Cavity surface residues of PAD4 and SAG101 contribute to EDS1 dimer signaling specificity in plant immunity.

Arabidopsis pathogen effector-triggered immunity (ETI) is controlled by a family of three lipase-like proteins (EDS1, PAD4, and SAG101) and two subfamilies of HET-S/LOB-B (HeLo)-domain "helper" nucleotide-binding/leucine-rich repeats (ADR1s and NRG1s). EDS1-PAD4 dimers cooperate with ADR1s, and EDS1-SAG101 dimers with NRG1s, in two separate defense-promoting modules. EDS1-PAD4-ADR1 and EDS1-SAG101-NRG1 complexes were detected in immune-activated leaf extracts but the molecular determinants for specific complex formation and function remain unknown. EDS1 signaling is mediated by a C-terminal EP domain (EPD) surface surrounding a cavity formed by the heterodimer. Here we investigated whether the EPDs of PAD4 and SAG101 contribute to EDS1 dimer functions. Using a structure-guided approach, we undertook a comprehensive mutational analysis of Arabidopsis PAD4. We identify two conserved residues (Arg314 and Lys380) lining the PAD4 EPD cavity that are essential for EDS1-PAD4-mediated pathogen resistance, but are dispensable for the PAD4-mediated restriction of green peach aphid infestation. Positionally equivalent Met304 and Arg373 at the SAG101 EPD cavity are required for EDS1-SAG101 promotion of ETI-related cell death. In a PAD4 and SAG101 interactome analysis of ETI-activated tissues, PAD4R314A and SAG101M304R EPD variants maintain interaction with EDS1 but lose association, respectively, with helper nucleotide-binding/leucine-rich repeats ADR1-L1 and NRG1.1, and other immune-related proteins. Our data reveal a fundamental contribution of similar but non-identical PAD4 and SAG101 EPD surfaces to specific EDS1 dimer protein interactions and pathogen immunity.

Adaptation of the heart to Frataxin depletion: Evidence that integrated stress response can predominate over mTORC1 activation.

Friedreich's ataxia (FRDA) is an inherited disorder caused by depletion of frataxin (FXN), a mitochondrial protein required for iron-sulfur cluster (ISC) biogenesis. Cardiac dysfunction is the main cause of death. Yet pathogenesis, and, more generally, how the heart adapts to FXN loss, remain poorly understood, though are expected to be linked to an energy deficit. We modified a transgenic (TG) mouse model of inducible FXN depletion that permits phenotypic evaluation of the heart at different FXN levels, and focused on substrate-specific bioenergetics and stress signaling. When FXN protein in the TG heart was 17% of normal, bioenergetics and signaling were not different from control. When, 8 weeks later, FXN was ~ 97% depleted in the heart, TG heart mass and cardiomyocyte cross-sectional area were less, without evidence of fibrosis or apoptosis. mTORC1 signaling was activated, as was the integrated stress response, evidenced by greater phosphorylation of eIF2α relative to total eIF2α, and decreased protein translation. We interpret these results to suggest that, in TG hearts, an anabolic stimulus was constrained by eIF2α phosphorylation. Cardiac contractility was maintained in the 97%-FXN-depleted hearts, possibly contributed by an unexpected preservation of ß-oxidation, though pyruvate oxidation was lower. Bioenergetics alterations were matched by changes in the mitochondrial proteome, including a non-uniform decrease in abundance of ISC-containing proteins. Altogether, these findings suggest that the FXN depleted heart can suppress a major ATP demanding process such as protein translation, which, together with some preservation of ß-oxidation, could be adaptive, at least in the short term.

Salinity alleviates arsenic stress-induced oxidative damage via antioxidative defense and metabolic adjustment in the root of the halophyte Salvadora persica.

MAIN CONCLUSION: Arsenic tolerance in the halophyte Salvadora persica is achieved by enhancing antioxidative defense and modulations of various groups of metabolites like amino acids, organic acids, sugars, sugar alcohols, and phytohormones. Salvadora persica is a facultative halophyte that thrives under high saline and arid regions of the world. In present study, we examine root metabolic responses of S. persica exposed to individual effects of high salinity (750 mM NaCl), arsenic (600 µM As), and combined treatment of salinity and arsenic (250 mM NaCl + 600 µM As) to decipher its As and salinity resistance mechanism. Our results demonstrated that NaCl supplementation reduced the levels of reactive oxygen species (ROS) under As stress. The increased activities of antioxidant enzymes like superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPX), and glutathione reductase (GR) maintained appropriate levels of ROS [superoxide (O2•-) and hydrogen peroxide (H2O2)] under salinity and/or As stress. The metabolites like sugars, amino acids, polyphenols, and organic acids exhibited higher accumulations when salt was supplied with As. Furthermore, comparatively higher accumulations of glycine, glutamate, and cystine under combined stress of salt and As may indicate its role in glutathione and phytochelatins (PCs) synthesis in root. The levels of phytohormones such as salicylate, jasmonate, abscisic acid, and auxins were significantly increased under high As with and without salinity stress. The amino acid metabolism, glutathione metabolism, carbohydrate metabolism, tricarboxylic acid cycle (TCA cycle), phenylpropanoid biosynthesis, and phenylalanine metabolism are the most significantly altered metabolic pathways in response to NaCl and/or As stress. Our study decoded the important metabolites and metabolic pathways involved in As and/or salinity tolerance in root of the halophyte S. persica providing clues for development of salinity and As resistance crops.

Interplay between MYZUS PERSICAE-INDUCED LIPASE 1 and OPDA signaling in limiting green peach aphid infestation on Arabidopsis thaliana.

MYZUS PERSICAE-INDUCED LIPASE1 (MPL1) encodes a lipase in Arabidopsis thaliana that is required for limiting infestation by the green peach aphid (GPA; Myzus persicae), an important phloem sap-consuming insect pest. Previously, we demonstrated that MPL1 expression was up-regulated in response to GPA infestation, and GPA fecundity was higher on the mpl1 mutant, compared with the wild-type (WT), and lower on 35S:MPL1 plants that constitutively expressed MPL1 from the 35S promoter. Here, we show that the MPL1 promoter is active in the phloem and expression of the MPL1 coding sequence from the phloem-specific SUC2 promoter in mpl1 is sufficient to restore resistance to GPA. The GPA infestation-associated up-regulation of MPL1 requires CYCLOPHILIN 20-3 (CYP20-3), which encodes a 12-oxo-phytodienoic acid (OPDA)-binding protein that is involved in OPDA signaling, and is required for limiting GPA infestation. OPDA promotes MPL1 expression to limit GPA fecundity, a process that requires CYP20-3 function. These results along with our observation that constitutive expression of MPL1 from the 35S promoter restores resistance to GPA in the cyp20-3 mutant, and MPL1 acts in a feedback loop to limit OPDA levels in GPA-infested plants, suggest that an interplay between MPL1, OPDA, and CYP20-3 contributes to resistance to GPA.

Copper iodide nanoparticles (CuI NPs): an efficient catalyst for the synthesis of alkynyl esters.

An environmentally benign protocol for the synthesis of alkynyl esters, by the cross-coupling of diazoacetate with various substituted alkynes under neat reaction conditions, has been described. Copper iodide nanoparticles (CuI NPs) were found to promote the Sonogashira-type coupling to afford the corresponding alkynyl esters in good yields. The CuI nanoparticles were characterized by PXRD, FESEM, EDAX, and Raman techniques. The developed methodology has several advantages such as a broad substrate scope, less reaction time, atom economy, avoidance of an additive/base/solvent, and enhanced values of green chemistry. The catalyst was recycled up to threefold without the loss of its catalytic activity.

Bio-inspired polynorepinephrine based nanocoatings for reduced graphene oxide/gold nanoparticles composite for high-performance biosensing of Mycobacterium tuberculosis.

Polydopamine (PDA) has established itself as a promising grafting and coating material, particularly for functional group-deprived electrochemically active nanomaterials such as graphene, MXene, CNT, metal nanoparticles, and so on, and has proven its extensive applicability in the design and development of electrochemical biosensor devices. However, polynorepinephrine (PNE), a sister compound of PDA, having additional -OH groups and greater coating uniformity and biocompatibility, has never been studied in the field of biosensors. Herein, we investigated PNE as a coating material for reduced graphene oxide (RGO) and gold nanoparticles (Au) in order to build an electrochemical genosensor for Mycobacterium tuberculosis (MTB) detection. Biotin-Avidin chemistry was used to covalently immobilize probe DNA (ssDNA) specific to MTB to the nanocomposite surface on glassy carbon electrode (GCE) in order to construct biosensing electrodes. The formation of RGO/PNE and RGO/PNE/Au nanocomposite as well as the immobilization of ssDNA onto the bioelectrodes are both corroborated by UV-Visible, Raman, and XRD studies with FE-SEM and HR-TEM analysis. The electrochemical studies performed using cyclic voltammetry (CV) and linear sweep voltammetry (LSV) showed the significant enhancement in charge transfer kinetics of RGO/PNE/GCE and RGO/PNE/Au/GCE electrode compared to GO/GCE electrode. The biosensing investigations performed using ssDNA/avidin/RGO/PNE/Au/GCE bioelectrode showed high sensitivity (2.3 × 10-3 mA µM-1), low detection limit (0.1 × 10-7 µM), broad detection range (0.1 × 10-2 to 0.1 × 10-7 µM) with good selectivity and low response time (5 s) of the developed sensor. In comparison to the analogous RGO/PDA/Au material system, RGO/PNE/Au demonstrated increased enzyme loading, improved electrochemical responsiveness, and superior biosensing performance.

Ternary nanocomposite-based smart sensor: Reduced graphene oxide/polydopamine/alanine nanocomposite for simultaneous electrochemical detection of Cd2+, Pb2+, Fe2+, and Cu2+ ions.

Heavy metal ion (HMI) sensors are the most sought commercial devices for environmental monitoring and food analysis research due to serious health concerns associated with HMI overdosage. Herein, we developed an effective electrochemical sensor for simultaneous detection of four HMI (Cd2+, Pb2+, Fe2+, and Cu2+) using a ternary nanocomposite of reduced graphene oxide functionalized with polydopamine and alanine (ALA/pDA/rGO). Comprehensive spectroscopic and microscopic characterizations were performed to ensure the formation of the ternary nanocomposite. The developed nanocomposite on glassy carbon electrode (GCE) yields >2-fold higher current than GO/GCE electrode with excellent electrochemical stability and charge transfer rate. Using DPV, various chemical and electrochemical parameters, such as supporting electrolyte, buffer pH, metal deposition time, and potential, were optimized to achieve highly sensitive detection of targeted HMI. For Cd2+, Pb2+, Fe2+, and Cu2+ sensing devised sensor exhibited detection limits of 1.46, 2.86, 50.23, and 17.95 ppb and sensitivity of 0.0929, 0.0744, 0.0051, and 0.0394 µA/ppb, respectively, with <6% interference. The sensor worked similarly well for real water samples with HMI. This study demonstrates a novel strategy for concurrently detecting and quantifying multiple HMI in water and soil using a smart ternary nanocomposite-based electrochemical sensor, which can also detect HMI in food samples.

Recent applications of nanoparticles in organic transformations.

A variation in the size of metal nanoparticles leads to a difference in their properties. As the size of metal nanoparticles decreases, the surface area increases which leads to an increase in the reactivity of metal nanoparticles. Metals like Au, Ag, Pd, and Pt have interesting properties when used in nanometric dimensions. They function efficiently in significant industrial processes as electrocatalysts and photocatalysts in various organic reactions. Recently, the green biosynthesis of nanoparticles has attracted the attention of researchers. With environmental pollution rising over the past few decades, metal nanoparticle catalysts could be the key to subdue the toxic effects. Being versatile, they can be used to degrade pollutants, develop solar cells, convert toxic nitroaromatic compounds, significantly reduce CO2 emissions per unit of energy, and many more. Owing to their unique properties, nanoparticles have wide applications in biomedicine, for example, gold cages are promising agents for cancer diagnosis and therapy. Transition metal-oxide nanoparticles have been considered one of the best supercapacitor electrodes with high electrochemical performance. In this review, we have summarised fundamental concepts of metal nanoparticles over the last decade's main emphasis from 2010 to 2021. It focuses on the exceptional use of these nanocatalysts in various organic reactions. Additionally, we have also discussed the utility of these reactions and their crucial role in solving the problems of today. Through this article, we hope to provide the necessary framework needed to further advance the applications of metal nanoparticles as catalysts.

AlRab7 from Aeluropus lagopoides ameliorates ion toxicity in transgenic tobacco by regulating hormone signaling and reactive oxygen species homeostasis.

The plants endomembrane system of the cellular compartments with its complex membrane trafficking network facilitates transport of macromolecules. The endomembrane dynamics are essential for maintaining basic and specific cellular functions including adaptation to the extracellular environment. The plant vacuole serves as a reservoir for nutrients and toxic metabolites and performs detoxification processes to maintain cellular homeostasis. The overexpression of AlRab7, a vesicle trafficking gene from Aeluropus lagopoides, improved germination and growth and reduced ionic and oxidative stress in transgenics. Moreover, the root and shoot of transgenic tobacco showed differential accumulation of phytohormone ABA and IAA with different ionic stresses. The improved growth (root and shoot length) can be co-related with higher IAA accumulation with NaCl stress. The low Na+ /K+ ratio with different NaCl stress treatments indicates better ion homeostasis in transgenics. Furthermore, the increased stomatal density and higher number of open stomata on both leaf surfaces in transgenics during NaCl stress suggest better gaseous exchange/functioning of guard cells. The maintained or increased superoxide dismutase, catalase, ascorbate peroxidase, guaiacol peroxidase, and glutathione reductase antioxidative enzyme activities suggest that an extensive reactive oxygen species (ROS) scavenging system was triggered to detoxify cellular ROS, which remained at low levels in transgenics during the different stress treatments. Our results suggest that the AlRab7 transgenic tobacco ameliorates ionic stress by facilitating differential and selective ion transport at vacuolar membrane regulating hormone signaling, ROS homeostasis, stomatal development, and movement.

Convalescent plasma therapy for the treatment of patients with COVID-19: Assessment of methods available for antibody detection and their correlation with neutralising antibody levels.

INTRODUCTION: The lack of approved specific therapeutic agents to treat coronavirus disease (COVID-19) associated with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection has led to the rapid implementation of convalescent plasma therapy (CPT) trials in many countries, including the United Kingdom. Effective CPT is likely to require high titres of neutralising antibody (nAb) in convalescent donations. Understanding the relationship between functional neutralising antibodies and antibody levels to specific SARS-CoV-2 proteins in scalable assays will be crucial for the success of a large-scale collection. We assessed whether neutralising antibody titres correlated with reactivity in a range of enzyme-linked immunosorbent assays (ELISA) targeting the spike (S) protein, the main target for human immune response. METHODS: Blood samples were collected from 52 individuals with a previous laboratory-confirmed SARS-CoV-2 infection. These were assayed for SARS-CoV-2 nAbs by microneutralisation and pseudo-type assays and for antibodies by four different ELISAs. Receiver operating characteristic (ROC) analysis was used to further identify sensitivity and specificity of selected assays to identify samples containing high nAb levels. RESULTS: All samples contained SARS-CoV-2 antibodies, whereas neutralising antibody titres of greater than 1:20 were detected in 43 samples (83% of those tested) and >1:100 in 22 samples (42%). The best correlations were observed with EUROimmun immunoglobulin G (IgG) reactivity (Spearman Rho correlation coefficient 0.88; p < 0.001). Based on ROC analysis, EUROimmun would detect 60% of samples with titres of >1:100 with 100% specificity using a reactivity index of 9.1 (13/22). DISCUSSION: Robust associations between nAb titres and reactivity in several ELISA-based antibody tests demonstrate their possible utility for scaled-up production of convalescent plasma containing potentially therapeutic levels of anti-SARS-CoV-2 nAbs.

Antibodies to SARS-CoV-2 protect against re-infection during outbreaks in care homes, September and October 2020.

Two London care homes experienced a second COVID-19 outbreak, with 29/209 (13.9%) SARS-CoV-2 RT-PCR-positive cases (16/103 residents, 13/106 staff). In those with prior SARS-CoV-2 exposure, 1/88 (1.1%) individuals (antibody positive: 87; RT-PCR-positive: 1) became PCR-positive compared with 22/73 (30.1%) with confirmed seronegative status. After four months protection offered by prior infection against re-infection was 96.2% (95% confidence interval (CI): 72.7-99.5%) using risk ratios from comparison of proportions and 96.1% (95% CI: 78.8-99.3%) using a penalised logistic regression model.

Advancements in spontaneous microbial desalination technology for sustainable water purification and simultaneous power generation: A review.

Population growth and rapid urbanization have put a lot of pressure on the already scarce freshwater around the globe. The availability of freshwater is not only limited but it is non-uniform also. Available desalination technologies help mitigate water shortage; however, these techniques are energy-intensive and unsustainable. Desalination technologies utilizing renewable energy and bio-electrochemical systems have been developed to achieve limited sustainability. With technological advancements, microbial desalination cell (MDC) has been developed which is capable of desalination, wastewater treatment, and power generation simultaneously. This review critically examined the performance of various MDC techniques concerning their stimulus parameters including COD removal, total desalination rate, total dissolved solids reduction rate, Coulombic efficiency, and power density. Limitations of MDCs have also been incorporated in the review. Work on MDC coupled with other robust desalination techniques offering advantages such as better desalination and more water recovery e.g. osmotic-MDC etc. has been included. Researchers have tremendously worked on MDCs with different electro-catalysts. Few of these are not sustainable and costly. Authors have reviewed critically with belief that it will pave a way for the commercialization of this eco-friendly technology.

The Arabidopsis PAD4 Lipase-Like Domain Is Sufficient for Resistance to Green Peach Aphid.

Plants have evolved mechanisms to protect themselves against pathogenic microbes and insect pests. In Arabidopsis, the immune regulator PAD4 functions with its cognate partner EDS1 to limit pathogen growth. PAD4, independently of EDS1, reduces infestation by green peach aphid (GPA). How PAD4 regulates these defense outputs is unclear. By expressing the N-terminal PAD4 lipase-like domain (PAD4LLD) without its C-terminal EDS1-PAD4 (EP) domain, we interrogated PAD4 functions in plant defense. Here, we show that transgenic expression of PAD4LLD in Arabidopsis is sufficient for limiting GPA infestation but not for conferring basal and effector-triggered pathogen immunity. This suggests that the C-terminal PAD4 EP domain is necessary for EDS1-dependent immune functions but is dispensable for aphid resistance. Moreover, PAD4LLD is not sufficient to interact with EDS1, indicating the PAD4-EP domain is required for stable heterodimerization. These data provide molecular evidence that PAD4 has domain-specific functions.

Duration of infectiousness and correlation with RT-PCR cycle threshold values in cases of COVID-19, England, January to May 2020.

Severe acute respiratory syndrome coronavirus 2 viral load in the upper respiratory tract peaks around symptom onset and infectious virus persists for 10 days in mild-to-moderate coronavirus disease (n = 324 samples analysed). RT-PCR cycle threshold (Ct) values correlate strongly with cultivable virus. Probability of culturing virus declines to 8% in samples with Ct > 35 and to 6% 10 days after onset; it is similar in asymptomatic and symptomatic persons. Asymptomatic persons represent a source of transmissible virus.

Convalescent plasma treatment for SARS-CoV-2 infection: analysis of the first 436 donors in England, 22 April to 12 May 2020.

Serological reactivity was analysed in plasma from 436 individuals with a history of disease compatible with COVID-19, including 256 who had been laboratory-confirmed with SARS-CoV-2 infection. Over 99% of laboratory-confirmed cases developed a measurable antibody response (254/256) and 88% harboured neutralising antibodies (226/256). Antibody levels declined over 3 months following diagnosis, emphasising the importance of the timing of convalescent plasma collections. Binding antibody measurements can inform selection of convalescent plasma donors with high neutralising antibody levels.

Arabidopsis ACTIN-DEPOLYMERIZING FACTOR3 Is Required for Controlling Aphid Feeding from the Phloem.

The actin cytoskeleton network has an important role in plant cell growth, division, and stress response. Actin-depolymerizing factors (ADFs) are a group of actin-binding proteins that contribute to reorganization of the actin network. Here, we show that the Arabidopsis (Arabidopsis thaliana) ADF3 is required in the phloem for controlling infestation by Myzus persicae Sülzer, commonly known as the green peach aphid (GPA), which is an important phloem sap-consuming pest of more than fifty plant families. In agreement with a role for the actin-depolymerizing function of ADF3 in defense against the GPA, we show that resistance in adf3 was restored by overexpression of the related ADF4 and the actin cytoskeleton destabilizers, cytochalasin D and latrunculin B. Electrical monitoring of the GPA feeding behavior indicates that the GPA stylets found sieve elements faster when feeding on the adf3 mutant compared to the wild-type plant. In addition, once they found the sieve elements, the GPA fed for a more prolonged period from sieve elements of adf3 compared to the wild-type plant. The longer feeding period correlated with an increase in fecundity and population size of the GPA and a parallel reduction in callose deposition in the adf3 mutant. The adf3-conferred susceptibility to GPA was overcome by expression of the ADF3 coding sequence from the phloem-specific SUC2 promoter, thus confirming the importance of ADF3 function in the phloem. We further demonstrate that the ADF3-dependent defense mechanism is linked to the transcriptional up-regulation of PHYTOALEXIN-DEFICIENT4, which is an important regulator of defenses against the GPA.

Pseudoaneurysm of the Greater Palatine Vessel Following Subepithelial Connective Tissue Graft.

Subepithelial gingival connective tissue grafts are a common surgical procedure performed in periodontal and implant surgery. This versatile procedure has many indications including tooth root coverage, thickening of gingiva, and improvement of the quality of the crestal gingiva. Several techniques have been described for graft harvest from the palate. Reported complications from these techniques include pain, inflammation, bleeding, flap necrosis, and infection in the donor site. We report a previously unpublished complication following subepithelial gingival connective tissue graft from the palate: pseudoaneurysm of the greater palatine vessel.

Elicitors and defense gene induction in plants with altered lignin compositions.

A reduction in the lignin content in transgenic plants induces the ectopic expression of defense genes, but the importance of altered lignin composition in such phenomena remains unclear. Two Arabidopsis lines with similar lignin contents, but strikingly different lignin compositions, exhibited different quantitative and qualitative transcriptional responses. Plants with lignin composed primarily of guaiacyl units overexpressed genes responsive to oomycete and bacterial pathogen attack, whereas plants with lignin composed primarily of syringyl units expressed a far greater number of defense genes, including some associated with cis-jasmone-mediated responses to aphids; these plants exhibited altered responsiveness to bacterial and aphid inoculation. Several of the defense genes were differentially induced by water-soluble extracts from cell walls of plants of the two lines. Glycome profiling, fractionation and enzymatic digestion studies indicated that the different lignin compositions led to differential extractability of a range of heterogeneous oligosaccharide epitopes, with elicitor activity originating from different cell wall polymers. Alteration of lignin composition affects interactions with plant cell wall matrix polysaccharides to alter the sequestration of multiple latent defense signal molecules with an impact on biotic stress responses.

Base-Mediated Hydroamination of Alkynes.

Inter- or intramolecular hydroamination reactions are a paradigmatic example of modern sustainable organic chemistry, as they are a catalytic, 100% atom-economical, and waste-free process of fundamental simplicity in which an amine is added to an alkyne substrate. Many enamines are found in many natural and synthetic compounds possessing interesting physiological and biological activities. The development of synthetic protocols for such molecules and their transformation is a persistent research topic in pharmaceutical and organic chemistry. Hydroamination is conspicuously superior to the other accessible methods, such as the imination of ketones or the aminomercuration/demercuration of alkynes, that involve the stoichiometric use of toxic reagents. Additionally, the hydroamination of alkyne substrates has been successfully employed as a key step in synthesizing target molecules through total syntheses containing substituted indoles, pyrroles, imidazoles, and other heterocycles as core moieties. Many research groups have explored inter- or intramolecular hydroamination of alkynes for the synthesis of diversely substituted nitrogen heterocycles using expensive metal catalysts. However, in contrast to metal-catalyzed hydroamination, the base-mediated hydroamination of alkynes has not been extensively studied. Various inorganic (such as hydroxides, phosphates, and carbonates) and organic bases have been proven to be valuable reagents for achieving the hydroamination process. This method represents an attractive strategy for the construction of a broad range of nitrogen-containing compounds that prevents the formation of byproducts in the creation of a C-N linkage. The presence of a base is thought to facilitate the attack of nitrogen nucleophiles, such as indoles, pyrroles, and imidazoles, on unsaturated carbon substrates through the activation of the triple bond and thus transforming the electron-rich alkyne into an electrophile. In the past few years, we have been involved in the development of methods for the nucleophilic addition of N-heterocycles onto terminal and internal alkynes using alkali base catalysts to achieve new carbon-nitrogen bond-forming reactions. During our study, we discovered the regioselective preferential nucleophilic addition of N-heterocycles onto the haloarylalkyne over N-arylation of the aryl halide. In this Account, we summarize our latest achievements in regio-, stereo-, and chemoselective hydroamination chemistry of N-nucleophiles with alkynes using a superbasic medium to produce a broad range of highly functionalized vinyl and styryl enamines, which are valuable and versatile synthetic intermediates for the synthesis of bioactive compounds. Interestingly, the stereoselectivity of the addition products (kinetically stable Z and thermodynamically stable E isomers) was found to be dependent upon time. It is worthwhile to note that hydroaminated products formed by the addition reaction can further be utilized for the synthesis of indolo-/pyrrolo[2,1-a]isoquinolines, naphthyridines, and bisindolo/pyrrolo[2,1-a]isoquinolinesvia tandem cyclization. This chemistry is expected to find application in organic synthesis for constructing a diverse variety of fused π-conjugated compounds, enaminones, and C-C coupled products.

Late Ebola virus relapse causing meningoencephalitis: a case report.

BACKGROUND: There are thousands of survivors of the 2014 Ebola outbreak in west Africa. Ebola virus can persist in survivors for months in immune-privileged sites; however, viral relapse causing life-threatening and potentially transmissible disease has not been described. We report a case of late relapse in a patient who had been treated for severe Ebola virus disease with high viral load (peak cycle threshold value 13.2). METHODS: A 39-year-old female nurse from Scotland, who had assisted the humanitarian effort in Sierra Leone, had received intensive supportive treatment and experimental antiviral therapies, and had been discharged with undetectable Ebola virus RNA in peripheral blood. The patient was readmitted to hospital 9 months after discharge with symptoms of acute meningitis, and was found to have Ebola virus in cerebrospinal fluid (CSF). She was treated with supportive therapy and experimental antiviral drug GS-5734 (Gilead Sciences, San Francisco, Foster City, CA, USA). We monitored Ebola virus RNA in CSF and plasma, and sequenced the viral genome using an unbiased metagenomic approach. FINDINGS: On admission, reverse transcriptase PCR identified Ebola virus RNA at a higher level in CSF (cycle threshold value 23.7) than plasma (31.3); infectious virus was only recovered from CSF. The patient developed progressive meningoencephalitis with cranial neuropathies and radiculopathy. Clinical recovery was associated with addition of high-dose corticosteroids during GS-5734 treatment. CSF Ebola virus RNA slowly declined and was undetectable following 14 days of treatment with GS-5734. Sequencing of plasma and CSF viral genome revealed only two non-coding changes compared with the original infecting virus. INTERPRETATION: Our report shows that previously unanticipated, late, severe relapses of Ebola virus can occur, in this case in the CNS. This finding fundamentally redefines what is known about the natural history of Ebola virus infection. Vigilance should be maintained in the thousands of Ebola survivors for cases of relapsed infection. The potential for these cases to initiate new transmission chains is a serious public health concern. FUNDING: Royal Free London NHS Foundation Trust.