SARS-CoV-2-related bat viruses evade human intrinsic immunity but lack efficient transmission capacity.

Circulating bat coronaviruses represent a pandemic threat. However, our understanding of bat coronavirus pathogenesis and transmission potential is limited by the lack of phenotypically characterized strains. We created molecular clones for the two closest known relatives of SARS-CoV-2, BANAL-52 and BANAL-236. We demonstrated that BANAL-CoVs and SARS-CoV-2 have similar replication kinetics in human bronchial epithelial cells. However, BANAL-CoVs have impaired replication in human nasal epithelial cells and in the upper airway of mice. We also observed reduced pathogenesis in mice and diminished transmission in hamsters. Further, we observed that diverse bat coronaviruses evade interferon and downregulate major histocompatibility complex class I. Collectively, our study demonstrates that despite high genetic similarity across bat coronaviruses, prediction of pandemic potential of a virus necessitates functional characterization. Finally, the restriction of bat coronavirus replication in the upper airway highlights that transmission potential and innate immune restriction can be uncoupled in this high-risk family of emerging viruses.

Antibody-assisted selective isolation of Purkinje cell nuclei from mouse cerebellar tissue.

We developed a method that utilizes fluorescent labeling of nuclear envelopes alongside cytometry sorting for the selective isolation of Purkinje cell (PC) nuclei. Beginning with SUN1 reporter mice, we GFP-tagged envelopes to confirm that PC nuclei could be accurately separated from other cell types. We then developed an antibody-based protocol to make PC nuclear isolation more robust and adaptable to cerebellar tissues of any genotypic background. Immunofluorescent labeling of the nuclear membrane protein RanBP2 enabled the isolation of PC nuclei from C57BL/6 cerebellum. By analyzing the expression of PC markers, nuclear size, and nucleoli number, we confirmed that our method delivers a pure fraction of PC nuclei. To demonstrate its applicability, we isolated PC nuclei from spinocerebellar ataxia type 7 (SCA7) mice and identified transcriptional changes in known and new disease-associated genes. Access to pure PC nuclei offers insights into PC biology and pathology, including the nature of selective neuronal vulnerability.

Clinicopathologic and molecular study of superficial CD34-positive fibroblastic tumours mimicking atypical fibrous histiocytoma (dermatofibroma).

AIMS: Superficial CD34-positive fibroblastic tumour (SCD34FT) is an uncommon but distinctive low-grade neoplasm of the skin and subcutis that shows frequent CADM3 expression by immunohistochemistry (IHC). In this study, prompted by an index case resembling 'atypical fibrous histiocytoma (FH)' that was positive for CADM3 IHC, we systematically examined a cohort of tumours previously diagnosed as 'atypical FH' by applying CADM3 and fluorescence in situ hybridization (FISH) for PRDM10 rearrangement, to investigate the overlap between these tumour types. METHODS AND RESULTS: Forty cases of atypical FH were retrieved, including CD34-positive tumours (n = 20) and CD34-negative tumours (n = 20). All tumours were stained for CADM3. All CADM3-positive tumours were evaluated by FISH to assess for PRDM10 rearrangement. Eleven CD34-positive tumours (11/20, 55%) coexpressed CADM3 and were reclassified as SCD34FT. None (0/20) of the CD34-negative atypical FH were CADM3-positive. Reclassified SCD34FT (10/11) arose on the lower extremity, with frequent involvement of the thigh (n = 8). Features suggestive of atypical FH were observed in many reclassified cases including variable cellularity, spindled morphology, infiltrative tumour margins, collagen entrapment, epidermal hyperpigmentation, and acanthosis. Variably prominent multinucleate giant cells, including Touton-like forms, were also present. An informative FISH result was obtained in 10/11 reclassified tumours, with 60% (6/10) demonstrating PRDM10 rearrangement. CONCLUSION: A significant subset of tumours that histologically resemble atypical FH, and are positive for CD34, coexpress CADM3 and harbour PRDM10 rearrangement, supporting their reclassification as SCD34FT. Awareness of this morphologic overlap and the application of CADM3 IHC can aid the distinction between SCD34FT and atypical FH.

Harnessing cold adaptation for postglacial colonisation: Galactinol synthase expression and raffinose accumulation in a polyploid and its progenitors.

Allotetraploid white clover (Trifolium repens) formed during the last glaciation through hybridisation of two European diploid progenitors from restricted niches: one coastal, the other alpine. Here, we examine which hybridisation-derived molecular events may have underpinned white clover's postglacial niche expansion. We compared the transcriptomic frost responses of white clovers (an inbred line and an alpine-adapted ecotype), extant descendants of its progenitor species and a resynthesised white clover neopolyploid to identify genes that were exclusively frost-induced in the alpine progenitor and its derived subgenomes. From these analyses we identified galactinol synthase, the rate-limiting enzyme in biosynthesis of the cryoprotectant raffinose, and found that the extant descendants of the alpine progenitor as well as the neopolyploid white clover rapidly accumulated significantly more galactinol and raffinose than the coastal progenitor under cold stress. The frost-induced galactinol synthase expression and rapid raffinose accumulation derived from the alpine progenitor likely provided an advantage during early postglacial colonisation for white clover compared to its coastal progenitor.

Development of a novel precision applicator for spot treatment of granular agrochemical in wild blueberry.

While spot spraying has gained increasing popularity in recent years, spot application of granule agrochemical has seen little development. Despite the potential for the technology, there currently exists no commercially available granular applicators capable of spot application. Therefore, the goal of this study was to design, build, and lab evaluate a precision applicator for spot applying granular agrochemical in wild blueberry. The design incorporated a John Deere RC2000 with a custom control box, recirculation system, and electrically actuated valves. All components were modified to fit a Valmar 1255 Twin-Roller. The system receives inputs from a predeveloped prescription map and can actuate each of the twelve valves separately to provide individual orifice control. Casoron® G4 was used as the testing agrochemical and in cycling the product pneumatically for 1 hour incurred no significant product degradation (p = 0.110). In lab evaluations, the applicator encountered zero errors in reading prescription maps and actuating the correct valves accordingly. Further, the granule recycling system had zero instances where product built up in the lines or jammed the valves. In all, this project represents the first successful development of a precision granular spot applicator for any cropping system.

Ephemeral stream water contributions to United States drainage networks.

Ephemeral streams flow only in direct response to precipitation and are ubiquitous landscape features. However, little is known about their influence on downstream rivers. Here, we modeled ephemeral stream water contributions to the contiguous United States network of more than 20 million rivers, lakes, and reservoirs, finding that ephemeral streams contribute, on average, 55% of the discharge exported from regional river systems, as defined by the United States Geological Survey. Our results show that ephemeral connectivity is a substantial pathway through which water and associated nutrients and pollution may enter the perennial drainage network and influence water quality. We provide quantitative insight into the implications of differing interpretations of regulatory jurisdiction under the United States Clean Water Act, including the current standard adopted by the Supreme Court of the United States in 2023.

What to do with an incidental finding of a fused sagittal suture: a modified Delphi study.

OBJECTIVE: As many as 5% of normocephalic children may have a prematurely fused sagittal suture, yet the clinical significance and best course of management of this finding remain unclear. Providers in the Synostosis Research Group were surveyed to create a multicenter consensus on an optimal treatment and monitoring algorithm for this condition. METHODS: A four-round modified Delphi method was utilized. The first two rounds consisted of anonymous surveys distributed to 10 neurosurgeons and 9 plastic surgeons with expertise in craniosynostosis across 9 institutions, and presented 3 patients (aged 3 years, 2 years, and 2 months) with incidentally discovered fused sagittal sutures, normal cephalic indices, and no parietal dysmorphology. Surgeons were queried about their preferred term for this entity and how best to manage these patients. Results were synthesized to create a treatment algorithm. The third and fourth feedback rounds consisted of open discussion of the algorithm until no further concerns arose. RESULTS: Most surgeons preferred the term "premature fusion of the sagittal suture" (93%). At the conclusion of the final round, all surgeons agreed to not operate on the 3- and 2-year-old patients unless symptoms of intracranial hypertension or papilledema were present. In contrast, 50% preferred to operate on the 2-month-old. However, all agreed to utilize shared decision-making, taking into account any concerns about future head shape and neurodevelopment. Panelists agreed that patients over 18 months of age without signs or symptoms suggesting elevated intracranial pressure (ICP) should not undergo surgical treatment. CONCLUSIONS: Through the Delphi method, a consensus regarding management of premature fusion of the sagittal suture was obtained from a panel of North American craniofacial surgeons. Without signs or symptoms of ICP elevation, surgery is not recommended in patients over 18 months of age. However, for children younger than 18 months, surgery should be discussed with caregivers using a shared decision-making process.

Widefield Super-Resolution Infrared Spectroscopy and Imaging of Autofluorescent Biological Materials and Photosynthetic Microorganisms Using Fluorescence Detected Photothermal Infrared (FL-PTIR).

We have demonstrated high-speed, super-resolution infrared (IR) spectroscopy and chemical imaging of autofluorescent biomaterials and organisms using camera-based widefield photothermal detection that takes advantage of temperature-dependent modulations of autofluorescent emission. A variety of biological materials and photosynthetic organisms exhibit strong autofluorescence emission under ultraviolet excitation and the autofluorescent emission has a very strong temperature dependence, of order 1%/K. Illuminating a sample with pulses of IR light from a wavelength-tunable laser source causes periodic localized sample temperature increases that result in a corresponding transient decrease in autofluorescent emission. A low-cost light-emitting diode-based fluorescence excitation source was used in combination with a conventional fluorescence microscopy camera to detect localized variations in autofluorescent emission over a wide area as an indicator of localized IR absorption. IR absorption image stacks were acquired over a range of IR wavelengths, including the fingerprint spectral range, enabling extraction of localized IR absorption spectra. We have applied widefield fluorescence detected photothermal IR (FL-PTIR) to an analysis of autofluorescent biological materials including collagen, leaf tissue, and photosynthetic organisms including diatoms and green microalgae cells. We have also demonstrated the FL-PTIR on live microalgae in water, demonstrating the potential for label-free dynamic chemical imaging of autofluorescent cells.

Can Adverse Event Patterns Inform Shared Decision-Making in ADHD Treatment? A Systematic Review of Evidence From Registration Trials for FDA-Approved Treatments in Adults.

INTRODUCTION: Adult patients and clinicians are faced with several pharmacological options to manage attention-deficit/hyperactivity disorder (ADHD). If types or rates of adverse experiences vary among these options, these differences could inform the shared decision-making process. METHODS: To discern differentiating evidence-based patterns of risk, we analyzed data from FDA package labels for drugs approved to treat adult ADHD and reports from the registration trials used to create these labels. Three analyses of adverse effects were conducted: placebo-corrected occurrence at rates of 1 in 5, 10, and 20 participants, association with discontinuation, and uniqueness of occurrence within the treatment options. RESULTS: Among the 7 agents approved to treat adult ADHD, the number of types of side effects experienced during a mix of fixed and flexible-dose studies was greatest among the nonstimulant medications, but the stimulant medications had higher rates of occurrence of side effects. The minimum frequency at which all medications had adverse events was 1 in 10 participants. Overall discontinuation rates did not differ among the stimulant medications nor between stimulants and nonstimulants. DISCUSSION: To our knowledge, this is the first study to compile and compare data from all FDA registration trials for medications approved to treat adult ADHD. This article describes a process by which readily available adverse event reporting data can be used as a tool to inform shared clinical decision-making. While differences in the methodology and outcome reporting of the trials included may limit generalizability, the number of individual patients included and the completeness of the discontinuation data can be used to inform discussions with patients about the relative likelihood of adverse experiences and other patient concerns.

A novel microporous biomaterial vaccine platform for long-lasting antibody mediated immunity against viral infection.

Current antigen delivery platforms, such as alum and nanoparticles, are not readily tunable, thus may not generate optimal adaptive immune responses. We created an antigen delivery platform by loading lyophilized Microporous Annealed Particle (MAP) with aqueous solution containing target antigens. Upon administration of antigen loaded MAP (VaxMAP), the biomaterial reconstitution forms an instant antigen-loaded porous scaffold area with a sustained release profile to maximize humoral immunity. VaxMAP induced CD4+ T follicular helper (Tfh) cells and germinal center (GC) B cell responses in the lymph nodes similar to Alum. VaxMAP loaded with SARS-CoV-2 spike protein improved the magnitude, neutralization, and duration of anti-receptor binding domain antibodies compared to Alum vaccinated mice. A single injection of Influenza specific HA1-loaded-VaxMAP enhanced neutralizing antibodies and elicited greater protection against influenza virus challenge than HA1-loaded-Alum. Thus, VaxMAP is a platform that can be used to promote adaptive immune cell responses to generate more robust neutralizing antibodies, and better protection upon pathogen challenge.

Oncogenic Control of Metabolism.

A cell committed to proliferation must reshape its metabolism to enable robust yet balanced production of building blocks for the assembly of proteins, lipids, nucleic acids, and other macromolecules, from which two functional daughter cells can be produced. The metabolic remodeling associated with proliferation is orchestrated by a number of pro-proliferative signaling nodes, which include phosphatidylinositol-3 kinase (PI3K), the RAS family of small GTPases, and transcription factor c-myc In metazoan cells, these signals are activated in a paracrine manner via growth factor-mediated activation of receptor (or receptor-associated) tyrosine kinases. Such stimuli are limited in duration and therefore allow the metabolism of target cells to return to the resting state once the proliferation demands have been satisfied. Cancer cells acquire activating genetic alterations within common pro-proliferative signaling nodes. These alterations lock cellular nutrient uptake and utilization into a perpetual progrowth state, leading to the aberrant accumulation and spread of cancer cells.

Impacts of groundwater dynamics around a macro-tidal river on agricultural soil salinity.

Estuaries are vulnerable to oceanic and atmospheric climate change. Much of the research investigating climate change impacts on estuaries is focused on saltwater intrusion within surface water due to drought and rising sea levels, with implications for ecosystems and humans. Groundwater and soil near estuaries may also be influenced, as estuary salinity and hydraulic head changes can impact soils and aquifers not previously at risk of salinization. This study was conducted to address knowledge gaps related to present and future groundwater salinity distribution in a groundwater system connected to a macro-tidal estuary. The studied estuary experiences a tidal bore due to its hydraulic connection to the Bay of Fundy in Nova Scotia, Canada. A parcel of agricultural land adjacent to the estuary was selected to assess the groundwater response to episodic fluctuations in estuary water levels and salinity. Groundwater monitoring and electromagnetic surveys were conducted to map soil and groundwater salinity patterns. A numerical model of groundwater flow and solute transport informed by field data was used to investigate how varying estuary salinity due to droughts and sea-level rise could impact groundwater salinity. Results showed that, in contrast to salt wedges observed along marine coasts, the saline groundwater existed as a plume immediately around the estuary. Model simulations showed that short-term droughts had an insignificant impact on the adjacent groundwater salinity. However, permanent increases in salinity caused by sea-level rise increased the plume volume by 86 %, or an additional ∼11 m horizontally and âˆ¼ 4.5 m vertically. Our results suggest that increased river salinity in this setting would not result in widespread salinization of porewater and agricultural soils, but more extensive salinization may be experienced in permeable aquifers or along more saline estuarine zones. Findings may inform land management decisions in regions exposed to increased salinity in the future.

Scoping review of the use of mesenchymal stem and stromal cell products in cats, Part 1: current logistics and safety.

Despite a pressing need for new therapies to address unmet veterinary medical need, no approved stem cell products are available for use in cats in the US. To evaluate the current state of mesenchymal stem or stromal cell (MSC) research in cats, a scoping review of published literature was performed, which identified 108 publications related to feline MSCs. Twenty-six of the articles described administration of MSC products to a total of 215 cats. Twelve of the studies included a control group. These experimental and clinical trials used 7 cell sources, 9 administration routes, 12 delivery vehicles, and a 300-fold range in dosages for initial studies in healthy cats and cats with 12 naturally occurring and induced diseases. The majority of studies administered 2 doses of allogeneic, adipose-derived MSC IV and monitored a median of 6.5 treated cats for a median of 90 days. The majority (150/215 [69.8%]) of cats had no reported adverse events associated with treatment. Although an increase in feline MSC publications in the past 10 years indicates progress, the wide variety and small number of studies using MSCs and MSC products in cats demonstrates that current evaluations are mostly still in the discovery phase, and several issues remain related to larger scale trials using MSC products in cats. The current available publications provide information to direct further clinical study development and informed owner consent for study enrollment.

Scoping review of the use of mesenchymal stem and stromal cell products in cats, Part 2: current scope and efficacy.

A scoping review of published literature found 108 articles related to mesenchymal stem or stromal cell (MSC) use in cats. Twenty-four of the publications summarized the treatment of 192 cats with MSC products for 12 naturally occurring and induced diseases. These trials used a variety of cell sources, administration routes, delivery vehicles, and dosages. The majority of studies did not have a control group. The disease with the largest number of cats administered MSCs thus far is chronic kidney disease (n = 59 cats). The majority of cats had no adverse events associated with treatment, which supports continued interest in the potential use of MSC products to address unmet medical needs. Treatment outcomes of the 192 cats have ranged from no response to long-term cure, depending on the disease being treated and the particular study. Some of these early studies show promise and provide significant information to direct both the design and focus of larger clinical trials investigating the safety and efficacy of MSC treatment for veterinary and human applications.

Intranasal neomycin evokes broad-spectrum antiviral immunity in the upper respiratory tract.

Respiratory virus infections in humans cause a broad-spectrum of diseases that result in substantial morbidity and mortality annually worldwide. To reduce the global burden of respiratory viral diseases, preventative and therapeutic interventions that are accessible and effective are urgently needed, especially in countries that are disproportionately affected. Repurposing generic medicine has the potential to bring new treatments for infectious diseases to patients efficiently and equitably. In this study, we found that intranasal delivery of neomycin, a generic aminoglycoside antibiotic, induces the expression of interferon-stimulated genes (ISGs) in the nasal mucosa that is independent of the commensal microbiota. Prophylactic or therapeutic administration of neomycin provided significant protection against upper respiratory infection and lethal disease in a mouse model of COVID-19. Furthermore, neomycin treatment protected Mx1 congenic mice from upper and lower respiratory infections with a highly virulent strain of influenza A virus. In Syrian hamsters, neomycin treatment potently mitigated contact transmission of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). In healthy humans, intranasal application of neomycin-containing Neosporin ointment was well tolerated and effective at inducing ISG expression in the nose in a subset of participants. These findings suggest that neomycin has the potential to be harnessed as a host-directed antiviral strategy for the prevention and treatment of respiratory viral infections.

Eyeing Risks: A Critical Analysis of the Use of Periorbital Steroids in Fronto-orbital Advancement.

OBJECTIVE: To evaluate the outcomes associated with two techniques of periorbital steroid administration in bilateral fronto-orbital advancement (FOA). DESIGN: Multi-institutional retrospective chart review. SETTING: Two high volume, tertiary US craniofacial centers. PATIENTS, PARTICIPANTS: Patients who underwent FOA between 2012 and 2021. INTERVENTIONS: Patients were divided into three cohorts based on method of steroid administration. Groups GEL and INJ represent those who received steroids in the form of triamcinolone soaked gelfoam or direct injection of dilute triamcinolone to the frontal/periorbital region, respectively. Group NON did not receive any periorbital steroids. MAIN OUTCOME MEASURE(S): Peri-operative outcomes including hospital length of stay and complications were evaluated based on method of periorbital steroid administration. Variables predictive of infectious complications were assessed using stepwise logistic regression. RESULTS: Four hundred and twelve patients were included in our sample (INJ:249, GEL:87, NON:76). Patients in the INJ group had a higher ASA class (P < .001) while patients in the NON group were significantly more likely to be syndromic (P < .001) and have multisuture craniosynostosis (P < .001). Rate of infectious complications for each cohort were NON: 2.6%, INJ: 4.4%, and GEL: 10.3%. There was no significant difference between groups in hospital length of stay (P = .654) or rate of post-operative infectious complications (P = .061). Increased ASA class (P = .021), increased length of stay (P = .016), and increased intraoperative narcotics (P = .011) were independent predictors of infectious complications. CONCLUSIONS: We identified a dose-dependent relationship between periorbital steroids and rate of postoperative infections, with key contributions from ASA class, hospital length of stay, and dose of intraoperative narcotics.

Intestinal tuft cell immune privilege enables norovirus persistence.

The persistent murine norovirus strain MNVCR6 is a model for human norovirus and enteric viral persistence. MNVCR6 causes chronic infection by directly infecting intestinal tuft cells, rare chemosensory epithelial cells. Although MNVCR6 induces functional MNV-specific CD8+ T cells, these lymphocytes fail to clear infection. To examine how tuft cells promote immune escape, we interrogated tuft cell interactions with CD8+ T cells by adoptively transferring JEDI (just EGFP death inducing) CD8+ T cells into Gfi1b-GFP tuft cell reporter mice. Unexpectedly, some intestinal tuft cells partially resisted JEDI CD8+ T cell-mediated killing-unlike Lgr5+ intestinal stem cells and extraintestinal tuft cells-despite seemingly normal antigen presentation. When targeting intestinal tuft cells, JEDI CD8+ T cells predominantly adopted a T resident memory phenotype with decreased effector and cytotoxic capacity, enabling tuft cell survival. JEDI CD8+ T cells neither cleared nor prevented MNVCR6 infection in the colon, the site of viral persistence, despite targeting a virus-independent antigen. Ultimately, we show that intestinal tuft cells are relatively resistant to CD8+ T cells independent of norovirus infection, representing an immune-privileged niche that can be leveraged by enteric microbes.

Genome-wide CRISPR activation screen identifies JADE3 as an antiviral activator of NF-kB-dependent IFITM3 expression.

The innate immune system features a web of interacting pathways that require exquisite regulation. To identify novel nodes in this immune landscape, we conducted a gain-of-function, genome-wide CRISPR activation screen with influenza A virus. We identified both appreciated and novel antiviral genes, including Jade family PHD zinc finger 3 (JADE3) a protein involved in directing the histone acetyltransferase histone acetyltransferase binding to ORC1 complex to modify chromatin and regulate transcription. JADE3 is both necessary and sufficient to restrict influenza A virus infection. Our results suggest a distinct function for JADE3 as expression of the closely related paralogs JADE1 and JADE2 does not confer resistance to influenza A virus infection. JADE3 is required for both constitutive and inducible expression of the well-characterized antiviral gene interferon-induced transmembrane protein 3 (IFITM3). Furthermore, we find JADE3 activates the NF-kB signaling pathway, which is required for the promotion of IFITM3 expression by JADE3. Therefore, we propose JADE3 activates an antiviral genetic program involving NF-kB-dependent IFITM3 expression to restrict influenza A virus infection.

Corrigendum: Iatrogenic air embolism: pathoanatomy, thromboinflammation, endotheliopathy, and therapies.

[This corrects the article DOI: 10.3389/fimmu.2023.1230049.].