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A modular approach to underexplored, unsymmetrical [1]benzothieno[3,2-b][1]benzothiophene (BTBT) scaffolds delivers a library of BTBT materials from readily available coupling partners by combining a transition-metal free Pummerer CH–CH-type cross-coupling and a Newman–Kwart reaction. This effective approach to unsymmetrical BTBT materials has allowed their properties to be studied. In particular, tuning the functional groups on the BTBT scaffold allows the solid-state assembly and molecular orbital energy levels to be modulated. Investigation of the charge transport properties of BTBT-containing small-molecule:polymer blends revealed the importance of molecular ordering during phase segregation and matching the highest occupied molecular orbital energy level with that of the semiconducting polymer binder, polyindacenodithiophene-benzothiadiazole (PIDTBT). The hole mobilities extracted from transistors fabricated using blends of PIDTBT with phenyl or methoxy functionalized unsymmetrical BTBTs were double those measured for devices fabricated using pristine PIDTBT. This study underscores the value of the synthetic methodology in providing a platform from which to study structure–property relationships in an underrepresented family of unsymmetrical BTBT molecular semiconductors., A modular approach to underexplored, unsymmetrical [1]benzothieno[3,2-b][1]benzothiophene (BTBT) scaffolds, combining a transition-metal free Pummerer CH–CH-type cross-coupling and a Newman–Kwart reaction, delivers a library of BTBT materials.

Turpentine wood is renowned for its resistance to attack by molluskan marine borers. This resistance is thought to be due to its high silica content, and possibly the presence of other, as yet unkn...

RAS mutations are the most common oncogenic drivers across human cancers, but there remains a paucity of clinically-validated pharmacological inhibitors of RAS, as druggable pockets have proven difficult to identify. Here, we identify two RAS-binding Affimer proteins, K3 and K6, that inhibit nucleotide exchange and downstream signaling pathways with distinct isoform and mutant profiles. Affimer K6 binds in the SI/SII pocket, whilst Affimer K3 is a non-covalent inhibitor of the SII region that reveals a conformer of wild-type RAS with a large, druggable SII/α3 pocket. Competitive NanoBRET between the RAS-binding Affimers and known RAS binding small-molecules demonstrates the potential to use Affimers as tools to identify pharmacophores. This work highlights the potential of using biologics with small interface surfaces to select unseen, druggable conformations in conjunction with pharmacophore identification for hard-to-drug proteins., Oncogenic RAS mutants remain difficult to target with small molecules. Here, the authors show that RAS-binding Affimer proteins inhibit RAS signaling while binding diverse regions on the RAS surface, suggesting the potential to use Affimers as tools to identify new binding pockets and pharmacophores.

Tuberculosis (TB) is a top-ten cause of death worldwide. Successful treatment is often limited by insufficient diagnostic capabilities, especially at the point of care in low-resource settings. The ideal diagnostic must be fast, be cheap, and require minimal clinical resources while providing high sensitivity, selectivity, and the ability to differentiate live from dead bacteria. We describe here the development of a fast, luminescent, and affordable sensor of Hip1 (FLASH) for detecting and monitoring drug susceptibility of Mycobacterium tuberculosis (Mtb). FLASH is a selective chemiluminescent substrate for the Mtb protease Hip1 that, when processed, produces visible light that can be measured with a high signal-to-noise ratio using inexpensive sensors. FLASH is sensitive to fmol of recombinant Hip1 enzyme in vitro and can detect as few as thousands of Mtb cells in culture or in human sputum samples within minutes. The probe is highly selective for Mtb compared to other nontuberculous mycobacteria and can distinguish live from dead cells. Importantly, FLASH can be used to measure antibiotic killing of Mtb in culture with greatly accelerated timelines compared to traditional protocols. Overall, FLASH has the potential to enhance both TB diagnostics and drug resistance monitoring in resource-limited settings.

Phenol oxidases are a group of extracellular enzymes that catalyze the oxidation of lignin and phenolic compounds in soil organic matter. Despite their importance, little is known about these enzymes in tropical soils, particularly the extent to which minerals like manganese (Mn) oxides contribute to the measured activity. We assayed phenol oxidase activity in surface and subsurface (A and B) horizons from 15 soil profiles representing a steep Mn gradient under a lowland tropical forest in Panama. Fresh and sterilized (autoclaved) soils were assayed to determine the abiotic contribution to activity. Assays were conducted using 5 mM l-DOPA substrate in bicarbonate buffer at pH 8.5. We also determined amorphous metal oxide concentrations, soil pH, organic matter, and effective cation exchange capacity (CEC). We found little difference in phenol oxidase activity between fresh and autoclaved soils, indicating a considerable abiotic contribution to substrate oxidation. Furthermore, phenol oxidase activity was similar in surface and subsurface soils, despite markedly lower organic matter at depth. Phenol oxidase activity was correlated positively with amorphous Mn oxide concentrations (extractable Mn determined by acidic ammonium oxalate extraction), suggesting a potential mechanism of abiotic oxidation. These results have important implications for understanding carbon dynamics in tropical soils, because they suggest that (1) a portion of organic matter turnover is mediated by abiotic oxidation via Mn oxides, and (2) conventional measures of phenol oxidase activity reflect abiotic oxidation rather than the activity of enzymes synthesized by the microbial community.

Monitoring crystallization events in real-time is challenging but crucial for understanding the molecular dynamics associated with nucleation and crystal growth, some of nature's most ubiquitous phenomena. Recent observations have suggested that the traditional nucleation model, which describes the nucleus having already the final crystal structure, may not be valid. It appears that the molecular assembly can range during nucleation from crystalline to partially ordered to totally amorphous phases, and can change its structure during the crystallization process. Therefore, it is of critical importance to develop methods that are able to provide real-time monitoring of the molecular interactions with high temporal resolution. Here, we demonstrate that a simple and scalable approach based on interdigitated electrode array sensors (IESs) is able to provide insights on the dynamics of the crystallization process with a temporal resolution of 15 ms., An Interdigitated array Electrode sensor (IES) is used for real-time monitoring of the crystallization dynamics of organic molecules, achieving a temporal resolution of 15 ms.

The synthesis of donor–acceptor di-, tri- and tetrablock copolymers via sequential ROMP of electron rich and electron deficient paracyclophanediene monomers using Grubbs ruthenium carbene initiator, G3, is discussed. Block copolymers were initially formed with a cis/trans alkene geometry and these can be readily photoisomerised to the all trans forms. The optical and electrochemical properties of these block copolymers were examined by theory and experiment, and it was found that the HOMO and LUMO energy levels are localised on the donor and acceptor blocks, respectively.

Root-released carboxylates enhance the availability of manganese (Mn), which enters roots through transporters with low substrate specificity. Leaf Mn concentration ([Mn]) has been proposed as a signature for phosphorus (P)-mobilising carboxylates in the rhizosphere. Here we test whether leaf [Mn] provides a signature for root functional types related to P acquisition. Across 727 species at 66 sites in Australia and New Zealand, we measured leaf [Mn] as related to root functional type, while also considering soil and climate variables. To further assess the specific situations under which leaf [Mn] is a suitable proxy for rhizosphere carboxylate concentration, we studied leaf [Mn] along a strong gradient in water availability on one representative site. In addition, we focused on two systems where a species produced unexpected results. Controlling for background site-specific variation in leaf [Mn] with soil pH and mean annual precipitation, we established that mycorrhizal species have significantly lower leaf [Mn] than non-mycorrhizal species with carboxylate-releasing root structures, e.g., cluster roots. In exception to the general tendency, leaf [Mn] did not provide information about root functional types under seasonally waterlogged conditions, which increase iron availability and thereby interfere with Mn-uptake capacity. Two further exceptions were scrutinised, leading to the conclusion that they were ‘anomalous’ in not functioning like typical species in their families, as expected according to the literature. Leaf [Mn] variation provides considerable insights on differences in belowground functioning among co-occurring species. Using this approach, we concluded that, within typical mycorrhizal families, some species actually depend on a carboxylate-releasing P-mobilising strategy. Likewise, within families that are known to produce carboxylate-releasing cluster roots, some do not produce functional cluster roots when mature. An analysis of leaf [Mn] can alert us to such ‘anomalous’ species.

Tropical soils contain one-third of the carbon stored in soils globally1, so destabilization of soil organic matter caused by the warming predicted for tropical regions this century2 could accelerate climate change by releasing additional carbon dioxide (CO2) to the atmosphere3–6. Theory predicts that warming should cause only modest carbon loss from tropical soils relative to those at higher latitudes5,7, but there have been no warming experiments in tropical forests to test this8. Here we show that in situ experimental warming of a lowland tropical forest soil on Barro Colorado Island, Panama, caused an unexpectedly large increase in soil CO2 emissions. Two years of warming of the whole soil profile by four degrees Celsius increased CO2 emissions by 55 per cent compared to soils at ambient temperature. The additional CO2 originated from heterotrophic rather than autotrophic sources, and equated to a loss of 8.2 ± 4.2 (one standard error) tonnes of carbon per hectare per year from the breakdown of soil organic matter. During this time, we detected no acclimation of respiration rates, no thermal compensation or change in the temperature sensitivity of enzyme activities, and no change in microbial carbon-use efficiency. These results demonstrate that soil carbon in tropical forests is highly sensitive to warming, creating a potentially substantial positive feedback to climate change. When tropical forest soils are warmed in situ, they release more CO2 than predicted by theory, creating a potentially substantial positive feedback to climate change.

Although broadly protective, stem-targeted Abs against the influenza A virus hemagglutinin (HA) have been well studied, very limited information is available on Abs that broadly recognize the head domain. We determined the crystal structure of the HA protein of the avian H7N9 influenza virus in complex with a pan-H7, non-neutralizing, protective human Ab. The structure revealed a B cell epitope in the HA head domain trimer interface (TI). This discovery of a second major protective TI epitope supports a model in which uncleaved HA trimers exist on the surface of infected cells in a highly dynamic state that exposes hidden HA head domain features.

Wood permeability has a major effect on industrial wood processing and utilization. Wood anatomy and resin influence the permeability of wood. Understanding and manipulating these influences is important to optimize the manufacture and use of forest products. This study investigated the relationships between wood anatomical traits, radial permeability, and resin content of samples collected from 19-year-old hybrid pines (Pinus elliottii var. elliottii × Pinus caribaea var. hondurensis) from Queensland, Australia. The earlywood tracheid tangential lumen diameter and axial resin canal diameter were statistically positively correlated with radial permeability. The heartwood proportion and the frequency of axial resin canals were statistically negatively correlated with radial permeability and positively correlated with resin content. The axial resin canal diameter, sapwood proportion, latewood content, ray frequency, and earlywood tracheid lumen diameter increased from pith to bark, whereas the axial resin canal frequency decreased. Resin was found distributed throughout the wood microstructure, from pith to bark in many samples, and in both heartwood and sapwood. However, there was a much greater quantity of resin in heartwood and wood from the middle (inner radius) of the tree, with widespread occurrence of resin impregnation in the axial tracheids.

Manganese (Mn) is toxic to plants at high concentrations with recent evidence indicating slowed tropical-tree growth by high soil Mn concentrations. However, little information exists about the physiological effects of high Mn on tropical trees. We investigated physiological responses and toxicity effects of Mn in several tropical tree species. Given potential physiological interactions between Mn and phosphorus (P), we also hypothesised that P could be used to detoxify Mn. We conducted two controlled-growth experiments on seedlings of five tree species from Panama. The first experiment tested the effects of increasing concentrations of soil Mn; the second examined whether P availability mitigates Mn toxicity. Severe Mn toxicity occurred at 5 mM Mn, with toxicity symptoms in some species at 1.5 mM Mn. Responses to Mn toxicity were species-specific and depended on P supply. One species intolerant of high Mn responded by using P for detoxification; others either tolerated or detoxified Mn by another mechanism. We conclude that Mn can be toxic to tropical tree species and Mn toxicity increases P demand in some species. Other species are broadly tolerant of high Mn, suggesting that Mn tolerance might contribute to differences in species distributions and growth of tropical trees.

Indoximod has shaped our understanding of the biology of IDO1 in the control of immune responses, though its mechanism of action has been poorly understood. Previous studies demonstrated that indoximod creates a tryptophan (Trp) sufficiency signal that reactivates mTOR in the context of low Trp concentrations, thus opposing the effects caused by IDO1. Here we extend the understanding of indoximod’s mechanism of action by showing that it has pleiotropic effects on immune regulation. Indoximod can have a direct effect on T cells, increasing their proliferation as a result of mTOR reactivation. Further, indoximod modulates the differentiation of CD4+ T cells via the aryl hydrocarbon receptor (AhR), which controls transcription of several genes in response to different ligands including kynurenine (Kyn). Indoximod increases the transcription of RORC while inhibiting transcription of FOXP3, thus favoring differentiation to IL-17-producing helper T cells and inhibiting the differentiation of regulatory T cells. These indoximod-driven effects on CD8+ and CD4+ T cells were independent from the activity of IDO/TDO and from the presence of exogenous Kyn, though they do oppose the effects of Kyn produced by these Trp catabolizing enzymes. Indoximod can also downregulate expression of IDO protein in vivo in murine lymph node dendritic cells and in vitro in human monocyte-derived dendritic cells via a mechanism that involves signaling through the AhR. Together, these data improve the understanding of how indoximod influences the effects of IDO, beyond and distinct from direct enzymatic inhibition of the enzyme.

In temperate and boreal forests, competition for soil resources between free-living saprotrophs and ectomycorrhizal (EcM) fungi has been suggested to restrict saprotrophic fungal dominance to the most superficial organic soil horizons in forests dominated by EcM trees. By contrast, lower niche overlap with arbuscular mycorrhizal (AM) fungi could allow fungal saprotrophs to maintain this dominance into deeper soil horizons in AM-dominated forests. Here we used a natural gradient of adjacent forest patches that were dominated by either AM or EcM trees, or a mixture of both to determine how fungal communities characterized with high-throughput amplicon sequencing change across organic and mineral soil horizons. We found a general shift from saprotrophic to mycorrhizal fungal dominance with increasing soil depth in all forest mycorrhizal types, especially in organic horizons. Vertical changes in soil chemistry, including pH, organic matter, exchangeable cations, and extractable phosphorus, coincided with shifts in fungal community composition. Although fungal communities and soil chemistry differed among adjacent forest mycorrhizal types, variations were stronger within a given soil profile, pointing to the importance of considering horizons when characterizing soil fungal communities. Our results also suggest that in temperate forests, vertical shifts from saprotrophic to mycorrhizal fungi within organic and mineral horizons occur similarly in both ectomycorrhizal and arbuscular mycorrhizal forests.

Silicon (Si) in plants confers a number of benefits, including resistance to herbivores and water or nutrient stress. However, the dynamics of Si during long-term ecosystem development remain poorly documented, especially the changes in soils in terms of plant availability. We studied a 2-million-year soil chronosequence to examine how long-term changes in soil properties influence soil Si pools. The chronosequence exhibits extreme mineralogical changes—from carbonate-rich to quartz-rich soils—where a carbonate weathering domain is succeeded by a silicate weathering domain. Plant-available Si concentrations were lowest in young soils (Holocene

Graphene oxide (GO) membranes have great potential for separation applications due to their low-friction water permeation combined with unique molecular sieving ability. However, the practical use of deposited GO membranes is limited by the inferior mechanical robustness of the membrane composite structure derived from conventional deposition methods. Here, we report a nanostructured GO membrane that possesses great permeability and mechanical robustness. This composite membrane consists of an ultrathin selective GO nanofilm (as low as 32 nm thick) and a postsynthesized macroporous support layer that exhibits excellent stability in water and under practical permeability testing. By utilizing thin-film lift off (T-FLO) to fabricate membranes with precise optimizations in both selective and support layers, unprecedented water permeability (47 L·m-2·hr-1·bar-1) and high retention (>98% of solutes with hydrated radii larger than 4.9 A) were obtained.

In this study, we developed a single helper-dependent adenovirus (HDAd) to deliver all of the components (donor DNA, CRISPR-associated protein 9 [Cas9], and guide RNA [gRNA]) needed to achieve high-efficiency gene targeting and homology-directed repair in transduced cells. We show that these “all-in-one” HDAds are up to 117-fold more efficient at gene targeting than donor HDAds that do not express CRISPR/Cas9 in human induced pluripotent stem cells (iPSCs). The vast majority (>90%) of targeted recombinants had only one allele targeted, and this was accompanied by high-frequency indel formation in the non-targeted allele at the site of Cas9 cleavage. These indels varied in size and nature, and included large deletions of ∼8 kb. The remaining minority of recombinants had both alleles targeted (so-called bi-allelic targeting). These all-in-one HDAds represent an important platform for accomplishing and expanding the utility of homology-directed repair, especially for difficult-to-transfect cells and for in vivo applications.

Despite their importance for global biogeochemical cycles and carbon sequestration, the microbiome of tropical peatlands remains under-determined. Microbial interactions within peatlands can regulate greenhouse gas production, organic matter turnover, and nutrient cycling. Here we analyze bacterial and fungal communities along a steep P gradient in a tropical peat dome and investigate community level traits and network analyses to better understand the composition and potential interactions of microorganisms in these understudied systems and their relationship to peatland biogeochemistry. We found that both bacterial and fungal community compositions were significantly different along the P gradient, and that the low-P bog plain was characterized by distinct fungal and bacterial families. At low P, the dominant fungal families were cosmopolitan parasites and endophytes, including Clavicipitaceae (19%) in shallow soils (0–4 cm), Hypocreaceae (50%) in intermediate-depth soils (4–8 cm), and Chaetothyriaceae (45%) in deep soils (24–30 cm). In contrast, high- and intermediate-P sites were dominated by saprotrophic families at all depths. Bacterial communities were consistently dominated by the acidophilic Koribacteraceae family, with the exception of the low-P bog site, which was dominated by Acetobacteraceae (19%) and Syntrophaceae (11%). These two families, as well as Rhodospirillaceae, Syntrophobacteraceae, Syntrophorhabdaceae, Spirochaetaceae, and Methylococcaceae appeared within low-P bacterial networks, suggesting the presence of a syntrophic-methanogenic consortium in these soils. Further investigation into the active microbial communities at these sites, when paired with CH4 and CO2 gas exchange, and the quantification of metabolic intermediates will validate these potential interactions and provide insight into microbially driven biogeochemical cycling within these globally important tropical peatlands.

Improved in situ hybridization methods for mRNA detection in tissues have been developed based on the hybridization chain reaction (HCR). We show that in situ HCR methods can be used for the detection of microRNAs in tissue sections from mouse retinas. In situ HCR can be used for the detection of two microRNAs simultaneously or for the combined detection of microRNA and mRNA. In addition, miRNA in situ HCR can be combined with immunodetection of proteins. We use these methods to characterize cells expressing specific microRNAs in the mouse retina. We find that miR-181a is expressed in amacrine cells during development and in adult retinas, and it is present in both GABAergic and glycinergic amacrine cells. The detection of microRNAs with in situ HCR should facilitate studies of microRNA function and gene regulation in the retina and other tissues.

Objectives:To examine the contributions of two aspects of executive functioning (executive cognitive functions and behavioral control) to changes in diabetes management across emerging adulthood.Methods:Two hundred and forty-seven high school seniors with type 1 diabetes were assessed at baseline and followed up for 3 years. The baseline assessment battery included performance-based measures of executive cognitive functions, behavioral control, IQ estimate (IQ-est), and psychomotor speed; self-report of adherence to diabetes regimen; and glycated hemoglobin (HbA1c) assay kits as a reflection of glycemic control.Results:Linear and quadratic growth curve models were used to simultaneously examine baseline performance on four cognitive variables (executive cognitive functions, behavioral control, IQ, and psychomotor speed) as predictors of indices of diabetes management (HbA1c and adherence) across four time points. Additionally, general linear regressions examined relative contributions of each cognitive variable at individual time points. The results showed that higher behavioral control at baseline was related to lower (better) HbA1c levels across all four time points. In contrast, executive cognitive functions at baseline were related to HbA1c trajectories, accounting for increasingly more HbA1c variance over time with increasing transition to independence. IQ-est was not related to HbA1c levels or changes over time, but accounted instead for HbA1c variance at baseline (while teens were still living at home), above and beyond all other variables. Cognition was unrelated to adherence.Conclusions:Different aspects of cognition play a different role in diabetes management at different time points during emerging adulthood years.

We describe a strategy to achieve footprintless bi-allelic homology-directed repair (HDR) using helper-dependent adenoviruses (HDAds). This approach utilizes two HDAds to deliver the donor DNA. These two HDAds are identical except for their selectable marker. One expresses the puromycin N-acetyltransferase-herpes simplex virus I thymidine kinase fusion gene (PACTk), while the other expresses the hygromycin phosphotransferase-herpes simplex virus I thymidine kinase fusion gene (HyTk). Therefore, puromycin and hygromycin double resistance can be used to select for targeted HDAd integration into both alleles. Subsequently, piggyBac-mediated excision of both PACTk and HyTk will confer resistance to gancyclovir, resulting in footprintless HDR at both alleles. However, gene-targeting frequency was not high enough to achieve simultaneous targeting at both alleles. Instead, sequential targeting, whereby the two alleles were targeted one at a time, was required in order to achieve bi-allelic HDR with HDAd. Keywords: homolog-directed repair, bi-allelic, CFTR, cystic fibrosis, helper-dependent, adenovirus, homologous recombination, gene targeting, gene editing

Borylated poly(fluorene-benzothiadiazoles) (PF8-BT) are π-conjugated polymers (CPs) with deep-red/near-infrared (NIR) absorption and emission profiles suitable for in vivo optical imaging. A fully ...

Organic phosphorus (P) accumulates in soil during pedogenesis, yet information on the composition and transformation of organic P during the early stages of soil development remains scarce. We studied the top 5 cm of mineral soil immediately beneath the organic horizon from six sites (0, 35, 45, 57, 85 and 125 years) along the Hailuogou glacier foreland chronosequence, on the southeastern edge of the Tibetan Plateau. Phosphorus compounds in the soils were determined by NaOH–EDTA extraction and solution phosphorus-31 nuclear magnetic resonance (31P NMR) spectroscopy. Extractable P was dominated by phosphomonoesters (up to 51.5%) and orthophosphate (37.9–44.6%) throughout the chronosequence. The phosphomonoesters were mainly hydrolysis products of RNA and phospholipids and followed a unimodal pattern with soil age, with maximum concentration at the 57-year-old site. myo-Inositol hexakisphosphate was not detected, although scyllo-inositol hexakisphosphate accounted for 4.7–9.3% of the extracted P and d -chiro- and neo-inositol hexakisphosphates occurred small amounts in a few soils (1.1–3.3% of the extracted P). DNA accounted for 4.0–8.3% of extracted P and increased continuously along the chronosequence, associated with increased inputs of plant and microbial residues and stronger sorption to soil surfaces during the rapid decline in soil pH. Pyrophosphate, an inorganic polyphosphate, occurred in small concentrations (up to 5.0%) that fluctuated with soil age. We conclude that organic P compounds accumulate rapidly in the top mineral soil during the early stages of pedogenesis, predominantly as relatively labile compounds from plant and microbial residues.

Inhibitory neurons in mammalian cortex exhibit diverse physiological, morphological, molecular, and connectivity signatures. While considerable work has measured the average connectivity of several interneuron classes, there remains a fundamental lack of understanding of the connectivity distribution of distinct inhibitory cell types with synaptic resolution, how it relates to properties of target cells, and how it affects function. Here, we used large-scale electron microscopy and functional imaging to address these questions for chandelier cells in layer 2/3 of the mouse visual cortex. With dense reconstructions from electron microscopy, we mapped the complete chandelier input onto 153 pyramidal neurons. We found that synapse number is highly variable across the population and is correlated with several structural features of the target neuron. This variability in the number of axo-axonic ChC synapses is higher than the variability seen in perisomatic inhibition. Biophysical simulations show that the observed pattern of axo-axonic inhibition is particularly effective in controlling excitatory output when excitation and inhibition are co-active. Finally, we measured chandelier cell activity in awake animals using a cell-type-specific calcium imaging approach and saw highly correlated activity across chandelier cells. In the same experiments, in vivo chandelier population activity correlated with pupil dilation, a proxy for arousal. Together, these results suggest that chandelier cells provide a circuit-wide signal whose strength is adjusted relative to the properties of target neurons.

A purchased lot of the antibiotic hygromycin B was found to be contaminated with a novel bacterial species, which we designate Pseudomonas hygromyciniae. Characteristics of P. hygromyciniae include its ability to use a variety of compounds as carbon sources, its pathogenicity towards lettuce and Galleria mellonella, and its ability to inhibit the growth of an E. coli strain. P. hygromyciniae is unlikely to be a human pathogen, as it did not survive at 37 °C and was not cytotoxic towards a mammalian cell line. The P. hygromyciniae strain harbors a novel 250 kb megaplasmid which confers resistance to hygromycin B and contains numerous other genes predicted to encode replication and conjugation machinery. These findings indicate that commercially manufactured antibiotics represent another extreme environment that may support the growth of novel bacterial species.IMPORTANCEMicrobial ecologists have surveyed numerous natural and manmade environments in search of new microbial species. In some instances, these microbes are discovered in harsh conditions, such as deep-sea vents, and their discovery leads to better understanding of how microbes adapt to their environment. Here, we have discovered a new species of bacteria from an extreme manmade environment: a lyophilized, commercially available bottle of the antibiotic hygromycin B.

Members of the Vaccinium genus bear fruits rich in anthocyanins, a class of red-purple flavonoid pigments that provide human health benefits, although the localization and concentrations of anthocyanins differ between species: blueberry (V. corymbosum) has white flesh, while bilberry (V. myrtillus) has red flesh. Comparative transcriptomics between blueberry and bilberry revealed that MYBPA1.1 and MYBA1 strongly correlated with the presence of anthocyanins, but were absent or weakly expressed in blueberry flesh. MYBPA1.1 had a biphasic expression profile, correlating with both proanthocyanidin biosynthesis early during fruit development and anthocyanin biosynthesis during berry ripening. MYBPA1.1 was unable to induce anthocyanin or proanthocyanidin accumulation in Nicotiana benthamiana, but activated promoters of flavonoid biosynthesis genes. The MYBPA1.1 promoter is directly activated by MYBA1 and MYBPA2 proteins, which regulate anthocyanins and proanthocyanidins, respectively. Our findings suggest that the lack of VcMYBA1 expression in blueberry flesh results in an absence of VcMYBPA1.1 expression, which are both required for anthocyanin regulation. In contrast, VmMYBA1 is well expressed in bilberry flesh, up-regulating VmMYBPA1.1, allowing coordinated regulation of flavonoid biosynthesis genes and anthocyanin accumulation. The hierarchal model described here for Vaccinium may also occur in a wider group of plants as a means to co-regulate different branches of the flavonoid pathway.

The antioxidant nuclear factor erythroid 2-related factor 2 (Nrf2) is a desirable therapeutic target for a broad range of pathologies, including chronic diseases of the lung and liver, and autoimmune, neurodegenerative, and cardiovascular disorders. However, current Nrf2 activators are limited by unwanted effects due to non-specificity, and systemic distribution and action. Here we report that a 1,2-dicarbonyl moiety masks the electrophilic reactivity of the Nrf2 activator monomethyl fumarate (MMF), otherwise responsible for its non-specific effects. The 1,2-dicarbonyl compound is highly susceptible to Baeyer-Villiger oxidation, with generation of MMF specifically on exposure to pathological levels of hydrogen peroxide or peroxynitrite. Oral treatment with the MMF generating 1,2-dicarbonyl compound reversed chronic neuropathic and osteoarthritis pain in mice, and selectively activated Nrf2 at sites of oxidative stress. This 1,2-dicarbonyl platform may be used to treat additional disorders of oxidative stress, and to selectively target other therapeutics to sites of redox imbalance.

Rationale Despite a long history of use in synaptic physiology, the lobster has been a neglected model for behavioral pharmacology. A restaurateur proposed that exposing lobster to cannabis smoke reduces anxiety and pain during the cooking process. It is unknown if lobster gill respiration in air would result in significant Δ9-tetrahydrocannabinol (THC) uptake and whether this would have any detectable behavioral effects. Objective The primary goal was to determine tissue THC levels in the lobster after exposure to THC vapor. Secondary goals were to determine if THC vapor altered locomotor behavior or nociception. Methods Tissue samples were collected (including muscle, brain and hemolymph) from Homarus americanus (N = 3 per group) following 30 or 60 min of exposure to vapor generated by an e-cigarette device using THC (100 mg/mL in a propylene glycol vehicle). Separate experiments assessed locomotor behavior and hot water nociceptive responses following THC vapor exposure. Results THC vapor produced duration-related THC levels in all tissues examined. Locomotor activity was decreased (distance, speed, time-mobile) by 30 min inhalation of THC. Lobsters exhibit a temperature-dependent withdrawal response to immersion of tail, antennae or claws in warm water; this is novel evidence of thermal nociception for this species. THC exposure for 60 min had only marginal effect on nociception under the conditions assessed. Conclusions Vapor exposure of lobsters, using an e-cigarette based model, produces dose-dependent THC levels in all tissues and reduces locomotor activity. Hot water nociception was temperature dependent, but only minimal anti-nociceptive effect of THC exposure was confirmed.

Puroindolines are small, amphipathic, wheat proteins that determine the hardness of the wheat kernel and protect crops from different pathogens. Puroindoline A (PinA) and puroindoline B (PinB) are two major isoforms of puroindolines. These proteins have antibacterial and antifungal properties mainly attributed to their characteristic tryptophan-rich domains (TRDs). In this in vitro study, we investigated the antimicrobial effect of PinA and PinB synthetic peptides against the growth and biofilm formation of Campylobacter jejuni. C. jejuni is an important microaerobic, foodborne pathogen that causes gastrointestinal and neurological diseases in humans. Our results showed that: (1) PinA, but not PinB, has strong antimicrobial activity against C. jejuni clinical strains 81-176 and F38011, Escherichia coli O157:H7, methicillin-resistant Staphylococcus aureus, Salmonella enterica serovar Typhimurium, and Listeria monocytogenes; (2) The substitution of two tryptophan residues to glycine (W→G) in the TRD of PinA abolishes its antimicrobial activity against these microorganisms; (3) PinA functions additively with two common antibiotics (ciprofloxacin and erythromycin) to inhibit or inactivate C. jejuni strains; (4) PinA damages the C. jejuni cellular membrane, (5) PinA is cytotoxic to human INT 407 cells at high concentrations; and (6) PinA inhibits C. jejuni biofilm formation. In summary, this study demonstrates the antimicrobial activity of PinA against C. jejuni growth and biofilm formation and further confirms the potential use of PinA as a therapeutic agent in health care or as preservatives in the agri-food industry.

Due to tightly coupled physical, chemical, and biological processes that often behave in nonlinear, counterintuitive ways, it is argued that soil is an archetype of a complex system. Unfortunately, human intuition and decision making has been shown to be inadequate when dealing with complex systems. This poses significant challenges for managers or policy makers responding to environmental externalities where soil dynamics play a central role (e.g., biogeochemical cycles) and where full ranges of outcomes result from numerous feedback processes not easily captured by reductionist approaches. In order to improve interpretation of these soil feedbacks, a dynamic systems framework is outlined (capturing feedback often excluded from investigation or left to intuition) and then applied to agroecosystem management problems related to irrigation or tillage practices that drive nutrient cycling (e.g., soil water, nitrogen, carbon, and sodium). Key soil feedbacks are captured via a variety of previously developed models simulating soil processes and their interactions. Results indicated that soil system trade-offs arising from conservation adoption (drip irrigation or no-tillage) provided reasonable supporting evidence (via compensating feedbacks) to managers justifying slow adoption of conservation practices. Modeling soils on the foundation provided in the complex systems sciences remains an area for innovations useful for improving soil system management.

RationaleDespite a long history of use in synaptic physiology, the lobster has been a neglected model for behavioral pharmacology. A restaurateur proposed that exposing lobster to cannabis smoke reduces anxiety and pain during the cooking process. It is unknown if lobster gill respiration in air would result in significant Δ9-tetrahydrocannabinol (THC) uptake and whether this would have any detectable behavioral effects.ObjectiveThe primary goal was to determine tissue THC levels in the lobster after exposure to THC vapor. Secondary goals were to determine if THC vapor altered locomotor behavior or nociception.MethodsTissue samples were collected (including muscle, brain and hemolymph) from Homarus americanus (N=3 per group) following 30 or 60 minutes of exposure to vapor generated by an e-cigarette device using THC (100 mg/mL in a propylene glycol vehicle). Separate experiments assessed locomotor behavior and hot water nociceptive responses following THC vapor exposure.ResultsTHC vapor produced duration-related THC levels in all tissues examined. Locomotor activity was decreased (distance, speed, time-mobile) by 30 min inhalation of THC. Lobsters exhibit a temperature-dependent withdrawal response to immersion of tail, antennae or claws in warm water; this is novel evidence of thermal nociception for this species. THC exposure for 60 minutes had only marginal effect on nociception under the conditions assessed.ConclusionsVapor exposure of lobsters, using an e-cigarette based model, produces dose-dependent THC levels in all tissues and reduces locomotor activity. Hot water nociception was temperature dependent, but only minimal anti-nociceptive effect of THC exposure was confirmed.

International audience; The resource availability hypothesis predicts that plants adapted to infertile soils have high levels of anti-herbivore leaf defenses. This hypothesis has been mostly explored for secondary metabolites such as phenolics, while it remains underexplored for silica-based defenses. We determined leaf concentrations of total phenols and silicon (Si) in plants growing along the 2million-year Jurien Bay chronosequence, exhibiting an extreme gradient of soil fertility. We found that nitrogen (N) limitation on young soils led to a greater expression of phenol-based defenses, whereas old, phosphorus (P)-impoverished soils favored silica-based defenses. Both defense types were negatively correlated at the community and individual species level. Our results suggest a tradeoff among these two leaf defense strategies based on the strength and type of nutrient limitation, thereby opening up new perspectives for the resource availability hypothesis and plant defense research. This study also highlights the importance of silica-based defenses under low P supply.

Electrogenic microorganisms possess unique redox biological features, being capable of transferring electrons to the cell exterior and converting highly toxic compounds into nonhazardous forms. These microorganisms have led to the development of Microbial Electrochemical Technologies (METs), which include applications in the fields of bioremediation and bioenergy production. The optimization of these technologies involves efforts from several different disciplines, ranging from microbiology to materials science. Geobacter bacteria have served as a model for understanding the mechanisms underlying the phenomenon of extracellular electron transfer, which is highly dependent on a multitude of multiheme cytochromes (MCs). MCs are, therefore, logical targets for rational protein engineering to improve the extracellular electron transfer rates of these bacteria. However, the presence of several heme groups complicates the detailed redox characterization of MCs. In this Review, the main characteristics of electroactive Geobacter bacteria, their potential to develop microbial electrochemical technologies and the main features of MCs are initially highlighted. This is followed by a detailed description of the current methodologies that assist the characterization of the functional redox networks in MCs. Finally, it is discussed how this information can be explored to design optimal Geobacter-mutated strains with improved capabilities in METs.

A library of closely related diacetylenes has been synthesised, comprising docosa-10,12-diynediamides with small n-alkyl or cycloalkyl substituents on the termini. Colour changes of these solids in response to ultraviolet (UV) radiation and heating were analysed using a range of analytical techniques, including UV–visible/near-infrared reflectance, hand-held Raman and solid-state nuclear magnetic resonance (ssNMR) spectroscopies. Colorimetric studies showed that this class of diacetylene undergoes clear colour changes that are very sensitive to even small alterations in molecular structure. The colour change transitions of diacetylenes bearing either cyclopentyl or cyclohexyl substituents were more similar to each other than to those of analogous diacetylenes bearing either n-pentyl or n-hexyl substituents. Differential scanning calorimetry (DSC) also reflected these differences, with the cycloalkyl substituted diacetylenes showing endothermic transitions at significantly higher temperatures than both acyclic analogues. From this library, an n-hexyl substituted diacetylene shows the greatest potential as a colour change pigment, with three distinct colours that are stable at room temperature (white, blue, red), as well as an unstable yellow colour at elevated temperatures.

Significance Forest soil carbon (C) storage plays a central role in sequestrating atmospheric CO2 on timescales from centuries to millennia. However, our current understanding of soil C sequestration in response to N deposition mainly focuses on mid-to-high latitudes in the Northern Hemisphere, where N supply typically constrains forest growth. We lack data about changes in soil C stocks in tropical forests, where most ecosystems are N-rich or N-saturated. Using more than a decade of continuous N addition experiment and a meta-analysis, we found that excess N deposition can significantly increase soil C in N-rich tropical forests. However, enhanced C sequestration in tropical soils is not a good reason to justify excess N emissions to the atmosphere., Terrestrial ecosystem carbon (C) sequestration plays an important role in ameliorating global climate change. While tropical forests exert a disproportionately large influence on global C cycling, there remains an open question on changes in below-ground soil C stocks with global increases in nitrogen (N) deposition, because N supply often does not constrain the growth of tropical forests. We quantified soil C sequestration through more than a decade of continuous N addition experiment in an N-rich primary tropical forest. Results showed that long-term N additions increased soil C stocks by 7 to 21%, mainly arising from decreased C output fluxes and physical protection mechanisms without changes in the chemical composition of organic matter. A meta-analysis further verified that soil C sequestration induced by excess N inputs is a general phenomenon in tropical forests. Notably, soil N sequestration can keep pace with soil C, based on consistent C/N ratios under N additions. These findings provide empirical evidence that below-ground C sequestration can be stimulated in mature tropical forests under excess N deposition, which has important implications for predicting future terrestrial sinks for both elevated anthropogenic CO2 and N deposition. We further developed a conceptual model hypothesis depicting how soil C sequestration happens under chronic N deposition in N-limited and N-rich ecosystems, suggesting a direction to incorporate N deposition and N cycling into terrestrial C cycle models to improve the predictability on C sink strength as enhanced N deposition spreads from temperate into tropical systems.

Dialkoxy- and dialkyl-substituted phenylene benzothiadiazole paracyclophane-1,9-diene monomers undergo living ring-opening metathesis polymerization using ruthenium carbene initiators (G3) to give ...

Tropical soils contain huge carbon stocks, which climate warming is projected to reduce by stimulating organic matter decomposition, creating a positive feedback that will promote further warming. Models predict that the loss of carbon from warming soils will be mediated by microbial physiology, but no empirical data are available on the response of soil carbon and microbial physiology to warming in tropical forests, which dominate the terrestrial carbon cycle. Here we show that warming caused a considerable loss of soil carbon that was enhanced by associated changes in microbial physiology. By translocating soils across a 3000 m elevation gradient in tropical forest, equivalent to a temperature change of ± 15 °C, we found that soil carbon declined over 5 years by 4% in response to each 1 °C increase in temperature. The total loss of carbon was related to its original quantity and lability, and was enhanced by changes in microbial physiology including increased microbial carbon‐use‐efficiency, shifts in community composition towards microbial taxa associated with warmer temperatures, and increased activity of hydrolytic enzymes. These findings suggest that microbial feedbacks will cause considerable loss of carbon from tropical forest soils in response to predicted climatic warming this century.

Tropical forest productivity is often thought to be limited by soil phosphorus (P) availability. Phosphorus availability might therefore constrain potential increases in growth as the atmospheric CO₂ concentration increases, yet there is little experimental evidence with which to evaluate this hypothesis. We hypothesized that while all species would respond more strongly to elevated CO₂ when supplied with extra P, the responses of individual species would also depend on their habitat associations with either high‐ or low‐P soils. We further hypothesized that this effect would be exacerbated by a reduction in transpiration rate under elevated CO₂, as transpiration may aid in P acquisition. We used a pot experiment to test the effects of P addition on the physiological and growth response to elevated CO₂ of eight tropical tree species with contrasting distributions across a soil P gradient in Panamanian lowland forests. Seedlings were grown in an ambient (400 ppm) or elevated (800 ppm) CO₂‐controlled glasshouse in either a high‐ or low‐P treatment to quantify the effects of P limitation on relative growth rate (RGR), transpiration, maximum photosynthetic rate and foliar nutrients. We found evidence of limitation by P and CO₂ on growth, photosynthesis, foliar nutrients and transpiration. However, the affinity of a species for P, defined as the species distribution relative to P availability, was not correlated with RGR or transpiration responses to elevated CO₂ in either the low‐P or high‐P treatments. Transpiration rates decreased under elevated CO₂, but foliar P was greater for some species under elevated CO₂, suggesting a greater capacity for upregulation of P acquisition in species associated with low‐P soils. Our results show that tropical forest responses to elevated CO₂ will be species‐specific and not necessarily explained by P affinities based on distribution, which poses challenges for predictions of community‐wide responses. A free Plain Language Summary can be found within the Supporting Information of this article.

Several next-generation (universal) influenza vaccines and broadly neutralizing antibodies (bNAbs) are in clinical development. Some of these mediate inhibitions of virus replication at the postentry stage or use Fc-dependent mechanisms. Nonneutralizing antibodies have the potential to mediate enhancement of viral infection or disease. In the current study, two monoclonal antibodies (MAbs) 72/8 and 69/1, enhanced respiratory disease (ERD) in mice following H3N2 virus challenge by demonstrating increased lung pathology and changes in lung cytokine/chemokine levels. MAb 78/2 caused changes in the lung viral loads in a dose-dependent manner. Both MAbs increased HA sensitivity to trypsin cleavage at a higher pH range, suggesting MAb-induced conformational changes. pHrodo-labeled virus particles' entry and residence time in the endocytic compartment were tracked during infection of Madin-Darby canine kidney (MDCK) cells. Both MAbs reduced H3N2 virus residence time in the endocytic pathway, suggesting faster virus fusion kinetics. Structurally, 78/2 and 69/1 Fabs bound the globular head or base of the head domain of influenza hemagglutinin (HA), respectively, and induced destabilization of the HA stem domain. Together, this study describes Mab-induced destabilization of the influenza HA stem domain, faster kinetics of influenza virus fusion, and ERD in vivo. The in vivo animal model and in vitro assays described could augment preclinical safety evaluation of antibodies and next-generation influenza vaccines that generate antibodies which do not block influenza virus-receptor interaction.

Dialkoxy and dialkyl substituted paracyclophane-1,9-dienes undergo bidirectional ring opening metathesis polymerisation (ROMP) on addition of bifunctional Hoveyda-Grubbs initiators. The living nature of the polymerisation was demonstrated for different [monomer]/[initiator] ratios and block copolymers were prepared by sequential ROMP and ROMP-ATRP to give fully conjugated rod-rod-rod and partially conjugated coil-rod-coil ABA and BAB triblock copolymers, respectively. These strategies were successfully extended to the synthesis of the corresponding pentablock copolymers. Optical and electrochemical properties of cis/trans and all trans homo and block copolymers are discussed.

Tungsten borides, such as tungsten tetraboride (WB4) exhibit a wide range of appealing physical properties, including superhardness, chemical inertness and electronic conductivity. Among the various tungsten borides, the most puzzling remains WB4, with its crystal structure to linger in question for over half a century (Lech et al. in Proc Natl Acad Sci USA 112:3223–3228, 2015). In the present investigation, polycrystalline WB4 samples have been synthesized with two different methods and characterized at the atomic level by combining X-ray diffraction, scanning electron microscopy and nuclear magnetic resonance spectroscopy. The 11B multiple quantum MAS experiment revealed a range of boron sites that were not resolved within the experiment. This result is in contrast to the 11B MAS spectrum of WB2 with four resolved, discernible boron resonances. However, despite the structural complexity and boron-site variety in WB4, the detection of a single exponential of 11B spin–lattice relaxation recovery suggested that all of the boron sites relaxed with a single time constant. The Knight shift (K) was found to be independent of temperature while the $$ T_{1}^{ - 1} $$ was governed by the Korringa law with a Korringa product T1T = 350 sK across the entire temperature range (168–437 K) of this study. The measured Korringa product was small, indicating substantial spin–lattice relaxation resulting from coupling with the conduction carriers. The abovementioned experimental results not only clearly rule out structures, such as the “MoB4-type phase” of WB4, with the resulting Fermi level in the pseudo-gap as has previously been predicted theoretically; but they also provide a comprehensible and valuable insight into the structural and electronic properties of WB4 at the atomic level.

B-type natriuretic peptide (BNP) is a cardiac-derived neurohormone regulated by intra-cardiac pressure. N-terminal pro-B-type natriuretic peptide (NT-proBNP) is a bio-inert product of BNP production identified as a blood biomarker for diagnosing cardiac distress. Rapid, point-of-care quantification of NT-proBNP concentration is challenging because accurate recognition is often masked by interference of native BNP in blood samples. Recently, monoclonal antibodies have been engineered with high specificity to the tail, i.e. cleaved region, of NT-proBNP; herein, we present a case study using two monoclonal antibodies for NT-proBNP, respectively, as detector and capture antibodies. Using the selected antibodies, we fabricated and characterized a lateral flow immunoassay for NT-proBNP using antibody-labeled quantum dots to improve limit of detection. The reported lateral flow immunoassay exhibited clinically-relevant linear response range: 50–1200 pg mL−1, good stability and repeatability with pooled variance of 8.8% across concentrations of NT-proBNP—highlighting a method for high-resolution, low-cost quantification of NT-proBNP as a point-of-care, single-step diagnostic sensor. Keywords: NT-proBNP, cardiac biomarker, lateral flow, immunoassay, quantum dot