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Advanced Node.js Development is a practical, project-based book that provides you with all you need to progress as a Node.js developer. Node is a ubiquitous technology on the modern web, and an essential part of any web developer’s toolkit. If you're looking to create real-world Node applications, or you want to switch careers or launch a side-project to generate some extra income, then you're in the right place. This book was written around a single goal: turning you into a professional Node developer capable of developing, testing, and deploying real-world production applications. There's no better time to dive in. According to the 2018 Stack Overflow Survey, Node is in the top ten for back-end popularity and back-end salary. This book is built from the ground up around the latest version of Node.js (version 9.x.x). You'll be learning all the cutting-edge features available only in the latest software versions. This book delivers advanced skills that you need to become a professional Node developer. Along this journey you'll create your own API, you'll build a full real-time web app and create projects that apply the latest Async and Await technologies. Andrew Mead maps everything out for you in this book so that you can learn how to build powerful Node.js projects in a comprehensive, easy-to-follow package designed to get you up and running quickly.

Learning Node.js Development is a practical, project-based book that provides you with all you need to get started as a Node.js developer. Node is a ubiquitous technology on the modern web, and an essential part of any web developers' toolkit. If you are looking to create real-world Node applications, or you want to switch careers or launch a side project to generate some extra income, then you're in the right place. This book has been written around a single goal—turning you into a professional Node developer capable of developing, testing, and deploying real-world production applications. Learning Node.js Development is built from the ground up around the latest version of Node.js (version 9.x.x). You'll be learning all the cutting-edge features available only in the latest software versions. This book cuts through the mass of information available around Node and delivers the essential skills that you need to become a Node developer. It takes you through creating complete apps and understanding how to build, deploy, and test your own Node apps. It maps out everything in a comprehensive, easy-to-follow package designed to get you up and running quickly.

Antimicrobial resistance is a global threat to both human and animal health, and transfer of resistance between these spheres is recognised as a key concern for all species. Selection for resistance at sub-inhibitory antimicrobial concentrations has been characterised for some bacteria-antimicrobial combinations but there is little data from non-laboratory strains, and veterinary antimicrobials and bacterial species. Here, we demonstrate a minimum selective concentration of 0.06 mg/L (1/6 xMIC) for florfenicol in wild-type Pasteurella multocida , through competition experiments between a susceptible strain and a floR -resistant mutant. We also show that sub-inhibitory concentrations of florfenicol do not appear to significantly select for de novo resistance in P. multocida and present the challenges with adapting experimental protocols between bacterial species. These results have important implications for antimicrobial resistance selection at sub-inhibitory concentrations, method development for within-species differentiation in novel bacterial species, and application to policy regarding antimicrobial contamination in animal-feed.

Improving photosynthesis is a major target for increasing crop yields and ensuring food security. Phenotyping of photosynthesis in the field is critical to understand the limits to crop performance in agricultural settings. Yet, detailed phenotyping of photosynthetic traits is relatively scarce in field-grown wheat, with previous studies focusing on narrow germplasm selections. Flag leaf photosynthetic traits, crop development, and yield traits were compared in 64 field-grown wheat cultivars in the UK. Pre-anthesis and post-anthesis photosynthetic traits correlated significantly and positively with grain yield and harvest index (HI). These traits included net CO₂ assimilation measured at ambient CO₂ concentrations and a range of photosynthetic photon flux densities, and traits associated with the light response of photosynthesis. In most cultivars, photosynthesis decreased post-anthesis compared with pre-anthesis, and this was associated with decreased Rubisco activity and abundance. Heritability of photosynthetic traits suggests that phenotypic variation can be used to inform breeding programmes. Specific cultivars were identified with traits relevant to breeding for increased crop yields in the UK: pre-anthesis photosynthesis, post-anthesis photosynthesis, light response of photosynthesis, and Rubisco amounts. The results indicate that flag leaf longevity and operating photosynthetic activity in the canopy can be further exploited to maximize grain filling in UK bread wheat.

High soil phosphorus (P) concentration is frequently shown to reduce root colonization by arbuscular mycorrhizal (AM) fungi, but the influence of P on the diversity of colonizing AM fungi is uncertain. We used terminal restriction fragment length polymorphism (T-RFLP) of 18S rDNA and cloning to assess diversity of AM fungi colonizing maize (Zea mays), soybean (Glycene max) and field violet (Viola arvensis) at three time points in one season along a P gradient of 10–280 mg l−1 in the field. Percentage AM colonization changed between sampling time points but was not reduced by high soil P except in maize. There was no significant difference in AM diversity between sampling time points. Diversity was reduced at concentrations of P > 25 mg l−1, particularly in maize and soybean. Both cloning and T-RFLP indicated differences between AM communities in the different host species. Host species was more important than soil P in determining the AM community, except at the highest P concentration. Our results show that the impact of soil P on the diversity of AM fungi colonizing plants was broadly similar, despite the fact that different plants contained different communities. However, subtle differences in the response of the AM community in each host were evident.

Ammonia-oxidising archaea of the phylum Thaumarchaeota are important organisms in the nitrogen cycle, but the mechanisms driving their radiation into diverse ecosystems remain underexplored. Here, existing thaumarchaeotal genomes are complemented with 12 genomes belonging to the previously under-sampled Nitrososphaerales to investigate the impact of lateral gene transfer (LGT), gene duplication and loss across thaumarchaeotal evolution. We reveal a major role for gene duplication in driving genome expansion subsequent to early LGT. In particular, two large LGT events are identified into Nitrososphaerales and the fate of these gene families is highly lineage-specific, being lost in some descendant lineages, but undergoing extensive duplication in others, suggesting niche-specific roles. Notably, some genes involved in carbohydrate transport or coenzyme metabolism were duplicated, likely facilitating niche specialisation in soils and sediments. Overall, our results suggest that LGT followed by gene duplication drives Nitrososphaerales evolution, highlighting a previously under-appreciated mechanism of genome expansion in archaea. Ammonia-oxidising archaea of the phylum Thaumarchaeota are important organisms in the nitrogen cycle. Using 12 new genomes, this study finds evidence that Nitrososphaerales evolution was marked by lateral gene transfer followed by gene duplication.

The globally distributed ectoparasite Varroa destructor is a vector for viral pathogens of the Western honeybee (Apis mellifera), in particular the Iflavirus Deformed Wing Virus (DWV). In the absence of Varroa low levels DWV occur, generally causing asymptomatic infections. Conversely, Varroa-infested colonies show markedly elevated virus levels, increased overwintering colony losses, with impairment of pupal development and symptomatic workers. To determine whether changes in the virus population were due Varroa amplifying and introducing virulent virus strains and/or suppressing the host immune responses, we exposed Varroa-naïve larvae to oral and Varroa-transmitted DWV. We monitored virus levels and diversity in developing pupae and associated Varroa, the resulting RNAi response and transcriptome changes in the host. Exposed pupae were stratified by Varroa association (presence/absence) and virus levels (low/high) into three groups. Varroa-free pupae all exhibited low levels of a highly diverse DWV population, with those exposed per os (group NV) exhibiting changes in the population composition. Varroa-associated pupae exhibited either low levels of a diverse DWV population (group VL) or high levels of a near-clonal virulent variant of DWV (group VH). These groups and unexposed controls (C) could be also discriminated by principal component analysis of the transcriptome changes observed, which included several genes involved in development and the immune response. All Varroa tested contained a diverse replicating DWV population implying the virulent variant present in group VH, and predominating in RNA-seq analysis of temporally and geographically separate Varroa-infested colonies, was selected upon transmission from Varroa, a conclusion supported by direct injection of pupae in vitro with mixed virus populations. Identification of a virulent variant of DWV, the role of Varroa in its transmission and the resulting host transcriptome changes furthers our understanding of this important viral pathogen of honeybees.

Background With ever increasing accessibility to high throughput technologies, more complex treatment structures can be assessed in a variety of ’omics applications. This adds an extra layer of complexity to the analysis and interpretation, in particular when inferential univariate methods are applied en masse. It is well-known that mass univariate testing suffers from multiplicity issues and although this has been well documented for simple comparative tests, few approaches have focussed on more complex explanatory structures. Results Two frameworks are introduced incorporating corrections for multiplicity whilst maintaining appropriate structure in the explanatory variables. Within this paradigm, a choice has to be made as to whether multiplicity corrections should be applied to the saturated model, putting emphasis on controlling the rate of false positives, or to the predictive model, where emphasis is on model selection. This choice has implications for both the ranking and selection of the response variables identified as differentially expressed. The theoretical difference is demonstrated between the two approaches along with an empirical study of lipid composition in Arabidopsis under differing levels of salt stress. Conclusions Multiplicity corrections have an inherent weakness when the full explanatory structure is not properly incorporated. Although a unifying ‘single best’ recommendation is not provided, two reasonable alternatives are provided and the applicability of these approaches is discussed for different scenarios where the aims of analysis will differ. The key result is that the point at which multiplicity is incorporated into the analysis will fundamentally change the interpretation of the results, and the choice of approach should therefore be driven by the specific aims of the experiment.

The essential product of the Duchenne muscular dystrophy (DMD) gene is dystrophin 1 , a rod-like protein 2 that protects striated myocytes from contraction-induced injury 3 , 4 . Dystrophin-related protein (or utrophin) retains most of the structural and protein binding elements of dystrophin 5 . Importantly, normal thymic expression in DMD patients 6 should protect utrophin by central immunologic tolerance. We designed a codon-optimized, synthetic transgene encoding a miniaturized utrophin (µUtro), deliverable by adeno-associated virus (AAV) vectors. Here, we show that µUtro is a highly functional, non-immunogenic substitute for dystrophin, preventing the most deleterious histological and physiological aspects of muscular dystrophy in small and large animal models. Following systemic administration of an AAV-µUtro to neonatal dystrophin-deficient mdx mice, histological and biochemical markers of myonecrosis and regeneration are completely suppressed throughout growth to adult weight. In the dystrophin-deficient golden retriever model, µUtro non-toxically prevented myonecrosis, even in the most powerful muscles. In a stringent test of immunogenicity, focal expression of µUtro in the deletional- null German shorthaired pointer model produced no evidence of cell-mediated immunity, in contrast to the robust T cell response against similarly constructed µDystrophin (µDystro). These findings support a model in which utrophin-derived therapies might be used to treat clinical dystrophin deficiency, with a favorable immunologic profile and preserved function in the face of extreme miniaturization. A gene therapy vector expressing micro-utrophin provides functional replacement of lost dystrophin, and lacks the adverse immunogenicity associated with direct dystrophin therapy, in rodent and canine models of Duchenne muscular dystrophy.

The dosage of colistin for the treatment of enteric E. coli in animals necessitates considering the heteroresistant (HR) nature of the targeted inoculum, described by the presence of a major susceptible population (S1, representing 99.95% of total population) mixed with an initial minor subpopulation of less susceptible bacteria (S2). Herein, we report the 1-compartment population pharmacokinetics (PK) of colistin in chicken intestine (jejunum and ileum) and combined it with a previously established pharmacodynamic (PD) model of HR in E. coli. We then computed probabilities of target attainment (PTA) with a pharmacodynamic target (AUC 24h /MIC) that achieves 50% of the maximal kill of bacterial populations (considering inoculums of pure S1, S2 or HR mixture of S1 + S2). For an MIC of 1 mg/L, PTA > 95% was achieved with the registered dose (75,000 IU/kg BW/day in drinking water) for the HR mixture of S1 + S2 E. coli , whether they harboured mcr or not. For an MIC of 2 mg/L (ECOFF), we predicted PTA > 90% against the dominant susceptible sub-population (S1) with this clinical dose given (i) over 24 h for mcr -negative isolates or (ii) over 6 h for mcr -positive isolates (pulse dosing). Colistin clinical breakpoint S ≤ 2 mg/L (EUCAST rules) should be confirmed clinically.