Discordant bioinformatic predictions of antimicrobial resistance from whole-genome sequencing data of bacterial isolates: an inter-laboratory study.

Antimicrobial resistance (AMR) poses a threat to public health. Clinical microbiology laboratories typically rely on culturing bacteria for antimicrobial-susceptibility testing (AST). As the implementation costs and technical barriers fall, whole-genome sequencing (WGS) has emerged as a 'one-stop' test for epidemiological and predictive AST results. Few published comparisons exist for the myriad analytical pipelines used for predicting AMR. To address this, we performed an inter-laboratory study providing sets of participating researchers with identical short-read WGS data from clinical isolates, allowing us to assess the reproducibility of the bioinformatic prediction of AMR between participants, and identify problem cases and factors that lead to discordant results. We produced ten WGS datasets of varying quality from cultured carbapenem-resistant organisms obtained from clinical samples sequenced on either an Illumina NextSeq or HiSeq instrument. Nine participating teams ('participants') were provided these sequence data without any other contextual information. Each participant used their choice of pipeline to determine the species, the presence of resistance-associated genes, and to predict susceptibility or resistance to amikacin, gentamicin, ciprofloxacin and cefotaxime. We found participants predicted different numbers of AMR-associated genes and different gene variants from the same clinical samples. The quality of the sequence data, choice of bioinformatic pipeline and interpretation of the results all contributed to discordance between participants. Although much of the inaccurate gene variant annotation did not affect genotypic resistance predictions, we observed low specificity when compared to phenotypic AST results, but this improved in samples with higher read depths. Had the results been used to predict AST and guide treatment, a different antibiotic would have been recommended for each isolate by at least one participant. These challenges, at the final analytical stage of using WGS to predict AMR, suggest the need for refinements when using this technology in clinical settings. Comprehensive public resistance sequence databases, full recommendations on sequence data quality and standardization in the comparisons between genotype and resistance phenotypes will all play a fundamental role in the successful implementation of AST prediction using WGS in clinical microbiology laboratories

Combined bezafibrate, medroxyprogesterone acetate and valproic acid treatment inhibits osteosarcoma cell growth without adversely affecting normal mesenchymal stem cells.

This document is the Accepted Manuscript version of a published work that appeared in final form in Bioscience Reports. To access the final edited and published work see http://dx.doi.org/10.1042/BSR20202505Drug repurposing is a cost effective means of targeting new therapies for cancer. We have examined the effects of the repurposed drugs, bezafibrate, medroxyprogesterone acetate and valproic acid on human osteosarcoma cells, i.e., SAOS2 and MG63 compared with their normal cell counterparts, i.e. mesenchymal stem/stromal cells (MSCs). Cell growth, viability and migration were measured by biochemical assay and live cell imaging, whilst levels of lipid-synthesising enzymes were measured by immunoblotting cell extracts. These drug treatments inhibited the growth and survival of SAOS2 and MG63 cells most effectively when used in combination (termed V-BAP). In contrast, V-BAP treated MSCs remained viable with only moderately reduced cell proliferation. V-BAP treatment also inhibited migratory cell phenotypes. MG63 and SAOS2 cells expressed much greater levels of fatty acid synthase and stearoyl CoA desaturase 1 than MSCs, but these elevated enzyme levels significantly decreased in the V-BAP treated osteosarcoma cells prior to cell death. Hence, we have identified a repurposed drug combination that selectively inhibits the growth and survival of human osteosarcoma cells in association with altered lipid metabolism without adversely affecting their non-transformed cell counterparts

“I’m a Red River local”: rock climbing mobilities and community hospitalities

With individuals continually on the move, mobility fosters constellations of places at which individuals collectively moor and perform community. By focusing on one climbing destination – the Red River Gorge – this paper works across scales to highlight the spatial politics of mobilizing hospitality. In so doing, it summarizes the ways hosting/guesting thresholds dissolve with the growth of particular rock climbing associated infrastructures and moves to examine the ways climbers performances of community result in the (semi-)privatization of public space and attempts at localization. Further, the paper highlights the ways mobility is employed to maintain a political voice from afar, as well as to forge “local” identities with The Red as place with distinct subcultural (in)hospitality practices. Hospitality practices affirm power relations, they communicate who is at “home” and who has the power in a particular space to extend hospitality. The decision to extend hospitality is not simply the difference between an ethical encounter and a conditional one; it takes place in the very performance of identity. Thus, integrating a mobilities perspective into hospitality studies further illuminates the spatial politics that are at play in an ethics of hospitality

The role of the genetic counsellor: a systematic review of research evidence

In Europe, genetic counsellors are employed in specialist genetic centres or other specialist units. According to the European Board of Medical Genetics, the genetic counsellor must fulfil a range of roles, including provision of information and facilitation of psychosocial adjustment of the client to their genetic status and situation. To evaluate the extent to which genetic counsellors fulfil their prescribed roles, we conducted a systematic review of the published relevant scientific evidence. We searched five relevant electronic databases (Medline, CINAHL, SocIndex, AMED and PsychInfo) using relevant search terms and handsearched four subject-specific journals for research-based papers published in English between 1 January 2000 and 30 June 2013. Of 419 potential papers identified initially, seven satisfied the inclusion criteria for the review. Themes derived from the thematic analysis of the data were: (i) rationale for genetic counsellors to provide care, (ii) appropriate roles and responsibilities and (iii) the types of conditions included in the genetic counsellor caseload. The findings of this systematic review indicate that where genetic counsellors are utilised in specialist genetic settings, they undertake a significant workload associated with direct patient care and this appears to be acceptable to patients. With the burden on genetic services, there is an argument for the increased use of genetic counsellors in countries where they are under-utilised. In addition, roles undertaken by genetic counsellors in specialist genetic settings could be adapted to integrate genetic counsellors into multi-disciplinary teams in other specialisms

Results of two multi-chord stellar occultations by dwarf planet (1) Ceres

We report the results of two multi-chord stellar occultations by the dwarf planet (1) Ceres that were observed from Brazil on 2010 August 17, and from the USA on 2013 October 25. Four positive detections were obtained for the 2010 occultation, and nine for the 2013 occultation. Elliptical models were adjusted to the observed chords to obtain Ceres' size and shape. Two limb fitting solutions were studied for each event. The first one is a nominal solution with an indeterminate polar aspect angle. The second one was constrained by the pole coordinates as given by Drummond et al. Assuming a Maclaurin spheroid, we determine an equatorial diameter of 972 $\pm$ 6 km and an apparent oblateness of 0.08 $\pm$ 0.03 as our best solution. These results are compared to all available size and shape determinations for Ceres made so far, and shall be confirmed by the NASA's Dawn space mission.Comment: 9 pages, 6 figures. Accepted for publication in MNRA

Key principle of the efficient running, swimming, and flying

Empirical observations indicate striking similarities among locomotion in terrestrial animals, birds, and fish, but unifying physical grounds are lacking. When applied to efficient locomotion, the analytical mechanics principle of minimum action yields two patterns of mechanical similarity via two explicit spatiotemporal coherent states. In steady locomotory modes, the slow muscles determining maximal optimum speeds maintain universal intrinsic muscular pressure. Otherwise, maximal speeds are due to constant mass-dependent stiffness of fast muscles generating a uniform force field, exceeding gravitation. Being coherent in displacements, velocities and forces, the body appendages of animals are tuned to natural propagation frequency through the state-dependent elastic muscle moduli. Key words: variational principle of minimum action (04.20.Fy), locomotion (87.19.ru), biomechanics (87.85.G-).Comment: Submitted to the Europhysical Letter

Thiamin and Riboflavin in Human Milk: Effects of Lipid-Based Nutrient Supplementation and Stage of Lactation on Vitamer Secretion and Contributions to Total Vitamin Content

While thiamin and riboflavin in breast milk have been analyzed for over 50 years, less attention has been given to the different forms of each vitamin. Thiamin-monophosphate (TMP) and free thiamin contribute to total thiamin content; flavin adenine-dinucleotide (FAD) and free riboflavin are the main contributors to total riboflavin. We analyzed milk collected at 2 (n = 258) or 6 (n = 104), and 24 weeks (n = 362) from HIV-infected Malawian mothers within the Breastfeeding, Antiretrovirals and Nutrition (BAN) study, randomly assigned at delivery to lipid-based nutrient supplements (LNS) or a control group, to investigate each vitamer’s contribution to total milk vitamin content and the effects of supplementation on the different thiamin and riboflavin vitamers at early and later stages of lactation, and obtain insight into the transport and distribution of these vitamers in human milk. Thiamin vitamers were derivatized into thiochrome-esters and analyzed by high-performance liquid-chromatography-fluorescence-detection (HPLC-FLD). Riboflavin and FAD were analyzed by ultra-performance liquid-chromatography-tandem-mass-spectrometry (ULPC-MS/MS). Thiamin-pyrophosphate (TPP), identified here for the first time in breast milk, contributed 1.9–4.5% to total thiamin. Free thiamin increased significantly from 2/6 to 24 weeks regardless of treatment indicating an active transport of this vitamer in milk. LNS significantly increased TMP and free thiamin only at 2 weeks compared to the control: median 170 versus 151μg/L (TMP), 13.3 versus 10.5μg/L (free thiamin, p<0.05 for both, suggesting an up-regulated active mechanism for TMP and free thiamin accumulation at early stages of lactation. Free riboflavin was consistently and significantly increased with LNS (range: 14.8–19.6μg/L (LNS) versus 5.0–7.4μg/L (control), p<0.001), shifting FAD:riboflavin relative amounts from 92–94:6–8% to 85:15%, indicating a preferred secretion of the free form into breast milk. The continuous presence of FAD in breast milk suggests an active transport and secretion system for this vitamer or possibly formation of this co-enymatic form in the mammary gland

The diversity of Klebsiella pneumoniae surface polysaccharides.

Klebsiella pneumoniae is considered an urgent health concern due to the emergence of multi-drug-resistant strains for which vaccination offers a potential remedy. Vaccines based on surface polysaccharides are highly promising but need to address the high diversity of surface-exposed polysaccharides, synthesized as O-antigens (lipopolysaccharide, LPS) and K-antigens (capsule polysaccharide, CPS), present in K. pneumoniae. We present a comprehensive and clinically relevant study of the diversity of O- and K-antigen biosynthesis gene clusters across a global collection of over 500 K. pneumoniae whole-genome sequences and the seroepidemiology of human isolates from different infection types. Our study defines the genetic diversity of O- and K-antigen biosynthesis cluster sequences across this collection, identifying sequences for known serotypes as well as identifying novel LPS and CPS gene clusters found in circulating contemporary isolates. Serotypes O1, O2 and O3 were most prevalent in our sample set, accounting for approximately 80 % of all infections. In contrast, K serotypes showed an order of magnitude higher diversity and differ among infection types. In addition we investigated a potential association of O or K serotypes with phylogenetic lineage, infection type and the presence of known virulence genes. K1 and K2 serotypes, which are associated with hypervirulent K. pneumoniae, were associated with a higher abundance of virulence genes and more diverse O serotypes compared to other common K serotypes

Lung Ultrasound As A Diagnostic Tool for Radiographically-Confirmed Pneumonia in Low Resource Settings

Background Pneumonia is a leading cause of morbidity and mortality in children worldwide; however, its diagnosis can be challenging, especially in settings where skilled clinicians or standard imaging are unavailable. We sought to determine the diagnostic accuracy of lung ultrasound when compared to radiographically-confirmed clinical pediatric pneumonia. Methods Between January 2012 and September 2013, we consecutively enrolled children aged 2–59 months with primary respiratory complaints at the outpatient clinics, emergency department, and inpatient wards of the Instituto Nacional de Salud del Niño in Lima, Peru. All participants underwent clinical evaluation by a pediatrician and lung ultrasonography by one of three general practitioners. We also consecutively enrolled children without respiratory symptoms. Children with respiratory symptoms had a chest radiograph. We obtained ancillary laboratory testing in a subset. Results Final clinical diagnoses included 453 children with pneumonia, 133 with asthma, 103 with bronchiolitis, and 143 with upper respiratory infections. In total, CXR confirmed the diagnosis in 191 (42%) of 453 children with clinical pneumonia. A consolidation on lung ultrasound, which is our primary endpoint for pneumonia, had a sensitivity of 88.5%, specificity of 100%, and an area under-the-curve of 0.94 (95% CI 0.92–0.97) when compared to radiographically-confirmed clinical pneumonia. When any abnormality on lung ultrasound was compared to radiographically-confirmed clinical pneumonia the sensitivity increased to 92.2% and the specificity decreased to 95.2%, with an area under-the-curve of 0.94 (95% CI 0.91–0.96). Conclusions Lung ultrasound had high diagnostic accuracy for the diagnosis of radiographically-confirmed pneumonia. Added benefits of lung ultrasound include rapid testing and high inter-rater agreement. Lung ultrasound may serve as an alternative tool for the diagnosis of pediatric pneumonia

COVID-19 and the potential long-term impact on antimicrobial resistance

The emergence of the Severe Acute Respiratory Syndrome coronavirus 2 (SARS-CoV-2) has required an unprecedented response to control the spread of the infection and protect the most vulnerable within society. Whilst the pandemic has focused society on the threat of emerging infections and hand hygiene, certain infection control and antimicrobial stewardship policies may have to be relaxed. It is unclear whether the unintended consequences of these changes will have a net-positive or -negative impact on rates of antimicrobial resistance. Whilst the urgent focus must be on controlling this pandemic, sustained efforts to address the longer-term global threat of antimicrobial resistance should not be overlooked