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Applied and Environmental Microbiology, February 2004, p. 913-920, Vol. 70, No. 2
0099-2240/04/$08.00+0     DOI: 10.1128/AEM.70.2.913-920.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

Multi-Virulence-Locus Sequence Typing of Listeria monocytogenes

Wei Zhang,1* Bhushan M. Jayarao,2 and Stephen J. Knabel1

Department of Food Science,1 Department of Veterinary Science, The Pennsylvania State University, University Park, Pennsylvania 168022

Received 18 June 2003/ Accepted 7 November 2003

A multi-virulence-locus sequence typing (MVLST) scheme was developed for subtyping Listeria monocytogenes, and the results obtained using this scheme were compared to those of pulsed-field gel electrophoresis (PFGE) and the published results of other typing methods, including ribotyping (RT) and multilocus sequence typing (MLST). A set of 28 strains (eight different serotypes and three known genetic lineages) of L. monocytogenes was selected from a strain collection (n > 1,000 strains) to represent the genetic diversity of this species. Internal fragments (ca. 418 to 469 bp) of three virulence genes (prfA, inlB, and inlC) and three virulence-associated genes (dal, lisR, and clpP) were sequenced and analyzed. Multiple DNA sequence alignment identified 10 (prfA), 19 (inlB), 13 (dal), 10 (lisR), 17 (inlC), and 16 (clpP) allelic types and a total of 28 unique sequence types. Comparison of MVLST with automated EcoRI-RT and PFGE with ApaI enzymatic digestion showed that MVLST was able to differentiate strains that were indistinguishable by RT (13 ribotypes; discrimination index = 0.921) or PFGE (22 profiles; discrimination index = 0.970). Comparison of MVLST with housekeeping-gene-based MLST analysis showed that MVLST provided higher discriminatory power for serotype 1/2a and 4b strains than MLST. Cluster analysis based on the intragenic sequences of the selected virulence genes indicated a strain phylogeny closely related to serotypes and genetic lineages. In conclusion, MVLST may improve the discriminatory power of MLST and provide a convenient tool for studying the local epidemiology of L. monocytogenes.

* Corresponding author. Mailing address: Department of Food Science, The Pennsylvania State University, 118 Borland Laboratory, University Park, PA 16802. Phone: (814) 865-9716. Fax: (814) 863-6132. E-mail: wuz100{at}psu.edu.

Applied and Environmental Microbiology, February 2004, p. 913-920, Vol. 70, No. 2
0099-2240/04/$08.00+0     DOI: 10.1128/AEM.70.2.913-920.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.



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