Development of stable reporter system cloning luxCDABE genes into chromosome of Salmonella enterica serotypes using Tn7 transposon

<p>Abstract</p> <p>Background</p> <p>Salmonellosis may be a food safety problem when raw food products are mishandled and not fully cooked. In previous work, we developed bioluminescent <it>Salmonella enterica </it>serotypes using a plasmid-based reporting system that can be used for real-time monitoring of the pathogen's growth on food products in short term studies. In this study, we report the use of a Tn7-based transposon system for subcloning of <it>luxCDABE </it>genes into the chromosome of eleven <it>Salmonella enterica </it>serotypes isolated from the broiler production continuum.</p> <p>Results</p> <p>We found that the <it>lux </it>operon is constitutively expressed from the chromosome post-transposition and the <it>lux </it>cassette is stable without external pressure, i.e. antibiotic selection, for all <it>Salmonella enterica </it>serotypes used. Bioluminescence expression is based on an active electron transport chain and is directly related with metabolic activity. This relationship was quantified by measuring bioluminescence against a temperature gradient in aqueous solution using a luminometer. In addition, bioluminescent monitoring of two serotypes confirmed that our chicken skin model has the potential to be used to evaluate pathogen mitigation strategies.</p> <p>Conclusions</p> <p>This study demonstrated that our new stable reporting system eliminates bioluminescence variation due to plasmid instability and provides a reliable real-time experimental system to study application of preventive measures for <it>Salmonella </it>on food products in real-time for both short and long term studies.</p

In Vivo Activity of Metal Complexes Containing 1,10-Phenanthroline and 3,6,9-Trioxaundecanedioate Ligands against Pseudomonas aeruginosa Infection in Galleria mellonella Larvae

Drug-resistant Pseudomonas aeruginosa is rapidly developing resulting in a serious global threat. Immunocompromised patients are specifically at risk, especially those with cystic fibrosis (CF). Novel metal complexes incorporating 1,10-phenanthroline (phen) ligands have previously demonstrated antibacterial and anti-biofilm effects against resistant P. aeruginosa from CF patients in vitro. Herein, we present the in vivo efficacy of {[Cu(3,6,9-tdda)(phen)2]·3H2O·EtOH}n (Cu-tdda-phen), {[Mn(3,6,9-tdda)(phen)2]·3H2O·EtOH}n (Mn-tdda-phen) and [Ag2(3,6,9-tdda)(phen)4]·EtOH (Ag-tdda-phen) (tddaH2 = 3,6,9-trioxaundecanedioic acid). Individual treatments of these metal-tdda-phen complexes and in combination with the established antibiotic gentamicin were evaluated in vivo in larvae of Galleria mellonella infected with clinical isolates and laboratory strains of P. aeruginosa. G. mellonella were able to tolerate all test complexes up to 10 µg/larva. In addition, the immune response was affected by stimulation of immune cells (hemocytes) and genes that encode for immune-related peptides, specifically transferrin and inducible metallo-proteinase inhibitor. The amalgamation of metal-tdda-phen complexes and gentamicin further intensified this response at lower concentrations, clearing a P. aeruginosa infection that were previously resistant to gentamicin alone. Therefore this work highlights the anti-pseudomonal capabilities of metal-tdda-phen complexes alone and combined with gentamicin in an in vivo model

In Vivo Activity of Copper(II), Manganese(II), and Silver(I) 1,10-Phenanthroline Chelates

Candida haemulonii is an emerging opportunistic pathogen resistant to most antifungal drugs currently used in clinical arena. Metal complexes containing 1,10-phenanthroline (phen) chelating ligands have well-established anti-Candida activity against different medically relevant species. This study utilized larvae of Galleria mellonella, a widely used model of in vivo infection, to examine C. haemulonii infection characteristics in response to different copper(II), manganese(II), and silver(I) chelates containing phen, which had demonstrated potent anti-C. haemulonii activity in a previous study. The results showed that C. haemulonii virulence was influenced by inoculum size and incubation temperature, and the host G. mellonella immune response was triggered in an inoculum-dependent manner reflected by the number of circulating immune cells (hemocytes) and observance of larval melanization process. All test chelates were non-toxic to the host in concentrations up to 10 μg/larva. The complexes also affected the G. mellonella immune system, affecting the hemocyte number and the expression of genes encoding antifungal and immune-related peptides (e.g., inducible metalloproteinase inhibitor protein, transferrin, galiomycin, and gallerimycin). Except for [Ag2(3,6,9-tdda)(phen)4].EtOH (3,6,9-tddaH2 = 3,6,9-trioxoundecanedioic acid), all chelates were capable of affecting the fungal burden of infected larvae and the virulence of C. haemulonii in a dose-dependent manner. This work shows that copper(II), manganese(II), and silver(I) chelates containing phen with anti-C. haemulonii activity are capable of (i) inhibiting fungal proliferation during in vivo infection, (ii) priming an immune response in the G. mellonella host and (iii) affecting C. haemulonii virulence

DNA Methylation Profiling of the Human Major Histocompatibility Complex: A Pilot Study for the Human Epigenome Project

The Human Epigenome Project aims to identify, catalogue, and interpret genome-wide DNA methylation phenomena. Occurring naturally on cytosine bases at cytosine–guanine dinucleotides, DNA methylation is intimately involved in diverse biological processes and the aetiology of many diseases. Differentially methylated cytosines give rise to distinct profiles, thought to be specific for gene activity, tissue type, and disease state. The identification of such methylation variable positions will significantly improve our understanding of genome biology and our ability to diagnose disease. Here, we report the results of the pilot study for the Human Epigenome Project entailing the methylation analysis of the human major histocompatibility complex. This study involved the development of an integrated pipeline for high-throughput methylation analysis using bisulphite DNA sequencing, discovery of methylation variable positions, epigenotyping by matrix-assisted laser desorption/ionisation mass spectrometry, and development of an integrated public database available at http://www.epigenome.org. Our analysis of DNA methylation levels within the major histocompatibility complex, including regulatory exonic and intronic regions associated with 90 genes in multiple tissues and individuals, reveals a bimodal distribution of methylation profiles (i.e., the vast majority of the analysed regions were either hypo- or hypermethylated), tissue specificity, inter-individual variation, and correlation with independent gene expression data

Surface plasmon resonance imaging of excitable cells

Surface plasmons (SPs) are surface charge density oscillations occuring at a metal/dieletric interface and are highly sensitive to refractive index variations adjacent to the surface. This sensitivity has been exploited successfully for chemical and biological assays. In these systems, a surface plasmon resonance (SPR)-based sensor detects temporal variations in the refractive index at a point. SPR has also been used in imaging systems where the spatial variations of refractive index in the sample provide the contrast mechanism. SPR imaging systems using high numerical aperture (NA) objective lenses have been designed to image adherent live cells with high magnification and near-diffraction limited spatial resolution. Addressing research questions in cell physiology and pharmacology often requires the development of a multimodal microscope where complementary information can be obtained.In this paper, we present the development of a multimodal microscope that combines SPR imaging with a number of additional imaging modalities including bright-field, epifluorescence, total internal reflection microscopy and SPR fluorescence microscopy. We used a high NA objective lens for SPR and TIR microscopy and the platform has been used to image live cell cultures demonstrating both fluorescent and label-free techniques. Both the SPR and TIR imaging systems feature a wide field of view (~300 µm) that allows measurements from multiple cells whilst maintaining a resolution sufficient to image fine cellular processes. The capability of the platform to perform label-free functional imaging of living cells was demonstrated by imaging the spatial variations in contractions from stem cell-derived cardiomyocytes. This technique shows promise for non-invasive imaging of cultured cells over very long periods of time during development

The Alliance of Genome Resources: Building a Modern Data Ecosystem for Model Organism Databases.

Model organisms are essential experimental platforms for discovering gene functions, defining protein and genetic networks, uncovering functional consequences of human genome variation, and for modeling human disease. For decades, researchers who use model organisms have relied on Model Organism Databases (MODs) and the Gene Ontology Consortium (GOC) for expertly curated annotations, and for access to integrated genomic and biological information obtained from the scientific literature and public data archives. Through the development and enforcement of data and semantic standards, these genome resources provide rapid access to the collected knowledge of model organisms in human readable and computation-ready formats that would otherwise require countless hours for individual researchers to assemble on their own. Since their inception, the MODs for the predominant biomedical model organisms [Mus sp (laboratory mouse), Saccharomyces cerevisiae, Drosophila melanogaster, Caenorhabditis elegans, Danio rerio, and Rattus norvegicus] along with the GOC have operated as a network of independent, highly collaborative genome resources. In 2016, these six MODs and the GOC joined forces as the Alliance of Genome Resources (the Alliance). By implementing shared programmatic access methods and data-specific web pages with a unified look and feel, the Alliance is tackling barriers that have limited the ability of researchers to easily compare common data types and annotations across model organisms. To adapt to the rapidly changing landscape for evaluating and funding core data resources, the Alliance is building a modern, extensible, and operationally efficient knowledge commons for model organisms using shared, modular infrastructure

The Vehicle, Fall 1983

Vol. 25, No. 1 Table of Contents Amish BoyDevon Flesorpage 3 SyllogismJ. Maura Davispage 3 Ten SecondsD.L. Lewispage 4 The Cedar ChestBridget M. Howepage 4 A Christmas With CarolSteve Longpage 5 TeethMichelle Mitchellpage 7 An I-Love-You PoemD.L. Lewispage 8 The Dragon SlayerSusan Gradypage 8 A DefinitionAmy J. Eadespage 9 FingernailsSuzanne Hornpage 10 The Liar\u27s TableBrook Wilsonpage 10 Fifi\u27s Last PartySteve Longpage 12 Absence/PresenceSuzanne Hornpage 13 From the Rantings of a Mad Astronomy StudentAmy J. Eadespage 13 In the Name of the Father, the Son, and MachiavelliF. Link Rapierpage 15 Errant LoverBecky Lawsonpage 16 DaddyKevin Lylespage 16 GhostsGary Ervinpage 17 TangoF. Link Rapierpage 17 Grandma\u27s SlippersBecky Lawsonpage 18 EdgesAmy J. Eadespage 19 Having ChildrenDevon Flesorpage 20 Young Black GirlKevin Lylespage 21 CatSuzanne Hornpage 22 Breakfast for OneMichelle Mitchellpage 22 A Modest ProposalBrooke Sanfordpage 23 Post MortemF. Link Rapierpage 26 Who Said I Forgot?Lynne Krausepage 27 The Corner Booth at StuckeysMaggie Kennedypage 28 The First DayDavis Brydenpage 29 DownLynne Krausepage 30 Fairie RingDevon Flesorpage 31 The LaundrymatKathy Fordpage 32 Sunday in OctoberBridget M. Howepage 32 The Kitchen WindowMaggie Kennedypage 33 UntitledChristina Maire Vitekpage 34 8th Grade Field Trip to SpringfieldMichelle Mitchellpage 34 Children of the FortiesF. Link Rapierpage 35 one winter and i was eightGary Ervinpage 35 Don\u27t we all know?Thomas B. Waltrippage 36 The TravelerMaggie Kennedypage 36 The VisitKathy Fordpage 40 CubismMaggie Kennedypage 40https://thekeep.eiu.edu/vehicle/1042/thumbnail.jp

The Vehicle, Fall 1983

Vol. 25, No. 1 Table of Contents Amish BoyDevon Flesorpage 3 SyllogismJ. Maura Davispage 3 Ten SecondsD.L. Lewispage 4 The Cedar ChestBridget M. Howepage 4 A Christmas With CarolSteve Longpage 5 TeethMichelle Mitchellpage 7 An I-Love-You PoemD.L. Lewispage 8 The Dragon SlayerSusan Gradypage 8 A DefinitionAmy J. Eadespage 9 FingernailsSuzanne Hornpage 10 The Liar\u27s TableBrook Wilsonpage 10 Fifi\u27s Last PartySteve Longpage 12 Absence/PresenceSuzanne Hornpage 13 From the Rantings of a Mad Astronomy StudentAmy J. Eadespage 13 In the Name of the Father, the Son, and MachiavelliF. Link Rapierpage 15 Errant LoverBecky Lawsonpage 16 DaddyKevin Lylespage 16 GhostsGary Ervinpage 17 TangoF. Link Rapierpage 17 Grandma\u27s SlippersBecky Lawsonpage 18 EdgesAmy J. Eadespage 19 Having ChildrenDevon Flesorpage 20 Young Black GirlKevin Lylespage 21 CatSuzanne Hornpage 22 Breakfast for OneMichelle Mitchellpage 22 A Modest ProposalBrooke Sanfordpage 23 Post MortemF. Link Rapierpage 26 Who Said I Forgot?Lynne Krausepage 27 The Corner Booth at StuckeysMaggie Kennedypage 28 The First DayDavis Brydenpage 29 DownLynne Krausepage 30 Fairie RingDevon Flesorpage 31 The LaundrymatKathy Fordpage 32 Sunday in OctoberBridget M. Howepage 32 The Kitchen WindowMaggie Kennedypage 33 UntitledChristina Maire Vitekpage 34 8th Grade Field Trip to SpringfieldMichelle Mitchellpage 34 Children of the FortiesF. Link Rapierpage 35 one winter and i was eightGary Ervinpage 35 Don\u27t we all know?Thomas B. Waltrippage 36 The TravelerMaggie Kennedypage 36 The VisitKathy Fordpage 40 CubismMaggie Kennedypage 40https://thekeep.eiu.edu/vehicle/1042/thumbnail.jp

Breast-Cancer-Specific Mortality in Patients Treated Based on the 21-Gene Assay: A SEER Population-Based Study

The 21-gene Recurrence Score assay is validated to predict recurrence risk and chemotherapy benefit in hormone-receptor-positive (HR+) invasive breast cancer. To determine prospective breast-cancer-specific mortality (BCSM) outcomes by baseline Recurrence Score results and clinical covariates, the National Cancer Institute collaborated with Genomic Health and 14 population-based registries in the the Surveillance, Epidemiology, and End Results (SEER) Program to electronically supplement cancer surveillance data with Recurrence Score results. The prespecified primary analysis cohort was 40–84 years of age, and had node-negative, HR+, HER2-negative, nonmetastatic disease diagnosed between January 2004 and December 2011 in the entire SEER population, and Recurrence Score results (N = 38,568). Unadjusted 5-year BCSM were 0.4% (n = 21,023; 95% confidence interval (CI), 0.3–0.6%), 1.4% (n = 14,494; 95% CI, 1.1–1.7%), and 4.4% (n = 3,051; 95% CI, 3.4–5.6%) for Recurrence Score \u3c 18, 18–30, and ≥ 31 groups, respectively (P \u3c 0.001). In multivariable analysis adjusted for age, tumor size, grade, and race, the Recurrence Score result predicted BCSM (P \u3c 0.001). Among patients with node-positive disease (micrometastases and up to three positive nodes; N = 4,691), 5-year BCSM (unadjusted) was 1.0% (n = 2,694; 95% CI, 0.5–2.0%), 2.3% (n = 1,669; 95% CI, 1.3–4.1%), and 14.3% (n = 328; 95% CI, 8.4–23.8%) for Recurrence Score \u3c 18, 18–30, ≥ 31 groups, respectively (P \u3c 0.001). Five-year BCSM by Recurrence Score group are reported for important patient subgroups, including age, race, tumor size, grade, and socioeconomic status. This SEER study represents the largest report of prospective BCSM outcomes based on Recurrence Score results for patients with HR+, HER2-negative, node-negative, or node-positive breast cancer, including subgroups often under-represented in clinical trials