Looking for a needle in a haystack: de novo phenotypic target identification reveals Hippo pathway-mediated miR-202 regulation of egg production.

Understanding microRNA (miRNA) functions has been hampered by major difficulties in identifying their biological target(s). Currently, the main limitation is the lack of a suitable strategy to identify biologically relevant targets among a high number of putative targets. Here we provide a proof of concept of successful de novo (i.e. without prior knowledge of its identity) miRNA phenotypic target (i.e. target whose de-repression contributes to the phenotypic outcomes) identification from RNA-seq data. Using the medaka mir-202 knock-out (KO) model in which inactivation leads to a major organism-level reproductive phenotype, including reduced egg production, we introduced novel criteria including limited fold-change in KO and low interindividual variability in gene expression to reduce the list of 2853 putative targets to a short list of 5. We selected tead3b, a member of the evolutionarily-conserved Hippo pathway, known to regulate ovarian functions, due to its remarkably strong and evolutionarily conserved binding affinity for miR-202-5p. Deleting the miR-202-5p binding site in the 3' UTR of tead3b, but not of other Hippo pathway members sav1 and vgll4b, triggered a reduced egg production phenotype. This is one of the few successful examples of de novo functional assignment of a miRNA phenotypic target in vivo in vertebrates.

Navigating PROSPERO4animals: 10 top tips for efficient pre-registration of your animal systematic review protocol.

Systematic reviews are an essential tool in identifying knowledge gaps and synthesizing evidence from in vivo animal research to improve human health. The review process follows an explicit and systematic methodology to minimize bias, but is not immune to biases or methodological flaws. Pre-registering a systematic review protocol has several benefits, including avoiding unplanned duplication of reviews, reducing reporting biases, and providing structure throughout the review process. It also helps to align the opinions of review team members and can shield researchers from post-hoc critique. PROSPERO4animals is the international prospective register of systematic reviews (PROSPERO) for the preregistration of systematic review of animal studies. As administrators, here we provide 10 tips to facilitate pre-registration in PROSPERO4animals. These tips address common difficulties that both beginners and experienced researchers may face when pre-registering their systematic review protocols. This article aims to help authors write and register a detailed systematic review protocol on PROSPERO4animals.

Healthcare professionals perspectives on tuberculosis barriers in Portuguese prisons-a qualitative study.

BACKGROUND: Tuberculosis (TB) remains a significant public health concern, particularly within prison settings, where the confluence of adverse health factors and high-risk behaviors contribute to a heightened risk of transmission. This study delves into the perspectives of medical doctors, regarding the implementation of the 2014 TB protocol in Portugal. METHODS: The study has a qualitative, descriptive design. Individual semi-structured interviews with medical doctors from TB outpatient centers in Porto and Lisbon were used for data collection. For the analysis thematic analysis method was used. RESULTS: The study population comprised 21 medical doctors with the majority being female (61.9%) and 57.1% specializing in pulmonology. The results indicate varied perceptions of the protocol's usefulness, with positive impacts on coordination reported by some participants. Improved communication and evolving collaboration between TB outpatient centers and prisons were highlighted, although challenges in contact tracing and resource constraints were acknowledged. The study also sheds light on the role of nurses in patient education. CONCLUSION: Despite overall positive perceptions, challenges such as sustaining therapy post-symptomatic improvement and delays in diagnostic methods were identified. The findings underscore the importance of continuous collaboration between prisons and TB control programs to address challenges, improve disease control and prevent TB transmission.

COVID-19 cases, vaccination, and SARS-CoV-2 in wastewater: insights from a Brazilian municipality.

Vaccines combatting COVID-19 demonstrate the ability to protect against disease and hospitalization, and reduce the likelihood of death caused by SARS-CoV-2. In addition, monitoring viral loads in sewage emerges as another crucial strategy in the epidemiological context, enabling early and collective detection of outbreaks. The study aimed to monitor the viral concentration of SARS-CoV-2 in untreated sewage in a Brazilian municipality. Also, it attempted to correlate these measurements with the number of clinical cases and deaths resulting from COVID-19 between July 2021 and July 2022. SARS-CoV-2 viral RNA was quantified by RT-qPCR. Pearson's correlation was performed to analyze the variables' relationship using the number of cases, deaths, vaccinated individuals, and viral concentration of SARS-CoV-2. The results revealed a significant negative correlation (p < 0.05) between the number of vaccinated individuals and the viral concentration of SARS-CoV-2, suggesting that after vaccination, the RNA viral load concentration was reduced in the sample population by the circulating concentration of wastewater. Consequently, wastewater monitoring, in addition to functioning as an early warning system for the circulation of SARS-CoV-2 and other pathogens, can offer a novel perspective that enhances decision-making, strengthens vaccination campaigns, and contributes to authorities establishing systematic networks for monitoring SARS-CoV-2.

A novel approach to automatic seizure detection using computer vision and independent component analysis.

OBJECTIVE: Epilepsy is a neurological disease that affects ~50 million people worldwide, 30% of which have refractory epilepsy and recurring seizures, which may contribute to higher anxiety levels and poorer quality of life. Seizure detection may contribute to addressing some of the challenges associated with this condition, by providing information to health professionals regarding seizure frequency, type, and/or location in the brain, thereby improving diagnostic accuracy and medication adjustment, and alerting caregivers or emergency services of dangerous seizure episodes. The main focus of this work was the development of an accurate video-based seizure-detection method that ensured unobtrusiveness and privacy preservation, and provided novel approaches to reduce confounds and increase reliability. METHODS: The proposed approach is a video-based seizure-detection method based on optical flow, principal component analysis, independent component analysis, and machine learning classification. This method was tested on a set of 21 tonic-clonic seizure videos (5-30 min each, total of 4 h and 36 min of recordings) from 12 patients using leave-one-subject-out cross-validation. RESULTS: High accuracy levels were observed, namely a sensitivity and specificity of 99.06% ± 1.65% at the equal error rate and an average latency of 37.45 ± 1.31 s. When compared to annotations by health care professionals, the beginning and ending of seizures was detected with an average offset of 9.69 ± 0.97 s. SIGNIFICANCE: The video-based seizure-detection method described herein is highly accurate. Moreover, it is intrinsically privacy preserving, due to the use of optical flow motion quantification. In addition, given our novel independence-based approach, this method is robust to different lighting conditions, partial occlusions of the patient, and other movements in the video frame, thereby setting the base for accurate and unobtrusive seizure detection.

Pre-injury performance is most important for predicting the level of match participation after Achilles tendon ruptures in elite soccer players: a study using a machine learning classifier.

PURPOSE: Achilles tendon ruptures (ATR) are career-threatening injuries in elite soccer players due to the decreased sports performance they commonly inflict. This study presents an exploratory data analysis of match participation before and after ATRs and an evaluation of the performance of a machine learning (ML) model based on pre-injury features to predict whether a player will return to a previous level of match participation. METHODS: The website transfermarkt.com was mined, between January and March of 2021, for relevant entries regarding soccer players who suffered an ATR while playing in first or second leagues. The difference between average minutes played per match (MPM) 1 year before injury and between 1 and 2 years after the injury was used to identify patterns in match participation after injury. Clustering analysis was performed using k-means clustering. Predictions of post-injury match participation were made using the XGBoost classification algorithm. The performance of this model was evaluated using the area under the receiver operating characteristic curve (AUROC) and Brier score loss (BSL). RESULTS: Two hundred and nine players were included in the study. Data from 32,853 matches was analysed. Exploratory data analysis revealed that forwards, midfielders and defenders increased match participation during the first year after injury, with goalkeepers still improving at 2 years. Players were grouped into four clusters regarding the difference between MPMs 1 year before injury and between 1 and 2 years after the injury. These groups ranged between a severe decrease (n = 34; - 59 ± 13 MPM), moderate decrease (n = 75; - 25 ± 8 MPM), maintenance (n = 70; 0 ± 8 MPM), or increase (n = 30; 32 ± 13 MPM). Regarding the predictive model, the average AUROC after cross-validation was 0.81 ± 0.10, and the BSL was 0.12, with the most important features relating to pre-injury match participation. CONCLUSION: Most players take 1 year to reach peak match participation after an ATR. Good performance was attained using a ML classifier to predict the level of match participation following an ATR, with features related to pre-injury match participation displaying the highest importance. LEVEL OF EVIDENCE: I.

Expression of genes containing tandem repeat patterns involved in the fungal-host interaction and in the response to antifungals in Trichophyton rubrum.

BACKGROUND: Trichophyton rubrum is the most common aetiological agent of human dermatophytoses. These infections mainly occur in keratinised layers such as skin, hair and nails because the fungus uses keratin as a nutrient source. Fluconazole and amphotericin are antifungal agents most commonly used to treat dermatophytoses and acts on cell membrane ergosterol. Despite the clinical importance of T rubrum, the mechanisms underlying the fungal-host relationship have not yet been clarified. Tandem repeats (TRs) are short DNA sequences that are involved in a variety of adaptive functions, including the process of fungal infection. It is known that the larger the number of TRs in the genome, the greater the capacity of cell-cell junction and surface adhesion, especially when these repeats are present in regions encoding cell surface proteins. OBJECTIVES: To identify in silico T rubrum genes containing TR patterns and to analyse the modulation of these genes in culture medium containing keratin (a model simulating skin infection) and antifungal drugs. METHODS: The Dermatophyte Tandem Repeats Database (DTRDB) and the FaaPred tool were used to identify four T rubrum genes containing TR patterns. Quantitative real-time (RT) PCR was used to evaluate the gene expression during the growth of T rubrum on keratin and in the presence of fluconazole, amphotericin B and Congo red (acts in the cell wall). RESULTS: The expression of these genes was found to be induced in culture medium containing keratin. In addition, these genes were induced in the presence of antifungal agents, especially fluconazole, indicating an adaptive response to the stress caused by this drug. CONCLUSION: The results suggest an important role of genes containing TRs in the fungal-host interaction and in the susceptibility to inhibitory compounds, indicating these sequences as new potential targets for the development of antifungal agents.

Two years into the Brazilian Reproducibility Initiative: reflections on conducting a large-scale replication of Brazilian biomedical science.

Scientists have increasingly recognised that low methodological and analytical rigour combined with publish-or-perish incentives can make the published scientific literature unreliable. As a response to this, large-scale systematic replications of the literature have emerged as a way to assess the problem empirically. The Brazilian Reproducibility Initiative is one such effort, aimed at estimating the reproducibility of Brazilian biomedical research. Its goal is to perform multicentre replications of a quasi-random sample of at least 60 experiments from Brazilian articles published over a 20-year period, using a set of common laboratory methods. In this article, we describe the challenges of managing a multicentre project with collaborating teams across the country, as well as its successes and failures over the first two years. We end with a brief discussion of the Initiative's current status and its possible future contributions after the project is concluded in 2021.

Variable Ventilation Associated With Recruitment Maneuver Minimizes Tissue Damage and Pulmonary Inflammation in Anesthetized Lung-Healthy Rats.

BACKGROUND: Recruitment maneuver and positive end-expiratory pressure (PEEP) can be used to counteract intraoperative anesthesia-induced atelectasis. Variable ventilation can stabilize lung mechanics by avoiding the monotonic tidal volume and protect lung parenchyma as tidal recruitment is encompassed within the tidal volume variability. METHODS: Forty-nine (7 per group) male Wistar rats were anesthetized, paralyzed, and mechanically ventilated. A recruitment maneuver followed by stepwise decremental PEEP titration was performed while continuously estimating respiratory system mechanics using recursive least squares. After a new recruitment, animals were ventilated for 2 hours in volume-control with monotonic (VCV) or variable (VV) tidal volumes. PEEP was adjusted at a level corresponding to the minimum elastance or 2 cm H2O above or below this level. Lungs were harvested for histologic analysis (left lung) and cytokines measurement (right lung). Seven animals were euthanized before the first recruitment as controls. RESULTS: A time-dependent increase in respiratory system elastance was observed and significantly minimized by PEEP (P < .001). Variable ventilation attenuated the amount of concentrations of proinflammatory mediators in lung homogenate: neutrophil cytokine-induced neutrophil chemoattractant 1 (VV = 40 ± 5 and VCV = 57 ± 8 pg/mg; P < .0001) and interleukin-1ß (VV = 59 ± 25 and VCV = 261 ± 113 pg/mg; P < .0001). Variable ventilation was also associated with lower structural lung parenchyma damage. Significant reductions in air fraction at dorsal and caudal lung regions were observed in all ventilated animals (P < .001). CONCLUSIONS: Variable ventilation was more protective than conventional ventilation within the applied PEEP levels.

Anticancer effects of seaweed compounds fucoxanthin and phloroglucinol, alone and in combination with 5-fluorouracil in colon cells.

Colorectal cancer therapy with 5-fluorouracil (5-Fu) frequently become ineffective due to resistance to this drug; and thus other effective compounds are essential for therapy. It is well-known marine brown seaweeds contain antioxidant compounds the carotenoid fucoxanthin (Fx) and polyphenolic compound phloroglucinol (Ph) which exerted diverse biological activities including antioxidant and anticancer. The aim of this study was to determine the anticancer activities of Fx or Ph alone as well as combination of each chemical with 5-Fu on two human colorectal cancer cell lines (HCT116 and HT29), with comparison to responses in a normal colon cell line (CCD-18Co). Effects of these compounds on cell viability, induction of DNA damage, and cell death were evaluated using MTT assay, comet assay, nuclear condensation assay, and Western blot. 5-Fu decreased cell viability in a concentration-dependent manner in HCT116 and HT29 cells but was not cytotoxic in CCD-18Co cells. 5-Fu induced DNA damage in HCT116 cells with induction of cell death, while no marked effects on DNA damage and cell death were observed in HT29 cells. Fx or Ph alone also reduced cell viability in both cancer cell lines but no apparent cytotoxic effect in CCD-18Co cells, except for Fx at 50 and 100 µM. Diminished cell viability was accompanied by induction of DNA damage (by Fx) and induction of cell death (by Ph). In combination with 5-Fu, Fx at 10 µM (in HCT116 and HT29 cells), and Ph at 300 µM (in HT29 cells) enhanced the cytotoxic effect of 5-Fu; however, no marked cytotoxicity was noted in CCD-18Co cells. Since Fx and Ph alone reduced cancer cell line viability without an effect on normal cells and when in combination enhanced the cytotoxic effect of 5-Fu only in colon cancer cells, these compounds seem promising as anticancer agents.