Toward an international standard for PCR-based detection of Escherichia coli O157 - Part 1. Assay development and multi-center validation

A. Abdulmawjood, M. Bulte, N. Cook, S. Roth, H. Schonenbrucher, Jeffrey Hoorfar

Research output: Contribution to journalJournal articleResearchpeer-review

Abstract

As part of a major European research project, a diagnostic PCR assay, including an internal amplification control, was developed and validated in a collaborative trial for the detection of Escherichia coli O157. The assay is based on amplification of sequences of the rJbE O157 gene. The collaborative trial, including 12 international laboratories, was carried out in two phases: phase (a) was performed with identical PCR reagents, including the internal control, provided by the sending laboratory; phase (b) was performed on the same samples and internal control but using in-house PCR reagents of own choice. Phase (a) showed an inclusivity (detection of target strains) of 96.8% and the exclusivity (negative response from nontarget strains) was 100%. The overall performance resulted of phase (a) in an accordance of 98.8, concordance of 98.6, and a concordance odds ratio of 1.11. Phase (b) results showed an accuracy of 100% with all partners and by using different polymerase types and thermocycler models. This indicates that the assay, under consideration as an international standard, was just as reproducible between laboratories, as repeatable within a laboratory. The assay is taken further for validation on carcass-rinse samples.
Original languageEnglish
JournalJournal of Microbiological Methods
Volume55
Issue number3
Pages (from-to)775-786
ISSN0167-7012
DOIs
Publication statusPublished - 2003

Keywords

  • Escherichia coli O157
  • standard
  • validation
  • rfbE gene

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