Single-Molecule Studies of Protein Folding with Optical Tweezers.

Manipulation of individual molecules with optical tweezers provides a powerful means of interrogating the structure and folding of proteins. Mechanical force is not only a relevant quantity in cellular protein folding and function, but also a convenient parameter for biophysical folding studies. Optical tweezers offer precise control in the force range relevant for protein folding and unfolding, from which single-molecule kinetic and thermodynamic information about these processes can be extracted. In this review, we describe both physical principles and practical aspects of optical tweezers measurements and discuss recent advances in the use of this technique for the study of protein folding. In particular, we describe the characterization of folding energy landscapes at high resolution, studies of structurally complex multidomain proteins, folding in the presence of chaperones, and the ability to investigate real-time cotranslational folding of a polypeptide.

Consensus on a netball video analysis framework of descriptors and definitions by the netball video analysis consensus group.

Using an expert consensus-based approach, a netball video analysis consensus (NVAC) group of researchers and practitioners was formed to develop a video analysis framework of descriptors and definitions of physical, technical and contextual aspects for netball research. The framework aims to improve the consistency of language used within netball investigations. It also aims to guide injury mechanism reporting and identification of injury risk factors. The development of the framework involved a systematic review of the literature and a Delphi process. In conjunction with commercially used descriptors and definitions, 19 studies were used to create the initial framework of key descriptors and definitions in netball. In a two round Delphi method consensus, each expert rated their level of agreement with each of the descriptors and associated definition on a 5-point Likert scale (1-strongly disagree; 2-somewhat disagree; 3-neither agree nor disagree; 4-somewhat agree; 5-strongly agree). The median (IQR) rating of agreement was 5.0 (0.0), 5.0 (0.0) and 5.0 (0.0) for physical, technical and contextual aspects, respectively. The NVAC group recommends usage of the framework when conducting video analysis research in netball. The use of descriptors and definitions will be determined by the nature of the work and can be combined to incorporate further movements and actions used in netball. The framework can be linked with additional data, such as injury surveillance and microtechnology data.

Evaluation of Hydrangea Cultivars for Tolerance Against Root Rot Caused by Fusarium oxysporum.

Root rot caused by Fusarium oxysporum Schltdl. is a newly identified disease in oakleaf hydrangea. Some cultivars such as Pee Wee and Queen of Hearts grown in pot-in-pot container systems showed root rot symptoms after late spring frost in May 2018 with 40 and 60% incidence in the infected nursery, respectively. This experiment was carried out to evaluate the tolerance among different hydrangea cultivars against root rot caused by F. oxysporum. Fifteen hydrangea cultivars from four different species were selected, and rooted cuttings were prepared from new spring flushes. Twelve plants from each cultivar were transplanted in a 1-gallon pot. Half of transplanted plants (six single plants) were inoculated by drenching 150 ml of F. oxysporum conidial suspension to maintain the concentration of 1 × 106 conidia/ml. Half of the plants remain noninoculated (control) and were drenched with sterile water. After 4 months, root rot was assessed using a scale of 0 to 100% root area affected, and recovery of F. oxysporum was recorded by plating 1-cm root sections in Fusarium selective medium. Fusaric acid (FA) and mannitol were extracted from the roots of inoculated and noninoculated plants to see the effect and role on pathogenesis. Further, mannitol concentration was analyzed using absorption wavelength in a spectrophotometer, and FA was analyzed using high-performance liquid chromatography (HPLC). Results indicated that no cultivars were resistant to F. oxysporum. Cultivars from Hydrangea arborescens, H. macrophylla, and H. paniculata were more tolerant to F. oxysporum compared to cultivars from H. quercifolia. Among H. quercifolia, cultivars Snowflake, John Wayne, and Alice were more tolerant to F. oxysporum.

PAs in the Republic of Ireland.

ABSTRACT: The physician associate/assistant (PA) profession was introduced into the Republic of Ireland following a 2-year pilot project with the Irish Department of Health between 2015 and 2017. Four PAs from North America were recruited into four designated surgical services at a large teaching hospital in Dublin. To date, the PA numbers are small in Ireland, with one university, in Dublin, running the program and 61 graduates working mostly in the hospital setting, with a small number in primary care. The cautious introduction of PAs partly is due to a delay in follow-up from the Department of Health after the pilot project and in the university's decision to increase the student intake slowly to ensure all graduates secure employment.

Management of Phytophthora cinnamomi Using Fungicides and Host Plant Defense Inducers Under Drought Conditions: A Case Study of Flowering Dogwood.

Phytophthora cinnamomi is considered one of the most destructive pathogens of ornamental crops. Different fungicides and host plant defense inducers were tested for their efficacy in managing Phytophthora root rot in drought conditions. In this study, the drought conditions were maintained by evaluating the moisture-holding capacity of the pine bark in a 10.2-cm nursery container. Four controls and nine different treatments were used in two trials for this greenhouse study. All treatments were drench-applied as a preventative or curative treatment. Seedlings were artificially inoculated with P. cinnamomi. Regular irrigation was carried out using overhead irrigation for 1 month after inoculation. Irrigation was regulated by drip irrigation after the first month. A moisture level of 15% to 18% of total moisture-holding capacity was maintained using the gravimetric method throughout the drought period. Physiological parameters of the seedlings were recorded a week after seedlings were drought stressed. In both trials of preventative and curative treatments, all treatments were able to suppress the disease significantly except Orkestra Intrinsic. Orkestra Intrinsic had a disease severity statistically similar to the inoculated and stressed control in trial 1 of the curative treatment. Net photosynthesis, stomatal conductance, and leaf moisture potential were significantly greater in seedlings treated with Subdue MAXX, Signature Xtra, and Empress Intrinsic in both trials of preventative and curative treatments. Effective quantum yield of Photosystem II was significantly lower in the inoculated stressed control in both trials of preventative and curative treatments. Net chlorophyll content through the SPAD meter showed higher values for Subdue MAXX treated seedlings compared with the noninoculated nonstressed controls in trial 1 as both a curative and preventative application. In trial 2, Subdue MAXX and Signature Xtra were the best curative treatments, whereas Empress Intrinsic, Interface, and Subdue MAXX were the best preventative treatments for higher chlorophyll content. This case study will help growers perform successful management of Phytophthora root rot in woody ornamental crops during drought or water deficit conditions.

Correction to: Changes in relative fit of human heat stress indices to cardiovascular, respiratory, and renal hospitalizations across five Australian urban populations.

The authors of the article would like to bring the following correction/corrigendum to attention: When recently investigating future changes in heat stress indices, we discovered an error in the use of the heatwave indices we compared in Goldie et al. (2017).

Changes in relative fit of human heat stress indices to cardiovascular, respiratory, and renal hospitalizations across five Australian urban populations.

Various human heat stress indices have been developed to relate atmospheric measures of extreme heat to human health impacts, but the usefulness of different indices across various health impacts and in different populations is poorly understood. This paper determines which heat stress indices best fit hospital admissions for sets of cardiovascular, respiratory, and renal diseases across five Australian cities. We hypothesized that the best indices would be largely dependent on location. We fit parent models to these counts in the summers (November-March) between 2001 and 2013 using negative binomial regression. We then added 15 heat stress indices to these models, ranking their goodness of fit using the Akaike information criterion. Admissions for each health outcome were nearly always higher in hot or humid conditions. Contrary to our hypothesis that location would determine the best-fitting heat stress index, we found that the best indices were related largely by health outcome of interest, rather than location as hypothesized. In particular, heatwave and temperature indices had the best fit to cardiovascular admissions, humidity indices had the best fit to respiratory admissions, and combined heat-humidity indices had the best fit to renal admissions. With a few exceptions, the results were similar across all five cities. The best-fitting heat stress indices appear to be useful across several Australian cities with differing climates, but they may have varying usefulness depending on the outcome of interest. These findings suggest that future research on heat and health impacts, and in particular hospital demand modeling, could better reflect reality if it avoided "all-cause" health outcomes and used heat stress indices appropriate to specific diseases and disease groups.

Opioid use disorder: A public health emergency.

The treatment of patients with acute and chronic pain not attributed to cancer or end-of-life conditions is a challenge for many clinicians. Although CDC guidelines that focus on the primary care setting have provided critical recommendations, evidence-based guidance is lacking on optimal duration of opioid treatment for postoperative and acute care in specialty settings. Over the last 2 decades, the liberal use of opioids has resulted in many unintended consequences, including dependence and abuse, illicit distribution of legally and illegally prescribed opioid medication, progression to IV heroin and an epidemic of overdoses, and most recently an increase in the incidence of HIV among patients sharing syringes, frequently in communities with historically low HIV rates. This article analyzes these complex issues and proposes strategies to help clinicians improve patient care through education and responsible prescribing.

Atypical chest discomfort in a 36-year-old man.

This article focuses on aortic aneurysms and the evaluation of patients with atypical chest pain. A complete and accurate family history is essential. Clinicians should pay close attention to persistence of symptoms and age in patients with atypical chest discomfort who do not have routine confirmatory cardiac findings.

Ribosomal proteins L7 and L8 function in concert with six A3 assembly factors to propagate assembly of domains I and II of 25S rRNA in yeast 60S ribosomal subunits.

Ribosome biogenesis is a complex multistep process that involves alternating steps of folding and processing of pre-rRNAs in concert with assembly of ribosomal proteins. Recently, there has been increased interest in the roles of ribosomal proteins in eukaryotic ribosome biogenesis in vivo, focusing primarily on their function in pre-rRNA processing. However, much less is known about participation of ribosomal proteins in the formation and rearrangement of preribosomal particles as they mature to functional subunits. We have studied ribosomal proteins L7 and L8, which are required for the same early steps in pre-rRNA processing during assembly of 60S subunits but are located in different domains within ribosomes. Depletion of either leads to defects in processing of 27SA(3) to 27SB pre-rRNA and turnover of pre-rRNAs destined for large ribosomal subunits. A specific subset of proteins is diminished from these residual assembly intermediates: six assembly factors required for processing of 27SA(3) pre-rRNA and four ribosomal proteins bound to domain I of 25S and 5.8S rRNAs surrounding the polypeptide exit tunnel. In addition, specific sets of ribosomal proteins are affected in each mutant: In the absence of L7, proteins bound to domain II, L6, L14, L20, and L33 are greatly diminished, while proteins L13, L15, and L36 that bind to domain I are affected in the absence of L8. Thus, L7 and L8 might establish RNP structures within assembling ribosomes necessary for the stable association and function of the A(3) assembly factors and for proper assembly of the neighborhoods containing domains I and II.

The latent symptom structure of the Beck depression inventory: second edition in Latina pregnant women.

Pregnancy represents a unique period of time when women are at an increased risk of developing depression. Although the Beck Depression Inventory-Second Edition (BDI-II) is one of the most widely used self-report measures of depression symptomology, its psychometric properties and underlying factor structures have not been determined for antenatal women and among Latinas. The current study evaluated the latent symptom structure of the BDI-II in a community-based sample of Latina pregnant women (N = 217) identified to be at high risk for depression. Exploratory factor analyses were used to identify underlying salient individual item loadings for two- and three-factor models. Confirmatory factor analyses then examined several different indices to determine the best model fit. Examination of exploratory and confirmatory factor analyses supports a three-factor oblique structure of the BDI-II composed of Cognitive-Affective, Somatic, and Pregnancy Symptoms. The three-factor model provides clinicians with the ability to target specific constellations of depressive symptoms instead of relying on the BDI-II total score that represents the overall severity of depression in this population.

Time to Pivot: A Guide to Holistic Admissions.

ABSTRACT: As an ever-increasing number of physician assistant (PA) programs moves toward holistic admissions, a better understanding of how to achieve their stated admission goals becomes more important. With the June 2023 US Supreme Court decision effectively ending affirmative action in higher education, navigating holistic admissions is now an even greater challenge. In this article, the PA Education Association's Presidents Commission offers a guide for programs to use in implementing holistic admissions at their institutions and key considerations. Is the process mission-driven? Does it follow principles of quality improvement and incorporate ongoing assessment of that process? Using data can be a constructive and insightful way to inform the process. The authors hope that tools, resources, and recommendations offered in this article will serve as valuable resources for any program attempting to institute or improve its holistic admissions process.

True functional ability of chronic stroke patients.

BACKGROUND: There is a paucity of information regarding visuospatial (VS) and visuomotor (VM) task performance in patients with chronic right fronto-parietal lobe stroke, as the majority of knowledge to date in this realm has been gleaned from acute stroke patients. The goal of this paper is to determine how VS and VM performance in chronic stroke patients compare to the performance of healthy participants. METHODS: Nine patients with stroke involving the right fronto-parietal region were evaluated against match controls on neuropsychological tests and a computerized visuomotor assessment task. RESULTS: Initial evaluation indicated that performance between participant groups were relatively similar on all measures. However, an in-depth analysis of variability revealed observable differences between participant groups. In addition, large effect sizes were also observed supporting the theory that using only conventional examination (e.g., p-values) measures may result in miss-identifying crucial stroke-related differences. CONCLUSION: Through conventional evaluation methods it would appear that the chronic stroke participants had made significant functional gains relatively to a control group many years post-stroke. It was shown that the type of evaluation used is essential to identifying group differences. Thus, supplementary methods of evaluation are required to unmask the true functional ability of individuals many years post-stroke.

Gram-scale synthesis and efficient purification of 13C-labeled levoglucosan from 13C glucose.

(13)C-Labeled levoglucosan has been synthesized and purified in good yield, and on the gram scale in one step from commercially available (13)C glucose. This one-step protocol uses 2-chloro-1,3-dimethylimidazolinium chloride that serves to selectively activate the anomeric carbon toward substitution reactions. The labeled glucose is then smoothly converted to the anhydroglucose. Purification is efficiently achieved on large scale by chromatography on silica gel.

A survey exploring factors affecting employment of physician associates in Ireland.

BACKGROUND: In the Republic of Ireland, the employment of physician associates (PAs) is growing. Following a pilot project in a hospital setting, PAs are now employed across primary and secondary care in public and private sectors. Most of the Irish PA graduates are working in hospital settings. AIMS: The aim of the study was to explore factors which supported or inhibited the employment of PAs in Irish hospital settings and the perceived supports or challenges for potential employers in recruiting PAs. METHODS: An online survey gathered data via human resources departments of public and private hospitals, with a 25% response rate. RESULTS: Similar to previous studies, the barriers included the lack of recognition and regulation of the role and the small number of PAs to fill available posts. Enablers, which influenced the employment of PAs, included improving workflow, continuity of care and helping to address junior doctors' working hours. CONCLUSIONS: Our data suggests that there is a keen interest and willingness to employ PAs and there is great potential to expand the role in Irish healthcare. However, there are some key issues around funding and recognition to be addressed at government level for this profession to highlight its worth.

Meeting the Accreditation Standard for Diversity.

ABSTRACT: While increasing diversity has been an ongoing concern in physician assistant (PA) education, there is now a concentrated focus on diversity, equity, inclusion, and social justice, elevating these to top priorities. To achieve the goal of diversifying the PA workforce, PA programs, with their institution's support, must systematically inculcate strategies for overcoming and dismantling barriers against students of color and students underrepresented in medicine (URiM). These strategies should disrupt the status quo and expand structural processes that ensure successful diversification of students, especially URiM students, LGBTQ students, students from medically underserved areas, and first-generation college students.

A Multimodal Intervention for Reducing Unnecessary Repeat Glycated Hemoglobin Testing.

OBJECTIVES: Reducing unnecessary tests that do not enhance quality can promote health-care value. Glycated hemoglobin (A1C) is often ordered at a frequency exceeding the recommendation of once every 3 months. We conducted a quality improvement (QI) initiative aimed to reduce unnecessary repeat testing by 75% at a tertiary care academic hospital. METHODS: A retrospective baseline analysis was conducted on laboratory data from 2019 that enumerated unnecessary A1C tests, defined as repeat tests ordered within 60 days. A multifaceted change intervention with iterative plan-do-study-act cycles was introduced in March 2019 to educate providers and to automatically cancel A1C tests requested within 60 days. Monthly totals of A1C testing processed were plotted on statistical process control charts. RESULTS: In 2019, 11% of all A1C tests ordered were unnecessary. Between March 2020 and January 2021, 11% of the tests (N=14,247 tests) were unnecessary, of which 84% were cancelled with our intervention. Providers in cardiology and nephrology accounted for over half (55%) of the unnecessary tests ordered. CONCLUSIONS: A 2-pronged approach informed by root-cause analysis, and comprised of gatekeeping and provider education, can effectively promote resource stewardship for reducing unnecessary A1C testing.

Enhanced multi-year predictability after El Niño and La Niña events.

Several aspects of regional climate including near-surface temperature and precipitation are predictable on interannual to decadal time scales. Despite indications that some climate states may provide higher predictability than others, previous studies analysing decadal predictions typically sample a variety of initial conditions. Here we assess multi-year predictability conditional on the phase of the El Niño-Southern Oscillation (ENSO) at the time of prediction initialisation. We find that predictions starting with El Niño or La Niña conditions exhibit higher skill in predicting near-surface air temperature and precipitation multiple years in advance, compared to predictions initialised from neutral ENSO conditions. This holds true in idealised prediction experiments with the Community Climate System Model Version 4 and to a lesser extent also real-world predictions using the Community Earth System Model and a multi-model ensemble of hindcasts contributed to the Coupled Model Intercomparison Project Phase 6 Decadal Climate Prediction Project. This enhanced predictability following ENSO events is related to phase transitions as part of the ENSO cycle, and related global teleconnections. Our results indicate that certain initial states provide increased predictability, revealing windows of opportunity for more skillful multi-year predictions.

Two haplotype-resolved genomes reveal important flower traits in bigleaf hydrangea (Hydrangea macrophylla) and insights into Asterid evolution.

The Hydrangea genus belongs to the Hydrangeaceae family, in the Cornales order of flowering plants, which early diverged among the Asterids, and includes several species that are commonly used ornamental plants. Of them, Hydrangea macrophylla is one of the most valuable species in the nursery trade, yet few genomic resources are available for this crop or closely related Asterid species. Two high-quality haplotype-resolved reference genomes of hydrangea cultivars 'Veitchii' and 'Endless Summer' [highest quality at 2.22 gigabase pairs (Gb), 396 contigs, N50 22.8 megabase pairs (Mb)] were assembled and scaffolded into the expected 18 pseudochromosomes. Utilizing the newly developed high-quality reference genomes along with high-quality genomes of other related flowering plants, nuclear data were found to support a single divergence point in the Asterids clade where both the Cornales and Ericales diverged from the euasterids. Genetic mapping with an F1 hybrid population demonstrated the power of linkage mapping combined with the new genomic resources to identify the gene for inflorescence shape, CYP78A5 located on chromosome 4, and a novel gene, BAM3 located on chromosome 17, for causing double flower. Resources developed in this study will not only help to accelerate hydrangea genetic improvement but also contribute to understanding the largest group of flowering plants, the Asterids.

Novel and Engineered Type II CRISPR Systems from Uncultivated Microbes with Broad Genome Editing Capability.

Type II Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-Cas9 nucleases have been extensively used in biotechnology and therapeutics. However, many applications are not possible owing to the size, targetability, and potential off-target effects associated with currently known systems. In this study, we identified thousands of CRISPR type II effectors by mining an extensive, genome-resolved metagenomics database encompassing hundreds of thousands of microbial genomes. We developed a high-throughput pipeline that enabled us to predict tracrRNA sequences, to design single guide RNAs, and to demonstrate nuclease activity in vitro for 41 newly described subgroups. Active systems represent an extensive diversity of protein sequences and guide RNA structures and require diverse protospacer adjacent motifs (PAMs) that collectively expand the known targeting capability of current systems. Several nucleases showed activity levels comparable to or significantly higher than SpCas9, despite being smaller in size. In addition, top systems exhibited low levels of off-target editing in mammalian cells, and PAM-interacting domain engineered chimeras further expanded their targetability. These newly discovered nucleases are attractive enzymes for translation into many applications, including therapeutics.