Spontaneous multifocal pyogenic granulomas.

Cutaneous pyogenic granulomas (PGs) are common, benign vascular tumors of uncertain pathogenesis; however, a growing body of literature suggests that the formation of PGs may be secondary to genetic alterations in both the Ras/Raf/MAPK and PI3K/Akt/mTOR pathways. We present three cases of spontaneous multifocal PGs that first presented in infancy, were not associated with other vascular anomalies or discernable etiology, harbored somatic genetic variants in the Ras/Raf/MAPK pathway (NRAS n = 2, FGFR1 n = 1), were refractory to treatment with beta-blockers and mTOR inhibitors, and responded best to pulsed dye laser. We propose the term "spontaneous multifocal PGs" to describe this entity.

Dual RNA-seq in filarial nematodes and Wolbachia endosymbionts using RNase H based ribosomal RNA depletion.

Lymphatic filariasis is caused by parasitic nematodes and is a leading cause of disability worldwide. Many filarial worms contain the bacterium Wolbachia as an obligate endosymbiont. RNA sequencing is a common technique used to study their molecular relationships and to identify potential drug targets against the nematode and bacteria. Ribosomal RNA (rRNA) is the most abundant RNA species, accounting for 80-90% of the RNA in a sample. To reduce sequencing costs, it is necessary to remove ribosomal reads through poly-A enrichment or ribosomal depletion. Bacterial RNA does not contain a poly-A tail, making it difficult to sequence both the nematode and Wolbachia from the same library preparation using standard poly-A selection. Ribosomal depletion can utilize species-specific oligonucleotide probes to remove rRNA through pull-down or degradation methods. While species-specific probes are commercially available for many commonly studied model organisms, there are currently limited depletion options for filarial parasites. Here, we performed total RNA sequencing from Brugia malayi containing the Wolbachia symbiont (wBm) and designed ssDNA depletion probes against their rRNA sequences. We compared the total RNA library to poly-A enriched, Terminator 5'-Phosphate-Dependent Exonuclease treated, NEBNext Human/Bacteria rRNA depleted and our custom nematode probe depleted libraries. The custom nematode depletion library had the lowest percentage of ribosomal reads across all methods, with a 300-fold decrease in rRNA when compared to the total RNA library. The nematode depletion libraries also contained the highest percentage of Wolbachia mRNA reads, resulting in a 16-1,000-fold increase in bacterial reads compared to the other enrichment and depletion methods. Finally, we found that the Brugia malayi depletion probes can remove rRNA from the filarial worm Dirofilaria immitis and the majority of rRNA from the more distantly related free living nematode Caenorhabditis elegans. These custom filarial probes will allow for future dual RNA-seq experiments between nematodes and their bacterial symbionts from a single sequencing library.

Neuromuscular electrical stimulation for physical function maintenance during hematopoietic stem cell transplantation: Study protocol.

Hematopoietic stem cell transplantation is a common life-saving treatment for hematologic malignancies, though can lead to long-term functional impairment, fatigue, muscle atrophy, with decreased quality of life. Although traditional exercise has helped reduce these effects, it is inconsistently recommended and infrequently maintained, and most patients remain sedentary during and after treatment. There is need for alternative rehabilitation strategies, like neuromuscular electrical stimulation, that may be more amenable to the capabilities of hematopoietic stem cell transplant recipients. Patients receiving autologous HCT are being enroled in a randomized controlled trial with 1:1 (neuromuscular electrical stimulation:sham) design stratified by diagnosis and sex. Physical function, body composition, quality of life, and fatigue are assessed prior to hematopoietic stem cell transplant (prior to initiating preparatory treatment) and 24±5 days post hematopoietic stem cell transplant (Follow-up 1); physical function and quality of life are also assessed 6-months post hematopoietic stem cell transplant (Follow-up 2). The primary outcome is between-group difference in the 6-minute walk test change scores (Follow-up 1-Pre-transplant; final enrolment goal N = 23/group). We hypothesize that 1) neuromuscular electrical stimulation will attenuate hematopoietic stem cell transplant-induced adverse effects on physical function, muscle mass, quality of life, and fatigue compared to sham at Follow-up 1, and 2) Pre-transplant physical function will significantly predict fatigue and quality of life at Follow-up 2. We will also describe feasibility and acceptability of neuromuscular electrical stimulation during hematopoietic stem cell transplant. This proposal will improve rehabilitative patient care and quality of life by determining efficacy and feasibility of a currently underutilized therapeutic strategy aimed at maintaining daily function and reducing the impact of a potent and widely used cancer treatment. This trial is registered with clinicaltrials.gov (NCT04364256).

Origin of the human malaria parasite Plasmodium vivax.

The geographic origin of Plasmodium vivax, a leading cause of human malaria, has been the subject of much speculation. Here we review the evolutionary history of P. vivax and P. vivax-like parasites in humans and non-human primates on three continents, providing overwhelming evidence for an African origin. This conclusion is consistent with recent reports showing that Duffy-negative humans in Africa are, in fact, susceptible to P. vivax, with parasites invading Duffy-antigen-expressing erythroid precursors. Thus, the African origin of P. vivax not only explains the distribution of the Duffy-negative genotype but also provides new insight into the history and status of P. vivax malaria in Africa and efforts geared toward its eradication.

Clinical Relevance of Physical Function Outcomes in Cancer Cachexia.

Managing clinical manifestations of cancer/treatment burden on functional status and quality of life remains paramount across the cancer trajectory, particularly for patients with cachexia who display reduced functional capacity. However, clinically relevant criteria for classifying functional impairment at a single point in time or for classifying meaningful functional changes subsequent to disease and/or treatment progression are lacking. This unmet clinical need remains a major obstacle to the development of therapies for cancer cachexia. This review aims to describe current literature-based evidence for clinically meaningful criteria for (1) functional impairment at a single timepoint between cancer patients with or without cachexia and (2) changes in physical function over time across interventional studies conducted in patients with cancer cachexia. The most common functional assessment in cross-sectional and interventional studies was hand grip strength (HGS). We observed suggestive evidence that an HGS deficit between 3 and 6 kg in cancer cachexia may display clinical relevance. In interventional studies, we observed that long-duration multimodal therapies with a focus on skeletal muscle may benefit HGS in patients with considerable weight loss. Future studies should derive cohort-specific clinically relevant criteria to confirm these observations in addition to other functional outcomes and investigate appropriate patient-reported anchors.

Evaluating the Disease-Related Experiences of TikTok Users With Lupus Erythematosus: Qualitative and Content Analysis.

BACKGROUND: Lupus erythematosus (LE) is an autoimmune condition that is associated with significant detriments to quality of life and daily functioning. TikTok, a popular social networking platform for sharing short videos, provides a unique opportunity to understand experiences with LE within a nonclinical sample, a population that is understudied in LE research. This is the first qualitative study that explores LE experiences using the TikTok platform. OBJECTIVE: This study aims to evaluate the disease-related experiences of TikTok users with LE using qualitative and content analysis. METHODS: TikTok videos were included if the hashtags included #lupus, were downloadable, were in English, and involved the personal experience of an individual with LE. A codebook was developed using a standardized inductive approach of iterative coding until saturation was reached. NVivo (Lumivero), a qualitative analysis software platform, was used to code videos and perform content analysis. Inductive thematic analysis was used to derive themes from the data. RESULTS: A total of 153 TikTok videos met the inclusion criteria. The most common codes were experiences with symptoms (106/153, 69.3%), mucocutaneous symptoms (61/153, 39.9%), and experiences with treatment (59/153, 38.6%). Experiences with symptoms and mucocutaneous symptoms had the greatest cumulative views (25,381,074 and 14,879,109 views, respectively). Five thematic conclusions were derived from the data: (1) mucocutaneous symptoms had profound effects on the mental health and body image of TikTok users with LE; (2) TikTok users' negative experiences with health care workers were often derived from diagnostic delays and perceptions of "medical gaslighting"; (3) TikTok users tended to portray pharmacologic and nonpharmacologic interventions, such as diet and naturopathic remedies, positively, whereas pharmacologic treatments were portrayed negatively or referred to as "chemotherapy"; (4) LE symptoms, particularly musculoskeletal symptoms and fatigue, interfered with users' daily functioning; and (5) although TikTok users frequently had strong support systems, feelings of isolation were often attributed to battling an "invisible illness." CONCLUSIONS: This study demonstrates that social media can provide important, clinically relevant information for health practitioners caring for patients with chronic conditions such as LE. As mucocutaneous symptoms were the predominant drivers of distress in our sample, the treatment of hair loss and rash is vital in this population. However, pharmacologic therapies were often depicted negatively, reinforcing the significance of discussions on the safety and effectiveness of these treatments. In addition, while TikTok users demonstrated robust support systems, feelings of having an "invisible illness" and "medical gaslighting" dominated negative interactions with others. This underscores the importance of providing validation in clinical interactions.

A Pipeline for Evaluation of Machine Learning/Artificial Intelligence Models to Quantify Programmed Death Ligand 1 Immunohistochemistry.

Immunohistochemistry (IHC) is used to guide treatment decisions in multiple cancer types. For treatment with checkpoint inhibitors, programmed death ligand 1 (PD-L1) IHC is used as a companion diagnostic. However, the scoring of PD-L1 is complicated by its expression in cancer and immune cells. Separation of cancer and noncancer regions is needed to calculate tumor proportion scores (TPS) of PD-L1, which is based on the percentage of PD-L1-positive cancer cells. Evaluation of PD-L1 expression requires highly experienced pathologists and is often challenging and time-consuming. Here, we used a multi-institutional cohort of 77 lung cancer cases stained centrally with the PD-L1 22C3 clone. We developed a 4-step pipeline for measuring TPS that includes the coregistration of hematoxylin and eosin, PD-L1, and negative control (NC) digital slides for exclusion of necrosis, segmentation of cancer regions, and quantification of PD-L1+ cells. As cancer segmentation is a challenging step for TPS generation, we trained DeepLab V3 in the Visiopharm software package to outline cancer regions in PD-L1 and NC images and evaluated the model performance by mean intersection over union (mIoU) against manual outlines. Only 14 cases were required to accomplish a mIoU of 0.82 for cancer segmentation in hematoxylin-stained NC cases. For PD-L1-stained slides, a model trained on PD-L1 tiles augmented by registered NC tiles achieved a mIoU of 0.79. In segmented cancer regions from whole slide images, the digital TPS achieved an accuracy of 75% against the manual TPS scores from the pathology report. Major reasons for algorithmic inaccuracies include the inclusion of immune cells in cancer outlines and poor nuclear segmentation of cancer cells. Our transparent and stepwise approach and performance metrics can be applied to any IHC assay to provide pathologists with important insights on when to apply and how to evaluate commercial automated IHC scoring systems.

Formative Evaluation of CLABSI Adoption and Sustainment Interventions in a Pediatric Intensive Care Unit.

Background: Pediatric patients require central venous catheters to maintain adequate hydration, nutritional status, and delivery of life-saving medications in the pediatric intensive care unit. Although central venous catheters provide critical medical therapies, their use increases the risk of severe infection, morbidity, and mortality. Adopting an evidence-based central line-associated bloodstream infection (CLABSI) bundle to guide nursing practice can decrease and sustain low CLABSI rates, but reliable and consistent implementation is challenging. This study aimed to conduct a mixed-methods formative evaluation to explore CLABSI bundle implementation strategies in a PICU. Methods: The team used The Consolidated Framework for Implementation Research to develop the interview guide and data analysis plan. Results: Facilitators and barriers for the CLABSI bundle occurred in four domains: inner setting, process, characteristics of individuals, and innovation characteristics in each cycle that led to recommended implementation strategy opportunities. The champion role was a major implementation strategy that facilitated the adoption and sustainment of the CLABSI bundle. Conclusions: Implementation Science Frameworks, such as Consolidated Framework for Implementation Research (CFIR), can be a beneficial framework to guide quality improvement efforts for evidence-based practices such as the CLABSI bundle. Using a champion role in the critical care setting may be an important implementation strategy for CLABSI bundle adoption and sustainment efforts.

Treatment of Subcorneal Pustular Dermatosis without Dapsone: A Case Report and Review of the Literature.

Subcorneal pustular dermatosis (SPD) is a rare neutrophilic dermatosis characterized by pustules on the trunk and intertriginous areas. While oral dapsone is the first-line treatment for SPD, alternative options are necessary for patients with glucose-6-phosphate dehydrogenase deficiency, drug hypersensitivity reactions, or refractory disease. To date, no consensus exists regarding next-best agents for SPD. In this report, we present a patient with significant SPD who developed dapsone hypersensitivity syndrome and then was successfully treated with colchicine and adalimumab. We propose that colchicine should be considered as a second-line treatment for SPD and present a therapeutic algorithm for clinicians to utilize when patients are not candidates for dapsone, have side effects requiring drug discontinuation, or have refractory disease.

Prevalence of FRAC Group 11 Fungicide Resistance in Stemphylium vesicarium Isolates, but Not S. beticola Isolates, Causing Stemphylium Leaf Spot of Spinach (Spinacia oleracea).

Stemphylium leaf spot of spinach, caused by Stemphylium beticola and S. vesicarium, is a disease of economic importance in fresh market, processing, and seed production. There have been increasing reports of difficulty managing the disease in the southern United States using fungicides in Fungicide Resistance Action Committee (FRAC) group 11. Isolates of S. beticola and S. vesicarium obtained from spinach leaves and seed from 2001 to 2020 were screened for resistance to azoxystrobin and pyraclostrobin in vitro, in vivo, and using PCR assays to detect mutations in cytochrome b associated with resistance in other fungi (F129L, G137R, and G143A). EC50 values for mycelial growth and conidial germination of S. vesicarium isolates in vitro were significantly less (mean of 0.35 µg/ml) than that of S. vesicarium (mean of 14.17 µg/ml) with both fungicides. All isolates were slightly more sensitive to pyraclostrobin than azoxystrobin in both assays. In vivo assays of plants inoculated with the isolates of S. vesicarium demonstrated poor efficacy of fungicides with each of the two active ingredients. Only the G143A mutation was detected in all spinach isolates of S. vesicarium, including an isolate of S. vesicarium collected in 2003 and 82.9% of isolates from spinach seed lots harvested from crops grown in or after 2017 in Europe, New Zealand, and the United States. The FRAC 11 mutations were not detected in any isolates of S. beticola. The in vitro, in vivo, and DNA mutation assays suggest FRAC group 11 fungicide resistance is widespread in spinach isolates of S. vesicarium but not S. beticola.

Diel flight activity in response to attractants in the predatory beetle, Cathartus quadricollis (Coleoptera: Silvanidae) in macadamia nut in Hawaii.

The diel flight activity in Cathartus quadricollis (Guerin-Meneville) (Coleoptera: Silvanidae), a predator of two important pests in Hawaii, coffee berry borer, Hypothenemus hampei (Ferrari) and tropical nut borer, Hypothenemus obscurus (F.) (Coleoptera: Curculionidae: Scolytinae) was studied in a macadamia nut orchard using yellow sticky traps baited with pheromone and fungal volatile attractants. The study was conducted at different months throughout the year and at different times during the lunar cycle (new moon and full moon). Flight activity peaked in the late hours of the photophase into the early hours of the scotophase, between 1830 and 2000 h; flight activity also occurred but to a lesser extent in the early morning hours between 0700 and 1030 h. Numbers of captured C. quadricollis during periods of flight activity were negatively correlated with wind speed. The implications of these findings for the development of optimal pest management strategies including biological control are discussed.

Improved metagenome assemblies through selective enrichment of bacterial genomic DNA from eukaryotic host genomic DNA using ATAC-seq.

Genomics can be used to study the complex relationships between hosts and their microbiota. Many bacteria cannot be cultured in the laboratory, making it difficult to obtain adequate amounts of bacterial DNA and to limit host DNA contamination for the construction of metagenome-assembled genomes (MAGs). For example, Wolbachia is a genus of exclusively obligate intracellular bacteria that live in a wide range of arthropods and some nematodes. While Wolbachia endosymbionts are frequently described as facultative reproductive parasites in arthropods, the bacteria are obligate mutualistic endosymbionts of filarial worms. Here, we achieve 50-fold enrichment of bacterial sequences using ATAC-seq (Assay for Transposase-Accessible Chromatin using sequencing) with Brugia malayi nematodes, containing Wolbachia (wBm). ATAC-seq uses the Tn5 transposase to cut and attach Illumina sequencing adapters to accessible DNA lacking histones, typically thought to be open chromatin. Bacterial and mitochondrial DNA in the lysates are also cut preferentially since they lack histones, leading to the enrichment of these sequences. The benefits of this include minimal tissue input (<1 mg of tissue), a quick protocol (<4 h), low sequencing costs, less bias, correct assembly of lateral gene transfers and no prior sequence knowledge required. We assembled the wBm genome with as few as 1 million Illumina short paired-end reads with >97% coverage of the published genome, compared to only 12% coverage with the standard gDNA libraries. We found significant bacterial sequence enrichment that facilitated genome assembly in previously published ATAC-seq data sets from human cells infected with Mycobacterium tuberculosis and C. elegans contaminated with their food source, the OP50 strain of E. coli. These results demonstrate the feasibility and benefits of using ATAC-seq to easily obtain bacterial genomes to aid in symbiosis, infectious disease, and microbiome research.

Cyclophosphamide induces the loss of taste bud innervation in mice.

Many common chemotherapeutics produce disruptions in the sense of taste which can lead to loss of appetite, nutritional imbalance, and reduced quality of life, especially if taste loss persists after treatment ends. Cyclophosphamide (CYP), an alkylating chemotherapeutic agent, affects taste sensitivity through its cytotoxic effects on mature taste receptor cells (TRCs) and on taste progenitor cell populations, retarding the capacity to replace TRCs. Mechanistic studies have focused primarily on taste cells, however, taste signaling requires communication between TRCs and the gustatory nerve fibers that innervate them. Here, we evaluate cyclophosphamide's effects on the peripheral gustatory nerve fibers that innervate the taste buds. Following histological analysis of tongue tissues, we find that CYP reduces innervation within the fungiform and circumvallates taste buds within 4 days after administration. To better understand the dynamics of the denervation process, we used 2-photon intravital imaging to visualize the peripheral gustatory nerve fibers within individual fungiform taste buds up to 20 days after CYP treatment. We find that gustatory fibers retract from the taste bud properly but are maintained within the central papilla core. These data indicate that in addition to TRCs, gustatory nerve fibers are also affected by CYP treatment. Because the connectivity between TRCs and gustatory neurons must be re-established for proper function, gustatory fibers should continue to be included in future studies to understand the mechanisms leading to chemotherapy-induced persistent taste loss.

Single-Cell Analysis of Rohon-Beard Neurons Implicates Fgf Signaling in Axon Maintenance and Cell Survival.

Peripheral sensory neurons are a critical part of the nervous system that transmit a multitude of sensory stimuli to the central nervous system. During larval and juvenile stages in zebrafish, this function is mediated by Rohon-Beard somatosensory neurons (RBs). RBs are optically accessible and amenable to experimental manipulation, making them a powerful system for mechanistic investigation of sensory neurons. Previous studies provided evidence that RBs fall into multiple subclasses; however, the number and molecular makeup of these potential RB subtypes have not been well defined. Using a single-cell RNA sequencing (scRNA-seq) approach, we demonstrate that larval RBs in zebrafish fall into three, largely nonoverlapping classes of neurons. We also show that RBs are molecularly distinct from trigeminal neurons in zebrafish. Cross-species transcriptional analysis indicates that one RB subclass is similar to a mammalian group of A-fiber sensory neurons. Another RB subclass is predicted to sense multiple modalities, including mechanical stimulation and chemical irritants. We leveraged our scRNA-seq data to determine that the fibroblast growth factor (Fgf) pathway is active in RBs. Pharmacological and genetic inhibition of this pathway led to defects in axon maintenance and RB cell death. Moreover, this can be phenocopied by treatment with dovitinib, an FDA-approved Fgf inhibitor with a common side effect of peripheral neuropathy. Importantly, dovitinib-mediated axon loss can be suppressed by loss of Sarm1, a positive regulator of neuronal cell death and axonal injury. This offers a molecular target for future clinical intervention to fight neurotoxic effects of this drug.

Open Access and Reproducibility in Plant Pathology Research: Guidelines and Best Practices.

The landscape of scientific publishing is experiencing a transformative shift toward open access, a paradigm that mandates the availability of research outputs such as data, code, materials, and publications. Open access provides increased reproducibility and allows for reuse of these resources. This article provides guidance for best publishing practices of scientific research, data, and associated resources, including code, in The American Phytopathological Society journals. Key areas such as diagnostic assays, experimental design, data sharing, and code deposition are explored in detail. This guidance aligns with that observed by other leading journals. We hope the information assembled in this paper will raise awareness of best practices and enable greater appraisal of the true effects of biological phenomena in plant pathology.

The impact of early direct-contact experiences on reducing mental health stigma among student pharmacists: A pilot study.

Introduction: Mental health-related stigma is a barrier to treatment and recovery for serious mental illnesses (SMIs). Educational training programs demonstrate positive changes in health professional students' attitudes and stigma toward SMI; however, student pharmacists have minimal opportunity to directly engage with the SMI population. This study aims to assess and compare student pharmacists' stigma related to SMI before and after participating in a pilot series of direct-contact workshop experiences. Methods: The 15-item Opening Minds Scale for Healthcare Providers survey was administered to student pharmacists before and after the workshop experiences to measure stigma toward SMI. Five 2-hour workshops were provided to members of a local nonprofit organization serving people with SMI by student pharmacist volunteers detailing a health and wellness topic. The postworkshop survey included free text responses to obtain student feedback. Results: Twenty-four complete preworkshop surveys were obtained, and most of them had positive attitudes and beliefs at baseline. Thirteen postworkshop surveys were obtained from student pharmacists who participated in a workshop event, and 9 were completed by student pharmacists who did not participate in a workshop event, which were used as a comparator group. Stigma decreased after participating in a workshop event, and those who participated demonstrated a lower degree of stigma versus the comparator group. Discussion: Direct-contact experiences allow student pharmacists to interact with people with SMI earlier in their training and help reduce stigma toward those with psychiatric disorders. Future research is needed to identify large-scale changes in pharmacy student stigma.

Expanded T lymphocytes in the cerebrospinal fluid of multiple sclerosis patients are specific for Epstein-Barr-virus-infected B cells.

Epstein-Barr virus (EBV) infection has long been associated with multiple sclerosis (MS), but the role of EBV in the pathogenesis of MS is not clear. Our hypothesis is that a major fraction of the expanded clones of T lymphocytes in the cerebrospinal fluid (CSF) are specific for autologous EBV-infected B cells. We obtained blood and CSF samples from eight relapsing-remitting patients in the process of diagnosis. We stimulated cells from the blood with autologous EBV-infected lymphoblastoid cell lines (LCL), EBV, varicella zoster virus, influenza, and candida and sorted the responding cells with flow cytometry after 6 d. We sequenced the RNA for T cell receptors (TCR) from CSF, unselected blood cells, and the antigen-specific cells. We used the TCR Vß CDR3 sequences from the antigen-specific cells to assign antigen specificity to the sequences from the CSF and blood. LCL-specific cells comprised 13.0 ± 4.3% (mean ± SD) of the total reads present in CSF and 13.3 ± 7.5% of the reads present in blood. The next most abundant antigen specificity was flu, which was 4.7 ± 1.7% of the reads in the CSF and 9.3 ± 6.6% in the blood. The prominence of LCL-specific reads was even more marked in the top 1% most abundant CSF clones with statistically significant 47% mean overlap with LCL. We conclude that LCL-specific sequences form a major portion of the TCR repertoire in both CSF and blood and that expanded clones specific for LCL are present in MS CSF. This has important implications for the pathogenesis of MS.