Development of Rapid Detection Methods for Fusarium oysporum f. sp. melonis in Melon Seeds.

Melon (Cucumis melo L.) is a global commercial crop that is sensitive to seed-borne wilt infections caused by Fusarium oxysporum f. sp. melonis (Fom). To address the challenge of detecting Fom contamination, we designed a probe-based real-time PCR method, TDCP2, in combination with rapid or column-based DNA extraction protocols to develop reliable molecular detection methods. Utilizing TDCP2, the detection rate reached 100% for both artificially Fom-inoculated (0.25-25%) and pod-inoculated melon seeds in conjunction with DNA samples from either the rapid or column-based extraction protocol. We performed analyses of precision, recall, and F1 scores, achieving a maximum F1 score of 1 with TDCP2, which highlights the robustness of the method. Additionally, intraday and interday assays were performed, which revealed the high reproducibility and stability of column-based DNA extraction protocols combined with TDCP2. These metrics confirm the reliability of our developed protocols, setting a foundation for future enhancements in seed pathology diagnostics and potentially broadening their applicability across various Fom infection levels. In the future, we hope that these methods will reduce food loss by improving the control and management of melon diseases.

The role of coagulopathy and subdural hematoma thickness at admission in predicting the prognoses of patients with severe traumatic brain injury: A multicenter retrospective cohort study from China.

BACKGROUND: Traumatic brain injury (TBI) is one of the diseases with high disability and mortality worldwide. Recent studies have shown that TBI-related factors may change the complex balance between bleeding and thrombosis, leading to coagulation disorders. The aim of this retrospective study was to investigate the prediction of coagulopathy and subdural hematoma thickness at admission using the Glasgow Outcome Scale (GOS) in patients with severe TBI at 6 months after discharge. METHODS: In this retrospective cohort study, a total of 1,006 patients with severe TBI in large medical centers in three different provinces of China from June 2015 to June 2021 were enrolled after the exclusion criteria, and 800 patients who met the enrollment criteria were included. A receiver operating characteristic (ROC) curve was used to determine the best cut-off values of platelet (PLT), international normalized ratio (INR), activated partial thromboplastin time (APTT), and subdural hematoma (SDH) thickness. The ROC curve, nomogram, calibration curve, and the decision curve were used to evaluate the predictive effect of the coagulopathy and Coagulopathy-SDH(X1) models on the prognoses of patients with severe TBI, and the importance of predictive indicators was ranked by machine learning. RESULTS: Among the patients with severe TBI on admission, 576/800 (72%) had coagulopathy, 494/800 (61%) had SDH thickness ≥14.05 mm, and 385/800 (48%) had coagulopathy combined with SDH thickness ≥14.05 mm. Multivariate logistic regression analyses showed that age, pupil, brain herniation, WBC, CRP, SDH, coagulopathy, and X1 were independent prognostic factors for GOS after severe TBI. Compared with other single indicators, X1 as a predictor of the prognosis of severe TBI was more accurate. The GOS of patients with coagulopathy and thick SDH (X1, 1 point) at 6 months after discharge was significantly worse than that of patients with coagulopathy and thin SDH (X1, 2 points), patients without coagulopathy and thick SDH (X1, 3 point), and patients without coagulopathy and thin SDH (X1, 4 points). In the training group, the C-index based on the coagulopathy nomogram was 0.900. The C-index of the X1-based nomogram was 0.912. In the validation group, the C-index based on the coagulopathy nomogram was 0.858. The C-index of the X1-based nomogram was 0.877. Decision curve analysis also confirmed that the X1-based model had a higher clinical net benefit of GOS at 6 months after discharge than the coagulopathy-based model in most cases, both in the training and validation groups. In addition, compared with the calibration curve based on the coagulopathy model, the prediction of the X1 model-based calibration curve for the probability of GOS at 6 months after discharge showed better agreement with actual observations. Machine learning compared the importance of each independent influencing factor in the evaluation of GOS prediction after TBI, with results showing that the importance of X1 was better than that of coagulopathy alone. CONCLUSION: Coagulopathy combined with SDH thickness could be used as a new, accurate, and objective clinical predictor, and X1, based on combining coagulopathy with SDH thickness could be used to improve the accuracy of GOS prediction in patients with TBI, 6 months after discharge.

SIRT1-Mediated HMGB1 Deacetylation Suppresses Neutrophil Extracellular Traps Related to Blood-Brain Barrier Impairment After Cerebral Venous Thrombosis.

Cerebral venous thrombosis (CVT) is a neurovascular disease with recently increasing incidence. Aseptic inflammatory responses play an important role in the pathology of CVT. Recent studies report that neutrophil extracellular traps (NETs) are major triggers of thrombosis and inflammation in stroke, but their effect on brain injury in CVT requires further validation. In this study, two CVT animal models were used to simulate superior sagittal sinus thrombosis and cortical vein thrombosis. The effects of brain tissue infiltration of NETs and the molecular mechanisms associated with NET formation were deeply explored in combination with proteomics, histology, and serology. The results showed that the cortical vein thrombosis model could be combined with more severe blood-brain barrier (BBB) disruption and showed more severe cerebral hemorrhage. Decreased Sirtuin 1 (SIRT1) expression promotes high mobility group box 1 (HMGB1) acetylation, causing increased cytosolic translocation and extracellular release, and HMGB1 can promote NET formation and recruitment. In addition, corticocerebral accumulation of NETs contributes to BBB damage. This establishes a vicious cycle between BBB damage and NET accumulation. SIRT1 mediated-HMGB1 deacetylation may play a critical role in attenuating BBB damage following CVT. This study employed a combined validation using models of venous sinus thrombosis and cortical vein thrombosis to investigate the deacetylation role of SIRT1, aiming to offer new insights into the pathological mechanisms of brain injury following CVT.

Analysis of the Pathogenicity and Phylogeny of Colletotrichum Species Associated with Brown Blight of Tea (Camellia sinensis) in Taiwan.

Brown blight, a destructive foliar disease of tea, has become a highly limiting factor for tea cultivation in Taiwan. To understand the population composition of the causal agents (Colletotrichum spp.), the fungal diversity in the main tea-growing regions all over Taiwan was surveyed from 2017 to 2019. A collection of 139 Colletotrichum isolates was obtained from 14 tea cultivars in 86 tea plantations. Phylogenic analysis using the ribosomal internal transcribed spacer, glutamine synthetase gene, Apn2-Mat1-2 intergenic spacer, ß-tubulin, actin, calmodulin, and glyceraldehyde-3-phosphate dehydrogenase genes together with morphological characterization revealed three species associated with brown blight of tea; namely, Colletotrichum camelliae (95.6% of all isolates), C. fructicola (3.7%), and C. aenigma (0.7%). This is the first report of C. aenigma in Taiwan. The optimal growth temperatures were 25°C for C. camelliae and 25 and 30°C for C. fructicola and C. aenigma. Although C. fructicola and C. aenigma were more adapted to high temperature, C. camelliae was the most pathogenic across different temperatures. Regardless of whether spore suspensions or mycelial discs were used, significantly larger lesions and higher disease incidences were observed for wounded than for nonwounded inoculation and for the third and fourth leaves than for the fifth leaves. Wounded inoculation of detached third and fourth tea leaves with mycelial discs was found to be a reliable and efficient method for assessing the pathogenicity of Colletotrichum spp. within 4 days. Preventive application of fungicides or biocontrol agents immediately after tea pruning and at a young leaf stage would help control the disease.

Brain functional activity of swallowing: A meta-analysis of functional magnetic resonance imaging.

BACKGROUND: Swallowing is one of the most important activities in our life and serves the dual roles of nutritional intake and eating enjoyment. OBJECTIVE: The study aimed to conduct a meta-analysis to investigate the brain activity of swallowing. METHODS: Studies of swallowing using functional magnetic resonance imaging were reviewed in PubMed, Web of Science, China National Knowledge Infrastructure (CNKI), Chinese Science and Technology Periodical Database (VIP) and Wan Fang before 30 November 2021. Two authors analysed the studies for eligibility criteria. The final inclusion of studies was decided by consensus. An activation likelihood estimation (ALE) meta-analysis of these studies was performed with GingerALE, including 16 studies. RESULTS: For swallowing, clusters with high activation likelihood were found in the bilateral insula, bilateral pre-central gyrus, bilateral post-central gyrus, left transverse temporal gyrus, right medial front gyrus, bilateral inferior frontal gyrus and bilateral cingulate gyrus. For water swallowing, clusters with high activation likelihood were found in the bilateral inferior frontal gyrus and the left pre-central gyrus. For saliva swallowing, clusters with high activation likelihood were found in the bilateral cingulate gyrus, bilateral pre-central gyrus, left post-central gyrus and left transverse gyrus. CONCLUSION: This meta-analysis reflects that swallowing is regulated by both sensory and motor cortex, and saliva swallowing activates more brain areas than water swallowing, which would promote our knowledge of swallowing and provide some direction for clinical and other research.

Identification of Cyprodinil + Fludioxonil to Manage Soybean Red Crown Rot Using the Microplate-Based High-Throughput Screening and Pot Assay.

Red crown rot (RCR), caused by the soilborne fungus Calonectria ilicicola, is an emerging soybean disease in Taiwan, and fungicide screening is desired to identify effective management for C. ilicicola. This study screened 11 fungicides, including azoxystrobin, boscalid, cyprodinil, cyprodinil + fludioxonil, difenoconazole, fluopyram, flutolanil, mancozeb, prochloraz, pyraclostrobin, and tebuconazole, for their inhibitory effects on the mycelial growth of 10 C. ilicicola field isolates. Subsequently, a microplate-based high-throughput screening (MHTS) method was established to measure the fungicide sensitivity in a population composed of 80 C. ilicicola isolates to three effective fungicides, cyprodinil + fludioxonil, fluopyram, and tebuconazole. The MHTS was optimized for multiple factors, including the optical scanning pattern, absorption wavelength, conidial concentration, and measurement timing based on the quality controls of Z' factor and the log-phase growth curve. The population mean EC50 estimated by MHTS were 0.14, 2.34, and 2.46 ppm to cyprodinil + fludioxonil, fluopyram, and tebuconazole, respectively. In addition to the in vitro assessment, fungicide efficacy was evaluated by coating cyprodinil + fludioxonil, fluopyram, or tebuconazole on soybean seeds in the pot assay. The results showed that cyprodinil + fludioxonil significantly reduced both postemergence damping-off and disease severity, while fluopyram and tebuconazole reduced only the postemergence damping-off but not disease severity. Based on the MHTS and the pot assay results, this study demonstrated cyprodinil + fludioxonil to be a potential fungicide to manage soybean RCR.

Rapid Detection of Fusarium oxysporum Using Insulated Isothermal PCR and a Rapid, Simple DNA Preparation Protocol.

We developed an insulated isothermal PCR (iiPCR) method for the efficient and rapid detection of Fusarium oxysporum (Fo), which is a fungus that infects various hosts and causes severe crop losses. The Fo iiPCR method was sensitive enough to detect up to 100 copies of standard DNA template and 10 fg of Fo genomic DNA. In addition, it could directly detect 1 pg of mycelium and 10 spores of Fo without DNA extraction. Our study compared the performance of Fo iiPCR to that of three published in planta molecular detection methods-conventional PCR, SYBR green-based real-time PCR, and hydrolysis probe-based real-time PCR-in field detection of Fo. All diseased field samples yielded positive detection results with high reproducibility when subjected to an Fo iiPCR test combined with a rapid DNA extraction protocol compared to Fo iiPCR with an automated magnetic bead-based DNA extraction protocol. Intraday and interday assays were performed to ensure the stability of this new rapid detection method. The results of detection of Fo in diseased banana pseudostem samples demonstrated that this new rapid detection method was suitable for field diagnosis of Fusarium wilt and had high F1 scores for detection (the harmonic mean of precision and recall of detection) for all asymptomatic and symptomatic Fo-infected banana samples. In addition, banana samples at four growth stages (seedling, vegetative, flowering and fruiting, and harvesting) with mild symptoms also showed positive detection results. These results indicate that this new rapid detection method is a potentially efficient procedure for on-site detection of Fo.

Metabolic Characteristics and M2 Macrophage Infiltrates in Invasive Nonfunctioning Pituitary Adenomas.

Objective: The aim of this study was to investigate the metabolic differences between invasive and non-invasive nonfunctioning pituitary adenomas (NFPAs), determine the expression of an M2 macrophage marker in NFPAs, and analyze the effects of metabolic changes in invasive NFPAs on M2 macrophage infiltrates. Methods: Tissue samples of NFPAs from patients who underwent transsphenoidal or craniotomy surgery from January 2021 to August 2021 were collected. NFPA tissues were analyzed based on a gas chromatography-mass spectrometry non-targeted metabolomics platform, and immunohistochemical staining for M2 macrophage marker CD206 was performed. Results: We evaluated 15 invasive and 21 non-invasive NFPAs. A total of 22 metabolites were identified through non-targeted metabolomics analysis. Among them, the expression of 1-octadecanol, inosine 5'-monophosphate, adenosine 5'-monophosphate, guanosine 5'-monophosphate, creatinine, desmosterol, taurine, hypotaurine, lactic acid, and succinic acid was upregulated in invasive NFPAs, while that of 1-oleoylglycerol, arachidonic acid, cis-11-eicosenoic acid, docosahexaenoic acid, glyceric acid, hypoxanthine, linoleic acid, lysine, oleic acid, uracil, valine, and xanthine was downregulated. Immunohistochemical analysis suggested that the number of CD206-positive cells was higher in invasive NFPAs than in non-invasive NFPAs. Conclusion: Invasive and non-invasive NFPAs showed distinct metabolite profiles. The levels of succinic acid and lactic acid were higher in invasive NFPAs, and the high expression of the M2 macrophage marker was verified in invasive NFPAs.

Novel Nomograms to Predict Delayed Hyponatremia After Transsphenoidal Surgery for Pituitary Adenoma.

Objective: This study aimed to develop a nomogram of clinical variables and magnetic resonance imaging scans to predict delayed hyponatremia after transsphenoidal surgery for pituitary adenoma. Methods: Patients who underwent transsphenoidal surgery for pituitary adenoma in Fuzong Clinical Medical College of Fujian Medical University between January 2012 and December 2020 were retrospectively investigated. Medical records, MRI findings, and laboratory examination results were recorded as candidate variable predictors of delayed hyponatremia. A nomogram to predict delayed hyponatremia was formulated based on the multivariable model of risk factors. The predictive accuracy and discriminative ability of the nomogram were assessed using the receiver operating characteristic (ROC) curve, calibration plot, and decision curve analyses. The model underwent prospective validation in three medical centers with patients who underwent transsphenoidal surgery for pituitary adenoma between January 2021 and February 2022. Results: The model that incorporated the postoperative length of "measurable pituitary stalk," pituitary stalk deviation angle difference, postoperative diabetes insipidus, sinking depth of diaphragma sellae, and blood sodium level on the second postoperative day was developed and presented as the nomogram of the training cohort. The nomogram achieved area under the ROC curve (AUCs) of 0.806 and 0.849 for the training cohort and the testing cohort, respectively, and displayed good calibration. Decision curve analysis showed that the nomogram was clinically useful when the threshold probability was 13-96%. Conclusions: We developed a nomogram to evaluate the individualized prediction of delayed hyponatremia after transsphenoidal surgery for pituitary adenomas.

A novel rat model for cerebral venous sinus thrombosis: verification of similarity to human disease via clinical analysis and experimental validation.

BACKGROUND: Cerebral venous sinus thrombosis (CVST) is a rare neurovascular disorder with highly variable manifestations and clinical courses. Animal models properly matched to the clinical form of CVST are necessary for elucidating the pathophysiology of the disease. In this study, we aimed to establish a rat model that accurately recapitulates the clinical features of CVST in human patients. METHODS: This study consisted of a clinical analysis and animal experiments. Clinical data for two centres obtained between January 2016 and May 2021 were collected and analysed retrospectively. In addition, a Sprague-Dawley rat model of CVST was established by inserting a water-swellable rubber device into the superior sagittal sinus, following which imaging, histological, haematological, and behavioural tests were used to investigate pathophysiological changes. Principal component analysis and hierarchical clustering heatmaps were used to evaluate the similarity between the animal models and human patients. RESULTS: The imaging results revealed the possibility of vasogenic oedema in animal models. Haematological analysis indicated an inflammatory and hypercoagulable state. These findings were mostly matched with the retrospective clinical data. Pathological and serological tests further revealed brain parenchymal damage related to CVST in animal models. CONCLUSIONS: We successfully established a stable and reproducible rat model of CVST. The high similarity between clinical patients and animal models was verified via cluster analysis. This model may be useful for the study of CVST pathophysiology and potential therapies.

First report of leaf brown blight caused by Neopestalotiopsis formicarum on jabuticaba in Taiwan.

Jabuticaba (Plinia cauliflora (Mart.) Kausel) was originated from Brazil (Lorenzi 2000). The production of jabuticaba is growing globally as its value in the food and pharmaceutical industries (Benvenutti 2021). In August 2019, jabuticaba plants with symptoms of leaf blight were observed in the field at the Meinong of Kaohsiung City, Taiwan. Disease incidence was 40%. Symptoms first presented as small, water-soaked lesions on young leaves, and then dark brown lesions of 1-3 cm in diameter on mature leaves. Six symptomatic leaves were collected from 6 jabuticaba plants for verifying the causal agents. Tissues (5 × 5 mm2) were cut from the margin of symptomatic leaf. Samples were sterilized in 1% sodium hypochlorite for 60 s, rinsed with sterile distilled water three times and then placed in 1% water agar in the dark for 5 days at room temperature. Resultant fungal colonies were purified by subculturing fungal hyphal tips on potato dextrose agar in a growth chamber (28°C, 12 h photoperiod) until fungal conidia appeared. The fungi initially produced white, cottony, aerial mycelium, after which concentric black conidiomata appeared on the plates after 7 days of incubation. The 5-celled conidia were fusiform to ellipsoid, straight to slightly curved, with sizes of 24.00-44.00 µm × 6.00-13.00 µm (avg. size, 32.00 × 9.37 µm, n = 120). The apical and basal cells were hyaline, and 3 median cells were pale brown and versicolorous. Conidia had 2-3 apical appendages and a conical basal cell with a truncate base. Based on the characteristics, which were common among isolates from diseased samples, the causal pathogen was identified as Neopestalotiopsis sp. (Solarte et al. 2018). Internal transcribed spacer (ITS), translation elongation factor 1α (TEF), ß-tubulin, and large ribosomal subunit (LSU) DNA sequences were obtained from these isolates and deposited in GenBank (MN723897, ITS; MN813055, TEF; MN813054, ß-tubulin; MN860104, LSU). Sequences demonstrated high sequence identity with those of Neopestalotiopsis formicarum ex-type cultures CBS 362.72 (Maharachchikumbura et al. 2014): 99.44% for ITS (KM199358), 99.38% for TEF (KM199517), 98.86% for ß-tubulin (KM199455), and 100.00% for LSU (KM116248). The phylogenetic relationship in Neopestalotiopsis species supported the identification of our isolates as N. formicarum. Three independent 3-isolate inoculation experiments were performed to fit Koch's postulates. Surface-sterilized leaves on live plants were punctured with a needle and inoculated with 5 µL of conidial suspension (1 × 105 conidia/mL). Inoculated plants were kept in a growth chamber (25°C, 70% relative humidity) for 7 days. Control plants were inoculated with sterile distilled water and kept under the same conditions. Inoculated leaves developed brown lesions around wounds after seven days. The pathogen was re-isolated from diseased plants, following the steps used for the original procedure, with identical characteristics as the initial isolates. This is the first report of leaf brown blight caused by N. formicarum on jabuticaba in Taiwan. N. formicarum was recently considered as a new threat to jabuticaba (Gualberto et al. 2021). In addition, it has a broad host range on many tropical crops, such as guarana and banana (Gualberto et al. 2021). Neopestalotiopsis spp. have been reported to cause important economic fruit diseases (Gualberto et al. 2021). Therefore, N. formicarum may become the potential risk for fruit production of tropical crops.

Superoxide Initiates the Hyphal Differentiation to Microsclerotia Formation of Macrophomina phaseolina.

The infection of Macrophomina phaseolina often results in a grayish appearance with numerous survival structures, microsclerotia, on the plant surface. Past works have studied the development of fungal survival structures, sclerotia and microsclerotia, in the Leotiomycetes and Sordariomycetes. However, M. phaseolina belongs to the Dothideomycetes, and it remains unclear whether the mechanism of microsclerotia formation remains conserved among these phylogenetic clades. This study applied RNA-sequencing (RNA-Seq) to profile gene expressions at four stages of microsclerotia formation, and the results suggested that reactive oxygen species (ROS)-related functions were significantly different between the microsclerotia stages and the hyphal stage. Microsclerotia formation was reduced in the plates amended with antioxidants such as ascorbic acid, dithiothreitol (DTT), and glutathione. Surprisingly, DTT drastically scavenged H2O2, but the microsclerotia amount remained similar to the treatment of ascorbic acid and glutathione that both did not completely eliminate H2O2. This observation suggested the importance of [Formula: see text] over H2O2 in initiating microsclerotia formation. To further validate this hypothesis, the superoxide dismutase 1 (SOD1) inhibitor diethyldithiocarbamate trihydrate (DETC) and H2O2 were tested. The addition of DETC resulted in the accumulation of endogenous [Formula: see text] and more microsclerotia formation, but the treatment of H2O2 did not. The expression of SOD1 genes were also found to be upregulated in the hyphae to the microsclerotia stage, which suggested a higher endogenous [Formula: see text] stress presented in these stages. In summary, this study not only showed that the ROS stimulation remained conserved for initiating microsclerotia formation of M. phaseolina but also highlighted the importance of [Formula: see text] in initiating the hyphal differentiation to microsclerotia formation. IMPORTANCE Reactive oxygen species (ROS) have been proposed as the key stimulus for sclerotia development by studying fungal systems such as Sclerotinia sclerotiorum, and the theory has been adapted for microsclerotia development in Verticillium dahliae and Nomuraea rileyi. While many studies agreed on the association between (micro)sclerotia development and the ROS pathway, which ROS type, superoxide ([Formula: see text]) or hydrogen peroxide (H2O2), plays a major role in initiating hyphal differentiation to the (micro)sclerotia formation remains controversial, and literature supporting either [Formula: see text] or H2O2 can be found. This study confirmed the association between ROS and microsclerotia formation for the charcoal rot fungus Macrophomina phaseolina. Moreover, the accumulation of [Formula: see text] but not H2O2 was found to induce higher density of microsclerotia. By integrating transcriptomic and phenotypic assays, this study presented the first conclusive case for M. phaseolina that [Formula: see text] is the main ROS stimulus in determining the amount of microsclerotia formation.

Systemic immune inflammation index and peripheral blood carbon dioxide concentration at admission predict poor prognosis in patients with severe traumatic brain injury.

Background: Recent studies have shown that systemic inflammation responses and hyperventilation are associated with poor outcomes in patients with severe traumatic brain injury (TBI). The aim of this retrospective study was to investigate the relationships between the systemic immune inflammation index (SII = platelet × neutrophil/lymphocyte) and peripheral blood CO2 concentration at admission with the Glasgow Outcome Score (GOS) at 6 months after discharge in patients with severe TBI. Methods: We retrospectively analyzed the clinical data for 1266 patients with severe TBI at three large medical centers from January 2016 to December 2021, and recorded the GOS 6 months after discharge. The receiver operating characteristic (ROC) curve was used to determine the best cutoff values for SII, CO2, neutrophil to lymphocyte ratio (NLR), platelet to lymphocyte ratio (PLR), and lymphocyte to monocyte ratio (LMR), and chi-square tests were used to evaluate the relationships among SII, CO2 and the basic clinical characteristics of patients with TBI. Multivariate logistic regression analysis was used to determine the independent prognostic factors for GOS in patients with severe TBI. Finally, ROC curve, nomogram, calibration curve and decision curve analyses were used to evaluate the value of SII and coSII-CO2 in predicting the prognosis of patients with severe TBI. And we used the multifactor regression analysis method to build the CRASH model and the IMPACT model. The CRASH model included age, GCS score (GCS, Glasgow Coma Scale) and Pupillary reflex to light: one, both, none. The IMPACT model includes age, motor score and Pupillary reflex to light: one, both, none. Results: The ROC curves indicated that the best cutoff values of SII, CO2, PLR, NLR and LMR were 2651.43×109, 22.15mmol/L, 190.98×109, 9.66×109 and 1.5×109, respectively. The GOS at 6 months after discharge of patients with high SII and low CO2 were significantly poorer than those with low SII and high CO2. Multivariate logistic regression analysis revealed that age, systolic blood pressure (SBP), pupil size, subarachnoid hemorrhage (SAH), SII, PLR, serum potassium concentration [K+], serum calcium concentration [Ca2+], international normalized ratio (INR), C-reactive protein (CRP) and co-systemic immune inflammation index combined with carbon dioxide (coSII-CO2) (P < 0.001) were independent prognostic factors for GOS in patients with severe TBI. In the training group, the C-index was 0.837 with SII and 0.860 with coSII-CO2. In the external validation group, the C-index was 0.907 with SII and 0.916 with coSII-CO2. Decision curve analysis confirmed a superior net clinical benefit with coSII-CO2 rather than SII in most cases. Furthermore, the calibration curve for the probability of GOS 6 months after discharge showed better agreement with the observed results when based on the coSII-CO2 rather than the SII nomogram. According to machine learning, coSII-CO2 ranked first in importance and was followed by pupil size, then SII. Conclusions: SII and CO2 have better predictive performance than NLR, PLR and LMR. SII and CO2 can be used as new, accurate and objective clinical predictors, and coSII-CO2, based on combining SII with CO2, can be used to improve the accuracy of GOS prediction in patients with TBI 6 months after discharge.

Establishment of a Rat Model of Superior Sagittal-Sinus Occlusion via a Thread-Embolism Method.

The mechanisms contributing to the natural onset of cerebral venous sinus thrombosis (CVST) are mostly unknown, and a variety of uncontrollable factors are involved in the course of the disease, resulting in great limitations in clinical research. Therefore, the establishment of stable CVST animal models that can standardize a variety of uncontrollable confounding factors have helped to circumvent shortcomings in clinical research. In recent decades, a variety of CVST animal models have been constructed, but the results based on these models have been inconsistent and incomplete. Hence, in order to further explore the pathophysiological mechanisms of CVST, it is necessary to establish a novel and highly compatible animal model, which has important practical value and scientific significance for the diagnosis and treatment of CVST. In the present study, a novel Sprague-Dawley (SD) rat model of superior sagittal sinus (SSS) thrombosis was established via a thread-embolization method, and the stability and reliability of the model were verified. Additionally, we evaluated changes in cerebral venous blood flow in rats after the formation of CVST. Collectively, the SD-rat SSS-thrombosis model represents a novel CVST animal model that is easily established, minimizes trauma, yields good stability, and allows for accurately controlling ischemic timing and location.

Outcomes of Preinjury Use of Statins in Patients with Traumatic Brain Injury: A Systematic Review and Meta-analysis.

BACKGROUND: No completely effective pharmacotherapies have been developed to improve the outcomes of traumatic brain injury (TBI). Given the reporting of cohort studies suggesting that preinjury statin use may reduce TBI-associated mortality, this study aimed to evaluate the effects of statin use in patients with TBI. METHODS: This study was performed according to the PRISMA guidelines. The PubMed, Embase, Cochrane Central, Web of Science, and China National Knowledge Infrastructure databases were searched from inception until April 13, 2021, using a search strategy that included 2 main terms: "statins" and "traumatic brain injury." The outcomes were mortality, hospital length of stay, and intensive care unit length of stay, which were evaluated using a random-effects model and represented by the pooled risk ratio with 95% confidence intervals. RESULTS: The search results identified 7 eligible studies, with a total of 111,935 patients with brain injury. Preinjury statin use in patients with TBI was associated with a significantly decreased risk of mortality compared with that in nonusers (risk ratio, 0.75; 95% confidence interval, 0.59-0.94; I2 = 53%). Subgroup analysis showed that statin withdrawal might increase mortality. Sensitivity analysis showed that the results were stable and robust. CONCLUSIONS: Preinjury statin use may contribute to mortality reduction in patients with TBI, whereas statin withdrawal might increase mortality. In clinical management, statin use should not be discontinued after TBI.

Whole-Genome Sequence Resource of Calonectria ilicicola, the Casual Pathogen of Soybean Red Crown Rot.

Calonectria ilicicola (anamorph: Cylindrocladium parasiticum) is a soilborne plant-pathogenic fungus with a broad host range, and it can cause red crown rot of soybean and Cylindrocladium black rot of peanut, which has become an emerging threat to crop production worldwide. Limited molecular studies have focused on Calonectria ilicicola and one of the possible difficulties is the lack of genomic resources. This study presents the first high quality and near-completed genome of C. ilicicola, using the Oxford Nanopore GridION sequencing platform. A total of 16 contigs were assembled and the genome of C. ilicicola isolate F018 was estimated to have 11 chromosomes. Currently, the C. ilicicola F018 genome represents the most contiguous assembly, which has the lowest contig number and the highest contig N50 among all Calonectria genome resources. Putative protein-coding sequences and secretory proteins were estimated to be 17,308 and 1,930 in the C. ilicicola F018 genome, respectively; and the prediction was close to other plant-pathogenic fungi, such as Fusarium species, within the Nectriaceae family. The availability of this high-quality genome resource is expected to facilitate research on fungal biology and genetics of C. ilicicola and to support advanced understanding of pathogen virulence and disease management.[Formula: see text] Copyright © 2021 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Degenerated Virulence and Irregular Development of Fusarium oxysporum f. sp. niveum Induced by Successive Subculture.

Successive cultivation of fungi on artificial media has been reported to cause the sectorization, which leads to degeneration of developmental phenotype, and virulence. Fusarium oxysporum f. sp. niveum (Fon), the causal agent of watermelon Fusarium wilt, forms degenerated sectors after successive cultivation. In the present research, we demonstrated that subculture with aged mycelia increased the incidence of degenerations. To further investigate the differences between the Fon wild type (sporodochial type, ST) and variants (MT: mycelial type and PT: pionnotal type), developmental phenotypes and pathogenicity to watermelon were examined. Results in variants (PT2, PT3, PT11, and MT6) were different from ST with mycelia growth, conidia production and chlamydospore formation. Virulence of degenerated variants on susceptible watermelon Grand Baby (GB) cultivar was determined after inoculation with Fon variants and Fon ST. In root dipping methods, Fon variants showed no significant differences in disease progress compared with ST. Fon variants showed a significant decrease in disease progression compared with ST through infested soil inoculation. The contrasting results of two inoculation methods suggest that the degenerative changes due to repeated successive cultivation may lead to the loss of pathogen virulence-related factors of the early stage of Fon infection process. Therefore, cell wall-degrading enzymes (CWDEs; cellulase, pectinase, and xylanase) activities of different variants were analyzed. All Fon degenerated variants demonstrated significant decreases of CWDEs activities compared with ST. Additionally, transcript levels of 9 virulence-related genes (fmk1, fgb1, pacC, xlnR, pl1, rho1, gas1, wc1, and fow1) were assessed in normal state. The degenerated variants demonstrated a significantly low level of tested virulence-related gene transcripts except for fmk1, xlnR, and fow1. In summary, the degeneration of Fon is triggered with successive subculture through aged mycelia. The degeneration showed significant impacts on virulence to watermelon, which was correlated with the reduction of CWDEs activities and declining expression of a set of virulence-related genes.

First report of anthracnose caused by Colletotrichum fructicola on tea in Taiwan.

Tea (Camellia sinensis (L.) O. Kuntze) is a very popular beverage and cash crop that is widely cultivated in tropical and subtropical areas. In November 2017, diseased tea plants that exhibiting brown blight disease were observed in Guanxi Township of Hsinchu County in Taiwan. In the plantation,15% of tea trees (about 4000 plants) had an average of 20% of the leaves with at least one lesion. The symptoms began as small, water-soaked lesions on young leaves and twigs and later became larger, dark brown, necrotic lesions of 1 to 3 cm in diameter on leaves and 2 to 5 cm in length on twigs. Symptomatic leaf tissue (1 cm2) from five samples per sample) was surface sterilized with 1% NaClO (from commercial bleach, Clorox) for 1 min, washed with sterilized water 3 times, plated onto potato dextrose agar (PDA), and incubated under 12h/12h cycles of light and darkness at 25°C until sporulation to determine the causal agent. A fungus was consistently isolated from symptomatic leaf samples (80% isolation rate). The fungus initially produced white-to-gray fluffy aerial hyphae, which subsequently exhibited dark pigmentation. Acervuli and setae were absent. The conidia were hyaline, aseptate, smooth-walled, and cylindrical with obtuse to slightly rounded ends, with sizes of 12.10 to 16.02 × 3.58 to 4.91 (average 13.77 × 4.05, n = 30) µm. The majority had two rounded guttules. The appressoria were brown to dark brown, ovoid and slightly obtuse at the tip in shape, had lengths ranging from 3.59 to 10.31 µm (with an average of 7.18 µm, n = 30), and had diameters of 3.14 to 6.43 µm (with an average of 5.10 µm, n = 30). Morphological characteristics matched the descriptions of Colletotrichum fructicola (Liu et al. 2015; Fuentes-Aragón et al. 2018). The internal transcribed spacer of nuclear ribosomal DNA (ITS), actin (ACT), chitin synthase (CHS-1), and Apn2-Mat1-2 intergenic spacer and partial mating-type Mat1-2 gene (ApMAT) sequences of the isolates were obtained to confirm this identification. The sequences showed close identity with those of C. fructicola ex-type cultures ICMP18581 and CBS 130416 (Weir et al. 2012) of 99.65% for the ITS (JX010165), 99.29% for the ACT (JX009501), and 100.00% for the CHS-1 (JX009866), as well as close identity with the other ex-type culture LF506 (Liu et al. 2015) of 99.59% for the ApMat (KJ954567), supporting the isolate's identification as C. fructicola. The sequences were deposited in GenBank, with the following accession Nos.: MN608177 (ITS), MN393175 (ACT), MT087546 (CHS-1), and MT087542 (ApMAT). Based on morphology and DNA sequence analysis, the associated fungus was identified as C. fructicola. Pathogenicity tests were performed next according to the procedures described in Chen et al. (2017). Healthy leaves on tea plants (Ca. sinensis 'Chin-shin Oolong') were wounded by pinpricking in the middle of each counterpart and inoculated with conidial suspension (1 × 107 conidia/ml, 10 µl). Both non-wounded and wounded healthy leaves were inoculated with the conidial suspension and sterile distilled water (a water control). The tea plants were covered with plastic bags to maintain high relative humidity for two days. One week after inoculation, anthracnose was observed on 40% of inoculated leaves, whereas all the control leaves remained healthy. The fungus was re-isolated from the diseased plants, and identified as C. fructicola by resequencing of the four genes. To the best of our knowledge, this is the first report of anthracnose caused by C. fructicola on tea in Taiwan although the pathogen has been present in China and Indonesia (Wang et al. 2016; Shi et al. 2017; Farr and Rossman, 2020).

Construction of Raman spectroscopic fingerprints for the detection of Fusarium wilt of banana in Taiwan.

Banana (Musa sp.) is cultivated worldwide and is one of the most popular fruits. The soil-borne fungal disease Fusarium wilt of banana (FWB), commonly known as Panama disease, is caused by Fusarium oxysporum f. sp. cubense (Foc) and is a highly lethal vascular fungal disease in banana plants. Raman spectroscopy, an emerging laser-based technology based on Raman scattering, has been used for the qualitative characterization of biological tissues such as foodborne pathogens, cancer cells, and melamine. In this study, we describe a Raman spectroscopic technique that could potentially be used as a method for diagnosing FWB. To that end, the Raman fingerprints of Foc (including mycelia and conidia) and Foc-infected banana pseudostems with varying levels of symptoms were determined. Our results showed that eight, eleven, and eleven characteristic surface-enhanced Raman spectroscopy peaks were observed in the mycelia, microconidia, and macroconidia of Foc, respectively. In addition, we constructed the Raman spectroscopic fingerprints of banana pseudostem samples with varying levels of symptoms in order to be able to differentiate Foc-infected bananas from healthy bananas. The rate at which FWB was detected in asymptomatic Foc-infected samples by using the spectral method was 76.2%, which was comparable to the rates previously reported for other FWB detection methods based on real-time PCR assays, suggesting that the spectral method described herein could potentially serve as an alternative tool for detecting FWB in fields. As such, we hope that the developed spectral method will open up new possibilities for the on-site diagnosis of FWB.

Molecular Detection Assays for Rapid Field-Detection of Rice Sheath Blight.

Rhizoctonia solani (Rs), a soil-borne fungal pathogen, can result in rice sheath blight (ShB), which causes yield loss. To prevent outbreaks of ShB and enhance the sustainability of rice production, it is critical to develop a rapid ShB detection method for specific, fast, and on-site disease management. In this study, a reagent for the rapid extraction of this pathogen was developed for on-site detection. The specificity and sensitivity of a novel SMS RS1-F/SMS RS1-R primer set and a ITS1/GMRS-3 reference primer set were tested, while four different extraction protocols for ShB were developed. Moreover, intraday and interday assays were performed to evaluate the reproducibility of the detection methods developed. The results indicated that all of the developed protocols are suitable for use in detecting ShB. In addition, all the samples of infected rice yielded positive Rs detection results when subjected to TaqMan probe-based real-time PCR and SYBR green-based real-time PCR (SMS RS1-F/SMS RS1-R) tests in which automatic magnetic bead-based DNA extraction was performed. These results indicated that the two molecular detection protocols were suitable for the field diagnosis of ShB for all asymptomatic and symptomatic rice samples.