Investigation of machine learning algorithms for taxonomic classification of marine metagenomes.

Microbial communities play key roles in ocean ecosystems through regulation of biogeochemical processes such as carbon and nutrient cycling, food web dynamics, and gut microbiomes of invertebrates, fish, reptiles, and mammals. Assessments of marine microbial diversity are therefore critical to understanding spatiotemporal variations in microbial community structure and function in ocean ecosystems. With recent advances in DNA shotgun sequencing for metagenome samples and computational analysis, it is now possible to access the taxonomic and genomic content of ocean microbial communities to study their structural patterns, diversity, and functional potential. However, existing taxonomic classification tools depend upon manually curated phylogenetic trees, which can create inaccuracies in metagenomes from less well-characterized communities, such as from ocean water. Herein, we explore the utility of deep learning tools-DeepMicrobes and a novel Residual Network architecture-that leverage natural language processing and convolutional neural network architectures to map input sequence data (k-mers) to output labels (taxonomic groups) without reliance on a curated taxonomic tree. We trained both models using metagenomic reads simulated from marine microbial genomes in the MarRef database. The performance of both models (accuracy, precision, and percent microbe predicted) was compared with the standard taxonomic classification tool Kraken2 using 10 complex metagenomic data sets simulated from MarRef. Our results demonstrate that time, compute power, and microbial genomic diversity still pose challenges for machine learning (ML). Moreover, our results suggest that high genome coverage and rectification of class imbalance are prerequisites for a well-trained model, and therefore should be a major consideration in future ML work. IMPORTANCE Taxonomic profiling of microbial communities is essential to model microbial interactions and inform habitat conservation. This work develops approaches in constructing training/testing data sets from publicly available marine metagenomes and evaluates the performance of machine learning (ML) approaches in read-based taxonomic classification of marine metagenomes. Predictions from two models are used to test accuracy in metagenomic classification and to guide improvements in ML approaches. Our study provides insights on the methods, results, and challenges of deep learning on marine microbial metagenomic data sets. Future machine learning approaches can be improved by rectifying genome coverage and class imbalance in the training data sets, developing alternative models, and increasing the accessibility of computational resources for model training and refinement.

Cooperative Self-Assembly of Dimer Junctions Driven by π Stacking Leads to Conductance Enhancement.

We demonstrate enhanced electronic transport through dimer molecular junctions, which self-assemble between two gold electrodes in π-π stabilized binding configurations. Single molecule junction conductance measurements show that benzimidazole molecules assemble into dimer junctions with a per-molecule conductance that is higher than that in monomer junctions. Density functional theory calculations reveal that parallel stacking of two benzimidazoles between electrodes is the most energetically favorable due to the large π system. Imidazole is smaller and has greater conformational freedom to access different stacking angles. Transport calculations confirm that the conductance enhancement of benzimidazole dimers results from the changed binding geometry of dimers on gold, which is stabilized and made energetically accessible by intermolecular π stacking. We engineer imidazole derivatives with higher monomer conductance than benzimidazole and large intermolecular interaction that promote cooperative in situ assembly of more transparent dimer junctions and suggest at the potential of molecular devices based on self-assembled molecular layers.

Studies on the Regioselective Rearrangement of Azanorbornanic Aminyl Radicals into 2,8-Diazabicyclo[3.2.1]oct-2-ene Systems.

Aminyl radicals are nitrogen-centered radicals of interest in synthetic strategies involving C-N bond formation due to their high reactivity. These intermediate radicals are generated by the reaction of an organic azide with tributyltin hydride (Bu3SnH) in the presence of substoichiometric amounts of azobisisobutyronitrile (AIBN). In this work, we report the regioselective rearrangement of azanorbornanic ([2.2.1]azabicyclic) aminyl radicals into 2,8-diazabicyclo[3.2.1]oct-2-ene systems. With the aim to establish the structural requirements for this ring expansion, we have studied the effect of different bridgehead atoms of the [2.2.1]bicyclic system and the presence of an alkyl substituent at C4. Attempts to perform this ring expansion on a monocyclic analogue have been also explored to evaluate the influence of the bicyclic skeleton on the rearrangement. A detailed mechanistic proposal supported by computational studies is reported.

Controlled masking and targeted release of redox-cycling ortho-quinones via a C-C bond-cleaving 1,6-elimination.

Natural products that contain ortho-quinones show great potential as anticancer agents but have been largely discarded from clinical development because their redox-cycling behaviour results in general systemic toxicity. Here we report conjugation of ortho-quinones to a carrier, which simultaneously masks their underlying redox activity. C-benzylation at a quinone carbonyl forms a redox-inactive benzyl ketol. Upon a specific enzymatic trigger, an acid-promoted, self-immolative C-C bond-cleaving 1,6-elimination mechanism releases the redox-active hydroquinone inside cells. By using a 5-lipoxygenase modulator, ß-lapachone, we created cathepsin-B-cleavable quinone prodrugs. We applied the strategy for intracellular release of ß-lapachone upon antibody-mediated delivery. Conjugation of protected ß-lapachone to Gem-IgG1 antibodies, which contain the variable region of gemtuzumab, results in homogeneous, systemically non-toxic and conditionally stable CD33+-specific antibody-drug conjugates with in vivo efficacy against a xenograft murine model of acute myeloid leukaemia. This protection strategy could allow the use of previously overlooked natural products as anticancer agents, thus extending the range of drugs available for next-generation targeted therapeutics.

Diverse nitrogen cycling pathways across a marine oxygen gradient indicate nitrogen loss coupled to chemoautotrophic activity.

Genetic markers and geochemical assays of microbial nitrogen cycling processes, including autotrophic and heterotrophic denitrification, anammox, ammonia oxidation, and nitrite oxidation, were examined across the oxycline, suboxic, and anoxic zones of the Cariaco Basin, Venezuela. Ammonia and nitrite oxidation genes were expressed through the entire gradient. Transcripts associated with autotrophic and heterotrophic denitrifiers were mostly confined to the suboxic zone and below but were also present in particles in the oxycline. Anammox genes and transcripts were detected over a narrow depth range near the bottom of the suboxic zone and coincided with secondary NO2 - maxima and available NH4 + . Dissolved inorganic nitrogen (DIN) amendment incubations and comparisons between our sampling campaigns suggested that denitrifier activity may be closely coupled with NO3 - availability. Expression of denitrification genes at depths of high rates of chemoautotrophic carbon fixation and phylogenetic analyses of nitrogen cycling genes and transcripts indicated a diverse array of denitrifiers, including chemoautotrophs capable of using NO3 - to oxidize reduced sulfur species. Thus, results suggest that the Cariaco Basin nitrogen cycle is influenced by autotrophic carbon cycling in addition to organic matter oxidation and anammox.

Cyclopropenylidenes as Strong Carbene Anchoring Groups on Au Surfaces.

The creation of stable molecular monolayers on metallic surfaces is a fundamental challenge of surface chemistry. N-Heterocyclic carbenes (NHCs) were recently shown to form self-assembled monolayers that are significantly more stable than the traditional thiols on Au system. Here we theoretically and experimentally demonstrate that the smallest cyclic carbene, cyclopropenylidene, binds even more strongly than NHCs to Au surfaces without altering the surface structure. We deposit bis(diisopropylamino)cyclopropenylidene (BAC) on Au(111) using the molecular adduct BAC-CO2 as a precursor and determine the structure, geometry, and behavior of the surface-bound molecules through high-resolution X-ray photoelectron spectroscopy, atomic force microscopy, and scanning tunneling microscopy. Our experiments are supported by density functional theory calculations of the molecular binding energy of BAC on Au(111) and its electronic structure. Our work is the first demonstration of surface modification with a stable carbene other than NHC; more broadly, it drives further exploration of various carbenes on metal surfaces.

Regioselectivity of the 1,3-Dipolar Cycloaddition of Organic Azides to 7-Heteronorbornadienes. Synthesis of ß-Substituted Furans/Pyrroles.

An efficient procedure for the preparation of ß-substituted furans/pyrroles is presented. The methodology is based on the use of 7-oxa/azanorbornadienes as dipolarophiles in 1,3-dipolar cycloaddition with benzyl azide. The triazoline cycloadduct thus formed spontaneously decomposes via a retro-Diels-Alder (rDA) reaction to afford a ß-substituted furan/pyrrole derivative and a stable triazole. The scope of this tandem 1,3-dipolar cycloaddition/rDA reaction was studied with thirteen 7-heteronorbornadienes. This study allowed a deep knowledge of the regioselectivity of the reaction, which can be tuned through the substituents of the heteronorbornadienic systems.

Stable Pyrrole-Linked Bioconjugates through Tetrazine-Triggered Azanorbornadiene Fragmentation.

An azanorbornadiene bromovinyl sulfone reagent for cysteine-selective bioconjugation has been developed. Subsequent reaction with dipyridyl tetrazine leads to bond cleavage and formation of a pyrrole-linked conjugate. The latter involves ligation of the tetrazine to the azanorbornadiene-tagged protein through inverse electron demand Diels-Alder cycloaddition with subsequent double retro-Diels-Alder reactions to form a stable pyrrole linkage. The sequence of site-selective bioconjugation followed by bioorthogonal bond cleavage was efficiently employed for the labelling of three different proteins. This method benefits from easy preparation of these reagents, selectivity for cysteine, and stability after reaction with a commercial tetrazine, which has potential for the routine preparation of protein conjugates for chemical biology studies.

Environmental DNA reveals seasonal shifts and potential interactions in a marine community.

Environmental DNA (eDNA) analysis allows the simultaneous examination of organisms across multiple trophic levels and domains of life, providing critical information about the complex biotic interactions related to ecosystem change. Here we used multilocus amplicon sequencing of eDNA to survey biodiversity from an eighteen-month (2015-2016) time-series of seawater samples from Monterey Bay, California. The resulting dataset encompasses 663 taxonomic groups (at Family or higher taxonomic rank) ranging from microorganisms to mammals. We inferred changes in the composition of communities, revealing putative interactions among taxa and identifying correlations between these communities and environmental properties over time. Community network analysis provided evidence of expected predator-prey relationships, trophic linkages, and seasonal shifts across all domains of life. We conclude that eDNA-based analyses can provide detailed information about marine ecosystem dynamics and identify sensitive biological indicators that can suggest ecosystem changes and inform conservation strategies.

Identifying the molecular adsorption site of a single molecule junction through combined Raman and conductance studies.

Single-molecule junctions are ideal test beds for investigating the fundamentals of charge transport at the nanoscale. Conducting properties are strongly dependent on the metal-molecule interface geometry, which, however, is very poorly characterized due to numerous experimental challenges. We report on a new methodology for characterizing the adsorption site of single-molecule junctions through the combination of surface enhanced Raman scattering (SERS), current-voltage (I-V) curve measurements, and density functional theory simulations. This new methodology discriminates between three different adsorption sites for benzenedithiol and aminobenzenethiol junctions, which cannot be identified by solo measurements of either SERS or I-V curves. Using this methodology, we determine the interface geometry of these two prototypical molecules at the junction and its time evolution. By modulating the applied voltage, we can change and monitor the distribution of adsorption sites at the junction.

Azabicyclic vinyl sulfones for residue-specific dual protein labelling.

We have developed [2.2.1]azabicyclic vinyl sulfone reagents that simultaneously enable cysteine-selective protein modification and introduce a handle for further bioorthogonal ligation. The reaction is fast and selective for cysteine relative to other amino acids that have nucleophilic side-chains, and the formed products are stable in human plasma and are moderately resistant to retro Diels-Alder degradation reactions. A model biotinylated [2.2.1]azabicyclic vinyl sulfone reagent was shown to efficiently label two cysteine-tagged proteins, ubiquitin and C2Am, under mild conditions (1-5 equiv. of reagent in NaPi pH 7.0, room temperature, 30 min). The resulting thioether-linked conjugates were stable and retained the native activity of the proteins. Finally, the dienophile present in the azabicyclic moiety on a functionalised C2Am protein could be fluorescently labelled through an inverse electron demand Diels-Alder reaction in cells to allow selective apoptosis imaging. The combined advantages of directness, site-specificity and easy preparation mean [2.2.1]azabicyclic vinyl sulfones can be used for residue-specific dual protein labelling/construction strategies with minimal perturbation of native function based simply on the attachment of an [2.2.1]azabicyclic moiety to cysteine.

High-Performance Field-Effect Transistors Based on αP and ßP.

The electronic properties of black and blue phosphorus nanoribbons are investigated, which enables the proposal of junction-free field-effect transistors that comprise metallic armchair nanoribbons as electrodes and, in between, a semiconducting zigzag nanoribbon as channel material (cut out of a single sheet of monolayer black or blue phosphorus). Using first-principles calculations and the nonequilibrium Green's function method, the proposed field-effect transistors are characterized. It is found that it is possible to achieve outstanding performance, with high on/off ratios, low subthreshold swings, and high transconductances.

Crossover in the inelastic electron tunneling spectra of conjugated molecules with direct Au-C links.

We use inelastic electron tunneling spectroscopy (IETS) first-principles simulations to identify and characterize the different vibrational modes of single conjugated molecules bonded to Au metal electrodes. The molecules are polyphenyls (with 1 to 4 benzene units) bonded to Au via highly conducting direct Au-C bonds. The short molecule shows near resonant elastic transmission, with a crossover to tunneling for the longer backbones. The calculated inelastic spectra exhibit dips in the IETS signal of the short molecule and peaks for the longer molecules. We characterize the symmetry of vibrational modes and scattering states and discuss the changes with increasing length, where the inelastic signal of different modes can be amplified, quenched or present a crossover as more benzene units are added to the molecular backbone. This analysis rationalizes the observed trends as a function of molecular length and illustrates the role of electronic and vibrational properties on Au-C bonded molecular junctions.

The Scientific Legacy of the CARIACO Ocean Time-Series Program.

The CARIACO (Carbon Retention in a Colored Ocean) Ocean Time-Series Program station, located at 10.50°N, 64.66°W, observed biogeochemical and ecological processes in the Cariaco Basin of the southwestern Caribbean Sea from November 1995 to January 2017. The program completed 232 monthly core cruises, 40 sediment trap deployment cruises, and 40 microbiogeochemical process cruises. Upwelling along the southern Caribbean Sea occurs from approximately November to August. High biological productivity (320-628 g C m-2 y-1) leads to large vertical fluxes of particulate organic matter, but only approximately 9-10 g C m-2 y-1 fall to the bottom sediments (∼1-3% of primary production). A diverse community of heterotrophic and chemoautotrophic microorganisms, viruses, and protozoa thrives within the oxic-anoxic interface. A decrease in upwelling intensity from approximately 2003 to 2013 and the simultaneous overfishing of sardines in the region led to diminished phytoplankton bloom intensities, increased phytoplankton diversity, and increased zooplankton densities. The deepest waters of the Cariaco Basin exhibited long-term positive trends in temperature, salinity, hydrogen sulfide, ammonia, phosphate, methane, and silica. Earthquakes and coastal flooding also resulted in the delivery of sediment to the seafloor. The program's legacy includes climate-quality data from suboxic and anoxic habitats and lasting relationships between international researchers.

In Situ Formation of N-Heterocyclic Carbene-Bound Single-Molecule Junctions.

Self-assembled monolayers (SAMs) formed using N-heterocyclic carbenes (NHCs) have recently emerged as thermally and chemically ultrastable alternatives to those formed from thiols. The rich chemistry and strong σ-donating ability of NHCs offer unique prospects for applications in nanoelectronics, sensing, and electrochemistry. Although stable in SAMs, free carbenes are notoriously reactive, making their electronic characterization challenging. Here we report the first investigation of electron transport across single NHC-bound molecules using the scanning tunneling microscope-based break junction (STM-BJ) technique. We develop a series of air-stable metal NHC complexes that can be electrochemically reduced in situ to form NHC-electrode contacts, enabling reliable single-molecule conductance measurements of NHCs under ambient conditions. Using this approach, we show that the conductance of an NHC depends on the identity of the single metal atom to which it is coordinated in the junction. Our observations are supported by density functional theory (DFT) calculations, which also firmly establish the contributions of the NHC linker to the junction transport characteristics. Our work demonstrates a powerful method to probe electron transfer across NHC-electrode interfaces; more generally, it opens the door to the exploitation of surface-bound NHCs in constructing novel, functionalized electrodes and/or nanoelectronic devices.

Evaluation of marine zooplankton community structure through environmental DNA metabarcoding.

Zooplankton dominate the abundance and biomass of multicellular animals in pelagic marine environments; however, traditional methods to characterize zooplankton communities are invasive and laborious. This study compares zooplankton taxonomic composition revealed through metabarcoding of the cytochrome oxidase I (COI) and 18S rRNA genes to traditional morphological identification by microscopy. Triplicates of three different sample types were collected from three coral reef sites in the Florida Keys National Marine Sanctuary: (1) 1 L surface seawater samples prefiltered through 3 µm filters and subsequently collected on 0.22 µm filters for eDNA (PF-eDNA); (2) 1 L surface seawater samples filtered on 0.22 µm pore-size filters (environmental DNA; eDNA), and (3) zooplankton tissue samples from 64 µm, 200 µm, and 500 µm mesh size net tows. The zooplankton tissue samples were split, with half identified morphologically and tissue DNA (T-DNA) extracted from the other half. The COI and 18S rRNA gene metabarcoding of PF-eDNA, eDNA, and T-DNA samples was performed using Illumina MiSeq. Of the families detected with COI and 18S rRNA gene metabarcoding, 40% and 32%, respectively, were also identified through morphological assessments. Significant differences in taxonomic composition were observed between PF-DNA, eDNA, and T-DNA with both genetic markers. PF-eDNA resulted in detection of fewer taxa than the other two sample types; thus, prefiltering is not recommended. All dominant copepod taxa (> 5% of total abundance) were detected with eDNA, T-DNA, and morphological assessments, demonstrating that eDNA metabarcoding is a promising technique for future biodiversity assessments of pelagic zooplankton in marine systems.

Salinity induction of recycling Crassulacean acid metabolism and salt tolerance in plants of Talinum triangulare.

Background and Aims: Crassulacean acid metabolism (CAM) can be induced by salinity, thus conferring the plant higher water-use efficiency. Talinum triangulare does not frequently encounter salt in its natural habitat but is cultivated in soils that may become salinized. Here we examined whether plants of T. triangulare can grow in saline soils and show salt-induced CAM. Methods: Leaf gas exchange, carbon isotopic ratio (δ13C), nocturnal acid accumulation (ΔH+), water relations, photosynthetic pigment and mineral contents, leaf anatomy and growth were determined in greenhouse in plants irrigated with 0, 150, 300 and 400 mm NaCl. Key Results: Salinity reduced gas exchange and induced CAM, ΔH+ reaching 50.2 µmol H+ g-1 fresh mass under 300 mm NaCl. No nocturnal CO2 uptake, but compensation, was observed. Values of δ13C were lowest under 0 and 400 mm NaCl, and highest under 150 and 300 mm. The difference in osmotic potential (ψs) between control and treated plants averaged 0.45 MPa for the three [NaCl] values, the decrease in ψs being accounted for by up to 63 % by Na+ and K+. Pigment contents were unaffected by treatment, suggesting lack of damage to the photosynthetic machinery. Changes in stomatal index with unchanged stomatal density in newly expanded leaves suggested inhibited differentiation of epidermal cells into stomata. Whole-leaf and parenchymata thickness increased under 150 and 300 mm NaCl. Only plants irrigated with 400 mm NaCl showed reductions in biomass (stems, 41 %; reproductive structures, 78 %). The K/Na molar ratio decreased with [NaCl] from 2.0 to 0.4. Conclusions: The operation of CAM in the recycling mode was evidenced by increased ΔH+ with no nocturnal CO2 uptake. Talinum triangulare can be classified as a halo-tolerant species based on its low K/Na molar ratio under salinity and the relatively small reduction in growth only at the highest [NaCl].

What is the potential for dark CO2 fixation in the facultative crassulacean acid metabolism species Talinum triangulare?

In obligate Crassulacean acid metabolism (CAM) plants, dark CO2 fixation is almost the sole route of CO2 fixation and, under drought, continues for long periods. In contrast, in plants of the facultative CAM species Talinum triangulare under experimental drought, dark CO2 fixation provides a small proportion of the daily assimilation observed in watered plants and occurs only for a few days, after which almost nil CO2 fixation is observed. Under field conditions, with a practically unlimited substrate volume, drought-induced CAM might operate for a longer period and make a higher contribution to daily CO2 fixation. Greenhouse-grown plants of T. triangulare were subjected to low and nearly constant soil water content; the operation of CAM was assessed through the measurement of nocturnal proton accumulation and dark CO2 fixation. Dark CO2 fixation appeared 19d after the onset of drought; its contribution during three months of experiment to daily CO2 assimilation ranged from 0.5 to 30.7% with a mean of 13.5%. Twenty days after the beginning of treatment, nocturnal proton accumulation increased six times and remained high for over three months. In spite of low soil water content, leaves did not engage in dark CO2 fixation all the time but dark CO2 fixation was large enough to produce an increase in relative (13)C composition of mature leaves compared to watered plants but not to the value in short-term drought experiments. Leaf anatomical characteristics may guarantee the achievement of higher rates of dark CO2 fixation but results evidence the occurrence of a limit to the expression of CAM that remains to be determined.

The necessary limits to temptation: the Turnkey Project.

In case of special engineering projects of important relevance it is interesting to pay attention to several possible risks; some of them are in the field of morality or ethics. Due to the social importance of these risks, additional considerations or even additional warranties are justified.

Tuberculosis pulmonar en relación con adalimumab: estudio de 3 casos / Pulmonary Tuberculosis Associated to Adalimumab: a Study of 3 Cases

Los fármacos antagonistas del factor de necrosis tumoral alfa representan un importante avance en el tratamiento de enfermedades inflamatorias como la artritis reumatoide, las espondiloartropatías y la enfermedad inflamatoria intestinal. Se reconoce el incremento de tuberculosis con infliximab, pero disponemos de menos datos que relacionen la tuberculosis específicamente con adalimumab. Presentamos los casos de 2 pacientes con artritis reumatoide y un paciente con espondilitis anquilopoyética en tratamiento con adalimumab, que desarrollaron tuberculosis pulmonar y diseminada a pesar de seguir las medidas de cribado y profilaxis recomendadas por las guías, y revisamos la asociación entre el tratamiento con antagonistas del factor de necrosis tumoral alfa y tuberculosis (AU)

Tumour necrosis factor-alpha antagonist drugs represent a significant advance in the treatment of inflammatory diseases, such as rheumatoid arthritis, spondyloarthropathies, and intestinal inflammatory disease. The increase in tuberculosis with infliximab is known, but there is less data available that specifically associates tuberculosis with adalimumab. We present the cases of 2 patients with rheumatoid arthritis and one patient with ankylopoietic spondylitis on treatment with adalimumab, who developed pulmonary and disseminated tuberculosis despite following the screening and prophylaxis measures recommended in guidelines. We also review the association between treatment with tumour necrosis factor-alpha antagonists and tuberculosis (AU)