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Abstract In a series of experiments involving beliefs and misinformation beliefs, we find that individuals who are prompted with a counterfactual mindset are significantly more likely to change their existing beliefs when presented with evidence that contradicts their beliefs. While research finds that beliefs that are considered part of one’s identity are highly resistant to change in the face of evidence that challenges these beliefs, four experiments provide evidence that counterfactual generation causes individuals to adjust beliefs and correct misinformation beliefs in response to contradicting evidence. Indeed, we find that a counterfactual mindset was effective in promoting incorporation of accurate facts and causing individuals to revise misinformation beliefs about COVID vaccination safety for a large sample of individuals who have rejected COVID vaccinations. Finally, the results of the psychophysiological experiment reveal that counterfactual generation alters decision makers’ search strategies, increases their cognitive arousal in response to evidence that challenges their beliefs, and increases their desire to seek out disconfirming evidence. Overall, the four experiments indicate that counterfactual generation can effectively activate mindsets that increase individuals’ willingness to evaluate evidence that contradicts their beliefs and adjust their beliefs in response to evidence.

Abstract Tests of the brain’s ability to process complex sounds (central auditory tests) correlate with overall measures of neurocognitive performance. In the low- middle-income countries where resources to conduct detailed cognitive testing is limited, tests that assess the central auditory system may provide a novel and useful way to track neurocognitive performance. This could be particularly useful for children living with HIV (CLWH). To evaluate this, we administered central auditory tests to CLWH and children living without HIV and examined whether central auditory tests given early in a child’s life could predict later neurocognitive performance. We used a machine learning technique to incorporate factors known to affect performance on neurocognitive tests, such as education. The results show that central auditory tests are useful predictors of neurocognitive performance and perform as well or in some cases better than factors such as education. Central auditory tests may offer an objective way to track neurocognitive performance in CLWH.

Circadian clocks are composed of transcription-translation negative feedback loops that pace rhythms of gene expression to the diurnal cycle. In the filamentous fungus Neurospora crassa, the proteins Frequency (FRQ), the FRQ-interacting RNA helicase (FRH), and Casein-Kinase I (CK1) form the FFC complex that represses expression of genes activated by the white-collar complex (WCC). FRQ orchestrates key molecular interactions of the clock despite containing little predicted tertiary structure. Spin labeling and pulse-dipolar electron spin resonance spectroscopy provide domain-specific structural insights into the 989-residue intrinsically disordered FRQ and the FFC. FRQ contains a compact core that associates and organizes FRH and CK1 to coordinate their roles in WCC repression. FRQ phosphorylation increases conformational flexibility and alters oligomeric state, but the changes in structure and dynamics are non-uniform. Full-length FRQ undergoes liquid–liquid phase separation (LLPS) to sequester FRH and CK1 and influence CK1 enzymatic activity. Although FRQ phosphorylation favors LLPS, LLPS feeds back to reduce FRQ phosphorylation by CK1 at higher temperatures. Live imaging of Neurospora hyphae reveals FRQ foci characteristic of condensates near the nuclear periphery. Analogous clock repressor proteins in higher organisms share little position-specific sequence identity with FRQ; yet, they contain amino acid compositions that promote LLPS. Hence, condensate formation may be a conserved feature of eukaryotic clocks.

Using logistic regression and neural networks, the aim of this study was to compare model performance when predicting player draft outcome during the 2021 AFL National Draft. Physical testing, in-game movement and technical involvements were collected from 708 elite-junior Australian Rules football players during consecutive seasons. Predictive models were generated using data from 465 players (2017 to 2020). Data from 243 players were then used to prospectively predict the 2021 AFL National Draft. Logistic regression and neural network models were compared for specificity, sensitivity and accuracy using relative cut-off thresholds from 5% to 50%. Using factored and unfactored data, and a range of relative cut-off thresholds, neural networks accounted for 73% of the 40 best performing models across positional groups and data configurations. Neural networks correctly classified more drafted players than logistic regression in 88% of cases at draft rate (15%) and convergence threshold (35%). Using individual variables across thresholds, neural networks (specificity = 79 ± 13%, sensitivity = 61 ± 24%, accuracy = 76 ± 8%) were consistently superior to logistic regression (specificity = 73 ± 15%, sensitivity = 29 ± 14%, accuracy = 66 ± 11%). Where the goal is to identify talented players with draft potential, model sensitivity is paramount, and neural networks were superior to logistic regression.

Introduction Invasive non-typhoidal Salmonella (iNTS) serovars are a major cause of community-acquired bloodstream infections in sub-Saharan Africa (SSA). In this setting, Salmonella enterica serovar Typhimurium accounts for two-thirds of infections and is associated with an estimated case fatality rate of 15%–20%. Several iNTS vaccine candidates are in early-stage assessment which—if found effective—would provide a valuable public health tool to reduce iNTS disease burden. The CHANTS study aims to develop a first-in-human Salmonella Typhimurium controlled human infection model, which can act as a platform for future vaccine evaluation, in addition to providing novel insights into iNTS disease pathogenesis.Methods and analysis This double-blind, safety and dose-escalation study will randomise 40–80 healthy UK participants aged 18–50 to receive oral challenge with one of two strains of S. Typhimurium belonging to the ST19 (strain 4/74) or ST313 (strain D23580) lineages. 4/74 is a global strain often associated with diarrhoeal illness predominantly in high-income settings, while D23580 is an archetypal strain representing invasive disease-causing isolates found in SSA. The primary objective is to determine the minimum infectious dose (colony-forming unit) required for 60%–75% of participants to develop clinical or microbiological features of systemic salmonellosis. Secondary endpoints are to describe and compare the clinical, microbiological and immunological responses following challenge. Dose escalation or de-escalation will be undertaken by continual-reassessment methodology and limited within prespecified safety thresholds. Exploratory objectives are to describe mechanisms of iNTS virulence, identify putative immune correlates of protection and describe host–pathogen interactions in response to infection.Ethics and dissemination Ethical approval has been obtained from the NHS Health Research Authority (London—Fulham Research Ethics Committee 21/PR/0051; IRAS Project ID 301659). The study findings will be disseminated in international peer-reviewed journals and presented at national/international stakeholder meetings. Study outcome summaries will be provided to both funders and participants.Trial registration number NCT05870150

BackgroundEffective negotiation in relationships is critical for successful long-duration space missions; inadequate conflict resolution has shown serious consequences. Less desirable forms of negotiation, including positional bargaining (eg, negotiating prices), can exacerbate conflicts. Traditional positional bargaining may work for simple, low-stakes transactions but does not prioritize ongoing relationships. High-stakes situations warrant interest-based negotiation, where parties with competing interests or goals collaborate in a mutually beneficial agreement. This is learnable but must be practiced. Refresher training during conflicts is important to prevent out-of-practice crew members from using less effective negotiation techniques. Training should be self-directed and not involve others because, on a space mission, the only other people available may be part of the conflict. ObjectiveWe aimed to develop and test an interactive module teaching principles and skills of interest-based negotiation in a way that users find acceptable, valuable for learning, and enjoyable. MethodsUsing a web-based, interactive-media approach, we scripted, filmed, and programmed an interest-based negotiation interactive training module. In the module, the program mentor introduces users to “The Circle of Value” approach to negotiation and highlights its key concepts through interactive scenarios requiring users to make selections at specific decision points. Each selection prompts feedback designed to reinforce a teaching point or highlight a particular negotiation technique. To evaluate the module, we sought populations experiencing isolation and confinement (an opportunistic design). This included 9 participants in isolated, confined environments in the Australian Antarctic Program and the Hawai'i Space Exploration Analog and Simulation Mars simulation, as well as a subset of people who self-identified as being isolated and confined during the COVID-19 pandemic. Feedback was collected from participants (n=54) through free-response answers and questionnaires with numerical scaling (0=strongly disagree to 4=strongly agree) at the end of the module. ResultsIn total, 51 of 54 (94%) participants found the activity valuable for learning about conflict management (identified by those who selected either “somewhat agree” or “strongly agree”), including 100% of participants in the isolated and confined environment subset (mode=3). In total, 79% (128/162) of participant responses indicated that the module was realistic (mode=3), including 85% (23/27) of responses from participants in isolated and confined environments (mode=3). Most participants felt that this would be particularly valuable for new team members in an isolated, confined environment (46/54, 85% of all participants, mode 4; 7/9, 78% of the isolated and confined environment subset, mode 3) as well as veterans. ConclusionsThis module offers a self-directed, consistent approach to interest-based negotiation training, which is well received by users. Although the data are limited due to the opportunistic study design, the module could be useful for individuals in isolated and confined environments and for anyone involved in high-stakes negotiations where sustaining relationships is essential.

Abstract Optical coherence tomography angiography (OCTA) is an emerging non-invasive technique for imaging the retinal vasculature. While there are many promising clinical applications for OCTA, determination of image quality remains a challenge. We developed a deep learning-based system using a ResNet152 neural network classifier, pretrained using ImageNet, to classify images of the superficial capillary plexus in 347 scans from 134 patients. Images were also manually graded by two independent graders as a ground truth for the supervised learning models. Because requirements for image quality may vary depending on the clinical or research setting, two models were trained—one to identify high-quality images and one to identify low-quality images. Our neural network models demonstrated outstanding area under the curve (AUC) metrics for both low quality image identification (AUC = 0.99, 95%CI 0.98–1.00, $$\kappa $$ κ = 0.90) and high quality image identification (AUC = 0.97, 95%CI 0.96–0.99, $$\kappa $$ κ = 0.81), significantly outperforming machine-reported signal strength (AUC = 0.82, 95%CI 0.77–0.86, $$\kappa $$ κ = 0.52 and AUC = 0.78, 95%CI 0.73–0.83, $$\kappa $$ κ = 0.27 respectively). Our study demonstrates that techniques from machine learning may be used to develop flexible and robust methods for quality control of OCTA images.

This study was designed to test the idea that the regulatory regions of human genes have evolved to be resistant to the effects of mutations in their primary function, the control of gene expression. It is proposed that the transcription factor/transcription factor binding site (TF/TFBS) pair having the greatest effect on control of a gene is the one with the highest abundance among the regulatory elements. Other pairs would have the same effect on gene expression and would predominate in the event of a mutation in the most abundant pair. It is expected that the overall regulatory design proposed here will be highly resistant to mutagenic change that would otherwise affect expression of the gene. The idea was tested beginning with a database of 42 human genes highly specific for expression in brain. For each gene, the five most abundant TF/TFBS pairs were identified and compared in their TFBS occupancy as measured by their ChIP-seq signal. A similar signal was observed and interpreted as evidence that the TF/TFBS pairs can substitute for one another. TF/TFBS pairs were also compared in their ability to substitute for one another in their effect on the level of gene expression. The study of brain specific genes was complemented with the same analysis performed with 31 human liver specific genes. Like the study of brain genes, the liver results supported the view that TF/TFBS pairs in liver specific genes can substitute for one another in the event of mutagenic damage. Finally, the TFBSs in the brain specific and liver specific gene populations were compared with each other with the goal of identifying any brain selective or liver selective TFBSs. Of the 89 TFBSs in the pooled population, 58 were found only in brain specific but not liver specific genes, 8 only in liver specific but not brain specific genes and 23 were found in both brain and liver specific genes. The results were interpreted to emphasize the large number of TFBS in brain specific genes.

Abstract Introduction The circadian circuit, a roughly 24 h molecular feedback loop, or clock, is conserved from bacteria to animals and allows for enhanced organismal survival by facilitating the anticipation of the day/night cycle. With circadian regulation reportedly impacting as high as 80% of protein coding genes in higher eukaryotes, the protein-based circadian clock broadly regulates physiology and behavior. Due to the extensive interconnection between the clock and other cellular systems, chronic disruption of these molecular rhythms leads to a decrease in organismal fitness as well as an increase of disease rates in humans. Importantly, recent research has demonstrated that proteins comprising the circadian clock network display a significant amount of intrinsic disorder. Main body In this work, we focus on the extent of intrinsic disorder in the circadian clock and its potential mechanistic role in circadian timing. We highlight the conservation of disorder by quantifying the extent of computationally-predicted protein disorder in the core clock of the key eukaryotic circadian model organisms Drosophila melanogaster, Neurospora crassa, and Mus musculus. We further examine previously published work, as well as feature novel experimental evidence, demonstrating that the core negative arm circadian period drivers FREQUENCY (Neurospora crassa) and PERIOD-2 (PER2) (Mus musculus), possess biochemical characteristics of intrinsically disordered proteins. Finally, we discuss the potential contributions of the inherent biophysical principals of intrinsically disordered proteins that may explain the vital mechanistic roles they play in the clock to drive their broad evolutionary conservation in circadian timekeeping. Conclusion The pervasive conservation of disorder amongst the clock in the crown eukaryotes suggests that disorder is essential for optimal circadian timing from fungi to animals, providing vital homeostatic cellular maintenance and coordinating organismal physiology across phylogenetic kingdoms. Video abstract Graphical abstract

Abstract Aedes aegypti is the primary vector for transmission of Dengue, Zika and chikungunya viruses. Previously it was shown that Dengue virus infection of the mosquito led to an in increased expression of the odorant binding protein 22 (AeOBP22) within the mosquito salivary gland and that siRNA mediated knockdown of AeOBP22 led to reduced mosquito feeding behaviors. Insect OBPs are implicated in the perception, storage and transport of chemosensory signaling molecules including air-borne odorants and pheromones. AeOBP22 is unusual as it is additionally expressed in multiple tissues, including the antenna, the male reproductive glands and is transferred to females during reproduction, indicating multiple roles in the mosquito life cycle. However, it is unclear what role it plays in these tissues and what ligands it interacts with. Here we present solution and X-ray crystallographic studies that indicate a potential role of AeOBP22 binding to fatty acids, and that the specificity for longer chain fatty acids is regulated by a conformational change in the C-terminal tail that leads to creation of an enlarged binding cavity that enhances binding affinity. This study sheds light onto the native ligands for AeOBP22 and provides insight into its potential functions in different tissues.

Arsenic is a carcinogenic groundwater contaminant that is toxic even at the parts-per-billion (ppb) level and its on-site determination remains challenging. Colorimetric test strips, though cheap and widely used, often fail to give reliable quantitative data. On the other hand, electrochemical detection is sensitive and accurate but considerably more expensive at the onset. Here, we present a study on arsenic detection in groundwater using a low-cost, open-source potentiostat based on Arduino technology. We tested different types of gold electrodes (screen-printed and microwire) with anodic stripping voltammetry (ASV), achieving low detection limits (0.7 μg L-1). In a study of arsenic contaminated groundwaters in Mexico, the microwire technique provides greater accuracy than test strips (reducing the median error from -50% to +2.9%) and greater precision (reducing uncertainties from ±25% to ±4.9%). Most importantly, the rate of false negatives versus the World Health Organisation’s 10 μg L-1 limit was reduced from 50% to 0% (N = 13 samples). Arsenic determination using open-source potentiostats may offer a low-cost option for research groups and NGOs wishing to perform arsenic analysis in-house, yielding superior quantitative data than the more widely used colorimetric test strips.

Circadian clocks in fungi and animals are driven by a functionally conserved transcription–translation feedback loop. In Neurospora crassa, negative feedback is executed by a complex of Frequency (FRQ), FRQ-interacting RNA helicase (FRH), and casein kinase I (CKI), which inhibits the activity of the clock’s positive arm, the White Collar Complex (WCC). Here, we show that the prd-2 (period-2) gene, whose mutation is characterized by recessive inheritance of a long 26 hr period phenotype, encodes an RNA-binding protein that stabilizes the ck-1a transcript, resulting in CKI protein levels sufficient for normal rhythmicity. Moreover, by examining the molecular basis for the short circadian period of upf-1prd-6 mutants, we uncovered a strong influence of the Nonsense-Mediated Decay pathway on CKI levels. The finding that circadian period defects in two classically derived Neurospora clock mutants each arise from disruption of ck-1a regulation is consistent with circadian period being exquisitely sensitive to levels of casein kinase I.

PURPOSE:Von Hippel-Lindau (VHL) disease is a hereditary disorder that can lead to ophthalmic manifestations, including retinal capillary hemangioma (RCH). The diagnosis of RCH is often guided by wide-field fluorescein angiography. In some cases, optical coherence tomography angiography (OCT-A) serves as a non-invasive alternative to FA. Herein, we used OCT-A to examine the macular microvasculature in patients with VHL disease. SUBJECTS:Subjects were selected from patients with a diagnosis of VHL. The control group included eyes without retinal diagnosis from patients with an episode of unilateral retinal detachment or trauma and age ≤ 50 years old. METHODS:Subjects were scanned on the Optovue RTVue-XR device to acquire 3mm x 3mm OCT-A images of the superficial (SCP) and deep capillary plexus (DCP). SCP and DCP vessel density (VD) were calculated after the images were binarized. Furthermore, for subjects with RCH, each OCT-A image was divided equally into four quadrants. SCP and DCP VD of quadrants with RCH were compared to those without RCH. T-tests were performed for statistical analysis. RESULTS:67 eyes with a history of VHL disease were included as study subjects, while 16 eyes were included as controls. Significant increases in VD were found in patients with VHL disease for both the SCP (p = 0.0441) and DCP (p = 0.0344). When comparing quadrants with associated RCH development to those without, we found no significant difference in SCP VD (p = 0.160) or DCP VD (p = 0.484). CONCLUSIONS:OCT-A can detect changes in the retinal microvasculature in the macula of patients with VHL disease. OCT-A imaging may be an additional tool for screening and early detection of patients at risk of developing ocular complications of VHL disease. Future studies should explore subtle progression on OCT-A associated with the pathogenesis and development of RCH, particularly with larger scan patterns.

Abstract Background The ST313 sequence type of Salmonella Typhimurium causes invasive non-typhoidal salmonellosis and was thought to be confined to sub-Saharan Africa. Two distinct phylogenetic lineages of African ST313 have been identified. Methods We analysed the whole genome sequences of S. Typhimurium isolates from UK patients that were generated following the introduction of routine whole-genome sequencing (WGS) of Salmonella enterica by Public Health England in 2014. Results We found that 2.7% (84/3147) of S. Typhimurium from patients in England and Wales were ST313 and were associated with gastrointestinal infection. Phylogenetic analysis revealed novel diversity of ST313 that distinguished UK-linked gastrointestinal isolates from African-associated extra-intestinal isolates. The majority of genome degradation of African ST313 lineage 2 was conserved in the UK-ST313, but the African lineages carried a characteristic prophage and antibiotic resistance gene repertoire. These findings suggest that a strong selection pressure exists for certain horizontally acquired genetic elements in the African setting. One UK-isolated lineage 2 strain that probably originated in Kenya carried a chromosomally located bla CTX-M-15, demonstrating the continual evolution of this sequence type in Africa in response to widespread antibiotic usage. Conclusions The discovery of ST313 isolates responsible for gastroenteritis in the UK reveals new diversity in this important sequence type. This study highlights the power of routine WGS by public health agencies to make epidemiologically significant deductions that would be missed by conventional microbiological methods. We speculate that the niche specialisation of sub-Saharan African ST313 lineages is driven in part by the acquisition of accessory genome elements.

Abstract Glial scar is a significant barrier to neural implant function. Micromotion between the implant and tissue is suspected to be a key driver of glial scar formation around neural implants. This study explores the ability of soft hydrogel coatings to modulate glial scar formation by reducing local strain. PEG hydrogels with controllable thickness and elastic moduli were formed on the surface of neural probes. These coatings significantly reduced the local strain resulting from micromotion around the implants. Coated implants were found to significantly reduce scarring in vivo, compared to hard implants of identical diameter. Increasing implant diameter was found to significantly increase scarring for glass implants, as well as increase local BBB permeability, increase macrophage activation, and decrease the local neural density. These results highlight the tradeoff in mechanical benefit with the size effects from increasing the overall diameter following the addition of a hydrogel coating. This study emphasizes the importance of both mechanical and geometric factors of neural implants on chronic timescales.

BackgroundAs a result of decades of effort by many investigators we now have an advanced level of understanding about several molecular systems involved in the control of gene expression. Examples include CpG islands, promoters, mRNA splicing and epigenetic signals. It is less clear, however, how such systems work together to integrate the functions of a living organism. Here I describe the results of a study to test the idea that a contribution might be made by focusing on genes specifically expressed in a particular tissue, the human testis.Experimental designA database of 239 testis-specific genes was accumulated and each was examined for the presence of features relevant to control of gene expression. These include: (1) the presence of a promoter, (2) the presence of a CpG island (CGI) within the promoter, (3) the presence in the promoter of a transcription factor binding site near the transcription start site, (4) the level of gene expression, and (5) the above features in genes of testis-specific cell types such as spermatocyte and spermatid that differ in their extent of differentiation.ResultsOf the 107 database genes with an annotated promoter, 56 were found to have one or more transcription factor binding sites near the transcription start site. Three of the binding sites observed, Pax-5, AP-2αA and GRα, stand out in abundance suggesting they may be involved in testis-specific gene expression. Compared to less differentiated testis-specific cells, genes of more differentiated cells were found to be (1) more likely to lack a CGI, (2) more likely to lack introns and (3) higher in expression level. The results suggest genes of more differentiated cells have a reduced need for CGI-based regulatory repression, reduced usage of gene splicing and a smaller set of expressed proteins.

Although drusen have long been acknowledged as a primary hallmark of dry age-related macular degeneration (AMD) their role in the disease remains unclear. We hypothesize that drusen accumulation increases the barrier to metabolite transport ultimately resulting in photoreceptor cell death. To investigate this hypothesis, a computational model was developed to evaluate steady-state oxygen distribution in the retina. Optical coherence tomography images from fifteen AMD patients and six control subjects were segmented and translated into 3D in silico representations of retinal morphology. A finite element model was then used to determine the steady-state oxygen distribution throughout the retina for both generic and patient-specific retinal morphology. Oxygen levels were compared to the change in retinal thickness at a later time point to observe possible correlations. The generic finite element model of oxygen concentration in the retina agreed closely with both experimental measurements from literature and clinical observations, including the minimal pathological drusen size identified by AREDS (64 μm). Modeling oxygen distribution in the outer retina of AMD patients showed a substantially stronger correlation between hypoxia and future retinal thinning (Pearson correlation coefficient, r = 0.2162) than between drusen height and retinal thinning (r = 0.0303) indicating the potential value of this physiology-based approach. This study presents proof-of-concept for the potential utility of finite element modeling in evaluating retinal health and also suggests a potential link between transport and AMD pathogenesis. This strategy may prove useful as a prognostic tool for predicting the clinical risk of AMD progression.

Objective: To compare the effects of 3% hypertonic saline (HS) and 0.9% normal saline with nebulized 0.9% normal saline with salbutamol in patients of acute viral bronchiolitis. Materials and Methods: Participants were divided into three groups, that is, 3% HS group, 0.9% normal saline group and 0.9% saline with salbutamol group. Four doses at interval of 6 h were given daily until discharge. Average CS score and length of hospital stay were compared. One-way analysis of variance paired t-test and Chi-square test were utilized for statistical analysis. Results: The mean ages of the patients in three groups were 6.03 ± 3.71, 5.69 ± 3.34 and 5.48 ± 3.35 respectively. The 3rd day CS scores for all the groups were 1.0 ± 1.1, 1.9 ± 1.1 and 3.3 ± 0.5 respectively (P = 0.000). The average length of hospital stay was 3.4 ± 1.7, 3.7 ± 1.9 and 4.9 ± 1.4 days respectively (P = 0.001). Conclusion: The present study concludes that 3% HS nebulization (without additional bronchodilators) is an effective and safe treatment for nonasthmatic, moderately ill patients of acute bronchiolitis. The economic benefit of this comparably priced modality of treatment can be enormous in terms of hospital costs with parents returning to work sooner.

BACKGROUND:DNA sequence elements in the core promoter can play a central role in regulation of gene expression. Core elements (e.g. INR and TATA box) are located within ~50bp of the transcription start site and both upstream and downstream elements are known. Although all can affect the level of gene expression, their mechanism of action has yet to be fully defined. The studies described here are focused on two core promoter elements, INR and BRE, in the human genome. The locations of the two elements were determined in a large number of human promoters and the results were interpreted in terms of overall promoter function. RESULTS:A total of 13,406 promoters were collected from the reference version of the human genome and found to contain 62,891 INR sequences and 32,290 BRE. An INR sequence was found in the core region of 1231 (9.2%) promoters and a BRE in 2592 (19.3%); 121 promoters (0.9%) have both INR and BRE elements. Counts support the view that most human promoters lack an INR or BRE element in the core promoter. Further analysis was carried out with the aligned aggregate of promoters from each chromosome. The results showed distinct INR distributions in separate chromosome groups indicating a degree of chromosome specificity to the way core promoter elements are deployed in the genome. The rare promoters with both INR and BRE elements were found to be enriched among the genes with divergent transcription. Enrichment raises the possibility that core promoter elements can have a function in chromosome organization as well as in initiation of transcription.

Longitudinal investigation of the neural correlates of reward processing in depression may represent an important step in defining effective biomarkers for antidepressant treatment outcome prediction, but the reliability of reward-related activation is not well understood. Thirty-seven healthy control participants were scanned using fMRI while performing a reward-related guessing task on two occasions, approximately one week apart. Two main contrasts were examined: right ventral striatum (VS) activation fMRI BOLD signal related to signed prediction errors (PE) and reward expectancy (RE). We also examined bilateral visual cortex activation coupled to outcome anticipation. Significant VS PE-related activity was observed at the first testing session, but at the second testing session, VS PE-related activation was significantly reduced. Conversely, significant VS RE-related activity was observed at time 2 but not time 1. Increases in VS RE-related activity from time 1 to time 2 were significantly associated with decreases in VS PE-related activity from time 1 to time 2 across participants. Intraclass correlations (ICCs) in VS were very low. By contrast, visual cortex activation had much larger ICCs, particularly in individuals with high quality data. Dynamic changes in brain activation are widely predicted, and failure to account for these changes could lead to inaccurate evaluations of the reliability of functional MRI signals. Conventional measures of reliability cannot distinguish between changes specified by algorithmic models of neural function and noisy signal. Here, we provide evidence for the former possibility: reward-related VS activations follow the pattern predicted by temporal difference models of reward learning but have low ICCs.

In October 2001, the first inhalational anthrax case in the United States since 1976 was identified in a media company worker in Florida. A national investigation was initiated to identify additional cases and determine possible exposures to Bacillus anthracis. Surveillance was enhanced through health-care facilities, laboratories, and other means to identify cases, which were defined as clinically compatible illness with laboratory-confirmed B. anthracis infection. From October 4 to November 20, 2001, 22 cases of anthrax (11 inhalational, 11 cutaneous) were identified; 5 of the inhalational cases were fatal. Twenty (91%) case-patients were either mail handlers or were exposed to worksites where contaminated mail was processed or received. B. anthracis isolates from four powder-containing envelopes, 17 specimens from patients, and 106 environmental samples were indistinguishable by molecular subtyping. Illness and death occurred not only at targeted worksites, but also along the path of mail and in other settings. Continued vigilance for cases is needed among health-care providers and members of the public health and law enforcement communities.

The biological attacks with powders containing Bacillus anthracis sent through the mail during September and October 2001 led to unprecedented public health and law enforcement investigations, which involved thousands of investigators from federal, state, and local agencies. Following recognition of the first cases of anthrax in Florida in early October 2001, investigators from Centers for Disease Control and Prevention (CDC) and the Federal Bureau of Investigation (FBI) were mobilized to assist investigators from state and local public health and law enforcement agencies. Although public health and criminal investigations have been conducted in concert in the past, the response to the anthrax attacks required close collaboration because of the immediate and ongoing threat to public safety. We describe the collaborations between CDC and FBI during the investigation of the 2001 anthrax attacks and highlight the challenges and successes of public health and law enforcement collaborations in general.

After public notification of confirmed cases of bioterrorism-related anthrax, the Centers for Disease Control and Prevention’s Emergency Operations Center responded to 11,063 bioterrorism-related telephone calls from October 8 to November 11, 2001. Most calls were inquiries from the public about anthrax vaccines (58.4%), requests for general information on bioterrorism prevention (14.8%), and use of personal protective equipment (12.0%); 882 telephone calls (8.0%) were referred to the state liaison team for follow-up investigation. Of these, 226 (25.6%) included reports of either illness clinically confirmed to be compatible with anthrax or direct exposure to an environment known to be contaminated with Bacillus anthracis. The remaining 656 (74.4%) included no confirmed illness but reported exposures to “suspicious” packages or substances or the receipt of mail through a contaminated facility. Emergency response staff must handle high call volumes following suspected or actual bioterrorist attacks. Standardized health communication protocols that address contact with unknown substances, handling of suspicious mail, and clinical evaluation of suspected cases would allow more efficient follow-up investigations of clinically compatible cases in high-risk groups.

Functional Magnetic Resonance Imagine (fMRI) is an important assessment tool in longitudinal studies of mental illness and its treatment. Understanding the psychometric properties of fMRI-based metrics, and the factors that influence them, will be critical for properly interpreting the results of these efforts. The current study examined whether the choice among alternative model specifications affects estimates of test-retest reliability in key emotion processing regions across a 6-month interval. Subjects (N = 46) performed an emotional-faces paradigm during fMRI in which neutral faces dynamically morphed into one of four emotional faces. Median voxelwise intraclass correlation coefficients (mvICCs) were calculated to examine stability over time in regions showing task-related activity as well as in bilateral amygdala. Four modeling choices were evaluated: a default model that used the canonical hemodynamic response function (HRF), a flexible HRF model that included additional basis functions, a modified CompCor (mCompCor) model that added corrections for physiological noise in the global signal, and a final model that combined the flexible HRF and mCompCor models. Model residuals were examined to determine the degree to which each pipeline met modeling assumptions. Results indicated that the choice of modeling approaches impacts both the degree to which model assumptions are met and estimates of test-retest reliability. ICC estimates in the visual cortex increased from poor (mvICC = 0.31) in the default pipeline to fair (mvICC = 0.45) in the full alternative pipeline - an increase of 45%. In nearly all tests, the models with the fewest assumption violations generated the highest ICC estimates. Implications for longitudinal treatment studies that utilize fMRI are discussed.

Group A streptococcus (GAS, Streptococcus pyogenes) is a strict human pathogen that causes severe, invasive diseases. GAS does not produce catalase, but has an ability to resist killing by reactive oxygen species (ROS) through novel mechanisms. The peroxide response regulator (PerR), a member of ferric uptake regulator (Fur) family, plays a key role for GAS to cope with oxidative stress by regulating the expression of multiple genes. Our previous studies have found that expression of an iron-binding protein, Dpr, is under the direct control of PerR. To elucidate the molecular interactions of PerR with its cognate promoter, we have carried out structural studies on PerR and PerR-DNA complex. By combining crystallography and small-angle X-ray scattering (SAXS), we confirmed that the determined PerR crystal structure reflects its conformation in solution. Through mutagenesis and biochemical analysis, we have identified DNA-binding residues suggesting that PerR binds to the dpr promoter at the per box through a winged-helix motif. Furthermore, we have performed SAXS analysis and resolved the molecular architecture of PerR-DNA complex, in which two 30 bp DNA fragments wrap around two PerR homodimers by interacting with the adjacent positively-charged winged-helix motifs. Overall, we provide structural insights into molecular recognition of DNA by PerR and define the hollow structural arrangement of PerR-30bpDNA complex, which displays a unique topology distinct from currently proposed DNA-binding models for Fur family regulators.

Advances in personalized medicine require the identification of variables that predict differential response to treatments as well as the development and refinement of methods to transform predictive information into actionable recommendations.To illustrate and test a new method for integrating predictive information to aid in treatment selection, using data from a randomized treatment comparison.Data from a trial of antidepressant medications (N = 104) versus cognitive behavioral therapy (N = 50) for Major Depressive Disorder were used to produce predictions of post-treatment scores on the Hamilton Rating Scale for Depression (HRSD) in each of the two treatments for each of the 154 patients. The patient's own data were not used in the models that yielded these predictions. Five pre-randomization variables that predicted differential response (marital status, employment status, life events, comorbid personality disorder, and prior medication trials) were included in regression models, permitting the calculation of each patient's Personalized Advantage Index (PAI), in HRSD units.For 60% of the sample a clinically meaningful advantage (PAI≥3) was predicted for one of the treatments, relative to the other. When these patients were divided into those randomly assigned to their "Optimal" treatment versus those assigned to their "Non-optimal" treatment, outcomes in the former group were superior (d = 0.58, 95% CI .17-1.01).This approach to treatment selection, implemented in the context of two equally effective treatments, yielded effects that, if obtained prospectively, would rival those routinely observed in comparisons of active versus control treatments.

To estimate the effectiveness of pneumococcal polysaccharide vaccine, we serotyped isolates submitted to the Pneumococcal Sentinel Surveillance System from 1984 to 1996 from 48 vaccinated and 125 unvaccinated children 2 to 5 years of age. Effectiveness against invasive disease caused by serotypes included in the vaccine was 63%. Effectiveness against serotypes in the polysaccharide vaccine but not in a proposed seven-valent protein conjugate vaccine was 94%.

The filamentous fungi are an ecologically important group of organisms which also have important industrial applications but devastating effects as pathogens and agents of food spoilage. Protein kinases have been implicated in the regulation of virtually all biological processes but how they regulate filamentous fungal specific processes is not understood. The filamentous fungus Aspergillus nidulans has long been utilized as a powerful molecular genetic system and recent technical advances have made systematic approaches to study large gene sets possible. To enhance A. nidulans functional genomics we have created gene deletion constructs for 9851 genes representing 93.3% of the encoding genome. To illustrate the utility of these constructs, and advance the understanding of fungal kinases, we have systematically generated deletion strains for 128 A. nidulans kinases including expanded groups of 15 histidine kinases, 7 SRPK (serine-arginine protein kinases) kinases and an interesting group of 11 filamentous fungal specific kinases. We defined the terminal phenotype of 23 of the 25 essential kinases by heterokaryon rescue and identified phenotypes for 43 of the 103 non-essential kinases. Uncovered phenotypes ranged from almost no growth for a small number of essential kinases implicated in processes such as ribosomal biosynthesis, to conditional defects in response to cellular stresses. The data provide experimental evidence that previously uncharacterized kinases function in the septation initiation network, the cell wall integrity and the morphogenesis Orb6 kinase signaling pathways, as well as in pathways regulating vesicular trafficking, sexual development and secondary metabolism. Finally, we identify ChkC as a third effector kinase functioning in the cellular response to genotoxic stress. The identification of many previously unknown functions for kinases through the functional analysis of the A. nidulans kinome illustrates the utility of the A. nidulans gene deletion constructs.

Pneumococcal pneumonia causes significant morbidity and mortality among adults. Given limitations of diagnostic tests for non-bacteremic pneumococcal pneumonia, most studies report the incidence of bacteremic or invasive pneumococcal disease (IPD), and thus, grossly underestimate the pneumococcal pneumonia burden. We aimed to develop a conceptual and quantitative strategy to estimate the non-bacteremic disease burden among adults with community-acquired pneumonia (CAP) using systematic study methods and the availability of a urine antigen assay.We performed a systematic literature review of studies providing information on the relative yield of various diagnostic assays (BinaxNOW® S. pneumoniae urine antigen test (UAT) with blood and/or sputum culture) in diagnosing pneumococcal pneumonia. We estimated the proportion of pneumococcal pneumonia that is bacteremic, the proportion of CAP attributable to pneumococcus, and the additional contribution of the Binax UAT beyond conventional diagnostic techniques, using random effects meta-analytic methods and bootstrapping. We included 35 studies in the analysis, predominantly from developed countries. The estimated proportion of pneumococcal pneumonia that is bacteremic was 24.8% (95% CI: 21.3%, 28.9%). The estimated proportion of CAP attributable to pneumococcus was 27.3% (95% CI: 23.9%, 31.1%). The Binax UAT diagnosed an additional 11.4% (95% CI: 9.6, 13.6%) of CAP beyond conventional techniques. We were limited by the fact that not all patients underwent all diagnostic tests and by the sensitivity and specificity of the diagnostic tests themselves. We address these resulting biases and provide a range of plausible values in order to estimate the burden of pneumococcal pneumonia among adults.Estimating the adult burden of pneumococcal disease from bacteremic pneumococcal pneumonia data alone significantly underestimates the true burden of disease in adults. For every case of bacteremic pneumococcal pneumonia, we estimate that there are at least 3 additional cases of non-bacteremic pneumococcal pneumonia.

RNA-dependent RNA polymerases play a vital role in the growth of RNA viruses where they are responsible for genome replication, but do so with rather low fidelity that allows for the rapid adaptation to different host cell environments. These polymerases are also a target for antiviral drug development. However, both drug discovery efforts and our understanding of fidelity determinants have been hampered by a lack of detailed structural information about functional polymerase-RNA complexes and the structural changes that take place during the elongation cycle. Many of the molecular details associated with nucleotide selection and catalysis were revealed in our recent structure of the poliovirus polymerase-RNA complex solved by first purifying and then crystallizing stalled elongation complexes. In the work presented here we extend that basic methodology to determine nine new structures of poliovirus, coxsackievirus, and rhinovirus elongation complexes at 2.2-2.9 Å resolution. The structures highlight conserved features of picornaviral polymerases and the interactions they make with the template and product RNA strands, including a tight grip on eight basepairs of the nascent duplex, a fully pre-positioned templating nucleotide, and a conserved binding pocket for the +2 position template strand base. At the active site we see a pre-bound magnesium ion and there is conservation of a non-standard backbone conformation of the template strand in an interaction that may aid in triggering RNA translocation via contact with the conserved polymerase motif B. Moreover, by engineering plasticity into RNA-RNA contacts, we obtain crystal forms that are capable of multiple rounds of in-crystal catalysis and RNA translocation. Together, the data demonstrate that engineering flexible RNA contacts to promote crystal lattice formation is a versatile platform that can be used to solve the structures of viral RdRP elongation complexes and their catalytic cycle intermediates.

Chlamydia pneumoniae commonly causes respiratory tract infections in children, and epidemiological investigations strongly link infection to the pathogenesis of asthma. The immune system in early life is immature and may not respond appropriately to pathogens. Toll-like receptor (TLR)2 and 4 are regarded as the primary pattern recognition receptors that sense bacteria, however their contribution to innate and adaptive immunity in early life remains poorly defined. We investigated the role of TLR2 and 4 in the induction of immune responses to Chlamydia muridarum respiratory infection, in neonatal wild-type (Wt) or TLR2-deficient ((-/-)), 4(-/-) or 2/4(-/-) BALB/c mice. Wt mice had moderate disease and infection. TLR2(-/-) mice had more severe disease and more intense and prolonged infection compared to other groups. TLR4(-/-) mice were asymptomatic. TLR2/4(-/-) mice had severe early disease and persistent infection, which resolved thereafter consistent with the absence of symptoms in TLR4(-/-) mice. Wt mice mounted robust innate and adaptive responses with an influx of natural killer (NK) cells, neutrophils, myeloid (mDCs) and plasmacytoid (pDCs) dendritic cells, and activated CD4(+) and CD8(+) T-cells into the lungs. Wt mice also had effective production of interferon (IFN)γ in the lymph nodes and lung, and proliferation of lymph node T-cells. TLR2(-/-) mice had more intense and persistent innate (particularly neutrophil) and adaptive cell responses and IL-17 expression in the lung, however IFNγ responses and T-cell proliferation were reduced. TLR2/4(-/-) mice had reduced innate and adaptive responses. Most importantly, neutrophil phagocytosis was impaired in the absence of TLR2. Thus, TLR2 expression, particularly on neutrophils, is required for effective control of Chlamydia respiratory infection in early life. Loss of control of infection leads to enhanced but ineffective TLR4-mediated inflammatory responses that prolong disease symptoms. This indicates that TLR2 agonists may be beneficial in the treatment of early life Chlamydia infections and associated diseases.

We identified an amino acid change (p.G92E) in the Bone Morphogenetic Protein antagonist NOGGIN in a 22-month-old boy who presented with a unilateral brachydactyly type B phenotype. Brachydactyly type B is a skeletal malformation that has been associated with increased Bone Morphogenetic Protein pathway activation in other patients. Previously, the amino acid change p.G92E in NOGGIN was described as causing fibrodysplasia ossificans progressiva, a rare genetic disorder characterized by limb malformations and progressive heterotopic bone formation in soft tissues that, like Brachydactyly type B, is caused by increased activation of Bone Morphogenetic Protein signaling. To determine whether G92E-NOGGIN shows impaired antagonism that could lead to increased Bone Morphogenetic Protein signaling, we performed functional assays to evaluate inhibition of BMP signaling. Interestingly, wt-NOGGIN shows different inhibition efficacies towards various Bone Morphogenetic Proteins that are known to be essential in limb development. However, comparing the biological activity of G92E-NOGGIN with wt-NOGGIN, we observed that G92E-NOGGIN inhibits activation of bone morphogenetic protein signaling with equal efficiency as wt-NOGGIN, supporting that G92E-NOGGIN does not cause pathological effects. Genetic testing of the child's parents revealed the same amino acid change in the healthy father, further supporting that p.G92E is a neutral amino acid substitution in NOGGIN. We conclude that p.G92E represents a rare polymorphism of the NOGGIN gene-- causing neither brachydactyly nor fibrodysplasia ossificans progressiva. This study highlights that a given genetic variation should not be considered pathogenic unless supported by functional analyses.

BACKGROUND: Viral and bacterial respiratory tract infections in early-life are linked to the development of allergic airway inflammation and asthma. However, the mechanisms involved are not well understood. We have previously shown that neonatal and infant, but not adult, chlamydial lung infections in mice permanently alter inflammatory phenotype and physiology to increase the severity of allergic airway disease by increasing lung interleukin (IL)-13 expression, mucus hyper-secretion and airway hyper-responsiveness. This occurred through different mechanisms with infection at different ages. Neonatal infection suppressed inflammatory responses but enhanced systemic dendritic cell:T-cell IL-13 release and induced permanent alterations in lung structure (i.e., increased the size of alveoli). Infant infection enhanced inflammatory responses but had no effect on lung structure. Here we investigated the role of hematopoietic cells in these processes using bone marrow chimera studies. METHODOLOGY/PRINCIPAL FINDINGS: Neonatal (

In comparison to the comprehensive analyses performed on virulence gene expression, regulation and action, the intracellular metabolism of Salmonella during infection is a relatively under-studied area. We investigated the role of the tricarboxylic acid (TCA) cycle in the intracellular replication of Salmonella Typhimurium in resting and activated macrophages, epithelial cells, and during infection of mice.We constructed deletion mutations of 5 TCA cycle genes in S. Typhimurium including gltA, mdh, sdhCDAB, sucAB, and sucCD. We found that the mutants exhibited increased net intracellular replication in resting and activated murine macrophages compared to the wild-type. In contrast, an epithelial cell infection model showed that the S. Typhimurium ΔsucCD and ΔgltA strains had reduced net intracellular replication compared to the wild-type. The glyoxylate shunt was not responsible for the net increased replication of the TCA cycle mutants within resting macrophages. We also confirmed that, in a murine infection model, the S. Typhimurium ΔsucAB and ΔsucCD strains are attenuated for virulence.Our results suggest that disruption of the TCA cycle increases the ability of S. Typhimurium to survive within resting and activated murine macrophages. In contrast, epithelial cells are non-phagocytic cells and unlike macrophages cannot mount an oxidative and nitrosative defence response against pathogens; our results show that in HeLa cells the S. Typhimurium TCA cycle mutant strains show reduced or no change in intracellular levels compared to the wild-type. The attenuation of the S. Typhimurium ΔsucAB and ΔsucCD mutants in mice, compared to their increased net intracellular replication in resting and activated macrophages suggest that Salmonella may encounter environments within the host where a complete TCA cycle is advantageous.

Mechanisms that underlie oscillatory transcriptional activity of nuclear receptors (NRs) are incompletely understood. Evidence exists for rapid, cyclic recruitment of coregulatory complexes upon activation of nuclear receptors. RIP140 is a NR coregulator that represses the transactivation of agonist-bound nuclear receptors. Previously, we showed that RIP140 is inducible by all-trans retinoic acid (RA) and mediates limiting, negative-feedback regulation of retinoid signaling.Here we report that in the continued presence of RA, long-paced oscillations of retinoic acid receptor (RAR) activity occur with a period ranging from 24 to 35 hours. Endogenous expression of RIP140 and other RA-target genes also oscillate in the presence of RA. Cyclic retinoid receptor transactivation is ablated by constitutive overexpression of RIP140. Further, depletion of RIP140 disrupts cyclic expression of the RA target gene HOXA5. Evidence is provided that RIP140 may limit RAR signaling in a selective, non-redundant manner in contrast to the classic NR coregulators NCoR1 and SRC1 that are not RA-inducible, do not cycle, and may be partially redundant in limiting RAR activity. Finally, evidence is provided that RIP140 can repress and be induced by other nuclear receptors in a manner that suggests potential participation in other NR oscillations.We provide evidence for novel, long-paced oscillatory retinoid receptor activity and hypothesize that this may be paced in part, by RIP140. Oscillatory NR activity may be involved in mediating hormone actions of physiological and pathological importance.

This study investigates aspects of masculinity that may relate to African American men's health-related attitudes and behaviors. Two hundred and eight men completed measures of traditional masculinity ideology and health-related attitudes and behaviors. Results indicated that after accounting for participants' age, education, income, and employment status, traditional masculinity norms of self-reliance and aggression were associated with behaviors conducive to personal wellness and certain health-related psychological tendencies. Restrictive emotionality was associated with anxiety about one's health and the belief that one's health status is outside of one's personal control. Implications of the findings for the health-related attitudes and behaviors of African American men are discussed.

Caffeine, commonly found in coffee and tea, affects various aspects of eye health as it blocks adenosine receptors, impacting tear production, intraocular pressure, macular perfusion, and choroidal thickness. However, its connection with eye conditions like glaucoma and cataracts remains uncertain due to conflicting research findings. Some studies suggest potential benefits for cataracts, while others warn against frequent caffeine intake in glaucoma and surgical scenarios due to possible increases in intraocular pressure. Conflicting evidence also exists regarding its effects on dry eye, macular degeneration, myopia/hyperopia, diabetic retinopathy, retinopathy of prematurity, and central serous retinopathy. Caffeine does not seem to be a risk factor for dry eye, although studies have shown that caffeine may offer protection against wet age-related macular degeneration, and the metabolite 7-methylxanthine could be a more promising treatment for myopia. Moreover, caffeine can potentially cause tremors and might hinder surgical performance, especially in less experienced surgeons. Recommendations from experts vary, highlighting the need for further research to fully understand how caffeine affects the eye. Individuals genetically predisposed to glaucoma should be cautious due to the possibility of clinically significant elevations in intraocular pressure with caffeine consumption. For delicate procedures like microsurgery, where tremors can be detrimental, caution should be exercised with caffeine. This review underscores the importance of additional studies to provide clearer insights and prudent recommendations regarding caffeine’s impact on eye health.

Abstract This study aimed to investigate the association between non-traditional lipid profiles and the risk of 1-year vascular events in patients who were already using statins before stroke and had admission LDL-C

Abstract To investigate the variability in subthreshold laser treatment plans for patients with diabetic macular edema or central serous chorioretinopathy. Diagnostic images from 20 patients were utilized, and 25 retina specialists generated subthreshold treatment plans along with a self-rated experience level. Evaluation comprised of i) Area Variability (AV): quantifies the consensus regarding the covered area and is the difference between the areas planned by 75% and 25% of the participants ii) mean Centroid Distance (CD): level of agreement on the localization of a treatment. Subgroup analysis investigated the impact of participants’ experience levels, utilizing the Mann-Whitney-Wilcoxon test. The predominant plan style is a targeted treatment approach (92%) and avoidance of subfoveal region (89%). Mean CD is 71.0 ± 37.5 pixels (≈half disc diameter) and mean AV is 9.8%±8.9%. A slight difference is observed between the 50-75% areas, but a notable distinction exists between the 25-50% areas. Subgroup analysis revealed CD and AV value of 75.9 pixels and 24% in the lower experience level group as opposed to 55.9 pixels and 8.6% in the higher experience level group. There is significant variability in treatment planning which reduces with increased experience of retina specialists. While consensus is observed around focal points, differences in the surrounding extents persist.