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This case discusses a 10-year-old boy who presented in significant respiratory distress with cardiac tamponade with associated gross ascites and hepatomegaly, requiring urgent transfer for pericardiocentesis. On further investigation, he was found to have multiple pulmonary emboli and evidence of panserositis. An underlying rheumatological cause was suspected in the absence of any evidence of infection or malignancy, and blood tests were positive for anti-double stranded DNA, anticardiolipin and antiglycoprotein antibodies as well as HLA B51. In his medical history, he has previously had mouth ulcers, chronic anaemia of undetermined cause and erythema multiforme. These symptoms, along with clinical presentation, mean a diagnosis of Behcet’s disease and associated antiphospholipid syndrome was felt to be most likely. Anticoagulation therapy was commenced for treatment of the emboli, and colchicine was started for management of Behcet’s disease. The patient was discharged clinically well from the hospital and continues under specialist rheumatological and haematological follow-up.

During the COVID-19 pandemic, SARS-CoV-2 surveillance efforts integrated genome sequencing of clinical samples to identify emergent viral variants and to support rapid experimental examination of genome-informed vaccine and therapeutic designs. Given the broad range of methods applied to generate new viral genomes, it is critical that consensus and variant calling tools yield consistent results across disparate pipelines. Here we examine the impact of sequencing technologies (Illumina and Oxford Nanopore) and 7 different downstream bioinformatic protocols on SARS-CoV-2 variant calling as part of the NIH Accelerating COVID-19 Therapeutic Interventions and Vaccines (ACTIV) Tracking Resistance and Coronavirus Evolution (TRACE) initiative, a public-private partnership established to address the COVID-19 outbreak. Our results indicate that bioinformatic workflows can yield consensus genomes with different single nucleotide polymorphisms, insertions, and/or deletions even when using the same raw sequence input datasets. We introduce the use of a specific suite of parameters and protocols that greatly improves the agreement among pipelines developed by diverse organizations. Such consistency among bioinformatic pipelines is fundamental to SARS-CoV-2 and future pathogen surveillance efforts. The application of analysis standards is necessary to more accurately document phylogenomic trends and support data-driven public health responses.

An array of SARS-CoV-2 virus variants have been isolated, propagated and used in in vitro assays, in vivo animal studies and human clinical trials. Observations of working stocks of SARS-CoV-2 suggest that sequential propagation in Vero cells leads to critical changes in the region of the furin cleavage site, which significantly reduce the value of the working stock for critical research studies. Serially propagating SARS-CoV-2 in Vero E6 cells leads to rapid increases in genetic variants while propagation in other cell lines (e.g. Vero/hSLAM) appears to mitigate this risk thereby improving the overall genetic stability of working stocks. From these observations, investigators are urged to monitor genetic variants carefully when propagating SARS-CoV-2 in Vero cells.

Objectives The Quality Improvement Project (QIP) aims to implement Family Integrated Care (FiCare) in our Neonatal Unit ensuring that a positive parental experience is provided. The principle objective was to gain feedback from previous parents in order to identify barriers and facilitators to a positive parental experience, areas for improvement and to consider specific changes in practice to implement. Method Data were collected in our tertiary level Unit using the following mixed methods: 1) Questionnaires posted to parents who had a baby admitted to the Neonatal Unit in the preceding two years. 2) One open-ended question. 3) A focus group inclusive of staff and parents. Quantitative data were analysed to identify areas of both success and for improvement. Qualitative data were analysed using framework analysis to identify common themes. Results From 37 completed questionnaires parents felt: •Involved in their baby's care. •Staff were trustworthy. •Good feeding support was provided. Areas for improvement included providing more information about a baby's condition and about support groups. Seven emerging themes from the analysis, including communication and staff support, identified a number of facilitators and barriers to a positive parental experience. Conclusion The data analysis suggested key areas for improvement include communication, providing more information about support groups and better preparation of discharge. Based on these findings, changes in our Neonatal Unit have been implemented to improve parental experience. Regular committee groups will continue and subsequently feedback will be re-assessed.

As SARS-CoV-2 variants emerge, there is a critical need to understand the effectiveness of serum elicited by different SARS-CoV-2 vaccines. A reference reagent comprised of post-vaccination sera from recipients of different vaccines allows evaluation of in vitro variant neutralization, and provides a reference for comparing assay results across laboratories. We prepared and pooled >1 L serum from donors who received the SARS-CoV-2 mRNA vaccines (BNT162b2, Pfizer and mRNA-1273, Moderna), a replication-incompetent adenovirus type 26 vaccine (Ad26.COV2.S, Johnson and Johnson), or recombinant spike protein expressed by baculovirus incorporated into a nanoparticle vaccine plus Matrix-M adjuvant (NVX-CoV2373, Novavax). Twice frozen sera were aliquoted and are available for distribution to the research community (BEI Resources). The calculated WHO titer of pooled sera to spike protein was 1,312, 1,447, 1,936, and 587 and the reciprocal RBD binding to ACE-2 IC90-titers were 60, 64, 118, and 46 for BNT162b2, mRNA1273, Ad26.CoV2373, and NVX-CoV2373 sera, respectively.

Building upon the foundational research of Robert Koch, who demonstrated the ability to grow Mycobacterium tuberculosis for the first time in 1882 using media made of coagulated bovine serum, microbiologists have continued to develop new and more efficient ways to grow mycobacteria. Presently, all known mycobacterial species can be grown in the laboratory using either axenic culture techniques or in vivo passage in laboratory animals. This chapter provides conventional protocols to grow mycobacteria for diagnostic purposes directly from clinical specimens, as well as in research laboratories for scientific purposes. Detailed protocols used for production of M. tuberculosis in large scale (under normoxic and hypoxic conditions) in bioreactors and for production of obligate intracellular pathogens such as Mycobacterium leprae and "Mycobacterium lepromatosis" using athymic nude mice and armadillos are provided.

The function of the SARS-CoV-2 accessory protein p6, encoded by ORF6, is not fully known. Based upon its similarity to p6 from SARS-CoV, it may play a similar role, namely as an antagonist of type I interferon (IFN) signaling. Here we report the sequencing of a SARS-CoV-2 strain passaged six times after original isolation from a clinical patient in Hong Kong. The genome sequence shows a 27 nt in-frame deletion (Δ27,264-27,290) within ORF6, predicted to result in a 9 aa deletion (ΔFKVSIWNLD) from the central portion of p6. This deletion is predicted to result in a dramatic alteration in the three-dimensional structure of the resultant protein (p6Δ22-30), possibly with significant functional implications. Analysis of the original clinical sample indicates that the deletion was not present, while sequencing of subsequent passages of the strain identifies the deletion as a majority variant. This suggests that the deletion originated ab initio during passaging and subsequently propagated into the majority, possibly due to the removal of selective pressure through the IFN-deficient Vero E6 cell line. The specific function of the SARS-CoV-2 p6 N-terminus, if any, has not yet been determined. However, this deletion is predicted to cause a shift from N-endo to N-ecto in the transmembrane localization of the SARS-CoV-2 p6Δ22-30 N-terminus, possibly leading to the ablation of its native function.

Objectives Our group has previously shown in a study presented at the Preventing Overdiagnosis 2018 conference (DOI: 10.1136/bmjebm-2018-111070.47) that lower respiratory tract infections (LRTIs) in paediatric patients are not only over diagnosed, but also over and inappropriately investigated and treated. The next stage of our projected, presented here, focuses on: Understanding the factors driving this overdiagnosis to prevent unnecessary investigations, notably Chest X–rays (CXRs) and blood tests, and reduce inappropriate antibiotic use Applying lessons learnt using a Plan, Do, Study, Act (PDSA) cycle prior to fully implementing a change of practice and re–auditing Method A mixed quantitative-qualitative analysis was conducted involving Paediatric departments across South East England involving multidisciplinary team (MDT) members. Using triangulation, data was collected using the following methods: Questionnaires (sent out via SurveyMonkey) to assess use of investigations and antibiotic prescribing practices of doctors in cases of suspected LRTI, and adherence to local and national guidelines Multiple focus groups including staff from Paediatrics, Accident and Emergency (A&E), Microbiology, and Pharmacy departments to consider factors driving overdiagnosis and over management, as well as to identify barriers and facilitators in preventing overdiagnosis Further focus groups to gather views on the full range of proposed interventions Re–audit of local sepsis pathway following changes made based on focus group discussions Focus group data was analysed using Framework analysis to identify common themes. Results Preliminary data gathered from questionnaires suggest that doctors are more likely to follow local guidelines for the management of LRTI, but that not every hospital has these. Over half of clinicians reported they would order blood tests in a child with suspected LRTI, and some would order CXRs. The majority of survey respondents said they would rely on a CRP result to decide if antibiotics are required, particularly intravenous (IV) antibiotics, and almost half would sometimes prescribe antibiotics if a parent or patient insists on it. Following the focus group discussions, the local sepsis pathway was identified as being ‘over-sensitive’, and thought to result in the over treatment of children with IV antibiotics. The pathway was therefore modified and audit data from before and after these changes will be presented. Other perceived barriers and facilitators to preventing overdiagnosis which emerged from the thematic analysis will also be presented. Conclusions This quantitative-qualitative analysis has highlighted some of the factors driving the overdiagnosis and over management of LRTIs in children. Findings suggest that faster availability of infection marker results, better clinical guidelines, regular antibiotic prescription audits, and educational sessions involving senior staff who influence junior prescribing habits would be useful in guiding management of suspected LRTI in children. As a result, the following interventions are being considered: 1) Auditing local sepsis tools and changes to current pathways 2) Discouraging unnecessary investigations by using posters in doctors’ offices, paediatric assessment units and A&E 3) Updating local guidelines to reflect national ones 4) Encouraging nurses to decline ordering CXRs if patients have not been assessed by doctors 5) Educational sessions on investigation and antibiotic choices We hope that implementing these changes will results in fewer unnecessary investigations and help promote better antimicrobial stewardship. We plan to fully re-audit at a future date.

Objectives The aim of this project was to test our clinical impression that a significant number of children and young people are diagnosed with and treated for ‘chest sepsis’ unnecessarily. We also wanted to investigate the hypothesis that children received unecessary investigations, including Chest X-rays (CXRs) and measurement of acute phase reactants such as C-Reactive Protein (CRP), which are not recommended by British Thoracic Society (BTS) guidelines and would have been treated inappropriately with intravenous (IV) antibiotics. Our main objectives were thus: To design a Quality Improvement Project to investigate whether we overdiagnose chest sepsis, over investigate children presenting with possible signs and symptoms of chest sepsis and over use IV antibiotics To evaluate adherence to BTS and NICE sepsis guidelines To suggest measures to implement to prevent over investigation and unnecessary use of antibiotics in children with chest sepsis. Method An audit was undertaken within the Paediatric Department in an NHS District General Hospital setting in the United Kingdom. Data was collected retrospectively from May 2017 to January 2018. The inclusion criteria were any child >10 days old admitted to the ward and treated with antibiotics for suspected lower respiratory tract infection (LRTI). Patients with underlying chronic lung disorders (except asthma/wheeze), cardiac conditions, immunodeficiency or those who are immunosuppressed were excluded. Patients were categorised by age ( 12) and then stratified according to sepsis risk, as per NICE and BTS guidelines. The data were then analysed assessing adherence to these guidelines during clinical assessment and management following presentation with signs and symptoms of a LRTI. Results • Data from 156 patients were collected: 132 aged under 5, 21 aged 5–11 and 3 aged 12–17. • 10% were low, 21% moderate and 69% high risk for sepsis on presentation. • 36 patients were noted to have low oxygen saturations on admission, of which 64% had co-existing wheeze. • 66% of patients, of which 73% were high risk and 27% were low or moderate risk for sepsis were treated with IV antibiotics, most commonly Ceftriaxone. • 33% were treated with oral antibiotics, most commonly Co-amoxiclav. Only 9.8% of this group were treated with oral Amoxicillin as per BTS guidelines. • The majority (88%) had a CXR. Half were reported as normal, of which 71% were treated with IV antibiotics. 78% of those treated with oral antibiotics had a CXR. • 78% of patients had blood tests, which all included a CRP, which is not recommended to be routinely measured. Conclusions First phase audit results support the hypothesis that children presenting with signs and symptoms of a possible LRTI are overdiagnosed and over investigated. 61% of children were under the age of two and treated with antibiotics even though BTS guidelines suggest this age group is much less likely to have a bacterial infection. Over half of those with low oxygen saturations had co-existing wheeze, suggesting over-treatment of patients with wheeze for LRTI. 88% of our patients had CXRs, which is contrary to BTS guidelines, that suggest routine CXRs are unnecessary. 71% of children reported to have a normal CXR received IV antibiotics. We plan to implement a change of practice within the Paediatric department to prevent unnecessary investigations, notably CXRs and blood tests and to reduce unnecessary antibiotic use that comes with long term risks for the wider population. Following this we plan to re-audit.

There are many resources available to mycobacterial researchers, including culture collections around the world that distribute biomaterials to the general scientific community, genomic and clinical databases, and powerful bioinformatics tools. However, many of these resources may be unknown to the research community. This review article aims to summarize and publicize many of these resources, thus strengthening the quality and reproducibility of mycobacterial research by providing the scientific community access to authenticated and quality-controlled biomaterials and a wealth of information, analytical tools and research opportunities.

Pseudomonas aeruginosa and Candida albicans (are opportunistic pathogens that cause systemic infections in immune-suppressed patients. They show important bacterial-fungal interactions including quorum sensing. This involves cell signaling to communicate between the cells of their own colony and the cells of rival microbes or the host. It is thought that this phenomenon is vital in the potential competition and virulence of the organisms. We report a case of a previously healthy 2-year-old boy, where an accidental injury had been sustained resulting in a closed fracture of femur. He subsequently developed sepsis related to co-infection by C. albicans and P. aeruginosa. Trauma may result in a transient immune-suppression and predispose to sepsis caused by opportunistic microorganisms. They can engage in bacterial-fungal interaction. Clinicians should consider invasive co-infection when initial cultures show evidence for only 1 pathogen.

FASPAX (Fermi-Argonne Semiconductor Pixel Array X-ray detector) has been planned as a fast integrating area detector with wide dynamic range for time resolved applications at the upgraded Advanced Photon Source (APS). The detector will achieve a burst image rate of 13 MHz, matching the bunch rate of the proposed storage ring timing mode. A unique three gain charge splitting integration circuit will permit wide dynamic range — from single photon sensitivity to 105 photons/pixel (100 μm × 100 μm) in a single exposure. Incorporation of a novel interposer layer in the hybrid stack will permit large area sensors without the usual coverage gaps associated with hybrid pixel devices. The detector will provide access to the full tem-poral resolution of the APS, and enable novel science, such as pump-probe studies of irreversible systems. The APS Upgrade is currently funding development of small prototypes (1.2 cm × 1.2 cm) that will be tested in the next few years. This work will present an overview of the detector, and current status of the detector development.

The number of children with congenital heart disease graduating to adult care is increasing rapidly as treatment becomes more successful. A significant proportion has pulmonary hypertension, which will lead to Eisenmenger’s syndrome, having an associated high morbidity and mortality rate. This article discusses the pathophysiology of Eisenmenger’s syndrome, its clinical course and management. Medical management of these complex patients is evolving quickly and rapidly changing. We discuss each of the therapies available for the treatment of this disease, use of phosphodiesterase inhibitors, endothelin receptor antagonists and prostanoids. Transplantation is now less likely with these new medications. We also discuss the important role of the nurse in Eisenmenger’s syndrome and administration of various therapies.

It is estimated that around 75% of patients with Borderline Personality Disorder (BPD) are prescribed psychotropic medication during their treatment course, although this is not recommended as first line therapy. In the UK, there are no guidelines to advise which drug treatments to use in BPD, however, numerous, but mostly small scale studies, show evidence that different medications target specific core symptoms. We report a case of a 25 year old woman with BPD, who has received treatment with five different psychotropic medications. We go on to assess not only the efficacy of these treatments in this individual case, but also whether the use of these treatments is in line with best evidence according to currently available research.

Coagulase-negative staphylococci (CoNS) are the most common cause of biofilm-associated sepsis in very low birth weight infants (VLBW). Standard biofilm assays may not predict the pathogenic potential of CoNS since biofilm production is regulated by diverse environmental stimuli. Staphylococcus epidermidis isolated from blood cultures from VLBW infants were evaluated for biofilm production in response to various environmental stimuli, including intravenous solutions and skin preparations. While responses to environmental stimuli were variable for individual isolates and products, some trends were observed. Biofilm production by hospital S. epidermidis isolates (predominantly ica and biofilm-positive) was most commonly increased at 30°C and decreased in the presence of intravenous solutions and moisturisers. Commensals (mainly biofilm-negative and lacking the ica gene) were more often induced to produce biofilm than hospital isolates. These results indicate that biofilm production in S. epidermidis can vary in response to environmental stimuli encountered in the clinical setting, that standard biofilm assays are unlikely to predict clinical outcome, and that harmless skin commensals may be induced to produce biofilm by some of the products used in neonatal units.