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Journal of Clinical Microbiology, October 2009, p. 3121-3128, Vol. 47, No. 10
0095-1137/09/$08.00+0     doi:10.1128/JCM.00267-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Improved Multiple-Locus Variable-Number Tandem-Repeat Assay for Staphylococcus aureus Genotyping, Providing a Highly Informative Technique Together with Strong Phylogenetic Value {triangledown} ,{dagger}

Christine Pourcel,1* Katia Hormigos,1 Lucie Onteniente,1 Olga Sakwinska,2 Ruud H. Deurenberg,3 and Gilles Vergnaud1,4

Université Paris-Sud 11, CNRS, UMR8621, Institut de Génétique et Microbiologie, 91405 Orsay, France,1 Department of Fundamental Microbiology, University of Lausanne, 1015 Lausanne, Switzerland,2 Department of Medical Microbiology, Maastricht University Medical Center, P.O. Box 5800, 6202 AZ Maastricht, The Netherlands,3 DGA/MRIS, Mission pour la Recherche et l'Innovation Scientifique, 92221 Bagneux, France4

Received 6 February 2009/ Returned for modification 23 April 2009/ Accepted 19 August 2009

We describe an improved multiple-locus variable-number tandem-repeat (VNTR) analysis (MLVA) scheme for genotyping Staphylococcus aureus. We compare its performance to those of multilocus sequence typing (MLST) and spa typing in a survey of 309 strains. This collection includes 87 epidemic methicillin-resistant S. aureus (MRSA) strains of the Harmony collection, 75 clinical strains representing the major MLST clonal complexes (CCs) (50 methicillin-sensitive S. aureus [MSSA] and 25 MRSA), 135 nasal carriage strains (133 MSSA and 2 MRSA), and 13 published S. aureus genome sequences. The results show excellent concordance between the techniques' results and demonstrate that the discriminatory power of MLVA is higher than those of both MLST and spa typing. Two hundred forty-two genotypes are discriminated with 14 VNTR loci (diversity index, 0.9965; 95% confidence interval, 0.9947 to 0.9984). Using a cutoff value of 45%, 21 clusters are observed, corresponding to the CCs previously defined by MLST. The variability of the different tandem repeats allows epidemiological studies, as well as follow-up of the evolution of CCs and the identification of potential ancestors. The 14 loci can conveniently be analyzed in two steps, based upon a first-line simplified assay comprising a subset of 10 loci (panel 1) and a second subset of 4 loci (panel 2) that provides higher resolution when needed. In conclusion, the MLVA scheme proposed here, in combination with available on-line genotyping databases (including http://mlva.u-psud.fr/), multiplexing, and automatic sizing, can provide a basis for almost-real-time large-scale population monitoring of S. aureus.

* Corresponding author. Mailing address: Bâtiment 400, Laboratoire GPMS, Institut de Génétique et Microbiologie, Université Paris Sud, 91405 Orsay cedex, France. Phone: 33 1 69 15 30 01. Fax: 33 1 69 15 66 78. E-mail: christine.pourcel{at}u-psud.fr

{triangledown} Published ahead of print on 26 August 2009.

{dagger} Supplemental material for this article may be found at http://jcm.asm.org/.

Journal of Clinical Microbiology, October 2009, p. 3121-3128, Vol. 47, No. 10
0095-1137/09/$08.00+0     doi:10.1128/JCM.00267-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.





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