Quantitative Imaging of Protein-Protein Interactions by Multiphoton Fluorescence Lifetime Imaging Microscopy using a Streak camera

Fluorescence Lifetime Imaging Microscopy (FLIM) using multiphoton excitation techniques is now finding an important place in quantitative imaging of protein-protein interactions and intracellular physiology. We review here the recent developments in multiphoton FLIM methods and also present a description of a novel multiphoton FLIM system using a streak camera that was developed in our laboratory. We provide an example of a typical application of the system in which we measure the fluorescence resonance energy transfer between a donor/acceptor pair of fluorescent proteins within a cellular specimen.Comment: Overview of FLIM techniques, StreakFLIM instrument, FRET application

Genome analysis of Clostridium difficile PCR ribotype 014 lineage in Australian pigs and humans reveals a diverse genetic repertoire and signatures of long-range interspecies transmission

Clostridium difficile PCR ribotype (RT) 014 is well-established in both human and porcine populations in Australia, raising the possibility that C. difficile infection (CDI) may have a zoonotic or foodborne etiology. Here, whole genome sequencing and high-resolution core genome phylogenetics were performed on a contemporaneous collection of 40 Australian RT014 isolates of human and porcine origin. Phylogenies based on MLST (7 loci, STs 2, 13, and 49) and core orthologous genes (1260 loci) showed clustering of human and porcine strains indicative of very recent shared ancestry. Core genome single nucleotide variant (SNV) analysis found 42 % of human strains showed a clonal relationship (separated by ≤ 2 SNVs in their core genome) with one or more porcine strains, consistent with recent inter-host transmission. Clones were spread over a vast geographic area with 50 % of the human cases occurring without recent healthcare exposure. These findings suggest a persistent community reservoir with long-range dissemination, potentially due to agricultural recycling of piggery effluent. We also provide the first pan-genome analysis for this lineage, characterizing its resistome, prophage content, and in silico virulence potential. The RT014 is defined by a large “open” pan-genome (7587 genes) comprising a core genome of 2296 genes (30.3 % of the total gene repertoire) and an accessory genome of 5291 genes. Antimicrobial resistance genotypes and phenotypes varied across host populations and ST lineages and were characterized by resistance to tetracycline [tetM, tetA(P), tetB(P) and tetW], clindamycin/erythromycin (ermB), and aminoglycosides (aph3-III-Sat4A-ant6-Ia). Resistance was mediated by clinically important mobile genetic elements, most notably Tn6194 (harboring ermB) and a novel variant of Tn5397 (harboring tetM). Numerous clinically important prophages (Siphoviridae and Myoviridae) were identified as well as an uncommon accessory gene regulator locus (agr3). Conservation in the pathogenicity locus and S-layer correlated with ST affiliation, further extending the concept of clonal C. difficile lineages. This study provides novel insights on the genetic variability and strain relatedness of C. difficile RT014, a lineage of emerging One Health importance. Ongoing molecular and genomic surveillance of strains in humans, animals, food, and the environment is imperative to identify opportunities to reduce the overall CDI burden

Capture of particles of dust by convective flow

Interaction of particles of dust with vortex convective flows is under theoretical consideration. It is assumed that the volume fraction of solid phase is small, variations of density due to nonuniform distribution of particles and those caused by temperature nonisothermality of medium are comparable. Equations for the description of thermal buoyancy convection of a dusty medium are developed in the framework of the generalized Boussinesq approximation taking into account finite velocity of particle sedimentation. The capture of a cloud of dust particles by a vortex convective flow is considered, general criterion for the formation of such a cloud is obtained. The peculiarities of a steady state in the form of a dust cloud and backward influence of the solid phase on the carrier flow are studied in detail for a vertical layer heated from the sidewalls. It is shown that in the case, when this backward influence is essential, a hysteresis behavior is possible. The stability analysis of the steady state is performed. It turns out that there is a narrow range of governing parameters, in which such a steady state is stable.Comment: 14 pages, 10 figures, published in Physics of Fluid

A Benefit of Context Reinstatement to Recognition Memory in Aging: The Role of Familiarity Processes

Reinstatement of encoding context facilitates memory for targets in young and older individuals (e.g., a word studied on a particular background scene is more likely to be remembered later if it is presented on the same rather than a different scene or no scene), yet older adults are typically inferior at recalling and recognizing target–context pairings. This study examined the mechanisms of the context effect in normal aging. Age differences in word recognition by context condition (original, switched, none, new), and the ability to explicitly remember target–context pairings were investigated using word–scene pairs (Experiment 1) and word–word pairs (Experiment 2). Both age groups benefited from context reinstatement in item recognition, although older adults were significantly worse than young adults at identifying original pairings and at discriminating between original and switched pairings. In Experiment 3, participants were given a three-alternative forced-choice recognition task that allowed older individuals to draw upon intact familiarity processes in selecting original pairings. Performance was age equivalent. Findings suggest that heightened familiarity associated with context reinstatement is useful for boosting recognition memory in aging

A three-dimensional model of wave attenuation in the marginal ice zone

Extent: 17p.A three-dimensional model of wave scattering by a large array of floating thin elastic plates is used to predict the rate of ocean wave attenuation in the marginal ice zone in terms of the properties of the ice cover and the incoming wavefield. This is regarded as a small step toward assimilating interactions of ocean waves with areas of sea ice into oceanic general circulation models. Numerical results confirm previous findings that attenuation is predominantly affected by wave period and by the average thickness of the ice cover. It is found that the shape and distribution of the floes and the inclusion of an Archimedean draft has little impact on the attenuation produced. The model demonstrates a linear relationship between ice cover concentration and attenuation. An additional study is conducted into the directional evolvement of the wavefield, where collimation and spreading can both occur, depending on the physical circumstances. Finally, the attenuation predicted by the new three-dimensional model is compared with an existing two-dimensional model and with two sets of experimental data, with the latter producing convincing agreement.L. G. Bennetts, M. A. Peter, V. A. Squire, and M. H. Meyla

PAin SoluTions In the Emergency Setting (PASTIES)--patient controlled analgesia versus routine care in emergency department patients with non-traumatic abdominal pain: randomised trial

© Smith et al 2015. OBJECTIVE: To determine whether patient controlled analgesia (PCA) is better than routine care in providing effective analgesia for patients presenting to emergency departments with moderate to severe non-traumatic abdominal pain.DESIGN: Pragmatic, multicentre, parallel group, randomised controlled trialSETTING: Five English hospitals.PARTICIPANTS: 200 adults (66% (n=130) female), aged 18 to 75 years, who presented to the emergency department requiring intravenous opioid analgesia for the treatment of moderate to severe non-traumatic abdominal pain and were expected to be admitted to hospital for at least 12 hours.INTERVENTIONS: Patient controlled analgesia or nurse titrated analgesia (treatment as usual).MAIN OUTCOME MEASURES: The primary outcome was total pain experienced over the 12 hour study period, derived by standardised area under the curve (scaled from 0 to 100) of each participant's hourly pain scores, captured using a visual analogue scale. Pre-specified secondary outcomes included total morphine use, percentage of study period in moderate or severe pain, percentage of study period asleep, length of hospital stay, and satisfaction with pain management.RESULTS: 196 participants were included in the primary analyses (99 allocated to PCA and 97 to treatment as usual). Mean total pain experienced was 35.3 (SD 25.8) in the PCA group compared with 47.3 (24.7) in the treatment as usual group. The adjusted between group difference was 6.3 (95% confidence interval 0.7 to 11.9). Participants in the PCA group received significantly more morphine (mean 36.1 (SD 22.4) v 23.6 (13.1) mg; mean difference 12.3 (95% confidence interval 7.2 to 17.4) mg), spent less of the study period in moderate or severe pain (32.6% v 46.9%; mean difference 14.5% (5.6% to 23.5%)), and were more likely to be perfectly or very satisfied with the management of their pain (83% (73/88) v 66% (57/87); adjusted odds ratio 2.56 (1.25 to 5.23)) in comparison with participants in the treatment as usual group.CONCLUSIONS: Significant reductions in pain can be achieved by PCA compared with treatment as usual in patients presenting to the emergency department with non-traumatic abdominal pain. Trial registration European Clinical Trials Database EudraCT2011-000194-31; Current Controlled Trials ISRCTN25343280