Fast, Easy, and Comprehensive Techniques for Microscopic Observations of Fungal and Oomycete Organisms Inside the Roots of Herbaceous and Woody Plants.

The roots of herbaceous and woody plants growing in soil are complex structures that are affected by both natural and artificial fungal colonization to various extents. To obtain comprehensive information about the overall distribution of fungi or oomycetes inside a plant root system, rapid, effective, and reliable screening methods are required. To observe both fine roots, i.e., a common site for penetration of fungi and oomycetes, and mature roots, different techniques are required to overcome visual barriers, such as root browning or tissue thickening. In our protocol, we propose using fast, cost-effective, and non-harmful methods to localize fungal or oomycete structures inside plant roots. Root staining with a fluorescent dye provides a quick initial indication of the presence of fungal structures on the root surfaces. The protocol is followed by clearing and staining steps, resulting in a deeper insight into the root tissue positioning, abundance, and characteristic morphological/reproductive features of fungal or oomycete organisms. If required, the stained samples can be prepared by using freeze-drying for further observations, including advanced microscopic techniques. Key features • The protocol enhances tissue-clearing techniques employing KOH or NaOH and is applicable to a broad range of roots from different plant species. • Hydroxides are mixed with hydrogen peroxide to obtain an efficient bleaching solution, which effectively clears roots without causing significant tissue damage. • The protocol could also be used for staining of fungi or oomycetes localized both on the root surface or inside the root tissues. • Simple combination of non-fluorescent methyl blue and fluorescent solophenyl flavine dyes allows the observation of fungal organisms in both brightfield and fluorescence microscopy.

The fungus Acremonium alternatum enhances salt stress tolerance by regulating host redox homeostasis and phytohormone signaling.

While endophytic fungi offer promising avenues for bolstering plant resilience against abiotic stressors, the molecular mechanisms behind this biofortification remain largely unknown. This study employed a multifaceted approach, combining plant physiology, proteomic, metabolomic, and targeted hormonal analyses to illuminate the early response of Brassica napus to Acremonium alternatum during the nascent stages of their interaction. Notably, under optimal growth conditions, the initial reaction to fungus was relatively subtle, with no visible alterations in plant phenotype and only minor impacts on the proteome and metabolome. Interestingly, the identified proteins associated with the Acremonium response included TUDOR 1, Annexin D4, and a plastidic K+ efflux antiporter, hinting at potential processes that could counter abiotic stressors, particularly salt stress. Subsequent experiments validated this hypothesis, showcasing significantly enhanced growth in Acremonium-inoculated plants under salt stress. Molecular analyses revealed a profound impact on the plant's proteome, with over 50% of salt stress response proteins remaining unaffected in inoculated plants. Acremonium modulated ribosomal proteins, increased abundance of photosynthetic proteins, enhanced ROS metabolism, accumulation of V-ATPase, altered abundances of various metabolic enzymes, and possibly promoted abscisic acid signaling. Subsequent analyses validated the accumulation of this hormone and its enhanced signaling. Collectively, these findings indicate that Acremonium promotes salt tolerance by orchestrating abscisic acid signaling, priming the plant's antioxidant system, as evidenced by the accumulation of ROS-scavenging metabolites and alterations in ROS metabolism, leading to lowered ROS levels and enhanced photosynthesis. Additionally, it modulates ion sequestration through V-ATPase accumulation, potentially contributing to the observed decrease in chloride content.

Leaflet modification before transcatheter aortic valve implantation in patients at risk for coronary obstruction: the ShortCut study.

BACKGROUND AND AIMS: This trial sought to assess the safety and efficacy of ShortCut, the first dedicated leaflet modification device, prior to transcatheter aortic valve implantation (TAVI) in patients at risk for coronary artery obstruction. METHODS: This pivotal prospective study enrolled patients with failed bioprosthetic aortic valves scheduled to undergo TAVI and were at risk for coronary artery obstruction. The primary safety endpoint was procedure-related mortality or stroke at discharge or 7 days, and the primary efficacy endpoint was per-patient leaflet splitting success. Independent angiographic, echocardiographic, and computed tomography core laboratories assessed all images. Safety events were adjudicated by a clinical events committee and data safety monitoring board. RESULTS: Sixty eligible patients were treated (77.0 ± 9.6 years, 70% female, 96.7% failed surgical bioprosthetic valves, 63.3% single splitting and 36.7% dual splitting) at 22 clinical sites. Successful leaflet splitting was achieved in all (100%; 95% confidence interval [CI] 94-100.0%, p<0.001) patients. Procedure time, including imaging confirmation of leaflet splitting, was 30.6 ± 17.9 min. Freedom from the primary safety endpoint was achieved in 59 (98.3%; 95% CI [91.1-100%]) patients, with no mortality and one (1.7%) disabling stroke. At 30 days, freedom from coronary obstruction was 95% (95% CI 86.1-99.0%). Within 90 days, freedom from mortality was 95% (95% CI 86.1-99.0%]), without any cardiovascular deaths. CONCLUSIONS: Modification of failed bioprosthetic aortic valve leaflets using ShortCut was safe, achieved successful leaflet splitting in all patients, and was associated with favorable clinical outcomes in patients at risk for coronary obstruction undergoing TAVI.

Safety and feasibility of early discharge after transcatheter aortic valve implantation with ACURATE Neo-the POLESTAR trial.

BACKGROUND: Transcatheter aortic valve implantation (TAVI) serves a growing range of patients with severe aortic stenosis (AS). TAVI has evolved to a streamlined procedure minimizing length of hospital stay. AIMS: To evaluate the safety and efficacy of an early discharge (ED) strategy after TAVI. METHODS: We performed an international, multi-center, prospective observational single-arm study in AS patients undergoing TAVI with the ACURATE valve platform. Eligibility for ED was assessed prior to TAVI and based on prespecified selection criteria. Discharge ≤ 48 h was defined as ED. Primary Valve Academic Research Consortium (VARC)-3-defined 30-day safety and efficacy composite endpoints were landmarked at 48 h and compared between ED and non-ED groups. RESULTS: A total of 252 patients were included. The median age was 82 [25th-75th percentile, 78-85] years and the median Society of Thoracic Surgeons Predicted Risk of Mortality (STS-PROM) score was 2.2% [25th-75th percentile, 1.6-3.3]. ED and non-ED were achieved in 173 (69%) and 79 (31%) patients respectively. Monitoring for conduction disturbances was the principal reason for non-ED (33%). Overall, at 30 days, all-cause mortality was 1%, new permanent pacemaker rate was 4%, and valve- or procedure-related rehospitalization was 4%. There was no difference in the primary safety and efficacy endpoint between the ED and non-ED cohorts (OR 0.84 [25th-75th percentile, 0.31-2.26], p = 0.73, and OR 0.97 [25th-75th percentile, 0.46-2.06], p = 0.94). The need for rehospitalization was similarly low for ED and non-ED groups. CONCLUSION: Early discharge after TAVI with the ACURATE valve is safe and feasible in selected patients. Rhythm monitoring and extended clinical observation protracted hospital stay.

Left Atrial Appendage Occlusion: British Cardiovascular Intervention Society and British Heart Rhythm Society Position Statement.

Percutaneous left atrial appendage occlusion aims to reduce the risk of stroke in patients with AF, particularly those who are not good candidates for systemic anticoagulation. The procedure has been studied in large international randomised trials and registries and was approved by the National Institute for Health and Care Excellence in 2014 and by NHS England in 2018. This position statement summarises the evidence for left atrial appendage occlusion and presents the current indications. The options and consensus on best practice for pre-procedure planning, undertaking a safe and effective implant and appropriate post-procedure management and follow-up are described. Standards regarding procedure volume for implant centres and physicians, the role of multidisciplinary teams and audits are highlighted.

Coronary artery disease and outcomes following transcatheter aortic valve implantation.

BACKGROUND: Aortic stenosis is a life-limiting condition for which transcatheter aortic valve implantation (TAVI) is an established therapy. Coronary artery disease (CAD) is frequently found in this patient group and optimal management in these patients remains uncertain. OBJECTIVES: We sought to examine the association of coexistent CAD on mortality and hospital readmission in patients undergoing TAVI. METHODS: In this observational cohort study, we examined patients who underwent TAVI and segregated them by the presence of obstructive epicardial CAD. The primary outcome was 3-year mortality with secondary outcomes being readmission for (1) all-causes, (2) a MACE (Major Adverse Cardiovascular Event) composite endpoint and (3) acute coronary syndrome. Subsidiary outcomes included patient angina and breathlessness scores. RESULTS: 898 patients underwent TAVI, of which 488 (54.3%) had unobstructed coronary arteries and 410 (45.7%) had obstructive CAD. Overall, n=298 (33.2%) patients experienced the primary mortality endpoint with no significant difference when stratified according to CAD (n=160 (32.9%) vs n=136 (33.2%), HR 0.98, CI 0.78 to 1.24). After multivariate analysis, the presence of CAD had no effect on the primary outcome (HR 0.98, CI 0.68 to 1.40). There was no significant difference in readmission for any cause (n=181, 37.1% (CAD) vs n=169, 41.2% (no CAD), p=0.23), including no significant difference on readmission for MACE (n=48, 9.8% (CAD) vs n=45, 11.0% (no CAD), p=0.11). CAD at the time of TAVI also did not alter breathlessness or angina scores before/after TAVI (p>0.05). CONCLUSION: Coexistent CAD had no significant association with mortality, any-cause readmission or symptoms for patients undergoing TAVI in our cohort.

Left Atrial Appendage Occlusion: British Cardiovascular Intervention Society and British Heart Rhythm Society Position Statement.

Percutaneous left atrial appendage occlusion aims to reduce the risk of stroke in patients with AF, particularly those who are not good candidates for systemic anticoagulation. The procedure has been studied in large international randomised trials and registries and was approved by the National Institute for Health and Care Excellence in 2014 and by NHS England in 2018. This position statement summarises the evidence for left atrial appendage occlusion and presents the current indications. The options and consensus on best practice for pre-procedure planning, undertaking a safe and effective implant and appropriate post-procedure management and follow-up are described. Standards regarding procedure volume for implant centres and physicians, the role of multidisciplinary teams and audits are highlighted.

Practical guide on left atrial appendage closure for the non-implanting physician: an international consensus paper.

A significant proportion of patients who suffer from atrial fibrillation (AF) and are in need of thromboembolic protection are not treated with oral anticoagulation or discontinue this treatment shortly after its initiation. This undertreatment has not improved sufficiently despite the availability of direct oral anticoagulants which are associated with less major bleeding than vitamin K antagonists. Multiple reasons account for this, including bleeding events or ischaemic strokes whilst on anticoagulation, a serious risk of bleeding events, poor treatment compliance despite best educational attempts, or aversion to drug therapy. An alternative interventional therapy, which is not associated with long-term bleeding and is as effective as vitamin K anticoagulation, was introduced over 20 years ago. Because of significant improvements in procedural safety over the years, left atrial appendage closure, predominantly achieved using a catheter-based, device implantation approach, is increasingly favoured for the prevention of thromboembolic events in patients who cannot achieve effective anticoagulation. This management strategy is well known to the interventional cardiologist/electrophysiologist but is not more widely appreciated within cardiology or internal medicine. This article introduces the devices and briefly explains the implantation technique. The indications and device follow-up are more comprehensively described. Almost all physicians who care for adult patients will have many with AF. This practical guide, written within guideline/guidance boundaries, is aimed at those non-implanting physicians who may need to refer patients for consideration of this new therapy, which is becoming increasingly popular.

Bioadhesive and Injectable Hydrogels and Their Correlation with Mesenchymal Stem Cells Differentiation for Cartilage Repair: A Mini-Review.

Injectable bioadhesive hydrogels, known for their capacity to carry substances and adaptability in processing, offer great potential across various biomedical applications. They are especially promising in minimally invasive stem cell-based therapies for treating cartilage damage. This approach harnesses readily available mesenchymal stem cells (MSCs) to differentiate into chondrocytes for cartilage regeneration. In this review, we investigate the relationship between bioadhesion and MSC differentiation. We summarize the fundamental principles of bioadhesion and discuss recent trends in bioadhesive hydrogels. Furthermore, we highlight their specific applications in conjunction with stem cells, particularly in the context of cartilage repair. The review also encompasses a discussion on testing methods for bioadhesive hydrogels and direct techniques for differentiating MSCs into hyaline cartilage chondrocytes. These approaches are explored within both clinical and laboratory settings, including the use of genetic tools. While this review offers valuable insights into the interconnected aspects of these topics, it underscores the need for further research to fully grasp the complexities of their relationship.

Seminal Bacterioflora of Two Rooster Lines: Characterization, Antibiotic Resistance Patterns and Possible Impact on Semen Quality.

This study aimed to characterize the bacterial profiles and their association with selected semen quality traits among two chicken breeds. Thirty Lohmann Brown and thirty ROSS 308 roosters were selected for semen quality estimation, including sperm motility, membrane and acrosome integrity, mitochondrial activity, and DNA fragmentation. The oxidative profile of the semen, including the production of reactive oxygen species (ROS), antioxidant capacity, protein, and lipid oxidation, were assessed as well. Moreover, the levels of pro-inflammatory cytokines, including tumor necrosis factor alpha (TNF-α), interleukins 1 and 6 (IL-1, IL-6) and C-reactive protein, as well as the concentrations of selected antibacterial proteins (cathelicidin, ß-defensin and lysozyme) in the seminal plasma were evaluated with the enzyme-linked immunosorbent assay. The prevailing bacterial genera identified by the matrix-assisted laser desorption/ionization time-of-flight mass spectrometry were Citrobacter spp., Enterococcus spp., Escherichia spp. and Staphylococcus spp. While the bacterial load was significantly higher in the ROSS 308 line (p < 0.05), a higher number of potentially uropathogenic bacteria was found in the Lohmann Brown roosters. Antimicrobial susceptibility tests revealed a substantial resistance of randomly selected bacterial strains, particularly to ampicillin, tetracycline, chloramphenicol, and tobramycin. Furthermore, Lohmann Brown ejaculates containing an increased proportion of Escherichia coli presented with significantly (p < 0.05) elevated levels of TNF-α and IL-6, as well as ROS overproduction and lipid peroxidation. Inversely, significantly (p < 0.05) higher levels of ß-defensin and lysozyme were found in the semen collected from the ROSS 308 roosters, which was characterized by a higher quality in comparison to the Lohmann Brown roosters. In conclusion, we emphasize the criticality of bacteriospermia in the poultry industry and highlight the need to include a more complex microbiological screening of semen samples designated for artificial insemination.

Endophytic yeast protect plants against metal toxicity by inhibiting plant metal uptake through an ethylene-dependent mechanism.

Toxic metal pollution requires significant adjustments in plant metabolism. Here, we show that the plant microbiota plays an important role in this process. The endophytic Sporobolomyces ruberrimus isolated from a serpentine population of Arabidopsis arenosa protected plants against excess metals. Coculture with its native host and Arabidopsis thaliana inhibited Fe and Ni uptake. It had no effect on host Zn and Cd uptake. Fe uptake inhibition was confirmed in wheat and rape. Our investigations show that, for the metal inhibitory effect, the interference of microorganisms in plant ethylene homeostasis is necessary. Application of an ethylene synthesis inhibitor, as well as loss-of-function mutations in canonical ethylene signalling genes, prevented metal uptake inhibition by the fungus. Coculture with S. ruberrimus significantly changed the expression of Fe homeostasis genes: IRT1, OPT3, OPT6, bHLH38 and bHLH39 in wild-type (WT) A. thaliana. The expression pattern of these genes in WT plants and in the ethylene signalling defective mutants significantly differed and coincided with the plant accumulation phenotype. Most notably, down-regulation of the expression of IRT1 solely in WT was necessary for the inhibition of metal uptake in plants. This study shows that microorganisms optimize plant Fe and Ni uptake by fine-tuning plant metal homeostasis.

The mechanism of enhanced lignin regulating foliar Cd absorption and yield in rice (Oryza sativa L.).

The impact of atmospheric deposition of cadmium (Cd) in cereal crops has become a global concern. Enhanced lignin content was expected to benefit the plant performance against Cd exposure. To date, however, the underlying mechanisms of lignin regulating foliar Cd absorption in rice (Oryza sativa L.) and its effect on grain yield remains unclear. In present study, the effect and mechanism of rice in response to leaf Cd exposure were investigated using 113Cd stable isotope and a lignin-increased rice mutant. The highest Cd uptake efficiency and uptake amount was observed in wild type (WT) plant grown in the maturity period, which were 3-fold higher than in mutant plant. Compared to WT, the mutant exhibited 14.75% and 25.43% higher contents in G- and S-unit of lignin monomers. Lignin biosynthesis and polymerization related genes (OsPAL/OsCOMT/Os4CL3/OsLAC5/OsLAC15) were significantly up-regulated in mutants. In addition, the enzyme activities involved in the above process were also significantly increased by 1.24-1.49-fold. The increased Cd retention in cell wall and decreased gene expression levels of OsNRAMP5, OsHMA3 and OsIRT1 in mutant indicated that lignin effectively inhibited Cd transportion in plant tissues. Moreover, the antioxidant capacity and photosynthesis efficiency in mutant plant were obviously improved, leading to higher Cd tolerance and increased grain yield. Our results revealed the molecular and physiological mechanisms of enhanced lignin regulating foliar Cd absorption and yield in rice, and provided the valuable rice genotype to ensure food safety.

Quercetin Ameliorates Testicular Damage in Zucker Diabetic Fatty Rats through Its Antioxidant, Anti-Inflammatory and Anti-Apoptotic Properties.

The aim of this study was to investigate the effects of quercetin (QUE) on the testicular architecture as well as markers of oxidative, inflammatory, and apoptotic profile of male gonads in Zucker diabetic fatty (ZDF) rats suffering from Type 2 diabetes mellitus in the absence or presence of obesity. QUE was administered orally at a dose of 20 mg/kg/day for 6 weeks. Morphometric analysis revealed that QUE treatment led to an improvement in testicular appearance, particularly in the case of Obese ZDF rats. Furthermore, a significant stabilization of the antioxidant capacity (p < 0.05), superoxide dismutase and catalase activity (p < 0.01), with a concomitant decrease in lipid peroxidation (p < 0.05) were observed in Obese ZDF animals exposed to QUE. Our data also indicate a significant decline in the levels of interleukin (IL)-1 (p < 0.05), IL-6 (p < 0.01) and tumor necrosis factor alpha (p < 0.001) following QUE supplementation to Obese ZDF rats in comparison with their respective control. Finally, a significant down-regulation of the pro-apoptotic BAX protein (p < 0.0001) was observed in Obese ZDF rats administered with QUE, while a significant Bcl-2 protein overexpression (p < 0.0001) was recorded in Lean ZDF animals when compared to their untreated control. As such, our results suggest that QUE is a potentially beneficial agent to reduce testicular damage in ZDF rats with Type 2 diabetes mellitus by decreasing oxidative stress, chronic inflammation, and excessive cell loss through apoptosis.

Characterization of the structural, oxidative, and immunological features of testis tissue from Zucker diabetic fatty rats.

The purpose of this study was to characterize the testicular profile of Zucker diabetic fatty (ZDF) rats presenting with type 2 diabetes mellitus (DM2) in the absence or presence of obesity. To achieve this, testes were collected from 270-day-old male Wistar (n = 15), ZDF nonobese (n = 15), and ZDF obese rats (n = 16). Changes to the testicular structure were quantified morphometrically, while immunocytochemistry was employed to assess caspase-3 activity. Reactive oxygen species (ROS) production, fluctuations of major antioxidant molecules, and the extent of damage to the proteins and lipids were assessed in tissue lysates. Levels of selected interleukins (ILs) were determined by enzyme-linked immunosorbent assay. The results reveal significant alterations to the testicular structure accompanied by caspase-3 overexpression, particularly in ZDF obese rats. The most notable disruption of the oxidative balance, characterized by ROS overproduction, antioxidant deficiency, protein, and lipid deterioration was recorded in ZDF rats suffering from both DM2 and obesity. Accordingly, the highest concentrations of pro-inflammatory IL-1, IL-6, and IL-18 accompanied by reduced levels of the anti-inflammatory IL-10 were found in testicular tissue collected from ZDF obese rats. This study highlights the vulnerability of male gonads to pathophysiological changes caused by hyperglycemia, which are further exacerbated by excessive adipose tissue.

The consumption of sea buckthorn (Hippophae rhamnoides L.) effectively alleviates type 2 diabetes symptoms in spontaneous diabetic rats.

Sea buckthorn (Hippophae rhamnoides L.) is described by various beneficial effects as it contains several bioactive substances characterized by antioxidant effects. These effects are closely related to the reduction of oxidative stress that is involved in the development of the disease. One such diseases is Diabetes mellitus, the prevalence of which is growing and is associated primarily with diet, lack of exercise and/or genetics. This study intends to examine the effects of sea buckthorn and metformin on body weight, water and feed intake, glycaemia, insulinemia, sorbitol accumulation and cataract development in Zucker diabetic fatty rats, which represent an animal model of type 2 Diabetes mellitus, as well as to characterize the individual content of bioactive substances and the antioxidant activity of sea buckthorn. Particular concentrations were applied (500 and 1000 mg.kg-1 body weight of sea buckthorn, and combinations with 150 mg.kg-1 body weight of metformin) by gastric gavage. The total antioxidant capacity and bioactive compounds were determined by spectrophotometric analysis. The best results of the study showed suppression of hyperglycaemia, water intake, decreased sorbitol levels in the lens of the eyes after sea buckthorn treatment. Determination of bioactive compounds showed significantly higher values in dry berries when compared to fresh berries of sea buckthorn and high total antioxidant capacity. Our results represent an interest in sea buckthorn and its potential use in the treatment of Diabetes mellitus as well as other experimental studies.

The effects of caffeine on the motility and viability of stallion spermatozoa at different temperature conditions.

The purpose of this study was to evaluate the dose- and time-dependent effect of caffeine treatment on the motility and viability of stallion spermatozoa at different temperatures. Six dose groups (A to F) were established with changing caffeine concentrations (from 0.625 to 10 mg/mL). The control samples were prepared by diluting the ejaculate only with physiological salt solution. The samples were examined after 0, 1, 2 and 3 h of incubation at 5 °C and 37 °C. The motility parameters were evaluated by Computer Assisted Semen Analyzer (CASA) system, and the viability was assessed by the mitochondrial toxicity test at the end of the incubation. A positive effect of the lowest tested caffeine concentration on the motility parameters was observed throughout the incubation period at 5 °C. At the end of the 3h incubation, the viability in every sample in these groups, treated with any caffeine concentration, showed lower values compared to the control. At the higher incubation temperature (37 °C), caffeine positively affected the motility in samples B (P < 0.05) and D, E, F (P < 0.001) after 3 h of incubation; however, the viability showed a slightly decreasing tendency. Our results suggest that caffeine, in an optimal concentration, may be used as a component of stallion semen extenders.

Can Interventional Cardiologists Help Deliver the UK Mechanical Thrombectomy Interventional Programme for Patients with Acute Ischaemic Stroke? A Discussion Paper from the British Cardiovascular Interventional Society Stroke Thrombectomy Focus Group.

There is a willingness among UK interventional cardiologists to contribute to provision of a 24/7 mechanical thrombectomy (MT) service for all suitable stroke patients if given the appropriate training. This highly effective intervention remains unavailable to the majority of patients who might benefit, partly because there is a limited number of trained specialists. As demonstrated in other countries, interdisciplinary working can be the solution and an opportunity to achieve this is outlined in this article.