Stiffness transitions in new walls post-cell division differ between Marchantia polymorpha gemmae and Arabidopsis thaliana leaves.

Plant morphogenesis is governed by the mechanics of the cell wall-a stiff and thin polymeric box that encloses the cells. The cell wall is a highly dynamic composite material. New cell walls are added during cell division. As the cells continue to grow, the properties of cell walls are modulated to undergo significant changes in shape and size without breakage. Spatial and temporal variations in cell wall mechanical properties have been observed. However, how they relate to cell division remains an outstanding question. Here, we combine time-lapse imaging with local mechanical measurements via atomic force microscopy to systematically map the cell wall's age and growth, with their stiffness. We make use of two systems, Marchantia polymorpha gemmae, and Arabidopsis thaliana leaves. We first characterize the growth and cell division of M. polymorpha gemmae. We then demonstrate that cell division in M. polymorpha gemmae results in the generation of a temporary stiffer and slower-growing new wall. In contrast, this transient phenomenon is absent in A. thaliana leaves. We provide evidence that this different temporal behavior has a direct impact on the local cell geometry via changes in the junction angle. These results are expected to pave the way for developing more realistic plant morphogenetic models and to advance the study into the impact of cell division on tissue growth.

Risk factors and outcomes of melanoma in children and adolescents: A retrospective multicenter study.

BACKGROUND: Pediatric melanoma presents with distinct clinical features compared to adult disease. OBJECTIVE: Characterize risk factors and negative outcomes in pediatric melanoma. METHODS: Multicenter retrospective study of patients under 20 years diagnosed with melanoma between January 1, 1995 and June 30, 2015 from 11 academic medical centers. RESULTS: Melanoma was diagnosed in 317 patients, 73% of whom were diagnosed in adolescence (age ≥11). Spitzoid (31%) and superficial spreading (26%) subtypes were most common and 11% of cases arose from congenital nevi. Sentinel lymph node biopsy was performed in 68% of cases and positive in 46%. Fatality was observed in 7% of cases. Adolescent patients with melanoma were more likely to have family history of melanoma (P = .046) compared to controls. LIMITATIONS: Retrospective nature, cohort size, control selection, and potential referral bias. CONCLUSION: Pediatric melanoma has diverse clinical presentations. Better understanding of these cases and outcomes may facilitate improved risk stratification of pediatric melanoma.

A case of STING-associated vasculopathy with onset in infancy with novel STING1 variant.

STING-associated vasculopathy with onset in infancy (SAVI) is a rare, monogenic interferonopathy caused by gain-of-function variants in STING1 (TMEM173) characterized by systemic inflammation, cutaneous vasculopathy, and interstitial lung disease. We report a case of SAVI attributed to a novel STING1 p.R284T variant who demonstrated characteristic cutaneous features including telangiectasias, livedo and acrocyanotic changes on face and extremities, as well as saddle nose deformity, failure to thrive, inflammatory arthritis and notable lack of pulmonary disease or autoantibody positivity. Due to the risk for progressive and irreversible lung and tissue damage and evolving therapeutic landscape involving the use of Janus kinase inhibitors, it is critical to recognize variable clinical phenotypes to diagnose and consider treatment options for SAVI patients early in their disease course.

Rat Hepatitis E Virus in Norway Rats, Ontario, Canada, 2018-2021.

We tested liver samples from 372 Norway rats (Rattus norvegicus) from southern Ontario, Canada, during 2018-2021 to investigate presence of hepatitis E virus infection. Overall, 21 (5.6%) rats tested positive for the virus. Sequence analysis demonstrated all infections to be rat hepatitis E virus (Rocahepevirus ratti genotype C1).

Class I HDAC inhibitor entinostat synergizes with PLK1 inhibitors in MYC-amplified medulloblastoma cells.

PURPOSE: We and others have demonstrated that MYC-amplified medulloblastoma (MB) cells are susceptible to class I histone deacetylase inhibitor (HDACi) treatment. However, single drug treatment with HDACi has shown limited clinical efficacy. We hypothesized that addition of a second compound acting synergistically with HDACi may enhance efficacy. METHODS: We used a gene expression dataset to identify PLK1 as a second target in MB cells and validated the relevance of PLK1 in MB. We measured cell metabolic activity, viability, and cycle progression in MB cells after treatment with PLK1-specific inhibitors (PLK1i). Chou-Talalay synergy calculations were used to determine the nature of class I HDACi entinostat and PLK1i interaction which was validated. Finally, the clinical potential of the combination was assessed in the in vivo experiment. RESULTS: MYC-amplified tumor cells are highly sensitive towards treatment with ATP-competitive PLK1i as a monotherapy. Entinostat and PLK1i in combination act synergistically in MYC-driven MB cells, exerting cytotoxic effects at clinically relevant concentrations. The downstream effect is exerted via MYC-related pathways, pointing out the potential of MYC amplification as a clinically feasible predictive biomarker for patient selection. While entinostat significantly extended survival of mice implanted with orthotopic MYC-amplified MB PDX, there was no evidence of the improvement of survival when treating the animals with the combination. CONCLUSION: The combination of entinostat and PLK1i showed synergistic interaction in vitro, but not in vivo. Therefore, further screening of blood-brain barrier penetrating PLK1i is warranted to determine the true potential of the combination as no on-target activity was observed after PLK1i volasertib treatment in vivo.

Ectoparasitic mites exert non-consumptive effects on the larvae of a fruit fly host.

The mere presence of predators or parasites can negatively impact the fitness of prey or hosts. Exposure to predators during an organism's development can have deleterious effects on juvenile survival and the subsequent adult stage. Currently, it is unknown if parasites have analogous impacts on host larval stages and whether these effects carry over into other subsequent life stages. However, parasites may be exerting widespread yet underestimated non-consumptive effects (NCEs). We tested if Drosophila nigrospiracula larvae avoid pupating near mite cues (caged Macrocheles subbadius) in arena experiments, and measured the rate of pupation in arenas with mites and arenas without mites. Larvae disproportionately pupated on the side of arenas that lacked mite cues. Furthermore, fewer larvae successfully pupated in arenas containing mites cues compared to arenas without mite cues. We found that ectoparasitic mites exert NCEs on Drosophila larvae, even though the larval stage is not susceptible to infection. We discuss these results in the context of parasite impacts on host population growth in an infectious world.

Determining Clinical Judgment Among Emergency Nurses During a Complex Simulation.

INTRODUCTION: Clinical judgment is imperative for the emergency nurse caring for the acutely ill patients often seen in the emergency department. Without optimal clinical judgment in the emergency department, patients are at risk of medical errors and a failure to rescue. METHODS: A descriptive observational approach using the Lasater Clinical Judgment Rubric evaluated nurses during a task that required recognition of clinical signs of deterioration and appropriate clinical care for simulated patients. RESULTS: A total of 18 practicing emergency nurses completed only 44.6% of the patient assessments leading to low levels of clinical judgment throughout the simulation. Nurses expressed 4 levels of clinical judgment: exemplary (n = 1), accomplishing (n = 6), developing (n = 9), and beginning (n = 2). On average, nurses completed 69% of required tasks. DISCUSSION: Assessments were completed less than half the time, demonstrating a breakdown in the noticing phase of clinical judgment. The nurses shifted to task completion focus with minimal use of clinical judgment. As the nurses remained task oriented, several medication and medical errors were noted while caring for the simulated patients. Experience and education did not influence observed clinical judgment among the participants. Given the extreme demands placed on the emergency nurse, it cannot be assumed that nurses have developed or can use clinical judgment when caring for their patients. Time and training targeting clinical judgment are essential for emergency nurse development.

Engineered Cytochrome P450-Catalyzed Oxidative Biaryl Coupling Reaction Provides a Scalable Entry into Arylomycin Antibiotics.

We report herein the first example of a cytochrome P450-catalyzed oxidative carbon-carbon coupling process for a scalable entry into arylomycin antibiotic cores. Starting from wild-type hydroxylating cytochrome P450 enzymes and engineered Escherichia coli, a combination of enzyme engineering, random mutagenesis, and optimization of reaction conditions generated a P450 variant that affords the desired arylomycin core 2d in 84% assay yield. Furthermore, this process was demonstrated as a viable route for the production of the arylomycin antibiotic core on the gram scale. Finally, this new entry affords a viable, scalable, and practical route for the synthesis of novel Gram-negative antibiotics.

Epitope profiling using computational structural modelling demonstrated on coronavirus-binding antibodies.

Identifying the epitope of an antibody is a key step in understanding its function and its potential as a therapeutic. Sequence-based clonal clustering can identify antibodies with similar epitope complementarity, however, antibodies from markedly different lineages but with similar structures can engage the same epitope. We describe a novel computational method for epitope profiling based on structural modelling and clustering. Using the method, we demonstrate that sequence dissimilar but functionally similar antibodies can be found across the Coronavirus Antibody Database, with high accuracy (92% of antibodies in multiple-occupancy structural clusters bind to consistent domains). Our approach functionally links antibodies with distinct genetic lineages, species origins, and coronavirus specificities. This indicates greater convergence exists in the immune responses to coronaviruses than is suggested by sequence-based approaches. Our results show that applying structural analytics to large class-specific antibody databases will enable high confidence structure-function relationships to be drawn, yielding new opportunities to identify functional convergence hitherto missed by sequence-only analysis.

Incorporating NOE-Derived Distances in Conformer Generation of Cyclic Peptides with Distance Geometry.

Nuclear magnetic resonance (NMR) data from NOESY (nuclear Overhauser enhancement spectroscopy) and ROESY (rotating frame Overhauser enhancement spectroscopy) experiments can easily be combined with distance geometry (DG) based conformer generators by modifying the molecular distance bounds matrix. In this work, we extend the modern DG based conformer generator ETKDG, which has been shown to reproduce experimental crystal structures from small molecules to large macrocycles well, to include NOE-derived interproton distances. In noeETKDG, the experimentally derived interproton distances are incorporated into the distance bounds matrix as loose upper (or lower) bounds to generate large conformer sets. Various subselection techniques can subsequently be applied to yield a conformer bundle that best reproduces the NOE data. The approach is benchmarked using a set of 24 (mostly) cyclic peptides for which NOE-derived distances as well as reference solution structures obtained by other software are available. With respect to other packages currently available, the advantages of noeETKDG are its speed and that no prior force-field parametrization is required, which is especially useful for peptides with unnatural amino acids. The resulting conformer bundles can be further processed with the use of structural refinement techniques to improve the modeling of the intramolecular nonbonded interactions. The noeETKDG code is released as a fully open-source software package available at www.github.com/rinikerlab/customETKDG.

Wastewater surveillance for SARS-CoV-2 in a small coastal community: Effects of tourism on viral presence and variant identification among low prevalence populations.

Wastewater-based epidemiology has been used to measure SARS-CoV-2 prevalence in cities worldwide as an indicator of community health, however, few longitudinal studies have followed SARS-CoV-2 in wastewater in small communities from the start of the pandemic or evaluated the influence of tourism on viral loads. Therefore the objective of this study was to use measurements of SARS-CoV-2 in wastewater to monitor viral trends and variants in a small island community over a twelve-month period beginning May 1, 2020, before the community re-opened to tourists. Wastewater samples were collected weekly and analyzed to detect and quantify SARS-CoV-2 genome copies. Sanger sequencing was used to determine genome sequences from total RNA extracted from wastewater samples positive for SARS-CoV-2. Visitor data was collected from the local Chamber of Commerce. We performed Poisson and linear regression to determine if visitors to the Cedar Key Chamber of Commerce were positively associated with SARS-CoV-2-positive wastewater samples and the concentration of SARS-CoV-2 RNA. Results indicated that weekly wastewater samples were negative for SARS-CoV-2 until mid-July when positive samples were recorded in four of five consecutive weeks. Additional positive results were recorded in November and December 2020, as well as January, March, and April 2021. Tourism data revealed that the SARS-CoV-2 RNA concentration in wastewater increased by 1.06 Log10 genomic copies/L per 100 tourists weekly. Sequencing from six positive wastewater samples yielded two complete sequences of SARS-CoV-2, two overlapping sequences, and two low yield sequences. They show arrival of a new variant SARS-CoV-2 in January 2021. Our results demonstrate the utility of wastewater surveillance for SARS-CoV-2 in a small community. Wastewater surveillance and viral genome sequencing suggest that population mobility likely plays an important role in the introduction and circulation of SARS-CoV-2 variants among communities experiencing high tourism and who have a small population size.

Identification of transcription factors controlling floral morphology in wild Petunia species with contrasting pollination syndromes.

Adaptation to different pollinators is an important driver of speciation in the angiosperms. Genetic approaches such as QTL mapping have been successfully used to identify the underlying speciation genes. However, these methods are often limited by widespread suppression of recombination due to divergence between species. While the mutations that caused the interspecific differences in floral color and scent have been elucidated in a variety of plant genera, the genes that are responsible for morphological differences remain mostly unknown. Differences in floral organ length determine the pollination efficiency of hawkmoths and hummingbirds, and therefore the genes that control these differences are potential speciation genes. Identifying such genes is challenging, especially in non-model species and when studying complex traits for which little prior genetic and biochemical knowledge is available. Here we combine transcriptomics with detailed growth analysis to identify candidate transcription factors underlying interspecific variation in the styles of Petunia flowers. Starting from a set of 2284 genes, stepwise filtering for expression in styles, differential expression between species, correlation with growth-related traits, allele-specific expression in interspecific hybrids, and/or high-impact polymorphisms resulted in a set of 43 candidate speciation genes. Validation by virus-induced gene silencing identified two MYB transcription factors, EOBI and EOBII, that were previously shown to regulate floral scent emission, a trait associated with pollination by hawkmoths.

Mechanobiology of cell division in plant growth.

Cell division in plants is particularly important as cells cannot rearrange. It therefore determines the arrangement of cells (topology) and their size and shape (geometry). Cell division reduces mechanical stress locally by producing smaller cells and alters mechanical properties by reinforcing the mechanical wall network, both of which can alter overall tissue morphology. Division orientation is often regarded as following geometric rules, however recent work has suggested that divisions align with the direction of maximal tensile stress. Mechanical stress has already been shown to feed into many processes of development including those that alter mechanical properties. Such an alignment may enable cell division to selectively reinforce the cell wall network in the direction of maximal tensile stress. Therefore there exists potential feedback between cell division, mechanical stress and growth. Improving our understanding of this topic will help to shed light on the debated role of cell division in organ scale growth.

Spatiotemporal coordination of cell division and growth during organ morphogenesis.

A developing plant organ exhibits complex spatiotemporal patterns of growth, cell division, cell size, cell shape, and organ shape. Explaining these patterns presents a challenge because of their dynamics and cross-correlations, which can make it difficult to disentangle causes from effects. To address these problems, we used live imaging to determine the spatiotemporal patterns of leaf growth and division in different genetic and tissue contexts. In the simplifying background of the speechless (spch) mutant, which lacks stomatal lineages, the epidermal cell layer exhibits defined patterns of division, cell size, cell shape, and growth along the proximodistal and mediolateral axes. The patterns and correlations are distinctive from those observed in the connected subepidermal layer and also different from the epidermal layer of wild type. Through computational modelling we show that the results can be accounted for by a dual control model in which spatiotemporal control operates on both growth and cell division, with cross-connections between them. The interactions between resulting growth and division patterns lead to a dynamic distributions of cell sizes and shapes within a deforming leaf. By modulating parameters of the model, we illustrate how phenotypes with correlated changes in cell size, cell number, and organ size may be generated. The model thus provides an integrated view of growth and division that can act as a framework for further experimental study.

Mind the clinical-analytic gap: Electronic health records and COVID-19 pandemic response.

Data quality is essential to the success of the most simple and the most complex analysis. In the context of the COVID-19 pandemic, large-scale data sharing across the US and around the world has played an important role in public health responses to the pandemic and has been crucial to understanding and predicting its likely course. In California, hospitals have been required to report a large volume of daily data related to COVID-19. In order to meet this need, electronic health records (EHRs) have played an important role, but the challenges of reporting high-quality data in real-time from EHR data sources have not been explored. We describe some of the challenges of utilizing EHR data for this purpose from the perspective of a large, integrated, mixed-payer health system in northern California, US. We emphasize some of the inadequacies inherent to EHR data using several specific examples, and explore the clinical-analytic gap that forms the basis for some of these inadequacies. We highlight the need for data and analytics to be incorporated into the early stages of clinical crisis planning in order to utilize EHR data to full advantage. We further propose that lessons learned from the COVID-19 pandemic can result in the formation of collaborative teams joining clinical operations, informatics, data analytics, and research, ultimately resulting in improved data quality to support effective crisis response.

Transcriptome dynamics at Arabidopsis graft junctions reveal an intertissue recognition mechanism that activates vascular regeneration.

The ability for cut tissues to join and form a chimeric organism is a remarkable property of many plants; however, grafting is poorly characterized at the molecular level. To better understand this process, we monitored genome-wide gene expression changes in grafted Arabidopsis thaliana hypocotyls. We observed a sequential activation of genes associated with cambium, phloem, and xylem formation. Tissues above and below the graft rapidly developed an asymmetry such that many genes were more highly expressed on one side than on the other. This asymmetry correlated with sugar-responsive genes, and we observed an accumulation of starch above the graft junction. This accumulation decreased along with asymmetry once the sugar-transporting vascular tissues reconnected. Despite the initial starvation response below the graft, many genes associated with vascular formation were rapidly activated in grafted tissues but not in cut and separated tissues, indicating that a recognition mechanism was activated independently of functional vascular connections. Auxin, which is transported cell to cell, had a rapidly elevated response that was symmetric, suggesting that auxin was perceived by the root within hours of tissue attachment to activate the vascular regeneration process. A subset of genes was expressed only in grafted tissues, indicating that wound healing proceeded via different mechanisms depending on the presence or absence of adjoining tissues. Such a recognition process could have broader relevance for tissue regeneration, intertissue communication, and tissue fusion events.

Outpatient Urinary Tract Infections in an Era of Virtual Healthcare: Trends From 2008 to 2017.

BACKGROUND: Urinary tract infections (UTIs) occur commonly, but recent data on UTI rates are scarce. It is unknown how the growth of virtual healthcare delivery affects outpatient UTI management and trends in the United States. METHODS: From 1 January 2008 to 31 December 2017, UTIs from outpatient settings (office, emergency, and virtual visits) were identified from electronic health records at Kaiser Permanente Southern California using multiple UTI definitions. Annual rates estimated by Poisson regression were stratified by sex, care setting, age, and race/ethnicity. Annual trends were estimated by linear or piecewise Poisson regression. RESULTS: UTIs occurred in 1 065 955 individuals. Rates per 1000 person-years were 53.7 (95% confidence interval [CI], 50.6-57.0) by diagnosis code with antibiotic and 25.8 (95% CI, 24.7-26.9) by positive culture. Compared to office and emergency visits, UTIs were increasingly diagnosed in virtual visits, where rates by diagnosis code with antibiotic increased annually by 21.2% (95% CI, 16.5%-26.2%) in females and 29.3% (95% CI, 23.7%-35.3%) in males. Only 32% of virtual care diagnoses had a culture order. Overall, UTI rates were highest and increased the most in older adults. Rates were also higher in Hispanic and white females and black and white males. CONCLUSIONS: Outpatient UTI rates increased from 2008 to 2017, especially in virtual care and among older adults. Virtual care is important for expanding access to health services, but strategies are needed in all outpatient care settings to ensure accurate UTI diagnosis and reduce inappropriate antibiotic treatment.

Risk of melanoma following keratinocyte malignancies.

Patients diagnosed with keratinocyte cancer experience heightened risk for melanoma, yet patients who go on to develop this malignancy have not been well-characterized. We followed a population-based cohort of 2243 participants with histologically confirmed KC identified from dermatology and pathology practices who did not have a history of internal malignancy (1363 BCC, 880 SCC). A total of 77 participants went on to develop melanoma. Individual-level data were collected via personal interviews including demographic information and skin cancer risk factors, as well as KC tumor characteristics such as anatomic site and histologic subtype. Using adjusted Cox proportionate hazards models, older patients (age 61 or older vs 60 or younger) were at twofold increased risk for developing melanoma following KC (age 61-65 HR = 2.5; 95% CI = 1.3-4.6) (age > 65 HR = 2.0; 95% CI = 1.2-3.4) and women were at reduced risk compared to men (HR = 0.5; 95% CI = 0.3-0.8). Among patients with BCC, those with tumors on the trunk/limbs compared to the head/neck were at greater risk for subsequent melanoma (HR = 2.7; 95% CI = 1.3-5.7). Subsequent risk of melanoma also related to established risk factors including blond/red vs dark hair (HR = 1.9; 95% CI = 1.1-3.4), tendency to burn rather than tan (HR = 1.7; 95% CI = 1.0-2.7), ≥1 nevi on their back compared to no nevi (HR = 2.2; 95% CI = 1.2-3.8) and a history of ≥1 painful childhood sunburns vs none (HR = 2.1; 95% CI = 1.2-3.6). Thus, in addition to pigmentary traits, ultraviolet radiation (UVR)-related factors and clinical features of KC such as anatomic site may be useful in identifying patients at increased risk for melanoma after KC.

Plant cell divisions: variations from the shortest symmetric path.

In plants, the spatial arrangement of cells within tissues and organs is a direct consequence of the positioning of the new cell walls during cell division. Since the nineteenth century, scientists have proposed rules to explain the orientation of plant cell divisions. Most of these rules predict the new wall will follow the shortest path passing through the cell centroid halving the cell into two equal volumes. However, in some developmental contexts, divisions deviate significantly from this rule. In these situations, mechanical stress, hormonal signalling, or cell polarity have been described to influence the division path. Here we discuss the mechanism and subcellular structure required to define the cell division placement then we provide an overview of the situations where division deviates from the shortest symmetric path.

Patient-derived orthotopic xenografts of pediatric brain tumors: a St. Jude resource.

Pediatric brain tumors are the leading cause of cancer-related death in children. Patient-derived orthotopic xenografts (PDOX) of childhood brain tumors have recently emerged as a biologically faithful vehicle for testing novel and more effective therapies. Herein, we provide the histopathological and molecular analysis of 37 novel PDOX models generated from pediatric brain tumor patients treated at St. Jude Children's Research Hospital. Using a combination of histopathology, whole-genome and whole-exome sequencing, RNA-sequencing, and DNA methylation arrays, we demonstrate the overall fidelity and inter-tumoral molecular heterogeneity of pediatric brain tumor PDOX models. These models represent frequent as well as rare childhood brain tumor entities, including medulloblastoma, ependymoma, atypical teratoid rhabdoid tumor, and embryonal tumor with multi-layer rosettes. PDOX models will be valuable platforms for evaluating novel therapies and conducting pre-clinical trials to accelerate progress in the treatment of brain tumors in children. All described PDOX models and associated datasets can be explored using an interactive web-based portal and will be made freely available to the research community upon request.