title: Rapid detection of health-care-associated bloodstream infection in critical care using multipathogen real-time polymerase chain reaction technology: a diagnostic accuracy study and systematic review.
abstract: BACKGROUND
  There is growing interest in the potential utility of real-time polymerase chain reaction (PCR) in diagnosing bloodstream infection by detecting pathogen deoxyribonucleic acid (DNA) in blood samples within a few hours. SeptiFast (Roche Diagnostics GmBH, Mannheim, Germany) is a multipathogen probe-based system targeting ribosomal DNA sequences of bacteria and fungi. It detects and identifies the commonest pathogens causing bloodstream infection. As background to this study, we report a systematic review of Phase III diagnostic accuracy studies of SeptiFast, which reveals uncertainty about its likely clinical utility based on widespread evidence of deficiencies in study design and reporting with a high risk of bias.
  
  
  OBJECTIVE
  Determine the accuracy of SeptiFast real-time PCR for the detection of health-care-associated bloodstream infection, against standard microbiological culture.
  
  
  METHODS
  Prospective multicentre Phase III clinical diagnostic accuracy study using the standards for the reporting of diagnostic accuracy studies criteria.
  
  
  METHODS
  Critical care departments within NHS hospitals in the north-west of England.
  
  
  METHODS
  Adult patients requiring blood culture (BC) when developing new signs of systemic inflammation.
  
  
  METHODS
  SeptiFast real-time PCR results at species/genus level compared with microbiological culture in association with independent adjudication of infection. Metrics of diagnostic accuracy were derived including sensitivity, specificity, likelihood ratios and predictive values, with their 95% confidence intervals (CIs). Latent class analysis was used to explore the diagnostic performance of culture as a reference standard.
  
  
  RESULTS
  Of 1006 new patient episodes of systemic inflammation in 853 patients, 922 (92%) met the inclusion criteria and provided sufficient information for analysis. Index test assay failure occurred on 69 (7%) occasions. Adult patients had been exposed to a median of 8 days (interquartile range 4-16 days) of hospital care, had high levels of organ support activities and recent antibiotic exposure. SeptiFast real-time PCR, when compared with culture-proven bloodstream infection at species/genus level, had better specificity (85.8%, 95% CI 83.3% to 88.1%) than sensitivity (50%, 95% CI 39.1% to 60.8%). When compared with pooled diagnostic metrics derived from our systematic review, our clinical study revealed lower test accuracy of SeptiFast real-time PCR, mainly as a result of low diagnostic sensitivity. There was a low prevalence of BC-proven pathogens in these patients (9.2%, 95% CI 7.4% to 11.2%) such that the post-test probabilities of both a positive (26.3%, 95% CI 19.8% to 33.7%) and a negative SeptiFast test (5.6%, 95% CI 4.1% to 7.4%) indicate the potential limitations of this technology in the diagnosis of bloodstream infection. However, latent class analysis indicates that BC has a low sensitivity, questioning its relevance as a reference test in this setting. Using this analysis approach, the sensitivity of the SeptiFast test was low but also appeared significantly better than BC. Blood samples identified as positive by either culture or SeptiFast real-time PCR were associated with a high probability (> 95%) of infection, indicating higher diagnostic rule-in utility than was apparent using conventional analyses of diagnostic accuracy.
  
  
  CONCLUSIONS
  SeptiFast real-time PCR on blood samples may have rapid rule-in utility for the diagnosis of health-care-associated bloodstream infection but the lack of sensitivity is a significant limiting factor. Innovations aimed at improved diagnostic sensitivity of real-time PCR in this setting are urgently required. Future work recommendations include technology developments to improve the efficiency of pathogen DNA extraction and the capacity to detect a much broader range of pathogens and drug resistance genes and the application of new statistical approaches able to more reliably assess test performance in situation where the reference standard (e.g. blood culture in the setting of high antimicrobial use) is prone to error.
  
  
  BACKGROUND
  The systematic review is registered as PROSPERO CRD42011001289.
  
  
  BACKGROUND
  The National Institute for Health Research Health Technology Assessment programme. Professor Daniel McAuley and Professor Gavin D Perkins contributed to the systematic review through their funded roles as codirectors of the Intensive Care Foundation (UK).
authors.affiliation: Infection, Injury and Inflammation Research Group, Salford Royal NHS Foundation Trust, Manchester Academic Health Sciences Centre (MAHSC), University of Manchester, Salford, UK.
authors.forename: Geoffrey
authors.initials: G
authors.lastname: Warhurst
authors.affiliation: Centre for Biostatistics, Institute of Population Health, Faculty of Medical and Human Sciences, Manchester Academic Health Sciences Centre (MAHSC), University of Manchester, Salford, UK.
authors.forename: Graham
authors.initials: G
authors.lastname: Dunn
authors.affiliation: Microbiology Department, Salford Royal NHS Foundation Trust, Manchester Academic Health Sciences Centre (MAHSC), University of Manchester, Salford, UK.
authors.forename: Paul
authors.initials: P
authors.lastname: Chadwick
authors.affiliation: Centre for Infection and Immunity, Queen's University Belfast, Belfast, UK.
authors.forename: Bronagh
authors.initials: B
authors.lastname: Blackwood
authors.affiliation: Centre for Infection and Immunity, Queen's University Belfast, Belfast, UK.
authors.forename: Daniel
authors.initials: D
authors.lastname: McAuley
authors.affiliation: Clinical Trials Unit, Medical School, Warwick University, Coventry, UK.
authors.forename: Gavin D
authors.initials: GD
authors.lastname: Perkins
authors.affiliation: Medical Microbiology, Royal Victoria Hospital, Belfast, UK.
authors.forename: Ronan
authors.initials: R
authors.lastname: McMullan
authors.affiliation: Clinical Trials Unit, Medical School, Warwick University, Coventry, UK.
authors.forename: Simon
authors.initials: S
authors.lastname: Gates
authors.affiliation: Institue of Inflammation and Repair, Faculty of Medical and Human Sciences, Manchester Academic Health Sciences Centre (MAHSC), University of Manchester, Salford, UK.
authors.forename: Andrew
authors.initials: A
authors.lastname: Bentley
authors.affiliation: Nuffield Department of Clinical Neurosciences, John Radcliffe Hospital, Oxford, UK.
authors.forename: Duncan
authors.initials: D
authors.lastname: Young
authors.affiliation: Infection, Injury and Inflammation Research Group, Salford Royal NHS Foundation Trust, Manchester Academic Health Sciences Centre (MAHSC), University of Manchester, Salford, UK.
authors.forename: Gordon L
authors.initials: GL
authors.lastname: Carlson
authors.affiliation: Infection, Injury and Inflammation Research Group, Salford Royal NHS Foundation Trust, Manchester Academic Health Sciences Centre (MAHSC), University of Manchester, Salford, UK.
authors.forename: Paul
authors.initials: P
authors.lastname: Dark
mesh_terms: D016470:Bacteremia / Q000175:diagnosis*; D003422:Critical Care; D003428:Cross Infection / Q000175:diagnosis*; D004739:England; D005188:False Negative Reactions; D005189:False Positive Reactions; D006801:Humans; D011446:Prospective Studies; D060888:Real-Time Polymerase Chain Reaction / Q000191:economics / Q000592:standards*; D012680:Sensitivity and Specificity; D013222:State Medicine; D013673:Technology Assessment, Biomedical; D013997:Time Factors