity over standard RT-PCR is necessary, for example in the early detection of infection from bait-test soils and for quarantine and breeding purposes.

Infrared brings to mind spectra with narrow, well-separated peaks; anybody expecting something similar from near-IR needs to think again.

In April 1999, severe soilborne wheat mosaic virus (SBWMV) symptoms were observed in five fields of winter wheat (Triticum aestivum, cvs. Consort, Equinox, and Savannah) on one farm in Wiltshire, UK. Affected plants were markedly stunted and had a pale mosaic on their leaf sheaths that developed into bright yellow, parallel streaks on the leaves as they unfolded. Symptomatic plants were found in discrete, elliptical patches ranging in size from a few square meters to nearly a hectare. During May and June, symptoms became less marked as temperatures increased and were restricted to lower leaves. SBWMV was positively identified in all five fields (60 to 170 plants per field) by double (W. Huth, BBA-Braunschweig, Germany; Sanofi Phyto-Diagnostics, Paris) and triple (T. Wilson, SCRI, Dundee, UK) antibody sandwich enzyme-linked immunosorbent assay and by reversetranscription polymerase chain reaction (2). Identification was confirmed by immunoelectron microscopy, including protein-A gold labeling, which revealed bipartite, rod-shaped particles typical of SBWMV. Neither wheat spindle streak mosaic virus nor barley yellow dwarf virus was detected in the field samples, nor was SBWMV detected in any other field subsequently sampled, despite a survey of the surrounding area. Wheat is the most important economic crop in the United Kingdom (≈1.9 million hectares are grown annually, yielding ≈16 million tonnes), but its position is threatened by the economic impact of SBWMV, which has decreased yields by up to 50% in the United States (1). References: (1) T. A. Kucharek and J. H. Walker. Plant Dis. Rep. 58:763, 1974. (2) R. E. Pennington et al. Plant Dis. 77:1202, 1993.

Gene expression analysis promises to change toxicology.

GC/MS systems are on the bench but still in the game.

Miniaturization is on track to take MS into space and the doctor's office.

Orthogonal acceleration brings the advantages of time of flight to electrospray.

Oat chlorotic stunt virus (OCSV) is a 35 nm icosahedral plant virus comprising of a single capsid protein with a Mr of 48.2 kDa and a 4.1 kb single stranded, positive sense genomic RNA. Northern blot analysis detected a single 3' terminal subgenomic RNA in extracts from infected plants, which was also found to be encapsidated. Virion RNA directs the synthesis of a 23 kDa polypeptide in a rabbit reticulocyte in vitro translation system. Primer extension analysis has been used to map the end of both the genomic and subgenomic RNA's, and has shown the genomic size to be 4115 nucleotides in length. The results have enabled a model for the genome expression to be proposed.

The complete genomic sequence of a new virus, first found infecting oats in Wales, UK, has been determined. The genome is a positive-sense ssRNA molecule, 4114 nucleotides in length, examination of which indicates the presence of four ORFs. The first ORF initiating at the 5' terminus (ORF1) encodes a protein with a predicted M(r) of 23476 (p23). ORF2 extends through the amber termination codon of ORF1 to give a protein with a predicted M(r) of 84355 (p84). The readthrough domain of p84 contains amino acid sequence similarities with a number of putative RNA-dependent RNA polymerases. ORF3 is in a different reading frame from ORF1/2 and encodes a protein with an M(r) of 48231 (p48), identified as the coat protein by direct peptide sequencing. ORF4 nests within ORF3 but is in a different frame from it and codes for a protein with a predicted M(r) of 8220 (p8). Comparisons of peptide sequence, particularly within the putative polymerase region and within the S domain of the coat protein, highlight similarities with members of both the tombusvirus and carmovirus groups. The coat protein region shows most similarity with members of the tombusvirus group, whilst the size and predicted strategy of the genome seem to be intermediate between that of the carmovirus and tombusvirus groups. These features highlight possible evolutionary links with each group whilst being distinct from both. We propose the name of oat chlorotic stunt for this new virus.

A diagnostic test based on reverse transcription followed by the polymerase chain reaction (RT-PCR) was developed for the detection of beet necrotic yellow vein virus (BNYVV). A specific 500-base-pair fragment was amplified from the read-through region of the coat protein gene located on RNA-2. The viral origin of the amplified product was confirmed by sequencing, with the sequence obtained having 94.5% homology with published sequence data for BNYVV. The assay gave a sensitivity of 800 times that of a TAS-ELISA and 50 times that of an amplified TAS-ELISA method. A range of BNYVV isolates from the UK and worldwide could be detected by this test, either as mechanically inoculated Chenopodium quinoa leaves or infected sugar beet roots. Use of the assay in routine diagnostic tests allowed a reduction of time needed for the detection of Rhizomania in soils from 7 to 4 weeks.