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Dipeptidyl aminopeptidases (DPAPs) are cysteine proteases that cleave dipeptides from the N-terminus of protein substrates and have been shown to play important roles in many pathologies including parasitic diseases such as malaria, toxoplasmosis and Chagas's disease. Inhibitors of the mammalian homologue cathepsin C have been used in clinical trials as potential drugs to treat chronic inflammatory disorders, thus proving that these enzymes are druggable. In Plasmodium species, DPAPs play important functions at different stages of parasite development, thus making them potential antimalarial targets. Most DPAP inhibitors developed to date are peptide-based or peptidomimetic competitive inhibitors. Here, we used a high throughput screening approach to identify novel inhibitor scaffolds that block the activity of Plasmodium falciparum DPAP1. Most of the hits identified in this screen also inhibit Plasmodium falciparum DPAP3, cathepsin C, and to a lesser extent other malarial clan CA proteases, indicating that these might be general DPAP inhibitors. Interestingly, our mechanism of inhibition studies indicate that most hits are allosteric inhibitors, which opens a completely new strategy to inhibit these enzymes, study their biological function, and potentially develop new inhibitors as starting points for drug development.

Parkinson's disease (PD) is a common neurodegenerative disorder which is mostly sporadic but familial-linked PD (FPD) cases have also been found. The first reported gene mutation that linked to PD is α-synuclein (α-syn). Studies have shown that mutations, increased expression or abnormal processing of α-syn can contribute to PD, but it is believed that multiple mechanisms are involved. One of the contributing factors is post-translational modification (PTM), such as phosphorylation of α-syn at serine 129 by G-protein-coupled receptor kinases (GRKs) and casein kinase 2α (CK2α). Another known important contributing factor to PD pathogenesis is oxidative and nitrosative stress. In this study, we found that GRK6 and CK2α can be S-nitrosylated by nitric oxide (NO) both in vitro and in vivo. S-nitrosylation of GRK6 and CK2α enhanced their kinase activity towards the phosphorylation of α-syn at S129. In an A53T α-syn transgenic mouse model of PD, we found that increased GRK6 and CK2α S-nitrosylation were observed in an age dependent manner and it was associated with an increased level of pSer129 α-syn. Treatment of A53T α-syn transgenic mice with Nω-Nitro-L-arginine (L-NNA) significantly reduced the S-nitrosylation of GRK6 and CK2α in the brain. Finally, deletion of neuronal nitric oxide synthase (nNOS) in A53T α-syn transgenic mice reduced the levels of pSer129 α-syn and α-syn in an age dependent manner. Our results provide a novel mechanism of how NO through S-nitrosylation of GRK6 and CK2α can enhance the phosphorylation of pSer129 α-syn in an animal model of PD.

[This corrects the article DOI: 10.1371/journal.pone.0232019.].

Dipeptidyl aminopeptidases (DPAPs) are cysteine proteases that cleave dipeptides from the N-terminus of protein substrates and have been shown to play important roles in many pathologies including parasitic diseases such as malaria, toxoplasmosis and Chagas's disease. Inhibitors of the mammalian homologue cathepsin C have been used in clinical trials as potential drugs to treat chronic inflammatory disorders, thus proving that these enzymes are druggable. In Plasmodium species, DPAPs play important functions at different stages of parasite development, thus making them potential antimalarial targets. Most DPAP inhibitors developed to date are peptide-based or peptidomimetic competitive inhibitors. Here, we used a high throughput screening approach to identify novel inhibitor scaffolds that block the activity of Plasmodium falciparum DPAP1. Most of the hits identified in this screen also inhibit Plasmodium falciparum DPAP3, cathepsin C, and to a lesser extent other malarial clan CA proteases, indicating that these might be general DPAP inhibitors. Interestingly, our mechanism of inhibition studies indicate that most hits are allosteric inhibitors, which opens a completely new strategy to inhibit these enzymes, study their biological function, and potentially develop new inhibitors as starting points for drug development.

Auxinic herbicides (e.g. dicamba) are extensively used in agriculture to selectively control broadleaf weeds. Although cultivated species of Brassicaceae (e.g. Canola) are susceptible to auxinic herbicides, some biotypes of Sinapis arvensis (wild mustard) were found dicamba resistant in Canada. In this research, dicamba tolerance from wild mustard was introgressed into canola through embryo rescue followed by conventional breeding. Intergeneric hybrids between S. arvensis (2n = 18) and B. napus (2n = 38) were produced through embryo rescue. Embryo formation and hybrid plant regeneration was achieved. Transfer of dicamba tolerance from S. arvensis into the hybrid plants was determined by molecular analysis and at the whole plant level. Dicamba tolerance was introgressed into B. napus by backcrossing for seven generations. Homozygous dicamba-tolerant B. napus lines were identified. The ploidy of the hybrid progeny was assessed by flow cytometry. Finally, introgression of the piece of DNA possibly containing the dicamba tolerance gene into B. napus was confirmed using florescence in situ hybridization (FISH). This research demonstrates for the first time stable introgression of dicamba tolerance from S. arvensis into B. napus via in vitro embryo rescue followed by repeated backcross breeding. Creation of dicamba-tolerant B. napus varieties by this approach may have potential to provide options to growers to choose a desirable herbicide-tolerant technology. Furthermore, adoption of such technology facilitates effective weed control, less tillage, and possibly minimize evolution of herbicide resistant weeds.

Human Immunodeficiency Virus (HIV) infection and the resultant Acquired Immunodeficiency Syndrome (AIDS) epidemic are major global health challenges; hepatitis C virus (HCV) co-infection has made the HIV/AIDS epidemic even worse. Interleukin-27 (IL-27), a cytokine which inhibits HIV and HCV replication in vitro, associates with HIV infection and HIV/HCV co-infection in clinical settings. However, the impact of HIV and HCV viral loads on plasma IL-27 expression levels has not been well characterized. In this study, 155 antiretroviral therapy-naïve Chinese were recruited. Among them 80 were HIV- and HCV-negative healthy controls, 45 were HIV-mono-infected and 30 were HIV/HCV-co-infected. Plasma level HIV, HCV, IL-27 and CD4+ number were counted and their correlation, regression relationships were explored. We show that: plasma IL-27 level was significantly upregulated in HIV-mono-infected and HIV/HCV-co-infected Chinese; HIV viral load was negatively correlated with IL-27 titer in HIV-mono-infected subjects whereas the relationship was opposite in HIV/HCV-co-infected subjects; and the relationships between HIV viral loads, IL-27 titers and CD4+ T cell counts in the HIV mono-infection and HIV/HCV co-infection groups were dramatically different. Overall, our results suggest that IL-27 differs in treatment-naïve groups with HIV mono-infections and HIV/HCV co-infections, thereby providing critical information to be considered when caring and treating those with HIV mono-infection and HIV/HCV co-infection.

BACKGROUND AND OBJECTIVES: Many studies have shown the adverse effects of air pollution on respiratory health, but few have examined the effects of air pollution on service utilisation in the primary care setting. The aim of this study was to examine the association between air pollution and the daily number of consultations due to upper respiratory tract infections (URTIs) in general outpatient clinics (GOPCs) in Hong Kong. METHODS: Daily data on the numbers of consultations due to URTIs in GOPCs, the concentrations of major air pollutants, and the mean values of metrological variables were retrospectively collected over a 3-year period (2008-2010, inclusive). Generalised additive models were constructed to examine the association between air pollution and the daily number of consultations, and to derive the relative risks and 95% confidence intervals (95% CI) of GOPC consultations for a unit increase in the concentrations of air pollutants. RESULTS: The mean daily consultations due to URTIs in GOPCs ranged from 68.4 to 253.0 over the study period. The summary relative risks (and 95% CI) of daily consultations in all GOPCs for the air pollutants PM10, NO2, O3, and SO2 were 1.005 (1.002, 1.009), 1.010 (1.006, 1.013), 1.009 (1.006, 1.012), and 1.004 (1.000, 1.008) respectively, per 10 µg/m(3) increase in the concentration of each pollutant. CONCLUSION: Significant associations were found between the daily number of consultations due to URTIs in GOPCs and the concentrations of air pollutants, implying that air pollution incurs a substantial morbidity and increases the burden of primary health care services.

α-Synuclein (α-syn) is a synaptic protein in which four mutations (A53T, A30P, E46K and gene triplication) have been found to cause an autosomal dominant form of Parkinson's disease (PD). It is also the major component of intraneuronal protein aggregates, designated as Lewy bodies (LBs), a prominent pathological hallmark of PD. How α-syn contributes to LB formation and PD is still not well-understood. It has been proposed that aggregation of α-syn contributes to the formation of LBs, which then leads to neurodegeneration in PD. However, studies have also suggested that aggregates formation is a protective mechanism against more toxic α-syn oligomers. In this study, we have generated α-syn mutants that have increased propensity to form aggregates by attaching a CL1 peptide to the C-terminal of α-syn. Data from our cellular study suggest an inverse correlation between cell viability and the amount of α-syn aggregates formed in the cells. In addition, our animal model of PD indicates that attachment of CL1 to α-syn enhanced its toxicity to dopaminergic neurons in an age-dependent manner and induced the formation of Lewy body-like α-syn aggregates in the substantia nigra. These results provide new insights into how α-syn-induced toxicity is related to its aggregation.

BACKGROUND: Hepatic insulin resistance impairs insulin's ability to suppress hepatic glucose production (HGP) and contributes to the development of type 2 diabetes (T2D). Although the interests to discover novel genes that modulate insulin sensitivity and HGP are high, it remains challenging to have a human cell based system to identify novel genes. METHODOLOGY/PRINCIPAL FINDINGS: To identify genes that modulate hepatic insulin signaling and HGP, we generated a human cell line stably expressing beta-lactamase under the control of the human glucose-6-phosphatase (G6PC) promoter (AH-G6PC cells). Both beta-lactamase activity and endogenous G6PC mRNA were increased in AH-G6PC cells by a combination of dexamethasone and pCPT-cAMP, and reduced by insulin. A 4-gene High-Throughput-Genomics assay was developed to concomitantly measure G6PC and pyruvate-dehydrogenase-kinase-4 (PDK4) mRNA levels. Using this assay, we screened an siRNA library containing pooled siRNA targeting 6650 druggable genes and identified 614 hits that lowered G6PC expression without increasing PDK4 mRNA levels. Pathway analysis indicated that siRNA-mediated knockdown (KD) of genes known to positively or negatively affect insulin signaling increased or decreased G6PC mRNA expression, respectively, thus validating our screening platform. A subset of 270 primary screen hits was selected and 149 hits were confirmed by target gene KD by pooled siRNA and 7 single siRNA for each gene to reduce G6PC expression in 4-gene HTG assay. Subsequently, pooled siRNA KD of 113 genes decreased PEPCK and/or PGC1alpha mRNA expression thereby demonstrating their role in regulating key gluconeogenic genes in addition to G6PC. Last, KD of 61 of the above 113 genes potentiated insulin-stimulated Akt phosphorylation, suggesting that they suppress gluconeogenic gene by enhancing insulin signaling. CONCLUSIONS/SIGNIFICANCE: These results support the proposition that the proteins encoded by the genes identified in our cell-based druggable genome siRNA screen hold the potential to serve as novel pharmacological targets for the treatment of T2D.

Human Immunodeficiency Virus (HIV) infection and the resultant Acquired Immunodeficiency Syndrome (AIDS) epidemic are major global health challenges; hepatitis C virus (HCV) co-infection has made the HIV/AIDS epidemic even worse. Interleukin-27 (IL-27), a cytokine which inhibits HIV and HCV replication in vitro, associates with HIV infection and HIV/HCV co-infection in clinical settings. However, the impact of HIV and HCV viral loads on plasma IL-27 expression levels has not been well characterized. In this study, 155 antiretroviral therapy-naïve Chinese were recruited. Among them 80 were HIV- and HCV-negative healthy controls, 45 were HIV-mono-infected and 30 were HIV/HCV-co-infected. Plasma level HIV, HCV, IL-27 and CD4+ number were counted and their correlation, regression relationships were explored. We show that: plasma IL-27 level was significantly upregulated in HIV-mono-infected and HIV/HCV-co-infected Chinese; HIV viral load was negatively correlated with IL-27 titer in HIV-mono-infected subjects whereas the relationship was opposite in HIV/HCV-co-infected subjects; and the relationships between HIV viral loads, IL-27 titers and CD4+ T cell counts in the HIV mono-infection and HIV/HCV co-infection groups were dramatically different. Overall, our results suggest that IL-27 differs in treatment-naïve groups with HIV mono-infections and HIV/HCV co-infections, thereby providing critical information to be considered when caring and treating those with HIV mono-infection and HIV/HCV co-infection. [ABSTRACT FROM AUTHOR]

mRNA-based COVID-19 vaccines have played a critical role in reducing severe outcomes of COVID-19. Humoral immune responses against SARS-CoV-2 after vaccination have been extensively studied in blood; however, limited information is available on the presence and duration of SARS-CoV-2 specific antibodies in saliva and other mucosal fluids. Saliva offers a non-invasive sampling method that may also provide a better understanding of mucosal immunity at sites where the virus enters the body. Our objective was to evaluate the salivary immune response after vaccination with the COVID-19 Moderna mRNA-1273 vaccine. Two hundred three staff members of the U.S. Centers for Disease Control and Prevention were enrolled prior to receiving their first dose of the mRNA-1273 vaccine. Participants were asked to self-collect 6 saliva specimens at days 0 (prior to first dose), 14, 28 (prior to second dose), 42, and 56 using a SalivaBio saliva collection device. Saliva specimens were tested for anti-spike protein SARS-CoV-2 specific IgA and IgG enzyme immunoassays. Overall, SARS-CoV-2-specific salivary IgA titers peaked 2 weeks after each vaccine dose, followed by a sharp decrease during the following weeks. In contrast to IgA titers, IgG antibody titers increased substantially 2 weeks after the first vaccine dose, peaked 2 weeks after the second dose and persisted at an elevated level until at least 8 weeks after the first vaccine dose. Additionally, no significant differences in IgA/IgG titers were observed based on age, sex, or race/ethnicity. All participants mounted salivary IgA and IgG immune responses against SARS-CoV-2 after receiving the mRNA-1273 COVID-19 vaccine. Because of the limited follow-up time for this study, more data are needed to assess the antibody levels beyond 2 months after the first dose. Our results confirm the potential utility of saliva in assessing immune responses elicited by immunization and possibly by infection., Competing Interests: The authors have declared that no competing interests exist., (Copyright: This is an open access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 public domain dedication.)

Background: Children in custodial settings are a vulnerable group. Prior to the COVID-19 pandemic there were concerns about the safety of children in these settings. COVID-19 has had an impact on everyone but given the vulnerability of children in custody, there were concerns about the impact of COVID-19 restrictions. All custody settings for children are independently inspected and this research aimed to analyse data from inspection reports. Twenty-six inspection reports undertaken between March 2020 and October 2021 were analysed to understand the impact of COVID-19 on delivery of usual care/regime., Results: Data showed that across all site's children spent considerable amounts of time isolated and in some cases, this was deemed to amount to solitary confinement. There was evidence of some positive experiences, in the smaller sites, around COVID-19 slowing the pace of life allowing staff and children could foster relationships. However, in the larger sites, isolation was extreme and COVID-19 policies such as 'bubbles' appear to have created unintended consequences as sites have moved into recovery, leading to increased violence and stress. COVID-19 directly impacted staffing levels. This and the COVID-19 policies to reduce mixing also had an impact on how children's behaviour, welfare and safeguarding was managed. In some larger sites, being COVID-19 secure was prioritised over the needs of the children., Conclusions: This research highlights the importance of multi-site longitudinal research to understand how children, staff and institution's function. The experiences of children in custody during COVID-19 differed by site type. The research suggests that the larger sites are struggling to keep children safe and there should be a shift towards smaller, more therapeutic environments. More research is needed to understand the longer-term unintended consequences of COVID-19 policy in custody, for these vulnerable children., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Lennox et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Objectives: Soccer heading is adversely associated with neurocognitive performance, but whether greater neck strength or anthropometrics mitigates these outcomes is controversial. Here, we examine the effect of neck strength or anthropometrics on associations of soccer heading with neurocognitive outcomes in a large cohort of adult amateur players., Methods: 380 adult amateur league soccer players underwent standardized measurement of neck strength (forward flexion, extension, left lateral flexion, right lateral flexion) and head/neck anthropometric measures (head circumference, neck length, neck circumference and neck volume). Participants were assessed for heading (HeadCount) and cognitive performance (Cogstate) on up to 7 visits over a period of two years. Principal components analysis (PCA) was performed on 8 neck strength and anthropometric measures. We used generalized estimating equations to test the moderation effect of each of the three PCs on 8 previously identified adverse associations of 2-week and 12-month heading estimates with cognitive performance (psychomotor speed, immediate verbal recall, verbal episodic memory, attention, working memory) and of unintentional head impacts on moderate to severe central nervous system symptoms., Results: 3 principal components (PC's) account for 80% of the variance in the PCA. In men, PC1 represents head/neck anthropometric measures, PC2 represents neck strength measures, and PC3 represents the flexor/extensor (F/E) ratio. In women, PC1 represents neck strength, PC2 represents anthropometrics, and PC3 represents the F/E ratio. Of the 48 moderation effects tested, only one showed statistical significance after Bonferroni correction, which was not robust to extensive sensitivity analyses., Conclusion: Neither neck strength nor anthropometrics mitigate adverse associations of soccer heading with cognitive performance in adult amateur players., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Lin et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Many theories of communication claim that perspective-taking is a fundamental component of the successful design of utterances for a specific audience. In three experiments, we investigated perspective-taking in a constrained communication situation: Participants played a word guessing game where each trial required them to select a clue word to communicate a single target word to their partner. In many cases, the task requires participants to take the perspective of their partner when generating, evaluating, and selecting potential clue words. For example, if the target word was 'heart', the first word that came to mind might be 'love', but this would not in fact be a very useful clue word. Instead, a word like 'cardiovascular' is much more likely than 'love' to make the partner guess 'heart'. Pairs of participants took turns giving and receiving clues to guess target words, receiving feedback after each trial. In Experiment 1, participants appeared unable to improve their perspective-taking over repeated interactions, despite a baseline performance that suggested strong perspective-taking abilities. In Experiment 2, which included extensive feedback after each trial and only target words for which good clues existed and which required perspective-taking, some measures of perspective-taking showed modest improvements. In Experiment 3, which was conducted online, we used Experiment 2 feedback with Experiment 1 target words. As in Experiment 1, participants did not improve over the course of the game in Experiment 3. The results of these three experiments show quite strong limits on people's ability to adapt and improve perspective-taking without the context provided by interaction history and growing common ground., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Nedergaard, Smith. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Topological changes like sliding motion, sources and sinks are a significant challenge in image registration. This work proposes the use of the alternating direction method of multipliers as a general framework for constraining the registration of separate objects with individual deformation fields from overlapping in image registration. This constraint is enforced by introducing a collision detection algorithm from the field of computer graphics which results in a robust divide and conquer optimization strategy using Free-Form Deformations. A series of experiments demonstrate that the proposed framework performs superior with regards to the combination of intersection prevention and image registration including synthetic examples containing complex displacement patterns. The results show compliance with the non-intersection constraints while simultaneously preventing a decrease in registration accuracy. Furthermore, the application of the proposed algorithm to the DIR-Lab data set demonstrates that the framework generalizes to real data by validating it on a lung registration problem., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Alscher et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Background: With the recent legalization of cannabis in Canada, there is an urgent need to understand the effect of cannabis use in pregnancy. Our population-based study investigated the effects of prenatal cannabis use on maternal and newborn outcomes, and modification by infant sex., Methods: The cohort included 1,280,447 singleton births from the British Columbia Perinatal Data Registry, the Better Outcomes Registry & Network Ontario, and the Perinatal Program Newfoundland Labrador from April 1st, 2012 to March 31st, 2019. Logistic regression determined the associations between prenatal cannabis use and low birth weight, small-for-gestational age, large-for-gestational age, spontaneous and medically indicated preterm birth, very preterm birth, stillbirth, major congenital anomalies, caesarean section, gestational diabetes and gestational hypertension. Models were adjusted for other substance use, socio-demographic and-economic characteristics, co-morbidities. Interaction terms were included to investigate modification by infant sex., Results: The prevalence of cannabis use in our cohort was approximately 2%. Prenatal cannabis use is associated with increased risks of spontaneous and medically indicated preterm birth (1.80[1.68-1.93] and 1.94[1.77-2.12], respectively), very preterm birth (1.73[1.48-2.02]), low birth weight (1.90[1.79-2.03]), small-for-gestational age (1.21[1.16-1.27]) and large-for-gestational age (1.06[1.01-1.12]), any major congenital anomaly (1.71[1.49-1.97]), caesarean section (1.13[1.09-1.17]), and gestational diabetes (1.32[1.23-1.42]). No association was found for stillbirth or gestational hypertension. Only small-for-gestational age (p = 0.03) and spontaneous preterm birth (p = 0.04) showed evidence of modification by infant sex., Conclusions: Prenatal cannabis use increases the likelihood of preterm birth, low birth weight, small-for-gestational age and major congenital anomalies with prenatally exposed female infants showing evidence of increased susceptibility. Additional measures are needed to inform the public and providers of the inherent risks of cannabis exposure in pregnancy., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2022 Luke et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

We investigate the accuracy of intensity-based deformable image registration (DIR) for tumor localization in liver stereotactic body radiotherapy (SBRT). We included 4DCT scans to capture the breathing motion of eight patients receiving SBRT for liver metastases within a retrospective clinical study. Each patient had three fiducial markers implanted. The liver and the tumor were delineated in the mid-ventilation phase, and their positions in the other phases were estimated with deformable image registration. We tested referenced and sequential registrations strategies. The fiducial markers were the gold standard to evaluate registration accuracy. The registration errors related to measured versus estimated fiducial markers showed a mean value less than 1.6mm. The positions of some fiducial markers appeared not stable on the 4DCT throughout the respiratory phases. Markers' center of mass tends to be a more reliable measurement. Distance errors of tumor location based on registration versus markers center of mass were less than 2mm. There were no statistically significant differences between the reference and the sequential registration, i.e., consistency and errors were comparable to resolution errors. We demonstrated that intensity-based DIR is accurate up to resolution level for locating the tumor in the liver during breathing motion., Competing Interests: The authors LBS, MJ, SN and IRV received grants and/or educational fees from Varian Medical Systems.

Introduction: Systems modelling and simulation can improve understanding of complex systems to support decision making, better managing system challenges. Advances in technology have facilitated accessibility of modelling by diverse stakeholders, allowing them to engage with and contribute to the development of systems models (participatory modelling). However, despite its increasing applications across a range of disciplines, there is a growing need to improve evaluation efforts to effectively report on the quality, importance, and value of participatory modelling. This paper aims to identify and assess evaluation frameworks, criteria, and/or processes, as well as to synthesize the findings into a comprehensive multi-scale framework for participatory modelling programs., Materials and Methods: A scoping review approach was utilized, which involved a systematic literature search via Scopus in consultation with experts to identify and appraise records that described an evaluation framework, criteria, and/or process in the context of participatory modelling. This scoping review is registered with the Open Science Framework., Results: The review identified 11 studies, which varied in evaluation purposes, terminologies, levels of examination, and time points. The review of studies highlighted areas of overlap and opportunities for further development, which prompted the development of a comprehensive multi-scale evaluation framework to assess participatory modelling programs across disciplines and systems modelling methods. The framework consists of four categories (Feasibility, Value, Change/Action, Sustainability) with 30 evaluation criteria, broken down across project-, individual-, group- and system-level impacts., Discussion & Conclusion: The presented novel framework brings together a significant knowledge base into a flexible, cross-sectoral evaluation effort that considers the whole participatory modelling process. Developed through the rigorous synthesis of multidisciplinary expertise from existing studies, the application of the framework can provide the opportunity to understand practical future implications such as which aspects are particularly important for policy decisions, community learning, and the ongoing improvement of participatory modelling methods., Competing Interests: Dr Louise Freebairn is currently employed part-time by the Brain and Mind Centre, University of Sydney; ACT Health as Director of Knowledge Translation and Health Outcomes, Epidemiology Section, ACT Health & CSART as Director of Policy Applications & Translational Science. A/Professor Jo-An Occhipinti is both Head of Systems Modelling, Simulation & Data Science at the University of Sydney’s Brain and Mind Centre and Managing Director of Computer Simulation & Advanced Research Technologies (CSART). Professor Ian Hickie is the Co-Director, Health and Policy at the Brain and Mind Centre (BMC) University of Sydney. The BMC operates an early-intervention youth services at Camperdown under contract to headspace. He is the Chief Scientific Advisor to, and a 5% equity shareholder in, InnoWell Pty Ltd. InnoWell was formed by the University of Sydney (45% equity) and PwC (Australia; 45% equity) to deliver the $30 M Australian Government-funded Project Synergy (2017-20; a three-year program for the transformation of mental health services) and to lead transformation of mental health services internationally through the use of innovative technologies.

Objective: To determine the significance of dysphagia on clinical outcomes of geriatric trauma patients., Methods: This is a retrospective population-based study of geriatric trauma patients 65 years and older utilizing the Florida Agency for Health Care Administration dataset from 2010 to 2019. Patients with pre-admission dysphagia were excluded. Multivariable regression was used to create statistical adjustments. Primary outcomes included mortality and the development of dysphagia. Secondary outcomes included length of stay and complications. Subgroup analyses included patients with dementia, patients who received transgastric feeding tubes (GFTs) or tracheostomies, and speech language therapy consultation., Results: A total of 52,946 geriatric patients developed dysphagia after admission during a 9-year period out of 1,150,438 geriatric trauma admissions. In general, patients who developed dysphagia had increased mortality, length of stay, and complications. When adjusted for traumatic brain and cervical spine injuries, the addition of mechanical ventilation decreased the mortality odds. This was also observed in the subset of patients with dysphagia who had GFTs placed. Of the three primary risk factors for dysphagia investigated, mechanical ventilation was the most strongly associated with later development of dysphagia and mortality., Conclusion: The geriatric trauma population is vulnerable to dysphagia with a large number associated with traumatic brain injury, cervical spine injury, and polytraumatic injuries that lead to mechanical ventilation. Earlier intubation/mechanical ventilation in association with GFTs was found to be associated with decreased inpatient hospital mortality. Tracheostomy placement was shown to be an independent risk factor for the development of dysphagia. The utilization of speech language therapy was found to be inconsistently utilized., Competing Interests: The authors have declared that no competing interests exist.

Background: With the introduction of the novel COVID-19 vaccine, public hesitancy is being experienced with many turning to healthcare professionals for advice. As future physicians, medical students play a critical role in the public's view of the vaccine., Objectives: To determine the attitude of U.S. medical students toward mandating the COVID-19 vaccine to healthcare workers and patients, as well as whether their knowledge of the vaccine plays a role in their view., Methods: The authors emailed a survey link to all U.S. medical schools with request to distribute it to their medical students. The survey remained open from 02/09/2021 to 03/15/2021 and included questions to determine the attitude of the medical students toward recommending the COVID-19 vaccine, and general knowledge questions about the vaccine. Chi square, Fisher's exact test, and linear regression were conducted to determine associations between willingness to recommend the COVID-19 vaccine and general knowledge of the vaccine., Results: Among the 1,899 responses from medical students representing 151 U.S. medical schools, 57.82% approved of making the COVID-19 vaccine mandatory to healthcare workers, and 16.27% approved of making it mandatory to patients. Additionally, those who tested most knowledgeable of the vaccine were less likely to approve of making the vaccine mandatory for patients (66.67% vs. 72.70). Those that tested most knowledgeable were also more likely to personally receive the vaccine (72.35% vs 62.99%) as opposed to those that tested the least knowledgeable who were less willing to personally receive the vaccine (4.12% vs 14.17%)., Conclusions: The data revealed that a slight majority of medical students support a vaccine mandate toward healthcare workers while a minority of medical students support a vaccine mandate toward healthcare workers. Additionally, medical students that had relatively high knowledge of the vaccine correlated with not approving of making the vaccine mandatory for patients. However increased knowledge of the vaccine correlated with increased willingness to personally receive the vaccine., Competing Interests: The authors have declared that no competing interests exist.

Studying different types of tooth movements can help us to better understand the force systems used for tooth position correction in orthodontic treatments. This study considers a more realistic force system in tooth movement modeling across different patients and investigates the effect of the couple force direction on the position of the center of rotation (CRot). The finite-element (FE) models of human mandibles from three patients are used to investigate the position of the CRots for different patients' teeth in 3D space. The CRot is considered a single point in a 3D coordinate system and is obtained by choosing the closest point on the axis of rotation to the center of resistance (CRes). A force system, consisting of a constant load and a couple (pair of forces), is applied to each tooth, and the corresponding CRot trajectories are examined across different patients. To perform a consistent inter-patient analysis, different patients' teeth are registered to the corresponding reference teeth using an affine transformation. The selected directions and applied points of force on the reference teeth are then transformed into the registered teeth domains. The effect of the direction of the couple on the location of the CRot is also studied by rotating the couples about the three principal axes of a patient's premolar. Our results indicate that similar patterns can be obtained for the CRot positions of different patients and teeth if the same load conditions are used. Moreover, equally rotating the direction of the couple about the three principal axes results in different patterns for the CRot positions, especially in labiolingual direction. The CRot trajectories follow similar patterns in the corresponding teeth, but any changes in the direction of the force and couple cause misalignment of the CRot trajectories, seen as rotations about the long axis of the tooth., Competing Interests: Authors TG and PS are affiliated with 3Shape A/S, which provided support in the form of salaries and data for this study. The commercial affiliation of the aforementioned authors does not alter our adherence to PLOS ONE policies on sharing data and materials. Additionally, there are no patents, products in development or marketed products associated with this research to declare.

Social anxiety is associated with reduced educational achievement. Given that concentration is a predictor of educational achievement, and social anxiety symptoms are associated with reduced concentration in class, this prospective study examined the possibility that social anxiety may impair educational achievement through reduced classroom concentration. A sample of 509 participants (53.8% female; M age: 12.77 years [SD = 0.81]) recruited from secondary schools completed questionnaires assessing social anxiety symptoms, depressive symptoms, and concentration in class. Educational achievement was assessed by internal grades within schools. An indirect effect of social anxiety on later educational achievement via concentration was observed, over and above baseline achievement and depression symptoms; adolescents with higher levels of social anxiety tend to have more difficulties concentrating in class, which in turn is associated with poorer academic outcomes. Findings underscore the challenges socially anxious adolescents will face trying to learn in school, and the need for education providers and clinicians to consider the effect of social anxiety symptoms on concentration and learning., Competing Interests: The authors have declared that no competing interests exist.

Background: Self-focused attention and safety behaviours are both associated with adolescent social anxiety. In adults, experimental studies have indicated that the processes are causally implicated in social anxiety, but this hypothesis has not yet been tested in a youth sample., Methods: This experiment explored this possibility by asking high and low socially anxious adolescents (N = 57) to undertake conversations under different conditions. During one conversation they were instructed to focus on themselves and use safety behaviours, and in the other they focused externally and did not use safety behaviours. Self-report, conversation partner report and independent assessor ratings were taken., Results: Self-focus and safety behaviours increased feelings and appearance of anxiety and undermined performance for all participants, but only high socially anxious participants reported habitually using self-focus and safety behaviours., Conclusions: The findings provide support for the causal role of self-focus and safety behaviours in adolescent social anxiety and point to the potential clinical value of techniques reversing them to treat the disorder., Competing Interests: The authors have declared that no competing interests exist.

The demand for food will outpace productivity of conventional agriculture due to projected growth of the human population, concomitant with shrinkage of arable land, increasing scarcity of freshwater, and a rapidly changing climate. While aquaponics has potential to sustainably supplement food production with minimal environmental impact, there is a need to better characterize the complex interplay between the various components (fish, plant, microbiome) of these systems to optimize scale up and productivity. Here, we investigated how the commonly-implemented practice of continued microbial community transfer from pre-existing systems might promote or impede productivity of aquaponics. Specifically, we monitored plant growth phenotypes, water chemistry, and microbiome composition of rhizospheres, biofilters, and fish feces over 61-days of lettuce (Lactuca sativa var. crispa) growth in nitrogen-limited aquaponic systems inoculated with bacteria that were either commercially sourced or originating from a pre-existing aquaponic system. Lettuce above- and below-ground growth were significantly reduced across replicates treated with a pre-existing aquaponic system inoculum when compared to replicates treated with a commercial inoculum. Reduced productivity was associated with enrichment in specific bacterial genera in plant roots, including Pseudomonas, following inoculum transfer from pre-existing systems. Increased productivity was associated with enrichment of nitrogen-fixing Rahnella in roots of plants treated with the commercial inoculum. Thus, we show that inoculation from a pre-existing system, rather than from a commercial inoculum, is associated with lower yields. Further work will be necessary to test the putative mechanisms involved., Competing Interests: The authors have declared that no competing interests exist.

Background: Analysis of cell-free tumour DNA, a liquid biopsy, is a promising biomarker for cancer. We have performed a proof-of principle study to test the applicability in the clinical setting, analysing copy number alterations (CNAs) in plasma and tumour tissue from 44 patients with gastro-oesophageal cancer., Methods: DNA was isolated from blood plasma and a tissue sample from each patient. Array-CGH was applied to the tissue DNA. The cell-free plasma DNA was sequenced by low-coverage whole-genome sequencing using a clinical pipeline for non-invasive prenatal testing. WISECONDOR and ichorCNA, two bioinformatic tools, were used to process the output data and were compared to each other., Results: Cancer-associated CNAs could be seen in 59% (26/44) of the tissue biopsies. In the plasma samples, a targeted approach analysing 61 regions of special interest in gastro-oesophageal cancer detected cancer-associated CNAs with a z-score >5 in 11 patients. Broadening the analysis to a whole-genome view, 17/44 patients (39%) had cancer-associated CNAs using WISECONDOR and 13 (30%) using ichorCNA. Of the 26 patients with tissue-verified cancer-associated CNAs, 14 (54%) had corresponding CNAs in plasma. Potentially clinically actionable amplifications overlapping the genes VEGFA, EGFR and FGFR2 were detected in the plasma from three patients., Conclusions: We conclude that low-coverage whole-genome sequencing without prior knowledge of the tumour alterations could become a useful tool for cell-free tumour DNA analysis of total CNAs in plasma from patients with gastro-oesophageal cancer., Competing Interests: The authors have declared that no competing interests exist.

Digital Light Processing (DLP) stereolithography (SLA) as a high-resolution 3D printing process offers a low-cost alternative for prototyping of microfluidic geometries, compared to traditional clean-room and workshop-based methods. Here, we investigate DLP-SLA printing performance for the production of micro-chamber chip geometries suitable for Polymerase Chain Reaction (PCR), a key process in molecular diagnostics to amplify nucleic acid sequences. A DLP-SLA fabrication protocol for printed micro-chamber devices with monolithic micro-channels is developed and evaluated. Printed devices were post-processed with ultraviolet (UV) light and solvent baths to reduce PCR inhibiting residuals and further treated with silane coupling agents to passivate the surface, thereby limiting biomolecular adsorption occurences during the reaction. The printed devices were evaluated on a purpose-built infrared (IR) mediated PCR thermocycler. Amplification of 75 base pair long target sequences from genomic DNA templates on fluorosilane and glass modified chips produced amplicons consistent with the control reactions, unlike the non-silanized chips that produced faint or no amplicon. The results indicated good functionality of the IR thermocycler and good PCR compatibility of the printed and silanized SLA polymer. Based on the proposed methods, various microfluidic designs and ideas can be validated in-house at negligible costs without the requirement of tool manufacturing and workshop or clean-room access. Additionally, the versatile chemistry of 3D printing resins enables customised surface properties adding significant value to the printed prototypes. Considering the low setup and unit cost, design flexibility and flexible resin chemistries, DLP-SLA is anticipated to play a key role in future prototyping of microfluidics, particularly in the fields of research biology and molecular diagnostics. From a system point-of-view, the proposed method of thermocycling shows promise for portability and modular integration of funcitonalitites for diagnostic or research applications that utilize nucleic acid amplification technology., Competing Interests: The authors have read the journal’s policy and have the following potential competing interests: QuantumDx had a role in the study design. This does not alter our adherence to PLOS ONE policies on sharing data and materials. There are no patents, products in development or marketed products associated with this research to declare.

The rapid global spread of the coronavirus disease (COVID-19) has inflicted significant health and socioeconomic burden on affected countries. As positive cases continued to rise in Malaysia, public health laboratories experienced an overwhelming demand for COVID-19 screening. The confirmation of positive cases of COVID-19 has solely been based on the detection of the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) using real-time reverse transcription polymerase chain reaction (qRT-PCR). In efforts to increase the cost-effectiveness and efficiency of COVID-19 screening, we evaluated the feasibility of pooling clinical Nasopharyngeal/Oropharyngeal (NP/OP) swab specimens during nucleic acid extraction without a reduction in sensitivity of qRT-PCR. Pools of 10 specimens were extracted and subsequently tested by qRT-PCR according to the WHO-Charité protocol. We demonstrated that the sample pooling method showed no loss of sensitivity. The effectiveness of the pooled testing strategy was evaluated on both retrospective and prospective samples, and the results showed a similar detection sensitivity compared to testing individual sample alone. This study demonstrates the feasibility of using a pooled testing strategy to increase testing capacity and conserve resources, especially when there is a high demand for disease testing., Competing Interests: The authors have declared that no competing interests exist.

Background: The identification of negative self-imagery as a maintenance factor in adult social anxiety has led to enhanced treatments for this population. Whilst intrusive negative imagery is commonly reported by socially anxious adolescents, no studies have demonstrated that it plays a causal role in maintaining symptoms. To assess this possibility, we undertook an experimental study manipulating social self-imagery in high socially anxious adolescents., Methods: High socially anxious adolescents undertook two conversations under different conditions. During one conversation they held a negative social self-image in mind, and in the other they held a benign social self-image in mind. Self-report, conversation partner report and independent assessor ratings were taken., Results: When participants held a negative self-image in mind, they reported feeling more anxious, and believed they looked more anxious and performed more poorly. Furthermore, they overestimated how anxious they looked compared to conversation partner ratings. As well as distorting participants' perception of their anxious appearance, holding a negative image in mind also had observable effects on the interaction. Participants were rated as looking more anxious and performing less well by their conversation partner when they held such images in mind, and the conversation was rated more critically by conversation partners and independent observers. Finally, a preliminary mediation analysis suggested that the detrimental effect of negative imagery on the social interaction may be partly due to the spontaneous use of avoidant safety behaviours., Conclusions: The findings provide support for a causal role of negative self-imagery in adolescent social anxiety and point to the potential clinical value of techniques targeting imagery to treat the disorder., Competing Interests: The authors have declared that no competing interests exist.

Introduction: Endometriosis has a significant cost of illness burden in Europe, UK and the USA, with the majority of costs coming from reductions in productivity. However, information is scarce on if there is a differing impact between endometriosis and other causes of chronic pelvic pain, and if there are modifiable factors, such as pain severity, that may be significant contributors to the overall burden., Methods: An online survey was hosted by SurveyMonkey and the link was active between February to April 2017. Women aged 18-45, currently living in Australia, who had either a confirmed diagnosis of endometriosis via laparoscopy or chronic pelvic pain without a diagnosis of endometriosis were included. The retrospective component of the WERF EndoCost tool was used to determine direct healthcare costs, direct non-healthcare costs (carers) and indirect costs due to productivity loss. Estimates were extrapolated to the Australian population using published prevalence estimates., Results: 407 valid responses were received. The cost of illness burden was significant in women with chronic pelvic pain (Int $16,970 to $ 20,898 per woman per year) irrespective of whether they had a diagnosis of endometriosis. The majority of costs (75-84%) were due to productivity loss. Both absolute and relative productivity costs in Australia were higher than previous estimates based on data from Europe, UK and USA. Pain scores showed the strongest relationship to productivity costs, a 12.5-fold increase in costs between minimal to severe pain. The total economic burden per year in Australia in the reproductive aged population (at 10% prevalence) was 6.50 billion Int $., Conclusion: Similar to studies in European, British and American populations, productivity costs are the greatest contributor to overall costs. Given pain is the most significant contributor, priority should be given to improving pain control in women with pelvic pain., Competing Interests: I have read the journal's policy and the authors of this manuscript have the following competing interests: MA,CS, AT: As a medical research institute, NICM Health Research Institute receives research grants and donations from foundations, universities, government agencies and industry. Sponsors and donors provide untied and tied funding for work to advance the vision and mission of the Institute. This study was not specifically supported by donor or sponsor funding to NICM. JA: reports he is a committee member for the Endometriosis National Action plan implementation committee in Australia and the president of the Australasian Gynaecological Endoscopy and Surgery (AGES) Society. This does not alter our adherence to PLOS ONE policies on sharing data and materials.

Viruses have different strategies for infecting their hosts. Fast and acute infections result in the development of severe symptoms and may cause the death of the plant. By contrast, in a persistent interaction, the virus can survive within its host for a long time, inducing only mild symptoms. In this study, we investigated the gene expression changes induced in CymRSV-, crTMV-, and TCV-infected Nicotiana benthamiana and in PVX- and TMV-U1-infected Solanum lycopersicum plants after the systemic spread of the virus by two different high-throughput methods: microarray hybridization or RNA sequencing. Using these techniques, we were able to clearly differentiate between acute and persistent infections. We validated the gene expression changes of selected genes by Northern blot hybridization or by qRT-PCR. We show that, in contrast to persistent infections, the drastic shut-off of housekeeping genes, downregulation of photosynthesis-related transcripts and induction of stress genes are specific outcomes with acute infections. We also show that these changes are not a consequence of host necrosis or the presence of a viral silencing suppressor. Thermal imaging data and chlorophyll fluorescence measurements correlated very well with the molecular changes. We believe that the molecular and physiological changes detected during acute infections mostly contribute to virus symptom development. The observed characteristic physiological changes associated with economically more dangerous acute infections could serve as a basis for the elaboration of remote monitoring systems suitable for detecting developing virus infections in crops. Moreover, as molecular and physiological changes are characteristics of different types of virus lifestyles, this knowledge can support risk assessments of recently described novel viruses., Competing Interests: The authors have declared that no competing interests exist.

In contemporary society, it is essential to have adequate mathematical skills. Being numerate has been linked to positive life outcomes and well-being in adults. It is also acknowledged that math anxiety (MA) hampers mathematical skills increasingly with age. Still, the mechanisms by which MA affect performance remain debated. Using structural equation modeling (SEM), we contrast the different ways in which MA has been suggested to interfere with math abilities. Our models indicate that MA may affect math performance through three pathways: (1) indirectly through working memory ability, giving support for the 'affective drop' hypothesis of MA's role in mathematical performance, (2) indirectly through symbolic number processing, corroborating the notion of domain-specific mechanisms pertaining to number, and (3) a direct effect of MA on math performance. Importantly, the pathways vary in terms of their relative strength depending on what type of mathematical problems are being solved. These findings shed light on the mechanisms by which MA may interfere with mathematical performance., Competing Interests: The authors have declared that no competing interests exist.

The African lion (Panthera leo), listed as a vulnerable species on the IUCN Red List of Threatened Species (Appendix II of CITES), is mainly impacted by indiscriminate killing and prey base depletion. Additionally, habitat loss by land degradation and conversion has led to the isolation of some subpopulations, potentially decreasing gene flow and increasing inbreeding depression risks. Genetic drift resulting from weakened connectivity between strongholds can affect the genetic health of the species. In the present study, we investigated the evolutionary history of the species at different spatiotemporal scales. Therefore, the mitochondrial cytochrome b gene (N = 128), 11 microsatellites (N = 103) and 9,103 SNPs (N = 66) were investigated in the present study, including a large sampling from Tanzania, which hosts the largest lion population among all African lion range countries. Our results add support that the species is structured into two lineages at the continental scale (West-Central vs East-Southern), underlining the importance of reviewing the taxonomic status of the African lion. Moreover, SNPs led to the identification of three lion clusters in Tanzania, whose geographical distributions are in the northern, southern and western regions. Furthermore, Tanzanian lion populations were shown to display good levels of genetic diversity with limited signs of inbreeding. However, their population sizes seem to have gradually decreased in recent decades. The highlighted Tanzanian African lion population genetic differentiation appears to have resulted from the combined effects of anthropogenic pressure and environmental/climatic factors, as further discussed., Competing Interests: The authors have declared that no competing interests exist.

Scleria subgen. Hypoporum (Cyperaceae), with 68 species, is the second largest subgenus in Scleria. Species of this pantropically distributed subgenus generally occur in seasonally or permanently wet grasslands or on shallow soils over sandstone or lateritic outcrops, less often they can be found in (open) woodlands. Previous studies established the monophyly of the subgenus, but the relationships between the species remained uncertain. In this study, DNA sequence data of 61 taxa of Scleria subgen. Hypoporum, where possible represented by multiple accessions from across their distributional range, were obtained for four molecular markers: the coding chloroplast marker ndhF, the chloroplast intron rps16 and the nuclear ribosomal regions ETS and ITS. Phylogenetic trees were constructed using Bayesian inference and maximum likelihood approaches. A species tree was constructed to summarise the results. The results indicate the existence of three sections: the monotypic, pantropically occurring, Scleria sect. Lithospermae, a new section from central and south America containing two species, and Scleria sect. Hypoporum, also pantropically distributed, containing the remainder of the species of the subgenus. Relationships in the latter section are not fully resolved. However, three or four different clades can be distinguished supported by some morphological characters. Our results indicate at least six new species in Scleria sect. Hypoporum. The new section and species are described in a taxonomical treatment. Their morphology is compared with (morphologically) closely related species., Competing Interests: The authors have declared that no competing interests exist.

A single-injection vaccine formulation that provides for both a prime and a boost immunization would have various advantages over a multiple-injection regime. For such a vaccine formulation, it is essential that the booster dose is released after a certain, preferably adjustable, lag time. In this study we investigated whether a core-shell based implant, containing ovalbumin as core material and poly(DL-lactic-co-glycolic acid) of various monomer ratios as shell material can be used to obtain such a booster release. An in vitro release study showed that the lag time after which the ovalbumin was released from the core-shell implant increased with increasing lactic to glycolic acid ratio of the polymer and ranged from 3-6 weeks. Fluorescence spectroscopy showed minimal differences between native ovalbumin and ovalbumin from core-shell implants that were incubated until just before the observed in vitro release. In addition, mice immunized with a subcutaneous inserted core-shell implant containing ovalbumin showed an ovalbumin-specific IgG1 antibody response after a lag time of 4 or 6-8 weeks. Moreover, delayed release of ovalbumin caused higher IgG1 antibody titers than conventional subcutaneous vaccination with ovalbumin dissolved in PBS. Collectively, these findings could contribute to the further development of a single-injection vaccine, making multiple injections of the vaccine superfluous., Competing Interests: Katie Amssoms, Ben De Clerck, Ellen Van Gulck, Guenter Kraus, Roger Sutmuller, and Kenny Simmen are or were at the time of the project employees of a Johnson & Johnson affiliate. Lieven Baert is managing director at Jalima Pharma and acted as consultant for Janssen. This does not alter our adherence to PLOS ONE policies on sharing data and materials.

The Triple Code Model (TCM) of numerical cognition argues for the existence of three representational codes for number: Arabic digits, verbal number words, and analog nonsymbolic magnitude representations, each subserved by functionally dissociated neural substrates. Despite the popularity of the TCM, no study to date has explored all three numerical codes within one fMRI paradigm. We administered three tasks, associated with each of the aforementioned numerical codes, in order to explore the neural correlates of numerosity processing in a sample of adults (N = 46). Independent task-control contrast analyses revealed task-dependent activity in partial support of the model, but also highlight the inherent complexity of a distributed and overlapping fronto-parietal network involved in all numerical codes. The results indicate that the TCM correctly predicts the existence of some functionally dissociated neural substrates, but requires an update that accounts for interactions with attentional processes. Parametric contrasts corresponding to differences in task difficulty revealed specific neural correlates of the distance effect, where closely spaced numbers become more difficult to discriminate than numbers spaced further apart. A conjunction analysis illustrated overlapping neural correlates across all tasks, in line with recent proposals for a fronto-parietal network of number processing. We additionally provide tentative results suggesting the involvement of format-independent numerosity-sensitive retinotopic maps in the early visual stream, extending previous findings of nonsymbolic stimulus selectivity. We discuss the functional roles of the components associated with the model, as well as the purported fronto-parietal network, and offer arguments in favor of revising the TCM., Competing Interests: The authors have declared that no competing interests exist.

High rates of sexual partner concurrency have been shown to facilitate the spread of various sexually transmitted infections. Assessments of explicit attitudes to concurrency have however found little difference between populations. Implicit attitudes to concurrency may vary between populations and play a role in generating differences in the prevalence of concurrency. We developed a concurrency implicit associations test (C-IAT) to assess if implicit attitudes towards concurrency may vary between individuals and populations and what the correlates of these variations are. A sample of 869 Belgian students (mean age 23, SD 5.1) completed an online version of the C-IAT together with a questionnaire concerning sexual behavior and explicit attitudes to concurrency. The study participants C-IATs demonstrated a strong preference for monogamy (-0.78, SD = 0.41). 93.2% of participants had a pro-monogamy C-IAT. There was no difference in this implicit preference for monogamy between heterosexual men and women. Men who have sex with men and women who have sex with women were more likely to exhibit implicit but not explicit preferences for concurrency compared to heterosexual men and women. Correlates of the C-IAT varied between men and women.

Introduction: HIV-1 dual infection is a condition that results from infection with at least two HIV-1 variants from different sources. The scarceness of information on this condition is partly due to the fact that its detection is technically challenging. Using next-generation sequencing we defined the extent of HIV-1 dual infection in a cohort of men who have sex with men (MSM)., Material & Methods: Eighty-six MSM, diagnosed with HIV-1 subtype B infection between 2008 and 2013 were selected for next-generation sequencing of the HIV-1 envelope V3. Sequencing was performed on 2 plasma samples collected with an interval of > 6 months before the initiation of antiretroviral therapy. Maximum likelihood phylogenetic trees were inspected for dual infection, defined as the presence of two or more monophyletic clusters with ≥ 90% bootstrap support and a mean between-cluster genetic distance of ≥ 10%. To confirm dual infection, deep V3 sequencing of intermediate samples was performed as well as clonal sequencing of the HIV-1 protease-reverse transcriptase gene., Results: Five of the 74 patients (6.8%) for whom deep sequencing was successful, showed clear evidence of dual infection. In 4 of them, the second strain was absent in the first sample but occurred in subsequent samples. This was highly suggestive for superinfection. In 3 patients both virus variants were of subtype B, in 2 patients at least one of the variants was a subtype B/non-B recombinant virus., Conclusions: Dual infection was confirmed in 6.8% of MSM diagnosed with HIV-1 in Belgium. This prevalence is probably an underestimation, because stringent criteria were used to classify viral variants as originating from a new infection event.

[This corrects the article DOI: 10.1371/journal.pone.0177266.].

MicroRNAs are endogenous, small non-coding RNAs that control gene expression by directing their target mRNAs for degradation and/or posttranscriptional repression. Abnormal expression of microRNAs is thought to contribute to the development and progression of cancer. A history of benign breast disease (BBD) is associated with increased risk of subsequent breast cancer. However, no large-scale study has examined the association between microRNA expression in BBD tissue and risk of subsequent invasive breast cancer (IBC). We conducted discovery and validation case-control studies nested in a cohort of 15,395 women diagnosed with BBD in a large health plan between 1971 and 2006 and followed to mid-2015. Cases were women with BBD who developed subsequent IBC; controls were matched 1:1 to cases on age, age at diagnosis of BBD, and duration of plan membership. The discovery stage (316 case-control pairs) entailed use of the Illumina MicroRNA Expression Profiling Assay (in duplicate) to identify breast cancer-associated microRNAs. MicroRNAs identified at this stage were ranked by the strength of the correlation between Illumina array and quantitative PCR results for 15 case-control pairs. The top ranked 14 microRNAs entered the validation stage (165 case-control pairs) which was conducted using quantitative PCR (in triplicate). In both stages, linear regression was used to evaluate the association between the mean expression level of each microRNA (response variable) and case-control status (independent variable); paired t-tests were also used in the validation stage. None of the 14 validation stage microRNAs was associated with breast cancer risk. The results of this study suggest that microRNA expression in benign breast tissue does not influence the risk of subsequent IBC.

Classical or M1 activity of microglia/macrophages has been described in several neurodegenerative and brain inflammatory conditions and has also been linked to expansion of ischemic injury in post-stroke brain. While different pathways of M1 polarization have been suggested to occur in the post-stroke brain, the precise underlying mechanisms remain undefined. Using a transient middle cerebral artery occlusion (MCAO) rat model, we showed a progressive M2 to M1 polarization in the perilesional brain region with M1 cells becoming one of the dominant subsets by day 4 post-stroke. Comparing key receptors involved in M1 polarization (CD8, IFNγR, Clec4, FcγR, TLR3 and TLR4) and their signal transducers (Syk, Stat1, Irf3, and Traf6) at the day 4 time point, we showed a strong upregulation of CD8 along with SYK transducer in dissected perilesional brain tissue. We further showed that CD8 expression in the post-stroke brain was associated with activated (CD68+) macrophages and that progressive accumulation of CD8+CD68+ cells in the post-stroke brain coincided with increased iNOS (M1 marker) and reduced Arg1 (M2 marker) expression on these cells. In vitro ligand-based stimulation of the CD8 receptor caused increased iNOS expression and an enhanced capacity to phagocytose E. coli particles; and interestingly, CD8 stimulation was also able to repolarize IL4-treated M2 cells to an M1 phenotype. Our data suggest that increased CD8 signaling in the post-stroke brain is primarily associated with microglia/macrophages and can independently drive M1 polarization, and that modulation of CD8 signaling could be a potential target to limit secondary post-stroke brain damage.

Objectives: Ischemia-reperfusion injury (IRI) is a major cause of acute kidney injury and chronic kidney disease. Two promising preconditioning methods for the kidney, intermittent arterial clamping (IC) and treatment with the hypoxia mimetic cobalt chloride, have never been directly compared. Furthermore, the protective efficacy of the chemically related transition metal Zn2+ against renal IRI is unclear. Although Co2+ ions have been shown to protect the kidney via hypoxia inducible factor (HIF), the effect of Zn2+ ions on the induction of HIF1α, HIF2α and HIF3α has not been investigated previously., Materials and Methods: The efficacy of different preconditioning techniques was assessed using a Sprague-Dawley rat model of renal IRI. Induction of HIF proteins following Zn2+ treatment of the human kidney cell lines HK-2 (immortalized normal tubular cells) and ACHN (renal cancer) was measured using Western Blot., Results: Following 40 minutes of renal ischemia in rats, cobalt preconditioning offered greater protection against renal IRI than IC as evidenced by lower peak serum creatinine and urea concentrations. ZnCl2 (10 mg/kg) significantly lowered the creatinine and urea concentrations compared to saline-treated control rats following a clinically relevant 60 minutes of ischemia. Zn2+ induced expression of HIF1α and HIF2α but not HIF3α in HK-2 and ACHN cells., Conclusion: ZnCl2 preconditioning protects against renal IRI in a dose-dependent manner. Further studies are warranted to determine the possible mechanisms involved, and to assess the benefit of ZnCl2 preconditioning for clinical applications.

With increased production of genomic data since the advent of next-generation sequencing (NGS), there has been a need to develop new bioinformatics tools and areas, such as comparative genomics. In comparative genomics, the genetic material of an organism is directly compared to that of another organism to better understand biological species. Moreover, the exponentially growing number of deposited prokaryote genomes has enabled the investigation of several genomic characteristics that are intrinsic to certain species. Thus, a new approach to comparative genomics, termed pan-genomics, was developed. In pan-genomics, various organisms of the same species or genus are compared. Currently, there are many tools that can perform pan-genomic analyses, such as PGAP (Pan-Genome Analysis Pipeline), Panseq (Pan-Genome Sequence Analysis Program) and PGAT (Prokaryotic Genome Analysis Tool). Among these software tools, PGAP was developed in the Perl scripting language and its reliance on UNIX platform terminals and its requirement for an extensive parameterized command line can become a problem for users without previous computational knowledge. Thus, the aim of this study was to develop a web application, known as PanWeb, that serves as a graphical interface for PGAP. In addition, using the output files of the PGAP pipeline, the application generates graphics using custom-developed scripts in the R programming language. PanWeb is freely available at http://www.computationalbiology.ufpa.br/panweb.