Evaluating the clinical and cost-effectiveness of a conservative approach to oxygen therapy for invasively ventilated adults in intensive care: Protocol for the UK-ROX trial.

Background: In the United Kingdom, around 184,000 adults are admitted to an intensive care unit (ICU) each year with over 30% receiving mechanical ventilation. Oxygen is the commonest therapeutic intervention provided to these patients but it is unclear how much oxygen should be administered for the best clinical outcomes. Methods: The UK-ROX trial will evaluate the clinical and cost-effectiveness of conservative oxygen therapy (the minimum oxygen concentration required to maintain an oxygen saturation of 90% ± 2%) versus usual oxygen therapy in critically ill adults receiving supplemental oxygen when invasively mechanically ventilated in ICUs in England, Wales and Northern Ireland. The trial will recruit 16,500 patients from approximately 100 UK adult ICUs. Using a deferred consent model, enrolled participants will be randomly allocated (1:1) to conservative or usual oxygen therapy until ICU discharge or 90 days after randomisation. Objectives: The primary clinical outcome is all cause mortality at 90 days following randomisation. Discussion: The UK-ROX trial has received ethical approval from the South Central - Oxford C Research Ethics Committee (Reference: 20/SC/0423) and the Confidentiality Advisory Group (Reference: 22/CAG/0154). The trial commenced in May 2021 and, at the time of publication, 95 sites had opened to recruitment.

Assessment of the comparative agreement between chest radiographs and CT scans in intensive care units.

PURPOSE: Chest radiographs in critically ill patients can be difficult to interpret due to technical and clinical factors. We sought to determine the agreement of chest radiographs and CT scans, and the inter-observer variation of chest radiograph interpretation, in intensive care units (ICUs). METHODS: Chest radiographs and corresponding thoracic computerised tomography (CT) scans (as reference standard) were collected from 45 ICU patients. All radiographs were analysed by 20 doctors (radiology consultants, radiology trainees, ICU consultants, ICU trainees) from 4 different centres, blinded to CT results. Specificity/sensitivity were determined for pleural effusion, lobar collapse and consolidation/atelectasis. Separately, Fleiss' kappa for multiple raters was used to determine inter-observer variation for chest radiographs. RESULTS: The median sensitivity and specificity of chest radiographs for detecting abnormalities seen on CTs scans were 43.2% and 85.9% respectively. Diagnostic sensitivity for pleural effusion was significantly higher among radiology consultants but no specialty/experience distinctions were observed for specificity. Median inter-observer kappa coefficient among assessors was 0.295 ("fair"). CONCLUSIONS: Chest radiographs commonly miss important radiological features in critically ill patients. Inter-observer agreement in chest radiograph interpretation is only "fair". Consultant radiologists are least likely to miss thoracic radiological abnormalities. The consequences of misdiagnosis by chest radiographs remain to be determined.

Tissue factor expression in monocyte subsets during human immunothrombosis, endotoxemia and sepsis.

INTRODUCTION: Tissue factor expression on monocytes is implicated in the pathophysiology of sepsis-induced coagulopathy. How tissue factor is expressed by monocyte subsets (classical, intermediate and non-classical) is unknown. METHODS: Monocytic tissue factor surface expression was investigated during three conditions. Primary human monocytes and microvascular endothelial cell co-cultures were used for in vitro studies. Volunteers received a bolus of lipopolysaccharide (2 ng/kg) to induce endotoxemia. Patients with sepsis, or controls with critical illness unrelated to sepsis, were recruited from four intensive care units. RESULTS: Contact with endothelium and stimulation with lipopolysaccharide reduced the proportion of intermediate monocytes. Lipopolysaccharide increased tissue factor surface expression on classical and non-classical monocytes. Endotoxemia induced profound, transient monocytopenia, along with activation of coagulation pathways. In the remaining circulating monocytes, tissue factor was up-regulated in intermediate monocytes, though approximately 60 % of individuals (responders) up-regulated tissue factor across all monocyte subsets. In critically ill patients, tissue factor expression on intermediate and non-classical monocytes was significantly higher in patients with established sepsis than among non-septic patients. Upon recovery of sepsis, expression of tissue factor increased significantly in classical monocytes. CONCLUSION: Tissue factor expression in monocyte subsets varies significantly during health, endotoxemia and sepsis.

Repair of Acute Respiratory Distress Syndrome in COVID-19 by Stromal Cells (REALIST-COVID Trial): A Multicenter, Randomized, Controlled Clinical Trial.

Rationale: Mesenchymal stromal cells (MSCs) may modulate inflammation, promoting repair in coronavirus disease (COVID-19)-related acute respiratory distress syndrome (ARDS). Objectives: We investigated the safety and efficacy of ORBCEL-C (CD362 [cluster of differentiation 362]-enriched, umbilical cord-derived MSCs) in COVID-19-related ARDS. Methods: In this multicenter, randomized, double-blind, allocation-concealed, placebo-controlled trial (NCT03042143), patients with moderate to severe COVID-19-related ARDS were randomized to receive ORBCEL-C (400 million cells) or placebo (Plasma-Lyte 148). The primary safety and efficacy outcomes were the incidence of serious adverse events and oxygenation index at Day 7, respectively. Secondary outcomes included respiratory compliance, driving pressure, PaO2:FiO2 ratio, and Sequential Organ Failure Assessment score. Clinical outcomes relating to duration of ventilation, lengths of ICU and hospital stays, and mortality were collected. Long-term follow-up included diagnosis of interstitial lung disease at 1 year and significant medical events and mortality at 2 years. Transcriptomic analysis was performed on whole blood at Days 0, 4, and 7. Measurements and Main Results: Sixty participants were recruited (final analysis: n = 30 received ORBCEL-C, n = 29 received placebo; 1 participant in the placebo group withdrew consent). Six serious adverse events occurred in the ORBCEL-C group and three in the placebo group (risk ratio, 2.9 [95% confidence interval, 0.6-13.2]; P = 0.25). Day 7 mean (SD) oxygenation index did not differ (ORBCEL-C, 98.3 [57.2] cm H2O/kPa; placebo, 96.6 [67.3] cm H2O/kPa). There were no differences in secondary surrogate outcomes or in mortality at Day 28, Day 90, 1 year, or 2 years. There was no difference in the prevalence of interstitial lung disease at 1 year or significant medical events up to 2 years. ORBCEL-C modulated the peripheral blood transcriptome. Conclusion: ORBCEL-C MSCs were safe in subjects with moderate to severe COVID-19-related ARDS but did not improve surrogates of pulmonary organ dysfunction.

Tracheostomy in children is associated with neutrophilic airway inflammation.

BACKGROUND: Tracheostomies in children are associated with significant morbidity, poor quality of life, excess healthcare costs and excess mortality. The underlying mechanisms facilitating adverse respiratory outcomes in tracheostomised children are poorly understood. We aimed to characterise airway host defence in tracheostomised children using serial molecular analyses. METHODS: Tracheal aspirates, tracheal cytology brushings and nasal swabs were prospectively collected from children with a tracheostomy and controls. Transcriptomic, proteomic and metabolomic methods were applied to characterise the impact of tracheostomy on host immune response and the airway microbiome. RESULTS: Children followed up serially from the time of tracheostomy up to 3 months postprocedure (n=9) were studied. A further cohort of children with a long-term tracheostomy were also enrolled (n=24). Controls (n=13) comprised children without a tracheostomy undergoing bronchoscopy. Long-term tracheostomy was associated with airway neutrophilic inflammation, superoxide production and evidence of proteolysis when compared with controls. Reduced airway microbial diversity was established pre-tracheostomy and sustained thereafter. CONCLUSIONS: Long-term childhood tracheostomy is associated with a inflammatory tracheal phenotype characterised by neutrophilic inflammation and the ongoing presence of potential respiratory pathogens. These findings suggest neutrophil recruitment and activation as potential exploratory targets in seeking to prevent recurrent airway complications in this vulnerable group of patients.

A human model of bilateral pulmonary vein sampling to assess the effects of one-lung ventilation on neutrophil function.

BACKGROUND: Neutrophil activation drives lung complications after cardiopulmonary bypass (CPB). Evidence suggests the healthy, ventilated lung may beneficially re-condition pro-inflammatory neutrophils. However, evidence in humans is lacking, due to a paucity of good models. CPB with simultaneous central venous and bilateral pulmonary vein sampling provides an opportunity to model effects of one-lung ventilation. The study's primary objectives were to establish a model of intra-operative, bilateral pulmonary vein sampling and to determine whether neutrophil function differed after passing through inflated or deflated lungs. METHODS: Seventeen patients having "on pump" coronary artery bypass grafting (CABG) with one-lung ventilation (in two cohorts with tidal volume 2ml kg-1 and FiO2 0.21, or tidal volume 4 ml kg-1 and FiO2 0.5 respectively) were recruited. Cohort 1 consisted of 9 patients (7 male, median age 62.0 years) and Cohort 2 consisted of 8 male patients (median age 65.5 years). Recruitment was via prospective screening of scheduled elective and non-elective CABG procedures with cardiopulmonary bypass. Each patient had five blood samples taken-central venous blood pre-operatively; central venous blood pre-CPB; central venous blood post-CPB; pulmonary venous blood draining the ventilated lung post-CPB; and pulmonary venous blood draining the deflated lung post-CPB. Neutrophil phagocytosis and priming status were quantified. Plasma cytokines were measured. RESULTS: Phagocytosis and priming were not significantly different in neutrophils returning from the ventilated lung as compared to the non-ventilated lung. Plasma IL-6, IL-8 and IL-10 were significantly elevated by CPB. CONCLUSIONS: The intra-operative, bilateral pulmonary vein sampling model provides unique opportunities to assess biological effects of interventions to one lung, with the other lung acting as an internal control. Single-lung ventilation during CPB had no significant effects on neutrophil function.

Hypoxia shapes the immune landscape in lung injury and promotes the persistence of inflammation.

Hypoxemia is a defining feature of acute respiratory distress syndrome (ARDS), an often-fatal complication of pulmonary or systemic inflammation, yet the resulting tissue hypoxia, and its impact on immune responses, is often neglected. In the present study, we have shown that ARDS patients were hypoxemic and monocytopenic within the first 48 h of ventilation. Monocytopenia was also observed in mouse models of hypoxic acute lung injury, in which hypoxemia drove the suppression of type I interferon signaling in the bone marrow. This impaired monopoiesis resulted in reduced accumulation of monocyte-derived macrophages and enhanced neutrophil-mediated inflammation in the lung. Administration of colony-stimulating factor 1 in mice with hypoxic lung injury rescued the monocytopenia, altered the phenotype of circulating monocytes, increased monocyte-derived macrophages in the lung and limited injury. Thus, tissue hypoxia altered the dynamics of the immune response to the detriment of the host and interventions to address the aberrant response offer new therapeutic strategies for ARDS.

Effect of Noninvasive Respiratory Strategies on Intubation or Mortality Among Patients With Acute Hypoxemic Respiratory Failure and COVID-19: The RECOVERY-RS Randomized Clinical Trial.

Importance: Continuous positive airway pressure (CPAP) and high-flow nasal oxygen (HFNO) have been recommended for acute hypoxemic respiratory failure in patients with COVID-19. Uncertainty exists regarding the effectiveness and safety of these noninvasive respiratory strategies. Objective: To determine whether either CPAP or HFNO, compared with conventional oxygen therapy, improves clinical outcomes in hospitalized patients with COVID-19-related acute hypoxemic respiratory failure. Design, Setting, and Participants: A parallel group, adaptive, randomized clinical trial of 1273 hospitalized adults with COVID-19-related acute hypoxemic respiratory failure. The trial was conducted between April 6, 2020, and May 3, 2021, across 48 acute care hospitals in the UK and Jersey. Final follow-up occurred on June 20, 2021. Interventions: Adult patients were randomized to receive CPAP (n = 380), HFNO (n = 418), or conventional oxygen therapy (n = 475). Main Outcomes and Measures: The primary outcome was a composite of tracheal intubation or mortality within 30 days. Results: The trial was stopped prematurely due to declining COVID-19 case numbers in the UK and the end of the funded recruitment period. Of the 1273 randomized patients (mean age, 57.4 [95% CI, 56.7 to 58.1] years; 66% male; 65% White race), primary outcome data were available for 1260. Crossover between interventions occurred in 17.1% of participants (15.3% in the CPAP group, 11.5% in the HFNO group, and 23.6% in the conventional oxygen therapy group). The requirement for tracheal intubation or mortality within 30 days was significantly lower with CPAP (36.3%; 137 of 377 participants) vs conventional oxygen therapy (44.4%; 158 of 356 participants) (absolute difference, -8% [95% CI, -15% to -1%], P = .03), but was not significantly different with HFNO (44.3%; 184 of 415 participants) vs conventional oxygen therapy (45.1%; 166 of 368 participants) (absolute difference, -1% [95% CI, -8% to 6%], P = .83). Adverse events occurred in 34.2% (130/380) of participants in the CPAP group, 20.6% (86/418) in the HFNO group, and 13.9% (66/475) in the conventional oxygen therapy group. Conclusions and Relevance: Among patients with acute hypoxemic respiratory failure due to COVID-19, an initial strategy of CPAP significantly reduced the risk of tracheal intubation or mortality compared with conventional oxygen therapy, but there was no significant difference between an initial strategy of HFNO compared with conventional oxygen therapy. The study may have been underpowered for the comparison of HFNO vs conventional oxygen therapy, and early study termination and crossover among the groups should be considered when interpreting the findings. Trial Registration: isrctn.org Identifier: ISRCTN16912075.

Feasibility of conservative fluid administration and deresuscitation compared with usual care in critical illness: the Role of Active Deresuscitation After Resuscitation-2 (RADAR-2) randomised clinical trial.

PURPOSE: Fluid overload is common in critical illness and is associated with mortality. This study investigated the feasibility of a randomised trial comparing conservative fluid administration and deresuscitation (active removal of accumulated fluid using diuretics or ultrafiltration) with usual care in critical illness. METHODS: Open-label, parallel-group, allocation-concealed randomised clinical feasibility trial. Mechanically ventilated adult patients expected to require critical care beyond the next calendar day were enrolled between 24 and 48 h following admission to the intensive care unit (ICU). Patients were randomised to either a 2-stage fluid strategy comprising conservative fluid administration and, if fluid overload was present, active deresuscitation, or usual care. The primary endpoint was fluid balance in the 24 h up to the start of study day 3. Secondary endpoints included cumulative fluid balance, mortality, and duration of mechanical ventilation. RESULTS: One hundred and eighty patients were randomised. After withdrawal of 1 patient, 89 patients assigned to the intervention were compared with 90 patients assigned to the usual care group. The mean plus standard deviation (SD) 24-h fluid balance up to study day 3 was lower in the intervention group (- 840 ± 1746 mL) than the usual care group (+ 130 ± 1401 mL; P < 0.01). Cumulative fluid balance was lower in the intervention group at days 3 and 5. Overall, clinical outcomes did not differ significantly between the two groups, although the point estimate for 30-day mortality favoured the usual care group [intervention arm: 19 of 90 (21.6%) versus usual care: 14 of 89 (15.6%), P = 0.32]. Baseline imbalances between groups and lack of statistical power limit interpretation of clinical outcomes. CONCLUSIONS: A strategy of conservative fluid administration and active deresuscitation is feasible, reduces fluid balance compared with usual care, and may cause benefit or harm. In view of wide variations in contemporary clinical practice, large, adequately powered trials investigating the clinical effectiveness of conservative fluid strategies in critically ill patients are warranted.

Phosphoinositide 3-Kinase δ Inhibition Improves Neutrophil Bacterial Killing in Critically Ill Patients at High Risk of Infection.

Acquired neutrophil dysfunction frequently develops during critical illness, independently increasing the risk for intensive care unit-acquired infection. PI3Kδ is implicated in driving neutrophil dysfunction and can potentially be targeted pharmacologically. The aims of this study were to determine whether PI3Kδ inhibition reverses dysfunction in neutrophils from critically ill patients and to describe potential mechanisms. Neutrophils were isolated from blood taken from critically ill patients requiring intubation and mechanical ventilation, renal support, or blood pressure support. In separate validation experiments, neutrophil dysfunction was induced pharmacologically in neutrophils from healthy volunteers. Phagocytosis and bacterial killing assays were performed, and activity of RhoA and protein kinase A (PKA) was assessed. Inhibitors of PI3Kδ, 3-phosphoinositide-dependent protein kinase-1 (PDK1), and PKA were used to determine mechanisms of neutrophil dysfunction. Sixty-six patients were recruited. In the 27 patients (40.9%) with impaired neutrophil function, PI3Kδ inhibition consistently improved function and significantly increased bacterial killing. These findings were validated in neutrophils from healthy volunteers with salbutamol-induced dysfunction and extended to demonstrate that PI3Kδ inhibition restored killing of clinical isolates of nine pathogens commonly associated with intensive care unit-acquired infection. PI3Kδ activation was associated with PDK1 activation, which in turn phosphorylated PKA, which drove phosphorylation and inhibition of the key regulator of neutrophil phagocytosis, RhoA. These data indicate that, in a significant proportion of critically ill patients, PI3Kδ inhibition can improve neutrophil function through PDK1- and PKA-dependent processes, suggesting that therapeutic use of PI3Kδ inhibitors warrants investigation in this setting.

Activation of the hypothalamic-pituitary-adrenal axis by exogenous and endogenous GDF15.

An acute increase in the circulating concentration of glucocorticoid hormones is essential for the survival of severe somatic stresses. Circulating concentrations of GDF15, a hormone that acts in the brain to reduce food intake, are frequently elevated in stressful states. We now report that GDF15 potently activates the hypothalamic-pituitary-adrenal (HPA) axis in mice and rats. A blocking antibody to the GDNF-family receptor α-like receptor completely prevented the corticosterone response to GDF15 administration. In wild-type mice exposed to a range of stressful stimuli, circulating levels of both corticosterone and GDF15 rose acutely. In the case of Escherichia coli or lipopolysaccharide injections, the vigorous proinflammatory cytokine response elicited was sufficient to produce a near-maximal HPA response, regardless of the presence or absence of GDF15. In contrast, the activation of the HPA axis seen in wild-type mice in response to the administration of genotoxic or endoplasmic reticulum toxins, which do not provoke a marked rise in cytokines, was absent in Gdf15-/- mice. In conclusion, consistent with its proposed role as a sentinel hormone, endogenous GDF15 is required for the activation of the protective HPA response to toxins that do not induce a substantial cytokine response. In the context of efforts to develop GDF15 as an antiobesity therapeutic, these findings identify a biomarker of target engagement and a previously unrecognized pharmacodynamic effect, which will require monitoring in human studies.

COVID-19 and pneumothorax: a multicentre retrospective case series.

INTRODUCTION: Pneumothorax and pneumomediastinum have both been noted to complicate cases of coronavirus disease 2019 (COVID-19) requiring hospital admission. We report the largest case series yet described of patients with both these pathologies (including nonventilated patients). METHODS: Cases were collected retrospectively from UK hospitals with inclusion criteria limited to a diagnosis of COVID-19 and the presence of either pneumothorax or pneumomediastinum. Patients included in the study presented between March and June 2020. Details obtained from the medical record included demographics, radiology, laboratory investigations, clinical management and survival. RESULTS: 71 patients from 16 centres were included in the study, of whom 60 had pneumothoraces (six with pneumomediastinum in addition) and 11 had pneumomediastinum alone. Two of these patients had two distinct episodes of pneumothorax, occurring bilaterally in sequential fashion, bringing the total number of pneumothoraces included to 62. Clinical scenarios included patients who had presented to hospital with pneumothorax, patients who had developed pneumothorax or pneumomediastinum during their inpatient admission with COVID-19 and patients who developed their complication while intubated and ventilated, either with or without concurrent extracorporeal membrane oxygenation. Survival at 28 days was not significantly different following pneumothorax (63.1±6.5%) or isolated pneumomediastinum (53.0±18.7%; p=0.854). The incidence of pneumothorax was higher in males. 28-day survival was not different between the sexes (males 62.5±7.7% versus females 68.4±10.7%; p=0.619). Patients aged ≥70 years had a significantly lower 28-day survival than younger individuals (≥70 years 41.7±13.5% survival versus <70 years 70.9±6.8% survival; p=0.018 log-rank). CONCLUSION: These cases suggest that pneumothorax is a complication of COVID-19. Pneumothorax does not seem to be an independent marker of poor prognosis and we encourage continuation of active treatment where clinically possible.

Human lipopolysaccharide models provide mechanistic and therapeutic insights into systemic and pulmonary inflammation.

Inflammation is a key feature in the pathogenesis of sepsis and acute respiratory distress syndrome (ARDS). Sepsis and ARDS continue to be associated with high mortality. A key contributory factor is the rudimentary understanding of the early events in pulmonary and systemic inflammation in humans, which are difficult to study in clinical practice, as they precede the patient's presentation to medical services. Lipopolysaccharide (LPS), a constituent of the outer membrane of Gram-negative bacteria, is a trigger of inflammation and the dysregulated host response in sepsis. Human LPS models deliver a small quantity of LPS to healthy volunteers, triggering an inflammatory response and providing a window to study early inflammation in humans. This allows biological/mechanistic insights to be made and new therapeutic strategies to be tested in a controlled, reproducible environment from a defined point in time. We review the use of human LPS models, focussing on the underlying mechanistic insights that have been gained by studying the response to intravenous and pulmonary LPS challenge. We discuss variables that may influence the response to LPS before considering factors that should be considered when designing future human LPS studies.

Biomarker-guided antibiotic stewardship in suspected ventilator-associated pneumonia (VAPrapid2): a randomised controlled trial and process evaluation.

BACKGROUND: Ventilator-associated pneumonia is the most common intensive care unit (ICU)-acquired infection, yet accurate diagnosis remains difficult, leading to overuse of antibiotics. Low concentrations of IL-1ß and IL-8 in bronchoalveolar lavage fluid have been validated as effective markers for exclusion of ventilator-associated pneumonia. The VAPrapid2 trial aimed to determine whether measurement of bronchoalveolar lavage fluid IL-1ß and IL-8 could effectively and safely improve antibiotic stewardship in patients with clinically suspected ventilator-associated pneumonia. METHODS: VAPrapid2 was a multicentre, randomised controlled trial in patients admitted to 24 ICUs from 17 National Health Service hospital trusts across England, Scotland, and Northern Ireland. Patients were screened for eligibility and included if they were 18 years or older, intubated and mechanically ventilated for at least 48 h, and had suspected ventilator-associated pneumonia. Patients were randomly assigned (1:1) to biomarker-guided recommendation on antibiotics (intervention group) or routine use of antibiotics (control group) using a web-based randomisation service hosted by Newcastle Clinical Trials Unit. Patients were randomised using randomly permuted blocks of size four and six and stratified by site, with allocation concealment. Clinicians were masked to patient assignment for an initial period until biomarker results were reported. Bronchoalveolar lavage was done in all patients, with concentrations of IL-1ß and IL-8 rapidly determined in bronchoalveolar lavage fluid from patients randomised to the biomarker-based antibiotic recommendation group. If concentrations were below a previously validated cutoff, clinicians were advised that ventilator-associated pneumonia was unlikely and to consider discontinuing antibiotics. Patients in the routine use of antibiotics group received antibiotics according to usual practice at sites. Microbiology was done on bronchoalveolar lavage fluid from all patients and ventilator-associated pneumonia was confirmed by at least 104 colony forming units per mL of bronchoalveolar lavage fluid. The primary outcome was the distribution of antibiotic-free days in the 7 days following bronchoalveolar lavage. Data were analysed on an intention-to-treat basis, with an additional per-protocol analysis that excluded patients randomly assigned to the intervention group who defaulted to routine use of antibiotics because of failure to return an adequate biomarker result. An embedded process evaluation assessed factors influencing trial adoption, recruitment, and decision making. This study is registered with ISRCTN, ISRCTN65937227, and ClinicalTrials.gov, NCT01972425. FINDINGS: Between Nov 6, 2013, and Sept 13, 2016, 360 patients were screened for inclusion in the study. 146 patients were ineligible, leaving 214 who were recruited to the study. Four patients were excluded before randomisation, meaning that 210 patients were randomly assigned to biomarker-guided recommendation on antibiotics (n=104) or routine use of antibiotics (n=106). One patient in the biomarker-guided recommendation group was withdrawn by the clinical team before bronchoscopy and so was excluded from the intention-to-treat analysis. We found no significant difference in the primary outcome of the distribution of antibiotic-free days in the 7 days following bronchoalveolar lavage in the intention-to-treat analysis (p=0·58). Bronchoalveolar lavage was associated with a small and transient increase in oxygen requirements. Established prescribing practices, reluctance for bronchoalveolar lavage, and dependence on a chain of trial-related procedures emerged as factors that impaired trial processes. INTERPRETATION: Antibiotic use remains high in patients with suspected ventilator-associated pneumonia. Antibiotic stewardship was not improved by a rapid, highly sensitive rule-out test. Prescribing culture, rather than poor test performance, might explain this absence of effect. FUNDING: UK Department of Health and the Wellcome Trust.