This Article Full Text Full Text (PDF) Other Versions of this Article: JCM.01361-06v1 45/2/370    most recent Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Ballmer, K. Articles by Hächler, H. Search for Related Content PubMed PubMed Citation Articles by Ballmer, K. Articles by Hächler, H.

 Previous Article  |  Next Article 

Journal of Clinical Microbiology, February 2007, p. 370-379, Vol. 45, No. 2
0095-1137/07/$08.00+0     doi:10.1128/JCM.01361-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Fast DNA Serotyping of Escherichia coli by Use of an Oligonucleotide Microarray

Institute of Veterinary Bacteriology, NENT, Vetsuisse-Faculty, University of Berne, CH-3001 Berne, Switzerland,¹ Clondiag Chip Technologies GmbH, D-07743 Jena, Germany²

Received 3 July 2006/ Returned for modification 9 August 2006/ Accepted 6 November 2006

Classical antibody-based serotyping of Escherichia coli is an important method in diagnostic microbiology for epidemiological purposes, as well as for a rough virulence assessment. However, serotyping is so tedious that its use is restricted to a few reference laboratories. To improve this situation we developed and validated a genetic approach for serotyping based on the microarray technology. The genes encoding the O-antigen flippase (wzx) and the O-antigen polymerase (wzy) were selected as target sequences for the O antigen, whereas fliC and related genes, which code for the flagellar monomer, were chosen as representatives for the H phenotype. Starting with a detailed bioinformatic analysis and oligonucleotide design, an ArrayTube-based assay was established: a fast and robust DNA extraction method was coupled with a site-specific, linear multiplex labeling procedure and hybridization analysis of the biotinylated amplicons. The microarray contained oligonucleotide DNA probes, each in duplicate, representing 24 of the epidemiologically most relevant of the over 180 known O antigens (O antigens 4, 6 to 9, 15, 26, 52, 53, 55, 79, 86, 91, 101, 103, 104, 111, 113, 114, 121, 128, 145, 157, and 172) as well as 47 of the 53 different H antigens (H antigens 1 to 12, 14 to 16, 18 to 21, 23 to 34, 37 to 43, 45, 46, 48, 49, 51 to 54, and 56). Evaluation of the microarray with a set of defined strains representing all O and H serotypes covered revealed that it has a high sensitivity and a high specificity. All of the conventionally typed 24 O groups and all of the 47 H serotypes were correctly identified. Moreover, strains which were nonmotile or nontypeable by previous serotyping assays yielded unequivocal results with the novel ArrayTube assay, which proved to be a valuable alternative to classical serotyping, allowing processing of single colonies within a single working day.

Published ahead of print on 15 November 2006.

This article has been cited by other articles:

• Anjum, M. F., Mafura, M., Slickers, P., Ballmer, K., Kuhnert, P., Woodward, M. J., Ehricht, R. (2007). Pathotyping Escherichia coli by Using Miniaturized DNA Microarrays. Appl. Environ. Microbiol. 73: 5692-5697 [Abstract] [Full Text]