Synthetic chemical inducers and genetic decoupling enable orthogonal control of the rhaBAD promoter

External control of gene expression is crucial in synthetic biology and biotechnology research and applications, and is commonly achieved using inducible promoter systems. The E. coli rhamnose-inducible rhaBAD promoter has properties superior to more commonly-used inducible expression systems, but is marred by transient expression caused by degradation of the native inducer, L-rhamnose. To address this problem, 35 analogs of L-rhamnose were screened for induction of the rhaBAD promoter, but no strong inducers were identified. In the native configuration, an inducer must bind and activate two transcriptional activators, RhaR and RhaS. Therefore, the expression system was reconfigured to decouple the rhaBAD promoter from the native rhaSR regulatory cascade so that candidate inducers need only activate the terminal transcription factor RhaS. Re-screening the 35 compounds using the modified rhaBAD expression system revealed several promising inducers. These were characterised further to determine the strength, kinetics and concentration-dependence of induction; whether the inducer was used as a carbon source by E. coli; and the modality (distribution) of induction among populations of cells. L-Mannose was found to be the most useful orthogonal inducer, providing an even greater range of induction than the native inducer Lrhamnose, and crucially, allowing sustained induction instead of transient induction. These findings address the key limitation of the rhaBAD expression system, and suggest it may now be the most suitable system for many applications

Team teaching as an agent for change

While the traditional image of the university or college course is often one professor delivering lectures and other course elements alone to a relatively small class, the modern course can be quite different. Large multi-sectioned courses may have a different instructor for each section, often with one instructor serving as coordinator. In other cases two or more faculty members may be involved in delivering a single course section. Courses with labs typically include a lab coordinator as well. These groups form different types of “teaching teams”, and the dynamics of these teams often stimulate change – in pedagogy, teaching philosophy, curriculum, student experience, scholarship and other elements. This panel will explore the ways in which team teaching can act as an agent of change. Panel members will reflect on their own experiences as members of different types of teaching teams, and hope to stimulate a broad and lively exchange with the audience, gathering and synthesizing different perspectives towards building a collective understanding of the potential impact of this teaching format. The session outcome will be an enhanced appreciation for the power and promise of collaborative teaching in generating transformation and change

A microfluidic-multiwell platform for rapid phase mapping of surfactant solutions

Measurement of the phase behavior and (meta)stability of liquid formulations, including surfactant solutions, is required for the understanding of mixture thermodynamics, as well as their practical utilization. We report a microfluidic platform with a stepped temperature profile, imposed by a dual Peltier module, connected to an automated multiwell plate injector and optical setup, for rapid solution phase mapping. The measurement protocol is defined by the temperature step ΔT ≡ T1 − T2 (≲100 ○C), volumetric flow rate Q ≡ ΔV/Δt (≲50 μl/min), which implicitly set the thermal gradient ΔT/Δt (≃0.1–50 ○C/min), and measurement time (which must exceed the intrinsic timescale of the relevant phase transformation). Furthermore, U-shaped microchannels can assess the reversibility of such transformations, yielding a facile measurement of the metastable zone width of the phase diagram. By contrast with traditional approaches, the platform precisely controls the cooling and heating rates by tuning the flow rate, and the absolute temperature excursion by the hot and cold thermal profile, which remain stationary during operation, thus allowing the sequential and reproducible screening of large sample arrays. As a model system, we examined the transition from the micellar (L1) to the liquid crystalline lamellar phase (Lα), upon cooling, of aqueous solutions of sodium linear alkylbenzene sulfonate, a biodegradable anionic surfactant extensively employed in industry. Our findings are validated with quiescent optical microscopy and small angle neutron scattering data

Potent antiviral agents fail to elicit genetically-stable resistance mutations in either enterovirus 71 or Coxsackievirus A16

Enterovirus 71 (EV71) and Coxsackievirus A16 (CVA16) are the two major causative agents 13 of hand, foot and mouth disease (HFMD), for which there are currently no licenced 14 treatments. Here, the acquisition of resistance towards two novel capsid-binding compounds, 15 NLD and ALD, was studied and compared to the analogous compound GPP3. During serial 16 passage, EV71 rapidly became resistant to each compound and mutations at residues I113 17 and V123 in VP1 were identified. A mutation at residue 113 was also identified in CVA16 18 after passage with GPP3. The mutations were associated with reduced thermostability and 19 were rapidly lost in the absence of inhibitors. In silico modelling suggested that the mutations 20 prevented the compounds from binding the VP1 pocket in the capsid. Although both viruses 21 developed resistance to these potent pocket-binding compounds, the acquired mutations were 22 associated with large fitness costs and reverted to WT phenotype and sequence rapidly in the 23 absence of inhibitors. The most effective inhibitor, NLD, had a very large selectivity index, 24 showing interesting pharmacological properties as a novel anti-EV71 agent

Dynamical Autler-Townes control of a phase qubit

Routers, switches, and repeaters are essential components of modern information-processing systems. Similar devices will be needed in future superconducting quantum computers. In this work we investigate experimentally the time evolution of Autler-Townes splitting in a superconducting phase qubit under the application of a control tone resonantly coupled to the second transition. A three-level model that includes independently determined parameters for relaxation and dephasing gives excellent agreement with the experiment. The results demonstrate that the qubit can be used as a ON/OFF switch with 100 ns operating time-scale for the reflection/transmission of photons coming from an applied probe microwave tone. The ON state is realized when the control tone is sufficiently strong to generate an Autler-Townes doublet, suppressing the absorption of the probe tone photons and resulting in a maximum of transmission.Comment: 8 pages, 8 figure

Geographic Coincidence of Increased Malaria Transmission Hazard and Vulnerability Occurring at the Periphery of two Tanzanian Villages.

The goal of malaria elimination necessitates an improved understanding of any fine-scale geographic variations in transmission risk so that complementary vector control tools can be integrated into current vector control programmes as supplementary measures that are spatially targeted to maximize impact upon residual transmission. This study examines the distribution of host-seeking malaria vectors at households within two villages in rural Tanzania. Host-seeking mosquitoes were sampled from 72 randomly selected households in two villages on a monthly basis throughout 2008 using CDC light-traps placed beside occupied nets. Spatial autocorrelation in the dataset was examined using the Moran's I statistic and the location of any clusters was identified using the Getis-Ord Gi* statistic. Statistical associations between the household characteristics and clusters of mosquitoes were assessed using a generalized linear model for each species. For both Anopheles gambiae sensu lato and Anopheles funestus, the density of host-seeking females was spatially autocorrelated, or clustered. For both species, houses with low densities were clustered in the semi-urban village centre while houses with high densities were clustered in the periphery of the villages. Clusters of houses with low or high densities of An. gambiae s.l. were influenced by the number of residents in nearby houses. The occurrence of high-density clusters of An. gambiae s.l. was associated with lower elevations while An. funestus was also associated with higher elevations. Distance from the village centre was also positively correlated with the number of household occupants and having houses constructed with open eaves. The results of the current study highlight that complementary vector control tools could be most effectively targeted to the periphery of villages where the households potentially have a higher hazard (mosquito densities) and vulnerability (open eaves and larger households) to malaria infection

Extra patient movement during mammographic imaging : an experimental study

Objectives: To determine if movement external to the patient occurring during mammography may be a source of image blur. Methods: Four mammography machines with seven flexible and nine fixed paddles were evaluated. In the first stage, movement at the paddle was measured mechanically using two calibrated linear potentiometers. A deformable breast phantom was used to mimic a female breast. For each paddle, the movement in millimeters and change in compression force in Newton was recorded at 0.5 and 1 second intervals respectively for 40 seconds with the phantom in an initially compressed state under a load of 80N. In the second stage, clinical audit on 28 females was conducted on one mammography machine with the 18x24cm and 24x29cm flexible paddles. Results: Movement at the paddle followed an exponential decay with a settling period of approximately 40 seconds. The compression force readings for both fixed and flexible paddles decreased exponentially with time while fixed paddles have a larger drop in compression force than flexible paddles. There is a linear relationship between movement at the paddle and change in compression force. Conclusions: Movement measured at the paddle during an exposure can be represented by a second order system. The amount of extra-patient movement during the actual exposure can be estimated using the linear relationship between movement at the paddle and the change in compression force. Advances in knowledge: This research provides a possible explanation to mammography image blurring caused by extra patient movement and proposes a theoretical model to analyze the movement

TGF-beta 1 induces human alveolar epithelial to mesenchymal cell transition (EMT)

Background: Fibroblastic foci are characteristic features in lung parenchyma of patients with idiopathic pulmonary fibrosis (IPF). They comprise aggregates of mesenchymal cells which underlie sites of unresolved epithelial injury and are associated with progression of fibrosis. However, the cellular origins of these mesenchymal phenotypes remain unclear. We examined whether the potent fibrogenic cytokine TGF-β1 could induce epithelial mesenchymal transition (EMT) in the human alveolar epithelial cell line, A549, and investigated the signaling pathway of TGF-β1-mediated EMT. Methods: A549 cells were examined for evidence of EMT after treatment with TGF-β1. EMT was assessed by: morphology under phase-contrast microscopy; Western analysis of cell lysates for expression of mesenchymal phenotypic markers including fibronectin EDA (Fn-EDA), and expression of epithelial phenotypic markers including E-cadherin (E-cad). Markers of fibrogenesis, including collagens and connective tissue growth factor (CTGF) were also evaluated by measuring mRNA level using RT-PCR, and protein by immunofluorescence or Western blotting. Signaling pathways for EMT were characterized by Western analysis of cell lysates using monoclonal antibodies to detect phosphorylated Erk1/2 and Smad2 after TGF-β1 treatment in the presence or absence of MEK inhibitors. The role of Smad2 in TGF-β1-mediated EMT was investigated using siRNA. Results: The data showed that TGF-β1, but not TNF-α or IL-1β, induced A549 cells with an alveolar epithelial type II cell phenotype to undergo EMT in a time-and concentration-dependent manner. The process of EMT was accompanied by morphological alteration and expression of the fibroblast phenotypic markers Fn-EDA and vimentin, concomitant with a downregulation of the epithelial phenotype marker E-cad. Furthermore, cells that had undergone EMT showed enhanced expression of markers of fibrogenesis including collagens type I and III and CTGF. MMP-2 expression was also evidenced. TGF-β1-induced EMT occurred through phosphorylation of Smad2 and was inhibited by Smad2 gene silencing; MEK inhibitors failed to attenuate either EMT-associated Smad2 phosphorylation or the observed phenotypic changes. Conclusion: Our study shows that TGF-β1 induces A549 alveolar epithelial cells to undergo EMT via Smad2 activation. Our data support the concept of EMT in lung epithelial cells, and suggest the need for further studies to investigate the phenomenon

Prognostic Factors in Arthroplasty in the Rheumatoid Shoulder

Total shoulder arthroplasty is commonly considered a good option for treatment of the rheumatoid shoulder. However, when the rotator cuff and glenoid bone stock are not preserved, the clinical outcome of arthroplasty in the rheumatoid patients remains unclear. Aim of the study is to explore the prognostic value of multiple preoperative and peroperative variables in total shoulder arthroplasty and shoulder hemiarthroplasty in rheumatoid patients. Clinical Hospital for Special Surgery Shoulder score was determined at different time points over a mean period of 6.5 years in 66 rheumatoid patients with total shoulder arthroplasty and 75 rheumatoid patients with shoulder hemiarthroplasty. Moreover, radiographic analysis was performed to assess the progression of humeral head migration and glenoid loosening. Advanced age and erosions or cysts at the AC joint at time of surgery were associated with a lower postoperative Clinical Hospital for Special Surgery Shoulder score. In total shoulder arthroplasty, status of the rotator cuff and its repair at surgery were predictive of postoperative improvement. Progression of proximal migration during the period after surgery was associated with a lower clinical score over time. However, in hemiarthroplasty, no relation was observed between the progression of proximal or medial migration during follow-up and the clinical score over time. Status of the AC joint and age at the time of surgery should be taken into account when considering shoulder arthroplasty in rheumatoid patients. Total shoulder arthroplasty in combination with good cuff repair yields comparable clinical results as total shoulder arthroplasty when the cuff is intact