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Innovative and rapid antimicrobial susceptibility testing systems

Antimicrobial resistance (AMR) is a major threat to human health worldwide, and the rapid detection and quantification of resistance, combined with antimicrobial stewardship, are key interventions to combat the spread and emergence of AMR. Antimicrobial susceptibility testing (AST) systems are the collective set of diagnostic processes that facilitate the phenotypic and genotypic assessment of AMR and antibiotic susceptibility. Over the past 30 years, only a few high-throughput AST methods have been developed and widely implemented. By contrast, several studies have established proof of principle for various innovative AST methods, including both molecular-based and genome-based methods, which await clinical trials and regulatory review. In this Review, we discuss the current state of AST systems in the broadest technical, translational and implementation-related scope

A Selenium-Dependent Xanthine Dehydrogenase Triggers Biofilm Proliferation in Enterococcus faecalis through Oxidant Production

Selenium has been shown to be present as a labile cofactor in a small class of molybdenum hydroxylase enzymes in several species of clostridia that specialize in the fermentation of purines and pyrimidines. This labile cofactor is poorly understood, yet recent bioinformatic studies have suggested that Enterococcus faecalis could serve as a model system to better understand the way in which this enzyme cofactor is built and the role of these metalloenzymes in the physiology of the organism. An mRNA that encodes a predicted selenium-dependent molybdenum hydroxylase (SDMH) has also been shown to be specifically increased during the transition from planktonic growth to biofilm growth. Based on these studies, we examined whether this organism produces an SDMH and probed whether selenoproteins may play a role in biofilm physiology. We observed a substantial increase in biofilm density upon the addition of uric acid to cells grown in a defined culture medium, but only when molybdate (Mo) and selenite (Se) were also added. We also observed a significant increase in biofilm density in cells cultured in tryptic soy broth with 1% glucose (TSBG) when selenite was added. In-frame deletion of selD, which encodes selenophosphate synthetase, also blocked biofilm formation that occurred upon addition of selenium. Moreover, mutation in the gene encoding the molybdoenzyme (xdh) prevented the induction of biofilm proliferation upon supplementation with selenium. Tungstate or auranofin addition also blocked this enhanced biofilm density, likely through inhibition of molybdenum or selenium cofactor synthesis. A large protein complex labeled with Se-75 is present in higher concentrations in biofilms than in planktonic cells, and the same complex is formed in TSBG. Xanthine dehydrogenase activity correlates with the presence of this labile selenoprotein complex and is absent in a selD or an xdh mutant. Enhanced biofilm density correlates strongly with higher levels of extracellular peroxide, which is produced upon the addition of selenite to TSBG. Peroxide levels are not increased in either the selD or the xdh mutant upon addition of selenite. Extracellular superoxide production, a phenomenon well established to be linked to clinical isolates, is abolished in both mutant strains. Taken together, these data provide evidence that an SDMH is involved in biofilm formation in Enterococcus faecalis, contributing to oxidant production either directly or alternatively through its involvement in redox-dependent processes linked to oxidant production

Mid-infrared interferometry with high spectral resolution

The Infrared Spatial Interferometer (ISI) is a three telescope interferometer system that operates near 11 microns wavelength using heterodyne detection with CO2 lasers as local oscillators. Stellar measurements have been made using consistent instrumentation for 20 years, allowing comparisons of stellar sizes of red giant and Mira stars over time intervals which are long in comparison to stellar luminosity periods. Recent visibility and closure phase measurements of the star Betelgeuse have been fitted to simple image models and these results have been added to the 17 year record of stellar observations. A new area of investigation of stellar properties at very high spectral resolution will begin in the 2010-2011 observing season. The design of a new digital spectrometer-correlator system is discussed. This system will obtain visibility measurements on-and-off individual spectral lines and the continuum, simultaneously

Observations of Late-Type Stars with the Infrared Spatial Interferometer

The Infrared Spatial Interferometer (ISI) has been conducting mid-IR measurements of red supergiant and AGB stars for about 20 years. This paper reviews the ISI system and results with an emphasis on measurements of changes in stellar sizes and shapes, and those of the surrounding dust shells, over time scales of weeks to decades. The long-term measurement record provides important new observables for stellar theory. A new spectrometer-correlator system is discussed where this system will measure visibilities on-and-off individual molecular spectral lines

Loss of Biological Diversity: A Global Crisis Requiring International Solutions: A Report to the National Science Board

Executive Summary Biological diversity refers to the variety and variability among living organisms and the ecological complexes in which they occur. Diversity can be defined as the number of different items and their relative frequency. For biological diversity, these items are organized at many levels, ranging from complete ecosystems to the chemical structures that are the molecular basis of heredity. Thus, the term encompasses different ecosystems, species, genes, and their relative abundance (OTA, 1987). There is an ongoing, unprecedented loss of the variety as well as absolute numbers of organisms-from the smallest microorganism to the largest and most spectacular of mammals. Loss of tropical moist forests, which contain over half the total species of organisms, has been well documented by scientists and is now widely reported in the media. Many other ecosystems are also threatened; as human populations and their support systems expand, natural ecosystems at all latitudes are altered or converted. At its meeting on October 15, 1987, the National Science Board concluded that the world\u27s decreasing biological diversity is a critical scientific issue requiring immediate attention. The National Science Board\u27s Committee on International Science was asked to study the scientific and international aspects of the decline of biological diversity and to recommend a course of action. This report describes what the National Science Foundation (NSF) can do to influence the U.S. science and education base, articulates where international scientific cooperation is needed, and suggests roles for other agencies and organizations (both national and international) which have scientific, educational, and management responsibilities. The current disappearance of biota has several causes: the destruction or degradation of entire ecosystems; the accelerating loss of individual species from communities or ecosystems as a result of human disturb;mce; and the loss of genetically distinct parts of populations due to human-induced selective pressures. Although not all parts of the planet are equally affected, the problem is global, and human activities are the primary cause. The loss of biological diversity is important because human existence depends on the biological resources of 1 the earth. Human prosperity is based very largely on the ability to utilize biological diversity: to take advantage of the properties of plants, animals, fungi, and microorganisms for food, clothing, medicine, and shelter. Scientific knowledge about the earth\u27s biological diversity has huge gaps. This lack of information hampers society\u27s ability either to estimate the magnitude of the problem or to prevent further losses. It is impossible to identify all the biological resources at risk, since there is no complete inventory of all the life forms on earth. Approximately 1.4 million species have been given scientific names, but estimates of actual numbers range from 5 million to 80 million species. Although knowledge of some taxa is extensive, the vast majority of groups are largely unknown. The current wave of extinction is destroying both known biotic resources and those still undiscovered. As is proving to be the case with most environmental problems, neither the loss of biological diversity nor its solution is the exclusive province of any one nation. International cooperation is necessary to develop both scientific knowledge and successful mitigation and management strategies. The root causes of the problem include sociological and economic processes which operate on an global scale; a thorough understanding will require investigation and elucidation of both biological and non-biological components. There are several reasons for increasing National Science Foundation (NSF) involvement in biodiversity studies: the economic and social importance of biodiversity (and the risk of opportunity lost due to accelerating extinction); the contributions such leadership can make toward to conservation of biological diversity; the important role of such studies in the international growth of science, especially in tropical countries; the potential impact of such studies on the future course of biology as a whole; and enhancing public awareness of the issues. NSF should assume a scientific leadership position with respect to agencies in the U.S. and throughout the world. By insisting on the central importance of biodiversity, the NSF could encourage collaborative support for the actions recommended below. 1. The Committee believes that the role of the NSF is clear-NSF should, as a matter of National Science Board Policy, provide leadership to undertake the inventory of the world\u27s biodiversity. 2. The scientific basis for conservation biology, restoration ecology, and environmental management must be strengthened. 3. Educational and public awareness programs related to biodiversity need increased support. 4. The economic and social aspects of the biodiversity crisis need additional study. 5. Enhance support for developing country scientists and institutions for biodiversity research and conservation

A distant trophoblast-specific enhancer controls HLA-G expression at the maternal–fetal interface

HLA-G, a nonclassical HLA molecule uniquely expressed in the placenta, is a central component of fetus-induced immune tolerance during pregnancy. The tissue-specific expression of HLA-G, however, remains poorly understood. Here, systematic interrogation of the HLA-G locus using massively parallel reporter assay (MPRA) uncovered a previously unidentified cis-regulatory element 12 kb upstream of HLA-G with enhancer activity, Enhancer L. Strikingly, clustered regularly-interspaced short palindromic repeats (CRISPR)/Cas9-mediated deletion of this enhancer resulted in ablation of HLA-G expression in JEG3 cells and in primary human trophoblasts isolated from placenta. RNA-seq analysis demonstrated that Enhancer L specifically controls HLA-G expression. Moreover, DNase-seq and chromatin conformation capture (3C) defined Enhancer L as a cell type-specific enhancer that loops into the HLA-G promoter. Interestingly, MPRA-based saturation mutagenesis of Enhancer L identified motifs for transcription factors of the CEBP and GATA families essential for placentation. These factors associate with Enhancer L and regulate HLA-G expression. Our findings identify long-range chromatin looping mediated by core trophoblast transcription factors as the mechanism controlling tissue-specific HLA-G expression at the maternal–fetal interface. More broadly, these results establish the combination of MPRA and CRISPR/Cas9 deletion as a powerful strategy to investigate human immune gene regulation

The Firre locus produces a trans -acting RNA molecule that functions in hematopoiesis

Abstract: RNA has been classically known to play central roles in biology, including maintaining telomeres, protein synthesis, and in sex chromosome compensation. While thousands of long noncoding RNAs (lncRNAs) have been identified, attributing RNA-based roles to lncRNA loci requires assessing whether phenotype(s) could be due to DNA regulatory elements, transcription, or the lncRNA. Here, we use the conserved X chromosome lncRNA locus Firre, as a model to discriminate between DNA- and RNA-mediated effects in vivo. We demonstrate that (i) Firre mutant mice have cell-specific hematopoietic phenotypes, and (ii) upon exposure to lipopolysaccharide, mice overexpressing Firre exhibit increased levels of pro-inflammatory cytokines and impaired survival. (iii) Deletion of Firre does not result in changes in local gene expression, but rather in changes on autosomes that can be rescued by expression of transgenic Firre RNA. Together, our results provide genetic evidence that the Firre locus produces a trans-acting lncRNA that has physiological roles in hematopoiesis

Genome-wide Association Meta-analysis of Childhood and Adolescent Internalizing Symptoms

Objective: To investigate the genetic architecture of internalizing symptoms in childhood and adolescence. Method: In 22 cohorts, multiple univariate genome-wide association studies (GWASs) were performed using repeated assessments of internalizing symptoms, in a total of 64,561 children and adolescents between 3 and 18 years of age. Results were aggregated in meta-analyses that accounted for sample overlap, first using all available data, and then using subsets of measurements grouped by rater, age, and instrument. Results: The meta-analysis of overall internalizing symptoms (INToverall) detected no genome-wide significant hits and showed low single nucleotide polymorphism (SNP) heritability (1.66%, 95% CI = 0.84-2.48%, n(effective) = 132,260). Stratified analyses indicated rater-based heterogeneity in genetic effects, with self-reported internalizing symptoms showing the highest heritability (5.63%, 95% CI = 3.08%-8.18%). The contribution of additive genetic effects on internalizing symptoms appeared to be stable over age, with overlapping estimates of SNP heritability from early childhood to adolescence. Genetic correlations were observed with adult anxiety, depression, and the well-being spectrum (vertical bar r(g)vertical bar > 0.70), as well as with insomnia, loneliness, attention-deficit/hyperactivity disorder, autism, and childhood aggression (range vertical bar r(g)vertical bar = 0.42-0.60), whereas there were no robust associations with schizophrenia, bipolar disorder, obsessive-compulsive disorder, or anorexia nervosa. Conclusion: Genetic correlations indicate that childhood and adolescent internalizing symptoms share substantial genetic vulnerabilities with adult internalizing disorders and other childhood psychiatric traits, which could partially explain both the persistence of internalizing symptoms over time and the high comorbidity among childhood psychiatric traits. Reducing phenotypic heterogeneity in childhood samples will be key in paving the way to future GWAS success.Peer reviewe