3D Bioprinting of Mature Bacterial Biofilms for Antimicrobial Resistance Drug Testing

The potential to bioprint and study 3D bacterial biofilm constructs could have great clinical significance at a time when antimicrobial resistance is rising to dangerously high levels worldwide. In this study, clinically relevant bacterial species including Escherichia coli, Staphylococcus aureus (MSSA), Methicillin-resistant Staphylococcus aureus (MRSA) and Pseudomonas aeruginosa were 3D bioprinted using a double-crosslinked alginate bioink to form mature bacteria biofilms, characterized by confocal laser scanning microscopy (CLSM) and fluorescent staining. Solid and porous bacteria-laden constructs were reproducibly bioprinted with thicknesses ranging from 0.25 to 4 mm. We demonstrated 3D bioprinting of thicker biofilms (>4 mm) than found in currently available in vitro models. Bacterial viability was excellent in the bioprinted constructs, with CLSM observation of bacterial biofilm production and maturation possible for at least 28 d in culture. Importantly, we observed the complete five-step biofilm life cycle in vitro following 3D bioprinting for the first time, suggesting the formation of mature 3D bioprinted biofilms. Bacterial growth was faster in thinner, more porous constructs whilst constructs crosslinked with BaCl 2 concentrations of above 10 mM had denser biofilm formation. 3D MRSA and MSSA biofilm constructs were found to show greater resistance to antimicrobials than corresponding two-dimensional (2D) cultures. Thicker 3D E. coli biofilms had greater resistance to tetracycline than thinner constructs over 7 d of treatment. Our methodology allowed for the precise 3D bioprinting of self-supporting 3D bacterial biofilm structures that developed biofilms during extended culture. 3D biofilm constructs containing bacterial biofilms produce a model with much greater clinical relevance compared to 2D culture models and we have demonstrated their use in antimicrobial testing

"Feeding Hungry Children": the growth of charitable breakfast clubs and holiday hunger projects in the UK

This paper reports the findings of a recent scoping study on the nature, evidence base and policy context of breakfast clubs and school holiday projects in the UK and aims to do three things. Firstly, bring child feeding initiatives firmly into the critical food and poverty debate. Secondly, critically assess the state of evidence on these projects and their perceived role in the lives of children at risk of, or experiencing, poverty. Thirdly, set out the next steps which will be required to build a systematic body of research on child feeding projects within the food and poverty research agenda

The geographies of food banks in the meantime

The authors gratefully acknowledge the support given by the British Academy for this research (grant no. SG131950). The ‘Emergency Food Provision in the UK’ research includes: over eighteen months of ethnographic research in a Trussell Trust Foodbank; a national survey of the Trussell Trust Network and Independent food banks (and other food aid providers); and in-depth interviews with food bank managers, volunteers and service-users in London, Bristol, Leicestershire, South Wales, Devon, and Cornwall

Clinical outcomes and response to treatment of patients receiving topical treatments for pyoderma gangrenosum: a prospective cohort study

Background: pyoderma gangrenosum (PG) is an uncommon dermatosis with a limited evidence base for treatment. Objective: to estimate the effectiveness of topical therapies in the treatment of PG. Methods: prospective cohort study of UK secondary care patients with a clinical diagnosis of PG suitable for topical treatment (recruited July 2009 to June 2012). Participants received topical therapy following normal clinical practice (mainly Class I-III topical corticosteroids, tacrolimus 0.03% or 0.1%). Primary outcome: speed of healing at 6 weeks. Secondary outcomes: proportion healed by 6 months; time to healing; global assessment; inflammation; pain; quality-of-life; treatment failure and recurrence. Results: Sixty-six patients (22 to 85 years) were enrolled. Clobetasol propionate 0.05% was the most commonly prescribed therapy. Overall, 28/66 (43.8%) of ulcers healed by 6 months. Median time-to-healing was 145 days (95% CI: 96 days, ∞). Initial ulcer size was a significant predictor of time-to-healing (hazard ratio 0.94 (0.88;80 1.00); p = 0.043). Four patients (15%) had a recurrence. Limitations: No randomised comparator Conclusion: Topical therapy is potentially an effective first-line treatment for PG that avoids possible side effects associated with systemic therapy. It remains unclear whether more severe disease will respond adequately to topical therapy alone

Case Reports1. A Late Presentation of Loeys-Dietz Syndrome: Beware of TGFβ Receptor Mutations in Benign Joint Hypermobility

Background: Thoracic aortic aneurysms (TAA) and dissections are not uncommon causes of sudden death in young adults. Loeys-Dietz syndrome (LDS) is a rare, recently described, autosomal dominant, connective tissue disease characterized by aggressive arterial aneurysms, resulting from mutations in the transforming growth factor beta (TGFβ) receptor genes TGFBR1 and TGFBR2. Mean age at death is 26.1 years, most often due to aortic dissection. We report an unusually late presentation of LDS, diagnosed following elective surgery in a female with a long history of joint hypermobility. Methods: A 51-year-old Caucasian lady complained of chest pain and headache following a dural leak from spinal anaesthesia for an elective ankle arthroscopy. CT scan and echocardiography demonstrated a dilated aortic root and significant aortic regurgitation. MRA demonstrated aortic tortuosity, an infrarenal aortic aneurysm and aneurysms in the left renal and right internal mammary arteries. She underwent aortic root repair and aortic valve replacement. She had a background of long-standing joint pains secondary to hypermobility, easy bruising, unusual fracture susceptibility and mild bronchiectasis. She had one healthy child age 32, after which she suffered a uterine prolapse. Examination revealed mild Marfanoid features. Uvula, skin and ophthalmological examination was normal. Results: Fibrillin-1 testing for Marfan syndrome (MFS) was negative. Detection of a c.1270G > C (p.Gly424Arg) TGFBR2 mutation confirmed the diagnosis of LDS. Losartan was started for vascular protection. Conclusions: LDS is a severe inherited vasculopathy that usually presents in childhood. It is characterized by aortic root dilatation and ascending aneurysms. There is a higher risk of aortic dissection compared with MFS. Clinical features overlap with MFS and Ehlers Danlos syndrome Type IV, but differentiating dysmorphogenic features include ocular hypertelorism, bifid uvula and cleft palate. Echocardiography and MRA or CT scanning from head to pelvis is recommended to establish the extent of vascular involvement. Management involves early surgical intervention, including early valve-sparing aortic root replacement, genetic counselling and close monitoring in pregnancy. Despite being caused by loss of function mutations in either TGFβ receptor, paradoxical activation of TGFβ signalling is seen, suggesting that TGFβ antagonism may confer disease modifying effects similar to those observed in MFS. TGFβ antagonism can be achieved with angiotensin antagonists, such as Losartan, which is able to delay aortic aneurysm development in preclinical models and in patients with MFS. Our case emphasizes the importance of timely recognition of vasculopathy syndromes in patients with hypermobility and the need for early surgical intervention. It also highlights their heterogeneity and the potential for late presentation. Disclosures: The authors have declared no conflicts of interes

Genetic mechanisms of critical illness in COVID-19.

Host-mediated lung inflammation is present1, and drives mortality2, in the critical illness caused by coronavirus disease 2019 (COVID-19). Host genetic variants associated with critical illness may identify mechanistic targets for therapeutic development3. Here we report the results of the GenOMICC (Genetics Of Mortality In Critical Care) genome-wide association study in 2,244 critically ill patients with COVID-19 from 208 UK intensive care units. We have identified and replicated the following new genome-wide significant associations: on chromosome 12q24.13 (rs10735079, P = 1.65 × 10-8) in a gene cluster that encodes antiviral restriction enzyme activators (OAS1, OAS2 and OAS3); on chromosome 19p13.2 (rs74956615, P = 2.3 × 10-8) near the gene that encodes tyrosine kinase 2 (TYK2); on chromosome 19p13.3 (rs2109069, P = 3.98 ×  10-12) within the gene that encodes dipeptidyl peptidase 9 (DPP9); and on chromosome 21q22.1 (rs2236757, P = 4.99 × 10-8) in the interferon receptor gene IFNAR2. We identified potential targets for repurposing of licensed medications: using Mendelian randomization, we found evidence that low expression of IFNAR2, or high expression of TYK2, are associated with life-threatening disease; and transcriptome-wide association in lung tissue revealed that high expression of the monocyte-macrophage chemotactic receptor CCR2 is associated with severe COVID-19. Our results identify robust genetic signals relating to key host antiviral defence mechanisms and mediators of inflammatory organ damage in COVID-19. Both mechanisms may be amenable to targeted treatment with existing drugs. However, large-scale randomized clinical trials will be essential before any change to clinical practice

Whole-genome sequencing reveals host factors underlying critical COVID-19

Critical COVID-19 is caused by immune-mediated inflammatory lung injury. Host genetic variation influences the development of illness requiring critical care1 or hospitalization2–4 after infection with SARS-CoV-2. The GenOMICC (Genetics of Mortality in Critical Care) study enables the comparison of genomes from individuals who are critically ill with those of population controls to find underlying disease mechanisms. Here we use whole-genome sequencing in 7,491 critically ill individuals compared with 48,400 controls to discover and replicate 23 independent variants that significantly predispose to critical COVID-19. We identify 16 new independent associations, including variants within genes that are involved in interferon signalling (IL10RB and PLSCR1), leucocyte differentiation (BCL11A) and blood-type antigen secretor status (FUT2). Using transcriptome-wide association and colocalization to infer the effect of gene expression on disease severity, we find evidence that implicates multiple genes—including reduced expression of a membrane flippase (ATP11A), and increased expression of a mucin (MUC1)—in critical disease. Mendelian randomization provides evidence in support of causal roles for myeloid cell adhesion molecules (SELE, ICAM5 and CD209) and the coagulation factor F8, all of which are potentially druggable targets. Our results are broadly consistent with a multi-component model of COVID-19 pathophysiology, in which at least two distinct mechanisms can predispose to life-threatening disease: failure to control viral replication; or an enhanced tendency towards pulmonary inflammation and intravascular coagulation. We show that comparison between cases of critical illness and population controls is highly efficient for the detection of therapeutically relevant mechanisms of disease

Rehabilitation versus surgical reconstruction for non-acute anterior cruciate ligament injury (ACL SNNAP): a pragmatic randomised controlled trial

BackgroundAnterior cruciate ligament (ACL) rupture is a common debilitating injury that can cause instability of the knee. We aimed to investigate the best management strategy between reconstructive surgery and non-surgical treatment for patients with a non-acute ACL injury and persistent symptoms of instability.MethodsWe did a pragmatic, multicentre, superiority, randomised controlled trial in 29 secondary care National Health Service orthopaedic units in the UK. Patients with symptomatic knee problems (instability) consistent with an ACL injury were eligible. We excluded patients with meniscal pathology with characteristics that indicate immediate surgery. Patients were randomly assigned (1:1) by computer to either surgery (reconstruction) or rehabilitation (physiotherapy but with subsequent reconstruction permitted if instability persisted after treatment), stratified by site and baseline Knee Injury and Osteoarthritis Outcome Score—4 domain version (KOOS4). This management design represented normal practice. The primary outcome was KOOS4 at 18 months after randomisation. The principal analyses were intention-to-treat based, with KOOS4 results analysed using linear regression. This trial is registered with ISRCTN, ISRCTN10110685, and ClinicalTrials.gov, NCT02980367.FindingsBetween Feb 1, 2017, and April 12, 2020, we recruited 316 patients. 156 (49%) participants were randomly assigned to the surgical reconstruction group and 160 (51%) to the rehabilitation group. Mean KOOS4 at 18 months was 73·0 (SD 18·3) in the surgical group and 64·6 (21·6) in the rehabilitation group. The adjusted mean difference was 7·9 (95% CI 2·5–13·2; p=0·0053) in favour of surgical management. 65 (41%) of 160 patients allocated to rehabilitation underwent subsequent surgery according to protocol within 18 months. 43 (28%) of 156 patients allocated to surgery did not receive their allocated treatment. We found no differences between groups in the proportion of intervention-related complications.InterpretationSurgical reconstruction as a management strategy for patients with non-acute ACL injury with persistent symptoms of instability was clinically superior and more cost-effective in comparison with rehabilitation management