Electrical signalling in prokaryotes and its convergence with quorum sensing in Bacillus.

The importance of electrical signalling in bacteria is an emerging paradigm. Bacillus subtilis biofilms exhibit electrical communication that regulates metabolic activity and biofilm growth. Starving cells initiate oscillatory extracellular potassium signals that help even the distribution of nutrients within the biofilm and thus help regulate biofilm development. Quorum sensing also regulates biofilm growth and crucially there is convergence between electrical and quorum sensing signalling axes. This makes B. subtilis an interesting model for cell signalling research. SpoOF is predicted to act as a logic gate for signalling pathway convergence, raising interesting questions about the functional nature of this gate and the relative importance of these disparate signals on biofilm behaviour. How is an oscillating signal integrated with a quorum signal? The model presented offers rich opportunities for future experimental and theoretical modelling research. The importance of direct cell-to-cell electrical signalling in prokaryotes, so characteristic of multicellular eukaryotes, is also discussed.

Recommendation of a practical guideline for safe tracheostomy during the COVID-19 pandemic.

PURPOSE: The COVID-19 pandemic is placing unprecedented demand upon critical care services for invasive mechanical ventilation. There is current uncertainty regarding the role of tracheostomy for weaning ventilated patients with COVID-19 pneumonia. This is due to a number of factors including prognosis, optimal healthcare resource utilisation, and safety of healthcare workers when performing such a high-risk aerosol-generating procedure. METHODS: Literature review and proposed practical guideline based on the experience of a tertiary healthcare institution with 195 critical care admissions for COVID-19 up until 4th April 2020. RESULTS: A synthesis of the current international literature and reported experience is presented with respect to prognosis, viral load and staff safety, thus leading to a pragmatic recommendation that tracheostomy is not performed until at least 14 days after endotracheal intubation in COVID-19 patients. Practical steps to minimise aerosol generation in percutaneous tracheostomy are outlined and we describe the process and framework for setting up a dedicated tracheostomy team. CONCLUSION: In selected COVID-19 patients, there is a role for tracheostomy to aid in weaning and optimise healthcare resource utilisation. Both percutaneous and open techniques can be performed safely with careful modifications to technique and appropriate enhanced personal protective equipment. ORL-HNS surgeons can play a valuable role in forming tracheostomy teams to support critical care teams during this global pandemic.

Current Status of Simulation in Otolaryngology: A Systematic Review.

OBJECTIVE: Otolaryngology is a highly technical and demanding specialty and the requirements for surgical trainees to acquire proficiency remains challenging. Simulation has been purported to be an effective tool in assisting with this. The aim of this systematic review is to identify the available otolaryngology simulators, their status of validation, and evaluation the level of evidence behind each training model and thereby establish a level of recommendation. DESIGN: PubMed, ERIC, and Google Scholar databases were searched for articles that described otolaryngology simulators or training models between 1980 and April 2016. Any validation studies for simulators were also retrieved. Titles and abstracts were screened for relevance using the preferred reporting items for systematic reviews and meta-analysis (PRISMA) guidelines. Level of evidence (LoE) and Level of recommendation (LoR) was awarded to each study and model, respectively. RESULTS: A total of 70 studies were identified describing 64 simulators. Out of these, at least 54 simulators had 1 validation study. Simulators for the ear and temporal bone surgery were the most common (n = 32), followed by laryngeal and throat (n = 20) and endoscopic sinus surgery (n = 12). Face validity was evaluated by 29 studies, 20 attempted to show construct, 20 assessed content, 20 transfer, and only 2 assessed concurrent validity. Of the validation assessments, 2 were classified as Level 1b, 10 Level 2a, and 48 Level 2b. No simulators received the highest LoR, but 8 simulators received a LoR of 2. CONCLUSIONS: Despite the lack of evidence in outcome studies and limited number of high-validity otolaryngology simulators, the role of simulation continues to grow across surgical specialties Hence, it is imperative that the simulators are of high validity and construct for trainees to practice and rehearse surgical skills to develop confidence.

Current practice in management of the neck after chemoradiotherapy for patients with locally advanced oropharyngeal squamous cell carcinoma.

Patients whose necks respond completely to chemoradiation are unlikely to have residual viable tumour, which questions the need for planned neck dissection. Partial responders often need further assessment. Positron emission tomography/computed tomography (PET/CT) is becoming the standard method of assessing the response of both the primary site and neck to chemoradiation. There is debate, however, about the timing of assessment, the best imaging technique, and the extent of neck dissection, and emerging evidence supports more selective procedures with their attendant reductions in morbidity. Various trials have tried to settle these controversies, but we hypothesised that current practice varies across the United Kingdom (UK), so we set out to establish what it is. A total of 219 questionnaires were sent to head and neck surgeons of varying disciplines and their oncology counterparts, which outlined a clinical picture of a patient with persistent nodal disease after chemoradiotherapy, and requested information about the respondents' preferred choice and timing of investigations in addition to the type of neck dissection, if indicated. There were noticeable variations in practice, with a tendency towards personal choice rather than a multidisciplinary approach. Although there were some items of broad agreement, there was disparity about the timing of imaging and operation. There is inconsistency in the management of the neck in these patients in the UK, which may reflect an absence of guidelines and paucity of evidence-based information. We need to unify practice to improve the care of patients.

Squamous cell carcinoma of the tonsillar remnant--clinical presentation and oncological outcome.

BACKGROUND: Squamous cell carcinoma (SCC) of the tonsil is the most common malignant tumour of the oropharynx. Paediatric tonsillectomy is one of the most commonly performed procedures in Otorhinolaryngology. SCC of the tonsil remnant (SCCTR) in a previously tonsillectomised patient is rare. METHODS: Retrospective review of patients with SCCTR presenting to the Otorhinolaryngology, Head and Neck Unit January 2000 to December 2007. RESULTS: Two hundred and fifty patients with tonsil SCC were identified. Ten (4%) of these had previously undergone tonsillectomy in childhood. Nine patients underwent radical treatment including surgery, radiotherapy and in four cases concomitant chemotherapy. Eight patients are alive with no signs of recurrence with follow-up of a minimum of 24 months. One has been lost from follow-up. CONCLUSIONS: Clinicians should be aware that SCC can arise from a tonsillar remnant. SCCTR has similar oncological outcomes as tonsillar tumours.

Expectant management of vestibular schwannoma: a retrospective multivariate analysis of tumor growth and outcome.

We conducted a retrospective observational study to assess the consequences of conservative management of vestibular schwannoma (VS). Data were collected from tertiary neuro-otological referral units in United Kingdom. The study included 59 patients who were managed conservatively with radiological diagnosis of VS. The main outcome measures were growth rate and rate of failure of conservative management. Multivariate analysis sought correlation between tumor growth and (i) demographic features, (ii) tumor characteristics. The mean tumor growth was 0.66 mm/y. 11 patients (19%) required intervention. Mean time to intervention was 37 months with two notable late "failures" occurring at 75 and 84 months. Tumors extending into the cerebellopontine angle (CPA) grew significantly faster than intracanalicular tumors (p = 0.0045). No association was found between growth rate and age, sex, tumor laterality, facial nerve function, and grade of hearing loss. Conservative management is acceptable for a subset of patients. Tumors extending into the CPA at diagnosis grow significantly faster than intracanalicular tumors. No growth within 5 years of surveillance does not guarantee a continued indolent growth pattern; surveillance must therefore continue.

Good correlation between original and modified House Brackmann facial grading systems.

OBJECTIVES/HYPOTHESIS: Subjective scales of facial function were plagued with reporting variations until the House-Brackmann scale was described in 1985. Despite its utility, weaknesses were identified, including noninclusion of synkinesis phenomena and insensitivity to segmental weakness. Therefore, the scale was recently revised to address these weaknesses. The objective of this investigation was to determine agreement between the original and the updated House-Brackmann scales. STUDY DESIGN: Prospective clinical trial. METHODS: Fifty consecutive new facial paralysis patients underwent standardized facial videography while performing facial movements. Video clips were scored by three independent facial nerve clinicians. The time it took to produce a score for each method was tracked. Interobserver correlations were calculated, and comparisons were made between scores by using the original and modified House-Brackmann scales. RESULTS: : Interobserver correlation was high for both House-Brackmann scales. Overall scores were in excellent agreement (difference of 0.1 ± 0.5, no statistical difference), although the modified scale took substantially longer to calculate (120 seconds ± 20 seconds vs. 30 seconds ± 10 seconds, P < .001). CONCLUSIONS: We found substantial grading correlation between the original and the newly modified House-Brackmann scales. Because of specific zonal and synkinesis scoring, the modified scale took longer to score.

Delayed presentation of a swallowed partial denture.

We report a unique case of a young patient who accidentally swallowed his partial denture and alarmingly only presented to our ear, nose and throat (ENT) department 4 weeks later despite several previous presentations to primary and secondary care. The partial denture was successfully removed under general anaesthetic using direct laryngoscopy following admission. He was discharged on a normal diet 6 days later after oesophageal perforation was excluded using a contrast swallow.

Partial adenoidectomy by suction diathermy in children with cleft palate, to avoid velopharyngeal insufficiency.

OBJECTIVES: Adenoidectomy is indicated for the relief of paediatric nasal obstruction, sleep-disordered breathing and otitis media with effusion (OME). Velopharyngeal insufficiency (VPI) is a rare complication. The main risk factor is the presence of pre-existing velopharyngeal closure-impaired mechanisms, including submucosal or overt cleft palate. Despite possible benefits, adenoidectomy is frequently withheld in such children to avoid VPI. This study aims to demonstrate the efficacy and safety of partial adenoidectomy using suction diathermy in children who previously underwent overt cleft palate repair during infancy, to allow selective resection of tissue and symptom resolution without producing VPI. METHODS: Since 1994, 18 patients with previously corrected overt cleft palate have undergone partial adenoidectomy at this centre, for the treatment of nasal obstruction or sleep-disordered breathing, with or without OME. Three had existing VPI following their cleft correction surgery. Selective resection of the adenoid was performed transorally under indirect vision, using a malleable suction coagulator. This allowed exposure of the posterior choanae, leaving the remaining adenoid bulk intact. RESULTS: Patients were followed up at 4 weeks, and subsequently at regular intervals (total follow up 30-180 months, median 92 months), including perceptual speech assessment in all cases. All demonstrated symptomatic improvement with respect to the original indications for surgery. None developed worsening hypernasal speech or other features of VPI, and there were no cases of symptomatic adenoidal re-growth. CONCLUSIONS: Partial adenoidectomy, employing a variety of methods, has been used successfully in children with submucosal cleft palate. This study demonstrates the safe and effective use of suction diathermy to enable partial adenoidectomy in children who have previously undergone surgical correction of overt cleft palate, allowing symptom resolution without producing VPI.

Presence of quorum-sensing systems associated with multidrug resistance and biofilm formation in Bacteroides fragilis.

Bacteroides fragilis constitutes 1-2% of the natural microbiota of the human digestive tract and is the predominant anaerobic opportunistic pathogen in gastrointestinal infections. Most bacteria use quorum sensing (QS) to monitor cell density in relation to other cells and their environment. In Gram-negative bacteria, the LuxRI system is common. The luxR gene encodes a transcriptional activator inducible by type I acyl-homoserine lactone autoinducers (e.g., N-[3-oxohexanoyl] homoserine lactone and hexanoyl homoserine lactone [C6-HSL]). This study investigated the presence of QS system(s) in B. fragilis. The genome of American-type culture collection strain no. ATCC25285 was searched for QS genes. The strain was grown to late exponential phase in the presence or absence of synthetic C6-HSL and C8-HSL or natural homoserine lactones from cell-free supernatants from spent growth cultures of other bacteria. Growth, susceptibility to antimicrobial agents, efflux pump gene (bmeB) expression, and biofilm formation were measured. Nine luxR and no luxI orthologues were found. C6-HSL and supernatants from Yersinia enterocolitica, Vibrio cholerae, and Pseudomonas aeruginosa caused a significant (1) reduction in cellular density and (2) increases in expression of four putative luxR genes, bmeB3, bmeB6, bmeB7, and bmeB10, resistance to various antibiotics, which was reduced by carbonyl cyanide-m-chlorophenyl hydrazone (CCCP, an uncoupler that dissipates the transmembrane proton gradient, which is also the driving force of resistance nodulation division efflux pumps) and (3) increase in biofilm formation. Susceptibility of ATCC25285 to C6-HSL was also reduced by CCCP. These data suggest that (1) B. fragilis contains putative luxR orthologues, which could respond to exogenous homoserine lactones and modulate biofilm formation, bmeB efflux pump expression, and susceptibility to antibiotics, and (2) BmeB efflux pumps could transport homoserine lactones.

Impact of anatomic site on growth, efflux-pump expression, cell structure, and stress responsiveness of Bacteroides fragilis.

This study investigated whether B. fragilis from various human sites acquired stable traits enabling it to express certain efflux pumps (EPs), adopt a particular cell structure, and tolerate certain stressors. Isolates from blood, abscess, and stool (n = 11 each) were investigated. Bacteria from various sites portrayed different ultrastructres and EP expression. Blood isolates were tolerant to nutrient limitation and stool isolates to NaCl and bile salt stress. Stressors significantly increased EP expression. These data demonstrate that (1) B. fragilis acquires stable traits from various in vivo microenvironments; (2) that EPs are involved in stress responsiveness; and (3) that EP expression is tightly controlled and site dependent.

Induction of multiple antibiotic resistance in Bacteroides fragilis by benzene and benzene-derived active compounds of commonly used analgesics, antiseptics and cleaning agents.

OBJECTIVES: To determine the potential of active compounds (ACs) present in commonly used analgesics/antiseptics and cleaning agents (detergents and disinfectants) to induce multiple antibiotic resistance (MAR) in Bacteroides fragilis. METHODS: B. fragilis ATCC 25285 untreated or pretreated with sublethal concentrations of ACs (n = 25) was grown for 12 h. Susceptibility of cells pre-treated with various ACs to antibiotics and expression of resistance nodulation division family (bmeB) efflux pumps and putative marA-like global activators (PGAs) were measured. RESULTS: Twelve aromatic ACs containing benzene or its activated derivatives (salicylate, acetaminophen, gingerol, benzoate, phenol, chlorhexidine gluconate, capsaicin, juglone, cinnamaldehyde, benzene, ibuprofen and Triton X-100) induced MAR, which was reduced by carbonyl cyanide m-chlorophenylhydrazone. There was a positive correlation between the predicted degree of benzene activation and the level of induction. Deactivated benzene or non-aromatic ACs were either poor inducers or non-inducers. Efflux pumps bmeB1, 3, 4, 7 and two PGAs bfrA1 and bfrA2 were overexpressed. Expression of bfrA1 or bfrA2 in Escherichia coli caused a >2-fold increase in the MAR and overexpression of acrB, suggesting that they were putative marA orthologues. CONCLUSIONS: These data demonstrate (i) the presence of an MarA-like system(s) in B. fragilis and (ii) the propensity of benzene or its activated derivatives present in pharmaceutical products to induce MAR.

Bile salts enhance bacterial co-aggregation, bacterial-intestinal epithelial cell adhesion, biofilm formation and antimicrobial resistance of Bacteroides fragilis.

Bacteroides fragilis is the most common anaerobic bacterium isolated from human intestinal tract infections. Before B. fragilis interacts with the intestinal epithelial cells, it is exposed to bile salts at physiological concentrations of 0.1-1.3%. The aim of this study was to determine how pre-treatment with bile salts affected B. fragilis cells and their interaction with intestinal epithelial cells. B. fragilis NCTC9343 was treated with conjugated bile salts (BSC) or non-conjugated bile salts (BSM). Cellular ultrastructure was assessed by electron microscopy, gene expression was quantified by comparative quantitative real-time RT-PCR. Adhesion to the HT-29 human intestinal cell line and to PVC microtitre plates (biofilm formation) was determined. Exposure to 0.15% BSC or BSM resulted in overproduction of fimbria-like appendages and outer membrane vesicles, and increased expression of genes encoding RND-type efflux pumps and the major outer membrane protein, OmpA. Bile salt-treated bacteria had increased resistance to structurally unrelated antimicrobial agents and showed a significant increase in bacterial co-aggregation, adhesion to intestinal epithelial cells and biofilm formation. These data suggest that bile salts could enhance intestinal colonization by B. fragilis via several mechanisms, and could therefore be significant to host-pathogen interactions.

The Bacteroides fragilis cell envelope: quarterback, linebacker, coach-or all three?

Bacteroides fragilis is an anaerobic commensal constituting only 1-2% of the micro-flora of the human gastrointestinal tract, yet it is the predominant anaerobic isolate in cases of intraabdominal sepsis and bacteremia. B. fragilis can play two roles in the host: in its role as friendly commensal, it must be able to establish itself in the host intestinal mucosa, to utilize and process polysaccharides for use by the host, and to resist the noxious effects of bile salts. In its role as pathogen, it must be able to attach itself to the site of infection, evade killing mechanisms by host defense, withstand antimicrobial treatment and produce factors that damage host tissue. The cell envelope of B. fragilis, likewise, must be able to function in the roles of aggressor, defender and strategist in allowing the organism to establish itself in the host--whether as friend or foe. Recent studies of the genomes and proteomes of the genus Bacteroides suggest that these organisms have evolved strategies to survive and dominate in the overcrowded gastrointestinal neighborhood. Analysis of the proteomes of B. fragilis and Bacteroides thetaiotaomicron demonstrates both a tremendous capacity to use a wide range of dietary polysaccharides, and the capacity to create variable surface antigenicities by multiple DNA inversion systems. The latter characteristic is particularly pronounced in the species B. fragilis, which is more frequently found at the mucosal surface (i.e., often the site of attack by host defenses). The B. fragilis cell envelope undergoes major protein expression and ultrastructural changes in response to stressors such as bile or antimicrobial agents. These agents may also act as signals for attachment and colonization. Thus the bacterium manages its surface characteristics to enable it to bind to its target, to use the available nutrients, and to avoid or evade hostile forces (host-derived or external) in its multiple roles.

Capture of flowing human neutrophils by immobilised immunoglobulin: roles of Fc-receptors CD16 and CD32.

We investigated capture and activation of flowing human neutrophils through their Fc-receptors, FcRgammaIIIB (CD16) and FcRgammaIIA (CD32). Immobilised platelets bearing murine monoclonal antibody against glycoprotein IIbIIIA were able to capture and activate flowing neutrophils. The activation response was inhibited by antibody blockade of neutrophil CD32. However, capture only occurred efficiently at wall shear stress below 0.1 Pa if platelet P-selectin was blocked. If neutrophils were perfused over immobilised human IgG, many adhered at 0.025 or 0.05 Pa, but not at 0.1 Pa. Adhesion was reduced by blockade of CD16 or CD32, but blockade of CD16 had the greater effect. When neutrophils were perfused over a combination of purified P-selectin and IgG, blockade of CD16 and CD32 inhibited activation of captured cells. Immunoglobulin deposited in tissue could capture and activate slow-flowing neutrophils. It might also potentiate inflammatory responses at higher stress if presented along with selectins. The dominant FcR for capture of neutrophils was CD16, but with murine antibody, CD32 played a greater role.

The fist model for nasal packing.

Epistaxis is a common ear, nose and throat emergency. A variety of nasal packs are available to control the bleeding by tamponade. Training of junior doctors to insert nasal packs is difficult when dealing with a bleeding patient. We discovered a readily available and simple model to enable trainees to learn the method of nasal packing.

Disturbed flow promotes deposition of leucocytes from flowing whole blood in a model of a damaged vessel wall.

Departure from simple laminar flow in arteries may promote the local attachment of leucocytes either to intact endothelium or platelet thrombi. We perfused blood through a chamber with a backward facing step, to observe whether adhesion from whole blood to P-selectin was indeed localized to a region of recirculating flow, and whether platelets binding to collagen in such a region could capture leucocytes. Blood flowing over the step established a stable vortex, a reattachment point where forward and backward flow separated, and a simple laminar flow with wall shear rate c. 400/s further downstream. Fluorescently labelled leucocytes were observed to attach to P-selectin immediately upstream or downstream of the reattachment point, and to roll back towards the step or away from it, respectively. There was negligible adhesion further downstream. When a P-selectin-Fc chimaera was used to coat the chamber, stable attachment occurred, again preferentially in the disturbed flow region. Numerous platelets adhered to a collagen coating throughout the chamber, although there were local maxima either side of the reattachment point. The adherent platelets captured flowing leucocytes in these regions alone. Leucocytes may adhere from flowing blood in vessels with high shear rate if the flow is disturbed. While platelets can adhere over a wider range of shear rates, their ability to capture leucocytes may be restricted to regions of disturbed flow.