Breaking Down Silos in the Workplace: A Framework to Foster Collaboration.

INTRODUCTION: Employees are often placed within an organization based on their respective roles or duties, which can lead to vertical and horizontal organizational silos. Organizational silos may restrict information, resources, and stymie progress and innovation. This analysis presents a framework to mitigate silos and overcome communication barriers within an organization by increasing collaboration. METHODS: The project team examined results from the Centers for Disease Control and Prevention (CDC), the National Center for Chronic Disease Prevention and Health Promotion (NCCDPHP) 2020 Employee Viewpoint Survey Results and conducted 19 key informant discussions with NCCDPHP employees. Participants were asked to provide feedback on (1) understanding silos in the workplace and (2) best practices for reducing silos and fostering collaboration. A thematic analysis was conducted to understand organizational silos, the motivation to reduce silos, and identify best practices and strategies. RESULTS: Respondents felt that siloing exists at the division and branch levels; however, 95% of respondents were motivated to reduce silos. Fifty-eight percent of respondents identified that institutional factors such as the organizational structure (n = 8) and red tape/bureaucracy (n = 3) contribute to siloing. Additional behaviors and actions that perpetuate silos were identified, and efforts to reduce silos were categorized to propose a model: Framework to Foster Collaboration for improving organizational collaborative efforts. DISCUSSION AND CONCLUSION: Key themes included inclusion, shared goals and vision, bi-directional communication, and relationship building and developing trust as critical elements for improving collaboration and creating synergy across teams in efforts to reduce silos in the workplace.

Tandem High-Pressure Crystallography-Optical Spectroscopy Unpacks Noncovalent Interactions of Piezochromic Fluorescent Molecular Rotors.

To develop luminescent molecular materials with predictable and stimuli-responsive emission, it is necessary to correlate changes in their geometries, packing structures, and noncovalent interactions with the associated changes in their optical properties. Here, we demonstrate that high-pressure single-crystal X-ray diffraction can be combined with high-pressure UV-visible absorption and fluorescence emission spectroscopies to elucidate how subtle changes in structure influence optical outputs. A piezochromic aggregation-induced emitter, sym-heptaphenylcycloheptatriene (Ph7C7H), displays bathochromic shifts in its absorption and emission spectra at high pressure. Parallel X-ray measurements identify the pressure-induced changes in specific phenyl-phenyl interactions responsible for the piezochromism. Pairs of phenyl rings from neighboring molecules approach the geometry of a stable benzene dimer, while conformational changes alter intramolecular phenyl-phenyl interactions correlated with a relaxed excited state. This tandem crystallographic and spectroscopic analysis provides insights into how subtle structural changes relate to the photophysical properties of Ph7C7H and could be applied to a library of similar compounds to provide general structure-property relationships in fluorescent organic molecules with rotor-like geometries.

Advanced airway management and drug-assisted intubation skills in an advanced critical care practitioner team.

INTRODUCTION: Airway management, including endotracheal intubation, is one of the cornerstones of care of critically ill patients. Internationally, health professionals from varying backgrounds deliver endotracheal intubation as part of their critical care role. This article considers the development of airway management skills within a single advanced critical care practitioner (ACCP) team and uses case series data to analyse the safety profile in performing this aspect of critical care. Skills were acquired during and after the ACCP training pathway. A combination of theoretical teaching, theatre experience, simulation and work-based practice was used. Case series data of all critical care intubations by ACCPs were collected. Audit results: Data collection identified 675 intubations carried out by ACCPs, 589 of those being supervised, non-cardiac arrest intubations requiring drugs. First pass success was achieved in 89.6% of cases. A second intubator was required in 4.3% of cases. Some form of complication was experienced by 42.3% of patients; however, the threshold for complications was set at a low level. CONCLUSIONS: This ACCP service developed a process to acquire advanced airway management skills including endotracheal intubation. Under medical supervision, ACCPs delivered advanced airway management achieving a first pass success rate of 89.6%, which compares favourably with both international and national success rates. Although complications were experienced in 48.3% of patients, when similar complication cut-offs are compared with published data, ACCPs also matched favourably.

Photophysics of Azobenzene Constrained in a UiO Metal-Organic Framework: Effects of Pressure, Solvation and Dynamic Disorder.

Photophysical studies of chromophoric linkers in metal-organic frameworks (MOFs) are undertaken commonly in the context of sensing applications, in search of readily observable changes of optical properties in response to external stimuli. The advantages of the MOF construct as a platform for investigating fundamental photophysical behaviour have been somewhat overlooked. The linker framework offers a unique environment in which the chromophore is geometrically constrained and its structure can be determined crystallographically, but it exists in spatial isolation, unperturbed by inter-chromophore interactions. Furthermore, high-pressure studies enable the photophysical consequences of controlled, incremental changes in local environment or conformation to be observed and correlated with structural data. This approach is demonstrated in the present study of the trans-azobenzene chromophore, constrained in the form of the 4,4'-azobenzenedicarboxylate (abdc) linker, in a UiO topology framework. Previously unobserved effects of pressure-induced solvation and conformational distortion on the lowest energy, nπ* transition are reported, and interpreted the light of crystallographic data. It was found that trans-azobenzene remains non-fluorescent (with a quantum yield less than 10-4 ) despite the prevention of trans-cis isomerization by the constraining MOF structure. We propose that efficient non-radiative decay is mediated by the local, pedal-like twisting of the azo group that is evident as dynamic disorder in the crystal structure.

Evaluation of the safety of inter-hospital transfers of critically ill patients led by advanced critical care practitioners.

INTRODUCTION: Ten thousand inter-hospital transfers of critically ill adults take place annually in the UK. Studies highlight deficiencies in experience and training of staff, equipment, stabilisation before departure, and logistical difficulties. This article is a quality improvement review of an advanced critical care practitioner (ACCP)-led inter-hospital transfer service. METHODS: The tool Standards for Quality Improvement Reporting Excellence was used as the format for the review, combined with clinical audit of advanced critical care practitioner-led transfers over a period of more than 3 years. RESULTS: The transfer service has operated for 8 years; ACCPs conducted 934 critical care transfers of mechanically ventilated patients, including 286 inter-hospital transfers, between January 2017 and September 2020. The acuity of transfer patients was high, 82.2% required support of more than one organ, 49% required more than 50% oxygen. Uneventful transfer occurred in 81.4% of cases; the most common patient-related complication being hypotension, logistical issues were responsible for half of the complications. CONCLUSION: This quality improvement project provides an example of safe and effective advanced practice in an area that is traditionally a medically led domain. ACCPs can provide an alternative process of care for critically ill adults who require external transfer, and a benchmark for audit and quality improvement.

Transcriptome analysis of the uterus of hens laying eggs differing in cuticle deposition.

BACKGROUND: Avian eggs have a proteinaceous cuticle. The quantity of cuticle varies and the deposition of a good cuticle in the uterus (Shell-gland) prevents transmission of bacteria to the egg contents. RESULTS: To understand cuticle deposition, uterus transcriptomes were compared between hens with i) naturally good and poor cuticle and, ii) where manipulation of the hypothalamo-pituitary-gonadal-oviduct axis produced eggs with or without cuticle. The highest expressed genes encoded eggshell matrix and cuticle proteins, e.g. MEPE (OC-116), BPIFB3 (OVX-36), RARRES1 (OVX-32), WAP (OVX-25), and genes for mitochondrial oxidative phosphorylation, active transport and energy metabolism. Expression of a number of these genes differed between hens laying eggs with or without cuticle. There was also a high expression of clock genes. PER2, CRY2, CRY1, CLOCK and BMAL1 were differentially expressed when cuticle deposition was prevented, and they also changed throughout the egg formation cycle. This suggests an endogenous clock in the uterus may be a component of cuticle deposition control. Cuticle proteins are glycosylated and glycosaminoglycan binding genes had a lower expression when cuticle proteins were deposited on the egg. The immediate early genes, JUN and FOS, were expressed less when the cuticle had not been deposited and changed over the egg formation cycle, suggesting they are important in oviposition and cuticle deposition. The uterus transcriptome of hens with good and poor cuticle deposition did not differ. CONCLUSIONS: We have gained insights into the factors that can affect the production of the cuticle especially clock genes and immediate early genes. We have demonstrated that these genes change their expression over the period of eggshell formation supporting their importance. The lack of differences in expression between the uterus of hens laying eggs with the best and worse cuticle suggest the genetic basis of the trait may lie outside the oviduct.

Correlating Pressure-Induced Emission Modulation with Linker Rotation in a Photoluminescent MOF.

Conformational changes of linker units in metal-organic frameworks (MOFs) are often responsible for gate-opening phenomena in selective gas adsorption and stimuli-responsive optical and electrical sensing behaviour. Herein, we show that pressure-induced bathochromic shifts in both fluorescence emission and UV/Vis absorption spectra of a two-fold interpenetrated Hf MOF, linked by 1,4-phenylene-bis(4-ethynylbenzoate) ligands (Hf-peb), are induced by rotation of the central phenyl ring of the linker, from a coplanar arrangement to a twisted, previously unseen conformer. Single-crystal X-ray diffraction, alongside in situ fluorescence and UV/Vis absorption spectroscopies, measured up to 2.1 GPa in a diamond anvil cell on single crystals, are in excellent agreement, correlating linker rotation with modulation of emission. Topologically isolating the 1,4-phenylene-bis(4-ethynylbenzoate) units within a MOF facilitates concurrent structural and spectroscopic studies in the absence of intermolecular perturbation, allowing characterisation of the luminescence properties of a high-energy, twisted conformation of the previously well-studied chromophore. We expect the unique environment provided by network solids, and the capability of combining crystallographic and spectroscopic analysis, will greatly enhance understanding of luminescent molecules and lead to the development of novel sensors and adsorbents.

Dynamic and static quenching of 2-aminopurine fluorescence by the natural DNA nucleotides in solution.

2-aminopurine (2AP) is a responsive fluorescent base analogue that is used widely as a probe of the local molecular environment in DNA. The ability of 2AP to report changes in local conformation and base-stacking interactions arises from the efficient quenching of its fluorescence by the natural DNA bases. However, the mechanism of this inter-base quenching remains imperfectly understood. Two previous studies of the collisional quenching of 2AP by the natural bases, in different buffer solutions, showed that dynamic quenching efficiency depends on the identity of the natural base, but disagreed on the relative quenching efficiencies of the bases. We report a comprehensive investigation of inter-base quenching of 2AP by the natural nucleoside monophosphates (NMPs), replicating the buffer conditions used in the previous studies. Using time-resolved fluorescence measurements to distinguish between dynamic and static quenching, we find that the dynamic quenching rate constants of the different bases show a consistent trend across both buffers, and this is in line with a charge-transfer mechanism. Time-resolved measurements also provide insight into static quenching, revealing formation of 2AP-NMP ground-state complexes in which 2AP displays a very short fluorescence lifetime, comparable to that seen in oligonucleotides. In these complexes, the dependence of the rate of quenching on the partner base also supports a charge-transfer mechanism.

The provision of central venous access, transfer of critically ill patients and advanced airway management.: Are advanced critical care practitioners safe and effective?

Advanced critical care practitioners are a new and growing component of the critical care multidisciplinary team in the United Kingdom. This audit considers the safety profile of advanced critical care practitioners in the provision of central venous catheterisation and transfer of ventilated critical care patients without direct supervision and supervised drug assisted intubation of critically ill patients. The audit showed that advanced critical care practitioners can perform central venous cannulation, transfer of critically ill ventilated patients and intubation with parity to published UK literature.

Single-Molecule Detection of a Fluorescent Nucleobase Analogue via Multiphoton Excitation.

The ability to routinely detect fluorescent nucleobase analogues at the single-molecule level would create a wealth of opportunities to study nucleic acids. We report the multiphoton-induced fluorescence and single-molecule detection of a dimethylamine-substituted extended-6-aza-uridine (DMAthaU). We show that DMAthaU can exist in a highly fluorescent form, emitting strongly in the visible region (470-560 nm). Using pulse-shaped broadband Ti:sapphire laser excitation, DMAthaU undergoes two-photon (2P) absorption at low excitation powers, switching to three-photon (3P) absorption at high incident intensity. The assignment of a 3P process is supported by cubic response calculations. Under both 2P and 3P excitation, the single-molecule brightness was over an order of magnitude higher than reported previously for any fluorescent base analogue, which facilitated the first single-molecule detection of an emissive nucleoside with multiphoton excitation.

Genetic variation and potential for genetic improvement of cuticle deposition on chicken eggs.

BACKGROUND: The cuticle is an invisible glycosylated protein layer that covers the outside of the eggshell and forms a barrier to the transmission of microorganisms. Cuticle-specific staining and in situ absorbance measurements have been used to quantify cuticle deposition in several pure breeds of chicken. For brown eggs, a pre-stain and a post-stain absorbance measurement is required to correct for intrinsic absorption by the natural pigment. For white eggs, a post-stain absorbance measurement alone is sufficient to estimate cuticle deposition. The objective of the research was to estimate genetic parameters and provide data to promote adoption of the technique to increase cuticle deposition and reduce vertical transmission of microorganisms. RESULTS: For all pure breeds examined here, i.e. Rhode Island Red, two White Leghorns, White Rock and a broiler breed, the estimate of heritability for cuticle deposition from a meta-analysis was moderately high (0.38 ± 0.04). In the Rhode Island Red breed, the estimate of the genetic correlation between measurements recorded at early and late times during the egg-laying period was ~ 1. There was no negative genetic correlation between cuticle deposition and production traits. Estimates of the genetic correlation of cuticle deposition with shell color ranged from negative values or 0 in brown-egg layers to positive values in white- or tinted-egg layers. Using the intrinsic fluorescence of tryptophan in the cuticle proteins to quantify the amount of cuticle deposition failed because of complex quenching processes. Tryptophan fluorescence intensity at 330 nm was moderately heritable, but there was no evidence of a non-zero genetic correlation with cuticle deposition. This was complicated furthermore by a negative genetic correlation of fluorescence with color in brown eggs, due to the quenching of tryptophan fluorescence by energy transfer to protoporphyrin pigment. We also confirmed that removal of the cuticle increased reflection of ultraviolet wavelengths from the egg. CONCLUSIONS: These results provide additional evidence for the need to incorporate cuticle deposition into breeding programs of egg- and meat-type birds in order to reduce vertical and horizontal transmission of potentially pathogenic organisms and to help improve biosecurity in poultry.

Cuticle deposition improves the biosecurity of eggs through the laying cycle and can be measured on hatching eggs without compromising embryonic development.

The cuticle is part of the egg's natural defense and it can be improved by genetic selection. Prior to adoption of this measurement in breeding programs, questions that need to be addressed are whether improved cuticle deposition will result in a reduced risk of eggs becoming contaminated and whether selection for this trait will have any unintended consequences on the incubation process. Bacterial penetration experiments were carried out using eggs from a pedigree line of broiler breeders (BB) and Rhode Island Red (RIR) layers. Within the natural variation in cuticle deposition in each line, a good cuticle was shown to reduce an egg's susceptibility to penetration by Escherichia coli (BB, P = 0.023) and Salmonella typhimurium (RIR, P < 0.001). Deglycosylation of cuticle proteins had little effect on their antimicrobial activity. The effect of bird age on cuticle deposition was also examined. Shell color decreased with age as anticipated; however, we found no evidence that cuticle deposition decreases with age, at least up to 50 wk. A thicker cuticle could affect the water vapor conductance (WPC) of hatching eggs. The WPC of eggs was, therefore, measured on eggs selected from the top and tail of the cuticle distribution, this time in a Lohmann Selected Leghorn (LSL) pedigree line. Broiler breeder eggs were also tested. No evidence of a relationship between cuticle deposition and WPC was found for LSL or BB eggs. Cuticle deposition measurements require eggs to be stained. Here, we show that this has no adverse effect on embryo development at d 12 of incubation. Thus, we conclude that cuticle deposition is important in preventing bacterial penetration of eggs in genetically divergent breeds of chicken and that the measurement can be practically incorporated into breeding programs. This will contribute to improving the biosecurity of eggs by reducing vertical and horizontal transmission of potentially zoonotic and pathogenic organisms from parent to offspring.

Pulse-shaped two-photon excitation of a fluorescent base analogue approaches single-molecule sensitivity.

Fluorescent nucleobase analogues (FBAs) have many desirable features in comparison to extrinsic fluorescent labels, but they are yet to find application in ultrasensitive detection. Many of the disadvantages of FBAs arise from their short excitation wavelengths (often in the ultraviolet), making two-photon excitation a potentially attractive approach. Pentacyclic adenine (pA) is a recently developed FBA that has an exceptionally high two-photon brightness. We have studied the two-photon-excited fluorescence properties of pA and how they are affected by incorporation in DNA. We find that pA is more photostable under two-photon excitation than via resonant absorption. When incorporated in an oligonucleotide, pA has a high two-photon cross section and emission quantum yield, varying with sequence context, resulting in the highest reported brightness for such a probe. The use of a two-photon microscope with ultrafast excitation and pulse shaping has allowed the detection of pA-containing oligonucleotides in solution with a limit of detection of ∼5 molecules, demonstrating that practical single-molecule detection of FBAs is now within reach.

Pentacyclic adenine: a versatile and exceptionally bright fluorescent DNA base analogue.

Emissive base analogs are powerful tools for probing nucleic acids at the molecular level. Herein we describe the development and thorough characterization of pentacyclic adenine (pA), a versatile base analog with exceptional fluorescence properties. When incorporated into DNA, pA pairs selectively with thymine without perturbing the B-form structure and is among the brightest nucleobase analogs reported so far. Together with the recently established base analog acceptor qAnitro, pA allows accurate distance and orientation determination via Förster resonance energy transfer (FRET) measurements. The high brightness at emission wavelengths above 400 nm also makes it suitable for fluorescence microscopy, as demonstrated by imaging of single liposomal constructs coated with cholesterol-anchored pA-dsDNA, using total internal reflection fluorescence microscopy. Finally, pA is also highly promising for two-photon excitation at 780 nm, with a brightness (5.3 GM) that is unprecedented for a base analog.

Analysis of time-correlated single photon counting data: a comparative evaluation of deterministic and probabilistic approaches.

We review various methods for analysing time-resolved fluorescence data acquired using the time-correlated single photon counting method in an attempt to evaluate their benefits and limitations. We have applied these methods to both experimental and simulated data. The relative merits of using deterministic approaches, such as the commonly used iterative reconvolution method, and probabilistic approaches, such as the smoothed exponential series method, the maximum entropy method and recently proposed basis pursuit denoising (compressed sensing) method, are outlined. In particular, we show the value of using multiple methods to arrive at the most appropriate choice of model. We show that the use of probabilistic analysis methods can indicate whether a discrete component or distribution analysis provides the better representation of the data.

Understanding avian egg cuticle formation in the oviduct: a study of its origin and deposition.

The cuticle is a unique invisible oviduct secretion that protects avian eggs from bacterial penetration through gas exchange pores. Despite its importance, experimental evidence is lacking for where, when, and what is responsible for its deposition. By using knowledge about the ovulatory cycle and oviposition, we have manipulated cuticle deposition to obtain evidence on these key points. Cuticle deposition was measured using staining and spectrophotometry. Experimental evidence supports the location of cuticle deposition to be the shell gland pouch (uterus), not the vagina, and the time of deposition to be within the final hour before oviposition. Oviposition induced by arginine vasotocin or prostaglandin, the penultimate and ultimate factors for the induction of oviposition, produces an egg with no cuticle; therefore, these factors are not responsible for cuticle secretion. Conversely, oviposition induced by GNRH, which mimics the normal events of ovulation and oviposition, results in a normal cuticle. There is no evidence that cuticle deposition differs at the end of a clutch and, therefore, there is no evidence that the ovulatory surge of progesterone affects cuticle deposition. Overall, the results demonstrate that the cuticle is a specific secretion and is not merely an extension of the organic matrix of the shell. Cuticle deposition was found to be reduced by an environmental stressor, and there is no codependence of the deposition of pigment and cuticle. Defining the basic facts surrounding cuticle deposition will help reduce contamination of hen's eggs and increase understanding of the strategies birds use to protect their eggs.

A DFT study of 2-aminopurine-containing dinucleotides: prediction of stacked conformations with B-DNA structure.

The fluorescence properties of dinucleotides incorporating 2-aminopurine (2AP) suggest that the simplest oligonucleotides adopt conformations similar to those found in duplex DNA. However, there is a lack of structural data for these systems. We report a density functional theory (DFT) study of the structures of 2AP-containing dinucleotides (deoxydinucleoside monophosphates), including full geometry optimisation of the sugar-phosphate backbone. Our DFT calculations employ the M06-2X functional for reliable treatment of dispersion interactions and include implicit aqueous solvation. Dinucleotides with 2AP in the 5'-position and each of the natural bases in the 3'-position are examined, together with the analogous 5'-adenine-containing systems. Computed structures are compared in detail with typical B-DNA base-step parameters, backbone torsional angles and sugar pucker, derived from crystallographic data. We find that 2AP-containing dinucleotides adopt structures that closely conform to B-DNA in all characteristic parameters. The structures of 2AP-containing dinucleotides closely resemble those of their adenine-containing counterparts, demonstrating the fidelity of 2AP as a mimic of the natural base. As a first step towards exploring the conformational heterogeneity of dinucleotides, we also characterise an imperfectly stacked conformation and one in which the bases are completely unstacked.

A bend, flip and trap mechanism for transposon integration.

Cut-and-paste DNA transposons of the mariner/Tc1 family are useful tools for genome engineering and are inserted specifically at TA target sites. A crystal structure of the mariner transposase Mos1 (derived from Drosophila mauritiana), in complex with transposon ends covalently joined to target DNA, portrays the transposition machinery after DNA integration. It reveals severe distortion of target DNA and flipping of the target adenines into extra-helical positions. Fluorescence experiments confirm dynamic base flipping in solution. Transposase residues W159, R186, F187 and K190 stabilise the target DNA distortions and are required for efficient transposon integration and transposition in vitro. Transposase recognises the flipped target adenines via base-specific interactions with backbone atoms, offering a molecular basis for TA target sequence selection. Our results will provide a template for re-designing mariner/Tc1 transposases with modified target specificities.

Analysis of the Thermal Degradation of the Individual Anthocyanin Compounds of Black Carrot (Daucus carota L.): A New Approach Using High-Resolution Proton Nuclear Magnetic Resonance Spectroscopy.

The black carrot dye is a mixture of cyanidin molecules, the nuclear magnetic resonance (NMR) spectrum of which shows a highly overlapped aromatic region. In this study, the (1)H NMR (800 MHz) aromatic chemical shifts of the mixture were fully assigned by overlaying them with the characterized (1)H NMR chemical shifts of the separated compounds. The latter were isolated using reverse-phase high-performance liquid chromatography (RP-HPLC), and their chemical shifts were identified using (1)H and two-dimensional (2D) correlation spectroscopy (COSY) NMR spectroscopy. The stability of the black carrot mixture to heat exposure was investigated at pH 3.6, 6.8, and 8.0 by heat-treating aqueous solutions at 100 °C and the powdered material at 180 °C. From integration of high-resolution (1)H NMR spectra, it was possible to follow the relative degradation of each compound, offering advantages over the commonly used ultraviolet/visible (UV/vis) and HPLC approaches. UV/vis spectroscopy and CIE color measurements were used to determine thermally induced color changes, under normal cooking conditions.

2-Aminopurine as a fluorescent probe of DNA conformation and the DNA-enzyme interface.

Nearly 50 years since its potential as a fluorescent base analogue was first recognized, 2-aminopurine (2AP) continues to be the most widely used fluorescent probe of DNA structure and the perturbation of that structure by interaction with enzymes and other molecules. In this review, we begin by considering the origin of the dramatic and intriguing difference in photophysical properties between 2AP and its structural isomer, adenine; although 2AP differs from the natural base only in the position of the exocyclic amine group, its fluorescence intensity is one thousand times greater. We then discuss the mechanism of interbase quenching of 2AP fluorescence in DNA, which is the basis of its use as a conformational probe but remains imperfectly understood. There are hundreds of examples in the literature of the use of changes in the fluorescence intensity of 2AP as the basis of assays of conformational change; however, in this review we will consider in detail only a few intensity-based studies. Our primary aim is to highlight the use of time-resolved fluorescence measurements, and the interpretation of fluorescence decay parameters, to explore the structure and dynamics of DNA. We discuss the salient features of the fluorescence decay of 2AP when incorporated in DNA and review the use of decay measurements in studying duplexes, single strands and other structures. We survey the use of 2AP as a probe of DNA-enzyme interaction and enzyme-induced distortion, focusing particularly on its use to study base flipping and the enhanced mechanistic insights that can be gained by a detailed analysis of the decay parameters, rather than merely monitoring changes in fluorescence intensity. Finally we reflect on the merits and shortcomings of 2AP and the prospects for its wider adoption as a fluorescence-decay-based probe.