Analysis of the cell surface layer ultrastructure of the oral pathogen Tannerella forsythia

The Gram-negative oral pathogen Tannerella forsythia is decorated with a 2D crystalline surface (S-) layer, with two different S-layer glycoprotein species being present. Prompted by the predicted virulence potential of the S-layer, this study focused on the analysis of the arrangement of the individual S-layer glycoproteins by a combination of microscopic, genetic, and biochemical analyses. The two S-layer genes are transcribed into mRNA and expressed into protein in equal amounts. The S-layer was investigated on intact bacterial cells by transmission electron microscopy, by immune fluorescence microscopy, and by atomic force microscopy. The analyses of wild-type cells revealed a distinct square S-layer lattice with an overall lattice constant of 10.1 ± 0.7 nm. In contrast, a blurred lattice with a lattice constant of 9.0 nm was found on S-layer single-mutant cells. This together with in vitro self-assembly studies using purified (glyco)protein species indicated their increased structural flexibility after self-assembly and/or impaired self-assembly capability. In conjunction with TEM analyses of thin-sectioned cells, this study demonstrates the unusual case that two S-layer glycoproteins are co-assembled into a single S-layer. Additionally, flagella and pilus-like structures were observed on T. forsythia cells, which might impact the pathogenicity of this bacterium

Reducing the environmental impact of surgery on a global scale: systematic review and co-prioritization with healthcare workers in 132 countries

Abstract Background Healthcare cannot achieve net-zero carbon without addressing operating theatres. The aim of this study was to prioritize feasible interventions to reduce the environmental impact of operating theatres. Methods This study adopted a four-phase Delphi consensus co-prioritization methodology. In phase 1, a systematic review of published interventions and global consultation of perioperative healthcare professionals were used to longlist interventions. In phase 2, iterative thematic analysis consolidated comparable interventions into a shortlist. In phase 3, the shortlist was co-prioritized based on patient and clinician views on acceptability, feasibility, and safety. In phase 4, ranked lists of interventions were presented by their relevance to high-income countries and low–middle-income countries. Results In phase 1, 43 interventions were identified, which had low uptake in practice according to 3042 professionals globally. In phase 2, a shortlist of 15 intervention domains was generated. In phase 3, interventions were deemed acceptable for more than 90 per cent of patients except for reducing general anaesthesia (84 per cent) and re-sterilization of ‘single-use’ consumables (86 per cent). In phase 4, the top three shortlisted interventions for high-income countries were: introducing recycling; reducing use of anaesthetic gases; and appropriate clinical waste processing. In phase 4, the top three shortlisted interventions for low–middle-income countries were: introducing reusable surgical devices; reducing use of consumables; and reducing the use of general anaesthesia. Conclusion This is a step toward environmentally sustainable operating environments with actionable interventions applicable to both high– and low–middle–income countries

CorneaNet : fast segmentation of cornea OCT scans of healthy and keratoconic eyes using deep learning

Deep learning has dramatically improved object recognition, speech recognition, medical image analysis and many other fields. Optical coherence tomography (OCT) has become a standard of care imaging modality for ophthalmology. We asked whether deep learning could be used to segment cornea OCT images. Using a custom-built ultrahigh-resolution OCT system, we scanned 72 healthy eyes and 70 keratoconic eyes. In total, 20,160 images were labeled and used for the training in a supervised learning approach. A custom neural network architecture called CorneaNet was designed and trained. Our results show that CorneaNet is able to segment both healthy and keratoconus images with high accuracy (validation accuracy: 99.56%). Thickness maps of the three main corneal layers (epithelium, Bowman’s layer and stroma) were generated both in healthy subjects and subjects suffering from keratoconus. CorneaNet is more than 50 times faster than our previous algorithm. Our results show that deep learning algorithm scan be used for OCT image segmentation and could be applied in various clinical settings. In particular, CorneaNet could be used for early detection of keratoconus and more generally to study other diseases altering corneal morphology.Published versio

Latissimus Dorsi Myocutaneous Flap Procedure in a Swine Model

Background As surgical research expands in both breadth and scope, translational models become increasingly important. The accessibility, reproducibility, and clinical applicability of translational models is of vital importance to ensure adequate and accurate research. Though different flap models have been described, the literature lacks an in-depth, technical description of an easy large-animal preclinical model. We here describe the procedure for elevation of a latissimus dorsi flap in a swine. This flap contains muscle and skin that can be isolated on a vascular pedicle, transferred as a free flap, perfused, or innervated/denervated as dictated by the needs of the experiment. Methods Five different latissimus dorsi flaps were elevated in miniature swine. Careful attention was paid to anatomical landmarks and optimal placement of incision, dissection, and retraction. Temporary ischemia with vascular clamping was performed along with serial digital and infrared imaging both intra- and postoperatively. In three of the flaps with induced ischemia, the animal was observed for a 30-day follow up with daily photodocumentation and intermittent biopsy. Results A reproducible latissimus flap model was designed with optimized conditions. In the animals in which flaps were followed postoperatively, complete healing was seen within 30 days without evidence of procedure-related ischemia or loss of motor function. Conclusion We have identified and described a pre-clinical large animal flap model that can be easily reproduced for translational studies of multiple scientific areas including flap-based repair, ischemia, ischemia reperfusion, and operative technique. This provides an important model for ready replication in preclinical studies of many varieties

Gluconate-Lactobionate-Dextran Perfusion Solutions Attenuate Ischemic Injury and Improve Function in a Murine Cardiac Transplant Model

Static cold storage is the cheapest and easiest method and current gold standard to store and preserve donor organs. This study aimed to compare the preservative capacity of gluconate-lactobionate-dextran (Unisol) solutions to histidine-tryptophan-ketoglutarate (HTK) solution. Murine syngeneic heterotopic heart transplantations (Balb/c-Balb/c) were carried out after 18 h of static cold storage. Cardiac grafts were either flushed and stored with Unisol-based solutions with high-(UHK) and low-potassium (ULK) ± glutathione, or HTK. Cardiac grafts were assessed for rebeating and functionality, histomorphologic alterations, and cytokine expression. Unisol-based solutions demonstrated a faster rebeating time (UHK 56 s, UHK + Glut 44 s, ULK 45 s, ULK + Glut 47 s) compared to HTK (119.5 s) along with a better contractility early after reperfusion and at the endpoint on POD 3. Ischemic injury led to a significantly increased leukocyte recruitment, with similar degrees of tissue damage and inflammatory infiltrate in all groups, yet the number of apoptotic cells tended to be lower in ULK compared to HTK. In UHK- and ULK-treated animals, a trend toward decreased expression of proinflammatory markers was seen when compared to HTK. Unisol-based solutions showed an improved preservative capacity compared with the gold standard HTK early after cardiac transplantation. Supplemented glutathione did not further improve tissue-protective properties