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Evaluation of the Polymorphisms Associated with Tandem Repeats for Pseudomonas aeruginosa Strain Typing

Lucie Onteniente,¹ Sylvain Brisse,^2, Panayotis T. Tassios,³ and Gilles Vergnaud^1,4*

Institut de Génétique et Microbiologie, Université Paris Sud, 91405 Orsay cedex,¹ Centre d'Etudes du Bouchet, BP3, 91710 Vert le Petit, France,⁴ Eijkman-Winkler Institute, Utrecht University, 3584 CX Utrecht, The Netherlands,² Department of Microbiology, Medical School, National and Kapodestrian University of Athens, M. Asias 75, 115 27 Athens, Greece³

Received 24 April 2003/ Returned for modification 23 June 2003/ Accepted 19 August 2003

We report on the development of a scheme for the typing of Pseudomonas aeruginosa, multiple-locus variable number of tandem repeat (VNTR) analysis (MLVA). We first evaluated the polymorphisms of 201 tandem repeat loci selected from more than 3,000 such sequences present in strain PAO1 with a test collection of 12 genotypically distinct clinical strains. Seven VNTR loci which can be easily scored with the technology used here were identified and used to genotype a collection of 89 clinical isolates that had previously been classified into 46 ribotypes, including 2 widespread ribotypes. Seventy-one different MLVA genotypes could be distinguished. With only two exceptions, strains with identical ribotypes were grouped together upon cluster analysis of the MLVA data. The 27 isolates with the most frequent ribotype were divided into 14 MLVA types, and the 18 isolates with the second most frequent ribotype were divided into 15 MLVA types. Analysis of a subset of 17 strains belonging to the major ribotype by pulsed-field gel electrophoresis with the enzyme SpeI distinguished seven types, identical to the number of MLVA types in this subset. Our data show that MLVA typing of P. aeruginosa based on the first set of loci has a high discriminatory power. Because MLVA is highly reproducible and easily portable among laboratories, it represents a very promising tool for the molecular surveillance of P. aeruginosa. A free, online strain identification service based on the genotyping data produced herein has been developed.

Present address: Biodiversity of Emerging Bacterial Pathogens Unit, Institut Pasteur, 75724 Paris Cedex 15, France.

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