State dependence explains individual variation in nest defence behaviour in a long-lived bird

Parental care, such as nest or offspring defence, is crucial for offspring survival in many species. Yet, despite its obvious fitness benefits, the level of defence can consistently vary between individuals of the same species. One prominent adaptive explanation for consistent individual differences in behaviours involves state dependency: relatively stable differences in individual state should lead to the emergence of repeatable behavioural variation whereas changes in state should lead to a readjustment of behaviour. Therefore, empirical testing of adaptive state dependence requires longitudinal data where behaviour and state of individuals of the same population are repeatedly measured. Here, we test if variation in states predicts nest defence behaviour (a 'risky' behaviour) in a long-lived species, the barnacle goose Branta leucopsis. Adaptive models have predicted that an individual's residual reproductive value or 'asset' is an important state variable underlying variation in risk-taking behaviour. Hence, we investigate how nest defence varies as a function of time of the season and individual age, two state variables that can vary between and within individuals and determine asset. Repeated measures of nest defence towards a human intruder (flight initiation distance or FID) of females of known age were collected during 15 breeding seasons. Increasing values of FID represent increasing shyness. We found that females strongly and consistently differed in FID within- and between-years. As predicted by theory, females adjusted their behaviour to state by decreasing their FID with season and age. Decomposing these population patterns into within- and between-individual effects showed that the state-dependent change in FID was driven by individual plasticity in FID and that bolder females were more plastic than shyer females. This study shows that nest defence behaviour differs consistently among individuals and is adjusted to individual state in a direction predicted by adaptive personality theory

Improved detection of artifactual viral minority variants in high-throughput sequencing data

textabstractHigh-throughput sequencing (HTS) of viral samples provides important information on the presence of viral minority variants. However, detection and accurate quantification is limited by the capacity to distinguish biological from artificial variation. In this study, errors related to the Illumina HiSeq2000 library generation and HTS process were investigated by determining minority variant frequencies in an influenza A/WSN/1933(H1N1) virus reverse-genetics plasmid pool. Errors related to amplification and sequencing were determined using the same plasmid pool, by generation of infectious virus using reverse genetics followed by in duplo reverse-transcriptase PCR (RT-PCR) amplification and HTS in the same sequence run. Results showed that after "best practice" quality control (QC), within the plasmid pool, one minority variant with a frequency >0.5% was identified, while 84 and 139 were identified in the RT-PCR amplified samples, indicating RT-PCR amplification artificially increased variation. Detailed analysis showed that artifactual minority variants could be identified by two major technical characteristics: their predominant presence in a single read orientation and uneven distribution of mismatches over the length of the reads. We demonstrate that by addition of two QC steps 95% of the artifactual minority variants could be identified. When our analysis approach was applied to three clinical samples 68% of the initially identified minority variants were identified as artifacts. Our study clearly demonstrated that, without additional QC steps, overestimation of viral minority variants is very likely to occur, mainly as a consequence of the required RT-PCR amplification step. The improved ability to detect and correct for artifactual minority variants, increases data resolution and could aid both past and future studies incorporating HTS. The source code has been made available through Sourceforge (https://sourceforge.net/projects/mva-ngs)

Improved detection of artifactual viral minority variants in high-throughput sequencing data

High-throughput sequencing (HTS) of viral samples provides important information on the presence of viral minority variants. However, detection and accurate quantification is limited by the capacity to distinguish biological from artificial variation. In this study, errors related to the Illumina Hiseq2000 library generation and HTS process were investigated by determining minority variant frequencies in an influenza A/WSN/1933(H1N1) virus reversegenetics plasmid pool. Errors related to amplification and sequencing were determined using the same plasmid pool, by generation of infectious virus using reverse genetics followed by in duplo reverse-transcriptase PCR (RT-PCR) amplification and HTS in the same sequence run. Results showed that after 'best practice' quality control (QC), within the plasmid pool, 1 minority variant with a frequency >0.5% was identified, while 84 and 139 were identified in the RT-PCR amplified samples, indicating RT-PCR amplification artificially increased variation. Detailed analysis showed that artifactual minority variants could be identified by two major technical characteristics: their predominant presence in a single read orientation and uneven distribution of mismatches over the length of the reads. We demonstrate that by addi

Improved detection of artifactual viral minority variants in high-throughput sequencing data

High-throughput sequencing (HTS) of viral samples provides important information on the presence of viral minority variants. However, detection and accurate quantification is limited by the capacity to distinguish biological from artificial variation. In this study, errors related to the Illumina HiSeq2000 library generation and HTS process were investigated by determining minority variant frequencies in an influenza A/WSN/1933(H1N1) virus reverse-genetics plasmid pool. Errors related to amplification and sequencing were determined using the same plasmid pool, by generation of infectious virus using reverse genetics followed by in duplo reverse-transcriptase PCR (RT-PCR) amplification and HTS in the same sequence run. Results showed that after "best practice" quality control (QC), within the plasmid pool, one minority variant with a frequency >0.5% was identified, while 84 and 139 were identified in the RT-PCR amplified samples, indicating RT-PCR amplification artificially increased variation. Detailed analysis showed that artifactual minority variants could be identified by two major technical characteristics: their predominant presence in a single read orientation and uneven distribution of mismatches over the length of the reads. We demonstrate that by addition of two QC steps 95% of the artifactual minority variants could be identified. When our analysis approach was applied to three clinical samples 68% of the initially identified minority variants were identified as artifacts. Our study clearly demonstrated that, without additional QC steps, overestimation of viral minority variants is very likely to occur, m

Expression of the Serpin Serine Protease Inhibitor 6 Protects Dendritic Cells from Cytotoxic T Lymphocyte–Induced Apoptosis: Differential Modulation by T Helper Type 1 and Type 2 Cells

Dendritic cells (DCs) play a central role in the immune system as they drive activation of T lymphocytes by cognate interactions. However, as DCs express high levels of major histocompatibility complex class I, this intimate contact may also result in elimination of DCs by activated cytotoxic T lymphocytes (CTLs) and thereby limit induction of immunity. We show here that immature DCs are indeed susceptible to CTL-induced killing, but become resistant upon maturation with anti-CD40 or lipopolysaccharide. Protection is achieved by expression of serine protease inhibitor (SPI)-6, a member of the serpin family that specifically inactivates granzyme B and thereby blocks CTL-induced apoptosis. Anti-CD40 and LPS-induced SPI-6 expression is sustained for long periods of time, suggesting a role for SPI-6 in the longevity of DCs. Importantly, T helper 1 cells, which mature DCs and boost CTL immunity, induce SPI-6 expression and subsequent DC resistance. In contrast, T helper 2 cells neither induce SPI-6 nor convey protection, despite the fact that they trigger DC maturation with comparable efficiency. Our data identify SPI-6 as a novel marker for DC function, which protects DCs against CTL-induced apoptosis

Priors and Posteriors in Bayesian Timing of Divergence Analyses : The Age of Butterflies Revisited

The need for robust estimates of times of divergence is essential for downstream analyses, yet assessing this robustness is still rare. We generated a time-calibrated genus-level phylogeny of butterflies (Papilionoidea), including 994 taxa, up to 10 gene fragments and an unprecedented set of 12 fossils and 10 host-plant node calibration points. We compared marginal priors and posterior distributions to assess the relative importance of the former on the latter. This approach revealed a strong influence of the set of priors on the root age but for most calibrated nodes posterior distributions shifted from the marginal prior, indicating significant information in the molecular data set. Using a very conservative approach we estimated an origin of butterflies at 107.6 Ma, approximately equivalent to the latest Early Cretaceous, with a credibility interval ranging from 89.5 Ma (mid Late Cretaceous) to 129.5 Ma (mid Early Cretaceous). In addition, we tested the effects of changing fossil calibration priors, tree prior, different sets of calibrations and different sampling fractions but our estimate remained robust to these alternative assumptions. With 994 genera, this tree provides a comprehensive source of secondary calibrations for studies on butterflies.Peer reviewe

Facilitators and barriers for the implementation of exercise are medicine in routine clinical care in Dutch university medical centres:a mixed methodology study on clinicians' perceptions

Objectives Despite the many proven advantages of a physically active lifestyle in patient populations, prescription of exercise is currently not widely implemented in routine clinical practice. The aims of this study were twofold: (1) to assess perceptions of clinicians on the current practice of exercise is medicine (E=M) prescription in two Dutch university medical centres and (2) to determine their perceived barriers and facilitators for the implementation of E=M in routine clinical care in Dutch university medical centres. Design A mixed methodologies study, using both online questionnaires and semi-structured interviews. Setting Dutch university medical centres. Participants Clinicians working within the departments of medical oncology, orthopaedics and rehabilitation medicine of two university medical centres. Results Forty-five clinicians (response rate of 51%) completed the questionnaire, and 19 clinicians were interviewed. The results showed that even though clinicians had a positive attitude towards prescribing E=M, only a few reported to regularly prescribe E=M to their patients. The 52 identified facilitators and barriers for implementation of E=M were categorised into four main themes: (1) beliefs toward the implementation of E=M (eg, clinicians knowledge and skills, and social support), (2) factors related to the patient perspective (eg, patient priorities or motivation), (3) factors related to the referral options (eg, knowledge of and trust in local referral options) and (4) practical considerations when implementing E=M (eg, time constraints). Conclusions Our study showed that even though many clinicians have a positive attitude toward an active lifestyle, many are not prescribing E=M on a regular basis. In order for clinicians to effectively implement E=M, strategies should focus on increasing clinicians E=M referral skills, improving clinicians knowledge of E=M referral options and develop a support system to ensure that E=M is high on the priority list of clinicians

Implementing Individually Tailored Prescription of Physical Activity in Routine Clinical Care:Protocol of the Physicians Implement Exercise = Medicine (PIE=M) Development and Implementation Project

BACKGROUND: The prescription of physical activity (PA) in clinical care has been advocated worldwide. This "exercise is medicine" (E=M) concept can be used to prevent, manage, and cure various lifestyle-related chronic diseases. Due to several challenges, E=M is not yet routinely implemented in clinical care. OBJECTIVE: This paper describes the rationale and design of the Physicians Implement Exercise = Medicine (PIE=M) study, which aims to facilitate the implementation of E=M in hospital care. METHODS: PIE=M consists of 3 interrelated work packages. First, levels and determinants of PA in different patient and healthy populations will be investigated using existing cohort data. The current implementation status, facilitators, and barriers of E=M will also be investigated using a mixed-methods approach among clinicians of participating departments from 2 diverse university medical centers (both located in a city, but one serving an urban population and one serving a more rural population). Implementation strategies will be connected to these barriers and facilitators using a systematic implementation mapping approach. Second, a generic E=M tool will be developed that will provide tailored PA prescription and referral. Requirements for this tool will be investigated among clinicians and department managers. The tool will be developed using an iterative design process in which all stakeholders reflect on the design of the E=M tool. Third, we will pilot-implement the set of implementation strategies, including the E=M tool, to test its feasibility in routine care of clinicians in these 2 university medical centers. An extensive learning process evaluation will be performed among clinicians, department managers, lifestyle coaches, and patients using a mixed-methods design based on the RE-AIM framework. RESULTS: This project was approved and funded by the Dutch grant provider ZonMW in April 2018. The project started in September 2018 and continues until December 2020 (depending on the course of the COVID-19 crisis). All data from the first work package have been collected and analyzed and are expected to be published in 2021. Results of the second work package are described. The manuscript is expected to be published in 2021. The third work package is currently being conducted in clinical practice in 4 departments of 2 university medical hospitals among clinicians, lifestyle coaches, hospital managers, and patients. Results are expected to be published in 2021. CONCLUSIONS: The PIE=M project addresses the potential of providing patients with PA advice to prevent and manage chronic disease, improve recovery, and enable healthy ageing by developing E=M implementation strategies, including an E=M tool, in routine clinical care. The PIE=M project will result in a blueprint of implementation strategies, including an E=M screening and referral tool, which aims to improve E=M referral by clinicians to improve patients' health, while minimizing the burden on clinicians