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Point-of-care antibiotic susceptibility test for urinary tract infections.

This study investigated the experiences and perceptions of Saudi patients with insomnia disorder. A cross-sectional design study (July 2019) using a self-administered questionnaire survey. Sixty-eight patients with insomnia disorder participated (53% male, 47% female; 60% were younger than 45 years and a majority had a university-degree education level). Of the respondents, 64.7% were dissatisfied with their knowledge of insomnia and 97% wanted more information about its treatment. Approximately 59% used prescribed sleep medicines from the first instance and only 6% were referred to a psychologist for counselling. Benzodiazepines and Z-drugs were prescribed for 16% and 19% of respondents, respectively. A majority of respondents were given antihistamines or over-the-counter herbs. While 60% of patients used medications for long periods, 87% were not involved in treatment decisions. For those on long-term use, 80% were not seen regularly by doctors and 93% were provided with no plan for medicine discontinuations. More than 60% of respondents said that sleeping pills were best for treating insomnia and 31% believed that they are safe and would continue taking them. More than 45% would not prefer non-pharmacological treatments (e.g., counselling). Responses to the survey indicate a lack of knowledge about insomnia disorder and its treatment among Saudi patients. Prescribing hypnotics is the most common treatment, non-pharmacological treatments are seldom used, and responses indicated inappropriate and long-term use of hypnotics. Patients should be better educated about insomnia and its treatment. Physicians should consider using non-pharmacological treatments and the Saudi Ministry of Health should develop country-specific guidelines.

Antibiotic resistance is a major global challenge. Although microfluidic antibiotic susceptibility tests (AST) offer great potential for rapid and portable testing to inform correct antibiotic selection, the impact of miniaturisation on broth microdilution (BMD) is not fully understood. \ud We developed a 10-plex microcapillary based broth microdilution using resazurin as a colorimetric indicator for bacterial growth. Each capillary had a 1 microlitre capillary volume, 100 times smaller than microplate broth microdilution. The microcapillary BMD was compared to an in-house standard microplate AST and commercial Vitek 2 system. \ud When tested with 25 uropathogenic isolates (20 Escherichia coli and 5 Klebsiella pneumoniae) and 2 reference E. coli, these devices gave 96.1% (441/459 isolate/antibiotic combinations) categorical agreement, across 17 therapeutically beneficial antibiotics, compared to in-house microplate BMD with resazurin. A further 99 (50 E. coli and 49 K. pneumoniae) clinical isolates were tested against 10 antibiotics and showed 92.3% categorical (914/990 isolate/antibiotic combinations) compared to the Vitek 2 measurements. \ud These microcapillary tests showed excellent analytical agreement to existing AST methods. Furthermore, the small size and simple colour change can be recorded using a smartphone camera or it is feasible to follow growth kinetics using very simple, low-cost readers. The test strips used here are produced in large batches, allowing hundreds of multiplex tests to be made and tested rapidly. Demonstrating performance of miniaturised broth microdilution with clinical isolates paves the way for wider use of microfluidic AST.

Counting viable bacterial cells and functional bacteriophage is fundamental to microbiology underpinning research, surveillance, biopharmaceuticals and diagnostics. Colony forming unit (CFU) and plaque forming unit (PFU) counting still requires slow and laborious solid culture on agar in Petri dishes or plates. Here, we show that dip-stick microfluidic strips can be used without growth indicator dye for rapid and simple CFU ml-1 and PFU ml-1 measurement. We demonstrate for the first time that fluoropolymer microcapillaries combined with digital imaging allow bacteriophage plaques to be counted rapidly in a dip-and-test format. The microfluidic length scales offer a linear 1-dimensional alternative to a 2D solid agar medium surface, with colonies or plaques clearly visible as "dashes" or "gaps". An inexpensive open source darkfield biosensor system using Raspberry Pi imaging permits label-free detection and counting of colonies or plaques within 4-8 hours in a linear, liquid matrix within ∼200 μm inner diameter microcapillaries. We obtained full quantitative agreement between 1D microfluidic colony counting in dipsticks versus conventional 2D solid agar Petri dish plates for S. aureus and E. coli, and for T2 phage and phage K, but up to 6 times faster. Time-lapse darkfield imaging permitted detailed kinetic analysis of colony growth in the microcapillaries, providing new insight into microfluidic microbiology and colony growth, not possible with Petri dishes. Surprisingly, whilst E. coli colonies appeared earlier, subsequent colony expansion was faster along the microcapillaries for S. aureus. This may be explained by the microenvironment offered for 1D colony growth within microcapillaries, linked to a mass balance between nutrient (glucose) diffusion and bacterial growth kinetics. Counting individual colonies in liquid medium was not possible for motile strains that spread rapidly along the capillary, however inclusion of soft agar inhibited spreading, making this new simple dip-and-test counting method applicable to both motile and non-motile bacteria. Label-free dipstick colony and plaque counting has potential for many analytical microbial tasks, and the innovation of 1D colony counting has relevance to other microfluidic microbiology.

The diverse cerebral consequences of preterm birth create significant challenges for understanding pathogenesis or predicting later outcome. Instead of focusing on describing effects common to the group, comparing individual infants against robust normative data offers a powerful alternative to study brain maturation. Here we used Gaussian process regression to create normative curves characterising brain volumetric development in 274 term-born infants, modelling for age at scan and sex. We then compared 89 preterm infants scanned at termequivalent age to these normative charts, relating individual deviations from typical volumetric development to perinatal risk factors and later neurocognitive scores. To test generalisability, we used a second independent dataset comprising of 253 preterm infants scanned using different acquisition parameters and scanner. We describe rapid, non-uniform brain growth during the neonatal period. In both preterm cohorts, cerebral atypicalities were widespread, often multiple, and varied highly between individuals. Deviations from normative development were associated with respiratory support, nutrition, birth weight, and later neurocognition, demonstrating their clinical relevance. Group-level understanding of the preterm brain disguise a large degree of individual differences. We provide a method and normative dataset that offer a more precise characterisation of the cerebral consequences of preterm birth by profiling the individual neonatal brain.

In the mature brain, structural and functional 'fingerprints' of brain connectivity can be used to identify the uniqueness of an individual. However, whether the characteristics that make a given brain distinguishable from others already exist at birth remains unknown. Here, we used neuroimaging data from the developing Human Connectome Project (dHCP) of preterm born neonates who were scanned twice during the perinatal period to assess the developing brain fingerprint. We found that 62% of the participants could be identified based on the congruence of the later structural connectome to the initial connectivity matrix derived from the earlier timepoint. In contrast, similarity between functional connectomes of the same subject at different time points was low. Only 10% of the participants showed greater self-similarity in comparison to self-to-other-similarity for the functional connectome. These results suggest that structural connectivity is more stable in early life and can represent a potential connectome fingerprint of the individual: a relatively stable structural connectome appears to support a changing functional connectome at a time when neonates must rapidly acquire new skills to adapt to their new environment. ispartof: DEVELOPMENTAL COGNITIVE NEUROSCIENCE vol:55 ispartof: location:Netherlands status: published

Over the past few decades, neuroimaging has become a ubiquitous tool in basic research and clinical studies of the human brain. However, no reference standards currently exist to quantify individual differences in neuroimaging metrics over time, in contrast to growth charts for anthropometric traits such as height and weight1. Here we assemble an interactive open resource to benchmark brain morphology derived from any current or future sample of MRI data ( http://www.brainchart.io/ ). With the goal of basing these reference charts on the largest and most inclusive dataset available, acknowledging limitations due to known biases of MRI studies relative to the diversity of the global population, we aggregated 123,984 MRI scans, across more than 100 primary studies, from 101,457 human participants between 115 days post-conception to 100 years of age. MRI metrics were quantified by centile scores, relative to non-linear trajectories2 of brain structural changes, and rates of change, over the lifespan. Brain charts identified previously unreported neurodevelopmental milestones3, showed high stability of individuals across longitudinal assessments, and demonstrated robustness to technical and methodological differences between primary studies. Centile scores showed increased heritability compared with non-centiled MRI phenotypes, and provided a standardized measure of atypical brain structure that revealed patterns of neuroanatomical variation across neurological and psychiatric disorders. In summary, brain charts are an essential step towards robust quantification of individual variation benchmarked to normative trajectories in multiple, commonly used neuroimaging phenotypes., R.A.I.B. was supported by a British Academy Postdoctoral fellowship and by the Autism Research Trust. J. Seidlitz was supported by NIMH T32MH019112-29 and K08MH120564. S.R.W. was funded by UKRI Medical Research Council MC_UU_00002/2 and was supported by the NIHR Cambridge Biomedical Research Centre (BRC-1215-20014). E.T.B. was supported by an NIHR Senior Investigator award and the Wellcome Trust collaborative award for the Neuroscience in Psychiatry Network. A.F.A.-B. was supported by NIMH K08MH120564. Data were curated and analysed using a computational facility funded by an MRC research infrastructure award (MR/M009041/1) to the School of Clinical Medicine, University of Cambridge and supported by the mental health theme of the NIHR Cambridge Biomedical Research Centre.

Purpose Diffusion magnetic resonance imaging (dMRI) studies report altered white matter (WM) development in preterm infants. Neurite orientation dispersion and density imaging (NODDI) metrics provide more realistic estimations of neurite architecture in vivo compared with standard diffusion tensor imaging (DTI) metrics. This study investigated microstructural maturation of WM in preterm neonates scanned between 25 and 45 weeks postmenstrual age (PMA) with normal neurodevelopmental outcomes at 2 years using DTI and NODDI metrics. Methods Thirty-one neonates (n = 17 male) with median (range) gestational age (GA) 32+1 weeks (24+2–36+4) underwent 3 T brain MRI at median (range) post menstrual age (PMA) 35+2 weeks (25+3–43+1). WM tracts (cingulum, fornix, corticospinal tract (CST), inferior longitudinal fasciculus (ILF), optic radiations) were delineated using constrained spherical deconvolution and probabilistic tractography in MRtrix3. DTI and NODDI metrics were extracted for the whole tract and cross-sections along each tract to assess regional development. Results PMA at scan positively correlated with fractional anisotropy (FA) in the CST, fornix and optic radiations and neurite density index (NDI) in the cingulum, CST and fornix and negatively correlated with mean diffusivity (MD) in all tracts. A multilinear regression model demonstrated PMA at scan influenced all diffusion measures, GA and GAxPMA at scan influenced FA, MD and NDI and gender affected NDI. Cross-sectional analyses revealed asynchronous WM maturation within and between WM tracts.). Conclusion We describe normal WM maturation in preterm neonates with normal neurodevelopmental outcomes. NODDI can enhance our understanding of WM maturation compared with standard DTI metrics alone.

BACKGROUND AND PURPOSE: Brain MR imaging at term-equivalent age is a useful tool to define brain injury in preterm infants. We report pragmatic clinical radiological assessment of images from a large unselected cohort of preterm infants imaged at term and document the spectrum and frequency of acquired brain lesions and their relation to outcomes at 20 months. MATERIALS AND METHODS: Infants born at

A range of biosensing techniques including immunoassays are routinely used for quantitation of analytes in biological samples and available in a range of formats, from centralized lab testing (e.g., microplate enzyme-linked immunosorbent assay (ELISA)) to automated point-of-care (POC) and lateral flow immunochromatographic tests. High analytical performance is intrinsically linked to the use of a sequence of reagent and washing steps, yet this is extremely challenging to deliver at the POC without a high level of fluidic control involving, e.g., automation, fluidic pumping, or manual fluid handling/pipetting. Here we introduce a microfluidic siphon concept that conceptualizes a multistep ″dipstick″ for quantitative, enzymatically amplified immunoassays using a strip of microporous or microbored material. We demonstrated that gravity-driven siphon flow can be realized in single-bore glass capillaries, a multibored microcapillary film, and a glass fiber porous membrane. In contrast to other POC devices proposed to date, the operation of the siphon is only dependent on the hydrostatic liquid pressure (gravity) and not capillary forces, and the unique stepwise approach to the delivery of the sample and immunoassay reagents results in zero dead volume in the device, no reagent overlap or carryover, and full start/stop fluid control. We demonstrated applications of a 10-bore microfluidic siphon as a portable ELISA system without compromised quantitative capabilities in two global diagnostic applications: (1) a four-plex sandwich ELISA for rapid smartphone dengue serotype identification by serotype-specific dengue virus NS1 antigen detection, relevant for acute dengue fever diagnosis, and (2) quantitation of anti-SARS-CoV-2 IgG and IgM titers in spiked serum samples. Diagnostic siphons provide the opportunity for high-performance immunoassay testing outside sophisticated laboratories, meeting the rapidly changing global clinical and public health needs.

The emerging field of geometric deep learning extends the application of convolutional neural networks to irregular domains such as graphs, meshes and surfaces. Several recent studies have explored the potential for using these techniques to analyse and segment the cortical surface. However, there has been no comprehensive comparison of these approaches to one another, nor to existing Euclidean methods, to date. This paper benchmarks a collection of geometric and traditional deep learning models on phenotype prediction and segmentation of sphericalised neonatal cortical surface data, from the publicly available Developing Human Connectome Project (dHCP). Tasks include prediction of postmenstrual age at scan, gestational age at birth and segmentation of the cortical surface into anatomical regions defined by the M-CRIB-S atlas. Performance was assessed not only in terms of model precision, but also in terms of network dependence on image registration, and model interpretation via occlusion. Networks were trained both on sphericalised and anatomical cortical meshes. Findings suggest that the utility of geometric deep learning over traditional deep learning is highly task-specific, which has implications for the design of future deep learning models on the cortical surface. The code, and instructions for data access, are available from https://github.com/Abdulah-Fawaz/Benchmarking-Surface-DL.

ObjectivesTo examine the association between maternal depressive symptoms in the immediate postnatal period and offspring’s mental health in a large cohort of term- and preterm-born toddlers.Design and ParticipantsData were drawn from the Developing Human Connectome Project. Maternal postnatal depressive symptoms were assessed at term, and children’s outcomes were evaluated at a median corrected age of 18.4 months (range 17.3 – 24.3).Exposure and outcomesPreterm birth was defined as 1/2-5 Total (CBCL) and Quantitative Checklist for Autism in Toddlers (Q-CHAT) scores. Toddlers’ cognition was assessed with the Bayley Scales of Infant and Toddler Development – Third Edition (Bayley-III).ResultsHigher maternal EPDS scores were associated with toddlers’ higher CBCL (B=0.93, 95% CI 0.43-1.44, p2=0.05) and Q-CHAT scores (B=0.27, 95% CI 0.03-0.52, p=.031, f2=0.01). Higher maternal EPDS scores were not associated with toddlers’ cognitive outcomes. Maternal EPDS, toddlers’ CBCL and Q-CHAT scores did not differ between preterm (n=97; 19.1% of the total sample) and term participants.ConclusionsOur findings indicate that children whose mothers had increased depressive symptoms in the early postnatal period, including subclinical symptoms, exhibit more maternally-reported behavioural problems in toddlerhood. These associations were independent of gestational age. Further research is needed to confirm the clinical significance of these findings.Strengths and limitations of this studyProspective study with a large sample, using multiple imputation to reduce non-response bias.Maternal depressive symptoms assessed as a continuous variable, providing more nuanced information about the significance of subclinical symptoms.Maternal depressive symptoms assessed earlier than in previous studies, enabling recognition of early screening opportunities for families.Potential shared method variance bias through parent-completed child behavioural assessments.Unknown paternal and parental factors, such as comorbid psychiatric conditions, that may confound our findings.

Developmental delays in infanthood often persist, turning into life-long difficulties, and coming at great cost for the individual and community. By examining the developing brain and its relation to developmental outcomes we can start to elucidate how the emergence of brain circuits is manifested in variability of infant motor, cognitive and behavioural capacities. In this study, we examined if cortical structural covariance at birth, indexing coordinated development, is related to later infant behaviour. We included 193 healthy term-born infants from the Developing Human Connectome Project (dHCP). An individual cortical connectivity matrix derived from morphological and microstructural features was computed for each subject (morphometric similarity networks, MSNs) and was used as input for the prediction of behavioural scores at 18 months using Connectome-Based Predictive Modeling (CPM). Neonatal MSNs successfully predicted social-emotional performance. Predictive edges were distributed between and within known functional cortical divisions with a specific important role for primary and posterior cortical regions. These results reveal that multi-modal neonatal cortical profiles showing coordinated maturation are related to developmental outcomes and that network organization at birth provides an early infrastructure for future functional skills.

Rapid and portable direct tests for antibiotic resistance in human clinical samples such as urine could reduce misuse of precious antimicrobials, by allowing treatment decisions to be informed by microfluidic diagnostic tests. We demonstrate that the variable composition of human urine can significantly affect the antibiotic minimum inhibitory concentration (MIC) measured using microfluidic devices. The urine sample matrix interference was not observed in pooled normal urine, emphasising the critical importance of assessing matrix interference with a wide range of individual urine samples, rather than a few standardised or pooled controls. Both dilution into assay medium and inclusion of buffer could reduce the matrix interference, but dilution may affect analytical sensitivity by increasing the minimum bacterial cell density needed in a sample for growth to be detected, especially for miniaturised devices that test small sample volumes. We conclude it is vital to fully assess and optimise novel analytical microbiology tools using multiple individual urine samples, otherwise the high variation in matrix interference will compromise the clinical performance of these rapid diagnostics that are urgently needed to tackle the global threat of antimicrobial resistance.

Multiple studies have demonstrated less favourable childhood outcomes in infants born in early term (37-38 weeks gestation) compared to those born at full term (40-41 weeks gestation). While this could be due to higher perinatal morbidity, gestational age at birth may also have a direct effect on the brain and subsequent neurodevelopment in term-born babies. Here we characterise structural brain correlates of gestational age at birth in healthy term-born neonates and their relationship to later neurodevelopmental outcome. We used T2 and diffusion weighted Magnetic Resonance Images acquired in the neonatal period from a cohort (n=454) of healthy babies born at term age (>37 weeks gestation) and scanned between 1 and 41 days after birth. Images were analysed using tensor based morphometry (TBM) and tract based spatial statistics (TBSS). Neurodevelopment was subsequently assessed at age 18 months using the Bayley-III Scales of Infant and Toddler Development, and the effects of gestational age at birth and related neuroimaging findings on outcome were analysed with linear regression. Infants born earlier had areas of higher relative ventricular volume, and lower relative brain volume in the basal ganglia, cerebellum and brainstem. Earlier birth was also associated with lower fractional anisotropy, higher mean, axial and radial diffusivity in major white matter tracts. Gestational age at birth was positively associated with all Bayley-III subscales at age 18 months. Linear regression models predicting outcome from gestational age at birth were significantly improved by adding neuroimaging features associated with gestational age at birth. This work adds to the growing body of evidence of the impact of early term birth and highlights the importance of considering the effect of gestational age at birth in future neuroimaging studies including term-born babies.

Diffusion MRI has the potential to provide important information about the connectivity and microstructure of the human brain during normal and abnormal development, non-invasively and in vivo. Recent developments in MRI hardware and reconstruction methods now permit the acquisition of large amounts of data within relatively short scan times. This makes it possible to acquire more informative multi-shell data, with diffusion-sensitisation applied along many directions over multiple b-value shells. Such schemes are characterised by the number of shells acquired, and the specific b-value and number of directions sampled for each shell. However, there is currently no clear consensus as to how to optimise these parameters. In this work, we propose a means of optimising multi-shell acquisition schemes by estimating the information content of the diffusion MRI signal, and optimising the acquisition parameters for sensitivity to the observed effects, in a manner agnostic to any particular diffusion analysis method that might subsequently be applied to the data. This method was used to design the acquisition scheme for the neonatal diffusion MRI sequence used in the developing Human Connectome Project, which aims to acquire high quality data and make it freely available to the research community. The final protocol selected by the algorithm, and currently in use within the dHCP, consists of b = 0, 400, 1000, 2600 s/mm2 with 20, 64, 88 & 128 DW directions per shell respectively.HighlightsA data driven method is presented to design multi-shell diffusion MRI acquisition schemes (b-values and no. directions).This method optimises the multi-shell scheme for maximum sensitivity to the information content in the signal.When applied in neonates, the data suggest that a b=0 + 3 shell strategy is appropriate

The Developing Human Connectome Project is an Open Science project that provides the first large sample of neonatal functional MRI data with high temporal and spatial resolution. These data enable mapping of intrinsic functional connectivity between spatially distributed brain regions under normal and adverse perinatal circumstances, offering a framework to study the ontogeny of large-scale brain organization in humans. Here, we characterize in unprecedented detail the maturation and integrity of resting state networks (RSNs) at term-equivalent age in 337 infants (including 65 born preterm). First, we applied group independent component analysis to define 11 RSNs in term-born infants scanned at 43.5–44.5 weeks postmenstrual age (PMA). Adult-like topography was observed in RSNs encompassing primary sensorimotor, visual and auditory cortices. Among six higher-order, association RSNs, analogues of the adult networks for language and ocular control were identified, but a complete default mode network precursor was not. Next, we regressed the subject-level datasets from an independent cohort of infants scanned at 37–43.5 weeks PMA against the group-level RSNs to test for the effects of age, sex and preterm birth. Brain mapping in term-born infants revealed areas of positive association with age across four of six association RSNs, indicating active maturation in functional connectivity from 37 to 43.5 weeks PMA. Female infants showed increased connectivity in inferotemporal regions of the visual association network. Preterm birth was associated with striking impairments of functional connectivity across all RSNs in a dose-dependent manner; conversely, connectivity of the superior parietal lobules within the lateral motor network was abnormally increased in preterm infants, suggesting a possible mechanism for specific difficulties such as developmental coordination disorder, which occur frequently in preterm children. Overall, we found a robust, modular, symmetrical functional brain organization at normal term age. A complete set of adult-equivalent primary RSNs is already instated, alongside emerging connectivity in immature association RSNs, consistent with a primary-to-higher order ontogenetic sequence of brain development. The early developmental disruption imposed by preterm birth is associated with extensive alterations in functional connectivity., Stroke survivors require therapies that are tailored to their unique brain injury, but it is currently difficult to assess the degree to which damaged brain networks are engaged in mental activities. Lorenz et al. reveal how a novel brain-scanning tool can overcome this challenge in patients, paving the way to individualised therapy.

Background: Miniaturised bioassays permit diagnostic testing near the patient, and the results can be recorded digitally using inexpensive cameras including smartphone and mobile phone cameras. Although digital cameras are now inexpensive and portable, the minimum performance required for microfluidic diagnostic bioassays has not been defined. We present a systematic comparison of a wide range of different digital cameras for capturing and measuring results of microfluidic bioassays and describe a framework to specify performance requirements to quantify immunoassays. Methods: A set of 200 µm diameter microchannels was filled with a range of concentrations of dyes used in colorimetric and fluorometric enzyme immunoassays. These were imaged in parallel using cameras of varying cost and performance ranging from £500. Results: Higher resolution imaging allowed larger numbers of microdevices to be resolved and analysed in a single image. In contrast, low quality cameras were still able to quantify results but for fewer samples. In some cases, an additional macro lens was added to focus closely. If image resolution was sufficient to identify individual microfluidic channels as separate lines, all cameras were able to quantify a similar range of concentrations of both colorimetric and fluorometric dyes. However, the mid-range cameras performed better, with the lowest cost cameras only allowing one or two samples to be quantified per image. Consistent with these findings, we demonstrate that quantitation (to determine endpoint titre) of antibodies against dengue and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) viruses is possible using a wide range of digital imaging devices including the mid-range smartphone iPhone 6S and a budget Android smartphone costing Conclusions: In conclusion, while more expensive and higher quality cameras allow larger numbers of devices to be simultaneously imaged, even the lowest resolution and cheapest cameras were sufficient to record and quantify immunoassay results.

There is an increasing need in decentralized high-performance analytical testing in many fields, including human healthcare, environmental, food and water monitoring, to name a few. High-performance testing has mostly remained so far limited to bulky and expensive analytical equipment in centralized labs. Smartphone are able to break that paradigm by providing a quantitation equipment (e.g. a smartphone camera) attached to portable digital platform. This chapter provides an overview on the opportunities and challenges for smartphone-based testing in modern point-of-need applications.

The performance of biosensors is often optimized in buffers, which brings inconsistencies during applications with biological samples. Current strategies for minimizing sample (matrix) interference are complex to automate and miniaturize, involving, e.g., sample dilution or recovery of serum/plasma. This study shows the first systematic analysis using hundreds of actual microfluidic immunoassay fluoropolymer strips to understand matrix interference in microflow systems. As many interfering factors are assay-specific, we have explored matrix interference for a range of enzymatic immunoassays, including a direct mIgG/anti-mIgG, a sandwich cancer biomarker PSA, and a sandwich inflammatory cytokine IL-1β. Serum matrix interference was significantly affected by capillary antibody surface coverage, suggesting for the first time that the main cause of the serum matrix effect is low-affinity serum components (e.g., autoantibodies) competing with high-affinity antigens for the immobilized antibody. Additional experiments carried out with different capillary diameters confirmed the importance of antibody surface coverage in managing matrix interference. Building on these findings, we propose a novel analytical approach where antibody surface coverage and sample incubation times are key for eliminating and/or minimizing serum matrix interference, consisting in bioassay optimization carried out in serum instead of buffer, without compromising the performance of the bioassay or adding extra cost or steps. This will help establishing a new route toward faster development of modern point-of-care tests and effective biosensor development.

Background: This study investigated the experiences and perceptions of Saudi patients with primary insomnia.Method: A cross-sectional design study (July 2019) using a self-administered questionnaire survey.Results: Sixty-eight patients with primary insomnia participated (53% male, 47% female; 60% were younger than 45 years and a majority had a university-degree education level). Of the respondents, 64.7% were dissatisfied with their knowledge of insomnia and 97% wanted more information about its treatment. Approximately 59% used prescribed sleep medicines from the first instance and only 6% were referred to a psychologist for counselling. Benzodiazepines and Z-drugs were prescribed for 16% and 19% of respondents, respectively. A majority of respondents were given antihistamines or over-the-counter herbs. While 60% of patients used medications for long periods, 87% were not involved in treatment decisions. For those on long-term use, 80% were not seen regularly by doctors and 93% were provided with no plan for medicine discontinuations. More than 60% of respondents said that sleeping pills were best for treating insomnia and 31% believed they are safe and would continue taking them. More than 45% would not prefer non-pharmacological treatments (e.g., counselling).Conclusions: Responses to the survey indicate a lack of knowledge about primary insomnia and its treatment among Saudi patients. Prescribing hypnotics is the most common treatment, non-pharmacological treatments are seldom used, and responses indicated inappropriate and long-term use of hypnotics. Patients should be better educated about insomnia and its treatment. Physicians should consider using non-pharmacological treatments and the Saudi Ministry of Health should develop country-specific guidelines.

The developing Human Connectome Project (dHCP) aims to create a detailed 4-dimensional connectome of early life spanning 20–45 weeks post-menstrual age. This is being achieved through the acquisition of multi-modal MRI data from over 1000 in- and ex-utero subjects combined with the development of optimised pre-processing pipelines. In this paper we present an automated and robust pipeline to minimally pre-process highly confounded neonatal resting-state fMRI data, robustly, with low failure rates and high quality-assurance. The pipeline has been designed to specifically address the challenges that neonatal data presents including low and variable contrast and high levels of head motion. We provide a detailed description and evaluation of the pipeline which includes integrated slice-to-volume motion correction and dynamic susceptibility distortion correction, a robust multimodal registration approach, bespoke ICA-based denoising, and an automated QC framework. We assess these components on a large cohort of dHCP subjects and demonstrate that processing refinements integrated into the pipeline provide substantial reduction in movement related distortions, resulting in significant improvements in SNR, and detection of high quality RSNs from neonates., NeuroImage, 223, ISSN:1053-8119, ISSN:1095-9572

A key challenge in medical imaging is determining a precise correspondence between image properties and tissue microstructure. This comparison is hindered by disparate scales and resolutions between medical imaging and histology. We present a new technique, 3D Bridging of Optically-clear histology with Neuroimaging Data (3D-BOND), for registering medical images with 3D histology to overcome these limitations. Ex vivo 120 × 120 × 200 μm resolution diffusion-MRI (dMRI) data was acquired at 7 T from adult C57Bl/6 mouse hippocampus. Tissue was then optically cleared using CLARITY and stained with cellular markers and confocal microscopy used to produce high-resolution images of the 3D-tissue microstructure. For each sample, a dense array of hippocampal landmarks was used to drive registration between upsampled dMRI data and the corresponding confocal images. The cell population in each MRI voxel was determined within hippocampal subregions and compared to MRI-derived metrics. 3D-BOND provided robust voxel-wise, cellular correlates of dMRI data. CA1 pyramidal and dentate gyrus granular layers had significantly different mean diffusivity (p > 0.001), which was related to microstructural features. Overall, mean and radial diffusivity correlated with cell and axon density and fractional anisotropy with astrocyte density, while apparent fibre density correlated negatively with axon density. Astrocytes, axons and blood vessels correlated to tensor orientation.

Incubation at controlled temperature is a key step in culture based microbiological tests. Access to culture-based microbiological testing requires access to conventional incubators in a laboratory. Portable incubators allow microbiological testing in the field and in resource-limited settings, and can eliminate the challenge of sample transportation, minimising the chance of sample degradation. Recent studies have reported low-cost portable incubator designs suitable for field or off-grid use, but these either need an external power supply (e.g. mains AC or 12 V DC), or rely on passive heating without thermostatic control. Here we report that small inexpensive uninterruptable power supply (UPS) products manufactured for consumer electronics and powered by lithium-ion battery packs allowing thermostatic temperature control in small portable incubators that can maintain precise temperatures with or without external power. We present an open-source design for a Microbiological Mobile Incubator (MicroMI) in two sizes for field use. The MicroMI is built from simple and widely available components and is easy to set up. The open source design can be customised for different numbers of samples. The smallest and most efficient design uses a vacuum insulated food flask that allows longer operation with smaller, lower capacity UPS. The larger flight case design has space for more samples, but depletes the battery faster. The UPS maintains a typical microbiology incubation temperature for up to 24 h without external power- ideal for typical incubation needed for culture methods. The battery capacity, incubator design, and external ambient temperature all affected duration of operation without requiring external power. We validated the MicroMI by conducting classical microbiological tests using agar petri dishes, slant cultures and dip slides, and biochemical tests. We conclude the MicroMI design allows inexpensive lithium battery products to be used to simplify field microbiology and increase access to vital analytical microbiology testing.

Viable bacterial cell counting is fundamental to analytical microbiology and agar plate colony counting remains common yet laborious and slow. Here, we demonstrate two methods for counting bacteria using commercially available microfluidic devices. We show that accurate viable cell counting is possible using simple and easy ‘dip and test’ arrays of microcapillaries. Colorimetric and fluorescent growth detection both permit viable cell counting in microcapillaries either by limiting dilution into multiple microfluidic compartments using a single endpoint measurement, or alternatively by quantifying growth kinetics. The microcapillary devices are compatible with conventional 96 well plates and multichannel pipettes, expanding each microplate row into 120 individual 1 or 2 microlitre samples. At limiting dilution, counting the proportion of positive compartments permitted accurate calculation of gram-negative and gram-positive bacteria (E. coli and S. saprophyticus) at concentrations down to as low as 10 CFU/mL with almost 1:1 agreement with agar plate colony counts over four orders of magnitude. A smartphone camera was sufficient to record endpoint images of resazurin growth detection both colorimetrically and fluorescently. Viable cell counting of E. coli and S. saprophyticus was also possible through recording growth kinetics and determining the time taken to detect resazurin conversion. However, only the limiting dilution method remained consistent in the presence of urine matrix, as some interference in growth rate was observed when bacteria were spiked into higher concentrations of normal urine to simulate urinary tract infection patient samples. However, with the limiting dilution counting method endpoint growth was always detected even in the presence of 90% urine matrix, suggesting that this method might permit bacterial pathogen counting directly in clinical samples without agar plating.

BACKGROUND AND PURPOSE: Assessing the response to treatment in infiltrative brain tumors by using lesion volume–based response criteria is challenging. We hypothesized that in such tumors, volume measurements alone may not accurately capture changes in actual tumor burden during treatment. We longitudinally evaluated volume changes in both normal-appearing supratentorial white matter and the brain stem lesions in patients treated for diffuse intrinsic pontine glioma to determine to what extent adjuvant systemic therapies may skew the accuracy of tumor response assessments based on volumetric analysis. MATERIALS AND METHODS: The anatomic MR imaging and diffusion tensor imaging data of 26 patients with diffuse intrinsic pontine glioma were retrospectively analyzed. Treatment included conformal radiation therapy in conjunction with vandetanib and dexamethasone. Volumetric and diffusion data were analyzed with time, and differences between time points were evaluated statistically. RESULTS: Normalized brain stem lesion volume decreased during combined treatment (slope = −0.222, P CONCLUSIONS: Longitudinal changes in brain stem lesion volume are robust; less pronounced but measurable changes occur in the supratentorial white matter. Volume changes in nonirradiated supratentorial white matter during the disease course reflect the effects of systemic medication on the water homeostasis of normal parenchyma. Our data suggest that adjuvant nontumor–targeted therapies may have a more substantial effect on lesion volume changes than previously thought; hence, an apparent volume decrease in infiltrative tumors receiving combined therapies may lead to overestimation of the actual response and tumor control.

Growth in open-source hardware designs combined with the low-cost of high performance optoelectronic and robotics components has supported a resurgence of in-house custom lab equipment development. We describe a low cost (below USD700), open-source, fully customizable high-throughput imaging system for analytical microbiology applications. The system comprises a Raspberry Pi camera mounted on an aluminium extrusion frame with 3D-printed joints controlled by an Arduino microcontroller running open-source Repetier Host Firmware. The camera position is controlled by simple G-code scripts supplied from a Raspberry Pi singleboard computer and allow customized time-lapse imaging of microdevices over a large imaging area. Open-source OctoPrint software allows remote access and control. This simple yet effective design allows high-throughput microbiology testing in multiple formats including formats for bacterial motility, colony growth, microtitre plates and microfluidic devices termed ‘lab-on-a-comb’ to screen the effects of different culture media components and antibiotics on bacterial growth. The open-source robot design allows customization of the size of the imaging area; the current design has an imaging area of ~420 × 300mm, which allows 29 ‘lab-on-a-comb’ devices to be imaged which is equivalent 3480 individual 1μl samples. The system can also be modified for fluorescence detection using LED and emission filters embedded on the PiCam for more sensitive detection of bacterial growth using fluorescent dyes.

Purpose This study aims to obtain consensus statements required for the development of clinical guidelines for the use of benzodiazepines (BZDs) and Z-drugs for the management of primary insomnia in adults in Saudi Arabia. Methods Three rounds of the e-Delphi technique using a Bristol Online Survey (BOS) were conducted between May and August 2018. The Director of the Saudi Sleep Medicine Group helped recruit the country’s sleep medicine experts. Snowballing was used to forward invitation emails, information sheets, and the survey to known sleep medicine experts and physicians deemed to be interested in the field. All participants’ details were anonymised except to the researcher. Results Fifteen experts from four different regions and specialities in Saudi Arabia participated in Round 1. Twenty-one statements originated from participants’ responses. In Round 2, there were 17 respondents and 16 of the statements obtained the required consensus of 70% or higher. Eleven experts participated in Round 3 and eight statements received 100% agreement, two received 91%, and six received 82%. Having obtained the required consensus of 80% or higher in Round 3, these 16 statements fulfilled the criteria to be included in future guidelines. The five statements that failed to attain the required consensus were rejected as inappropriate for inclusion in Saudi Arabian clinical guidelines. Conclusions The items that achieved the required consensus can be included in future guidelines for the use of BZDs and Z-drugs in the treatment of primary insomnia in adults to standardize best practices in sleep medicine in Saudi Arabia.

Based on clinical observations, we hypothesized that in infiltrative high-grade brainstem neoplasms, such as diffuse intrinsic pontine glioma (DIPG), longitudinal metabolic evaluation of the tumor by magnetic resonance spectroscopy (MRS) may be more accurate than volumetric data for monitoring the tumor’s biological evolution during standard treatment. We evaluated longitudinal MRS data and corresponding tumor volumes of 31 children with DIPG. We statistically analyzed correlations between tumor volume and ratios of Cho/NAA, Cho/Cr, and NAA/Cr at key time points during the course of the disease through the end of the progression-free survival period. By the end of RT, tumor volume had significantly decreased from the baseline (P

BACKGROUND AND PURPOSE: The conventional MR imaging appearance of diffuse intrinsic pontine glioma suggests intralesional histopathologic heterogeneity, and various distinct lesion components, including T2-hypointense foci, have been described. Here we report the prevalence, conventional MR imaging semiology, and advanced MR imaging features of non-necrotic T2-hyperintense foci in diffuse intrinsic pontine glioma. MATERIALS AND METHODS: Twenty-five patients with diffuse intrinsic pontine gliomas were included in this study. MR imaging was performed at 3T by using conventional and advanced MR imaging sequences. Perfusion (CBV), vascular permeability (ve, Ktrans), and diffusion (ADC) metrics were calculated and used to characterize non-necrotic T2-hyperintense foci in comparison with other lesion components, namely necrotic T2-hyperintense foci, T2-hypointense foci, peritumoral edema, and normal brain stem. Statistical analysis was performed by using Kruskal-Wallis and Wilcoxon rank sum tests. RESULTS: Sixteen non-necrotic T2-hyperintense foci were found in 12 tumors. In these foci, ADC values were significantly higher than those in either T2-hypointense foci (P = .002) or normal parenchyma (P = .0002), and relative CBV values were significantly lower than those in either T2-hypointense (P = .0002) or necrotic T2-hyperintense (P = .006) foci. Volume transfer coefficient values in T2-hyperintense foci were lower than those in T2-hypointense (P = .0005) or necrotic T2-hyperintense (P = .0348) foci. CONCLUSIONS: Non-necrotic T2-hyperintense foci are common, distinct lesion components within diffuse intrinsic pontine gliomas. Advanced MR imaging data suggest low cellularity and an early stage of angioneogenesis with leaky vessels resulting in expansion of the extracellular space. Because of the lack of biopsy validation, the underlying histoarchitectural and pathophysiologic changes remain unclear; therefore, these foci may correspond to a poorly understood biologic event in tumor evolution.

Innovative and coordinated strategies to address weight bias among health professionals are urgently needed. We conducted a systematic literature review of empirical peer-reviewed published studies to assess the impact of interventions designed to reduce weight bias in students or professionals in a health-related field. Combination sets of keywords based on three themes (1: weight bias/stigma; 2: obesity/overweight; 3: health professional) were searched within nine databases. Our search yielded 1447 individual records, of which 17 intervention studies satisfied the inclusion criteria. Most studies (n = 15) included medical, dietetic, health promotion, psychology and kinesiology students, while the minority included practicing health professionals (n = 2). Studies utilized various bias-reduction strategies. Many studies had methodological weaknesses, including short assessment periods, lack of randomization, lack of control group and small sample sizes. Although many studies reported changes in health professionals' beliefs and knowledge about obesity aetiology, evidence of effectiveness is poor, and long-term effects of intervention strategies on weight bias reduction remain unknown. The findings highlight the lack of experimental research to reduce weight bias among health professionals. Although changes in practice will likely require multiple strategies in various sectors, well-designed trials are needed to test the impact of interventions to decrease weight bias in healthcare settings.

Conventional methods for the detection and quantitation of bacteria are slow, laborious and require a laboratory. Microfluidic systems offer faster and portable testing, and smartphone cameras can record colorimetric or fluorometric bioassays, but this requires dye addition. Here, we demonstrate for the first time label-free smartphone detection of bacterial light scattering by darkfield microfluidic imaging to measure bacteria and bacteriophage lysis. A single LED and portable 3D printed imaging box allowed bacterial concentration and growth to be measured by direct imaging of bacterial light scattering. Bacteriophage lysis was detected within a 10-channel microfluidic device made from melt-extruded fluoropolymer micro capillary film, allowing rapid detection of host specificity. Elimination of unwanted reflections and optimising illumination angle are critical for successful darkfield bacterial imaging, with 15° giving maximal intensity. Bacterial sedimentation was directly observed within microfluidic devices, and detection sensitivity significantly increased by allowing bacteria to sediment for 30 min. With this simple, low-cost, 3D printed system bacterial concentrations down to an optical density of 0.1 could be measured corresponding to 8 × 104 colony forming units (CFU) per microdevice, approaching the sensitivity of conventional spectrophotometers. Bacteriophage lysis could be detected at a range of starting cell concentrations. With a low starting cell concentration, the increase in light scatter signal with incubation was prevented in the presence of bacteriophage. Conversely, with high starting cell concentration, the light scatter signal detected at the start was clearly eliminated when phage were added, indicating this simple system allows direct visualisation of bacteriophage eliminating light scattering by lysis.

Both home sample collection and home testing using rapid point-of-care diagnostic devices can offer benefits over attending a clinic/hospital to be tested by a healthcare professional. Usability is critical to ensure that in-home sampling or testing by untrained users does not compromise analytical performance. Usability studies can be laborious and rely on participants attending a research location or a researcher visiting homes; neither has been appropriate during COVID-19 outbreak control restrictions. We therefore developed a remote research usability methodology using videolink observation of home users. This avoids infection risks from home visits and ensures the participant follows the test protocol in their home environment. In this feasibility study, volunteers were provided with models of home blood testing and home blood sampling kits including a model lancet, sampling devices for dried blood spot collection, and model lateral flow device. After refining the study protocol through an initial pilot (n = 7), we compared instructions provided either as written instructions (n = 5), vs addition of video instructions (n = 5), vs written and video instructions plus videolink supervision by the researcher (n = 5). All users were observed via video call to define which test elements could be assessed remotely. All 22 participants in the study accessed the video call and configured their videolink allowing the researcher to clearly observe all testing tasks. The video call allowed the researcher to assess distinct errors during use including quantitative (volume of blood) and qualitative (inaccurate interpretation of results) errors many of which could compromise test accuracy. All participants completed the tasks and returned images of their completed tests (22/22) and most returned completed questionnaires (20/22). We suggest this remote observation via videolink methodology is a simple, rapid and powerful methodology to assess and optimise usability of point-of-care testing methods in the home setting.

Docosahexaenoic acid (DHA) and arachidonic acid (ArA) are essential brain specific fatty acids (BSFA) for mammalian central nervous system development. Human brains have accelerated growth with significant increase in cerebral content of ArA and DHA during the last trimester of pregnancy and first postnatal months. This randomized double blind placebo controlled single centre trial assessed the impact of BSFA supplementation in pregnancy on newborn infants' brain volumes. Eighty six infants born to study mothers had brain magnetic resonance imaging (MRI) scans soon after birth. Total and regional brain volumes were analyzed and related to maternal supplementation group. Males born to the BSFA supplemented mothers had significantly larger total brain volumes, total gray matter, corpus callosum and cortical volumes when compared to the placebo group. This is the first study to show maternal BSFA supplementation enhances newborn infants' brain size and suggests differential sex sensitivity of fetal brains to pregnancy BSFA status.

Microglia-mediated neuroinflammation is key in numerous brain diseases including encephalopathy of the preterm born infant. Microglia of the still-developing brain have unique properties but little is known of how they regulate their inflammatory activation. This is important information as every year 9 million preterm born infants acquire persisting neurological injuries associated with encephalopathy and we lack strategies to prevent and treat these injuries. Our study of activation state regulators in immature brain microglia found a robust down-regulation of Wnt/β-catenin pathway receptors, ligands and intracellular signalling members in pro-inflammatory microglia. We undertook our studies initially in a mouse model of microglia-mediated encephalopathy including the clinical hallmarks of oligodendrocyte injury and hypomyelination. We purified microglia from this model and applied a genome-wide transcriptomics analysis validated with quantitative profiling. We then verified that down-regulation of the Wnt/β-catenin signalling cascade is sufficient and necessary to drive microglia into an oligodendrocyte-damaging phenotype using multiple pharmacological and genetic approaches in vitro and in vivo in mice and in humans and zebrafish. We also demonstrated that genomic variance in the WNT/β-catenin pathway is associated with the anatomical connectivity phenotype of the human preterm born infant. This integrated analysis of genomics and connectivity, as a surrogate for oligodendrocyte function/myelination, is agnostic to cell type. However, this data indicates that the WNT pathway is relevant to human brain injury and specifically that WNT variants may be useful clinically for injury stratification and prognosis. Finally, we performed a translational experiment using a BBB penetrant microglia-specific targeting 3DNA nanocarrier to deliver a Wnt agonist specifically and directly to microglia in vivo. Increasing the activity of the Wnt/β-catenin pathway specifically in microglia in our model of microglia-mediated encephalopathy was able to reduce microglial pro-inflammatory activation, prevent the typical hypomyelination and also prevent the long-term memory deficit associated with this hypomyelination. In summary, the canonical Wnt/β-catenin pathway regulates microglial activation and up-regulation of this pathway could be a viable neurotherapeutic strategy.

Conjugation of small molecule agonists of Toll-like receptor 7 (TLR7) to proteins, lipids, or polymers is known to modulate potency, and the physical form or formulation of these conjugates is likely to have a major effect on their immunostimulatory activity. Here, we studied the effect of formulation on potency of a 1,2‑di‑(9Z‑octadecenoyl)‑sn‑glycero‑3‑phosphoethanolamine (DOPE) conjugated TLR7 agonist (DOPE-TLR7a) alongside assessing physical form using Dynamic Light Scattering (DLS), Nanosight Particle Tracking (NTA) analysis and Small Angle X-ray Scattering (SAXS). A very high potency of DOPE-TLR7a conjugate (EC50 around 9 nM) was observed either when prepared by direct dilution from DMSO or when formulated into 400–700 nm large multilamella liposomes containing dimethyldioctadecylammonium bromide salt (DDA) and DOPE. When prepared by dissolution in DMSO followed by dilution in aqueous culture medium, 93 ± 5 nm nanoparticles were formed. Without dilution from solution in DMSO, no nanoparticles were observed and no immunostimulatory activity could be detected without this formulation step. SAXS analysis of the conjugate after DMSO dissolution/water dilution revealed a lamellar order with a layer spacing of 68.7 Å, which correlates with arrangement in groups of 3 bilayers. The addition of another immunostimulatory glycolipid, trehalose‑6,6‑dibehenate (TDB), to DOPE:DDA liposomes gave no further increase in immunostimulatory activity beyond that provided by incorporating DOPE-TLR7a. Given the importance of nanoparticle or liposomal formulation for activity, we conclude that the major mechanism for increased potency when TLR7 agonists are conjugated to macromolecules is through alteration of physical form., Graphical abstract Unlabelled Image

Language acquisition appears to rely at least in part on recruiting pre-existing brain structures. We hypothesized that the neural substrate for language can be characterized by distinct, non-trivial network properties of the brain, that modulate language acquisition early in development. We tested whether these brain network properties present at the normal age of birth predicted later language abilities, and whether these were robust against perturbation by studying infants exposed to the extreme environmental stress of preterm birth.We found that brain network controllability and integration predicted respectively phonological, ‘bottom-up’ and syntactical, ‘top-down’ language skills at 20 months, and that syntactical but not phonological functions were modulated by premature extrauterine life. These data show that the neural substrate for language acquisition is a network property present at term corrected age. These distinct developmental trajectories may be relevant to the emergence of social interaction after birth.

Toll like receptors (TLR) are critical mediators of the immune system with their activation linked to infection, inflammation and the pathogenesis of immune diseases including autoimmunity and cancer. For this reason, over the last two decades, TLR and their associated signalling pathways have been targeted therapeutically to enhance innate and adaptive immunity. Several TLR ligands, both endogenous and synthetic are at various phases of clinical testing, and new ligands are continually emerging. Agonists of TLR7 are known immune response modifiers, simultaneously stimulating several cell types, resulting in immune cell activation and cytokine and chemokine release. The immune stimulating properties of the TLR7 agonist Imiquimod has also been exploited for use in the treatment of malignant superficial tumours of the skin. Here, we investigated a novel TLR7 agonist UC-1V150 and demonstrate it activates both human and mouse myeloid cells in vitro and in vivo, to deliver potent FcγR mediated engulfment of opsonised target cells.

Premature birth increases the risk of developing neurocognitive and neurobe-havioural disorders. The mechanisms of altered brain development causing these disorders are yet unknown. Studying the morphology and function of the brain during maturation provides us not only with a better understanding of normal development, but may help us to identify causes of abnormal development and their consequences. A particular difficulty is to distinguish abnormal patterns of neurodevelopment from normal variation. The Developing Human Connectome Project (dHCP) seeks to create a detailed four-dimensional (4D) connectome of early life. This connectome may provide insights into normal as well as abnormal patterns of brain development. As part of this project, more than a thousand healthy fetal and neonatal brains will be scanned in vivo. This requires computational methods which scale well to larger data sets. We propose a novel groupwise method for the construction of a spatio-temporal model of mean morphology from cross-sectional brain scans at different gestational ages. This model scales linearly with the number of images and thus improves upon methods used to build existing public neonatal atlases, which derive correspondence between all pairs of images. By jointly estimating mean shape and longitudinal change, the atlas created with our method overcomes temporal inconsistencies, which are encountered when mean shape and intensity images are constructed separately for each time point. Using this approach, we have constructed a spatio-temporal atlas from 275 healthy neonates between 35 and 44 weeks post-menstrual age (PMA). The resulting atlas qualitatively preserves cortical details significantly better than publicly available atlases. This is moreover confirmed by a number of quantitative measures of the quality of the spatial normalisation and sharpness of the resulting template brain images.

Ambient temperature filter dried vaccine formulations have been proposed to simultaneously achieve thermostability and offer a ready-to-use immunisation device that combines reconstitution and injection. Vaccine concentration should be uniform at the point of injection, but the uniformity following direct reconstitution of filter-dried vaccines has not been reported. We present here a study of vaccine mixing and release following dissolution of filter-dried model protein and toxoid antigens within a single syringe, filter and needle unit. Release was better for filters made from glass than cellulose. Without additional mixing, uniformity was poor and only 41% of input protein was released from protein filter-dried onto glass fibre. In contrast, adding a simple glass bead and mixing by inversion, 100% release antigen solution was achieved, with uniform concentration at exit from the needle throughout a simulated injection. Adsorption onto alum adjuvant had no detectable effect on vaccine dissolution and mixing. The uniformity and yield of low doses of diphtheria and tetanus toxoid was also improved by mixing, albeit with a lower yield of 60-68%. We conclude that uniformity and mixing should be studied to ensure safety and efficacy of directly reconstituted filter-dried vaccine formulations.

We present a novel but simple enteric coated sphere formulation containing probiotic bacteria (Lactobacillus casei). Oral delivery of live bacterial cells (LBC) requires live cells to survive firstly manufacturing processes and secondly GI microbicidal defenses including gastric acid. We incorporated live L. casei directly in the granulation liquid, followed by granulation, extrusion, spheronization, drying and spray coating to produce dried live probiotic spheres. A blend of MCC, calcium-crosslinked alginate, and lactose was developed that gave improved live cell survival during manufacturing, and gave excellent protection from gastric acid plus rapid release in intestinal conditions. No significant loss of viability was observed in all steps except drying, which resulted in approximately 1 log loss of viable cells. Eudragit coating was used to protect dried live cells from acid, and microcrystalline cellulose (MCC) was combined with sodium alginate to achieve efficient sphere disintegration leading to rapid and complete bacterial cell release in intestinal conditions. Viability and release of L. casei was evaluated in vitro in simulated GI conditions. Uncoated spheres gave partial acid protection, but enteric coated spheres effectively protected dried probiotic LBC from acid for 2 h, and subsequently released all viable cells within 1h of transfer into simulated intestinal fluid.

In the mature mammalian brain, the primary somatosensory and motor cortices are known to be spatially organized such that neural activity relating to specific body parts can be somatopically mapped onto an anatomical “homunculus”. This organization creates an internal body representation which is fundamental for precise motor control, spatial awareness and social interaction. Although it is unknown when this organization develops in humans, animal studies suggest that it may emerge even before the time of normal birth. We therefore characterized the somatotopic organization of the primary sensorimotor cortices using functional MRI and a set of custom-made robotic tools in 35 healthy preterm infants aged from 31 + 6 to 36 + 3 weeks postmenstrual age. Functional responses induced by somatosensory stimulation of the wrists, ankles, and mouth had a distinct spatial organization as seen in the characteristic mature homunculus map. In comparison to the ankle, activation related to wrist stimulation was significantly larger and more commonly involved additional areas including the supplementary motor area and ipsilateral sensorimotor cortex. These results are in keeping with early intrinsic determination of a somatotopic map within the primary sensorimotor cortices. This may explain why acquired brain injury in this region during the preterm period cannot be compensated for by cortical reorganization and therefore can lead to long-lasting motor and sensory impairment.

Antibiotic resistance in urinary tract infection is a major global challenge, and improved cost-effective and rapid antibiotic susceptibility tests (AST) are urgently needed to inform correct antibiotic selection. Although microfluidic technology can miniaturise AST, the high dynamic range of pathogen density found in clinical urine samples makes direct testing of clinical samples – rather than testing colonies from overnight agar plates – extremely challenging. We evaluated for the first time how pathogen concentration in urine affects microfluidic AST using a novel microplate-compatible high-throughput microfluidic AST system termed “Lab-on-a-Comb”. When tested with clinical E. coli isolates at standardised density, these devices gave identical antibiotic susceptibility profiles to standard disc diffusion and microtitre plate tests. Bacterial detection directly in synthetic urine spiked with clinical E. coli UTI isolates was possible over a very large dynamic range of starting cell densities, from 103 – 108 CFU/mL which covers the range of pathogen cell densities found in patient urine. The lowest cell density where cell growth was reproducibly detected optically was 9.6x102 CFU/mL, corresponding to one single CFU detected in a 1 μL microcapillary-an unprecedented level of sensitivity. Cell growth kinetics followed a simple Monod model with fast growth limited by the substrate availability and an estimated doubling time of 24.5 min, indicating optimal E. coli growth conditions within these microfluidic devices. There was a trade-off between sensitivity and speed of detection, with 105 CFU/mL detection possible within 2h, but 6h incubation required at 103 CFU/mL.

This study reports for the first time the sensitive colorimetric and fluorescence detection of clinically relevant protein biomarkers by sandwich immunoassays using the covalent immobilisation of antibodies onto the fluoropolymer surface inside Teflon®-FEP microfluidic devices. Teflon®-FEP has outstanding optical transparency ideal for high-sensitivity colorimetric and fluorescence bioassays, however this thermoplastic is regarded as chemically inert and very hydrophobic. Covalent immobilisation can offer benefits over passive adsorption to plastic surfaces by allowing better control over antibody density, orientation and analyte binding capacity, and so we tested a range of different and novel covalent immobilisation strategies. We first functionalised the inner surface of a 10-bore, 200 μm internal diameter FEP microcapillary film with high-molecular weight polyvinyl alcohol (PVOH) without changing the outstanding optical transparency of the device delivered by the matched refractive index of FEP and water. Glutaraldehyde immobilisation was compared with the use of photoactivated linkers and NHS-ester crosslinkers for covalently immobilising capture antibodies onto PVOH. Three clinically relevant sandwich ELISAs were tested against the cytokine IL-1β, the myocardial infarct marker cardiac troponin I (cTnI), and the chronic heart failure marker brain natriuretic peptide (BNP). Overall, glutaraldehyde immobilisation was effective for BNP assays, but yielded unacceptable background for IL-1β and cTnI assays caused by direct binding of the biotinylated detection antibody to the modified PVOH surface. We found NHS-ester groups reacted with APTES-treated PVOH coated fluoropolymers. This facilitated a novel method for capture antibody immobilisation onto fluoropolymer devices using a bifunctional NHS-maleimide crosslinker. The density of covalently immobilised capture antibodies achieved using PVOH/APTES/NHS/maleimide approached levels seen with passive adsorption, and sensitive and quantitative assay performance was achieved using this method. Overall, the PVOH coating provided an excellent surface for controlled covalent antibody immobilisation onto Teflon®-FEP for performing high-sensitivity immunoassays.

Live bacterial cells (LBC) are administered orally as attenuated vaccines, to deliver biopharmaceutical agents, and as probiotics to improve gastrointestinal health. However, LBC present unique formulation challenges and must survive gastrointestinal antimicrobial defenses including gastric acid after administration. We present a simple new formulation concept, termed Polymer Film Laminate (PFL). LBC are ambient dried onto cast acid-resistant enteric polymer films that are then laminated together to produce a solid oral dosage form. LBC of a model live bacterial vaccine and a probiotic were dried directly onto a cast film of enteric polymer. The effectiveness at protecting dried cells in a simulated gastric fluid (pH 2.0) depended on the composition of enteric polymer film used, with a blend of ethylcellulose plus Eudragit L100 55 providing greater protection from acid than Eudragit alone. However, although PFL made from blended polymers films completely released low molecular weight dye into intestinal conditions (pH 7.0), they failed to release LBC. In contrast, PFL made from Eudragit alone successfully protected dried probiotic or vaccine LBC from simulated gastric fluid for 2h, and subsequently released all viable cells within 60min of transfer into simulated intestinal fluid. Release kinetics could be controlled by modifying the lamination method.

AIM: Studies have explored how mothers and premature babies make the transition from a neonatal unit (NNU) to home, but little is known about how mothers cope with urban life with a vulnerable baby. This controlled trial investigated how first-time mothers with singleton preterm babies handled that experience in the first few months after discharge from a NNU, compared to the first-time mothers of healthy, singleton term-born infants.METHODS: This parent-led, qualitative study was carried out in London, UK, from 2013-2015 and used diaries and walking interviews with researchers. Thematic network analysis was performed to provide insights into the experiences of these 19 mothers.RESULTS: The two groups had similar experiences and needs in the urban environment and these common experiences of city life shaped their new identities as mothers. However, the mothers of preterm babies had difficulties developing supportive relationships and seeking affirming social environments.CONCLUSION: This study highlights what the mothers of preterm babies experienced in the first few months after their infant was discharged from hospital. It stresses the need to understand the complexity of the mothers' social, emotional and psychological processes when they make the transition from home to visit the city with a vulnerable baby. This article is protected by copyright. All rights reserved.