Identification of the CRP regulon using in vitro and in vivo transcriptional profiling

The Escherichia coli cyclic AMP receptor protein (CRP) is a global regulator that controls transcription initiation from more than 100 promoters by binding to a specific DNA sequence within cognate promoters. Many genes in the CRP regulon have been predicted simply based on the presence of DNA-binding sites within gene promoters. In this study, we have exploited a newly developed technique, run-off transcription/microarray analysis (ROMA) to define CRP-regulated promoters. Using ROMA, we identified 176 operons that were activated by CRP in vitro and 16 operons that were repressed. Using positive control mutants in different regions of CRP, we were able to classify the different promoters into class I or class II/III. A total of 104 operons were predicted to contain Class II CRP-binding sites. Sequence analysis of the operons that were repressed by CRP revealed different mechanisms for CRP inhibition. In contrast, the in vivo transcriptional profiles failed to identify most CRP-dependent regulation because of the complexity of the regulatory network. Analysis of these operons supports the hypothesis that CRP is not only a regulator of genes required for catabolism of sugars other than glucose, but also regulates the expression of a large number of other genes in E.coli. ROMA has revealed 152 hitherto unknown CRP regulons

Hybrid assembly of an agricultural slurry virome reveals a diverse and stable community with the potential to alter the metabolism and virulence of veterinary pathogens

Background: Viruses are the most abundant biological entities on Earth, known to be crucial components of microbial ecosystems. However, there is little information on the viral community within agricultural waste. There are currently ~ 2.7 million dairy cattle in the UK producing 7–8% of their own bodyweight in manure daily, and 28 million tonnes annually. To avoid pollution of UK freshwaters, manure must be stored and spread in accordance with guidelines set by DEFRA. Manures are used as fertiliser, and widely spread over crop fields, yet little is known about their microbial composition. We analysed the virome of agricultural slurry over a 5-month period using short and long-read sequencing. Results: Hybrid sequencing uncovered more high-quality viral genomes than long or short-reads alone; yielding 7682 vOTUs, 174 of which were complete viral genomes. The slurry virome was highly diverse and dominated by lytic bacteriophage, the majority of which represent novel genera (~ 98%). Despite constant influx and efflux of slurry, the composition and diversity of the slurry virome was extremely stable over time, with 55% of vOTUs detected in all samples over a 5-month period. Functional annotation revealed a diverse and abundant range of auxiliary metabolic genes and novel features present in the community, including the agriculturally relevant virulence factor VapE, which was widely distributed across different phage genera that were predicted to infect several hosts. Furthermore, we identified an abundance of phage-encoded diversity-generating retroelements, which were previously thought to be rare on lytic viral genomes. Additionally, we identified a group of crAssphages, including lineages that were previously thought only to be found in the human gut. Conclusions: The cattle slurry virome is complex, diverse and dominated by novel genera, many of which are not recovered using long or short-reads alone. Phages were found to encode a wide range of AMGs that are not constrained to particular groups or predicted hosts, including virulence determinants and putative ARGs. The application of agricultural slurry to land may therefore be a driver of bacterial virulence and antimicrobial resistance in the environment. [MediaObject not available: see fulltext.

Complete Genome Sequence and Comparative Metabolic Profiling of the Prototypical Enteroaggregative Escherichia coli Strain 042

Background \ud Escherichia coli can experience a multifaceted life, in some cases acting as a commensal while in other cases causing intestinal and/or extraintestinal disease. Several studies suggest enteroaggregative E. coli are the predominant cause of E. coli-mediated diarrhea in the developed world and are second only to Campylobacter sp. as a cause of bacterial-mediated diarrhea. Furthermore, enteroaggregative E. coli are a predominant cause of persistent diarrhea in the developing world where infection has been associated with malnourishment and growth retardation. \ud \ud Methods \ud In this study we determined the complete genomic sequence of E. coli 042, the prototypical member of the enteroaggregative E. coli, which has been shown to cause disease in volunteer studies. We performed genomic and phylogenetic comparisons with other E. coli strains revealing previously uncharacterised virulence factors including a variety of secreted proteins and a capsular polysaccharide biosynthetic locus. In addition, by using Biolog™ Phenotype Microarrays we have provided a full metabolic profiling of E. coli 042 and the non-pathogenic lab strain E. coli K-12. We have highlighted the genetic basis for many of the metabolic differences between E. coli 042 and E. coli K-12. \ud \ud Conclusion \ud This study provides a genetic context for the vast amount of experimental and epidemiological data published thus far and provides a template for future diagnostic and intervention strategies

Impact of contrasting poultry exposures on human, poultry, and wastewater antibiotic resistomes in Bangladesh

Interactions between humans, animals, and the environment are considered critical foci for addressing antimicrobial resistance (AMR). However, One Health data on AMR in low- and middle-income countries are presently scarce. Using metagenomics, we investigated whether and how the fecal antibiotic resistomes of humans are influenced by exposure to intensive and non-intensively reared poultry within contrasting settings of urban wet markets (n = 13) and rural households (n = 7) in Bangladesh. We also considered poultry (n = 10) and wastewater (n = 10) resistomes in these settings. We found that occupational poultry exposures did not significantly alter the human fecal resistome. In contrast, macrolide-lincosamide-streptogramin and streptothricin antibiotic resistance genes (ARGs) were enriched in poultry from urban wet markets relative to rural household chickens. Wastewater had the highest ARG richness, though this was only partially explained by poultry cecal and human fecal sources. Wastewater also contained clinically significant carbapenem ARGs. This study therefore provides critical insight into the distribution of ARGs in Bangladesh

DNA binding shifts the redox potential of the transcription factor SoxR

Electrochemistry measurements on DNA-modified electrodes are used to probe the effects of binding to DNA on the redox potential of SoxR, a transcription factor that contains a [2Fe-2S] cluster and is activated through oxidation. A DNA-bound potential of +200 mV versus NHE (normal hydrogen electrode) is found for SoxR isolated from Escherichia coli and Pseudomonas aeruginosa. This potential value corresponds to a dramatic shift of +490 mV versus values found in the absence of DNA. Using Redmond red as a covalently bound redox reporter affixed above the SoxR binding site, we also see, associated with SoxR binding, an attenuation in the Redmond red signal compared with that for Redmond red attached below the SoxR binding site. This observation is consistent with a SoxR-binding-induced structural distortion in the DNA base stack that inhibits DNA-mediated charge transport to the Redmond red probe. The dramatic shift in potential for DNA-bound SoxR compared with the free form is thus reconciled based on a high-energy conformational change in the SoxR–DNA complex. The substantial positive shift in potential for DNA-bound SoxR furthermore indicates that, in the reducing intracellular environment, DNA-bound SoxR is primarily in the reduced form; the activation of DNA-bound SoxR would then be limited to strong oxidants, making SoxR an effective sensor for oxidative stress. These results more generally underscore the importance of using DNA electrochemistry to determine DNA-bound potentials for redox-sensitive transcription factors because such binding can dramatically affect this key protein property

Antimicrobial resistance in dairy slurry tanks: A critical point for measurement and control

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Shifting Home Energy Consumption Through a Holistic Understanding of the Home System of Practice

This chapter describes the concept of the home system of practice, which can be used to provide a deeper understanding of the social system of the home and inform long-term solutions for enabling domestic energy reduction. More traditional methods have attempted to persuade occupants to change behaviour through the use of information campaigns and feedback technology. However, these interventions are usually short lived as they ignore the underlying reasons for practices to occur. A more effective solution is through practice-oriented design, which co-creates innovative technology with the user. In addition, the emergence and use of automated technology enables practices to act independently of the user. Yet, the success of automation is also reliant on an understanding of the home system of practice, occupant needs and skills

Does proactive personality matter in leadership transitions? Effects of proactive personality on new leader identification and responses to new leaders and their change agendas

Despite the growing frequency of leadership transitions and their significant impact on team and organizational performance, little research has examined why and how teams develop an identification with a new leader or their subsequent receptiveness to the new leader’s change initiatives. Drawing from the contrast and congruence effects and the theoretical perspectives of leader identification, this study empirically tests a model in which the congruence of new leaders’ and their teams’ proactive personalities foster new leader identification, as well as the team’s behavioral responses to the new leader’s change agenda. This effect is strongest when the new leader’s proactive personality is higher than that of the former leader’s proactive personality (positive contrast). Our findings of a four-wave “before-and-after” transition survey of 155 hotel employees and 51 new leaders, achieved through polynomial regression analyses, proved very insightful. Essentially, we found that the congruence between a new leader’s and his/her team’s proactive personalities and the positive contrast between a former leader’s and the new leader’s proactive personalities enhanced new leader identification and the team’s shared identification with the new leader’s change agenda, and, thereby led the team to exhibit more behavioral engagement with, and voice behavior about, the new leader’s change agenda

Intelligence, reason of state and the art of governing risk and opportunity in early modern Europe

Drawing upon primary and secondary historical material, this paper explores the role of intelligence in early modern government. It focuses upon developments in seventeenth- and early-eighteenth-century England, a site-specific genealogical moment in the broader history of state power/knowledges. Addressing a tendency in Foucauldian work to neglect pre-eighteenth-century governance, the analysis reveals a set of interrelated processes which gave rise to an innovative technique for anticipating hazard and opportunity for the state. At the intersection of raison d’État, the evolving art of government, widespread routines of secrecy and a post-Westphalia field of European competition and exchange, intelligence was imagined as a fundamental solution to the concurrent problems of ensuring peace and stability while improving state forces. In the administrative offices of the English Secretary of State, an assemblage of complex and interrelated procedures sought to produce and manipulate information in ways which exposed both possible risks to the state and potential opportunities for expansion and gain. As this suggests, the art of intelligence played an important if largely unacknowledged role in the formation and growth of the early modern state. Ensuring strategic advantage over rivals, intelligence also limited the ability of England's neighbours to dominate trade, control the seas and master the colonies, functioning as a constitutive feature of European balance and equilibrium. As the analysis concludes, understanding intelligence as a form of governmental technique – a way of doing something – reveals an entirely novel way of thinking about and investigating its myriad (historical and contemporary) formations