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Applied and Environmental Microbiology, February 2006, p. 1569-1578, Vol. 72, No. 2
0099-2240/06/$08.00+0     doi:10.1128/AEM.72.2.1569-1578.2006
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

Multiple-Locus Sequence Typing Analysis of Bacillus cereus and Bacillus thuringiensis Reveals Separate Clustering and a Distinct Population Structure of Psychrotrophic Strains

Alexei Sorokin,^1* Benjamin Candelon,^1, Kévin Guilloux,¹ Nathalie Galleron,¹ Natalia Wackerow-Kouzova,^1, S. Dusko Ehrlich,¹ Denis Bourguet,^2, and Vincent Sanchis²

Génétique Microbienne, INRA, Domaine de Vilvert, 78352 Jouy en Josas cedex,¹ Génétique Microbienne et Environnement, INRA La Minière, 78285 Guyancourt cedex, France²

Received 6 September 2005/ Accepted 8 November 2005

We used multilocus sequence typing (MLST) to characterize phylogenetic relationships for a collection of Bacillus cereus group strains isolated from forest soil in the Paris area during a mild winter. This collection contains multiple strains isolated from the same soil sample and strains isolated from samples from different sites. We characterized 115 strains of this collection and 19 other strains based on the sequences of the clpC, dinB, gdpD, panC, purF, and yhfL loci. The number of alleles ranged from 36 to 53, and a total of 93 allelic profiles or sequence types were distinguished. We identified three major strain clusters—C, T, and W—based on the comparison of individual gene sequences or concatenated sequences. Some less representative clusters and subclusters were also distinguished. Analysis of the MLST data using the concept of clonal complexes led to the identification of two, five, and three such groups in clusters C, T, and W, respectively. Some of the forest isolates were closely related to independently isolated psychrotrophic strains. Systematic testing of the strains of this collection showed that almost all the strains that were able to grow at a low temperature (6°C) belonged to cluster W. Most of these strains, including three independently isolated strains, belong to two clonal complexes and are therefore very closely related genetically. These clonal complexes represent strains corresponding to the previously identified species Bacillus weihenstephanensis. Most of the other strains of our collection, including some from the W cluster, are not psychrotrophic. B. weihenstephanensis (cluster W) strains appear to comprise an effectively sexual population, whereas Bacillus thuringiensis (cluster T) and B. cereus (cluster C) have clonal population structures.

Present address: Section of Infectious Diseases, Department of Internal Medicine, Yale University School of Medicine, 1 Gilbert Street, TAC S140, New Haven, CT 06510.

Present address: Agrophysics Research Institute, Grazhdansky pr. 14, 195220 St. Petersburg, Russia.

Present address: Centre de Biologie et de Gestion des Populations (CBGP), Campus International de Baillarguet, CS 30 016, 34988 Montferrier/Lez cedex, France.

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