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ABSTRACTThe central dogma of molecular biology is a key concept for undergraduate students in the life sciences as it describes the flow of information in living systems from gene-to-gene product. However, despite often being covered in many introductory life science courses, students may still have misconceptions surrounding the central dogma even as they move on to advanced courses. Active learning strategies such as laboratory activities can be useful in addressing such misconceptions. In the laboratory exercise presented here, senior undergraduate students explore the intricacies of nonsense suppressor mutations to challenge their understanding of the central dogma. The students introduce a plasmid carrying a nonfunctional chromogenic protein gene due to a nonsense mutation in a codon encoding the chromophore to various nonsense suppressor strains of Escherichia coli. Students then observe distinct chromogenic phenotypes, depending on the suppressor strain. Students showed a moderate increase in understanding of the central dogma. While the central dogma remains a challenging concept, active learning strategies like the one presented here can help reduce conceptual errors.

The Ensembl project (https://www.ensembl.org) makes key genomic data sets available to the entire scientific community without restrictions. Ensembl seeks to be a fundamental resource driving scientific progress by creating, maintaining and updating reference genome annotation and comparative genomics resources. This year we describe our new and expanded gene, variant and comparative annotation capabilities, which led to a 50% increase in the number of vertebrate genomes we support. We have also doubled the number of available human variants and added regulatory regions for many mouse cell types and developmental stages. Our data sets and tools are available via the Ensembl website as well as a through a RESTful webservice, Perl application programming interface and as data files for download.

Objectives: Clostridium difficile infection (CDI) is the most common cause of healthcare-associated infections in the United States. Despite well-established risk factors, little research has focused on use of these variables to identify a patient population at high risk for CDI to target with primary prevention strategies. A predictive index for healthcare-associated CDI could improve clinical care and guide research for primary prevention trials. Our objective was to develop a predictive index to identify patients at high risk for healthcare-associated CDI., Methods: We performed a secondary database analysis in a five-hospital health system in Houston, Texas. Our cohort consisted of 97 130 hospitalized patients admitted for more than 48 hours between October 2014 and September 2016. The derivation cohort consisted of the initial 80% of admissions (75 545 patients), with the remainder being used in the validation cohort., Results: CDI rates in the derivation and validation cohorts were 1.55% and 1.43%, respectively. Thirty-day predictors of CDI were increased number of high-risk antibiotics, Charlson comorbidity index score, age and receipt of a proton pump inhibitor. These variables were incorporated into a simple risk index with a score range of 0 to 10. The final model demonstrated good discrimination and calibration with the observed CDI incidence ranging from 0.1% to 20.4%., Conclusions: We developed a predictive index for 30-day risk of healthcare-associated CDI using readily available and clinically useful variables. This simple predictive risk index may be used to improve clinical decision making and resource allocation for CDI stewardship initiatives, and guide research design., (Copyright © 2018 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All rights reserved.)

The Ensembl project has been aggregating, processing, integrating and redistributing genomic datasets since the initial releases of the draft human genome, with the aim of accelerating genomics research through rapid open distribution of public data. Large amounts of raw data are thus transformed into knowledge, which is made available via a multitude of channels, in particular our browser (http://www.ensembl.org). Over time, we have expanded in multiple directions. First, our resources describe multiple fields of genomics, in particular gene annotation, comparative genomics, genetics and epigenomics. Second, we cover a growing number of genome assemblies; Ensembl Release 90 contains exactly 100. Third, our databases feed simultaneously into an array of services designed around different use cases, ranging from quick browsing to genome-wide bioinformatic analysis. We present here the latest developments of the Ensembl project, with a focus on managing an increasing number of assemblies, supporting efforts in genome interpretation and improving our browser., (© The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Ensembl Genomes (http://www.ensemblgenomes.org) is an integrating resource for genome-scale data from non-vertebrate species, complementing the resources for vertebrate genomics developed in the Ensembl project (http://www.ensembl.org). Together, the two resources provide a consistent set of programmatic and interactive interfaces to a rich range of data including genome sequence, gene models, transcript sequence, genetic variation, and comparative analysis. This paper provides an update to the previous publications about the resource, with a focus on recent developments and expansions. These include the incorporation of almost 20 000 additional genome sequences and over 35 000 tracks of RNA-Seq data, which have been aligned to genomic sequence and made available for visualization. Other advances since 2015 include the release of the database in Resource Description Framework (RDF) format, a large increase in community-derived curation, a new high-performance protein sequence search, additional cross-references, improved annotation of non-protein-coding genes, and the launch of pre-release and archival sites. Collectively, these changes are part of a continuing response to the increasing quantity of publicly-available genome-scale data, and the consequent need to archive, integrate, annotate and disseminate these using automated, scalable methods., (© The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.)

The Ensembl Genome Browser provides a wealth of freely available genomic data that can be accessed for many purposes by genetics, genomics, and molecular biology researchers. Herein we present two protocols for exploring different aspects of these data: a phenotype and its associated variants and genes, and a promoter and the epigenetic marks and protein-binding activity associated with it. These workflows illustrate a subset of the data types available through the Ensembl Browser, and can be considered a springboard for further exploration.

Ensembl (www.ensembl.org) is a database and genome browser for enabling research on vertebrate genomes. We import, analyse, curate and integrate a diverse collection of large-scale reference data to create a more comprehensive view of genome biology than would be possible from any individual dataset. Our extensive data resources include evidence-based gene and regulatory region annotation, genome variation and gene trees. An accompanying suite of tools, infrastructure and programmatic access methods ensure uniform data analysis and distribution for all supported species. Together, these provide a comprehensive solution for large-scale and targeted genomics applications alike. Among many other developments over the past year, we have improved our resources for gene regulation and comparative genomics, and added CRISPR/Cas9 target sites. We released new browser functionality and tools, including improved filtering and prioritization of genome variation, Manhattan plot visualization for linkage disequilibrium and eQTL data, and an ontology search for phenotypes, traits and disease. We have also enhanced data discovery and access with a track hub registry and a selection of new REST end points. All Ensembl data are freely released to the scientific community and our source code is available via the open source Apache 2.0 license., (© The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Objective: To describe the characteristics of patients who underwent a cranial operation and postoperatively suffered an intracranial hemorrhage significant enough to require evacuation. MATERIALS & METHODS : 3,109 cranial operations were performed at Houston Methodist Hospital (Texas Medical Center campus) between January 2009 and December 2013. Of these, 59 cases required a second operation for evacuation of an intracranial hemorrhage. The information gathered included the patients' age, gender, past medical history, medications and laboratory data, initial diagnosis, date/type of first and second operations, duration of hospitalization, discharge condition, and discharge destination., Results: The study found a 1.90% rate of a postoperative hemorrhage significant enough to require evacuation after a cranial operation. The average age in the cohort requiring reoperation was 63 +/- 14 years with 42 male and 17 female. Hematoma evacuations were performed at various time intervals depending on the pathology treated at the initial operation. The time to second operation was 2.7 days after intraparenchymal hematoma evacuation, 6.0 days after cerebrovascular surgery, 6.2 days after tumor surgery and 9.7 days after subdural hematoma evacuation. The rate of postoperative hematoma development was 9.1% after a subdural hematoma evacuation, while it was only 1.1% in all other operations. Overall, those requiring hematoma evacuation had a 15% mortality rate, 64% were non-ambulatory, and 54% were discharged to long-term acute care facility, skilled nursing facility, rehabilitation facility or hospice. CONCLUSIONS : Neurological outcomes were poor in patients who underwent a cranial operation and required a second operation to remove a hematoma. This study suggests close observation of elderly males after a cranial operation, especially after subdural hematoma evacuation, and longer observation time for patients undergoing subdural hematoma evacuation than intraparenchymal hematoma evacuation, tumor surgery or cerebrovascular surgery.

Our recent study highlights the role of 2 glutathione transferases (GSTs) in the detoxification of the environmental pollutant, 2,4,6-trinitrotoluene (TNT) in Arabidopsis thaliana. TNT is toxic and highly resistant to biodegradation in the environment, raising both health and environmental concerns. Two GSTs, GST-U24 and GST-U25, are upregulated in response to TNT treatment, and expressed predominantly in the root tissues; the site of TNT location following uptake. Plants overexpressing GST-U24 and GST-U25 exhibited significantly enhanced ability to withstand and detoxify TNT, and remove TNT from contaminated soil. Analysis of the catalytic activities of these 2 enzymes revealed that they form 3 TNT-glutathionyl products. Of particular interest is 2-glutathionyl-4,6-dinitrotoluene as this represents a potentially favorable step toward subsequent degradation and mineralization of TNT. We demonstrate how GSTs fit into what is already known about pathways for TNT detoxification, and discuss the short and longer-term fate of TNT conjugates in planta.

The explosive 2,4,6-trinitrotoluene (TNT) is a major worldwide military pollutant. The presence of this toxic and highly persistent pollutant, particularly at military sites and former manufacturing facilities, presents various health and environmental concerns. Due to the chemically resistant structure of TNT, it has proven to be highly recalcitrant to biodegradation in the environment. Here, we demonstrate the importance of two glutathione transferases (GSTs), GST-U24 and GST-U25, from Arabidopsis (Arabidopsis thaliana) that are specifically up-regulated in response to TNT exposure. To assess the role of GST-U24 and GST-U25, we purified and characterized recombinant forms of both enzymes and demonstrated the formation of three TNT glutathionyl products. Importantly, GST-U25 catalyzed the denitration of TNT to form 2-glutathionyl-4,6-dinitrotoluene, a product that is likely to be more amenable to subsequent biodegradation in the environment. Despite the presence of this biochemical detoxification pathway in plants, physiological concentrations of GST-U24 and GST-U25 result in only a limited innate ability to cope with the levels of TNT found at contaminated sites. We demonstrate that Arabidopsis plants overexpressing GST-U24 and GST-U25 exhibit significantly enhanced ability to withstand and detoxify TNT, properties that could be applied for in planta detoxification of TNT in the field. The overexpressing lines removed significantly more TNT from soil and exhibited a corresponding reduction in glutathione levels when compared with wild-type plants. However, in the absence of TNT, overexpression of these GSTs reduces root and shoot biomass, and although glutathione levels are not affected, this effect has implications for xenobiotic detoxification., (© 2014 American Society of Plant Biologists. All Rights Reserved.)

Male Copenhagen rats implanted with Dunning R3327 prostate carcinoma were treated with regional hyperthermia and high energy shock waves (HESW). Twenty-four rats were divided into four equal groups. All rats in one group were matched with the corresponding rats in the other three groups according to tumor size. Hyperthermia was created by the resistive heating of a 22-gauge 1.5-cm needle; HESW were generated by the Dornier XL-1 experimental lithotripter. In two hyperthermia treatments administered 48 hours apart, the tumor was heated to 46.5 degrees C for 2 hours; HESW treatment involved 1,600 shocks at 18 kV. Twenty-nine days after initiation of this study, animals were sacrificed and tumor growth calculated. Tumor growth in group 1 (combined hyperthermia and HESW) was significantly delayed between the 7th and 29th day (p = 0.05). In group 2 (HESW alone), tumor growth was significantly delayed between the 7th and 22nd day (p = 0.01), while tumor growth in group 3 (hyperthermia alone) was delayed between the 3rd and 22nd day (p = 0.02). On day 22, mean percent change in tumor growth in the control group exceeded that of group 1 by 449% (p = 0.015) as well as that of groups 2 and 3, by 350% (p = 0.034) and by 268% (p = 0.049), respectively. As previous reports demonstrate the efficacy of hyperthermia in combination with irradiation or chemotherapy for the treatment of prostate carcinoma, our results indicate hyperthermia in combination with HESW may also be a useful treatment modality.