Multiplexed enrichment and genomic profiling of peripheral blood cells reveal subset-specific immune signatures

Specialized immune cell subsets are involved in autoimmune disease, cancer immunity, and infectious disease through a diverse range of functions mediated by overlapping pathways and signals. However, subset-specific responses may not be detectable in analyses of whole blood samples, and no efficient approach for profiling cell subsets at high throughput from small samples is available. We present a low-input microfluidic system for sorting immune cells into subsets and profiling their gene expression. We validate the system’s technical performance against standard subset isolation and library construction protocols and demonstrate the importance of subset-specific profiling through in vitro stimulation experiments. We show the ability of this integrated platform to identify subset-specific disease signatures by profiling four immune cell subsets in blood from patients with systemic lupus erythematosus (SLE) and matched control subjects. The platform has the potential to make multiplexed subset-specific analysis routine in many research laboratories and clinical settings.National Institute of Allergy and Infectious Diseases (U.S.) (Grant U24 AI118668

Digital MDA for enumeration of total nucleic acid contamination

Multiple displacement amplification (MDA) is an isothermal, sequence-independent method for the amplification of high molecular weight DNA that is driven by ϕ29 DNA polymerase (DNAP). Here we report digital MDA (dMDA), an ultrasensitive method for quantifying nucleic acid fragments of unknown sequence. We use the new assay to show that our custom ϕ29 DNAP preparation is free of contamination at the limit of detection of the dMDA assay (1 contaminating molecule per assay microliter). Contamination in commercially available preparations is also investigated. The results of the dMDA assay provide strong evidence that the so-called ‘template-independent’ MDA background can be attributed to high-molecular weight contaminants and is not primer-derived in the commercial kits tested. dMDA is orders of magnitude more sensitive than PCR-based techniques for detection of microbial genomic DNA fragments and opens up new possibilities for the ultrasensitive quantification of DNA fragments in a wide variety of application areas using MDA chemistry and off-the-shelf hardware developed for digital PCR

Single-cell and metagenomic analyses indicate a fermentative and saccharolytic lifestyle for members of the OP9 lineage

OP9 is a yet-uncultivated bacterial lineage found in geothermal systems, petroleum reservoirs, anaerobic digesters and wastewater treatment facilities. Here we use single-cell and metagenome sequencing to obtain two distinct, nearly complete OP9 genomes, one constructed from single cells sorted from hot spring sediments and the other derived from binned metagenomic contigs from an in situ-enriched cellulolytic, thermophilic community. Phylogenomic analyses support the designation of OP9 as a candidate phylum for which we propose the name ‘Atribacteria’. Although a plurality of predicted proteins is most similar to those from Firmicutes, the presence of key genes suggests a diderm cell envelope. Metabolic reconstruction from the core genome suggests an anaerobic lifestyle based on sugar fermentation by Embden–Meyerhof glycolysis with production of hydrogen, acetate and ethanol. Putative glycohydrolases and an endoglucanase may enable catabolism of (hemi)cellulose in thermal environments. This study lays a foundation for understanding the physiology and ecological role of the ‘Atribacteria’.United States. National Aeronautics and Space Administration (Exobiology Grant EXO-NNX11AR78G)National Science Foundation (U.S.) (Grant MCB 0546865)National Science Foundation (U.S.) (Grant OISE 0968421)United States. Dept. of Energy (Grant DE-EE-0000716)Nevada Renewable Energy ConsortiumUnited States. Dept. of Energy. Office of Science. Joint Genome Institute (Contract DE-AC02-05CH11231

Profesionalna astma u radnika izloženih prašinama iz biljnih i voćnih čajeva

We performed a cross-sectional study to detect occupational asthma (OA) in 63 subjects occupationally exposed to herbal and fruit tea dust and in 63 corresponding controls. The evaluation included a questionnaire, skin prick tests to workplace and common inhalant allergens, spirometry, and histamine challenge test. The evaluation of the work-relatedness of asthma in the exposed workers was based on serial peak expiratory flow rate (PEFR) measurements and bronchoprovocation tests. We found a higher prevalence of respiratory symptoms in the exposed workers, whereas spirometric parameters were significantly lower. The prevalence of sensitisation to allergens and of bronchial hyperresponsivenss (BHR) did not differ significantly between the groups. The prevalence of asthma was also similar in both groups (8.0 % vs. 6.4 %; P=0.540). Work-relatedness of symptoms was reported by all asthmatic tea workers and by no control with asthma. Significant work-related changes in PEFR diurnal variations and in non-specific BHR, suggesting allergic OA, were found in one tea worker with asthma (1.6 %). No specific workplace agent causing OA in the affected subject was identified. None of the tea workers with asthma met the criteria for medical case definition of the reactive airway dysfunction syndrome (RADS). Our data confirm workplace exposure to herbal and fruit tea dust as a risk factor for OA.Svrha je ovoga presječnog ispitivanja bila otkriti profesionalnu astmu u skupini od 63 ispitanika koji su na radnome mjestu bili izloženi prašinama biljnih i voćnih čajeva. Kao kontrola uzet je jednak broj uredskih radnika koji nisu bili izloženi ovim prašinama. Ocjena izloženih i kontrolnih ispitanika obuhvatila je upitnik, skin prick testove na uobičajene i profesionalne inhalacijske alergene, spirometriju te histaminski test. Povezanost astme s profesionalnom izloženosti u radnika utvrđena je prema kriterijima Američkog kolegija pulmologa (American College of Chest Physicians, krat. ACCP), a na temelju mjerenja niza vršnih ekspiratornih protoka (engl. peak expiratory flow rate, PEFR) i niza bronhoprovokativnih testova. Izloženi su radnici iskazali veću prevalenciju respiratornih simptoma odnosno niže spirometrijske vrijednosti od kontrole. Izloženi ispitanici nisu se značajno razlikovali od kontrole u prevalenciji senzibilizacije na profesionalne i uobičajene inhalacijske alergene te prevalenciji pretjerane bronhalne reaktivnosti (engl. bronchial hyperresponsiveness, krat. BHR). Isto vrijedi i za prevalenciju astme (8,0 % u izloženih radnika prema 6,4 % u kontrola; P=0,540). Povezanost simptoma s poslom prijavili su svi radnici u obradi čaja oboljeli od astme te ni jedan kontrolni ispitanik s astmom. U jednoga astmatičnog radnika na čaju utvrđene su značajne promjene u dnevnim varijacijama PEFR-a te u nespecifičnom BHR-u koji upućuju na profesionalnu astmu (1.6 %). Nije utvrđeno koja je to tvar uzrokovala profesionalnu astmu u ovog ispitanika. Nitko od izloženih radnika s astmom nije zadovoljio sve medicinske kriterije za dijagnozu sindroma reaktivne disfunkcije dišnih putova (engl. reactive airway dysfunction syndrome, RADS). Naši podaci potvrđuju da je profesionalna izloženost prašinama iz biljnih i voćnih čajeva čimbenik rizika od profesionalne astme

Searching for DNA Lesions: Structural Evidence for Lower- and Higher-Affinity DNA Binding Conformations of Human Alkyladenine DNA Glycosylase

To efficiently repair DNA, human alkyladenine DNA glycosylase (AAG) must search the million-fold excess of unmodified DNA bases to find a handful of DNA lesions. Such a search can be facilitated by the ability of glycosylases, like AAG, to interact with DNA using two affinities: a lower-affinity interaction in a searching process and a higher-affinity interaction for catalytic repair. Here, we present crystal structures of AAG trapped in two DNA-bound states. The lower-affinity depiction allows us to investigate, for the first time, the conformation of this protein in the absence of a tightly bound DNA adduct. We find that active site residues of AAG involved in binding lesion bases are in a disordered state. Furthermore, two loops that contribute significantly to the positive electrostatic surface of AAG are disordered. Additionally, a higher-affinity state of AAG captured here provides a fortuitous snapshot of how this enzyme interacts with a DNA adduct that resembles a one-base loop.National Institutes of Health (U.S.) (grant no. P30-ES002109)National Institutes of Health (U.S.) (grant no. GM65337)National Institutes of Health (U.S.) (grant no. GM65337-03S2)National Institutes of Health (U.S.) (grant no. CA055042)National Institutes of Health (U.S.) (grant no. CA092584)Repligen Corporation (KIICR Graduate Fellowship

Uplifting manhood to wonderful heights? News coverage of the human costs of military conflict from world war I to Gulf war Two

Domestic political support is an important factor constraining the use of American military power around the world. Although the dynamics of war support are thought to reflect a cost-benefit calculus, with costs represented by numbers of friendly war deaths, no previous study has examined how information about friendly, enemy, and civilian casualties is routinely presented to domestic audiences. This paper establishes a baseline measure of historical casualty reporting by examining New York Times coverage of five major wars that occurred over the past century. Despite important between-war differences in the scale of casualties, the use of conscription, the type of warfare, and the use of censorship, the frequency of casualty reporting and the framing of casualty reports has remained fairly consistent over the past 100 years. Casualties are rarely mentioned in American war coverage. When casualties are reported, it is often in ways that minimize or downplay the human costs of war

Different battlegrounds, similar concerns? The “history wars” and the teaching of history in Australia and England

Debates about the purpose and content of history education in schools have been prevalent in most Westernised democratic nations over the last thirty years. At expense are essential questions concerning national identity/ies, competing narratives and the aims of history education. The impact of “history wars” have been felt within both Australia and England, as conservative commentators – including politicians and historians – have raised concerns about the depth and effectiveness of history education and have sought to make significant changes to the history curriculum for schools. This analysis examines the history wars in Australia and England, exploring the view that history education has been in danger and/or crisis and examining the curricular implications of a move toward greater recognition of national narratives. It raises some essential tensions that remain regarding two aspects of history teaching in both nations – (i) historiography and (ii) chronological understanding

Different paths to the modern state in Europe: the interaction between domestic political economy and interstate competition

Theoretical work on state formation and capacity has focused mostly on early modern Europe and on the experience of western European states during this period. While a number of European states monopolized domestic tax collection and achieved gains in state capacity during the early modern era, for others revenues stagnated or even declined, and these variations motivated alternative hypotheses for determinants of fiscal and state capacity. In this study we test the basic hypotheses in the existing literature making use of the large date set we have compiled for all of the leading states across the continent. We find strong empirical support for two prevailing threads in the literature, arguing respectively that interstate wars and changes in economic structure towards an urbanized economy had positive fiscal impact. Regarding the main point of contention in the theoretical literature, whether it was representative or authoritarian political regimes that facilitated the gains in fiscal capacity, we do not find conclusive evidence that one performed better than the other. Instead, the empirical evidence we have gathered lends supports to the hypothesis that when under pressure of war, the fiscal performance of representative regimes was better in the more urbanized-commercial economies and the fiscal performance of authoritarian regimes was better in rural-agrarian economie

Minority youth, crime, conflict, and belonging in Australia

In recent decades, the size and diversity of the minority population of contemporary western societies has increased significantly. To the critics of immigration, minority youth have been increasingly linked to crime, criminal gangs, anti-social behaviour, and riots. In this article, we draw on fieldwork conducted in Sydney, Australia's largest and most ethnically diverse city, to probe aspects of the criminality, anti-social behaviour, national identity, and belonging of ethnic minority youth in Australia. We conclude that the evidence on minority youth criminality is weak and that the panic about immigrant youth crime and immigrant youth gangs is disproportionate to the reality, drawing on and in turn creating racist stereotypes, particularly with youth of 'Middle Eastern appearance'. A review of the events leading up to the Sydney Cronulla Beach riots of December 2005 suggests that the underlying cause of the riots were many years of international, national, and local anti-Arab, anti-Muslim media discourse, and political opportunism, embedded in changing but persistent racist attitudes and practises. Our argument is that such inter-ethnic conflict between minority and majority youth in Sydney is the exception, not the rule. Finally, we draw on a hitherto unpublished survey of youth in Sydney to explore issues of national identity and belonging among young people of diverse ethnic and religious background. We conclude that minority youth in Sydney do not live 'parallel lives' but contradictory, inter-connected cosmopolitan lives. They are connected to family and local place, have inter-ethnic friendships but are often disconnected to the nation and the flag. © 2009 Springer Science+Business Media B.V