Application of quasimetagenomics methods to define microbial diversity and subtype Listeria monocytogenes in dairy and seafood production facilities.

IMPORTANCE: In developed countries, the human diet is predominated by food commodities, which have been manufactured, processed, and stored in a food production facility. Little is known about the application of metagenomic sequencing approaches for detecting foodborne pathogens, such as L. monocytogenes, and characterizing microbial diversity in food production ecosystems. In this work, we investigated the utility of 16S rRNA amplicon and quasimetagenomic sequencing for the taxonomic and phylogenetic classification of Listeria culture enrichments of environmental swabs collected from dairy and seafood production facilities. We demonstrated that single-nucleotide polymorphism (SNP) analyses of L. monocytogenes metagenome-assembled genomes (MAGs) from quasimetagenomic data sets can achieve similar resolution as culture isolate whole-genome sequencing. To further understand the impact of genome coverage on MAG SNP cluster resolution, an in silico downsampling approach was employed to reduce the percentage of target pathogen sequence reads, providing an initial estimate of required MAG coverage for subtyping resolution of L. monocytogenes.

Gastrointestinal conditions related to tooth wear.

Gastro-oesophageal reflux disease (GORD) is a relatively common condition that occurs in adults and less commonly in children. It develops when the reflux of stomach contents into the oesophagus causes troublesome symptoms and/or complications. Signs and symptoms include heartburn, retrosternal discomfort, epigastric pain and hoarseness, dental erosion, chronic cough, burning mouth syndrome, halitosis and laryngitis. A proportion of patients will, however, have silent reflux. Strongly associated risk factors include family history, age, hiatus hernia, obesity and neurological conditions, such as cerebral palsy. There are different treatment options which may be considered for GORD, consisting of conservative, medical and surgical therapy. Dentists should be aware of the symptoms of GORD and dental signs of intrinsic erosion indicative of possible GORD so that they can question patients about this and, if appropriate, initiate a referral to a general medical practitioner.

Clinical and radiographic assessment and restoration of maxillary sinus function in relation to oro-antral fistula closure: a systematic review.

BACKGROUND: The purpose of this study was to determine current practice in assessment of maxillary sinus health during oro-antral fistula closure. METHOD: A systematic review using specific MeSH headings between 1990 and September 2021 on Medline, Ovid, and PubMed was performed. The inclusion criterion was closure of oro-antral fistula in adult patients. Exclusion criteria were oro-antral fistula arising in children; or from trauma, malignancy, or developmental abnormality; or any osteonecrotic lesion of bone. Risk of bias for individual papers was not assessed. Oxford Centre for Evidence Based Medicine levels of evidence were recorded. RESULTS: 4309 papers were identified, and 119 were eligible for review. Nineteen were from otolaryngology (16%), 25 from combined ear, nose, and throat (ENT) and oral/oral and maxillofacial (OS/OMF) surgery (21%), and 75 from OS/OMF surgery (63%). Preoperative rhinoscopy and cross-sectional imaging were reported in 33 papers (28%), reflecting the role of ENT with or without OS/OMFS. Sixty-eight did not record any formal assessment of maxillary sinus health (57%), of which 60 were from OS/OMF surgery (50%). Use of computed tomography was reported in 58 papers (49%), and use of cone beam computed tomography was reported in 8 papers (7%) but did not correlate with assessment of sinus health. DISCUSSION: Most surgery for oro-antral fistual closure was undertaken without objective assessment of maxillary sinus health, pre- or postoperatively. The involvement of ENT was associated with formal assessment of maxillary sinus health.

Modeling the catarrhal stage of Bordetella pertussis upper respiratory tract infections in mice.

Pertussis (whooping cough) is a highly transmissible human respiratory disease caused by Bordetella pertussis, a human-restricted pathogen. Animal models generally involve pneumonic infections induced by depositing large numbers of bacteria in the lungs of mice. These models have informed us about the molecular pathogenesis of pertussis and guided development of vaccines that successfully protect against severe disease. However, they bypass the catarrhal stage of the disease, when bacteria first colonize and initially grow in the upper respiratory tract. This is a critical and highly transmissible stage of the infection that current vaccines do not prevent. Here, we demonstrate a model system in which B. pertussis robustly and persistently infects the nasopharynx of TLR4-deficient mice, inducing localized inflammation, neutrophil recruitment and mucus production as well as persistent shedding and occasional transmission to cage mates. This novel experimental system will allow the study of the contributions of bacterial factors to colonization of and shedding from the nasopharynx, as occurs during the catarrhal stage of pertussis, and interventions that might better control the ongoing circulation of pertussis.

Bbvac: A Live Vaccine Candidate That Provides Long-Lasting Anamnestic and Th17-Mediated Immunity against the Three Classical Bordetella spp.

Acute pathogens such as Bordetella pertussis can cause severe disease but are ultimately cleared by the immune response. This has led to the accepted paradigm that convalescent immunity is optimal and therefore broadly accepted as the "gold standard" against which vaccine candidates should be compared. However, successful pathogens like B. pertussis have evolved multiple mechanisms for suppressing and evading host immunity, raising the possibility that disruption of these mechanisms could result in substantially stronger or better immunity. Current acellular B. pertussis vaccines, delivered in a 5-dose regimen, induce only short-term immunity against disease and even less against colonization and transmission. Importantly, they provide modest protection against other Bordetella species that cause substantial human disease. A universal vaccine that protects against the three classical Bordetella spp. could decrease the burden of whooping cough-like disease in humans and other animals. Our recent work demonstrated that Bordetella spp. suppress host inflammatory responses and that disrupting the regulation of immunosuppressive mechanisms can allow the host to generate substantially stronger sterilizing immunity against the three classical Bordetella spp. Here, we identify immune parameters impacted by Bordetella species immunomodulation, including the generation of robust Th17 and B cell memory responses. Disrupting immunomodulation augmented the immune response, providing strong protection against the prototypes of all three classical Bordetella spp. as well as recent clinical isolates. Importantly, the protection in mice lasted for at least 15 months and was associated with recruitment of high numbers of B and T cells in the lungs as well as enhanced Th17 mucosal responses and persistently high titers of antibodies. These findings demonstrate that disrupting bacterial immunomodulatory pathways can generate much stronger and more protective immune responses to infection, with important implications for the development of better vaccines. IMPORTANCE Infectious diseases are a major cause of morbidity and mortality in the United States, accounting for over 40 million hospitalizations since 1998. Therefore, novel vaccine strategies are imperative, which can be improved with a better understanding of the mechanisms that bacteria utilize to suppress host immunity, a key mechanism for establishing colonization. Bordetella spp., the causative agents of whooping cough, suppress host immunity, which allows for persistent colonization. We discovered a regulator of a bacterial immunosuppressive pathway, which, when mutated in Bordetella spp., allows for rapid clearance of infection and subsequent generation of protective immunity for at least 15 months. After infection with the mutant strain, mice exhibited sterilizing immunity against the three classical Bordetella spp., suggesting that the immune response can be both stronger and cross-protective. This work presents a strategy for vaccine development that can be applied to other immunomodulatory pathogens.

Modeling Immune Evasion and Vaccine Limitations by Targeted Nasopharyngeal Bordetella pertussis Inoculation in Mice.

Conventional pertussis animal models deliver hundreds of thousands of Bordetella pertussis bacteria deep into the lungs, rapidly inducing severe pneumonic pathology and a robust immune response. However, human infections usually begin with colonization and growth in the upper respiratory tract. We inoculated only the nasopharynx of mice to explore the course of infection in a more natural exposure model. Nasopharyngeal colonization resulted in robust growth in the upper respiratory tract but elicited little immune response, enabling prolonged and persistent infection. Immunization with human acellular pertussis vaccine, which prevents severe lung infections in the conventional pneumonic infection model, had little effect on nasopharyngeal colonization. Our infection model revealed that B. pertussis can efficiently colonize the mouse nasopharynx, grow and spread within and between respiratory organs, evade robust host immunity, and persist for months. This experimental approach can measure aspects of the infection processes not observed in the conventional pneumonic infection model.

Neuregulin-4 contributes to the establishment of cutaneous sensory innervation.

Recent work has shown that neuregulin-4 (NRG4) is a physiological regulator of the growth of sympathetic axons and CNS dendrites in the developing nervous system. Here, we have investigated whether NRG4 plays a role in sensory axon growth and the establishment of cutaneous sensory innervation. Imaging early nerve fibers in the well-characterized cutaneous trigeminal territory, the brachial plexus, and thorax revealed very marked and highly significant decreases in nerve fiber length and branching density in Nrg4-/- embryos compared with Nrg4+/+ littermates. NRG4 promoted neurotrophin-independent sensory axon growth from correspondingly early trigeminal ganglion and DRG neurons in culture but not from enteroceptive nodose ganglion neurons. High levels of Nrg4 mRNA were detected in cutaneous tissues but not in sensory ganglia. Our findings suggest that NRG4 is an important target-derived factor that participates in the establishment of early cutaneous sensory innervation.

BET 1: is the patient's perception of shortness of breath a useful triage tool in exacerbations of COPD?

A shortcut review was carried out to establish whether the degree of breathlessness in patients with an acute exacerbation of COPD is indicative of the severity of the exacerbation. Three hundred and forty-seven papers were found using the reported searches, of which five presented the best available evidence to answer the clinical question. The author, date and country of publication, patient group studied, study type, relevant outcomes, results and study weaknesses of these five papers are tabulated. It is concluded that increased shortness of breath is associated with a worse prognosis in patients with acute exacerbations of COPD. Dyspnoea assessment should be included in the triage process.

CD40 forward signalling is a physiological regulator of early sensory axon growth.

Multiple members of the tumour necrosis factor superfamily (TNFSF) regulate the growth and branching of neural processes late in development, when neurons are establishing and refining connections. Here, we present the first evidence that a TNFSF member acts much earlier in development, when axons are growing to their targets. CD40L transiently enhanced axon growth from embryonic mouse DRG neurons cultured at this early stage. Early spinal nerves of embryos lacking the CD40L receptor (Cd40-/- mice) were significantly shorter in vivo than those of Cd40+/+ littermates. CD40L was synthesized in early DRG targets and was co-expressed with CD40 in early DRG neurons. Whereas CD40L enhanced early axon growth independently of neurotrophins, disruption of a CD40L/CD40 autocrine loop impaired early neurotrophin-promoted axon growth. In marked contrast to the widespread regulation of axon and dendrite growth by CD40L reverse signalling later in development, CD40-Fc, which activates reverse signalling, had no effect on early sensory axon growth. These results suggest that CD40 forward signalling is a novel physiological regulator of early axon growth that acts by target-derived and autocrine mechanisms.

BET 1: The role of topical phenylephrine in non-traumatic epistaxis.

A short cut review of the literature was carried out to establish whether topical phenylephrine was an effective treatment for non-traumatic epistaxis. A single paper looked at this modality comparing it to another topical treatment. The author, date and country of publication, patient group studied, study type, relevant outcomes, results and study weaknesses are tabulated. It is concluded that further more robust studies are needed to answer this question.

Enhanced triage for patients with suspected cardiac chest pain: the History and Electrocardiogram-only Manchester Acute Coronary Syndromes decision aid.

OBJECTIVES: Several decision aids can 'rule in' and 'rule out' acute coronary syndromes (ACS) in the Emergency Department (ED) but all require measurement of blood biomarkers. A decision aid that does not require biomarker measurement could enhance risk stratification at triage and could be used in the prehospital environment. We aimed to derive and validate the History and ECG-only Manchester ACS (HE-MACS) decision aid using only the history, physical examination and ECG. METHODS: We undertook secondary analyses in three prospective diagnostic accuracy studies that included patients presenting to the ED with suspected cardiac chest pain. Clinicians recorded clinical features at the time of arrival using a bespoke form. Patients underwent serial troponin sampling and 30-day follow-up for the primary outcome of ACS. The model was derived by logistic regression in one cohort and validated in two similar prospective studies. RESULTS: The HE-MACS model was derived in 796 patients and validated in cohorts of 474 and 659 patients. HE-MACS incorporated age, sex, systolic blood pressure plus five historical variables to stratify patients into four risk groups. On validation, 5.5 and 12.1% (pooled total 9.4%) patients were identified as 'very low risk' (potential immediate rule out) with a pooled sensitivity of 99.5% (95% confidence interval: 97.1-100.0%). CONCLUSION: Using only the patient's history and ECG, HE-MACS could 'rule out' ACS in 9.4% of patients while effectively risk stratifying remaining patients. This is a very promising tool for triage in both the prehospital environment and ED. Its impact should be prospectively evaluated in those settings.

Enhancement of immune response against Bordetella spp. by disrupting immunomodulation.

Well-adapted pathogens must evade clearance by the host immune system and the study of how they do this has revealed myriad complex strategies and mechanisms. Classical bordetellae are very closely related subspecies that are known to modulate adaptive immunity in a variety of ways, permitting them to either persist for life or repeatedly infect the same host. Exploring the hypothesis that exposure to immune cells would cause bordetellae to induce expression of important immunomodulatory mechanisms, we identified a putative regulator of an immunomodulatory pathway. The deletion of btrS in B. bronchiseptica did not affect colonization or initial growth in the respiratory tract of mice, its natural host, but did increase activation of the inflammasome pathway, and recruitment of inflammatory cells. The mutant lacking btrS recruited many more B and T cells into the lungs, where they rapidly formed highly organized and distinctive Bronchial Associated Lymphoid Tissue (BALT) not induced by any wild type Bordetella species, and a much more rapid and strong antibody response than observed with any of these species. Immunity induced by the mutant was measurably more robust in all respiratory organs, providing completely sterilizing immunity that protected against challenge infections for many months. Moreover, the mutant induced sterilizing immunity against infection with other classical bordetellae, including B. pertussis and B. parapertussis, something the current vaccines do not provide. These findings reveal profound immunomodulation by bordetellae and demonstrate that by disrupting it much more robust protective immunity can be generated, providing a pathway to greatly improve vaccines and preventive treatments against these important pathogens.

Adipocyte-secreted BMP8b mediates adrenergic-induced remodeling of the neuro-vascular network in adipose tissue.

Activation of brown adipose tissue-mediated thermogenesis is a strategy for tackling obesity and promoting metabolic health. BMP8b is secreted by brown/beige adipocytes and enhances energy dissipation. Here we show that adipocyte-secreted BMP8b contributes to adrenergic-induced remodeling of the neuro-vascular network in adipose tissue (AT). Overexpression of bmp8b in AT enhances browning of the subcutaneous depot and maximal thermogenic capacity. Moreover, BMP8b-induced browning, increased sympathetic innervation and vascularization of AT were maintained at 28 °C, a condition of low adrenergic output. This reinforces the local trophic effect of BMP8b. Innervation and vascular remodeling effects required BMP8b signaling through the adipocytes to 1) secrete neuregulin-4 (NRG4), which promotes sympathetic axon growth and branching in vitro, and 2) induce a pro-angiogenic transcriptional and secretory profile that promotes vascular sprouting. Thus, BMP8b and NRG4 can be considered as interconnected regulators of neuro-vascular remodeling in AT and are potential therapeutic targets in obesity.

Microbiomes Associated With Foods From Plant and Animal Sources.

Food microbiome composition impacts food safety and quality. The resident microbiota of many food products is influenced throughout the farm to fork continuum by farming practices, environmental factors, and food manufacturing and processing procedures. Currently, most food microbiology studies rely on culture-dependent methods to identify bacteria. However, advances in high-throughput DNA sequencing technologies have enabled the use of targeted 16S rRNA gene sequencing to profile complex microbial communities including non-culturable members. In this study we used 16S rRNA gene sequencing to assess the microbiome profiles of plant and animal derived foods collected at two points in the manufacturing process; post-harvest/pre-retail (cilantro) and retail (cilantro, masala spice mixes, cucumbers, mung bean sprouts, and smoked salmon). Our findings revealed microbiome profiles, unique to each food, that were influenced by the moisture content (dry spices, fresh produce), packaging methods, such as modified atmospheric packaging (mung bean sprouts and smoked salmon), and manufacturing stage (cilantro prior to retail and at retail). The masala spice mixes and cucumbers were comprised mainly of Proteobacteria, Firmicutes, and Actinobacteria. Cilantro microbiome profiles consisted mainly of Proteobacteria, followed by Bacteroidetes, and low levels of Firmicutes and Actinobacteria. The two brands of mung bean sprouts and the three smoked salmon samples differed from one another in their microbiome composition, each predominated by either by Firmicutes or Proteobacteria. These data demonstrate diverse and highly variable resident microbial communities across food products, which is informative in the context of food safety, and spoilage where indigenous bacteria could hamper pathogen detection, and limit shelf life.

TWE-PRIL reverse signalling suppresses sympathetic axon growth and tissue innervation.

TWE-PRIL is a naturally occurring fusion protein of components of two TNF superfamily members: the extracellular domain of APRIL; and the intracellular and transmembrane domains of TWEAK with no known function. Here, we show that April-/- mice (which lack APRIL and TWE-PRIL) exhibited overgrowth of sympathetic fibres in vivo, and sympathetic neurons cultured from these mice had significantly longer axons than neurons cultured from wild-type littermates. Enhanced axon growth from sympathetic neurons cultured from April-/- mice was prevented by expressing full-length TWE-PRIL in these neurons but not by treating them with soluble APRIL. Soluble APRIL, however, enhanced axon growth from the sympathetic neurons of wild-type mice. siRNA knockdown of TWE-PRIL but not siRNA knockdown of APRIL alone also enhanced axon growth from wild-type sympathetic neurons. Our work reveals the first and physiologically relevant role for TWE-PRIL and suggests that it mediates reverse signalling.

Blood or Serum Exposure Induce Global Transcriptional Changes, Altered Antigenic Profile, and Increased Cytotoxicity by Classical Bordetellae.

The classical bordetellae sense and respond to a variety of environments outside and within their mammalian hosts. By causing inflammation and tissue damage, we reasoned that bordetellae are likely to encounter components of blood and/or serum during the course of a respiratory infection, and that detecting and responding to these would be advantageous. Therefore, we hypothesized that classical bordetellae have the ability to sense and respond to blood or serum. Blood or serum exposure resulted in substantial transcriptional changes in Bordetella bronchiseptica, including enhanced expression of many virulence-associated genes. Exposure to blood or serum additionally elicited production of multiple antigens not otherwise detectable, and led to increased bacterial cytotoxicity against macrophages. Transcriptional responses to blood/serum were observed in a Bvg- phase-locked mutant, indicating that the response is not solely dependent on a functional BvgAS system. Similar transcriptional responses to blood/serum were observed for the other classical bordetellae, Bordetella pertussis and Bordetella parapertussis. These data suggest the classical bordetellae respond to signals present in blood and serum by changing their behavior in ways that likely contribute to their remarkable success, via effects on pathogenesis, persistence and/or transmission between hosts.

How events in emergency medicine impact doctors' psychological well-being.

BACKGROUND: Emergency medicine is a high-pressured specialty with exposure to disturbing events and risk. We conducted a qualitative study to identify which clinical events resulted in emotional disruption and the impact of these events on the well-being of physicians working in an ED. METHODS: We used the principles of naturalistic inquiry to conduct narrative interviews with physicians working in the ED at Central Manchester University Hospitals NHS Foundation Trust, between September and October 2016. Participants were asked, 'Could you tell me about a time when an event at work has continued to play on your mind after the shift in which it occurred was over?' Data were analysed using framework analysis. The study had three a priori themes reported here. Other emergent themes were analysed separately. RESULTS: We interviewed 17 participants. Within the first a priori theme ('clinical events') factors associated with emotional disruption included young or traumatic deaths, patients or situations that physicians could relate to, witnessing the impact of death on relatives, the burden of responsibility (including medical error) and conflict in the workplace. Under theme 2 (psychological and physical effects), participants reported substantial upset leading to substance misuse, sleep disruption and neglecting their own physical needs through preoccupation with caring. Within theme 3 (impact on relationships), many interviewees described becoming withdrawn from personal relationships following clinical events, while others described feeling isolated because friends and family were non-medical. CONCLUSIONS: Clinical events encountered in the ED can affect a physician's psychological and physical well-being. For many participants these effects were negative and long lasting.

Social-media-enabled learning in emergency medicine: a case study of the growth, engagement and impact of a free open access medical education blog.

BACKGROUND: Clinicians are increasingly using social media for professional development and education. In 2012, we developed the St.Emlyn's blog, an open access resource dedicated to providing free education in the field of emergency medicine. OBJECTIVE: To describe the development and growth of this international emergency medicine blog. METHOD: We present a narrative description of the development of St.Emlyn's blog. Data on scope, impact and engagement were extracted from WordPress, Twitter and Google Analytics. RESULTS: The St.Emlyn's blog demonstrates a sustained growth in size and user engagement. Since inception in 2012, the site has been viewed over 1.25 million times with a linear year-on-year growth. We have published over 500 blog posts, each of which attracts a mean of 2466 views (range 382-69 671). The site has been viewed in nearly every country in the world, although the majority (>75%) of visitors come from the USA, UK and Australia. SUMMARY: This case study of an emergency medicine blog quantifies the reach and engagement of social-media-enabled learning in emergency medicine.