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Abstract Background Delayed graft function, the requirement for dialysis due to poor kidney function post-transplant, is a frequent complication of deceased donor kidney transplantation and is associated with inferior outcomes and higher costs. Intravenous fluids given during and after transplantation may affect the risk of poor kidney function after transplant. The most commonly used fluid, isotonic sodium chloride (0.9% saline), contains a high chloride concentration, which may be associated with acute kidney injury, and could increase the risk of delayed graft function. Whether using a balanced, low-chloride fluid instead of 0.9% saline is safe and improves kidney function after deceased donor kidney transplantation is unknown. Methods BEST-Fluids is an investigator-initiated, pragmatic, registry-based, multi-center, double-blind, randomized controlled trial. The primary objective is to compare the effect of intravenous Plasma-Lyte 148 (Plasmalyte), a balanced, low-chloride solution, with the effect of 0.9% saline on the incidence of delayed graft function in deceased donor kidney transplant recipients. From January 2018 onwards, 800 participants admitted for deceased donor kidney transplantation will be recruited over 3 years in Australia and New Zealand. Participants are randomized 1:1 to either intravenous Plasmalyte or 0.9% saline peri-operatively and until 48 h post-transplant, or until fluid is no longer required; whichever comes first. Follow up is for 1 year. The primary outcome is the incidence of delayed graft function, defined as dialysis in the first 7 days post-transplant. Secondary outcomes include early kidney transplant function (composite of dialysis duration and rate of improvement in graft function when dialysis is not required), hyperkalemia, mortality, graft survival, graft function, quality of life, healthcare resource use, and cost-effectiveness. Participants are enrolled, randomized, and followed up using the Australia and New Zealand Dialysis and Transplant (ANZDATA) Registry. Discussion If using Plasmalyte instead of 0.9% saline is effective at reducing delayed graft function and improves other clinical outcomes in deceased donor kidney transplantation, this simple, inexpensive change to using a balanced low-chloride intravenous fluid at the time of transplantation could be easily implemented in the vast majority of transplant settings worldwide. Trial registration Australian New Zealand Clinical Trials Registry: ACTRN12617000358347 . Registered on 8 March 2017. ClinicalTrials.gov: NCT03829488 . Registered on 4 February 2019.

Abstract Background Many of the commonly used methods for orthology detection start from mutually most similar pairs of genes (reciprocal best hits) as an approximation for evolutionary most closely related pairs of genes (reciprocal best matches). This approximation of best matches by best hits becomes exact for ultrametric dissimilarities, i.e., under the Molecular Clock Hypothesis. It fails, however, whenever there are large lineage specific rate variations among paralogous genes. In practice, this introduces a high level of noise into the input data for best-hit-based orthology detection methods. Results If additive distances between genes are known, then evolutionary most closely related pairs can be identified by considering certain quartets of genes provided that in each quartet the outgroup relative to the remaining three genes is known. A priori knowledge of underlying species phylogeny greatly facilitates the identification of the required outgroup. Although the workflow remains a heuristic since the correct outgroup cannot be determined reliably in all cases, simulations with lineage specific biases and rate asymmetries show that nearly perfect results can be achieved. In a realistic setting, where distances data have to be estimated from sequence data and hence are noisy, it is still possible to obtain highly accurate sets of best matches. Conclusion Improvements of tree-free orthology assessment methods can be expected from a combination of the accurate inference of best matches reported here and recent mathematical advances in the understanding of (reciprocal) best match graphs and orthology relations. Availability Accompanying software is available at https://github.com/david-schaller/AsymmeTree .

Abstract Three‐dimensional biofabrication using photo‐crosslinkable hydrogel bioscaffolds has the potential to revolutionize the need for transplants and implants in joints, with articular cartilage being an early target tissue. However, to successfully translate these approaches to clinical practice, several barriers must be overcome. In particular, the photo‐crosslinking process may impact on cell viability and DNA integrity, and consequently on chondrogenic differentiation. In this review, we primarily explore the specific sources of cellular cytotoxicity and genotoxicity inherent to the photo‐crosslinking reaction, the methods to analyze cell death, cell metabolism, and DNA damage within the bioscaffolds, and the possible strategies to overcome these detrimental effects.

A novel bismuth(III) complex with 2-amino-3-carbomethoxy-4,5,6,7-tetrahydrobenzo[b]thiophene (ACTT) as a ligand have been synthesized. The novel complex was characterized on the basis of its IR, NMR, elemental analysis and MS spectral data. It was found that the ligand behaves as a monodentate chelating agent and bonds to the metal ion through the nitrogen atom of the amino group to form the [BiIII(ACTT)6]Cl3 complex. The new complex compound displayed significant antimicrobial activity (MIC = 8–32 μg/mL) against Bacillus subtilis, Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa, Shigella flexneri, Candida albicans, Candida tropicalis and Cryptococcus neoformans.

Abstract Background Although education, exercise, and weight loss are recommended for management of knee osteoarthritis, the additional benefits of incorporating weight loss strategies into exercise interventions have not been well investigated. The aim of this study is to compare, in a private health insurance setting, the clinical- and cost-effectiveness of a remotely-delivered, evidence- and theory-informed, behaviour change intervention targeting exercise and self-management (Exercise intervention), with the same intervention plus active weight management (Exercise plus weight management intervention), and with an information-only control group for people with knee osteoarthritis who are overweight or obese. Methods Three-arm, pragmatic parallel-design randomised controlled trial involving 415 people aged ≥45 and ≤ 80 years, with body mass index ≥28 kg/m2 and

Abstract Despite the success of cross-sectional imaging in evaluating hepatopancreatobiliary system malignancies, several non-malignant disease processes may closely mimic malignancy. Differentiating these benign diseases from malignancy may be difficult, or even impossible, even in the hands of experienced imagers. In this manuscript, we present benign mimics involving the hepatopancreatobiliary system and try to increase awareness of these potential pitfalls.

The communication practices between educators, administrators, government officials, and students were key features of educational technology leadership in Western Canada. This paper presents the findings of an exhaustive study of all 75 large K-12 districts in Canada's three westernmost provinces: British Columbia, Alberta, and Saskatchewan. A data transformation model mixed methods triangulation design methodology was used in this study of over 1.1 million students across a geography of over 2.2 million square kilometers. Western Canadian K-12 relies heavily on cloud computing, and this approach enabled a successful shift to online education during COVID-19. What emerged from this research were several organizational collaboration practices that enabled inter-disciplinary communication 1i. These practices produced this highly robust IT infrastructure. This paper will present these collaborative communication practices and the formalized organizational initiatives that enable them. The extent and impact of these interdisciplinary collaborative communication practices were so profound that they nullified differences in school district size and the locus of authority for educational technology decision-making.

Abstract Background The survival of INA-6 human multiple myeloma cells is strictly dependent upon the Interleukin-6-activated transcription factor STAT3. Although transcriptional analyses have revealed many genes regulated by STAT3, to date no protein-coding STAT3 target gene is known to mediate survival in INA-6 cells. Therefore, the aim here was to identify and analyze non-protein-coding STAT3 target genes. In addition to the oncogenic microRNA-21, we previously described five long noncoding RNAs (lncRNAs) induced by STAT3, named STAiRs. Here, we focus on STAT3-induced RNA 18 (STAiR18), an mRNA-like, long ncRNA that is duplicated in the human lineage. One STAiR18 locus is annotated as the already well described LINC00152/CYTOR, however, the other harbors the MIR4435-2HG gene and is, up to now, barely described. Methods CAPTURE-RNA-sequencing was used to analyze STAiR18 transcript architecture. To identify the STAiR18 and STAT3 phenotype, siRNA-based knockdowns were performed and microarrays were applied to identify their target genes. RNA-binding partners of STAiR18 were determined by Chromatin-Isolation-by-RNA-Purification (ChIRP) and subsequent sequencing. STAT3 expression in dependence of STAiR18 was investigated by immunoblots, chromatin- and RNA-immunoprecipitations. Results As identified by CAPTURE-RNA sequencing, a complex splice pattern originates from both STAiR18 loci, generating different transcripts. Knockdown of the most abundant STAiR18 isoforms dramatically decreased INA-6 cell vitality, suggesting a functional role in myeloma cells. Additionally, STAiR18 and STAT3 knockdowns yielded overlapping changes of transcription patterns in INA-6 cells, suggesting a close functional interplay between the two factors. Moreover, Chromatin isolation by RNA purification (ChIRP), followed by genome-wide RNA sequencing showed that STAiR18 associates specifically with the STAT3 primary transcript. Furthermore, the knockdown of STAiR18 reduced STAT3 levels on both the RNA and protein levels, suggesting a positive feedback between both molecules. Furthermore, STAiR18 knockdown changes the histone methylation status of the STAT3 locus, which explains the positive feedback and indicates that STAiR18 is an epigenetic modulator. Conclusion Hence, STAiR18 is an important regulator of myeloma cell survival and is strongly associated with the oncogenic function of STAT3. The close functional interplay between STAT3 and STAiR18 suggests a novel principle of regulatory interactions between long ncRNAs and signaling pathways.

-point crossover operators and their recombination sets are studied from different perspectives. We show that transit functions of -point crossover generate, for all , the same convexity as the interval function of the underlying graph. This settles in the negative an open problem by Mulder about whether the geodesic convexity of a connected graph is uniquely determined by its interval function . The conjecture of Gitchoff and Wagner that for each transit set distinct from a hypercube there is a unique pair of parents from which it is generated is settled affirmatively. Along the way we characterize transit functions whose underlying graphs are Hamming graphs, and those with underlying partial cube graphs. For general values of it is shown that the transit sets of -point crossover operators are the subsets with maximal Vapnik–Chervonenkis dimension.

Abstract Background The domestic cat (Felis catus) is an important companion animal and is used as a large animal model for human disease. However, the comprehensive study of adaptive immunity in this species is hampered by the lack of data on lymphocyte antigen receptor genes and usage. The objectives of this study were to annotate the feline T cell receptor (TR) loci and to characterize the expressed repertoire in lymphoid organs of normal cats using high-throughput sequencing. Results The Felis catus TRG locus contains 30 genes: 12 TRGV, 12 TRGJ and 6 TRGC, the TRB locus contains 48 genes: 33 TRBV, 2 TRBD, 11 TRBJ, 2 TRBC, the TRD locus contains 19 genes: 11 TRDV, 2 TRDD, 5 TRDJ, 1 TRDC, and the TRA locus contains 127 genes: 62 TRAV, 64 TRAJ, 1 TRAC. Functional feline V genes form monophyletic clades with their orthologs, and clustering of multimember subgroups frequently occurs in V genes located at the 5′ end of TR loci. Recombination signal (RS) sequences of the heptamer and nonamer of functional V and J genes are highly conserved. Analysis of the TRG expressed repertoire showed preferential intra-cassette over inter-cassette rearrangements and dominant usage of the TRGV2–1 and TRGJ1–2 genes. The usage of TRBV genes showed minor bias but TRBJ genes of the second J-C-cluster were more commonly rearranged than TRBJ genes of the first cluster. The TRA/TRD V genes almost exclusively rearranged to J genes within their locus. The TRAV/TRAJ gene usage was relatively balanced while the TRD repertoire was dominated by TRDJ3. Conclusions This is the first description of all TR loci in the cat. The genomic organization of feline TR loci was similar to that of previously described jawed vertebrates (gnathostomata) and is compatible with the birth-and-death model of evolution. The large-scale characterization of feline TR genes provides comprehensive baseline data on immune repertoires in healthy cats and will facilitate the development of improved reagents for the diagnosis of lymphoproliferative diseases in cats. In addition, these data might benefit studies using cats as a large animal model for human disease.

Quantum computers with thousands or millions of qubits will require a scalable solution for qubit control and readout electronics. Colocating these electronics at millikelvin temperatures has been proposed and demonstrated, but there exist significant challenges with power dissipation, reproducibility, fidelity, and scalability. In this article, we experimentally demonstrate the use of a Josephson arbitrary waveform synthesizer (JAWS) to generate control signals at 4 K and perform spectroscopy of two components of a typical superconducting quantum information system: a linear resonator and a (nonlinear) transmon qubit. By locating the JAWS chip at 4 K and a qubit at 0.1 K, the direct path for quasi-particle poisoning from the JAWS chip to the qubit is broken. We demonstrate the stable, self-calibrated, and reproducible output signal of the JAWS when operated in its quantum locking range, a feature that allows these synthesizers to be replicated and scaled in the cryostat, all with identical on-chip, quantized, outputs. This is a proof-of-concept demonstration to generate signals at 4 K using driven superconducting electronics to control qubits at lower temperatures.

Current diagnostic criteria for schizophrenia place emphasis on delusions and hallucinations, whereas the classical descriptions of schizophrenia by Kraepelin and Bleuler emphasized disorganization and impoverishment of mental activity. Despite the availability of antipsychotic medication for treating delusions and hallucinations, many patients continue to experience persisting disability. Improving treatment requires a better understanding of the processes leading to persisting disability. We recently introduced the term classical schizophrenia to describe cases with disorganized and impoverished mental activity, cognitive impairment and predisposition to persisting disability. Recent evidence reveals that a polygenic score indicating risk for schizophrenia predicts severity of the features of classical schizophrenia: disorganization, and to a lesser extent, impoverishment of mental activity and cognitive impairment. Current understanding of brain function attributes a cardinal role to predictive coding: the process of generating models of the world that are successively updated in light of confirmation or contradiction by subsequent sensory information. It has been proposed that abnormalities of these predictive processes account for delusions and hallucinations. Here we examine the evidence provided by electrophysiology and fMRI indicating that imprecise predictive coding is the core pathological process in classical schizophrenia, accounting for disorganization, psychomotor poverty and cognitive impairment. Functional imaging reveals aberrant brain activity at network hubs engaged during encoding of predictions. We discuss the possibility that frequent prediction errors might promote excess release of the neurotransmitter, dopamine, thereby accounting for the occurrence of episodes of florid psychotic symptoms including delusions and hallucinations in classical schizophrenia. While the predictive coding hypotheses partially accounts for the time-course of classical schizophrenia, the overall body of evidence indicates that environmental factors also contribute. We discuss the evidence that chronic inflammation is a mechanism that might link diverse genetic and environmental etiological factors, and contribute to the proposed imprecision of predictive coding.

In the past 30 years there has been a significant increase in the number of publications on phospholipid (PL) metabolism, both for the medical purposes of detection and diagnosis of cancer and for the monitoring of the treatment of human cancers. Most of the work has focused on the pathway that produces phosphatidylcholine, the major component of human cell membranes. The trigger for this research was the advent of applications of NMR spectroscopy in vitro and in vivo in the 1980’s and observations that most cancer cells and tumors had significant increases in the water-soluble PL precursors and breakdown products. Increased phosphocholine (PC) has been focused on as a marker for cancer using Magnetic Resonance Spectroscopy (MRS) and Positron Emission Tomography (PET). MRS is now used clinically to aid in the diagnosis and severity of some brain tumors; and choline PET is used for the diagnosis and staging of recurrent prostate cancer, paid for by medical insurance companies. Another major area of research starting in the 1990’s was the development of specific choline kinase (CK) inhibitors aimed at the isoenzyme CK-a. This isoenzyme is markedly upregulated in cancer cells and unexpectedly was found to have a role in oncogenic transformation independent of its enzyme function.

In West Africa, kidney diseases are frequently seen, but diagnostic and therapeutic options are poor due to limited access to specialized facilities. To unravel the etiology and develop clinical guidelines, we collected clinical data and results of kidney biopsies in 121 pediatric and mostly young adult patients with edema and proteinuria in The Gambia. Workup included clinical examination, urine and serum analysis, and kidney biopsy findings. Selected cases were treated with steroids.ResultsThe median age was 14.9 years (range 1.8–52.0) at presentation. The most frequent underlying histologies were post-infectious glomerulonephritis (PIGN) in 38%, focal-segmental glomerulosclerosis (FSGS) in 30%, minimal change nephrotic syndrome (MCNS) in 15%, and membranous glomerulonephritis (MGN) in 10% of cases. Patients with PIGN were significantly younger and had less proteinuria and higher serum albumin levels than the other three. Infected scabies was seen more often in cases with PIGN. Clinical parameters could not distinguish patients with FSGS, MCNS, and MGN. Steroid response was prompt in patients with MCNS (remission in 10/10 cases) compared to FSGS (4/19) and MGN (0/4). In summary, the clinical histopathological correlation allows a better approach to therapy and can be the basis for urgently needed interventional studies in steroid-resistant cases.

Mitochondria are multifunctional organelles of which ultrastructure is tightly linked to cell physiology. Accumulating evidence shows that mitochondrial remodeling has an impact on immune responses, but our current understanding of the mitochondrial architecture, interactions, and morphological changes in immune cells, mainly in eosinophils, is still poorly known. Here, we applied transmission electron microscopy (TEM), single-cell imaging analysis, and electron tomography, a technique that provides three-dimensional (3D) views at high resolution, to investigate mitochondrial dynamics in mouse eosinophils developing in cultures as well as in the context of inflammatory diseases characterized by recruitment and activation of these cells (mouse models of asthma, H1N1 influenza A virus (IAV) infection, and schistosomiasis mansoni). First, quantitative analyses showed that the mitochondrial area decrease 70% during eosinophil development (from undifferentiated precursor cells to mature eosinophils). Mitophagy, a consistent process revealed by TEM in immature but not in mature eosinophils, is likely operating in mitochondrial clearance during eosinophilopoiesis. Events of mitochondria interaction (inter-organelle membrane contacts) were also detected and quantitated within developing eosinophils and included mitochondria-endoplasmic reticulum, mitochondria-mitochondria, and mitochondria-secretory granules, all of them significantly higher in numbers in immature compared to mature cells. Moreover, single-mitochondrion analyses revealed that as the eosinophil matures, mitochondria cristae significantly increase in number and reshape to lamellar morphology. Eosinophils did not change (asthma) or reduced (IAV and Schistosoma infections) their mitochondrial mass in response to inflammatory diseases. However, asthma and schistosomiasis, but not IAV infection, induced amplification of both cristae numbers and volume in individual mitochondria. Mitochondrial cristae remodeling occurred in all inflammatory conditions with the proportions of mitochondria containing only lamellar or tubular, or mixed cristae (an ultrastructural aspect seen just in tissue eosinophils) depending on the tissue/disease microenvironment. The ability of mitochondria to interact with granules, mainly mobilized ones, was remarkably captured by TEM in eosinophils participating in all inflammatory diseases. Altogether, we demonstrate that the processes of eosinophilopoiesis and inflammation-induced activation interfere with the mitochondrial dynamics within mouse eosinophils leading to cristae remodeling and inter-organelle contacts. The understanding of how mitochondrial dynamics contribute to eosinophil immune functions is an open interesting field to be explored.

Preexisting antibodies to endemic coronaviruses (CoV) that cross-react with SARS-CoV-2 have the potential to influence the antibody response to COVID-19 vaccination and infection for better or worse. In this observational study of mucosal and systemic humoral immunity in acutely infected, convalescent, and vaccinated subjects, we tested for cross-reactivity against endemic CoV spike (S) protein at subdomain resolution. Elevated responses, particularly to the β-CoV OC43, were observed in all natural infection cohorts tested and were correlated with the response to SARS-CoV-2. The kinetics of this response and isotypes involved suggest that infection boosts preexisting antibody lineages raised against prior endemic CoV exposure that cross-react. While further research is needed to discern whether this recalled response is desirable or detrimental, the boosted antibodies principally targeted the better-conserved S2 subdomain of the viral spike and were not associated with neutralization activity. In contrast, vaccination with a stabilized spike mRNA vaccine did not robustly boost cross-reactive antibodies, suggesting differing antigenicity and immunogenicity. In sum, this study provides evidence that antibodies targeting endemic CoV are robustly boosted in response to SARS-CoV-2 infection but not to vaccination with stabilized S, and that depending on conformation or other factors, the S2 subdomain of the spike protein triggers a rapidly recalled, IgG-dominated response that lacks neutralization activity.

Introduction: Violence against women (VAW) can result in long-term and varied sequela for survivors, making it difficult to evaluate healthcare intervention. This study seeks to improve understanding of the healthcare experiences of women survivors prior to a violence-related diagnosis, allowing healthcare systems to better design strategies to meet the needs of this population. Methods: Using population-based data from 2016 to 2019, this cross-sectional observational study presents healthcare spending, utilization, and diagnostic patterns of privately insured women, age 18 or older, in the 10-months prior to an episode of care for a documented experience of violence (DEV). Results: Of 12 624 764 women meeting enrollment criteria, 10 980 women had DEV. This group had higher general medical complexity, despite being 10 years younger than the comparison group (mean age 32.7 vs 43.5). These relationships held up when comparing participants in each cohort by age. Additional key findings including higher numbers of medical visits across clinical settings and higher total cost ($10 138-$4585). Conclusions: The study utilized population-based data, to describe specific areas of health and medical cost for women with DEV. Increased medical complexity and utilization patterns among survivors broaden the understanding of the health profiles and healthcare touchpoints of survivors to inform and optimize strategies for medical system engagement and resource allocation for this public health crisis.

Ionizing radiation (UV, X-ray and ɣ) administered at an appropriate dose to pathogenic organisms can prevent replication while preserving metabolic activity. We have established the GMP process for attenuation by ionizing radiation of the Plasmodium falciparum (Pf) sporozoites (SPZ) in Sanaria® PfSPZ Vaccine, a protective vaccine against malaria. Mosquitoes raised and infected aseptically with Pf were transferred into infected mosquito transport containers (IMTC) and ɣ-irradiated using a 60Co source. PfSPZ were then extracted, purified, vialed, and cryopreserved. To establish the appropriate radiation conditions, the irradiation field inside the IMTCs was mapped using radiochromic film and alanine transfer dosimeters. Dosimeters were irradiated for times calculated to provide 120-170 Gy at the minimum dose location inside the IMTC and regression analysis was used to determine the time required to achieve a lower 95% confidence interval for 150 Gy. A formula incorporating the half-life of 60Co was then used to construct tables of irradiation times for each calendar day. From the mapping studies, formulae were derived to estimate the minimum and maximum doses of irradiation received inside the IMTC from a reference dosimeter mounted on the outside wall. For PfSPZ Vaccine manufacture a dose of 150 Gy was targeted for each irradiation event, a dose known to completely attenuate PfSPZ. The reference dosimeters were processed by the National Institute of Standards and Technology. There have been 587 irradiation events to produce PfSPZ Vaccine during 13 years which generated multiple lots released for pre-clinical studies and clinical trials. The estimated doses at the minimum dose location (mean 154.3 ± 1.77 Gy; range 150.0-159.3 Gy), and maximum dose location (mean 166.3 ± 3.65 Gy, range 155.7 to 175.3 Gy), in IMTCs were normally distributed. Overall dose uniformity was 1.078 ± 0.012. There was no siginifcant change in measured dose over 13 years. As of January 2022, 21 clinical trials of PfSPZ Vaccine have been conducted, with 1,740 volunteers aged 5 months to 61 years receiving 5,648 doses of PfSPZ Vaccine totalling >5.3 billion PfSPZ administered. There have been no breakthrough infections, confirming the consistency and robustness of the radiation attenuation process.

ABSTRACT In response to infections, human immune cells release extracellular vesicles (EVs) that carry a situationally adapted cocktail of proteins and nucleic acids, including microRNAs (miRNAs), to coordinate the immune response. In this study, we identified hsa-miR-21-5p and hsa-miR-24-3p as the most common miRNAs in exosomes released by human monocytes in response to the pathogenic fungus Candida albicans. Functional analysis of miRNAs revealed that hsa-miR-24-3p, but not hsa-miR-21-5p, acted across species and kingdoms, entering C. albicans and inducing fungal cell growth by inhibiting translation of the cyclin-dependent kinase inhibitor Sol1. Packaging of hsa-miR-24-3p into monocyte exosomes required binding of fungal soluble β-glucan to complement receptor 3 (CR3) and binding of mannan to Toll-like receptor 4 (TLR4), resulting in receptor colocalization. Together, our in vitro and in vivo findings reveal a novel cross-species evasion mechanism by which C. albicans exploits a human miRNA to promote fungal growth and survival in the host. IMPORTANCE Over the last decade, communication between immune cells by extracellular vesicle-associated miRNAs has emerged as an important regulator of the coordinated immune response. Therefore, a thorough understanding of the conversation occurring via miRNAs, especially during infection, may provide novel insights into both the host reaction to the microbe as well as the microbial response. This study provides evidence that the pathogenic fungus C. albicans communicates with human monocytes and induces the release of a human miRNA that promotes fungal growth. This mechanism represents an unexpected cross-species interaction and implies that an inhibition of specific miRNAs offers new possibilities for the treatment of human fungal infections.

ABSTRACT Microbial community diversity is often correlated with physical environmental stresses like acidity, salinity, and temperature. For example, species diversity usually declines with increasing temperature above 20°C. However, few studies have examined whether the genetic functional diversity of community metagenomes varies in a similar way as species diversity along stress gradients. Here, we investigated bacterial communities in thermal spring sediments ranging from 21 to 88°C, representing communities of 330 to 3,800 bacterial and archaeal species based on 16S rRNA gene amplicon analysis. Metagenomes were sequenced, and Pfam abundances were used as a proxy for metagenomic functional diversity. Significant decreases in both species diversity and Pfam diversity were observed with increasing temperatures. The relationship between Pfam diversity and species diversity followed a power function with the steepest slopes in the high-temperature, low-diversity region of the gradient. Species additions to simple thermophilic communities added many new Pfams, while species additions to complex mesophilic communities added relatively fewer new Pfams, indicating that species diversity does not approach saturation as rapidly as Pfam diversity does. Many Pfams appeared to have distinct temperature ceilings of 60 to 80°C. This study suggests that temperature stress limits both taxonomic and functional diversity of microbial communities, but in a quantitatively different manner. Lower functional diversity at higher temperatures is probably due to two factors, including (i) the absence of many enzymes not adapted to thermophilic conditions, and (ii) the fact that high-temperature communities are comprised of fewer species with smaller average genomes and, therefore, contain fewer rare functions. IMPORTANCE Only recently have microbial ecologists begun to assess quantitatively how microbial species diversity correlates with environmental factors like pH, temperature, and salinity. However, still, very few studies have examined how the number of distinct biochemical functions of microbial communities, termed functional diversity, varies with the same environmental factors. Our study examined 18 microbial communities sampled across a wide temperature gradient and found that increasing temperature reduced both species and functional diversity, but in different ways. Initially, functional diversity increased sharply with increasing species diversity but eventually plateaued, following a power function. This pattern has been previously predicted in theoretical models, but our study validates this predicted power function with field metagenomic data. This study also presents a unique overview of the distribution of metabolic functions along a temperature gradient, demonstrating that many functions have temperature “ceilings” above which they are no longer found.

Human natural killer (NK) cells can target tumor cells in an antigen-specific manner by the recognition of cell bound antibodies. This process induces antibody-dependent cell-mediated cytotoxicity (ADCC) and is exclusively mediated by the low affinity IgG Fc receptor CD16A (FcγRIIIA). Exploiting ADCC by NK cells is a major area of emphasis for advancing cancer immunotherapies. CD64 (FcγRI) is the only high affinity IgG FcR and it binds to the same IgG isotypes as CD16A, but it is not expressed by human NK cells. We have generated engineered human NK cells expressing recombinant CD64 with the goal of increasing their ADCC potency. Preclinical testing of this approach is essential for establishing efficacy and safety of the engineered NK cells. The dog provides particular advantages as a model, which includes spontaneous development of cancer in the setting of an intact and outbred immune system. To advance this immunotherapy model, we cloned canine CD16A and CD64 and generated specific mAbs. We report here for the first time the expression patterns of these FcγRs on dog peripheral blood leukocytes. CD64 was expressed by neutrophils and monocytes, but not lymphocytes, while canine CD16A was expressed at high levels by a subset of monocytes and lymphocytes. These expression patterns are similar to that of human leukocytes. Based on phenotypic characteristics, the CD16A+ lymphocytes consisted of T cells (CD3+ CD8+ CD5dim α/β TCR+) and NK cells (CD3− CD5− CD94+), but not B cells. Interestingly, the majority of canine CD16A+ lymphocytes were from the T cell population. Like human CD16A, canine CD16A was downregulated by a disintegrin and metalloproteinase 17 (ADAM17) upon leukocyte activation, revealing a conserved means of regulation. We also directly demonstrate that both canine CD16A and CD64 can induce ADCC when expressed in the NK cell line NK-92. These findings pave the way to engineering canine NK cells or T cells with high affinity recombinant canine CD64 to maximize ADCC and to test their safety and efficacy to benefit both humans and dogs.

Abstract Bioenergy production often focuses on the aboveground feedstock production for conversion to fuel and other materials. However, the belowground component is crucial for soil carbon sequestration, greenhouse gas fluxes, and ecosystem function. Roots maximize feedstock production on marginal lands by acquiring soil resources and mediating soil ecosystem processes through interactions with the microbial community. This belowground world is challenging to observe and quantify; however, there are unprecedented opportunities using current methodologies to bring roots, microbes, and soil into focus. These opportunities allow not only breeding for increased feedstock production but breeding for increased soil health and carbon sequestration as well. A recent workshop hosted by the USDOE Bioenergy Research Centers highlighted these challenges and opportunities while creating a roadmap for increased collaboration and data interoperability through standardization of methodologies and data using F.A.I.R. principles. This article provides a background on the need for belowground research in bioenergy cropping systems, a primer on root system properties of major U.S. bioenergy crops, and an overview of the roles of root chemistry, exudation, and microbial interactions on sustainability. Crucially, we outline how to adopt standardized measures and databases to meet the most pressing methodological needs to accelerate root, soil, and microbial research to meet the pressing societal challenges of the century.

Background Human immunodeficiency virus (HIV), hepatitis C virus (HCV), and injection drug use are syndemic in the central Appalachian states. In Tennessee (TN), declines in HIV among persons who inject drugs (PWID) stalled, and HCV infection rates increased significantly from 2013–2017. To better target strategies to address the syndemic, county-level socioeconomic, opioid use, access to healthcare, and health factors were modeled to identify indicators predictive of vulnerability to an HIV/HCV outbreak among PWID in TN. Methods Newly reported chronic HCV cases among persons aged 13–39 years in 2016–2017 were used as a proxy for county-level HIV/HCV vulnerability among TN’s 95 counties. Seventy-five publicly available county-level measures from 2016–2017 were collected and reduced through multiple dimension reduction techniques. Negative binomial regression identified indicators associated with HCV which were used to calculate county-level vulnerability to a local HIV/HCV outbreak. Results Thirteen county-level indicators were identified as strongly predictive of HIV/HCV vulnerability with the statistically significant indicators being percentage of the population aged 20–44 years, per capita income, teen birth rate, percentage of clients in TDMHSAS-funded opioid treatment and recovery, syphilis case rate, and percentage of homes with at least one vehicle. Based on the 13 indicators, we identified the distribution of vulnerability to an HIV/HCV outbreak among TN’s counties. Eleven high vulnerability counties were identified, with the preponderance located in east and middle TN. Conclusion This analysis identified the county-level factors most associated with vulnerability to an HIV/HCV outbreak among PWID in TN. These results, alongside routine surveillance, will guide targeted prevention and linkage to care efforts for the most vulnerable communities.

Background. Convalescent plasma obtained from individuals who have recovered from severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) contains neutralizing antibodies to the virus and has been frequently used as a treatment in hospitalized patients with severe COVID-19. Methods. We conducted a retrospective, observational cohort study involving 96 hospitalized patients with severe COVID-19 who were allocated in a 1 : 1 ratio to having received either high antibody concentration convalescent plasma or low antibody concentration convalescent plasma. Quantitative measurements of IgG to the receptor-binding domain (RBD), the S1 subunit of the spike protein, and the SARS-CoV-2 nucleocapsid (N) protein were determined from donor plasma samples. The primary outcome was all-cause mortality within 30 days following convalescent plasma administration in regard to each of the three antibody domains. Results. Within the nucleocapsid antibody domain, death occurred in 22.2% of patients in the low antibody concentration group versus 23.5% in the high antibody concentration group (p=0.88). Within the RBD antibody domain, death occurred in 22.9% of patients in both the low and the high antibody concentration groups (p=1.0). Within the S1 subunit antibody domain, death occurred in 27.1% of patients in the low antibody concentration group versus 18.8% in the high antibody concentration group (p=0.33). Conclusions. No significant differences were observed between low and high concentration convalescent plasma in regard to overall mortality at 30 days, hospital length of stay, number of ventilator days, and subsequent receipt of invasive mechanical ventilation in patients who were previously not receiving mechanical ventilation. Trial Registration. This study was not associated with a clinical trial due to the retrospective nature of study design.

In this study, we investigated how word- and text-level processes contribute to different types of reading fluency measures. We aimed to increase our understanding of the underlying processes necessary for fluent reading. The sample included 73 Dutch Grade 3 children, who were assessed on serial word reading rate (familiar words), word-list reading fluency (increasingly difficult words), and sentence reading fluency. Word-level processes were individual word recognition speed (discrete word reading) and sequential processing efficiency (serial digit naming). Text-level processes were receptive vocabulary and syntactic skills. The results showed that word- and text-level processes combined accounted for a comparable amount of variance in all fluency outcomes. Both word-level processes were moderate predictors of all fluency outcomes. However, vocabulary only moderately predicted sentence reading fluency, and syntactic skills merely contributed to sentence reading fluency indirectly through vocabulary. The findings indicate that sequential processing efficiency has a crucial role in reading fluency across various measures besides individual word recognition speed. Additionally, text-level processes come into play when complexity and context availability of fluency measures increases, but the exact timing requires further study. Findings are discussed in terms of future directions and their possible value for diagnostic assessment and intervention of reading difficulties.

GAL08 are bacteria belonging to an uncultivated phylogenetic cluster within the phylum Acidobacteria. We detected a natural population of the GAL08 clade in sediment from a pH-neutral hot spring located in British Columbia, Canada. To shed light on the abundance and genomic potential of this clade, we collected and analyzed hot spring sediment samples over a temperature range of 24.2–79.8°C. Illumina sequencing of 16S rRNA gene amplicons and qPCR using a primer set developed specifically to detect the GAL08 16S rRNA gene revealed that absolute and relative abundances of GAL08 peaked at 65°C along three temperature gradients. Analysis of sediment collected over multiple years and locations revealed that the GAL08 group was consistently a dominant clade, comprising up to 29.2% of the microbial community based on relative read abundance and up to 4.7 × 105 16S rRNA gene copy numbers per gram of sediment based on qPCR. Using a medium quality threshold, 25 single amplified genomes (SAGs) representing these bacteria were generated from samples taken at 65 and 77°C, and seven metagenome-assembled genomes (MAGs) were reconstructed from samples collected at 45–77°C. Based on average nucleotide identity (ANI), these SAGs and MAGs represented three separate species, with an estimated average genome size of 3.17 Mb and GC content of 62.8%. Phylogenetic trees constructed from 16S rRNA gene sequences and a set of 56 concatenated phylogenetic marker genes both placed the three GAL08 bacteria as a distinct subgroup of the phylum Acidobacteria, representing a candidate order (Ca. Frugalibacteriales) within the class Blastocatellia. Metabolic reconstructions from genome data predicted a heterotrophic metabolism, with potential capability for aerobic respiration, as well as incomplete denitrification and fermentation. In laboratory cultivation efforts, GAL08 counts based on qPCR declined rapidly under atmospheric levels of oxygen but increased slightly at 1% (v/v) O2, suggesting a microaerophilic lifestyle.

ABSTRACT: OBJECTIVE: This phase I trial (NCT01912625) evaluated the safety and pharmacokinetics of definitive concurrent chemoradiotherapy (cCRT) and the radiosensitizer trametinib (MEK1/2 inhibitor) for KRAS-mutated nonmetastatic non-small cell lung cancer (NSCLC). METHODS: Patients received cCRT (carboplatin/paclitaxel and 60 Gy/30 fractions radiotherapy); oral trametinib (7 days/week) commenced on day 1 and completed on the final day of radiotherapy. Dose-finding of trametinib was done using the time-to-event continual reassessment method (TiTE-CRM); dose levels were 0.5mg (level -1), 1mg (initial, level 1), 1.5mg (level 2), and 2mg (level 3). Progression-free (PFS) and overall survival (OS) times were also recorded. RESULTS: Fifteen patients (stage III, variety of KRAS mutations) were treated, with 1/5/4/5 at dose levels -1/1/2/3, respectively. Five patients received dose reductions (n=2, levels 2 and 3; n=1, level 1). Twelve patients completed the full cCRT course. One patient (following 12d trametinib) was taken off protocol for an unrelated/unresolved grade 1 event and later experienced grade 5 sepsis/respiratory failure. There was one grade 4 retinal detachment; grade 3 events included skin rash (n=2) and ventricular dysfunction, pneumonitis, pain, fatigue, and diarrhea (n=1 each). The final dose selected by the TiTE-CRM of trametinib was 1.5 mg. Pharmacokinetic profiles were elucidated and extensively described. At median follow-up of 70 months, median PFS was 11 months and median OS was 38 months. CONCLUSIONS: The MTD for trametinib when combined with cCRT is 1.5 mg, with encouraging preliminary outcomes. This combination merits further study to combine with consolidation durvalumab in non-metastatic KRAS mutant NSCLC.

Objective We assessed trends and identified individual- and county-level factors associated with individual linkage to HIV care in Tennessee (TN). Methods TN residents diagnosed with HIV from 2012–2016 were included in the analysis (n = 3,751). Individuals were assigned county-level factors based on county of residence at the time of diagnosis. Linkage was defined by the first CD4 or HIV RNA test date after HIV diagnosis. We used modified Poisson regression to estimate probability of 30-day linkage to care at the individual-level and the contribution of individual and county-level factors to this outcome. Results Both MSM (aRR 1.23, 95%CI 0.98–1.55) and women who reported heterosexual sex risk factors (aRR 1.39, 95%CI 1.18–1.65) were more likely to link to care within 30-days than heterosexual males. Non-Hispanic Black individuals had poorer linkage than White individuals (aRR 0.77, 95%CI 0.71–0.83). County-level mentally unhealthy days were negatively associated with linkage (aRR 0.63, 95%CI: 0.40–0.99). Conclusions Racial disparities in linkage to care persist at both individual and county levels, even when adjusting for county-level social determinants of health. These findings suggest a need for structural interventions to address both structural racism and mental health needs to improve linkage to care and minimize racial disparities in HIV outcomes.

Objective. Medicolegal examination of an intervention as common as endotracheal intubation may be valuable to physicians in many specialties. Our objectives were to comprehensively detail the factors raised in litigation to better educate physicians on strategies for minimizing liability and augmenting patient safety. Methods. Publicly available court records were searched for pertinent litigation. Ultimately, 214 jury verdict and settlement reports were examined for various factors, including outcome, award, geographic location, defendant specialty, setting in which an injury occurred, patient demographics, and other causes of malpractice. Results. Ninety-two cases (43.0%) were resolved in the defendant’s favor, with the remaining cases resulting in out-of-court settlement or a plaintiff’s verdict. Payments from these cases were considerable, averaging $2.5 M. The most frequent physician defendants were anesthesiologists (59.8%) and emergency-physicians (19.2%), although other specialties were well represented. The most common setting of injury was the operating room (45.3%). Common factors included sustaining permanent deficits (89.2%), death (50.5%), and anoxic brain injury (37.4%). Injuries occurring in labor and delivery mostly involved newborns and had among the highest awards. Conclusions. Litigation involves injuries sustained in numerous settings. The most common factors present included sustaining permanent deficits, including anoxic brain injury. The presence of this latter injury increased the likelihood of a case being resolved with payment. Finally, deficits in informed consent were noted in numerous cases, stressing the importance of a clear process in which the physician explains specific risks (such as those detailed in this analysis), benefits, and alternatives.

The jarosite family of minerals are part of the alunite supergroup with the general formula AB3(TO4)2(OH)6. Jarosite family minerals are known to incorporate silver (Ag), but the extent to which this occurs, and at what temperature range, is not well constrained. To address this knowledge gap, jarosite compounds with the A site filled with K, Na, Ag and H3O were synthesised at 22 °C, 97 °C and 140 °C to simulate low-, moderate- and high-temperature environments, respectively. The compounds were characterised by XRD, SEM, chemical analysis and Raman spectroscopy. All of the synthesised compounds took up Ag. In general, higher temperatures of synthesis increased alkali and Ag occupancy of the A site of the products. Silver contents increased with the increasing concentration of Ag in the starting solutions at all temperatures. The order of preference for occupancy of the A site in the synthesised solids is K > Na > H3O > Ag at all temperatures, which is consistent with the reported order of ΔGf of −3309 kJ/mol, −3270 kJ/mol, −3247 kJ/mol and −2948 kJ/mol for jarosite, natrojarosite, hydroniumjarosite and argentojarosite, respectively. The results of this study show that Ag can be incorporated in jarosite and natrojarosite at low-to-high temperatures, and therefore, jarosite family minerals can be important stores of Ag in in natural and engineered environments.

The efficiency definition allows us to compare two machines with each other. In general, the efficiency is defined as the ratio of usable power to the required power. This raises the question: what is the usable power? Most engineers discuss efficiency on the basis of the energy balance, i.e., the first law of thermodynamics. In this paper, we derive the exegetic efficiency taking the second law of thermodynamics into account. Exergy analysis takes into account work and heat and is able to model reality very accurately. On this basis, a comparison between the isentropic and exergetic efficiencies is given. A high-pressure radial fan is used as an example, and the differences are discussed. Therefore, measurements of a non-adiabatic fan are evaluated, and the role of the heat flux in the environment is discussed. The investigations show that a relevant difference between the isentropic and exergetic efficiencies becomes apparent in the partial-load range with high-pressure build-up. The thermal energy contained in the flow belongs proportionally to the exergy, i.e., the working capacity of the gas relative to its environment. For a standard such as ISO 5801 “Fans—Performance testing using standardized airways”, the efficiency must not only be physically correct, it must also be simple and practical. Against this background, the outlook of this paper discusses when and which efficiency definition is appropriate and best suited for a standard.

The growing possibilities of non-invasive heart rate and blood pressure measurement with mobile devices allow vital data to be continuously collected and used to assess patients’ health status. When it comes to the risk assessment of abdominal aortic aneurysms (AAA), the continuous tracking of blood pressure and heart rate could enable a more patient-specific approach. The use of a load function and an energy function, with continuous blood pressure, heart rate, and aneurysm stiffness as input parameters, can quantify dynamic load on AAA. We hypothesise that these load functions correlate with aneurysm growth and outline a possible study procedure in which the hypothesis could be tested for validity. Subsequently, uncertainty quantification of input quantities and derived quantities is performed.

Elements of fractal analysis are widely used in scientific research, including several biological disciplines. In this study, we hypothesized that chicken breed biodiversity manifests not only at the phenotypic level, but also at the genetic-system level in terms of different profiles of fractal conformity and bioconsolidation in the early myogenesis gene expression. To demonstrate this effect, we developed two mathematical models that describe the fractal nature of the expression of seven key genes in the embryonic breast and thigh muscles in eight breeds of meat, dual purpose, egg and game types. In the first model, we produced breed-specific coefficients of gene expression conformity in each muscle type using the slopes of regression dependencies, as well as an integral myogenesis gene expression index (MGEI). Additionally, breed fractal dimensions and integral myogenesis gene expression fractal dimension index (MGEFDI) were determined. The second gene expression model was based on plotting fractal portraits and calculating indices of fractal bioconsolidation. The bioconsolidation index of myogenesis gene expression correlated with the chick growth rate and nitric oxide (NO) oxidation rate. The proposed fractal models were instrumental in interpreting the genetic diversity of chickens at the level of gene expression for early myogenesis, NO metabolism and the postnatal growth of chicks.

The ZrO B ^1 Π– X ^1 Σ ^+ , B ^1 Π– A ^1 Δ, and C ^1 Σ ^+ – X ^1 Σ ^+ band systems are important opacity sources in the near-infrared and optical spectra of S-type stars. A total of 21 rovibronic bands with v ″ ≤ 7 and $v^{\prime} \leqslant 5$ were observed and fit for the B ^1 Π– X ^1 Σ ^+ transition, five bands for the ^90 ZrO B ^1 Π– A ^1 Δ transition and one band for the ^90 ZrO C ^1 Σ ^+ – X ^1 Σ ^+ transition. All band systems were analyzed using high-temperature, high-resolution emission spectra collected at the National Solar Observatory (Kitt Peak). A modern spectroscopic analysis was performed using the PGOPHER program to provide updated spectroscopic constants. In general, we improve the accuracy of the line positions reported in the literature and slightly extend the vibrational analysis. Equilibrium molecular constants were then derived and combined with new ab initio calculations of transition dipole moment functions to produce line lists with line strengths.

We report a detailed analysis of a failed eruption and flare in active region 12018 on 2014 April 3 using multiwavelength observations from the Solar Dynamics Observatory/Atmospheric Imaging Assembly, IRIS, STEREO, and Hinode/Solar Optical Telescope. At least four jets were observed to emanate from the cusp of this small active region (large bright point) with a null-point topology during the 2 hr prior to the slow rise of a filament. During the filament slow rise multiple plasma blobs were seen, most likely formed in a null-point current sheet near the cusp. The subsequent filament eruption, which was outside the IRIS field of view, was accompanied by a flare but remained confined. During the explosive flare reconnection phase, additional blobs appeared repetitively and moved bidirectionally within the flaring region below the erupting filament. The filament kinked, rotated, and underwent leg–leg reconnection as it rose, yet it failed to produce a coronal mass ejection. Tiny jet-like features in the fan loops were detected during the filament slow rise/preflare phase. We interpret them as signatures of reconnection between the ambient magnetic field and the plasmoids leaving the null-point sheet and streaming along the fan loops. We contrast our interpretation of these tiny jets, which occur within the large-scale context of a failed filament eruption, with the local nanoflare-heating scenario proposed by Antolin et al.

Ornithine aminotransferase (OAT) is overexpressed in hepatocellular carcinoma (HCC), and we previously showed that inactivation of OAT inhibits the growth of HCC. Recently, we found that (3S,4S)-3-amino-4-fluorocyclopentenecarboxylic acid (5) was a potent inactivator of γ-aminobutyric acid aminotransferase (GABA-AT), proceeding by an enamine mechanism. Here we describe our investigations into the activity and mechanism of 5 as an inactivator of human OAT. We have found that 5 exhibits 10-fold less inactivation efficiency (kinact/KI) against hOAT than GABA-AT. A comprehensive mechanistic study was carried out to understand its inactivation mechanism with hOAT. pKa and electrostatic potential calculations were performed to further support the notion that the α,β-unsaturated alkene of 5 is critical for enhancing acidity and nucleophilicity of the corresponding intermediates and ultimately responsible for the improved inactivation efficiency of 5 over the corresponding saturated analogue (4). Intact protein mass spectrometry and the crystal structure complex with hOAT provide evidence to conclude that 5 mainly inactivates hOAT through noncovalent interactions, and that, unlike with GABA-AT, covalent binding with hOAT is a minor component of the total inhibition which is unique relative to other monofluoro-substituted derivatives. Furthermore, based on the results of transient-state measurements and free energy calculations, it is suggested that the α,β-unsaturated carboxylate group of PLP-bound 5 may be directly involved in the inactivation cascade by forming an enolate intermediate. Overall, compound 5 exhibits unusual structural conversions which are catalyzed by specific residues within hOAT, ultimately leading to an enamine mechanism-based inactivation of hOAT through noncovalent interactions and covalent modification.

Data were collected on patients admitted to the Queen Elizabeth Hospital Birmingham with type-1 myocardial infarction during 2016 and 2017 inclusively, who were treated by percutaneous intervention and had pre-discharge transthoracic echocardiography. The data were obtained from prospectively maintained hospital databases and records. Echocardiography was performed and reported contemporaneously by accredited echocardiographers. The purpose was to understand the prevalence and characteristics of mitral regurgitation (MR) after acute MI, including patients with ST-elevation (STEMI) and non-ST elevation MI (NSTEMI). MR was observed in 294/1000 patients with the following relative severities: mild = 76%, moderate = 21%, severe = 3% [1]. MR was graded by multiparametric quantification including proximal isolvelocity surface area (PISA), vena contracta (VC), effective regurgitant orifice area (EROA) and regurgitant volume (RVol). Amongst all patients with MR (n=294), PISA was performed in 89/294 (30%), VC 75/294 (26%), EROA in 53/294 (18%) and RVol in 26/294 (9%). Amongst patients with moderate or severe MR (n=70), PISA was performed in 57/70 (81%), VC in 55/70 (79%), EROA in 46/70 (66%) and RVol in 25/70 (36%). Characteristics of MR following acute MI were also assessed including frequency of reported leaflet thickness (259/294 = 88%) and mitral annular calcification (102/294 = 35%). Furthermore, the effect of MI on pre-existing MR was investigated and patients with pre-existing MR who continue to have MR after acute MI were found to have progression of MR by one grade in approximately 25% of cases. Finally, using Cox proportional hazards univariate analysis, significant factors associated with mortality in patients with MR post-MI include age (HR 1.065; 95% CI 1.035-1.096; p

Objective:. To investigate the factors underlying willingness to participate in a hypothetical trial among patients and surgeons, to inform the design of future placebo surgery controlled trials. Background:. Placebo surgery controlled trials are the gold standard for testing the efficacy of surgical procedures. However, these trials commonly fail to meet the target sample size and terminate underpowered. Methods:. From October 2019 to July 2020, eligible patients were identified from the orthopedic waiting list at a single tertiary hospital and surgeons were identified from orthopedic clinics at three tertiary hospitals in Australia. Qualitative interviews explored factors underlying willingness to participate in a hypothetical trial, including understanding of trial concepts; attitudes; and trial design preferences. Data collection and analysis were conducted in parallel. Recruitment ceased when no new concepts emerged. Interview data were analyzed using reflexive thematic analysis. Results:. The majority of surgeons and only a few patients indicated a willingness to participate in a placebo surgery controlled trial. Factors underlying willingness were captured in four themes: (1) Understanding and attitudes toward placebo; (2) Attitudes towards randomization/perception of equipoise; (3) Perception of risk; and (4) Ethical concerns. Conclusions:. To optimize recruitment in the future, trialists may consider embedding strategies into the recruitment process that validate patients’ symptoms, encourage an altruistic mindset, address surgeon biases, and involve surgeons in explaining trial concepts to patients. Trialists may also consider designing three arm trials that meet surgeons’ preferences for a “low” and “high” fidelity placebo.

PfSPZ Vaccine against malaria is composed of Plasmodium falciparum (Pf) sporozoites (SPZ) manufactured using aseptically reared Anopheles stephensi mosquitoes. Immune response genes of Anopheles mosquitoes such as Leucin-Rich protein (LRIM1), inhibit Plasmodium SPZ development (sporogony) in mosquitoes by supporting melanization and phagocytosis of ookinetes. With the aim of increasing PfSPZ infection intensities, we generated an A. stephensi LRIM1 knockout line, Δaslrim1, by embryonic genome editing using CRISPR-Cas9. Δaslrim1 mosquitoes had a significantly increased midgut bacterial load and an altered microbiome composition, including elimination of commensal acetic acid bacteria. The alterations in the microbiome caused increased mosquito mortality and unexpectedly, significantly reduced sporogony. The survival rate of Δaslrim1 mosquitoes and their ability to support PfSPZ development, were partially restored by antibiotic treatment of the mosquitoes, and fully restored to baseline when Δaslrim1 mosquitoes were produced aseptically. Deletion of LRIM1 also affected reproductive capacity: oviposition, fecundity and male fertility were significantly compromised. Attenuation in fecundity was not associated with the altered microbiome. This work demonstrates that LRIM1's regulation of the microbiome has a major impact on vector competence and longevity of A. stephensi. Additionally, LRIM1 deletion identified an unexpected role for this gene in fecundity and reduction of sperm transfer by males.

A key benefit of long-read nanopore sequencing technology is the ability to detect modified DNA bases, such as 5-methylcytosine. The lack of R/Bioconductor tools for the effective visualization of nanopore methylation profiles between samples from different experimental groups led us to develop the NanoMethViz R package. Our software can handle methylation output generated from a range of different methylation callers and manages large datasets using a compressed data format. To fully explore the methylation patterns in a dataset, NanoMethViz allows plotting of data at various resolutions. At the sample-level, we use dimensionality reduction to look at the relationships between methylation profiles in an unsupervised way. We visualize methylation profiles of classes of features such as genes or CpG islands by scaling them to relative positions and aggregating their profiles. At the finest resolution, we visualize methylation patterns across individual reads along the genome using the spaghetti plot and heatmaps, allowing users to explore particular genes or genomic regions of interest. In summary, our software makes the handling of methylation signal more convenient, expands upon the visualization options for nanopore data and works seamlessly with existing methylation analysis tools available in the Bioconductor project. Our software is available at https://bioconductor.org/packages/NanoMethViz. Author summary Recently developed nanopore sequencing technology enables DNA methylation measurement on long DNA molecules. This technology provides a new tool for investigating DNA methylation, a form of DNA modification that plays an essential role in early development, and is linked to some forms of cancer through adulthood. There is a lack of R/Bioconductor software for effective visualization of methylation calls based on nanopore platforms, which hinders the analysis and presentation of results. We developed NanoMethViz, the first R package to create visualizations for nanopore methylation data at various summary resolutions. NanoMethViz produces publication-quality plots to inspect the broad differences in methylation profiles of different samples, the aggregated methylation profiles of classes of genomic features, and the methylation profiles of individual long reads. Our software provides an efficient data format for storing methylation information and converts data from popular methylation calling software to formats recognized by statistical methods available in the Bioconductor toolkit for further analysis. NanoMethViz allows researchers to more quickly and effectively analyze their data and produce high-quality figures to present their results.

In recent years, there has been an exponential growth in sequencing projects due to accelerated technological advances, leading to a significant increase in the amount of data and resulting in new challenges for biological sequence analysis. Consequently, the use of techniques capable of analyzing large amounts of data has been explored, such as machine learning (ML) algorithms. ML algorithms are being used to analyze and classify biological sequences, despite the intrinsic difficulty in extracting and finding representative biological sequence methods suitable for them. Thereby, extracting numerical features to represent sequences makes it statistically feasible to use universal concepts from Information Theory, such as Tsallis and Shannon entropy. In this study, we propose a novel Tsallis entropy-based feature extractor to provide useful information to classify biological sequences. To assess its relevance, we prepared five case studies: (1) an analysis of the entropic index q; (2) performance testing of the best entropic indices on new datasets; (3) a comparison made with Shannon entropy and (4) generalized entropies; (5) an investigation of the Tsallis entropy in the context of dimensionality reduction. As a result, our proposal proved to be effective, being superior to Shannon entropy and robust in terms of generalization, and also potentially representative for collecting information in fewer dimensions compared with methods such as Singular Value Decomposition and Uniform Manifold Approximation and Projection.

BACKGROUND Naturally acquired immunity to malaria is incompletely understood. We used controlled human malaria infection (CHMI) to study the impact of past exposure on malaria in Kenyan adults in relation to infection with a non-Kenyan parasite strain.METHODS We administered 3.2 × 103 aseptic, purified, cryopreserved Plasmodium falciparum sporozoites (Sanaria PfSPZ Challenge, NF54 West African strain) by direct venous inoculation and undertook clinical monitoring and serial quantitative PCR (qPCR) of the 18S ribosomal RNA gene. The study endpoint was met when parasitemia reached 500 or more parasites per μL blood, clinically important symptoms were seen, or at 21 days after inoculation. All volunteers received antimalarial drug treatment upon meeting the endpoint.RESULTS One hundred and sixty-one volunteers underwent CHMI between August 4, 2016, and February 14, 2018. CHMI was well tolerated, with no severe or serious adverse events. Nineteen volunteers (11.8%) were excluded from the analysis based on detection of antimalarial drugs above the minimal inhibitory concentration or parasites genotyped as non-NF54. Of the 142 volunteers who were eligible for analysis, 26 (18.3%) had febrile symptoms and were treated; 30 (21.1%) reached 500 or more parasites per μL and were treated; 53 (37.3%) had parasitemia without meeting thresholds for treatment; and 33 (23.2%) remained qPCR negative.CONCLUSION We found that past exposure to malaria, as evidenced by location of residence, in some Kenyan adults can completely suppress in vivo growth of a parasite strain originating from outside Kenya.TRIAL REGISTRATION ClinicalTrials.gov NCT02739763.FUNDING Wellcome Trust.

People with schizophrenia (SZ) process emotions less accurately than do healthy comparators (HC), and emotion recognition have expanded beyond accuracy to performance variables like reaction time (RT) and confidence. These domains are typically evaluated independently, but complex inter-relationships can be evaluated through machine learning at an item-by-item level. Using a mix of ranking and machine learning tools, we investigated item-by-item discrimination of facial affect with two emotion recognition tests (BLERT and ER-40) between SZ and HC. The best performing multi-domain model for ER40 had a large effect size in differentiating SZ and HC (d = 1.24) compared to a standard comparison of accuracy alone (d = 0.48); smaller increments in effect sizes were evident for the BLERT (d = 0.87 vs. d = 0.58). Almost half of the selected items were confidence ratings. Within SZ, machine learning models with ER40 (generally accuracy and reaction time) items predicted severity of depression and overconfidence in social cognitive ability, but not psychotic symptoms. Pending independent replication, the results support machine learning, and the inclusion of confidence ratings, in characterizing the social cognitive deficits in SZ. This moderate-sized study (n = 372) included subjects with schizophrenia (SZ, n = 218) and healthy controls (HC, n = 154).

Background ApoAI (apolipoproteins AI) and apoAII (apolipoprotein AII) are structural and functional proteins of high‐density lipoproteins (HDL) which undergo post‐translational modifications at specific residues, creating distinct proteoforms. While specific post‐translational modifications have been reported to alter apolipoprotein function, the full spectrum of apoAI and apoAII proteoforms and their associations with cardiometabolic phenotype remains unknown. Herein, we comprehensively characterize apoAI and apoAII proteoforms detectable in serum and their post‐translational modifications and quantify their associations with cardiometabolic health indices. Methods and Results Using top‐down proteomics (mass‐spectrometric analysis of intact proteins), we analyzed paired serum samples from 150 CARDIA (Coronary Artery Risk Development in Young Adults) study participants from year 20 and 25 exams. Measuring 15 apoAI and 9 apoAII proteoforms, 6 of which carried novel post‐translational modifications, we quantified associations between percent proteoform abundance and key cardiometabolic indices. Canonical (unmodified) apoAI had inverse associations with HDL cholesterol and HDL‐cholesterol efflux, and positive associations with obesity indices (body mass index, waist circumference), and triglycerides, whereas glycated apoAI showed positive associations with serum glucose and diabetes mellitus. Fatty‐acid‒modified ApoAI proteoforms had positive associations with HDL cholesterol and efflux, and inverse associations with obesity indices and triglycerides. Truncated and dimerized proteoforms of apoAII were associated with HDL cholesterol (positively) and obesity indices (inversely). Several proteoforms had no significant associations with phenotype. Conclusions Associations between apoAI and AII and cardiometabolic indices are proteoform‐specific. These results provide “proof‐of‐concept” that precise chemical characterization of human apolipoproteins will yield improved insights into the complex pathways through which proteins signify and mediate health and disease.

Objectives The COVID-19 pandemic is unprecedented as a global crisis over the last century. How do specialist surgeons make decisions about patient care in these unprecedent times?Design Between April and May 2020, we conducted an international qualitative study. Sarcoma surgeons from diverse global settings participated in 60 min interviews exploring surgical decision making during COVID-19. Interview data were analysed using an inductive thematic analysis approach.Setting Participants represented public and private hospitals in 14 countries, in different phases of the first wave of the pandemic: Australia, Argentina, Canada, India, Italy, Japan, Nigeria, Singapore, Spain, South Africa, Switzerland, Turkey, UK and USA.Participants From 22 invited sarcoma surgeons, 18 surgeons participated. Participants had an average of 19 years experience as a sarcoma surgeon.Results 17/18 participants described a decision they had made about patient care since the start of the pandemic that was unique to them, that is, without precedence. Common to ‘unique’ decisions about patient care was uncertainty about what was going on and what would happen in the future (theme 1: the context of uncertainty), the impact of the pandemic on resources or threat of the pandemic to overwhelm resources (theme 2: limited resources), perceived increased risk to self (theme 3: duty of care) and least-worst decision making, in which none of the options were perceived as ideal and participants settled on the least-worst option at that point in time (theme 4: least-worst decision making).Conclusions In the context of rapidly changing standards of justice and beneficence in patient care, traditional decision-making frameworks may no longer apply. Based on the experiences of surgeons in this study, we describe a framework of least-worst decision making. This framework gives rise to actionable strategies that can support decision making in sarcoma and other specialised fields of surgery, both during the current crisis and beyond.