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Background: and Purpose: Outdoor play is widely acknowledged for its benefits to physical health and psychological well-being, yet its relationship with school readiness remains understudied in preschoolers. To address this gap in the literature, this study investigated how outdoor play relates to cognitive and psychosocial development among a nationally representative sample of preschoolers., Materials and Method: Data on the duration of outdoor play (on weekdays and weekends) and specific cognitive and psychosocial outcome variables (i.e., early learning skills, self-regulation, social-emotional development, and flourishing) were collected via questionnaires provided to the caregivers of preschoolers. Logistic regressions were performed to examine the associations between outdoor play with cognitive and psychosocial outcomes while adjusting for covariates and calculate the odds ratio with 95 % confidence intervals (CI)., Results: Among the 10,682 preschoolers (i.e., 3-5y) included in this study (i.e., 5558 boys, Mage = 3.98 ± 0.99 years), there was a lower percentage of preschoolers engaged in outdoor play for over 3 h per day on weekdays (33.39 %) compared to weekends (56.85 %). Outdoor play in preschoolers is positively associated with various domains of school readiness, and engaging in outdoor play for more than 3 h per day is associated with more beneficial outcomes., Conclusion: The findings of this cross-sectional study suggest that promoting outdoor play among preschoolers can be an effective strategy for enhancing cognitive, social, and emotional development in this age group although further intervention studies are required to buttress this assumption empirically., Competing Interests: Declaration of competing interest The authors declare that they have no competing interests., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

In addition to its role as cellular energy currency, adenosine triphosphate (ATP) serves as an extracellular messenger that mediates diverse cell-to-cell communication. Compelling evidence supports that ATP is released from cells through pannexins, a family of membrane proteins that form heptameric large-pore channels. However, the activation mechanisms that trigger ATP release by pannexins remain poorly understood. Here, we discover lysophospholipids as endogenous pannexin activators, using activity-guided fractionation of mouse tissue extracts combined with untargeted metabolomics and electrophysiology. We show that lysophospholipids directly and reversibly activate pannexins in the absence of other proteins. Secretomics experiments reveal that lysophospholipid-activated pannexin 1 leads to the release of not only ATP but also other signaling metabolites, such as 5'-methylthioadenosine, which is important for immunomodulation. We also demonstrate that lysophospholipids activate endogenous pannexin 1 in human monocytes, leading to the release of IL-1β through inflammasome activation. Our results provide a connection between lipid metabolism and purinergic signaling, both of which play major roles in immune responses.

Objective: The International Code of Marketing of Breast-Milk Substitutes is an important instrument to protect and promote appropriate infant and young child feeding and the safe use of commercial milk formulas. Ghana and Tanzania implemented the Code into national legislation in 2000 and 1994, respectively. We aimed to estimate the effects of the Code implementation on child mortality (CM) in both countries., Setting: The countries analysed were Ghana and Tanzania., Participants: For CM and HIV rates, data from the Institute for Health Metrics and Evaluation from up to 2019 were used. Data for income and skilled birth rates were retrieved from the World Bank, for fertility from the World Population Prospects, for vaccination from the Global Health Observatory and for employment from the International Labour Organization., Design: We used the synthetic control group method and performed placebo tests to assess statistical inference. The primary outcomes were CM by lower respiratory infections, mainly pneumonia, and diarrhoea and the secondary outcome was overall CM., Results: One-sided inference tests showed statistically significant treatment effects for child deaths by lower respiratory infections in Ghana ( P = 0·0476) and Tanzania ( P = 0·0476) and for diarrhoea in Tanzania ( P = 0·0476). More restrictive two-sided inference tests showed a statistically significant treatment effect for child deaths by lower respiratory infections in Ghana ( P = 0·0476). No statistically significant results were found for overall CM., Conclusion: The results suggest that the implementation of the Code in both countries had a potentially beneficial effect on CM due to infectious diseases; however, further research is needed to corroborate these findings.

Glycosylation of recombinant proteins is a post-translational modification that affects multiple physicochemical and biological properties of proteins. As such, it is a critical quality attribute that must be carefully controlled during protein production in the pharmaceutical industry. Glycosylation can be modulated by various conditions, including the composition of production media and feeds. In this study, the N-glycosylation-modulating effects of numerous compounds, including metal enzyme cofactors, enzyme inhibitors, and metabolic intermediates, were evaluated. Chinese hamster ovary cells producing three different IgG antibodies were cultivated in a fed-batch mode. First, a one-factor-at-a-time experiment was performed in 24-well deep well plates to identify the strongest modulators and appropriate concentration ranges. Then, a full response surface experiment was designed to gauge the effects and interactions of the 14 most effective hit compounds in an Ambr ® 15 bioreactor system. A wide range of glycoform content was achieved, with an up to eight-fold increase in individual glycoforms compared to controls. The resulting model can be used to determine modulator combinations that will yield desired glycoforms in the final product., Competing Interests: Authors LK, MM, and MB were employed by Novartis Pharmaceutical Manufacturing LLC. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The author(s) declared that they were an editorial board member of Frontiers, at the time of submission. This had no impact on the peer review process and the final decision., (Copyright © 2024 Kranjc, Kramer, Mikelj, Anderluh, Pišlar and Brinc.)

The microbial communities residing in the mosquito midgut play a key role in determining the outcome of mosquito pathogen infection. Elizabethkingia anophelis, originally isolated from the midgut of Anopheles gambiae possess a broad-spectrum antiviral phenotype, yet a gap in knowledge regarding the mechanistic basis of its interaction with viruses exists. The current study aims to identify pathways and genetic factors linked to E. anophelis antiviral activity. The understanding of E. anophelis antiviral mechanism could lead to novel transmission barrier tools to prevent arboviral outbreaks. We utilized a non-targeted multi-omics approach, analyzing extracellular lipids, proteins, metabolites of culture supernatants coinfected with ZIKV and E. anophelis. We observed a significant decrease in arginine and phenylalanine levels, metabolites that are essential for viral replication and progression of viral infection. This study provides insights into the molecular basis of E. anophelis antiviral phenotype. The findings lay a foundation for in-depth mechanistic studies., (© 2024. The Author(s).)

Abstract: Plasma pTau181, a marker of amyloid and tau burden, was evaluated as a prognostic predictor of clinical decline and Alzheimer's disease (AD) progression of amyloid-positive (Aβ+) patients with mild cognitive impairment (MCI). The training cohort for constructing the Bayesian prediction models comprised 135 Aβ+ MCI clinical trial placebo subjects. Performance was evaluated in two validation cohorts. An 18-month ≥1 increase in the Clinical Dementia Rating Sum of Boxes was the clinical decline criterion. Baseline plasma pTau181 concentration matched clinical assessments' prediction performance. Adding pTau181 to clinical assessments significantly improved the prediction of an 18-month clinical decline and the 36-month progression from Aβ+ MCI to AD. The area under the receiver operating characteristic curve for the latter increased from 71.8% to 79%, and the hazard ratio for time-to-progression improved from 2.26 to 3.11 ( p  < 0.0001). Baseline plasma pTau181 has the potential for identifying Aβ+ MCI subjects with faster clinical decline over time., Highlights: This study assessed pTau181 as a prognostic predictor of 18-month clinical decline and extended progression to Alzheimer's disease (AD) in amyloid-positive patients with mild cognitive impairment (Aβ+ MCI).The research findings underscore the promise of baseline plasma pTau181 as a screening tool for identifying Aβ+ MCI individuals with accelerated clinical decline within a standard 18-month clinical trial period. The predictive accuracy is notably enhanced when combined with clinical assessments.Similar positive outcomes were noted in forecasting the extended progression of Aβ+ MCI subjects to AD., Competing Interests: VD, YH, HH, LK, SD, and MI are employees of Eisai Inc. HH is a Reviewing Editor and previously a Senior Associate Editor for the journal Alzheimer's & Dementia, and he was not involved in the editorial process. There are no competing disclosures to report for DAL. Author disclosures are available in the Supporting Information., (© 2024 Eisai Inc. Alzheimer's & Dementia: Diagnosis, Assessment & Disease Monitoring published by Wiley Periodicals LLC on behalf of Alzheimer's Association.)

This systematic review addresses the gap in the literature regarding sibling relationship quality among Latinx families, a topic that has not been comprehensively examined to date. This study aimed to synthesize current research on sibling relationship quality in Latinx families, focusing on the influence of cultural factors, identifying key variables associated with sibling relationship quality, and evaluating the methodological approaches used. This paper is a systematic review based on a recently published evidence and gap map (EGM) that identified and visually presented all published studies investigating sibling relationship quality. Additional inclusion criteria were applied to select articles that specifically examined sibling relationship quality in the United States samples with at least 50% Latinx participants. The review included 12 articles representing 6 distinct studies, revealing significant findings on the roles of familism, simpatía, and gender in shaping sibling dynamics. Results indicated that cultural values such as familism and simpatía positively influence sibling intimacy and warmth, while gender dynamics further moderate these relationships. Methodologically, the articles employed longitudinal and cross-sectional designs, utilizing various quantitative measures. The findings underscore the importance of culturally sensitive approaches in studying sibling relationships and highlight the need for further research to explore these dynamics in diverse Latinx subgroups.

Background: Alzheimer disease (AD) is a major health problem of aging, with tremendous burden on healthcare systems, patients, and families globally. Lecanemab, an FDA-approved amyloid beta (Aβ)-directed antibody indicated for the treatment of early AD, binds with high affinity to soluble Aβ protofibrils, which have been shown to be more toxic to neurons than monomers or insoluble fibrils. Lecanemab has been shown to be well tolerated in multiple clinical trials, although risks include an increased rate of amyloid-related imaging abnormalities (ARIA) and infusion reactions relative to placebo., Methods: Clarity AD was an 18-month treatment (Core study), multicenter, double-blind, placebo-controlled, parallel-group study with open-label extension (OLE) in participants with early AD. Eligible participants were randomized 1:1 across 2 treatment groups (placebo and lecanemab 10 mg/kg biweekly). Safety evaluations included monitoring of vital signs, physical examinations, adverse events, clinical laboratory parameters, and 12-lead electrocardiograms. ARIA occurrence was monitored throughout the study by magnetic resonance imaging, read both locally and centrally., Results: Overall, 1795 participants from Core and 1612 participants with at least one dose of lecanemab (Core + OLE) were included. Lecanemab was generally well-tolerated in Clarity AD, with no deaths related to lecanemab in the Core study. There were 9 deaths during the OLE, with 4 deemed possibly related to study treatment. Of the 24 deaths in Core + OLE, 3 were due to intracerebral hemorrhage (ICH): 1 placebo in the Core due to ICH, and 2 lecanemab in OLE with concurrent ICH (1 on tissue plasminogen activator and 1 on anticoagulant therapy). In the Core + OLE, the most common adverse events in the lecanemab group (> 10%) were infusion-related reactions (24.5%), ARIA with hemosiderin deposits (ARIA-H) microhemorrhages (16.0%), COVID-19 (14.7%), ARIA with edema (ARIA-E; 13.6%), and headache (10.3%). ARIA-E and ARIA-H were largely radiographically mild-to-moderate. ARIA-E generally occurred within 3-6 months of treatment, was more common in ApoE e4 carriers (16.8%) and most common in ApoE ε4 homozygous participants (34.5%)., Conclusions: Lecanemab was generally well-tolerated, with the most common adverse events being infusion-related reactions, ARIA-H, ARIA-E. Clinicians, participants, and caregivers should understand the incidence, monitoring, and management of these events for optimal patient care., Trial Registration: ClinicalTrials.gov numbers: Clarity AD NCT03887455)., (© 2024. The Author(s).)

Introduction: Health-Related Quality of Life (HRQoL) data is frequently requested in early benefit assessment in Germany. To test the hypothesis that the importance of HRQoL in general and the significance of disease-specific instruments in particular has increased since the introduction of AMNOG in Germany, we analysed all early benefit assessments between 2011 and 2022., Methods: All 793 early benefit assessments completed between 01/01/2011 and 03/11/2022 were systematically analysed. The HRQoL instruments featured in the dossier submissions were extracted for all assessments and categorized into generic and specific instruments. All G-BA resolutions were likewise assessed for consideration and acceptance of generic and specific HRQoL instruments. In addition, it was examined whether there was an association between HRQoL data and the extent of additional benefit., Results: Since 2014 HRQoL data have continuously been submitted in 70% to 80% of assessments per year with the exception of 2022 (63%). Generally, disease-specific instruments are favoured, regarding submissions by industry but especially with higher acceptance by the G-BA in the resolution. Subgroup analyses revealed oncology as a major contributor to the submission and acceptance rates of disease-specific instruments. Disease-specific instruments were submitted in 81% of all oncology assessments and accepted in 53% of assessments. Overall, assessments with accepted HRQoL data tend to reach a higher overall benefit. Procedures with accepted HRQoL were most likely to receive a considerable benefit (31%), while for procedures in which HRQoL data were not accepted, a benefit was most often (65%) not proven., Discussion: Industry has followed the request for submission of specific HRQoL instruments early on. Higher submission rates of specific instruments over time which at the same time meet the methodological requirements indicate that industry has learned from early assessments. A potential reason for the high submission- and acceptance rates of specific HRQoL instruments in oncology might be the particularly high relevance of HRQoL in this indication. Possible effects of changes in legislature on the future development of submission and acceptance of HRQoL data need to be monitored., Conclusion: In Germany, HRQoL has gained a relevant position in early benefit assessment over time. Overall specific instruments are favoured, regarding submissions by industry but especially through consideration by the G-BA in the resolution. Furthermore, HRQoL data can be supportive for benefit assessments, in particular to show that advantages in morbidity and/or mortality are reflected in HRQoL and not at the expense of HRQoL., (Copyright © 2024. Published by Elsevier GmbH.)

Background: A severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection lasts longer in immunocompromised hosts than in immunocompetent patients. Prolonged infection is associated with a higher probability of selection for novel SARS-CoV-2 mutations, particularly in the spike protein, a critical target for vaccines and therapeutics., Methods: From December 2020 to September 2022, respiratory samples from 444 immunocompromised patients and 234 health care workers positive for SARS-CoV-2, diagnosed at 2 hospitals in Paris, France, were analyzed using whole-genome sequencing using Nanopore technology. Custom scripts were developed to assess the SARS-CoV-2 genetic diversity between the 2 groups and within the host., Results: Most infections were SARS-CoV-2 Delta or Omicron lineages. Viral genetic diversity was significantly higher in infections of immunocompromised patients than those of controls. Minor mutations were identified in viruses sequenced from immunocompromised individuals, which became signature mutations for newer SARS-CoV-2 variants as the epidemic progressed. Two patients were coinfected with Delta and Omicron variants. The follow-up of immunocompromised patients revealed that the SARS-CoV-2 genome evolution differed in the upper and lower respiratory tracts., Conclusions: This study found that SARS-CoV-2 infection in immunocompromised patients is associated with higher genetic diversity, which could lead to the emergence of new SARS-CoV-2 variants with possible immune evasion or different virulence characteristics., Competing Interests: Potential conflicts of interest. All authors: No reported conflicts. All authors have submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest. Conflicts that the editors consider relevant to the content of the manuscript have been disclosed., (© The Author(s) 2023. Published by Oxford University Press on behalf of Infectious Diseases Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Complete blood counts (CBCs) measure the abundance of individual immune cells, red blood cells, and related measures such as platelets in circulating blood. These measures can indicate the health status of an animal; thus, baseline circulating levels in a healthy animal may be related to the productive life, resilience, and production efficiency of cattle. The objective of this study is to determine the heritability of CBC traits and identify genomic regions that are associated with CBC measurements in lactating Holstein dairy cattle. The heritability of CBCs was estimated using a Bayes C0 model. The study population consisted of 388 cows with genotypes at roughly 75,000 markers and 16 different CBC phenotypes taken at one to three time points (n = 33, 131, and 224 for 1, 2, and 3 time points, respectively). Heritabilities ranged from 0.00 ± 0.00 (red cell distribution width) to 0.68 ± 0.06 (lymphocytes). A total of 96 different 1-Mb windows were identified that explained more than 1% of the genetic variance for at least one CBC trait, with 10 windows explaining more than 1% of the genetic variance for two or more traits. Multiple genes in the identified regions have functions related to immune response, cell differentiation, anemia, and disease. Positional candidate genes include RAD52 motif-containing protein 1 ( RDM1 ), which is correlated with the degree of immune infiltration of immune cells, and C-X-C motif chemokine ligand 12 ( CXCL12 ), which is critically involved in neutrophil bone marrow storage and release regulation and enhances neutrophil migration. Since animal health directly impacts feed intake, understanding the genetics of CBCs may be useful in identifying more disease-resilient and feed-efficient dairy cattle. Identification of genes responsible for variation in CBCs will also help identify the variability in how dairy cattle defend against illness and injury., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Siberski-Cooper, Mayes, Gorden, Kramer, Bhatia and Koltes.)

Background: Models for forecasting individual clinical progression trajectories in early Alzheimer's disease (AD) are needed for optimizing clinical studies and patient monitoring., Methods: Prediction models were constructed using a clinical trial training cohort (TC; n = 934) via a gradient boosting algorithm and then evaluated in two validation cohorts (VC 1, n = 235; VC 2, n = 421). Model inputs included baseline clinical features (cognitive function assessments, APOE ε4 status, and demographics) and brain magnetic resonance imaging (MRI) measures., Results: The model using clinical features achieved R 2 of 0.21 and 0.31 for predicting 2-year cognitive decline in VC 1 and VC 2, respectively. Adding MRI features improved the R 2 to 0.29 in VC 1, which employed the same preprocessing pipeline as the TC. Utilizing these model-based predictions for clinical trial enrichment reduced the required sample size by 20% to 49%., Discussion: Our validated prediction models enable baseline prediction of clinical progression trajectories in early AD, benefiting clinical trial enrichment and various applications., (© 2023 Eisai Inc. Alzheimer's & Dementia published by Wiley Periodicals LLC on behalf of Alzheimer's Association.)

Objective: Many factors contribute to inadequate diversity in Alzheimer disease (AD) clinical trials. We evaluated eligibility rates among racial and ethnic groups at US sites in large global multisite trials in early AD., Methods: Using screening data from 4 randomized, double-blind, placebo-controlled clinical trials in early AD, we assessed rates of eligibility among racial and ethnic groups controlling for other demographic covariates. Each trial incorporated positron emission tomography and/or cerebrospinal fluid to evaluate brain amyloid pathology, as well as typical eligibility criteria used in early AD trials., Results: Across the trials, 10,804 US participants were screened: 193 (2%) were of Hispanic ethnicity and Black race, 2,624 (25%) were of Hispanic ethnicity and White race, 118 (1%) were of non-Hispanic ethnicity (NH) and Asian race, 696 (7%) were of NH ethnicity and Black race, and 7,017 (65%) were of NH ethnicity and White race. Data from 156 participants who did not fit into these categories were excluded. Accounting for age, sex, and trial and using NH White participants as a reference group, we observed higher probabilities of ineligibility for amyloid biomarker criteria among Hispanic Black (odds ratio [OR] = 3.20, 95% confidence interval [CI] = 2.11-4.88), Hispanic White (OR = 4.15, 95% CI = 3.58-4.83), NH Asian (OR = 2.35, 95% CI = 1.23-4.55), and NH Black (OR = 3.75, 95% CI = 2.80-5.06) participants., Interpretation: Differential eligibility may contribute to underrepresentation of some minoritized racial and ethnic groups in early AD trials. Amyloid biomarker eligibility is a requirement to confirm the diagnosis of AD and for treatment with amyloid-lowering drugs and differed among racial and ethnic groups. ANN NEUROL 2024;95:288-298., (© 2023 The Authors. Annals of Neurology published by Wiley Periodicals LLC on behalf of American Neurological Association.)

Chemicals in the aquatic environment can be harmful to organisms and ecosystems. Knowledge on effect concentrations as well as on mechanisms and modes of interaction with biological molecules and signaling pathways is necessary to perform chemical risk assessment and identify toxic compounds. To this end, we developed criteria and a pipeline for harvesting and summarizing effect concentrations from the US ECOTOX database for the three aquatic species groups algae, crustaceans, and fish and researched the modes of action of more than 3,300 environmentally relevant chemicals in literature and databases. We provide a curated dataset ready to be used for risk assessment based on monitoring data and the first comprehensive collection and categorization of modes of action of environmental chemicals. Authorities, regulators, and scientists can use this data for the grouping of chemicals, the establishment of meaningful assessment groups, and the development of in vitro and in silico approaches for chemical testing and assessment., (© 2024. The Author(s).)

The Farm Animal Genotype-Tissue Expression (FarmGTEx) project has been established to develop a public resource of genetic regulatory variants in livestock, which is essential for linking genetic polymorphisms to variation in phenotypes, helping fundamental biological discovery and exploitation in animal breeding and human biomedicine. Here we show results from the pilot phase of PigGTEx by processing 5,457 RNA-sequencing and 1,602 whole-genome sequencing samples passing quality control from pigs. We build a pig genotype imputation panel and associate millions of genetic variants with five types of transcriptomic phenotypes in 34 tissues. We evaluate tissue specificity of regulatory effects and elucidate molecular mechanisms of their action using multi-omics data. Leveraging this resource, we decipher regulatory mechanisms underlying 207 pig complex phenotypes and demonstrate the similarity of pigs to humans in gene expression and the genetic regulation behind complex phenotypes, supporting the importance of pigs as a human biomedical model., (© 2024. The Author(s).)

Mutations in genes encoding epigenetic regulators are commonly observed at relapse in B cell acute lymphoblastic leukemia (B-ALL). Loss-of-function mutations in SETD2, an H3K36 methyltransferase, have been observed in B-ALL and other cancers. Previous studies on mutated SETD2 in solid tumors and acute myelogenous leukemia support a role in promoting resistance to DNA damaging agents. We did not observe chemoresistance, an impaired DNA damage response, nor increased mutation frequency in response to thiopurines using CRISPR-mediated knockout in wild-type B-ALL cell lines. Likewise, restoration of SETD2 in cell lines with hemizygous mutations did not increase sensitivity. SETD2 mutations affected the chromatin landscape and transcriptional output that was unique to each cell line. Collectively our data does not support a role for SETD2 mutations in driving clonal evolution and relapse in B-ALL, which is consistent with the lack of enrichment of SETD2 mutations at relapse in most studies.

Plasmodium berghei ANKA (PbA) infection in mice resembles several aspects of severe malaria in humans, such as cerebral malaria and acute respiratory distress syndrome. Herein, the effects of N -(coumarin-3-yl)cinnamamide (M220) against severe experimental malaria have been investigated. Treatment with M220 proved to protect cognitive abilities and lung function in PbA-infected mice, observed by an object recognition test and spirometry, respectively. In addition, treated mice demonstrated decreased levels of brain and lung inflammation. The production and accumulation of microglia, and immune cells that produce the inflammatory cytokines TNF and IFN-γ, decreased, while the production of the anti-inflammatory cytokine IL-10 by innate and adaptive immune cells was enhanced. Treatment with M220 promotes immunomodulatory, neuroprotective, and lung function-preserving effects during experimental severe malaria. Therefore, it may be an interesting therapeutic candidate to treat severe malaria effects.

Numerous biting and nuisance insects are a noted cause of discomfort and stress to horses. Pyrethrins and pyrethroids have been used for many years in numerous formulations for the control of insect pests in animals, humans and environment. There are, however, few studies reporting their field efficacy in horses. The aim of the present study was to evaluate the repellent activity of a spray formulation based on prallethrin and permethrin synergized with piperonyl butoxide (BRONCO® Equine Fly Spray, Farnam Companies, Inc., USA) against annoying and harmful insects for horses in field conditions. Nine horses of mixed breed were divided into 2 groups (treatment and control). Pre-treatment insect counts were compared to daily counts for 4 days post-treatment (pt). One minute after the administration of the product (day 0), all the horses were negative for the presence of insects. All counts up to the 6-h pt check remained negative for Hippobosca equina, tabanid flies and Simulium spp., showing 100% efficacy. This remained above 90% throughout the study. For the H. equina, the repellent efficacy remained > 99.7% for all 4 days pt, for tabanid flies > 93.3% and for Simulium spp. > 97.4%. The efficacy against Musca spp. decreased from 82.2% at day 0 to 62.2% at day 3. Treatment was well-tolerated. In conclusion, despite the low number of tested horses, Bronco® has demonstrated high insecticide and repellent efficacy and a good persistence, maintained for up to 4 days post-treatment, against the most common species of insects harmful for horses., (© 2023. The Author(s).)

The zoonotic nematodes Dirofilaria immitis and Dirofilaria repens are endemic in many European countries and their distribution is changing. A correct diagnosis is the starting point to reduce their spread and veterinary practitioners represent the first line of defense against these two vector-borne diseases in pets and humans. With the aim to better understand current knowledge by veterinarians and to compare them with the previous survey, an online questionnaire was sent to Italian companion animal veterinary facilities. The overall response rate was 22.3%. Approximately 50% and 5% of facilities have diagnosed at least one case of Dirofilaria spp. in the last year in dogs and cats, respectively. In agreement with international guidelines, of the 665 facilities that responded to the questionnaire, 16.8% used Knott's test in combination with serology (Antigenic test-Ag-) to diagnose heartworm infection in dogs. For the diagnosis of D. repens infection, 13.8% used Knott's test in dogs and cats, and 10.5% used Knott's test only in dogs. Most facilities recommended beginning prevention of canine heartworm disease in April and ending in November. Injectable moxidectin and oral ivermectin were the two most commonly used preventives for D. immitis in dogs, while selamectin was most frequently used in cats. The drug most commonly used for heartworm adulticide treatment in dogs was melarsomine followed by the combined ivermectin-doxycycline treatment. The results of this study suggest that, on the one hand, veterinarians are aware of the recent developments in adulticide therapy, while on the other hand, diagnosis is not always made according to what is recommended by international guidelines and that the start and end times of prevention may not be optimal. Furthermore, the data show an increase in cases of coinfection compared to the previous survey., Competing Interests: Declaration of Competing Interest The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.)

Protein tyrosine phosphatases (PTPs) are emerging drug targets for many diseases, including cancer, autoimmunity, and neurological disorders. A high degree of structural similarity between their catalytic domains, however, has hindered the development of selective pharmacological agents. Our previous research uncovered two unfunctionalized terpenoid inhibitors that selectively inhibit PTP1B over T-cell PTP (TCPTP), two PTPs with high sequence conservation. Here, we use molecular modeling, with supporting experimental validation, to study the molecular basis of this unusual selectivity. Molecular dynamics (MD) simulations suggest that PTP1B and TCPTP share a h-bond network that connects the active site to a distal allosteric pocket; this network stabilizes the closed conformation of the catalytically essential WPD loop, which it links to the L-11 loop and neighboring α3 and α7 helices on the other side of the catalytic domain. Terpenoid binding to either of two proximal C-terminal sites─an α site and a β site─can disrupt the allosteric network; however, binding to the α site forms a stable complex only in PTP1B. In TCPTP, two charged residues disfavor binding at the α site in favor of binding at the β site, which is conserved between the two proteins. Our findings thus indicate that minor amino acid differences at the poorly conserved α site enable selective binding, a property that might be enhanced with chemical elaboration, and illustrate more broadly how minor differences in the conservation of neighboring─yet functionally similar─allosteric sites can affect the selectivity of inhibitory scaffolds (e.g., fragments).

Calculus, the study of change in processes and systems, serves as the foundation for many STEM disciplines. Traditional, lecture-based calculus instruction may present a barrier for students seeking STEM degrees, limit their access to STEM professions, and block their potential to address society's challenges. A large-scale pragmatic trial with randomized student allocation was conducted to compare two calculus instruction styles: active student engagement (treatment condition) versus traditional, lecture-based instruction (control condition). A sample of 811 university students were studied across 32 sections taught by 19 instructors over three semesters at a large, US-based Hispanic-serving institution. Large effect sizes were consistently measured for student learning outcomes in the treatment condition, which demonstrates a new standard for calculus instruction and increased opportunities for completion of STEM degrees.

Coral reef ecosystems are being fundamentally restructured by local human impacts and climate-driven marine heatwaves that trigger mass coral bleaching and mortality 1 . Reducing local impacts can increase reef resistance to and recovery from bleaching 2 . However, resource managers lack clear advice on targeted actions that best support coral reefs under climate change 3 and sector-based governance means most land- and sea-based management efforts remain siloed 4 . Here we combine surveys of reef change with a unique 20-year time series of land-sea human impacts that encompassed an unprecedented marine heatwave in Hawai'i. Reefs with increased herbivorous fish populations and reduced land-based impacts, such as wastewater pollution and urban runoff, had positive coral cover trajectories predisturbance. These reefs also experienced a modest reduction in coral mortality following severe heat stress compared to reefs with reduced fish populations and enhanced land-based impacts. Scenario modelling indicated that simultaneously reducing land-sea human impacts results in a three- to sixfold greater probability of a reef having high reef-builder cover four years postdisturbance than if either occurred in isolation. International efforts to protect 30% of Earth's land and ocean ecosystems by 2030 are underway 5 . Our results reveal that integrated land-sea management could help achieve coastal ocean conservation goals and provide coral reefs with the best opportunity to persist in our changing climate., (© 2023. This is a U.S. Government work and not under copyright protection in the US; foreign copyright protection may apply.)

Neutral mutational drift is an important source of biological diversity that remains underexploited in fundamental studies of protein biophysics. This study uses a synthetic transcriptional circuit to study neutral drift in protein tyrosine phosphatase 1B (PTP1B), a mammalian signaling enzyme for which conformational changes are rate limiting. Kinetic assays of purified mutants indicate that catalytic activity, rather than thermodynamic stability, guides enrichment under neutral drift, where neutral or mildly activating mutations can mitigate the effects of deleterious ones. In general, mutants show a moderate activity-stability tradeoff, an indication that minor improvements in the activity of PTP1B do not require concomitant losses in its stability. Multiplexed sequencing of large mutant pools suggests that substitutions at allosterically influential sites are purged under biological selection, which enriches for mutations located outside of the active site. Findings indicate that the positional dependence of neutral mutations within drifting populations can reveal the presence of allosteric networks and illustrate an approach for using synthetic transcriptional systems to explore these mutations in regulatory enzymes., (© 2023 The Protein Society.)

Historically, in the United States, scientific racism has been used to justify differential treatment of Black, Indigenous, People of Color (BIPOC) populations compared with the White population. Discrimination toward BIPOC populations by the medical community has contributed to racial and ethnic disparities in health care that persist to this day. At the 2022 American Society of Clinical Psychopharmacology Annual Meeting, a panel of 5 experts from the areas of academia, advocacy, and clinical research discussed racial and ethnic disparities in mental health care. This Academic Highlights summarizes and expands on that discussion by tracing the history of scientific racism from the colonization of the US to present-day health inequities, illustrating the ongoing problem of low diversity in clinical trials, and outlining solutions through community engagement., (© Copyright 2023 Physicians Postgraduate Press, Inc.)

Background: The lungworm Aelurostrongylus abstrusus infects wild and domestic feline species worldwide and is considered a primary respiratory parasite of cats. Definitive diagnosis is based on the identification of first-stage larvae (L1s) released in faeces approximately 5 to 6 weeks after infection. More recently, serology has been shown to be a diagnostic alternative for A. abstrusus infection in cats. The present study aimed at evaluating the diagnostic performance of serological antibody detection compared to faecal examination for A. abstrusus infection in a population of cats with known infection status from endemic areas in Italy and to identify factors (larval scores, age, co-infections with other helminths) that may influence test sensitivity and specificity of serology., Methods: All cats resulting positive using the Baermann technique (n = 78) were tested with the A. abstrusus ELISA. An additional 90 serum samples from cats living in three geographical areas with infection prevalence > 10%, but that resulted negative on Baermann, were also tested., Results: Among 78 cats copromicroscopically positive for L1s of A. abstrusus (Group 1), 29 (37.2%) were seropositive in ELISA. Of the 90 cats from Group 2 (cats living in three geographical areas in Italy with A. abstrusus prevalence > than 10%, but negative on Baermann examination), 11 (12.2%) were positive on ELISA. The overall seroprevalence was 23.8%. There was no statistical difference either between average optical density (OD) values of cats excreting > 100 L1s vs. cats excreting < 100 L1s (0.84 vs. 0.66; P value = 0.3247) or comparing the OD values with age of infected cats. Few Baermann-negative cats positive for Toxocara cati or hookworms were seropositive, supporting lack of cross-reactivity to these nematodes., Conclusions: Results from the present study suggest that relying solely on faecal examination may underestimate prevalence of A. abstrusus infection in cats and that field surveys based on antibody detection are useful for establishing true prevalence of infected and/or exposed animals., (© 2023. The Author(s).)

Cefiderocol (FDC) is a siderophore cephalosporin now recognized as a new weapon in the treatment of difficult-to-treat-resistant (DTR) Gram-negative pathogens, including carbapenemase-producing enterobacterales and non-fermentative Gram-negative bacilli (GNB). This article reports our experience with an FDC-based regimen in the treatment of 16 extremely severe patients (invasive mechanical ventilation, 15/16; extracorporeal membrane oxygenation, 9/16; and renal replacement therapy, 8/16) infected with DTR GNB. Our case series provides detailed insight into the pharmacokinetic profile and the microbiological data in real-life conditions. In the narrative review, we discuss the interest of FDC in the treatment of non-fermentative GNB in critically ill patients. We reviewed the microbiological spectrum, resistance mechanisms, pharmacokinetics/pharmacodynamics, efficacy and safety profiles, and real-world evidence for FDC. On the basis of our experience and the available literature, we discuss the optimal FDC-based regimen, FDC dosage, and duration of therapy in critically ill patients with DTR non-fermentative GNB infections.

Background: Several studies in both experimentally and naturally infected dogs have reported the adulticide effect of a combination of macrocyclic lactones and doxycycline against Dirofilaria immitis, showing that these protocols can be used as an alternative to melarsomine. The present study evaluated the efficacy of oral, topical and extended-release injectable formulations of moxidectin when combined with doxycycline in dogs naturally infected with D. immitis from a shelter located in southern Italy., Methods: Thirty dogs with naturally acquired D. immitis infection were divided in three groups (G) and treated with oral moxidectin (G1) once a month for 9 consecutive months, topical moxidectin (G2) once a month for 9 consecutive months or extended release moxidectin injectable (G3) at enrolment and again at 6 months (Day 180). All treatment groups received doxycycline for the first 30 days. Microfilarial concentrations in 1 ml (mff/ml) blood were determined monthly for 9 months with the modified Knott's test. A clinical scoring system was employed for each dog enrolled in the study based on thoracic radiography and cardiac ultrasound (CU) examinations performed at Day - 15 (before treatment) and at Day 180., Results: Results from the present study suggest that the majority of dogs from all treatment groups became antigen negative, as evaluated at Day 270: 9/10 dogs (90.0%) from G1, 6/10 dogs (60.0%) from G2 and 8/10 dogs (80.0%) from G3. Improvement of radiographic alterations was observed in all treatment groups, and almost all dogs were cleared of pulmonary abnormalities by 6 months from the beginning of treatment (P = 0.000). Cardiac ultrasound examination showed a progressive improvement of cardiac function in a limited number of animals (4/30)., Conclusions: The combination of doxycycline and three different formulations of moxidectin leads to antigen-negative status in naturally infected dogs., (© 2023. The Author(s).)

The extracellular matrix (ECM) obtained by decellularization provides scaffolds with the natural complex architecture and biochemical composition of the target organ. Whole kidney decellularization by perfusion uses the vasculature to remove cells leaving a scaffold that can be recellularized with patient-specific cells. However, decellularization and recellularization are highly complex processes that require intensive optimization of various parameters. In pursuit of this, a huge number of animals must be sacrificed. Therefore, we used precision-cut kidney slices (PCKS) as a source of natural scaffolds, which were decellularized by immersion in chemical reagents allowing the examination of more parameters with less animals. However, chemical reagents have a damaging effect on the structure and components of the ECM. Therefore, this study aimed at investigating the effects of physical treatment methods on the effectiveness of PCKS decellularization by immersion in chemical reagents (CHEM). PCKS were treated physically before or during immersion in chemicals (CHEM) with high hydrostatic pressure (HHP), freezing-thawing cycles (FTC) or in an ultrasonic bath system (UBS). Biochemical and DNA quantification as well as structural evaluation with conventional histology and scanning electron microscopy (SEM) were performed. Compared to decellularization by CHEM alone, FTC treatment prior to CHEM was the most effective in reducing DNA while also preserving glycosaminoglycan (GAG) content. Moreover, while UBS resulted in a comparable reduction of DNA, it was the least effective in retaining GAGs. In contrast, despite the pretreatment with HHP with pressures up to 200 MPa, it was the least effective in DNA removal. Histological scoring showed that HHP scaffolds received the best score followed by UBS, FTC and CHEM scaffolds. However further analysis with SEM demonstrated a higher deterioration of the ultrastructure in UBS scaffolds. Altogether, pretreatment with FTC prior to CHEM resulted in a better balance between DNA removal and structural preservation., (Creative Commons Attribution license.)

Background: Evidence suggests that dental anxiety and phobia are frequently the result of direct associative fear conditioning but that pre-exposure to dental stimuli prior to conditioning results in latent inhibition of fear learning. The mechanisms underlying the pre-exposure effect in humans, however, are poorly understood. Moreover, pain sensitivity has been linked to dental fear conditioning in correlational investigations and theory suggests it may moderate the latent inhibition effect, but this hypothesis has not been directly tested. These gaps in our understanding are a barrier to the development of evidence-based dental phobia prevention efforts., Methods: Healthy volunteers between the ages of 6 and 35 years will be enrolled across two sites. Participants will complete a conditioning task in a novel virtual reality environment, allowing for control over pre-exposure and the examination of behaviour. A dental startle (a brief, pressurized puff of air to a tooth) will serve as the unconditioned stimulus. Using a within-subjects experimental design, participants will experience a pre-exposed to-be conditioned stimulus, a non-pre-exposed to-be conditioned stimulus, and a neutral control stimulus. Two hypothesized mechanisms, changes in prediction errors and attention, are expected to mediate the association between stimulus condition and fear acquisition, recall, and retention. To ascertain the involvement of pain sensitivity, this construct will be measured through self-report and the cold pressor task., Discussion: Dental phobia negatively affects the dental health and overall health of individuals. This study aims to determine the mechanisms through which pre-exposure retards conditioned dental fear acquisition, recall, and retention. A randomized control trial will be used to identify these mechanisms so that they can be precisely targeted and maximally engaged in preventative efforts., (© 2023. The Author(s).)

Proteases are an important class of drug targets that continue to drive inhibitor discovery. These enzymes are prone to resistance mutations, yet their promise for treating viral diseases and other disorders continues to grow. This study develops a general approach for detecting microbially synthesized protease inhibitors and uses it to screen terpenoid pathways for inhibitory compounds. The detection scheme relies on a bacterial two-hybrid (B2H) system that links protease inactivation to the transcription of a swappable reporter gene. This system, which can accomodate multiple biochemical outputs (i.e., luminescence and antibiotic resistance), permitted the facile incorporation of four disease-relevant proteases. A B2H designed to detect the inactivation of the main protease of severe acute respiratory syndrome coronavirus 2 enabled the identification of a terpenoid inhibitor of modest potency. An analysis of multiple pathways that make this terpenoid, however, suggested that its production was necessary but not sufficient to confer a survival advantage in growth-coupled assays. This finding highlights an important challenge associated with the use of genetic selection to search for inhibitors─notably, the influence of pathway toxicity─and underlines the value of including multiple pathways with overlapping product profiles in pathway screens. This study provides a detailed experimental framework for using microbes to screen libraries of biosynthetic pathways for targeted protease inhibitors.

Introduction: We have recently developed a novel T cell engager concept by utilizing γ9δ2TCR as tumor targeting domain, named gamma delta TCR anti-CD3 bispecific molecule (GAB), targeting the phosphoantigen-dependent orchestration of BTN2A1 and BTN3A1 at the surface of cancer cells. GABs are made by the fusion of the ectodomains of a γδTCR to an anti-CD3 single chain variable fragment (scFv) (γδECTO-αCD3), here we explore alternative designs with the aim to enhance GAB effectivity., Methods: The first alternative design was made by linking the variable domains of the γ and δ chain to an anti-CD3 scFv (γδVAR-αCD3). The second alternative design was multimerizing γδVAR-αCD3 proteins to increase the tumor binding valency. Both designs were expressed and purified and the potency to target tumor cells by T cells of the alternative designs was compared to γδECTO-αCD3, in T cell activation and cytotoxicity assays., Results and Discussion: The γδVAR-αCD3 proteins were poorly expressed, and while the addition of stabilizing mutations based on finding for αβ single chain formats increased expression, generation of meaningful amounts of γδVAR-αCD3 protein was not possible. As an alternative strategy, we explored the natural properties of the original GAB design (γδECTO-αCD3), and observed the spontaneous formation of γδECTO-αCD3-monomers and -dimers during expression. We successfully enhanced the fraction of γδECTO-αCD3-dimers by shortening the linker length between the heavy and light chain in the anti-CD3 scFv, though this also decreased protein yield by 50%. Finally, we formally demonstrated with purified γδECTO-αCD3-dimers and -monomers, that γδECTO-αCD3-dimers are superior in function when compared to similar concentrations of monomers, and do not induce T cell activation without simultaneous tumor engagement. In conclusion, a γδECTO-αCD3-dimer based GAB design has great potential, though protein production needs to be further optimized before preclinical and clinical testing., Competing Interests: JK reports grants from Gadeta, Novartis, and Miltenyi Biotech and is the inventor on patents dealing with γδT cell-related aspects, as well as the co-founder and shareholder of Gadeta. EvD and DB are inventor on patents dealing with γδT cell-related aspects. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 van Diest, Nicolasen, Kramer, Zheng, Hernández-López, Beringer and Kuball.)

Ascaris suum is one of the most important parasites of pigs. Apart from liver condemnation due to lesions caused by migrating larvae ("milk spots"), A. suum infections can compromise weight gain, feed conversion efficacy, as well as meat quality. The true prevalence of infection depends on the diagnostic test used and is often underestimated. We compared liver inspection at slaughter with serology, based on the recognition of a purified A. suum haemoglobin or complete homogenate of the 3rd stage larvae isolated from lungs, in nine pig farms in northern Italy. Liver lesions were found on all farms with prevalence ranging from 3.8% to 98.3%. All farms were also positive for circulating antibodies against As-Hb and As-Lung-L3, with prevalence among pigs on each farm ranging from 36.4-100% and 54.5-100%, respectively. Seroprevalence was consistently higher when compared to the prevalence of milk spots at slaughter. The higher sensitivity of the ELISA tests combined with their ease of use makes them an interesting tool to evaluate A. suum infection levels., Competing Interests: Declaration of Competing Interest Authors declared no conflict of interests, (Copyright © 2022 Elsevier B.V. All rights reserved.)

Background: Lecanemab is a humanized IgG1 monoclonal antibody binding with high affinity to amyloid-beta protein protofibrils. In phase 3 development, lecanemab has been shown to reduce markers of amyloid in early Alzheimer's disease and reduce decline on clinical endpoints of cognition and function at 18 months., Objectives: To describe the health-related quality-of-life (HRQoL) results from Clarity AD which were exploratory outcomes in this trial., Design: Clarity AD was an 18-month, multi-center, double-blind, phase 3 trial., Setting: Early Alzheimer's disease., Participants: Individuals 50-90 years of age with a diagnosis of mild cognitive impairment or mild dementia due to Alzheimer's disease and positron emission tomography or cerebrospinal fluid evidence of cerebral amyloid accumulation., Intervention: Placebo or lecanemab 10-mg/kg IV biweekly., Measurements: HRQoL was measured at baseline and every 6 months using the European Quality of Life-5 Dimensions (EQ-5D-5L; by subject) and Quality of Life in AD (QOL-AD; by subject and proxy). Study partner burden was measured using the Zarit Burden Interview (ZBI)., Results: A total of 1795 participants were enrolled (lecanemab:898; placebo:897). At month 18, adjusted mean change from baseline in EQ-5D-5L and QOL-AD by subject showed 49% and 56% less decline, respectively. QOL-AD rated by study partner as proxy resulted in 23% less decline. ZBI adjusted mean change from baseline at 18 months resulted in 38% less increase of care partner burden. Individual HRQoL test items and dimensions also showed lecanemab benefit., Conclusions: Lecanemab was associated with a relative preservation of HRQoL and less increase in caregiver burden, with consistent benefits seen across different quality of life scales and within scale subdomains. These benefits provide valuable patient reported outcomes which, together with previously reported benefits of lecanemab across multiple measures of cognition, function, disease progression, and biomarkers, demonstrate that lecanemab treatment may offer meaningful benefits to patients, care partners, and society., Competing Interests: SC is a consultant (no personal fees received) to Alnylam, Biogen, Cassava Sciences, Cognivue, Cogstate, Eisai, Eli Lily, INmune Bio, ProMIS Neuroscience, Roche, and RetiSpec and receives research support (paid to institution) from AbbVie, AgeneBio, Alnylam, Anavex, Biogen, Cassava, Eisai, Eli Lilly, Genentech, Green Valley, Janssen, RetiSpec, Roche, and Vielight. CvD is a consultant for Roche, Eisai, Cerevel, and Ono and receives research support from Biogen, Eisai, Roche, Genentech, Eli Lilly, Janssen, UCB, Cerevel, and Biohaven. MG, TD, MK, SD, DL, SH, MI, AND LDK are employees of Eisai.

Introduction: The lack of ideal measurement of treatment efficacy is a well acknowledged problem in the epilepsy community, both in clinical care and clinical trials. Whilst still the current gold-standard, self-reported seizure frequency significantly underestimates the true number of seizures and does not account for any other at least equally important outcome parameters, such as neurodevelopment and cognition. With the rise of disease modifying treatments, the need for more reliable endpoints in practice and clinical trials becomes more pressing. In this paper we assembled an expert panel to discuss the nature of these needs, current limitations, and obstacles based on a survey amongst these experts who were queried about the most important issues regarding the use of electroencephalography (EEG) parameters as endpoints in clinical drug and device development., Methods: A structured survey was sent to a group of experts in the design and conduct of epilepsy trials in adults and children. This was followed by a virtual in-person meeting discussing the results of the trial and identifying a list of most important issues., Results: Six clinical trialists and 5 individuals from pharmaceutical companies returned the survey containing 14 questions, and 8 clinical trialists and 10 pharma-representatives attended the meeting. Three main issues were identified (1) lack of accuracy of seizure diaries due to nocturnal seizures, subtle motor seizures, impairment of consciousness and lack of awareness of the seizure by the patient (2) inter-rater variability of EEG assessment (3) lack of standardization regarding definition(s) of seizures (clinical and electrographic), EEG recording methods and EEG data management. Recommended solutions included (1) validation of EEG parameters as biomarkers and use of wearables (2) development of a manual that describes EEG rating criteria, protocol for validation by > 1 central reader and use of a resolution of disagreements reporting template (3) standardization of EEG recording, data management and reporting., Discussion & Conclusion: Current developments in research and technology seem promising to advance the use of EEG parameters as potential endpoints and offer partial solutions to the current needs. However, continuous, focused and collaborative efforts of all stakeholders (academia, industry and regulatory agencies) are needed to formulate guidelines, validate emerging technologies and approve them for use in trials. It is the intent of this opinion "position paper" to stimulate those efforts., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Objective: To evaluate the exploratory time to exceed pre-randomization seizure count (T-PSC) in the determination of efficacy of adjunctive perampanel in participants with primary generalized tonic-clonic (PGTC) seizures in generalized-onset epilepsy., Methods: In this multicenter, double-blind study (ClinicalTrials.gov identifier: NCT01393743), participants ≥12 years of age with treatment-resistant idiopathic generalized epilepsy were randomized to receive placebo or adjunctive perampanel (≤8 mg/day) across a 17-week double-blind treatment phase (4-week titration; 13-week maintenance). We evaluated the pre-planned exploratory end point of the T-PSC using a Kaplan-Meier analysis. We also re-evaluated the correspondence of the primary end points of median percent seizure frequency change (MPC) and 50% responder rate (50RR) calculated at T-PSC and at the end of the trial., Results: The exploratory end point of median T-PSC on placebo was 43 days and >120 days on perampanel (log-rank p < .001). The primary end points calculated at T-PSC did not differ significantly from the end points at the end of the trial (MPC -31% vs -42% at T-PSC; 50RR 32% vs 51% at T-PSC). After T-PSC was reached, participants had a median (interquartile range) of 5 (3-13) additional seizures on placebo and 5 (2-10) on perampanel., Significance: The exploratory end point of T-PSC demonstrated the effectiveness of perampanel despite a shorter duration of monitoring. The seizures that occurred after T-PSC did not influence the conclusions of the trial; therefore, T-PSC may be a viable alternative to traditional trial end points that reduces the risk to participants., (© 2022 The Authors. Epilepsia published by Wiley Periodicals LLC on behalf of International League Against Epilepsy.)

Protein tyrosine phosphatases (PTPs) are promising drug targets for treating a wide range of diseases such as diabetes, cancer, and neurological disorders, but their conserved active sites have complicated the design of selective therapeutics. This study examines the allosteric inhibition of PTP1B by amorphadiene (AD), a terpenoid hydrocarbon that is an unusually selective inhibitor. Molecular dynamics (MD) simulations carried out in this study suggest that AD can stably sample multiple neighboring sites on the allosterically influential C-terminus of the catalytic domain. Binding to these sites requires a disordered α7 helix, which stabilizes the PTP1B-AD complex and may contribute to the selectivity of AD for PTP1B over TCPTP. Intriguingly, the binding mode of AD differs from that of the most well-studied allosteric inhibitor of PTP1B. Indeed, biophysical measurements and MD simulations indicate that the two molecules can bind simultaneously. Upon binding, both inhibitors destabilize the α7 helix by disrupting interactions at the α3-α7 interface and prevent the formation of hydrogen bonds that facilitate closure of the catalytically essential WPD loop. These findings indicate that AD is a promising scaffold for building allosteric inhibitors of PTP1B and illustrate, more broadly, how unfunctionalized terpenoids can engage in specific interactions with protein surfaces.

Monoclonal antibodies (MAbs) targeting the Spike glycoprotein of SARS-CoV-2 is a key strategy to prevent severe COVID-19. Here, the efficacy of two monoclonal antibody bitherapies against SARS-CoV-2 was assessed on 92 patients at high risk of severe COVID-19 between March and October 2021 (Bichat-Claude Bernard Hospital, Paris, France). Nine patients died despite appropriate management. From 14 days following treatment initiation, we observed a slower viral load decay for patients treated with the bitherapy Bamlanivimab/Etsevimab compared to the Casirivimab/Imdevimab association therapy ( P  = 0.045). The emergence of several mutations on the Spike protein known to diminish antiviral efficacy was observed from 1 to 3 weeks after infusion. The Q493R mutation was frequently selected, located in a region of joint structural overlap by Bamlanivimab/Etsevimab antibodies. Despite that this study was done on former SARS-CoV-2 variants (Alpha and Delta), the results provide new insights into resistance mechanisms in SARS-CoV-2 antibodies neutralization escape and should be considered for current and novel variants. IMPORTANCE Monoclonal antibody bitherapies (MAbs) are commonly prescribed to treat severe SARS-CoV-2-positive patients, and the rapid growth of resistance mutation emergence is alarming globally. To explore this issue, we conducted both clinical and genomic analyses of SARS-CoV-2 in a series of patients treated in 2021. We first noticed that the two dual therapies prescribed during the study had different kinetics of viral load decay. Rapidly after initiation of the treatments, resistance mutations emerged in the interface between the MAbs and the target Spike glycoprotein, demonstrating the importance to continuously screen the viral genome during treatment course. Taken together, the results highlight that viral mutations may emerge under selective pressure, conferring a putative competitive advantage, and could rapidly spread, as observed for the Omicron variant.

Terpenoids, the largest and most structurally diverse group of natural products, include a striking variety of biologically active compounds, from flavors to medicines. Despite their well-documented biochemical versatility, the evolutionary processes that generate new functional terpenoids are poorly understood and difficult to recapitulate in engineered systems. This study uses a synthetic biochemical objective─a transcriptional system that links the inhibition of protein tyrosine phosphatase 1B (PTP1B), a human drug target, to the expression of a gene for antibiotic resistance in Escherichia coli ( E. coli )─to evolve a terpene synthase to produce enzyme inhibitors. Site saturation mutagenesis of poorly conserved residues on γ-humulene synthase (GHS), a promicuous enzyme, yielded mutants that improved fitness (i.e., the antibiotic resistance of E. coli ) by reducing GHS toxicity and/or by increasing inhibitor production. Intriguingly, a combination of two mutations enhanced the titer of a minority product─a terpene alcohol that inhibits PTP1B─by over 50-fold, and a comparison of similar mutants enabled the identification of a site where mutations permit efficient hydroxylation. Findings suggest that the plasticity of terpene synthases enables an efficient sampling of structurally distinct starting points for building new functional molecules and provide an experimental framework for exploiting this plasticity in activity-guided screens.

The Hawaiian Archipelago experienced a moderate bleaching event in 2019-the third major bleaching event over a 6-year period to impact the islands. In response, the Hawai'i Coral Bleaching Collaborative (HCBC) conducted 2,177 coral bleaching surveys across the Hawaiian Archipelago. The HCBC was established to coordinate bleaching monitoring efforts across the state between academic institutions, non-governmental organizations, and governmental agencies to facilitate data sharing and provide management recommendations. In 2019, the goals of this unique partnership were to: 1) assess the spatial and temporal patterns of thermal stress; 2) examine taxa-level patterns in bleaching susceptibility; 3) quantify spatial variation in bleaching extent; 4) compare 2019 patterns to those of prior bleaching events; 5) identify predictors of bleaching in 2019; and 6) explore site-specific management strategies to mitigate future bleaching events. Both acute thermal stress and bleaching in 2019 were less severe overall compared to the last major marine heatwave events in 2014 and 2015. Bleaching observed was highly site- and taxon-specific, driven by the susceptibility of remaining coral assemblages whose structure was likely shaped by previous bleaching and subsequent mortality. A suite of environmental and anthropogenic predictors was significantly correlated with observed bleaching in 2019. Acute environmental stressors, such as temperature and surface light, were equally important as previous conditions (e.g. historical thermal stress and historical bleaching) in accounting for variation in bleaching during the 2019 event. We found little evidence for acclimation by reefs to thermal stress in the main Hawaiian Islands. Moreover, our findings illustrate how detrimental effects of local anthropogenic stressors, such as tourism and urban run-off, may be exacerbated under high thermal stress. In light of the forecasted increase in severity and frequency of bleaching events, future mitigation of both local and global stressors is a high priority for the future of corals in Hawai'i., Competing Interests: The authors have declared that no competing interests exist.

The anomalous past two years of the COVID-19 pandemic have been a test of human response to global crisis management as typical human activities were significantly altered. The COVID-instigated anthropause has illustrated the influence that humans and the biosphere have on each other, especially given the variety of national mobility interventions that have been implemented globally. These local COVID-19-era restrictions influenced human-ecosystem interactions through changes in accessibility of water systems and changes in ecosystem service demand. Four urban aquatic case studies in the Netherlands demonstrated shifts in human demand during the anthropause. For instance, reduced boat traffic in Amsterdam canals led to improved water clarity. In comparison, ongoing service exploitation from increased recreational fishing, use of bathing waters and national parks visitation are heightening concerns about potential ecosystem degradation. We distilled management lessons from both the case studies as well as from recent literature pertaining to ecological intactness and social relevance. Equally important to the lessons themselves, however, is the pace at which informed management practices are established after the pandemic ends, particularly as many communities currently recognize the importance of aquatic ecosystems and are amenable to their protection., Competing Interests: Declaration of Competing Interest The authors declare that there is no conflict of personal, professional or financial interest., (Copyright © 2022 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Dirofilaria immitis is a parasitic nematode and causes dirofilariosis, a potentially fatal pulmonary infection which primarily infects canids. Dirofilariosis infections are controlled via prophylactic macrocyclic lactone (ML) regimens. Recent evidence has confirmed the development of ML-resistant isolates in the USA, which are genetically distinct from wildtype populations. Single nucleotide polymorphisms (SNP) associated with ML-resistant phenotypes were clinically validated in USA populations. In this study, 3 USA laboratory-maintained isolates (Berkeley, Georgia II, and WildCat) and 11 randomly selected European clinical samples from 7 hosts were analyzed. The samples tested were fresh microfilaria (mf) in blood or adult worms preserved in ethanol. The samples underwent MiSeq sequencing of the top 9 SNP associated with ML resistance. The results provide the first genotypic analysis of the three USA laboratory-maintained isolates and any European samples. The European clinical samples show no genomic evidence of ML resistance based on the 9 SNP. The early adoption of genotyping of clinical D. immitis samples could provide an early indication of the potential development of ML resistance and aid to distinguish clinical cases of heartworm infection due to ML resistance from those due to a lack compliance with the recommended treatments, as has been seen in North America.