Protocol: precision engineering of plant gene loci by homologous recombination cloning in Escherichia coli.

Plant genome sequence data now provide opportunities to conduct molecular genetic studies at the level of the whole gene locus and above. Such studies will be greatly facilitated by adopting and developing further the new generation of genetic engineering tools, based on homologous recombination cloning in Escherichia coli, which are free from the constraints imposed by the availability of suitably positioned restriction sites. Here we describe the basis for homologous recombination cloning in E. coli, the available tools and resources, together with a protocol for long range cloning and manipulation of an Arabidopsis thaliana gene locus, to create constructs co-ordinately driven by locus-specific regulatory elements.RIGHTS : This article is licensed under the BioMed Central licence at http://www.biomedcentral.com/about/license which is similar to the 'Creative Commons Attribution Licence'. In brief you may : copy, distribute, and display the work; make derivative works; or make commercial use of the work - under the following conditions: the original author must be given credit; for any reuse or distribution, it must be made clear to others what the license terms of this work are

Assessing the validity and reliability and determining cut-points of the Actiwatch 2 in measuring physical activity

Objective: The Actiwatch 2 (AW2) is a wrist-worn accelerometer typically used to measure sleep. Although it can measure physical activity, there is limited evidence supporting its validity. We assessed the validity and reliability of the AW2 to measure sedentary behavior and physical activity (light, moderate, vigorous intensities), and reported their respective count cut-points. Approach: Twenty-eight males and 22 females completed a task battery comprising three sedentary tasks and six randomized physical activity tasks at varying intensities, whilst wearing the AW2, a reference accelerometry device (Actigraph GT3X) and a cardiopulmonary gas analyzer on two separate occasions. Validity was assessed using correlations (AW2 counts versus GT3X counts and metabolic equivalent (MET) values), reliability using Bland–Altman analyses, and cut-points were determined using receiver operating characteristic (ROC) area under the curve (AUC) analyses. Main results: AW2 counts were positively correlated with GT3X counts (rho = 0.902, p < 0.001) and METs (rho = 0.900, p < 0.001). AW2-derived counts were comparable across independent assessment periods. Sedentary (AUC = 0.99, cut-point: 256 cpm) and vigorous activity (AUC = 0.95, cut-point: 720 cpm) were strongly characterized, and moderate activity (AUC = 0.66, cut-point: 418 cpm) was weakly characterized. Significance: The use of the AW2 in physical activity monitoring looks promising for sedentary behavior, moderate and vigorous activity, however, further validation is needed

Associations Between Self-Reported Sleep Duration and Mortality in Employed Individuals:Systematic Review and Meta-Analysis

Objective: Sleeping less or more than the 7-8 h has been associated with mortality in the general population, which encompasses diversity in employment status, age and community settings. Since sleep patterns of employed individuals may differ to those of their unemployed counterparts, the nature of their sleep-mortality relationship may vary. We therefore investigated the association between self-reported sleep duration and all-cause mortality (ACM) or cardiovascular disease mortality (CVDM) in employed individuals. Data sources: Based on Preferred Reporting Items for Systematic Reviews and Meta-Analyses, searches between January 1990 and May 2020 were conducted in PubMed, Web of Science and Scopus. Inclusion/exclusion criteria: Included were prospective cohort studies of 18–64-year-old disease-free employed persons with sleep duration measured at baseline, and cause of death recorded prospectively as the outcome. Gray literature, case-control or intervention design studies were excluded. Data Extraction: Characteristics of the studies, participants, and study outcomes were extracted. The quality and risk of bias were assessed using the Newcastle-Ottawa Scale. Data synthesis: The pooled relative risks (RR) with 95% confidence intervals (CI) were obtained with a random-effects model and results presented as forest plots. Heterogeneity and sensitivity analysis were assessed. Results: Shorter sleep duration (less than or equal to 6 h) was associated with a higher risk for (ACM) (RR: 1.16, 95% CI: 1.11 -1.22) and CVDM (RR: 1.26, 95% CI: 1.12 -1.41) compared to 7-8 h of sleep, with no significant heterogeneity. The association between longer sleep (greater than or equal to 8 h) and ACM (RR: 1.18, 95% CI:1.12 -1.23, P < 0.001) needs to be interpreted cautiously owing to high heterogeneity (I2 ¼ 86.0%, P < 0.001). Conclusion: Interventions and education programs targeting sleep health in the workplace may be warranted, based on our findings that employed individuals who report shorter sleep appear to have a higher risk for ACM and CVDM

Cardiovascular implications and physical activity in middle-aged and older adults with a history of COVID-19 (CV COVID)::a protocol for a randomised controlled trial

Background: The clinical manifestation of COVID-19 is associated with infection and inflammation of the lungs, but there is evidence to suggest that COVID-19 may also affect the structure and function of the cardiovascular system. At present, it is not fully understood to what extent COVID-19 impacts cardiovascular function in the short- and long-term following infection. The aim of the present study is twofold: (i) to define the effect of COVID-19 on cardiovascular function (i.e. arterial stiffness, cardiac systolic and diastolic function) in otherwise healthy individuals and (ii) to evaluate the effect of a home-based physical activity intervention on cardiovascular function in people with a history of COVID-19. Methods: This prospective, single-centre, observational study will recruit 120 COVID-19-vaccinated adult participants aged between 50 and 85 years, i.e. 80 with a history of COVID-19 and 40 healthy controls without a history of COVID-19. All participants will undergo baseline assessments including 12-lead electrocardiography, heart rate variability, arterial stiffness, rest and stress echocardiography with speckle tracking imaging, spirometry, maximal cardiopulmonary exercise testing, 7-day physical activity and sleep measures and quality of life questionnaires. Blood samples will be collected to assess the microRNA expression profiles, cardiac and inflammatory biomarkers, i.e. cardiac troponin T; N-terminal pro B-type natriuretic peptide; tumour necrosis factor alpha; interleukins 1, 6 and 10; C-reactive protein; d-dimer; and vascular endothelial growth factors. Following baseline assessments, COVID-19 participants will be randomised 1:1 into a 12-week home-based physical activity intervention aiming to increase their daily number of steps by 2000 from baseline. The primary outcome is change in left ventricular global longitudinal strain. Secondary outcomes are arterial stiffness, systolic and diastolic function of the heart, functional capacity, lung function, sleep measures, quality of life and well-being (depression, anxiety, stress and sleep efficiency). Discussion: The study will provide insights into the cardiovascular implications of COVID-19 and their malleability with a home-based physical activity intervention. Trial registration: ClinicalTrials.gov NCT05492552. Registered on 7 April 2022

Associations between fears related to safety during sleep and self-reported sleep in men and women living in a low-socioeconomic status setting

South Africans living in low socioeconomic areas have self-reported unusually long sleep durations (approximately 9–10 h). One hypothesis is that these long durations may be a compensatory response to poor sleep quality as a result of stressful environments. This study aimed to investigate whether fear of not being safe during sleep is associated with markers of sleep quality or duration in men and women. South Africans (n = 411, 25–50 y, 57% women) of African-origin living in an urban township, characterised by high crime and poverty rates, participated in this study. Participants are part of a larger longitudinal cohort study: Modelling the Epidemiologic Transition Study (METS)–Microbiome. Customised questions were used to assess the presence or absence of fears related to feeling safe during sleep, and the Epworth Sleepiness Scale, Pittsburgh Sleep Quality Index (PSQI) and Insomnia Severity Index were used to assess daytime sleepiness, sleep quality and insomnia symptom severity respectively. Adjusted logistic regression models indicated that participants who reported fears related to safety during sleep were more likely to report poor sleep quality (PSQI &gt; 5) compared to participants not reporting such fears and that this relationship was stronger among men than women. This is one of the first studies outside American or European populations to suggest that poor quality sleep is associated with fear of personal safety in low-SES South African adults

Colorectal Cancers from Distinct Ancestral Populations Show Variations in BRAF Mutation Frequency

It has been demonstrated for some cancers that the frequency of somatic oncogenic mutations may vary in ancestral populations. To determine whether key driver alterations might occur at different frequencies in colorectal cancer, we applied a high-throughput genotyping platform (OncoMap) to query 385 mutations across 33 known cancer genes in colorectal cancer DNA from 83 Asian, 149 Black and 195 White patients. We found that Asian patients had fewer canonical oncogenic mutations in the genes tested (60% vs Black 79% (P = 0.011) and White 77% (P = 0.015)), and that BRAF mutations occurred at a higher frequency in White patients (17% vs Asian 4% (P = 0.004) and Black 7% (P = 0.014)). These results suggest that the use of genomic approaches to elucidate the different ancestral determinants harbored by patient populations may help to more precisely and effectively treat colorectal cancer

Comparison between an African town and a neighbouring village shows delayed, but not decreased, sleep during the early stages of urbanisation

The well-established negative health outcomes of sleep deprivation, and the suggestion that availability of electricity may enable later bed times without compensating sleep extension in the morning, have stimulated interest in studying communities whose sleep pattern may resemble a pre-industrial state. Here, we describe sleep and activity in two neighbouring communities, one urban (Milange) and one rural (Tengua), in a region of Mozambique where urbanisation is an ongoing process. The two communities differ in the amount and timing of daily activity and of light exposure, with later bedtimes (≈1 h) associated with more evening and less daytime light exposure seen in the town of Milange. In contrast to previous reports comparing communities with and without electricity, sleep duration did not differ between Milange (7.28 h) and Tengua (7.23 h). Notably, calculated sleep quality was significantly poorer in rural Tengua than in Milange, and poor sleep quality was associated with a number of attributes more characteristic of rural areas, including more intense physical labour and less comfortable sleeping arrangements. Thus, whilst our data support the hypothesis that access to electricity delays sleep timing, the higher sleep quality in the urban population also suggests that some aspects of industrialisation are beneficial to sleep

Differences in Physiological Responses to Cardiopulmonary Exercise Testing in Adults With and Without Type 1 Diabetes: A Pooled Analysis

OBJECTIVE To investigate physiological responses to cardiopulmonary exercise (CPX) testing in adults with type 1 diabetes compared with age-, sex-, and BMI-matched control participants without type 1 diabetes.RESEARCH DESIGN AND METHODS We compared results from CPX tests on a cycle ergometer in individuals with type 1 diabetes and control participants without type 1 diabetes. Parameters were peak and threshold variables of VO2, heart rate, and power output. Differences between groups were investigated through restricted maximum likelihood modeling and post hoc tests. Differences between groups were explained by stepwise linear regressions (P < 0.05).RESULTS Among 303 individuals with type 1 diabetes (age 33 [interquartile range 22; 43] years, 93 females, BMI 23.6 [22; 26] kg/m2, HbA1c 6.9% [6.2; 7.7%] [52 (44; 61) mmol/mol]), VO2peak (32.55 [26.49; 38.72] vs. 42.67 ± 10.44 mL/kg/min), peak heart rate (179 [170; 187] vs. 184 [175; 191] beats/min), and peak power (216 [171; 253] vs. 245 [200; 300] W) were lower compared with 308 control participants without type 1 diabetes (all P < 0.001). Individuals with type 1 diabetes displayed an impaired degree and direction of the heart rate-to-performance curve compared with control participants without type 1 diabetes (0.07 [−0.75; 1.09] vs. 0.66 [−0.28; 1.45]; P < 0.001). None of the exercise physiological responses were associated with HbA1c in individuals with type 1 diabetes.CONCLUSIONS Individuals with type 1 diabetes show altered responses to CPX testing, which cannot be explained by HbA1c. Intriguingly, the participants in our cohort were people with recent-onset type 1 diabetes; heart rate dynamics were altered during CPX testing

Research Directions in the Clinical Implementation of Pharmacogenomics: An Overview of US Programs and Projects

Response to a drug often differs widely among individual patients. This variability is frequently observed not only with respect to effective responses but also with adverse drug reactions. Matching patients to the drugs that are most likely to be effective and least likely to cause harm is the goal of effective therapeutics. Pharmacogenomics (PGx) holds the promise of precision medicine through elucidating the genetic determinants responsible for pharmacological outcomes and using them to guide drug selection and dosing. Here we survey the US landscape of research programs in PGx implementation, review current advances and clinical applications of PGx, summarize the obstacles that have hindered PGx implementation, and identify the critical knowledge gaps and possible studies needed to help to address them

Defence Responses of Arabidopsis thaliana to Infection by Pseudomonas syringae Are Regulated by the Circadian Clock

The circadian clock allows plants to anticipate predictable daily changes in abiotic stimuli, such as light; however, whether the clock similarly allows plants to anticipate interactions with other organisms is unknown. Here we show that Arabidopsis thaliana (Arabidopsis) has circadian clock-mediated variation in resistance to the virulent bacterial pathogen Pseudomonas syringae pv. tomato DC3000 (Pst DC3000), with plants being least susceptible to infection in the subjective morning. We suggest that the increased resistance to Pst DC3000 observed in the morning in Col-0 plants results from clock-mediated modulation of pathogen associated molecular pattern (PAMP)-triggered immunity. Analysis of publicly available microarray data revealed that a large number of Arabidopsis defence-related genes showed both diurnal- and circadian-regulation, including genes involved in the perception of the PAMP flagellin which exhibit a peak in expression in the morning. Accordingly, we observed that PAMP-triggered callose deposition was significantly higher in wild-type plants inoculated with Pst DC3000 hrpA in the subjective morning than in the evening, while no such temporal difference was evident in arrhythmic plants. Our results suggest that PAMP-triggered immune responses are modulated by the circadian clock and that temporal regulation allows plants to anticipate and respond more effectively to pathogen challenges in the daytime