Characterization of the amp genes involved in the regulation of beta-lactamase expression in Pseudomonas aeruginosa

Antibiotic resistance, production of alginate and virulence factors, and altered host immune responses are the hallmarks of chronic Pseudomonas aeruginosa infection. Failure of antibiotic therapy has been attributed to the emergence of P. aeruginosa strains that produce β-lactamase constitutively. In Enterobacteriaceae, β-lactamase induction involves four genes with known functions: ampC, ampR, ampD, and ampG, encoding the enzyme, transcriptional regulator, amidase and permease, respectively. In addition to all these amp genes, P. aeruginosa possesses two ampG paralogs, designated ampG and ampP. In this study, P. aeruginosa ampC, ampR, ampG and ampP were analyzed. Inactivation of ampC in the prototypic PAO1 failed to abolish the β-lactamase activity leading to the discovery of P. aeruginosa oxacillinase PoxB. Cloning and expression of poxB in Escherichia coli confers β-lactam resistance. Both AmpC and PoxB contribute to P. aeruginosa resistance against a wide spectrum of β-lactam antibiotics. The expression of PoxB and AmpC is regulated by a LysR-type transcriptional regulator AmpR that up-regulates AmpC but down-regulates PoxB activities. Analyses of P. aeruginosa ampR mutant demonstrate that AmpR is a global regulator that modulates the expressions of Las and Rhl quorum sensing (QS) systems, and the production of pyocyanin, LasA protease and LasB elastase. Introduction of the ampR mutation into an alginate-producing strain reveals the presence of a complex co-regulatory network between antibiotic resistance, QS alginate and other virulence factor production. Using phoA and lacZ protein fusion analyses, AmpR, AmpG and AmpP were localized to the inner membrane with one, 16 and 10 transmembrane helices, respectively. AmpR has a cytoplasmic DNA-binding and a periplasmic substrate binding domains. AmpG and AmpP are essential for the maximal expression of β-lactamase. Analysis of the murein breakdown products suggests that AmpG exports UDP-N-acetylmuramyl-L-alanine-γ-D-glutamate-meso-diaminopimelic acid-D-alanine-D-alanine (UDP-MurNAc-pentapeptide), the corepressor of AmpR, whereas AmpP imports N-acetylglucosaminyl-beta-1,4-anhydro-N-acetylmuramic acid-Ala-γ-D-Glu-meso-diaminopimelic acid (GlcNAc-anhMurNAc-tripeptide) and GlcNAc-anhMurNAc-pentapeptide, the co-inducers of AmpR. This study reveals a complex interaction between the Amp proteins and murein breakdown products involved in P. aeruginosa β-lactamase induction. In summary, this dissertation takes us a little closer to understanding the P. aeruginosa complex co-regulatory mechanism in the development of β-lactam resistance and establishment of chronic infection. ^

Feature recognition & tool path generation for 5 axis step-nc machining of free form / irregular contoured surfaces

This research paper presents a five step algorithm to generate tool paths for machining Free form / Irregular Contoured Surface(s) (FICS) by adopting STEP-NC (AP-238) format. In the first step, a parametrized CAD model with FICS is created or imported in UG-NX6.0 CAD package. The second step recognizes the features and calculates a Closeness Index (CI) by comparing them with the B-Splines / Bezier surfaces. The third step utilizes the CI and extracts the necessary data to formulate the blending functions for identified features. In the fourth step Z-level 5 axis tool paths are generated by adopting flat and ball end mill cutters. Finally, in the fifth step, tool paths are integrated with STEP-NC format and validated. All these steps are discussed and explained through a validated industrial component.

Enlivening, Invigorating & Enhancing Science:This session explores learning from a range of Primary Science Teaching Trust projects focused on invigorating the experience of children, teachers and whole school communities with primary science.

This session will provide you with opportunity to find out what is being achieved and explore the implications for your own practice.

WHAT DOES THE SOCIAL ACTION, RESPONSIBILITY & HEROISM ACT 2015 MEAN FOR SPORTS VOLUNTEERS AND NGBS?

By highlighting the context of sports coaching in the UK, this article reveals the considerable limitations of both section 1 of the Compensation Act 2006, and the Social Action, Responsibility and Heroism Act 2015, in safeguarding (volunteer) coaches from negligence liability.

Exchange protein directly activated by cyclic AMP (EPAC) activation reverses neutrophil dysfunction induced by β2-agonists, corticosteroids, and critical illness

Background Neutrophils play a role in the pathogenesis of asthma, chronic obstructive pulmonary disease, and pulmonary infection. Impaired neutrophil phagocytosis predicts hospital-acquired infection. Despite this, remarkably few neutrophil-specific treatments exist. 

Objectives We sought to identify novel pathways for the restoration of effective neutrophil phagocytosis and to activate such pathways effectively in neutrophils from patients with impaired neutrophil phagocytosis. 

Methods Blood neutrophils were isolated from healthy volunteers and patients with impaired neutrophil function. In healthy neutrophils phagocytic impairment was induced experimentally by using β₂-agonists. Inhibitors and activators of cyclic AMP (cAMP)-dependent pathways were used to assess the influence on neutrophil phagocytosis in vitro. 

Results β₂-Agonists and corticosteroids inhibited neutrophil phagocytosis. Impairment of neutrophil phagocytosis by β₂-agonists was associated with significantly reduced RhoA activity. Inhibition of protein kinase A (PKA) restored phagocytosis and RhoA activity, suggesting that cAMP signals through PKA to drive phagocytic impairment. However, cAMP can signal through effectors other than PKA, such as exchange protein directly activated by cyclic AMP (EPAC). An EPAC-activating analog of cAMP (8CPT-2Me-cAMP) reversed neutrophil dysfunction induced by β₂-agonists or corticosteroids but did not increase RhoA activity. 8CPT-2Me-cAMP reversed phagocytic impairment induced by Rho kinase inhibition but was ineffective in the presence of Rap-1 GTPase inhibitors. 8CPT-2Me-cAMP restored function to neutrophils from patients with known acquired impairment of neutrophil phagocytosis. 

Conclusions EPAC activation consistently reverses clinical and experimental impairment of neutrophil phagocytosis. EPAC signals through Rap-1 and bypasses RhoA. EPAC activation represents a novel potential means by which to reverse impaired neutrophil phagocytosis.

Emotions & Power:Habitus and Change in Late Modernity

(Under contract)

Reassembled, Slightly Askew:Composition & Sound Design

Reassembled, Slightly Askew is an autobiographical, immersive audio-based artwork based on Shannon Sickels’ experience of falling critically ill with a rare brain infection and her journey of rehabilitation with an acquired brain injury. Audience members experience Reassembled individually, listening to the audio via headphones while lying on a bed. The piece makes use of binaural microphone technology and spatial sound design techniques, causing listeners to feel they are inside Shannon’s head, viscerally experiencing her descent into coma, brain surgeries, early days in the hospital, and re-integration into the world with a hidden disability. It is a new kind of storytelling, never done before about this topic, that places the listener safely in the first-person perspective with the aim of increasing empathy and understanding. Reassembled… was made through a 5-year collaboration with an interdisciplinary team of artists led by Shannon Sickels (writer & performer), Paul Stapleton (composer & sound designer), Anna Newell (director), Hanna Slattne (dramaturgy), Stevie Prickett (choreography), and Shannon’s consultant neurosurgeon and head injury nurse. It’s development and production has been made possible with the support of a Wellcome Trust Arts Award, the Arts Council NI, Sonic Arts Research Centre, Belfast's Metropolitan Arts Centre, and grants from the Arts & Disability Award Ireland scheme. In its 2015 premiere year, Reassembled had 99 shows across Northern Ireland, including at the Cathedral Quarter Arts Festival (the MAC, Belfast) and BOUNCE Arts & Disability Forum Festival (Lyric Theatre, Belfast). It was awarded 5 stars in the Stage, a Hospital Club h100 Theatre & Performance Award, and been shared at medical conferences and trainings across the UK. It continues to be presented in diverse artistic and educational contexts, including as part of A Nation’s Theatre Festival in 2016 at Battersea Arts Centre in London where it was given 4 star reviews in the Guardian, Time Out London and the Evening Standard. "A real-life ordeal, captured by a daring, disorientating artistic collaboration, which works brilliantly on so many levels…It should be available on prescription.” — The Stage ★★★★★ www.reassembled.co.uk Audio clips and documentary footage available here: http://www.paulstapleton.net/portfolio/reassembled-slightly-askew

Lawyers & Truth Recovery Mechanisms: South Africa

Focusing on post-apartheid South Africa, the report explores the role of lawyers in truth recovery mechanisms.

The report was prepared by Dr Rachel Killean and draws on a series of interviews conducted in South Africa (with legal academics, ‘struggle’ lawyers, state lawyers, judges and human rights activists) as part of the wider Lawyers, Conflict and Transition project.

Dr Killean begins with an overview of the various roles the legal profession has played in South Africa, both during the apartheid era and post-transition.

The first half of the report then explores the role of lawyers as professional participants – firstly at the South African Truth and Reconciliation Commission and secondly in the Marikana Commission of Inquiry.

The report then considers the notion of lawyers as subjects of truth recovery, looking in particular at the Special Legal Hearing on the legal profession as part of the South African Truth and Reconciliation Commission.

In the concluding section Killean reflects on the extent to which lawyers influence the procedures and outcomes of truth recovery mechanisms and offers some concrete suggestions as to how the involvement of lawyers in such processes might be more effectively managed.

With regard to lawyers as subjects of truth recovery, she acknowledges the limitations of the South African model but posits that the endeavour must be applauded, not least because it demonstrated that it is possible to scrutinise the role of the legal profession in past conflict, and that it is worth wrestling with the associated challenges.

Det laborativa arbetsmaterialet i matematik i lågstadiet (åk1-3) – vad, vem, när, hur & varför?

Syftet med denna studie är att undersöka det laborativa arbetsmaterialets förekomst i matematikundervisningen och ta reda på hur lärare förhåller sig till materialet och användandet av detta i årskurserna 1-3. Undersökningen är huvudsakligen en kvalitativ studie som bygger på 7 enkätfrågor ställda till lärare i lågstadiet. Min undersökning visar att laborativt arbetsmaterial förekommer i matematikundervisningen och används vid genomgångar av lärarna samt vid behov av både elever och lärare. Materialet är både roligt att använda och ett stöd för lärandet. Vissa tyckte dock att det kunde vara jobbigt att plocka fram och städa undan och att eleverna ibland lekte med det. Lärarna gav exempel på kopplingar till läroplanen och de tillfrågade lärarna upplever sig ha tillräcklig kompetens för att kunna bedriva god undervisning för främjandet av elevernas kunskapsutveckling inom ämnet matematik. Det var dock viktigt med ett fungerande erfarenhetsutbyte i arbetslaget samt möjlighet till fortbildning vid behov.

Uppskattning av betalningsviljan för Peace & Love 2011

I denna uppsats skattas betalningsviljan hos besökarna på Peace & Love-festivalen år 2011. Med hjälp av enkätdata baserad på avslöjade och uttalade preferenser presenteras en regressionsanalys med olika oberoende variabler som karaktäriserar en festivalbesökare. Total budget är den beroende variabeln i regressionsanalysen och tolkas i uppsatsen som ekvivalent med besökarnas betalningsvilja. Analysen visar att män i genomsnitt spenderar 301 kronor mer än kvinnor, att turister i genomsnitt spenderar 1 124 kronor mer än en icke-turist samt att den genomsnittliga besökaren har en betalningsvilja på 4 183 kronor. Ett skattat konsumentöverskott har också värderats, vilket uppgick till 743 kronor per person och cirka 37 miljoner kronor totalt för de 50 000 festivalbesökarna. Uppsatsen tar inte hänsyn till de ekonomiska effekter som festivalen har på Borlänge som stad.

A importância da liderança no clima organizacional: estudo caso da AMP

Esta Monografia intitulada, “A importância da liderança no clima organizacional” realizada na Cidade do Mindelo, na Universidade do Mindelo, incide sobre a problemática da organização das empresas, como sendo um dos pilares base para a liderança dentro das empresas, centrando a sua importância na influência que esta tem no clima organizacional, sem descorar do seu impacto para o bom funcionamento de qualquer empresa. Por isso, apresentam-se ao longo deste trabalho os vários aspetos que podem ser determinantes para que possa haver um bom clima organizacional, e como é que a liderança pode contribuir para tal, como sendo determinante para o sucesso ou insucesso de uma empresa ou serviço. Ele revela ainda os diferentes modelos e finalidades de liderança, versando também a importância da formação e do conhecimento do líder para um melhor acompanhamento da sua empresa tendo como pano de fundo um bom clima organizacional. Realça-se também, as consequências que são causados por uma má liderança ou um clima organizacional instável, bem como as suas consequências para o ambiente profissional e as interferências que isto pode ter no sucesso ou insucesso de uma empresa ou serviço, conduzindo assim a performance deste ou aquele serviço. A partir da metodologia utilizada conseguiu-se avaliar o tipo de clima organizacional e a liderança existentes na empresa em estudo e estabelecer a uma comparação da forma como a teoria se articula com a prática. Os resultados alcançados demostram que a liderança influencia no clima organizacional de forma positiva ou negativa.