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Cancer progression and response to therapy are inextricably reliant on the co-evolution of a supportive tissue microenvironment. This is particularly evident in pancreatic ductal adenocarcinoma (PDAC), a tumor type characterized by expansive and heterogeneous stroma. Herein, we employed single cell RNAseq and spatial transcriptomics of normal, inflamed, and malignant pancreatic tissues to contextualize stromal dynamics associated with disease and treatment status, identifying temporal and spatial trajectories of fibroblast differentiation. Using analytical tools to infer cellular communication, together with a newly developed assay to annotate genomic alterations in cancer cells, we additionally explored the complex intercellular networks underlying tissue circuitry, highlighting a fibroblast-centric interactome that grows in strength and complexity in the context of malignant transformation. Our study yields new insights on the stromal remodeling events favoring the development of a tumor-supportive microenvironment and provides a powerful resource for the exploration of novel points of therapeutic intervention in PDAC.

Early detection of disseminating vancomycin-resistant Enterococcus faecium (VREfm) in ICU wards is crucial for outbreak identification and the implementation of prompt infection control measures. Genotypic methods like pulsed-field gel electrophoresis (PFGE) and whole-genome sequencing (WGS) are costly and time-consuming, hindering rapid response due to batch dependency. Fourier-transform infrared spectroscopy (FT-IR) offers the potential for real-time outbreak detection and reliable strain typing. We utilized FT-IR to identify clonal VREfm dissemination and compared its performance to PFGE and WGS. Between February through October 2023, an unusually high number of VREfm were recovered at a tertiary hospital in Barcelona. Isolates were examined for antimicrobial susceptibility, carriage of vanA/vanB genes and clonality was also studied using FT-IR, PFGE, and WGS. Routine FT-IR inspections revealed recurring VREfm clustering during the outbreak's initial weeks. In total, 104 isolates were recovered from 75 patients and from multiple wards. However, only one isolate was recovered from an environmental sample, suggesting the absence of environmental reservoirs. An ST80 vancomycin-resistant ( vanA ) E. faecium strain was the main strain responsible for the outbreak, although a few additional VREfm strains were also identified, all belonging to CC17. PFGE and cgMLST (WGS) yielded identical clustering results to FT-IR, and WGS confirmed vanA/vanB gene carriage in all VREfm isolates. Infection control measures led to a rapid decline in VREfm isolates, with no isolates detected in November. FT-IR spectroscopy offers rapid turnaround times, sensitivity, and reproducibility, comparable to standard typing methods. It proved as an effective tool for monitoring VREfm dissemination and early outbreak detection.

Tolerance of mouse kidney allografts arises in grafts that develop regulatory tertiary lymphoid organs (rTLOs). Single-cell RNA-seq (scRNA-seq) data and adoptive transfer of alloreactive T cells after transplantation showed that cytotoxic CD8+ T cells are reprogrammed within the accepted graft to an exhausted/regulatory-like phenotype mediated by IFN-γ. Establishment of rTLOs was required because adoptive transfer of alloreactive T cells prior to transplantation results in kidney allograft rejection. Despite the presence of intragraft CD8+ cells with a regulatory phenotype, they were not essential for the induction and maintenance of kidney allograft tolerance since renal allotransplantation into CD8-KO recipients resulted in acceptance and not rejection. Analysis of scRNA-seq data from allograft kidneys and malignant tumors identified similar regulatory-like cell types within the T cell clusters and trajectory analysis showed that cytotoxic CD8+ T cells are reprogrammed into an exhausted/regulatory-like phenotype intratumorally. Induction of cytotoxic CD8+ T cell dysfunction of infiltrating cells appears to be a beneficial mechanistic pathway that protects the kidney allotransplant from rejection through a process we call "defensive tolerance." This pathway has implications for our understanding of allotransplant tolerance and tumor resistance to host immunity.

Acinetobacter spp. are often isolated from natural sources, but knowledge about their presence in wild animals is fragmented and uncomplete. The present study aimed to characterize a series of Acinetobacter radioresistens isolated from Humboldt penguins (Spheniscus humboldti). Fifteen Humboldt penguins from an inhabited northern Peruvian island were sampled. Microorganisms were identified by MALDI-TOF MS. Antibiotic susceptibility to 12 antimicrobial agents was established, and clonal relationships were determined. A representative isolate was selected for whole genome sequencing (WGS). A. radioresistens were isolated from the feces of 12 (80%) Humboldt penguins, being susceptible to all the antimicrobial agents tested, except eight cefotaxime-intermediate isolates. All A. radioresistens were clonally related. WGS showed that the isolate belonged to ST1972, the presence of two chromosomal encoded carbapenemases (blaOXA-23 and a putative subclass B3 metallo-β-lactamase), and a series of point mutations in antibiotic-resistance related chromosomal genes, which were considered as polymorphisms. In addition, a few virulence factors, including a capsule-encoding operon, superoxide dismutases, catalases, phospholipases and a siderophore receptor were identified. The present results suggest that A. radioresistens may be a common member of the gut microbiota of Humboldt penguins, but further studies in other geographical areas are needed to establish this finding.

For a majority of patients with non-small cell lung cancer with EGFR mutations, treatment with EGFR inhibitors (EGFRi) induces a clinical response. Despite this initial reduction in tumor size, residual disease persists that leads to disease relapse. Elucidating the preexisting biological differences between sensitive cells and surviving drug-tolerant persister cells and deciphering how drug-tolerant cells evolve in response to treatment could help identify strategies to improve the efficacy of EGFRi. In this study, we tracked the origins and clonal evolution of drug-tolerant cells at a high resolution by using an expressed barcoding system coupled with single-cell RNA sequencing. This platform enabled longitudinal profiling of gene expression and drug sensitivity in response to EGFRi across a large number of clones. Drug-tolerant cells had higher expression of key survival pathways such as YAP and EMT at baseline and could also differentially adapt their gene expression following EGFRi treatment compared with sensitive cells. In addition, drug combinations targeting common downstream components (MAPK) or orthogonal factors (chemotherapy) showed greater efficacy than EGFRi alone, which is attributable to broader targeting of the heterogeneous EGFRi-tolerance mechanisms present in tumors. Overall, this approach facilitates thorough examination of clonal evolution in response to therapy that could inform the development of improved diagnostic approaches and treatment strategies for targeting drug-tolerant cells., Significance: The evolution and heterogeneity of EGFR inhibitor tolerance are identified in a large number of clones at enhanced cellular and temporal resolution using an expressed barcode technology coupled with single-cell RNA sequencing., (©2023 American Association for Cancer Research.)

Introduction: There are no data on community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) infections in the context of the chemsex phenomenon. This study aimed to characterize CA-MRSA-related infections in a cohort of people living with HIV (PLWH) who engage in chemsex., Methods: At the Hospital Clinic of Barcelona, from February 2018 to January 2022, we analyzed CA-MRSA infections diagnosed in a cohort of PLWH who engage in chemsex. Epidemiological, behavioral and clinical variables were assessed. Mass spectrometry identification and antimicrobial susceptibility testing were performed on MRSA isolates. Pulse field electrophoresis was used to assess the clonality of the MRSA strains. The presence of Panton-Valentine leukocidin was also investigated., Results: Among the cohort of 299 participants who engage in chemsex, 25 (8%) with CA-MRSA infections were identified, 9 at baseline and 16 with incident cases; the cumulative incidence was 5.5% (95% CI: 3.2%, 8.8%). The most common drugs were methamphetamine (96%) and GHB/GBL (92%). Poly-consumption and slamming were reported by 32% and 46%, respectively. CA-MRSA was isolated from the infection sites of 20 participants, and CA-MRSA colonization was confirmed in the remaining 5 persons. Seventy-one percent had used antibiotics in the previous year. All participants presented with skin and soft tissue infections, 28% required hospitalization, and 48% had recurrence. Of the 23 MRSA isolates further studied, 19 (82,6%) belonged to the same clone. Panton-Valentine leukocidin was detected in all isolates., Conclusion: PLWH who engage in chemsex may present with CA-MRSA infections. Clinical suspicion and microbiological diagnosis are required to provide adequate therapy, and CA-MRSA prevention interventions should be designed., (© 2023. The Author(s).)

Introduction: Obsessive-compulsive and related disorders (OCRDs) and disorders due to addictive behaviours (DABs) are prevalent conditions that share behavioural and neurobiological characteristics. The Research Domain Criteria lists a series of constructs whose dysfunctions may be present in both groups of disorders. The present study will describe the research protocol of a scoping review of the literature on self-report scales and questionnaires that tap dysfunctional constructs that underlie OCRDs and DABs., Methods and Analysis: This protocol outlines a scoping review on self-report tools and questionnaires that assess OCRDs and DABs-related constructs. The scoping review will select sources in MEDLINE, EMBASE, PsychINFO and Web of Science databases. Inclusion and exclusion criteria will be designed according to the Population, Concept, Context, Types of source framework. Two reviewers will screen independently titles, abstracts and full texts to determine the eligibility of articles. A methodological framework including six stages steps ((1) identifying a research question; (2) identifying relevant studies; (3) study selection; (4) charting the data; (5) collating, summarising and reporting the result) will be used, and the findings will be reported according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses extension for Scoping Reviews checklist. Information extracted will be collated, and quantitative results will be presented using descriptive statistics such as percentages, tables, charts and flow diagrams as appropriate., Ethics and Dissemination: Ethical approval for conducting this scoping review is not required, as this study will involve secondary analysis of existing literature. The researchers will disseminate the study results via conference presentations and publication in a peer-reviewed journal., Scoping Review Protocol Registration: DOI 10.17605/OSF.IO/UJ7G5., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2022. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

CD3-engaging bispecific antibodies (BsAbs) enable the formation of an immune synapse between T cells and tumor cells, resulting in robust target cell killing not dependent on a preexisting tumor specific T cell receptor. While recent studies have shed light on tumor cell-specific factors that modulate BsAb sensitivity, the T cell-intrinsic determinants of BsAb efficacy and response durability are poorly understood. To better clarify the genes that shape BsAb-induced T cell responses, we conducted targeted analyses and a large-scale unbiased in vitro CRISPR/Cas9-based screen to identify negative regulators of BsAb-induced T cell proliferation. These analyses revealed that CD8+ T cells are dependent on CD4+ T cell-derived signaling factors in order to achieve sustained killing in vitro . Moreover, the mammalian target of rapamycin (mTOR) pathway and several other candidate genes were identified as intrinsic regulators of BsAb-induced T cell proliferation and/or activation, highlighting promising approaches to enhancing the utility of these potent therapeutics., Competing Interests: Authors RM, TF, GT, EC, DR, MP, JL, HW, and HL are currently employed by the Novartis Institutes for Biomedical Research (NIBR). The following authors were previously employed by NIBR when they were involved with the work described herein: MV (currently employed by Arbor Biotechnologies), GE (currently employed by Foghorn Therapeutics), MM (currently employed by Bristol Myers Squibb), and SK (currently employed by 2seventy Bio)., (Copyright © 2022 Molony, Funk, Trabucco, Corcoran, Ruddy, Varadarajan, Elliot, Piquet, Lam, Meyer, Wang, Kurtulus and Lu.)

Asparagine deprivation by l-asparaginase (L-ASNase) is an effective therapeutic strategy in acute lymphoblastic leukemia, with resistance occurring due to upregulation of ASNS, the only human enzyme synthetizing asparagine ( Annu. Rev. Biochem. 2006 , 75 (1), 629-654). l-Asparaginase efficacy in solid tumors is limited by dose-related toxicities ( OncoTargets and Therapy 2017, pp 1413-1422). Large-scale loss of function genetic in vitro screens identified ASNS as a cancer dependency in several solid malignancies ( Cell 2017 , 170 (3), 564-576.e16. Cell 2017 , 170 (3), 577-592.e10). Here we evaluate the therapeutic potential of targeting ASNS in melanoma cells. While we confirm in vitro dependency on ASNS silencing, this is largely dispensable for in vivo tumor growth, even in the face of asparagine deprivation, prompting us to characterize such a resistance mechanism to devise novel therapeutic strategies. Using ex vivo quantitative proteome and transcriptome profiling, we characterize the compensatory mechanism elicited by ASNS knockout melanoma cells allowing their survival. Mechanistically, a genome-wide CRISPR screen revealed that such a resistance mechanism is elicited by a dual axis: GCN2-ATF4 aimed at restoring amino acid levels and MAPK-BCLXL to promote survival. Importantly, pharmacological inhibition of such nodes synergizes with l-asparaginase-mediated asparagine deprivation in ASNS deficient cells suggesting novel potential therapeutic combinations in melanoma., Competing Interests: The authors declare the following competing financial interest(s): Authors affiliated to Novartis Institute for Biomedical Research are employees of Novartis Pharma AG., (© 2021 American Chemical Society.)

Despite the increasing interest in targeting stromal elements of the tumor microenvironment, we still face tremendous challenges in developing adequate therapeutics to modify the tumor stromal landscape. A major obstacle to this is our poor understanding of the phenotypic and functional heterogeneity of stromal cells in tumors. Herein, we perform an unbiased interrogation of tumor mesenchymal cells, delineating the co-existence of distinct subsets of cancer-associated fibroblasts (CAFs) in the microenvironment of murine carcinomas, each endowed with unique phenotypic features and functions. Furthermore, our study shows that neutralization of TGFβ in vivo leads to remodeling of CAF dynamics, greatly reducing the frequency and activity of the myofibroblast subset, while promoting the formation of a fibroblast population characterized by strong response to interferon and heightened immunomodulatory properties. These changes correlate with the development of productive anti-tumor immunity and greater efficacy of PD1 immunotherapy. Along with providing the scientific rationale for the evaluation of TGFβ and PD1 co-blockade in the clinical setting, this study also supports the concept of plasticity of the stromal cell landscape in tumors, laying the foundation for future investigations aimed at defining pathways and molecules to program CAF composition for cancer therapy.

Pearl millet ( Pennisetum glaucum (L.) R. Br.) is a staple food and a drought-tolerant cereal well adapted to Sub-Saharan Africa agro-ecosystems. An important diversity of pearl millet landraces has been widely conserved by farmers and therefore could help copping with climate changes and contribute to future food security. Hence, characterizing its genetic diversity and population structure can contribute to better assist breeding programs for a sustainable agricultural productivity enhancement. Toward this goal, a comprehensive panel of 404 accessions were used that correspond to 12 improved varieties, 306 early flowering and 86 late-flowering cultivated landraces from Senegal. Twelve highly polymorphic SSR markers were used to study diversity and population structure. Two genes, PgMADS11 and PgPHYC , were genotyped to assess their association to flowering phenotypic difference in landraces. Results indicate a large diversity and untapped potential of Senegalese pearl millet germplasm as well as a genetic differentiation between early- and late-flowering landraces. Further, a fine-scale genetic difference of PgPHYC and PgMADS11 (SNP and indel, respectively) and co-variation of their alleles with flowering time were found among landraces. These findings highlight new genetic insights of pearl millet useful to define heterotic populations for breeding, genomic association panel, or crosses for trait-specific mapping.

Research is expected to produce knowledge, methods and tools to enhance stakeholders' adaptive capacity by helping them to anticipate and cope with the effects of climate change at their own level. Farmers face substantial challenges from climate change, from changes in the average temperatures and the precipitation regime to an increased variability of weather conditions and the frequency of extreme events. Such changes can have dramatic consequences for many types of agricultural production systems such as grassland-based livestock systems for which climate change influences the seasonality and productivity of fodder production. We present a participatory design method called FARMORE (FARM-Oriented REdesign) that allows farmers to design and evaluate adaptations of livestock systems to future climatic conditions. It explicitly considers three climate features in the design and evaluation processes: climate change, climate variability and the limited predictability of weather. FARMORE consists of a sequence of three workshops for which a pre-existing game-like platform was adapted. Various year-round forage production and animal feeding requirements must be assembled by participants with a computerized support system. In workshop 1, farmers aim to produce a configuration that satisfies an average future weather scenario. They refine or revise the previous configuration by considering a sample of the between-year variability of weather in workshop 2. In workshop 3, they explicitly take the limited predictability of weather into account. We present the practical aspects of the method based on four case studies involving twelve farmers from Aveyron (France), and illustrate it through an in-depth description of one of these case studies with three dairy farmers. The case studies shows and discusses how workshop sequencing (1) supports a design process that progressively accommodates complexity of real management contexts by enlarging considerations of climate change to climate variability and low weather predictability, and (2) increases the credibility and salience of the design method. Further enhancements of the method are outlined, especially the selection of pertinent weather scenarios., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Rangeland-based livestock systems have to deal with the significant instability and uncertainty of the agricultural context (policy changes, volatility of input prices, etc.), and especially of the climatic context. Thus, they are particularly concerned by adaptive management strategies. To support the development of such strategies, we developed a board game including a computer model called "Rangeland Rummy". It is to be used by groups of farmers and agricultural consultants in the context of short workshops (about 3 h). Rangeland Rummy builds upon five types of material object: (i) a game board; (ii) a calendar stick indicating the starting date of the game board; (iii) sticks marked with the feed resources available for combinations of vegetation types and their management practices; (iv) cards to define animal groups and their feeding requirements throughout the year; (v) cards related to types of feed that can be attributed to animal groups throughout the year. Using these material objects, farmers collectively design a rangeland-based livestock system. This system is immediately evaluated using a computer model, i.e. a spreadsheet providing graphs and indicators providing information on, among other things, the extent to which quantitative and qualitative animal feeding requirements are covered across the year. Playing the game thus consists in collectively and iteratively designing and evaluating rangeland-based livestock systems, while confronting the players with new contextual challenges (e.g. interannual variability of weather, volatility of input prices) or new farmers' objectives (e.g. being self-sufficient for animal feeding). An example of application of Rangeland Rummy with 3 farmers in southern France is reported. Applications show that it tends to develop farmers' adaptive capacity by stimulating their discussions and the exchange of locally-relevant knowledge on management strategies and practices in rangeland-based livestock systems., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Background: Transcutaneous neurostimulation (TNS) at extracephalic sites is a well known treatment of pain. Thanks to recent technical progress, the Cefaly® device now also allows supraorbital TNS. During observational clinical studies, several patients reported decreased vigilance or even sleepiness during a session of supraorbital TNS. We decided therefore to explore in more detail the potential sedative effect of supraorbital TNS, using standardized psychophysical tests in healthy volunteers., Methods: We performed a double-blind cross-over sham-controlled study on 30 healthy subjects. They underwent a series of 4 vigilance tests (Psychomotor Vigilance Task, Critical Flicker Fusion Frequency, Fatigue Visual Numeric Scale, d2 test). Each subject was tested under 4 different experimental conditions: without the neurostimulation device, with sham supraorbital TNS, with low frequency supraorbital TNS and with high frequency supraorbital TNS., Results: As judged by the results of three tests (Psychomotor Vigilance Task, Critical Flicker Fusion Frequency, Fatigue Visual Numeric Scale) there was a statistically significant (p < 0.001) decrease in vigilance and attention during high frequency TNS, while there were no changes during the other experimental conditions. Similarly, performance on the d2 test was impaired during high frequency TNS, but this change was not statistically significant., Conclusion: Supraorbital high frequency TNS applied with the Cefaly® device decreases vigilance in healthy volunteers. Additional studies are needed to determine the duration of this effect, the underlying mechanisms and the possible relation with the stimulation parameters. Meanwhile, this effect opens interesting perspectives for the treatment of hyperarousal states and, possibly, insomnia.