Combining Thermal Desorption with Selected Ion Flow Tube Mass Spectrometry for Analyses of Breath Volatile Organic Compounds

Supporting Information is available online at: https://pubs.acs.org/doi/10.1021/acs.analchem.3c04286 .An instrument integrating thermal desorption (TD) to selected ion flow tube mass spectrometry (SIFT-MS) is presented, and its application to analyze volatile organic compounds (VOCs) in human breath is demonstrated for the first time. The rationale behind this development is the need to analyze breath samples in large-scale multicenter clinical projects involving thousands of patients recruited in different hospitals. Following adapted guidelines for validating analytical techniques, we developed and validated a targeted analytical method for 21 compounds of diverse chemical class, chosen for their clinical and biological relevance. Validation has been carried out by two independent laboratories, using calibration standards and real breath samples from healthy volunteers. The merging of SIFT-MS and TD integrates the rapid analytical capabilities of SIFT-MS with the capacity to collect breath samples across multiple hospitals. Thanks to these features, the novel instrument has the potential to be easily employed in clinical practice.This project was supported by Royal Society of Chemistry through the Research Fund. This research was also supported by Wellcome Trust, Surgery and Cancer Department, Imperial College London and NIHR London In Vitro Diagnostics Co-operative

Pathotyping the Zoonotic Pathogen Streptococcus suis: Novel Genetic Markers To Differentiate Invasive Disease-Associated Isolates from Non-Disease-Associated Isolates from England and Wales.

Streptococcus suis is one of the most important zoonotic bacterial pathogens of pigs, causing significant economic losses to the global swine industry. S. suis is also a very successful colonizer of mucosal surfaces, and commensal strains can be found in almost all pig populations worldwide, making detection of the S. suis species in asymptomatic carrier herds of little practical value in predicting the likelihood of future clinical relevance. The value of future molecular tools for surveillance and preventative health management lies in the detection of strains that genetically have increased potential to cause disease in presently healthy animals. Here we describe the use of genome-wide association studies to identify genetic markers associated with the observed clinical phenotypes (i) invasive disease and (ii) asymptomatic carriage on the palatine tonsils of pigs on UK farms. Subsequently, we designed a multiplex PCR to target three genetic markers that differentiated 115 S. suis isolates into disease-associated and non-disease-associated groups, that performed with a sensitivity of 0.91, a specificity of 0.79, a negative predictive value of 0.91, and a positive predictive value of 0.79 in comparison to observed clinical phenotypes. We describe evaluation of our pathotyping tool, using an out-of-sample collection of 50 previously uncharacterized S. suis isolates, in comparison to existing methods used to characterize and subtype S. suis isolates. In doing so, we show our pathotyping approach to be a competitive method to characterize S. suis isolates recovered from pigs on UK farms and one that can easily be updated to incorporate global strain collections.This work was supported by a Biotechnology and Biological Sciences Research Council (BBSRC) Knowledge Transfer Network CASE studentship co-funded by Zoetis (previously Pfizer Animal Health UK) and with significant contribution from BQP Ltd (Award Reference: BB/L502479/1). Funding bodies provided scholarship support but had no part in study design, data collection, analysis and interpretation of data or in writing the manuscript. AWT is supported by a BBSRC Longer and Larger (LoLa) grant (Award Reference: BB/G019274/1). LAW is supported by a Dorothy Hodgkin Fellowship funded by the Royal Society (Grant Number: DH140195) and a Sir Henry Dale Fellowship co-funded by the Royal Society and Wellcome Trust (Grant Number: 109385/Z/15/Z)

The N-linking glycosylation system from Actinobacillus pleuropneumoniae is required for adhesion and has potential use in glycoengineering.

Actinobacillus pleuropneumoniae is a mucosal respiratory pathogen causing contagious porcine pleuropneumonia. Pathogenesis studies have demonstrated a major role for the capsule, exotoxins and outer membrane proteins. Actinobacillus pleuropneumoniae can also glycosylate proteins, using a cytoplasmic N-linked glycosylating enzyme designated NGT, but its transcriptional arrangement and role in virulence remains unknown. We investigated the NGT locus and demonstrated that the putative transcriptional unit consists of rimO, ngt and a glycosyltransferase termed agt. From this information we used the A. pleuropneumoniae glycosylation locus to decorate an acceptor protein, within Escherichia coli, with a hexose polymer that reacted with an anti-dextran antibody. Mass spectrometry analysis of a truncated protein revealed that this operon could add up to 29 repeat units to the appropriate sequon. We demonstrated the importance of NGT in virulence, by creating deletion mutants and testing them in a novel respiratory cell line adhesion model. This study demonstrates the importance of the NGT glycosylation system for pathogenesis and its potential biotechnological application for glycoengineering.This work was supported by a Longer and Larger (LoLa) grant from the UK Biotechnology and Biological Sciences Research Council (grant nos BB/G020744/1, BB/G019177/1, BB/G019274/1 and BB/G003203/1) and The Wellcome Trust (grant no. 102979/Z/13/Z)

Refining the transit-timing and photometric analysis of TRAPPIST-1: Masses, Radii, densities, dynamics, and ephemerides

We have collected transit times for the TRAPPIST-1 system with the Spitzer Space Telescope over four years. We add to these ground-based, HST and K2 transit time measurements, and revisit an N-body dynamical analysis of the seven-planet system using our complete set of times from which we refine the mass ratios of the planets to the star. We next carry out a photodynamical analysis of the Spitzer light curves to derive the density of the host star and the planet densities. We find that all seven planets' densities may be described with a single rocky mass-radius relation which is depleted in iron relative to Earth, with Fe 21 wt% versus 32 wt% for Earth, and otherwise Earth-like in composition. Alternatively, the planets may have an Earth-like composition, but enhanced in light elements, such as a surface water layer or a core-free structure with oxidized iron in the mantle. We measure planet masses to a precision of 3-5%, equivalent to a radial-velocity (RV) precision of 2.5 cm/sec, or two orders of magnitude more precise than current RV capabilities. We find the eccentricities of the planets are very small; the orbits are extremely coplanar; and the system is stable on 10 Myr timescales. We find evidence of infrequent timing outliers which we cannot explain with an eighth planet; we instead account for the outliers using a robust likelihood function. We forecast JWST timing observations, and speculate on possible implications of the planet densities for the formation, migration and evolution of the planet system

Trial baseline characteristics of a cluster randomised controlled trial of a school-located obesity prevention programme; the Healthy Lifestyles Programme (HeLP) trial

This is the final version of the article. Available from BioMed Central via the DOI in this record.Background We have developed a healthy lifestyles programme (HeLP) for primary school aged children (9–10 years), currently being evaluated in a definitive cluster randomised controlled trial. This paper descriptively presents the baseline characteristics of trial children (BMI, waist circumference, % body fat, diet and physical activity) by gender, cluster level socio-economic status, school size and time of recruitment into the trial. Methods Schools were recruited from across the South West of England and allocated 1:1 to either intervention (HeLP) or control (usual practice) stratified by the proportion of children eligible for free school meals (FSM, 1 Year 5 class). The primary outcome is change in body mass index standard deviation score (BMI sds) at 24 months post-randomisation. Secondary outcomes are BMI sds at 18 months, waist circumference and percentage body fat sds at 18 and 24 months, proportion of children classified as underweight, overweight and obese at 18 and 24 months, physical activity (for a sub-sample) and food intake at 18 months. Results At baseline 11.4% and 13.6% of children were categorised as overweight or obese respectively. A higher percentage of girls than boys (25.3% vs 24.8%) and children from schools in FSM category 2 (28.2% vs 23.2%) were overweight or obese. Children were consuming a mean (range) of 4.15 (0–13) energy dense snacks (EDS) and 3.23 (0–9) healthy snacks (HS) per day with children from schools in FSM category 2 consuming more EDS and negative food markers and less HS and positive food markers. Children spent an average 53.6 min per day (11.9 to 124.8) in MVPA and thirteen hours (779.3 min) per day (11 h to 15 h) doing less than ‘light’ intensity activity. Less than 5% of children achieved the Departments of Health’s recommendation of 60 min of MVPA every day. Conclusion We have excellent completeness of baseline data for all measures and have achieved compliance to accelerometry not seen before in other large scale studies. Our anthropometric baseline data is representative of local and national data for children this age and reflects the gender and socio-economic variations expected of children this age in relation to physical activity and weight status.The definitive trial of HeLP is funded by the UK National Institute for Health Research (NIHR) Public Health Research Programme (10/3010/01) and a full report will be published on the NIHR website. Intervention materials and delivery was funded by the Peninsula College of Medicine and Dentistry. PenCLAHRC provided methodological support during the transition from the exploratory trial to the definitive evaluation

Polymorphism in a lincRNA Associates with a Doubled Risk of Pneumococcal Bacteremia in Kenyan Children.

Bacteremia (bacterial bloodstream infection) is a major cause of illness and death in sub-Saharan Africa but little is known about the role of human genetics in susceptibility. We conducted a genome-wide association study of bacteremia susceptibility in more than 5,000 Kenyan children as part of the Wellcome Trust Case Control Consortium 2 (WTCCC2). Both the blood-culture-proven bacteremia case subjects and healthy infants as controls were recruited from Kilifi, on the east coast of Kenya. Streptococcus pneumoniae is the most common cause of bacteremia in Kilifi and was thus the focus of this study. We identified an association between polymorphisms in a long intergenic non-coding RNA (lincRNA) gene (AC011288.2) and pneumococcal bacteremia and replicated the results in the same population (p combined = 1.69 × 10(-9); OR = 2.47, 95% CI = 1.84-3.31). The susceptibility allele is African specific, derived rather than ancestral, and occurs at low frequency (2.7% in control subjects and 6.4% in case subjects). Our further studies showed AC011288.2 expression only in neutrophils, a cell type that is known to play a major role in pneumococcal clearance. Identification of this novel association will further focus research on the role of lincRNAs in human infectious disease.Wellcome Trust (Grant ID: 084716/Z/08/Z)This is the final version of the article. It first appeared from Cell Press/Elsevier via http://dx.doi.org/10.1016/j.ajhg.2016.03.02

The relationship between the three-dimensional structure of porous GaN distributed Bragg reflectors and their birefringence

Porous GaN DBRs offer an opportunity to provide the high reflectance, lattice-matched components required for efficient GaN VCSELs. The birefringence of these structures is therefore of key interest, as it could be used to control the polarization of the emitted light. Here we present a detailed analysis of the optical birefringence for both laterally etched, patterned structures and selfassembled radial porous structures. We correlate this with the 3D structure of the pores, which we measure through the use of FIB milling and serial block-face SEM imaging (SBI). This is a powerful method for imaging the internal nanostructure of the sample and allows the internal pore morphology to be viewed in a reconstruction of any 3D plane. We measure the birefringence of our porous GaN layers as Δ=0.14, with a lower refractive index parallel to the pores (∥) than perpendicular to them (⟂). Using finite element modelling we accurately reproduce the experimentally observed birefringence trends and find that this can be done by modelling GaN as a perfect dielectric. This indicates that the birefringence arises from the limited width across the pores. This also shows that standard modelling approaches can be used to design porous GaN birefringent devices effectively