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Emerging and re-emerging viruses are a global health concern. Genome sequencing as an approach for monitoring circulating viruses is currently hampered by complex and expensive methods. Untargeted, metagenomic nanopore sequencing can provide genomic information to identify pathogens, prepare for or even prevent outbreaks. SMART (Switching Mechanism at the 5′ end of RNA Template) is a popular approach for RNA-Seq but most current methods rely on oligo-dT priming to target polyadenylated mRNA molecules. We have developed two random primed SMART-Seq approaches, a sequencing agnostic approach ‘SMART-9N’ and a version compatible rapid adapters available from Oxford Nanopore Technologies ‘Rapid SMART-9N’. The methods were developed using viral isolates, clinical samples, and compared to a gold-standard amplicon-based method. From a Zika virus isolate the SMART-9N approach recovered 10kb of the 10.8kb RNA genome in a single nanopore read. We also obtained full genome coverage at a high depth coverage using the Rapid SMART-9N, which takes only 10 minutes and costs up to 45% less than other methods. We found the limits of detection of these methods to be 6 focus forming units (FFU)/mL with 99.02% and 87.58% genome coverage for SMART-9N and Rapid SMART-9N respectively. Yellow fever virus plasma samples and SARS-CoV-2 nasopharyngeal samples previously confirmed by RT-qPCR with a broad range of Ct-values were selected for validation. Both methods produced greater genome coverage when compared to the multiplex PCR approach and we obtained the longest single read of this study (18.5 kb) with a SARS-CoV-2 clinical sample, 60% of the virus genome using the Rapid SMART-9N method. This work demonstrates that SMART-9N and Rapid SMART-9N are sensitive, low input, and long-read compatible alternatives for RNA virus detection and genome sequencing and Rapid SMART-9N improves the cost, time, and complexity of laboratory work.

Abstract Blood‐brain‐barrier (BBB) disruption has been associated with a variety of central‐nervous‐system diseases. In vitro BBB models enable to investigate how the barrier reacts to external injury events, commonly referred to as insults. Here, a human‐cell‐based BBB platform with integrated, transparent electrodes to monitor barrier tightness in real time at high resolution is presented. The BBB model includes human cerebral endothelial cells and primary pericytes and astrocytes in a 3D arrangement within a pump‐free, open‐microfluidic platform. With this platform, this study demonstrates that oxygen‐glucose deprivation (OGD), which mimics the characteristics of an ischemic insult, induces a rapid remodeling of the cellular actin structures and subsequent morphological changes in the endothelial cells. High‐resolution live imaging shows the formation of large actin stress‐fiber bundles in the endothelial layer during OGD application, which ultimately leads to cell shrinkage and barrier breakage. Simultaneous electrical measurements evidence a rapid decrease of the barrier electrical resistance before the appearance of stress fibers, which indicates that the barrier function is compromised already before the appearance of drastic morphological changes. The results demonstrate that the BBB platform recapitulates the main barrier functions in vitro and can be used to investigate rapid reorganization of the BBB upon application of external stimuli.

Introduction: The adipose tissue secretes adipokines and influences the release of inflammatory mediators contributing to a state of low-grade systemic inflammation that may change lung function. Objective: To correlate levels of adipokines and inflammatory mediators with lung function in individuals with obesity and bronchial asthma symptoms. Materials and methods: A cross-sectional study, including women with obesity (grade II and III) with symptoms and clinical diagnosis of asthma. Anthropometric measurements (weight, height, BMI), pulmonary function test (spirometry), asthma control test questionnaire, collection of systemic inflammatory markers (blood collection) and pulmonary markers (sputum collection) were collected and were analyzed: IL-6, IL-8, TNF-α, adiponectin, resistin, leptin and C-reactive protein (CRP). The patients were stratified into two groups according to asthma control. Results: 80 women were analyzed and 40% had an ACT score greater than or equal to 18 and were classified as “controlled asthma”. More than half of the patients of ACT

Background: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is linked to high mortality, primarily through an intense inflammatory response. Diacerein has emerged as a potential therapy for COVID-19 due to its potential impact in decreasing the inflammasome activation and coronavirus replication. This study aims to explore diacerein's influence in inhibiting both viral replication and the inflammatory response after SARS-CoV-2 infection. Methods: Human peripheral blood mononuclear cells (PBMCs) were obtained from healthy volunteers and infected in vitro with SARS-CoV-2. Additionally, we carried out a pilot randomized, double-blind, placebo-controlled study with 14 participants allocated to diacerein (n = 7) or placebo (n = 7) therapies every 12 h for 10 days. The primary endpoint was change in plasma markers of inflammasome activation (NLRP3, caspase-1, and gasdermin-D). Results: In vitro protocols have shown that rhein, diacerein's primary metabolite, decreased IL-1ß secretion caused by SARS-CoV-2 infection in human PBMCs (p < 0.05), and suppressed viral replication when administered either before or after the virus incubation (p < 0.05). This later effect was, at least partially, attributed to its inhibitory effect on 3-chymotrypsin-like protease (SARS-CoV-2 3CLpro) and papain-like protease in the SARS-CoV-2 (SARS-CoV-2 PLpro) virus and in the phosphorylation of proteins related cytoskeleton network (p < 0.05). Diacereintreated COVID-19 patients presented a smaller area under the curve for NLRP3, caspase-1 and GSDM-D measured on days 2, 5, and 10 after hospitalization compared to those receiving a placebo (p < 0.05). Conclusion: The indicated mechanisms of action of diacerein/rhein can reduce viral replication and mitigate the inflammatory response related to SARS-CoV-2. These findings are preliminary and require confirmation in clinical trials. [ABSTRACT FROM AUTHOR]

In this work, naphthalenediimide (NDI) derivatives are widely studied for their semiconducting properties and the influence of the side-chain length on the crystal packing is reported, along with the thermal properties of three core-chlorinated NDIs with different fluoroalkyl side-chain lengths (CF3-NDI, C3F7-NDI and C4F9-NDI). The introduction of fluorinated substituents at the imide nitrogen and addition of strong electron-withdrawing groups at the NDI core are used to improve the NDI derivatives air stability. The new compound, CF3-NDI, was deeply analyzed and compared to the well-known C3F7-NDI and C4F9-NDI, leading to the discovery and solution of two different crystal phases, form α and solvate form, and a solid solution of CF3-NDI and CF3-NDI-OH, formed by the decomposition in DMSO. [ABSTRACT FROM AUTHOR]

COVID-19-associated invasive pulmonary aspergillosis (CAPA) is common and is associated with poor outcomes in critically ill patients. This prospective observational study aimed to explore the association between CAPA development and the incidence and prognosis of cytomegalovirus (CMV) reactivation in critically ill COVID-19 patients. We included all consecutive critically ill adult patients with confirmed COVID-19 infection who were admitted to three COVID-19 intensive care units (ICUs) in an Italian hospital from 25 February 2020 to 8 May 2022. A standardized procedure was employed for early detection of CAPA. Risk factors associated with CAPA and CMV reactivation and the association between CMV recurrence and mortality were estimated using adjusted Cox proportional hazard regression models. CAPA occurred in 96 patients (16.6%) of the 579 patients analyzed. Among the CAPA population, 40 (41.7%) patients developed CMV blood reactivation with a median time of 18 days (IQR 7–27). The CAPA+CMV group did not exhibit a significantly higher 90-day mortality rate (62.5% vs. 48.2%) than the CAPA alone group (p = 0.166). The CAPA+CMV group had a longer ICU stay, fewer ventilation-free days, and a higher rate of secondary bacterial infections than the control group of CAPA alone. In the CAPA population, prior immunosuppression was the only independent risk factor for CMV reactivation (HR 2.33, 95% C.I. 1.21–4.48, p = 0.011). In critically ill COVID-19 patients, CMV reactivation is common in those with a previous CAPA diagnosis. Basal immunosuppression before COVID-19 appeared to be the primary independent variable affecting CMV reactivation in patients with CAPA. Furthermore, the association of CAPA+CMV versus CAPA alone appears to impact ICU length of stay and secondary bacterial infections but not mortality.

Stefania Lucamante. Righteous Anger in Contemporary Literary and Cinematic Narratives. Toronto: University of Toronto Press, 2020. Righteous Anger in Contemporary Italian Literary and Cinematic Narratives presents an original and incisive [...]

Abstract As 3D in vitro tissue models become more pervasive, their built-in nutrient, metabolite, compound, and waste gradients increase biological relevance at the cost of analysis simplicity. Investigating these gradients and the resulting metabolic heterogeneity requires invasive and time-consuming methods. An alternative is using electrochemical biosensors and measuring concentrations around the tissue model to obtain size-dependent metabolism data. With our hanging-drop-integrated enzymatic glucose biosensors, we conducted current measurements within hanging-drop compartments hosting spheroids formed from the human colorectal carcinoma cell line HCT116. We developed a physics-based mathematical model of analyte consumption and transport, considering (1) diffusion and enzymatic conversion of glucose to form hydrogen peroxide (H2O2) by the glucose-oxidase-based hydrogel functionalization of our biosensors at the microscale; (2) H2O2 oxidation at the electrode surface, leading to amperometric H2O2 readout; (3) glucose diffusion and glucose consumption by cancer cells in a spherical tissue model at the microscale; (4) glucose and H2O2 transport in our hanging-drop compartments at the macroscale; and (5) solvent evaporation, leading to glucose and H2O2 upconcentration. Our model relates the measured currents to the glucose concentrations generating the currents. The low limit of detection of our biosensors (0.4 ± 0.1 μM), combined with our current-fitting method, enabled us to reveal glucose dynamics within our system. By measuring glucose dynamics in hanging-drop compartments populated by cancer spheroids of various sizes, we could infer glucose distributions within the spheroid, which will help translate in vitro 3D tissue model results to in vivo.

This article presents a multiplexed current sensitive readout for label-free zeptomolar-sensitive detectors realized with large-area electrolyte-gated organic thin-film transistors (EGOFETs). These highly capacitive biosensors are multiplexed using an organic thin-film transistor (OTFT) line driver and OTFT switches and interfaced to a 65-nm Si CMOS, low-power, pA-sensitive front-end. The Si chip performs analog-to-digital conversion and data transmission to a microcontroller too. A current domain interface is used to transmit the signals coming from multiple biosensors to the 1.2-V supply CMOS Si-IC via the 30-V supply OTFT electronics. Exploiting an analog module implemented in the Si-IC, the EGOFETs are precisely biased, even in the presence of a large OTFT multiplexer resistance. The CMOS current sensitive front-end achieves a dynamic range (DR) of 137 dB and a power consumption of 42- $\mu \text{W}$ per channel reaching a state-of-the-art DR-power-bandwidth FOM of 208 dB. The front-end has been designed with a first-stage programmable-gain, active-feedback transimpedance amplifier topology that, contrary to common current-sensitive front-end solutions, is not affected by the sensor capacitance. The system has been validated with different concentrations of human IgG and IgM proteins using both a single sensor and a 4 $\times $ 4 array of EGOFETs. Thanks to the multiplexing strategy and the low costs of its modules, the system here presented has the potential to enable widespread use of precision diagnostic with extreme sensitivity even in point-of-care and low-resource settings.

Abstract Electrolyte‐gated organic transistors (EGOTs) are promising and versatile devices for next‐generation biosensors, neuromorphic systems, and low‐voltage electronics. They are particularly indicated for applications where stable operation in aqueous environment and cost‐effective manufacturing are required. Indeed, EGOTs can be fabricated through low‐cost, large area, and scalable techniques, such as printing, from a large portfolio of solution processable organic materials, which are often able to stably operate in water or physiological solutions. Despite a large number of solution processable EGOTs have been reported in the literature so far, only a few are based on printed semiconductors, with no examples of digitally printed, i.e., inkjet printed, n‐type devices, which would easily enable complementary architectures. In this work, we propose the first example of a n‐type electrolyte gated organic transistor based on an inkjet printed polymer. The proposed device shows a high stability when operated in water and requires only 3 hours of conditioning to produce a stable response, a much faster dynamic than in the case of printed polymers currently tested for p‐type EGOTs. As a proof‐of‐concept, the proposed printed n‐type EGOT is successfully integrated with a printed single‐walled carbon‐nanotubes based p‐type device in a logic inverter, demonstrating the possibility to build simple water‐gated digital electronic circuits.

This study aimed to evaluate youth-to-senior transition and the relative age effect in Italian female football national teams. Birthdate data of 774 female players selected for Under 17 (N = 416), 19 (N = 265) and National Senior team (N = 93) were analysed. The youth-to-senior transition rate was determined by the number of youth players competing in the Senior National team (and vice versa), whilst birth quarter (Q) distributions with a chi-square goodness-of-fit test. Only 17.4% of youth players were able to be selected for the Senior National team, whereas 31.2% of the players reached the high-senior level without being selected for youth age groups. Data revealed a skewed birth date distribution in Under 17 and 19 teams (on average, Q1 = 35.6% vs Q4 = 18.5%) but not in the Senior National team. Youth players born in Q1 were two times more likely to be selected than in Q4. In Under 17, goalkeepers, defenders, and midfielders of Q1 players were overrepresented. However, Q4 players recorded higher conversion rates than Q1 (Q1 = 16.4% vs. Q4 = 25.0%). National youth experience may not be a prerequisite for being selected at the senior level. Moreover, this confers a higher probability of playing in the National Senior team than players not selected in youth rosters.

The breeding of captive wild species can play a crucial role in their preservation. Jaguars, the largest felines in the Americas, face challenges in captivity that hinder their breeding success in zoos and conservation institutions. One major barrier to successful captive breeding is the difficulty of replicating the natural environment, including nutritional, social, and habitat needs. Hence, any insights into the breeding behaviour and parental care of jaguars can inform and enhance breeding strategies for these captive felines. In this study, we aimed to examine maternal behaviours during the pre- and post-partum periods in the first successful case of natural reproduction of captive jaguars in Brazil. The study took place at the Mata Ciliar Association in Jundiaí , Brazil, where infrared cameras were used to observe maternal behaviours without human interference. The behaviours were analysed using an ethogram of the mother and Generalized Linear Models (GLM) to examine her behavioural repertoire during three different phases of parental care. Our findings suggest that there were differences in the behaviours exhibited by the female jaguar in the pre- and post-partum periods. Furthermore, the mother's interactions with her jaguar cubs reduced her stereotyped behaviours, implying an improvement in her welfare. Finally, we offer suggestions and recommendations to facilitate future breeding efforts for captive jaguars.

Il presente articolo propone due brevi percorsi didattici, sviluppati nell’ottica della didattica multimediale. Le attività presentate sono state adattate, in seguito alla pandemia, per essere flessibili e poter, quindi, venire proposte sia in presenza che a distanza (facendo sempre uso degli strumenti multimediali). Le due brevi unità si caratterizzano per la possibilità di essere proposte a studenti di diverso livello di conoscenza dell’italiano (dall’A1 al B1/B2). Inoltre, caratteristica centrale dal punto di vista della costruzione degli esercizi è l’utilizzo di piattaforme gratuite e la possibilità per gli studenti di interagire anche semplicemente attraverso uno smartphone, senza bisogno di un computer. Examples of multimedia teaching This paper offers two brief educational courses designed according to multimedia teaching. Following the global pandemic, the proposed activities have been adapted to be flexible so that they can be proposed both in the presence and at a distance (always using multimedia tools). The two brief activities stand out for their possibility of being proposed to students with different levels of knowledge of the Italian language (A1 to B1/B2 level). Moreover, in terms of structure, a central feature of the activities is the use of free platforms that allow students to interact simply through a smartphone without needing a computer.

IntroductionThe role of fluorodeoxyglucose (FDG) positron emission tomography/computed tomography (PET/CT) in early breast cancer treated with preoperative systemic therapy (PST) is not yet established in clinical practice. PET parameters have aroused great interest in the recent years, as non-invasive dynamic biological markers for predicting response to PST.MethodsIn this retrospective study, we included 141 patients with stage II-III breast cancer who underwent surgery after PST. Using ROC analysis, we set optimal cutoff of FDG-PET/CT parameters predictive for pathological complete response (pCR). We investigated the correlation between FDG-PET/CT parameters and pCR, median disease-free survival (DFS), and median overall survival (mOS).ResultsAt multivariable analysis, baseline SUVmax (high vs low: OR 9.00, CI 1.85 – 61.9, p=0.012) and Delta SUVmax (high vs low: OR 9.64, CI 1.84, 69.2, p=0.012) were significantly associated with pCR rates. Interestingly, we found that a combined analysis of the metabolic parameter Delta SUVmax with the volume-based parameter Delta MTV, may help to identify patients with pCR, especially in the subgroup of hormone receptor positive breast cancer. Delta SUVmax was also an independent predictive marker for both mDFS (high vs low: HR 0.17, 95%CI 0.05-0.58, p=0.004) and mOS (high vs. low: HR 0.19, 95%CI 0.04-0.95, p=0.029).DiscussionOur results suggest that Delta SUVmax may predict survival of early BC patients treated with PST.