Volume 13 Supplement 4

Sepsis 2009

Open Access

Performance evaluation and further development of the PCR and microarray-based Prove-it™ Sepsis assay

  • P Tissari1,
  • E Tarkka1,
  • S Mero1,
  • L Savolainen1,
  • M Vaara1,
  • A Zumla2,
  • J Huggett2,
  • C Carder2,
  • V Gant2,
  • A Aittakorpi3,
  • S Laakso3,
  • M Lindfors3,
  • P Piiparinen3,
  • N Kumlin3,
  • H Piiparinen3 and
  • M Mäki3
Critical Care200913(Suppl 4):P3

https://doi.org/10.1186/cc8059

Published: 11 November 2009

Introduction

The Prove-it™ Sepsis assay is a rapid, broad-range PCR and microarray-based assay designed to identify the majority of sepsis-causing bacteria from positive blood cultures. The pathogen panel covers 50 Gram-negative and Gram-positive bacterial species (Table 1). It also reports methicillin resistance by detecting the mecA gene. The assay time is 3 hours. Our objective was to conduct a performance evaluation study for Prove-it™ Sepsis according to the EN 13612-standard (Performance evaluation of in vitro diagnostic medical devices) and to compare obtained results with those of current culture-based diagnostics. We evaluated the sensitivity, specificity and time to result of Prove-it™ Sepsis in two major teaching hospitals in Helsinki and London.
Table 1

Bacteria and an antibacterial resistance marker identified by the Prove-it™ Sepsis assay

Gram-negative

Gram-positive

Antibacterial resistance

Neisseria meningitidis

Staphylococcus aureus

Methicillin resistance marker mecA

Enterobacter aerogenes

Staphylococcus epidermidis

 

Enterobacter cloacae

Coagulase-negative Staphylococcus d

 

Escherichia coli

Streptococcus pyogenes

 

Klebsiella oxytoca

Streptococcus agalactiae

 

Klebsiella pneumoniae

Streptococcus dysgalactiae subsp. equisimilis

 

Proteus mirabilis

Streptococcus pneumoniae

 

Proteus vulgaris

Enterococcus faecalis

 

Salmonella enterica subsp. entericaa

Enterococcus faecium

 

Serratia marcescens

Listeria monocytogenes

 

Enterobacteriaceae family b

Clostridium perfringens

 

Acinetobacter baumannii

  

Pseudomonas aeruginosa

  

Stenotrophomonas maltophilia

  

Haemophilus influenzae

  

Campylobacter jejuni/coli

  

Bacteroides fragilis groupc

  

aSalmonella enterica subsp. enterica covers at least the following serovars: Enteritidis, Oranienburg, Othmarschen, Paratyphi, Stanley, Typhi, Typhimurium, Virchow, Group A, B, C, D. bEnterobacteriaceae covers at least the following species: Citrobacter amalonaticus, Citrobacter freundii, Citrobacter koseri, Citrobacter braakii, Enterobacter hormaechei, Enterobacter sakazakii, Kluyvera intermedia, Morganella morganii, Pantoea agglomerans, Providencia rettgeri, Providencia stuartii, Yersinia enterocolitica, Yersinia pseudotuberculosis. cBacteroides fragilis covers at least the following species: B. fragilis, B. vulgatus, B. thetaiotaomicron. dCoagulase-negative Staphylococcus covers at least the following species: S. capitis, S. lugdunensis, S. haemolyticus, S. hominis, S. saprophyticus, S. warneri, S. xylosus.

Materials

A total of 3,318 blood samples from patients with suspected sepsis were collected. Blood culture bottles of BacT/ALERT 3D (bioMérieux) and BACTEC 924 (Becton Dickinson) were incubated for a total of 6 days or until flagged as positive.

Methods

DNA was extracted from blood cultures using the automated DNA extraction instrument easyMAG (bioMérieux) prior to the Prove-it™ Sepsis assay. Conventional blood culture was conducted in parallel and results were only revealed for comparison at the statistical analysis stage. Discordant results were studied by DNA sequencing and case-by-case review of original microbiology laboratory data.

Results

Of the analyzed 3,318 blood cultures, 2,107 yielded a pathogen by conventional techniques. Of these, 302 samples contained microbes not covered by Prove-it™ Sepsis, and an additional 137 cultures contained more than one organism. Sensitivity and specificity for Prove-it™ Sepsis were 94.7% and 98.7%, respectively. Of particular significance was the assay's faultless ability to differentiate MRSA from MSSA and from CNS. Furthermore, it provided results on average 1 day earlier than reference methods.

Conclusion

Prove-it™ Sepsis was considered to be a fast, robust, and high-performance diagnostic platform, which is easily implemented into everyday laboratory workflow. Both study sites identified cases where timely information provided by Prove-it™ Sepsis would have significantly improved patient management. Examples include more rational management and antibiotic choice subsequent to earlier differentiation of Gram-positive cocci in clumps into MRSA, MSSA, or CNS, and earlier speciation of Gram-negative organisms. Prove-it™ Sepsis is further configured for detection of Candida spp. and new bacterial targets. The assay now identifies 60 out of the 302 samples not covered during the evaluation, increasing the pathogen coverage from 86% to 89%. The earlier speciation provided by Prove-it™ Sepsis could contribute to faster, more evidence-based patient management and, thus, positive outcomes.

Authors’ Affiliations

(1)
Division of Clinical Microbiology, Helsinki University Hospital Laboratory HUSLAB
(2)
Department of Clinical Microbiology, , Centre for Infectious Diseases and International Health, University College London Hospitals NHS Foundation Trust, and University College London Medical School
(3)
Mobidiag Ltd

Copyright

© BioMed Central Ltd 2009

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