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Recent genome-wide studies of rare genetic variants have begun to implicate novel mechanisms for tetralogy of Fallot (TOF), a severe congenital heart defect (CHD). To provide statistical support for case-only data without parental genomes, we re-analyzed genome sequences of 231 individuals with TOF (n = 175) or related CHD. We adapted a burden test originally developed for de novo variants to assess ultra-rare variant burden in individual genes, and in gene-sets corresponding to functional pathways and mouse phenotypes, accounting for highly correlated gene-sets and for multiple testing. For truncating variants, the gene burden test confirmed significant burden in FLT4 (Bonferroni corrected p-value < 0.01). For missense variants, burden in NOTCH1 achieved genome-wide significance only when restricted to constrained genes (i.e., under negative selection, Bonferroni corrected p-value = 0.004), and showed enrichment for variants affecting the extracellular domain, especially those disrupting cysteine residues forming disulfide bonds (OR = 39.8 vs. gnomAD). Individuals with NOTCH1 ultra-rare missense variants, all with TOF, were enriched for positive family history of CHD. Other genes not previously implicated in CHD had more modest statistical support in gene burden tests. Gene-set burden tests for truncating variants identified a cluster of pathways corresponding to VEGF signaling (FDR = 0%), and of mouse phenotypes corresponding to abnormal vasculature (FDR = 0.8%); these suggested additional candidate genes not previously identified (e.g., WNT5A and ZFAND5). Results for the most promising genes were driven by the TOF subset of the cohort. The findings support the importance of ultra-rare variants disrupting genes involved in VEGF and NOTCH signaling in the genetic architecture of TOF, accounting for 11–14% of individuals in the TOF cohort. These proof-of-principle data indicate that this statistical methodology could assist in analyzing case-only sequencing data in which ultra-rare variants, whether de novo or inherited, contribute to the genetic etiopathogenesis of a complex disorder.

[This corrects the article DOI: 10.1371/journal.pbio.3000193.].

Vascular bypass graft infection with Mycobacterium bovis following Bacillus Calmette–Guérin (BCG) immunotherapy for bladder cancer is an incredibly rare complication. We present the case of an 85-year-old man with a history of femorofemoral bypass who developed this complication over a year after BCG treatment. He was successfully treated with explantation of the polytetrafluoroethylene (PTFE) graft, redo bypass with vein graft, and antituberculous medical therapy.

Metabolic pathways and inflammatory processes are under circadian regulation. Rhythmic immune cell recruitment is known to impact infection outcomes, but whether the circadian clock modulates immunometabolism remains unclear. We find that the molecular clock Bmal1 is induced by inflammatory stimulants, including Ifn-γ/lipopolysaccharide (M1) and tumor-conditioned medium, to maintain mitochondrial metabolism under metabolically stressed conditions in mouse macrophages. Upon M1 stimulation, myeloid-specific Bmal1 knockout (M-BKO) renders macrophages unable to sustain mitochondrial function, enhancing succinate dehydrogenase (SDH)-mediated mitochondrial production of reactive oxygen species as well as Hif-1α-dependent metabolic reprogramming and inflammatory damage. In tumor-associated macrophages, aberrant Hif-1α activation and metabolic dysregulation by M-BKO contribute to an immunosuppressive tumor microenvironment. Consequently, M-BKO increases melanoma tumor burden, whereas administering the SDH inhibitor dimethyl malonate suppresses tumor growth. Therefore, Bmal1 functions as a metabolic checkpoint that integrates macrophage mitochondrial metabolism, redox homeostasis and effector functions. This Bmal1-Hif-1α regulatory loop may provide therapeutic opportunities for inflammatory diseases and immunotherapy.

Background Animal and in vitro experiments implicate the Wnt pathway in cardiac development, fibrosis, vascular calcification, and atherosclerosis, but research in humans is lacking. We examined peripheral blood Wnt pathway gene expression and arterial stiffness in 369 healthy African ancestry men (mean age, 64 years). Methods and Results Gene expression was assessed using a custom Nanostring nCounter gene expression panel (N=43 genes) and normalized to housekeeping genes and background signal. Arterial stiffness was assessed via brachial‐ankle pulse‐wave velocity. Fourteen Wnt genes showed detectable expression and were tested individually as predictors of pulse‐wave velocity using linear regression, adjusting for age, height, weight, blood pressure, medication use, resting heart rate, current smoking, alcohol intake, and sedentary lifestyle. Adenomatous polyposis coli regulator of Wnt signaling pathway (APC), glycogen synthase kinase 3β (GSK3B), and transcription factor 4 (TCF4) were significantly associated with arterial stiffness (P

Sunk costs have been known to elicit violations of expected utility theory, in particular, the independence or cancellation axiom. Separately, violations of the stochastic dominance principle have been demonstrated in various settings despite the fact that descriptive models of choice favored in economics deem such violations irrational. However, it is currently unknown whether sunk costs also yield stochastic dominance violations. In two studies using a tri-colored roulette wheel choice task with non-equiprobable events yet equal payoffs, we observed that those who had sunk costs selected a stochastically dominated option significantly more than did those who had no costs. Moreover, a second study revealed that people chose a stochastically dominated option significantly more when the expected value was low compared to high. A model comparison of psychological explanations demonstrated that theories that incorporate a reference shift of the status quo could predict these sunk cost-based violations of stochastic dominance whereas other models could not.

Abstract Background Although several genetic variants for autism spectrum disorder (ASD) have now been identified, these largely occur sporadically or are de novo. Much less progress has been made in identifying inherited variants, even though the disorder itself is familial in the majority of cases. The objective of this study was to identify chromosomal regions that harbor inherited variants increasing the risk for ASD using an approach that examined both ASD and the broad autism phenotype (BAP) among a unique sample of extended pedigrees. Methods ASD and BAP were assessed using standardized tools in 28 pedigrees from Canada and the USA, each with at least three ASD-diagnosed individuals from two nuclear families. Genome-wide linkage analysis was performed using the posterior probability of linkage (PPL) statistic, a quasi-Bayesian method that provides strength of evidence for or against linkage in an essentially model-free manner, with outcomes on the probability scale. Results The results confirm appreciable interfamilial heterogeneity as well as a high level of intrafamilial heterogeneity. Both ASD and combined ASD/BAP specific loci are apparent. Conclusions Inclusion of subclinical phenotypes such as BAP should be more widely employed in genetic studies of ASD as a way of identifying inherited genetic variants for the disorder. Moreover, the results underscore the need for approaches to identifying genetic risk factors in extended pedigrees that are robust to high levels of inter/intrafamilial locus and allelic heterogeneity.

Summary: Throughout the developing nervous system, considerable synaptic re-organization takes place as postsynaptic neurons extend dendrites and incoming axons refine their synapses, strengthening some and eliminating others. It is well accepted that these processes rely on synaptic activity; however, the mechanisms that lead to this developmental reorganization are not fully understood. Here, we explore the regulation of cap-dependent translation, a mechanism known to play a role in synaptic growth and plasticity. Using sympathetic ganglia in α3 nicotinic acetylcholine receptor (nAChR)-knockout (KO) mice, we establish that electrophysiologically silent synapses between preganglionic axons and postsynaptic sympathetic neurons do not refine, and the growth of dendrites and the targeting of synapses on postsynaptic neurons are impaired. Remarkably, genetically removing 4E-BP, a suppressor of cap-dependent translation, from these α3 nAChR-KO mice largely restores these features. We conclude that synaptic connections can re-organize and refine without postsynaptic activity during post-natal development when 4E-BP-regulated cap-dependent translation is enhanced. : Synaptic activity is required for synaptic refinement and reorganization during post-natal development. Chong et al. find that silent synapses in superior cervical ganglia (SCG) refine when 4E-BP is genetically removed, suggesting that enhanced cap-dependent translation promotes synaptic refinement in the absence of postsynaptic activity. Keywords: sympathetic neurons, cap-dependent translation, synaptic refinement, synaptic activity, nicotinic receptors, silent synapses

Effective management of dairy farms requires an accurate prediction of pasture biomass. Generally, estimation of pasture biomass requires site-specific data, or often perfect world assumptions to model prediction systems when field measurements or other sensory inputs are unavailable. However, for small enterprises, regular measurements of site-specific data are often inconceivable. In this study, we approach the estimation of pasture biomass by predicting sward heights across the field. A convolution based sequential architecture is proposed for pasture height predictions using deep learning. We develop a process to create synthetic datasets that simulate the evolution of pasture growth over a period of 30 years. The deep learning based pasture prediction model (DeepPaSTL) is trained on this dataset while learning the spatiotemporal characteristics of pasture growth. The architecture purely learns from the trends in pasture growth through available spatial measurements and is agnostic to any site-specific data, or climatic conditions, such as temperature, precipitation, or soil condition. Our model performs within a 12% error margin even during the periods with the largest pasture growth dynamics. The study demonstrates the potential scalability of the architecture to predict any pasture size through a quantization approach during prediction. Results suggest that the DeepPaSTL model represents a useful tool for predicting pasture growth both for short and long horizon predictions, even with missing or irregular historical measurements.

The Metaverse is a pervasive expression of technological culture whose impact will be global. First, through knowledge, then through social, and now through geo-spatial, AI (the foundation of the Metaverse) will connect all entities on Earth through digital means thereby creating a three-dimensional informational and experiential layer across the world dubbed the Metaverse. The Metaverse has four characteristics: augmented reality, lifelogging, mirror worlds, and virtual reality. From the standpoint of Christian cultural engagement, a contextual theology has yet to be developed. In the work that follows, the Metaverse is engaged through a combination of contextualization and wholemaking from the standpoint of posthumanism and mysticism. The study focuses on evolutionary wholemaking as identified by Teilhard/Delio, while being guided by Bevans’ five (early) models of contextualization. The method of contextual wholemaking enables new ways of seeing, embracing, communing, complexifying, and creating within the four spheres of the Metaverse. After exploring the nature of the Metaverse in the first half of the paper, insights were gathered from the dialogue between contextual theology and culture and discussed in the second half of the paper.

Acinetobacter baumannii is currently classified as one of six pathogens that contribute to increased patient mortality. Thus, exploratory studies navigating alternative treatment strategies are of supreme interest. Herein, we completed minimum inhibitory concentration (MIC) testing, and time-kill analyses (TKA) on 50 carbapenem-resistant Acinetobacterbaumannii isolates including 28 colistin-resistant isolates. Upon testing of MEM or TGC in the presence of sub-inhibitory COL against the 50 isolates, there was a median 2-fold reduction in MEM and TGC MICs. In the TKAs, the COL+MEM combination was synergistic in 45 (90%) isolates and bactericidal in 43 (86%) isolates at 24 hours, whereas the COL+TGC combination TKAs demonstrated synergy in 32 (64%) isolates and bactericidal activity was shown in 28 (56%) isolates. Additionally, sulbactam (SUL) and TGC were added to the COL+MEM dual therapy regimen to assess the possible utility of a triple therapy regimen against five non-responsive isolates. The COL+MEM+SUL and COL+MEM+TGC regimens effectively restored synergy in (5/5) 100% of the isolates. The results of this study demonstrate the potential utility of COL combinations in the treatment of carbapenem-resistant isolates.

Abstract Background Autism spectrum disorder (ASD), a developmental disorder of early childhood onset, affects males four times more frequently than females, suggesting a role for the sex chromosomes. In this study, we describe a family with ASD in which a predicted pathogenic nonsense mutation in the X-chromosome gene RAB39B segregates with ASD phenotype. Methods Clinical phenotyping, microarray, and whole genome sequencing (WGS) were performed on the five members of this family. Maternal and female sibling X inactivation ratio was calculated, and phase was investigated. Mutant-induced pluripotent stem cells engineered for an exon 2 nonsense mutation were generated and differentiated into cortical neurons for expression and pathway analyses. Results Two males with an inherited RAB39B mutation both presented with macrocephaly, intellectual disability (ID), and ASD. Their female sibling with the same mutation presented with ID and a broad autism phenotype. In contrast, their transmitting mother has no neurodevelopmental diagnosis. Our investigation of phase indicated maternal preferential inactivation of the mutated allele, with no such bias observed in the female sibling. We offer the explanation that this bias in X inactivation may explain the absence of a neurocognitive phenotype in the mother. Our cellular knockout model of RAB39B revealed an impact on expression in differentiated neurons for several genes implicated in brain development and function, supported by our pathway enrichment analysis. Conclusions Penetrance for ASD is high among males but more variable among females with RAB39B mutations. A critical role for this gene in brain development and function is demonstrated.

Objective: Alternative activation (M2) of adipose tissue resident macrophage (ATM) inhibits obesity-induced metabolic inflammation. The underlying mechanisms remain unclear. Recent studies have shown that dysregulated lipid homeostasis caused by increased lipolysis in white adipose tissue (WAT) in the obese state is a trigger of inflammatory responses. We investigated the role of M2 macrophages in lipotoxicity-induced inflammation. Methods: We used microarray experiments to profile macrophage gene expression regulated by two M2 inducers, interleukin-4 (Il-4), and peroxisome proliferator-activated receptor delta/gamma (Pparδ/Pparγ) agonists. Functional validation studies were performed in bone marrow-derived macrophages and mice deprived of the signal transducer and activator of transcription 6 gene (Stat6; downstream effector of Il-4) or Pparδ/Pparγ genes (downstream effectors of Stat6). Palmitic acid (PA) and β-adrenergic agonist were employed to induce macrophage lipid loading in vitro and in vivo, respectively. Results: Profiling of genes regulated by Il-4 or Pparδ/Pparγ agonists reveals that alternative activation promotes the cell survival program, while inhibiting that of inflammation-related cell death. Deletion of Stat6 or Pparδ/Pparγ increases the susceptibility of macrophages to PA-induced cell death. NLR family pyrin domain containing 3 (Nlrp3) inflammasome activation by PA in the presence of lipopolysaccharide is also increased in Stat6−/− macrophages and to a lesser extent, in Pparδ/γ−/− macrophages. In concert, β-adrenergic agonist-induced lipolysis results in higher levels of cell death and inflammatory markers in ATMs derived from myeloid-specific Pparδ/γ−/− or Stat6−/− mice. Conclusions: Our data suggest that ATM cell death is closely linked to metabolic inflammation. Within WAT where concentrations of free fatty acids fluctuate, M2 polarization regulated by the Stat6-Ppar axis enhances ATM's tolerance to lipid-mediated stress, thereby maintaining the homeostatic state.

Summary: Lineage-specific transcription factors are critical for long-range enhancer interactions, but direct or indirect contributions of architectural proteins such as CCCTC-binding factor (CTCF) to enhancer function remain less clear. The LDB1 complex mediates enhancer-gene interactions at the β-globin locus through LDB1 self-interaction. We find that an LDB1-bound enhancer upstream of carbonic anhydrase 2 (Car2) activates its expression by interacting directly with CTCF at the gene promoter. Both LDB1 and CTCF are required for enhancer-Car2 looping, and the domain of LDB1 contacted by CTCF is necessary to rescue Car2 transcription in LDB1-deficient cells. Genome-wide studies and CRISPR/Cas9 genome editing indicate that LDB1-CTCF enhancer looping underlies activation of a substantial fraction of erythroid genes. Our results provide a mechanism by which long-range interactions of architectural protein CTCF can be tailored to achieve a tissue-restricted pattern of chromatin loops and gene expression. : It is unclear whether or how architectural proteins such as CTCF might contribute to interactions between enhancers and target genes. Lee et al. show that CTCF interacts directly with LDB1 and that a significant fraction of erythroid genes loop to LDB1-complex-occupied enhancers via CTCF sites within their promoters. Keywords: LDB1, CTCF, enhancer, looping, erythroid cells

Abstract Autism spectrum disorder is a developmental condition of early childhood onset, which impacts socio-communicative functioning and is principally genetic in etiology. Currently, more than 50 genomic loci are deemed to be associated with susceptibility to autism spectrum disorder, showing de novo and inherited unbalanced copy number variants and smaller insertions and deletions (indels), more complex structural variants, as well as single-nucleotide variants deemed of pathological significance. However, the phenotypes associated with many of these genes are variable, and penetrance is largely unelaborated in clinical descriptions. This case report describes a family harboring two copy number variant microdeletions, which affect regions of NRXN1 and MBD5—each well-established in association with risk of autism spectrum disorder and other neurodevelopmental disorders. Although each copy number variant would likely be categorized as pathologically significant, both genomic alterations are transmitted in this family from an unaffected father to the proband, and shared by an unaffected sibling. This family case illustrates the importance of recognizing that phenotype can vary among exon overlapping variants of the same gene, and the need to evaluate penetrance of such variants in order to properly inform on risks.

Severe appetite and weight loss define the eating disorder anorexia nervosa, and can also accompany the progression of some neurodegenerative disorders such as amyotrophic lateral sclerosis (ALS). Although acute loss of hypothalamic neurons that produce appetite-stimulating neuropeptide Y (Npy) and agouti-related peptide (Agrp) in adult mice or in mice homozygous for the anorexia (anx) mutation causes aphagia, our understanding of the factors that help maintain appetite regulatory circuitry is limited. Here we identify a mutation (C19T) that converts an arginine to a tryptophan (R7W) in the TYRO3 protein tyrosine kinase 3 (Tyro3) gene, which resides within the anx critical interval, as contributing to the severity of anx phenotypes. Our observation that, like Tyro3−/− mice, anx/anx mice exhibit abnormal secondary platelet aggregation suggested that the C19T Tyro3 variant might have functional consequences. Tyro3 is expressed in the hypothalamus and other brain regions affected by the anx mutation, and its mRNA localization appeared abnormal in anx/anx brains by postnatal day 19 (P19). The presence of wild-type Tyro3 transgenes, but not an R7W-Tyro3 transgene, doubled the weight and lifespans of anx/anx mice and near-normal numbers of hypothalamic Npy-expressing neurons were present in Tyro3-transgenic anx/anx mice at P19. Although no differences in R7W-Tyro3 signal sequence function or protein localization were discernible in vitro, distribution of R7W-Tyro3 protein differed from that of Tyro3 protein in the cerebellum of transgenic wild-type mice. Thus, R7W-Tyro3 protein localization deficits are only detectable in vivo. Further analyses revealed that the C19T Tyro3 mutation is present in a few other mouse strains, and hence is not the causative anx mutation, but rather an anx modifier. Our work shows that Tyro3 has prosurvival roles in the appetite regulatory circuitry and could also provide useful insights towards the development of interventions targeting detrimental weight loss.

The Canadian beaver (Castor canadensis) is the largest indigenous rodent in North America. We report a draft annotated assembly of the beaver genome, the first for a large rodent and the first mammalian genome assembled directly from uncorrected and moderate coverage (< 30 ×) long reads generated by single-molecule sequencing. The genome size is 2.7 Gb estimated by k-mer analysis. We assembled the beaver genome using the new Canu assembler optimized for noisy reads. The resulting assembly was refined using Pilon supported by short reads (80 ×) and checked for accuracy by congruency against an independent short read assembly. We scaffolded the assembly using the exon–gene models derived from 9805 full-length open reading frames (FL-ORFs) constructed from the beaver leukocyte and muscle transcriptomes. The final assembly comprised 22,515 contigs with an N50 of 278,680 bp and an N50-scaffold of 317,558 bp. Maximum contig and scaffold lengths were 3.3 and 4.2 Mb, respectively, with a combined scaffold length representing 92% of the estimated genome size. The completeness and accuracy of the scaffold assembly was demonstrated by the precise exon placement for 91.1% of the 9805 assembled FL-ORFs and 83.1% of the BUSCO (Benchmarking Universal Single-Copy Orthologs) gene set used to assess the quality of genome assemblies. Well-represented were genes involved in dentition and enamel deposition, defining characteristics of rodents with which the beaver is well-endowed. The study provides insights for genome assembly and an important genomics resource for Castoridae and rodent evolutionary biology.

ABSTRACT Carbapenem-resistant Klebsiella pneumoniae (CRKP) strains belonging to sequence type 258 (ST258) are frequent causes of hospital-associated outbreaks and are a major contributor to the spread of carbapenemases. This genetic lineage emerged several decades ago and remains a major global health care challenge. In this study, genomic epidemiology was used to investigate the emergence, evolution, and persistence of ST258 carbapenem-resistant K. pneumoniae outbreak-causing lineages at a large tertiary care hospital over 8 years. A time-based phylogenetic analysis of 136 ST258 isolates demonstrated the succession of multiple genetically distinct ST258 sublineages over the 8-year period. Ongoing genomic surveillance identified the emergence and persistence of several distinct clonal ST258 populations. Patterns of multidrug resistance determinants and plasmid replicons were consistent with continued evolution and persistence of these populations. Five ST258 outbreaks were documented, including three that were caused by the same clonal lineage. Mutations in genes encoding effectors of biofilm production and iron acquisition were identified among persistent clones. Two emergent lineages bearing K. pneumoniae integrative conjugative element 10 (ICEKp10) and harboring yersiniabactin and colibactin virulence factors were identified. The results show how distinct ST258 subpopulations have evolved and persisted within the same hospital over nearly a decade. IMPORTANCE The carbapenem class of antibiotics is invaluable for the treatment of selected multidrug-resistant Gram-negative pathogens. The continued transmission of carbapenem-resistant bacteria such as ST258 K. pneumoniae is of serious global public health concern, as treatment options for these infections are limited. This genomic epidemiologic investigation traced the natural history of ST258 K. pneumoniae in a single health care setting over nearly a decade. We found that distinct ST258 subpopulations have caused both device-associated and ward-associated outbreaks, and some of these populations remain endemic within our hospital to the present day. The finding of virulence determinants among emergent ST258 clones supports the idea of convergent evolution of drug-resistant and virulent CRKP strains and highlights the need for continued surveillance, prevention, and control efforts to address emergent and evolving ST258 populations in the health care setting.

We present a computationally inexpensive, flexible feature identification method which uses a comparison of time series to identify a rank-ordered set of features in geophysically-sourced data sets. Many physical phenomena perturb multiple physical variables nearly simultaneously, and so features are identified as time periods in which there are local maxima of absolute deviation in all time series. Unlike other available methods, this method allows the analyst to tune the method using their knowledge of the physical context. The method is applied to a data set from a moored array of instruments deployed in the coastal environment of Monterey Bay, California, and a data set from sensors placed within the submerged Yax Chen Cave System in Tulum, Quintana Roo, Mexico. These example data sets demonstrate that the method allows for the automated identification of features which are worthy of further study.

Despite abundant focus on responsible care of laboratory animals, we argue that inattention to the maltreatment of wildlife constitutes an ethical blind spot in contemporary animal research. We begin by reviewing significant shortcomings in legal and institutional oversight, arguing for the relatively rapid and transformational potential of editorial oversight at journals in preventing harm to vertebrates studied in the field and outside the direct supervision of institutions. Straightforward changes to animal care policies in journals, which our analysis of 206 journals suggests are either absent (34%), weak, incoherent, or neglected by researchers, could provide a practical, effective, and rapidly imposed safeguard against unnecessary suffering. The Animals in Research: Reporting On Wildlife (ARROW) guidelines we propose here, coupled with strong enforcement, could result in significant changes to how animals involved in wildlife research are treated. The research process would also benefit. Sound science requires animal subjects to be physically, physiologically, and behaviorally unharmed. Accordingly, publication of methods that contravenes animal welfare principles risks perpetuating inhumane approaches and bad science.

Gastrointestinal (GI) mucosal dysfunction predicts and likely contributes to non-infectious comorbidities and mortality in HIV infection and persists despite antiretroviral therapy. However, the mechanisms underlying this dysfunction remain incompletely understood. Neutrophils are important for containment of pathogens but can also contribute to tissue damage due to their release of reactive oxygen species and other potentially harmful effector molecules. Here we used a flow cytometry approach to investigate increased neutrophil lifespan as a mechanism for GI neutrophil accumulation in chronic, treated HIV infection and a potential role for gastrointestinal dysbiosis. We report that increased neutrophil survival contributes to neutrophil accumulation in colorectal biopsy tissue, thus implicating neutrophil lifespan as a new therapeutic target for mucosal inflammation in HIV infection. Additionally, we characterized the intestinal microbiome of colorectal biopsies using 16S rRNA sequencing. We found that a reduced Lactobacillus: Prevotella ratio associated with neutrophil survival, suggesting that intestinal bacteria may contribute to GI neutrophil accumulation in treated HIV infection. Finally, we provide evidence that Lactobacillus species uniquely decrease neutrophil survival and neutrophil frequency in vitro, which could have important therapeutic implications for reducing neutrophil-driven inflammation in HIV and other chronic inflammatory conditions.

Summary: Complement-mediated cytotoxicity may act as a selective pressure for tumor overexpression of complement regulators. We hypothesize that the same selective pressure could lead to complement alterations at the genetic level. We find that, when analyzed as a pathway, mutations in complement genes occur at a relatively high frequency and are associated with changes in overall survival across a number of cancer types. Analysis of pathways expressed in patients with complement mutations that are associated with poor overall survival reveals crosstalk between complement and hypoxia in colorectal cancer. The importance of this crosstalk is highlighted by two key findings: hypoxic signaling is increased in tumors harboring complement mutations, and hypoxic tumor cells are resistant to complement-mediated cytotoxicity due, in part, to hypoxia-induced expression of complement regulator CD55. The range of strategies employed by tumors to dysregulate the complement system testifies to the importance of this pathway in tumor progression. : Mutations in the complement system are prevalent across cancers. Olcina et al. find that colorectal cancers with complement component mutations are associated with increased hypoxic signaling and poor overall survival outcomes. Hypoxia-induced expression of complement regulator CD55 controls complement-mediated cytotoxicity. Keywords: innate immunity, cancer, mutations, hypoxia, complement system, complement-mediated cytotoxicity

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Research Highlights: Forest phenology and productivity were responsive to seasonal heating and chilling accumulation, but responses differed across the temperature range. Background and Objectives: Temperate forests have responded to recent climate change worldwide, but the pattern and magnitude of response have varied, necessitating additional studies at higher spatial and temporal resolutions. We investigated climatic drivers of inter-annual variation in forest phenology and productivity across the Algonquin-to-Adirondacks (A2A) conservation corridor of eastern North America. Methods: We used remotely sensed indices from the AVHRR sensor series and a suite of gridded climate data from the Daymet database spanning from 1989–2014. We used random forest regression to characterize forest–climate relationships between forest growth indices and climatological variables. Results: A large portion of the annual variation in phenology and productivity was explained by climate (pR2 > 80%), with variation largely driven by accumulated heating and chilling degree days. Only very minor relationships with precipitation-related variables were evident. Conclusions: Our results indicate that anthropogenic climate change in the A2A has not yet reached the point of triggering widespread changes in forest phenology and productivity, but the sensitivity of forest growth to inter-annual variation in seasonal temperature accumulation suggests that more temperate forest area will be affected by climate change as warming continues.

Coastal dissolved oxygen (DO) concentrations have a profound impact on nearshore ecosystems and, in recent years, there has been an increased prevalance of low DO hypoxic events that negatively impact nearshore organisms. Even with advanced numerical models, accurate prediction of coastal DO variability is challenging and computationally expensive. Here, we apply machine learning techniques in order to reconstruct and predict nearshore DO concentrations in a small coastal embayment while using a comprehensive set of nearshore and offshore measurements and easily measured input (training) parameters. We show that both random forest regression (RFR) and support vector regression (SVR) models accurately reproduce both the offshore DO and nearshore DO with extremely high accuracy. In general, RFR consistently peformed slightly better than SVR, the latter of which was more difficult to tune and took longer to train. Although each of the nearshore datasets were able to accurately predict DO values using training data from the same site, the model only had moderate success when using training data from one site to predict DO at another site, which was likely due to the the complexities in the underlying dynamics across the sites. We also show that high accuracy can be achieved with relatively little training data, highlighting a potential application for correcting time series with missing DO data due to quality control or sensor issues. This work establishes the ability of machine learning models to accurately reproduce DO concentrations in both offshore and nearshore coastal waters, with important implications for the ability to detect and indirectly measure coastal hypoxic events in near real-time. Future work should explore the ability of machine learning models in order to accurately forecast hypoxic events.

Abstract Background Ambulatory antibiotic prescribing contributes to the development of antibiotic resistance and increases societal costs. Here, we estimate the hidden societal cost of antibiotic resistance per antibiotic prescribed in the United States. Methods In an exploratory analysis, we used published data to develop point and range estimates for the hidden societal cost of antibiotic resistance (SCAR) attributable to each ambulatory antibiotic prescription in the United States. We developed four estimation methods that focused on the antibiotic-resistance attributable costs of hospitalization, second-line inpatient antibiotic use, second-line outpatient antibiotic use, and antibiotic stewardship, then summed the estimates across all methods. Results The total SCAR attributable to each ambulatory antibiotic prescription was estimated to be $13 (range: $3–$95). The greatest contributor to the total SCAR was the cost of hospitalization ($9; 69 % of the total SCAR). The costs of second-line inpatient antibiotic use ($1; 8 % of the total SCAR), second-line outpatient antibiotic use ($2; 15 % of the total SCAR) and antibiotic stewardship ($1; 8 %). This apperars to be an error.; of the total SCAR) were modest contributors to the total SCAR. Assuming an average antibiotic cost of $20, the total SCAR attributable to each ambulatory antibiotic prescription would increase antibiotic costs by 65 % (range: 15–475 %) if incorporated into antibiotic costs paid by patients or payers. Conclusions Each ambulatory antibiotic prescription is associated with a hidden SCAR that substantially increases the cost of an antibiotic prescription in the United States. This finding raises concerns regarding the magnitude of misalignment between individual and societal antibiotic costs.

Oropharyngeal squamous cell carcinoma (OPSCC) is increasing in incidence in the United States and in many countries worldwide primarily as a result of increasing rates of human papillomavirus (HPV) infection. HPV-positive OPSCC represents a distinct disease entity from head and neck squamous cell carcinoma caused by traditional risk factors such as tobacco and alcohol, with different epidemiology, patterns of failure, and expected outcomes. Because patients with HPV-positive OPSCC have a younger median age and superior prognosis compared with their HPV-negative counterparts, they live longer with the morbidity of treatment, which can be severe. Therefore, efforts are under way to de-escalate therapy in favorable-risk patients while maintaining treatment efficacy. Additional work is being undertaken to discover new therapies that may benefit both HPV-positive and HPV-negative patient subsets. Herein, we will review the available data for the evolving treatment paradigms in OPSCC as well as discuss ongoing clinical trials.

Abstract Background This paper contributes to research on the education-health association by extending the scope of inquiry to adult inmates. Not only are inmates excluded from most nationally representative studies of health but they also represent a highly select group in terms of both education and health. As such, our study provides new information about the health of incarcerated populations and it extends the generalizability of the education-health association beyond the non-institutionalized population. Methods We use a prison-level fixed-effects regression model with the 2004 Survey of Inmates in State Correctional Facilities (n = 287 facilities) to evaluate the effects of education on a standardized morbidity scale of 11 lifetime and current health conditions among incarcerated men (n = 10,493) and women (n = 2,797). Results Education prior to incarceration is negatively associated with lifetime health problems for both women and men and the association is stronger among women. Among inmates who enter prison with less than a GED level of education, attaining a GED in prison is associated with better current health outcomes for men, but not women. Conclusions The generalization of the education-health association among prisoners further highlights the fundamental nature of education as a health promotive resource. Discussed are the implications for the education-health literature in general and health promotion efforts among incarcerated adults specifically.

Abstract Background In the past two decades, rates of suicide mortality have declined among most OECD member states. Two notable exceptions are Japan and South Korea, where suicide mortality has increased by 20 % and 280 %, respectively. Methods Population and suicide mortality data were collected through national statistics organizations in Japan and South Korea for the period 1985 to 2010. Age, period of observation, and birth cohort membership were divided into five-year increments. We fitted a series of intrinsic estimator age-period-cohort models to estimate the effects of age-related processes, secular changes, and birth cohort dynamics on the rising rates of suicide mortality in the two neighboring countries. Results In Japan, elevated suicide rates are primarily driven by period effects, initiated during the Asian financial crisis of the late 1990s. In South Korea, multiple factors appear to be responsible for the stark increase in suicide mortality, including recent secular changes, elevated suicide risks at older ages in the context of an aging society, and strong cohort effects for those born between the Great Depression and the aftermath of the Korean War. Conclusion In spite of cultural, demographic and geographic similarities in Japan and South Korea, the underlying causes of increased suicide mortality differ across these societies—suggesting that public health responses should be tailored to fit each country’s unique situation.

This study presents a post-facto evaluation of the local capacity development processes used under co-management of fisheries and other resources of southern Bangladesh. It answers the question of how supportive were the capacity development tools used in implementing co-management. An 18 month study was conducted and six cases were investigated to understand the approaches to co-management programs used to develop local capacity. Founded in pragmatism and viewing co-management through a governance lens, a comparative case study method was used that combined both qualitative and quantitative research approaches for data collection and subsequent analysis. This study provides empirical evidence that co-management programs have applied a number of strategies (e.g. human resource and economic development) to enhance local capacities. However, these strategies have achieved mixed results with regard to developing governance that supports livelihoods. Training provided to develop human resources and economic capacity were not useful for fishers or had little lasting effects on fisheries development due to poor monitoring and a disconnection with the needs of local users. This study concludes that comanagement can facilitate local capacity but in order to realize the full potential of this approach we must address the issues of inappropriate technologies for training, the financial barriers to fishers with low cash income, and uneven power relationships among stakeholders, to create an enabling environment for effective modern governance of the fisheries commons. Our findings indicate a needsbased approach to capacity building is needed in order to support the livelihoods of local users through co-management

Abstract Objective To measure the benefits to household caregivers of a psychotherapeutic intervention for adolescents and young adults living in a war-affected area. Methods Between July 2012 and July 2013, we carried out a randomized controlled trial of the Youth Readiness Intervention - a cognitive-behavioural intervention for war-affected young people who exhibit depressive and anxiety symptoms and conduct problems - in Freetown, Sierra Leone. Overall, 436 participants aged 15-24 years were randomized to receive the intervention (n = 222) or care as usual ( n = 214). Household caregivers for the participants in the intervention arm (n = 101) or control arm ( n = 103) were interviewed during a baseline survey and again, if available (n = 155), 12 weeks later in a follow-up survey. We used a burden assessment scale to evaluate the burden of care placed on caregivers in terms of emotional distress and functional impairment. The caregivers' mental health - i.e. internalizing, externalizing and prosocial behaviour - was evaluated using the Oxford Measure of Psychosocial Adjustment. Difference-in-differences multiple regression analyses were used, within an intention-to-treat framework, to estimate the treatment effects. Findings Compared with the caregivers of participants of the control group, the caregivers of participants of the intervention group reported greater reductions in emotional distress (scale difference: 0.252; 95% confidence interval, CI: 0.026-0.4782) and greater improvements in prosocial behaviour (scale difference: 0.249; 95% CI: 0.012-0.486) between the two surveys. Conclusion A psychotherapeutic intervention for war-affected young people can improve the mental health of their caregivers.

The degree to which we can understand the multi-scale organization of cellular life is tied to how well our models can represent this organization and the processes that drive its evolution. This paper uses Vivarium—an engine for composing heterogeneous computational biology models into integrated, multi-scale simulations. Vivarium’s approach is demonstrated by combining several sub-models of biophysical processes into a model of chemotactic E. coli that exchange molecules with their environment, express the genes required for chemotaxis, swim, grow, and divide. This model is developed incrementally, highlighting cross-compartment mechanisms that link E. coli to its environment, with models for: (1) metabolism and transport, with transport moving nutrients across the membrane boundary and metabolism converting them to useful metabolites, (2) transcription, translation, complexation, and degradation, with stochastic mechanisms that read real gene sequence data and consume base pairs and ATP to make proteins and complexes, and (3) the activity of flagella and chemoreceptors, which together support navigation in the environment.

Both mammals and birds exhibit two sleep states, slow wave sleep (SWS) and rapid eye movement (REM) sleep. Studying certain aspects of sleep-related electrophysiology in freely behaving animals can present numerous methodological constraints, particularly when even fine body movements interfere with electrophysiological signals. Interestingly, under light general anesthesia, mammals and birds also exhibit slow waves similar to those observed during natural SWS. For these reasons, slow waves occurring under general anesthesia are commonly used in the investigation of sleep-related neurophysiology. However, how spectral properties of slow waves induced by anesthesia correspond to those occurring during natural SWS in birds has yet to be investigated systematically. In this study, we systematically analyzed spectral properties of electroencephalographic (EEG) patterns of pigeons (Columba livia) occurring under two commonly used anesthetics, isoflurane and urethane. These data were compared with EEG patterns during natural sleep. Slow waves occurring during spontaneous SWS, and those induced with isoflurane and urethane all showed greatest absolute power in the slowest frequencies (

Niemann-Pick disease, type C1 (NPC1) is a lysosomal disease characterized by progressive cerebellar ataxia. In NPC1, a defect in cholesterol transport leads to endolysosomal storage of cholesterol and decreased cholesterol bioavailability. Purkinje neurons are sensitive to the loss of NPC1 function. However, degeneration of Purkinje neurons is not uniform. They are typically lost in an anterior-to-posterior gradient with neurons in lobule X being resistant to neurodegeneration. To gain mechanistic insight into factors that protect or potentiate Purkinje neuron loss, we compared RNA expression in cerebellar lobules III, VI, and X from control and mutant mice. An unexpected finding was that the gene expression differences between lobules III/VI and X were more pronounced than those observed between mutant and control mice. Functional analysis of genes with anterior to posterior gene expression differences revealed an enrichment of genes related to neuronal cell survival within the posterior cerebellum. This finding is consistent with the observation, in multiple diseases, that posterior Purkinje neurons are, in general, resistant to neurodegeneration. To our knowledge, this is the first study to evaluate anterior to posterior transcriptome-wide changes in gene expression in the cerebellum. Our data can be used to not only explore potential pathological mechanisms in NPC1, but also to further understand cerebellar biology.

White matter hyperintensities (WMH) are common in acute ischemic stroke patients. Although WMH volume has been reported to influence post-stroke cognition, it is still not clear whether WMH location, independent of acute ischemic lesion (AIL) volume and location, contributes to cognitive impairment after stroke. Here, we proposed a multiple-lesion symptom mapping model that considers both the presence of WMH and AIL to measure the additional contribution of WMH locations to post-stroke cognitive impairment. Seventy-six first-ever stroke patients with AILs in the left hemisphere were examined by Montreal Cognitive Assessment (MoCA) at baseline and 1 year after stroke. The association between the location of AIL and WMH and global cognition was investigated by a multiple-lesion symptom mapping (MLSM) model based on support vector regression (SVR). To explore the relative merits of MLSM over the existing lesion-symptom mapping approaches with only AIL considered (mass-univariate VLSM and SVR-LSM), we measured the contribution of the significant AIL and/or WMH clusters from these models to post-stroke cognitive impairment. In addition, we compared the significant WMH locations identified by the optimal SVR-MLSM model for cognitive impairment at baseline and 1 year post stroke. The identified strategic locations of WMH significantly contributed to the prediction of MoCA at baseline (short-term) and 1 year (long-term) after stroke independent of the strategic locations of AIL. The significant clusters of WMH for short-term and long-term post-stroke cognitive impairment were mainly in the corpus callosum, corona radiata, and posterior thalamic radiation. We noted that in some regions, the AIL clusters that were significant for short-term outcome were no longer significant for long-term outcome, and interestingly more WMH clusters in these regions became significant for long-term outcome compared to short-term outcome. This indicated that there are some regions where local WMH burden has larger impact than AIL burden on the long-term post-stroke cognitive impairment. In consequence, SVR-MLSM was effective in identifying the WMH locations that have additional impact on post-stroke cognition on top of AIL locations. Such a method can also be applied to other lesion-behavior studies where multiple types of lesions may have potential contributions to a specific behavior.

Abstract Significant treeline advance can only occur with successful establishment, growth, and survival of new seedlings. Several studies have examined microsite factors at single locations to explain the presence/absence of seedlings at treeline. We conducted a much larger observational study and included multiple factors to determine (1) which variable(s) was/were most important, (2) whether their importance differed between aspects, and (3) whether the same variables explained variation in seedling growth and damage. We analyzed five biophysical and six shrub variables along four forest–tundra ecotones in southwest Yukon at 640 points. The model that best explained seedling occurrence was similar between north‐ and south‐facing slopes. Of all variables, seedling occurrence was best explained by the proximity, height, and upslope orientation of shrubs (relative to the seedling). The data indicate an optimal range of shrub cover, which differed with aspect. On north‐facing slopes, seedlings occurred most often when shrub cover exceeded 13%, while on south‐facing slopes seedlings occurred most when shrub cover was between 9% and 72%. We also found that with the exception of shrub‐related factors, very few biophysical variables explained size and growth characteristics and damage of tree seedlings, suggesting that the relative strength and importance of variables change depending on the life stage and size of the tree seedling. Collectively, our results demonstrate a non‐random distribution of seedlings in the forest–tundra ecotone, suggesting that as shrub distributions change with climate change, colonization sites for seedlings will also be influenced.

Drosophila Argonaute2 (AGO2) has been shown to regulate expression of certain loci in an RNA interference (RNAi)-independent manner, but its genome-wide function on chromatin remains unknown. Here, we identified the nuclear scaffolding protein LaminB as a novel interactor of AGO2. When either AGO2 or LaminB are depleted in Kc cells, similar transcription changes are observed genome-wide. In particular, changes in expression occur mainly in active or potentially active chromatin, both inside and outside LaminB-associated domains (LADs). Furthermore, we identified a somatic target of AGO2 transcriptional repression, no hitter (nht), which is immersed in a LAD located within a repressive topologically-associated domain (TAD). Null mutation but not catalytic inactivation of AGO2 leads to ectopic expression of nht and downstream spermatogenesis genes. Depletion of either AGO2 or LaminB results in reduced looping interactions within the nht TAD as well as ectopic inter-TAD interactions, as detected by 4C-seq analysis. Overall, our findings reveal coordination of AGO2 and LaminB function to dictate genome architecture and thereby regulate gene expression.

Purpose: To investigate differences between rating of perceived exertion (RPE) and percentage one-repetition maximum (1RM) load assignment in resistance-trained males (19–35 years) performing protocols with matched sets and repetitions differentiated by load-assignment.Methods: Participants performed squats then bench press 3x/weeks in a daily undulating format over 8-weeks. Participants were counterbalanced by pre-test 1RM then assigned to percentage 1RM (1RMG, n = 11); load-assignment via percentage 1RMs, or RPE groups (RPEG, n = 10); participant-selected loads to reach target RPE ranges. Ultrasonography determined pre and post-test pectoralis (PMT), and vastus lateralis muscle thickness at 50 (VLMT50) and 70% (VLMT70) femur-length.Results: Bench press (1RMG +9.64 ± 5.36; RPEG + 10.70 ± 3.30 kg), squat (1RMG + 13.91 ± 5.89; RPEG + 17.05 ± 5.44 kg) and their combined-total 1RMs (1RMG + 23.55 ± 10.38; RPEG + 27.75 ± 7.94 kg) increased (p < 0.05) in both groups as did PMT (1RMG + 1.59 ± 1.33; RPEG +1.90 ± 1.91 mm), VLMT50 (1RMG +2.13 ± 1.95; RPEG + 1.85 ± 1.97 mm) and VLMT70 (1RMG + 2.40 ± 2.22; RPEG + 2.31 ± 2.27 mm). Between-group differences were non-significant (p > 0.05). Magnitude-based inferences revealed 79, 57, and 72% chances of mean small effect size (ES) advantages for squat; ES 90% confidence limits (CL) = 0.50 ± 0.63, bench press; ES 90% CL = 0.28 ± 0.73, and combined-total; ES 90% CL = 0.48 ± 0.68 respectively, in RPEG. There were 4, 14, and 6% chances 1RMG had a strength advantage of the same magnitude, and 18, 29, and 22% chances, respectively of trivial differences between groups.Conclusions: Both loading-types are effective. However, RPE-based loading may provide a small 1RM strength advantage in a majority of individuals.

There is an established relationship between primary DNA sequence, secondary and tertiary chromatin structure, and transcriptional activity, suggesting that observed differences in one of these properties may reflect changes in the others. Here, we exploit these relationships to show that variations in DNA structure can be used to identify a wide range of genomic alterations in mammalian samples. In this proof-of-concept study we characterized and compared genome-wide histone occupancy by ChIP-Seq, DNA accessibility by ATAC-Seq, and chromosomal conformation by Hi-C for five CRISPR/Cas9-modified mammalian cell lines and their unmodified parent strains, as well as in one modified tissue sample and its parent strain. The results showed that the impact of genomic alterations on each of the levels of DNA organization varied depending on mutation type (insertion or deletion), size, and genomic location. The largest genomic alterations we identified included chromosomal rearrangements and deletions (greater than 200 Kb) in four of the modified cell lines, which can be difficult to identify by standard whole genome sequencing analysis. This multi-level DNA organizational analysis provides a sensitive approach for identifying a wide range of genomic and epigenomic perturbations that can be utilized for biomedical and biosecurity applications.

Growth responses of trees and shrubs to climate often exhibit unexplained variation in alpine regions, making it difficult to predict how they will respond to future changes in climate. We sought to characterize and explain this variability in southwest Yukon, a topographically complex region of subarctic Canada. We collected cores and sections from 360 spruce trees and 480 willow shrubs across treelines on north and south aspects in six valleys spanning two mountain ranges. We compared growth rates, growth patterns, and climate-growth responses between species and topographic factors. South aspects had wider tree rings and higher tree and shrub interseries correlations than north aspects, likely because of shallow active layers on the latter. Growth patterns and responses to climate did not vary between aspects or elevations but differed slightly between mountain ranges, likely because of differences in spring soil moisture content between ranges. Growth responses of both species to summer temperature were positive, but tree growth was negatively correlated to spring temperature and shrub growth was negatively correlated to summer precipitation, both of which are projected to increase along with summer temperature. Future changes in climate could therefore reduce the growth of one or both species in southwest Yukon.

Salmonella vertebral discitis/osteomyelitis is a rare manifestation of Salmonella infection. Here, we report a case of a 54-year-old Caucasian male who presented with five weeks of progressively worsening bilateral low back, buttock, and lower extremity pain following an 8-foot fall onto concrete from a ladder. Initial workup following the fall included hip X-ray and MRI of the lumbar spine and revealed only mild lumbar facet arthropathy and moderate left neural foraminal stenosis at L3-L4 without any concomitant hip or spine fracture. The patient’s pain continued to increase in severity over the next several weeks, and he was evaluated by multiple healthcare professionals with no discovered pathology. Approximately 5 weeks following the fall, repeat CT scan and MRI were conducted which then revealed extensive findings of discitis/osteomyelitis at L5–S1 as well as an epidural abscess resulting in severe narrowing of the central spinal canal. Patient underwent emergent decompression laminectomy and discectomy at L5–S1 with evacuation of the epidural abscess. Intraoperative tissue and wound cultures revealed Salmonella enterica serovar Agbeni. The patient recovered well and was discharged on an eight-week regimen of IV ceftriaxone. He has since recovered appropriately with no neurologic deficits. Important takeaways from this case include continuing to work up patients whose pain or condition is not consistent with radiographic findings and the importance of clinical intuition. This case also highlights the use of intraoperative cultures and sensitivities to correctly direct antibiotic management. Lastly, this report adds to the paucity of literature surrounding Salmonella Agbeni-related discitis and epidural abscesses and makes the suggestion that traumatic incidents such as a fall may instigate these infections.

It is widely accepted that cAMP regulates gene transcription principally by activating the protein kinase A (PKA)-targeted transcription factors. Here, we show that cAMP enhances the generation of 5-hydroxymethylcytosine (5hmC) in multiple cell types. 5hmC is converted from 5-methylcytosine (5mC) by Tet methylcytosine dioxygenases, for which Fe(II) is an essential cofactor. The promotion of 5hmC was mediated by a prompt increase of the intracellular labile Fe(II) pool (LIP). cAMP enhanced the acidification of endosomes for Fe(II) release to the LIP likely through RapGEF2. The effect of cAMP on Fe(II) and 5hmC was confirmed by adenylate cyclase activators, phosphodiesterase inhibitors, and most notably by stimulation of G protein-coupled receptors (GPCR). The transcriptomic changes caused by cAMP occurred in concert with 5hmC elevation in differentially transcribed genes. Collectively, these data show a previously unrecognized regulation of gene transcription by GPCR-cAMP signaling through augmentation of the intracellular labile Fe(II) pool and DNA hydroxymethylation.