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Emetic toxin formation of Bacillus cereus is restricted to a single evolutionary lineage of closely related strains

¹ Lehrstuhl für Mikrobielle Ökologie, Department für Grundlagen der Biowissenschaften, Technische Universität München, Weihenstephaner Berg 3, D-85354 Freising, Germany
² Swedish Dairy Association, Scheelevaegen 18, 223 63 Lund, Sweden
³ Institut National de la Recherche Agronomique, UMR A408 Sécurité et Qualité des Produits d'Origine Végétale, INRA, Domaine Saint-Paul, Site Agroparc, F-84914 Avignon Cedex 9, France
⁴ Department of Pharmacology, Microbiology and Food Hygiene, The Norwegian School of Veterinary Science, Ullevalsveien 72, PO Box 8146, Dep., N-0033 Oslo, Norway
⁵ Dept for Applied Chemistry and Microbiology, College of Agriculture and Forestry at the University of Helsinki, Biocenter PO Box 56, Viikinkaari 9, FIN 00014 Helsinki University, Finland
⁶ Institute of Hygiene and Technology of Food of Animal Origin, Ludwig-Maximilians-Universität München, Veterinaerstr 13, D-80539 Munich, Germany

Correspondence
Siegfried Scherer
Siegfried.Scherer{at}wzw.tum.de

An in-depth polyphasic approach was applied to study the population structure of the human pathogen Bacillus cereus. To assess the intraspecific biodiversity of this species, which is the causative agent of gastrointestinal diseases, a total of 90 isolates from diverse geographical origin were studied by genetic [M13-PCR, random amplification of polymorphic DNA (RAPD), multilocus sequence typing (MLST)] and phenetic [Fourier transform Infrared (FTIR), protein profiling, biochemical assays] methods. The strain set included clinical strains, isolates from food remnants connected to outbreaks, as well as isolates from diverse food environments with a well documented strain history. The phenotypic and genotypic analysis of the compiled panel of strains illustrated a considerable diversity among B. cereus connected to diarrhoeal syndrome and other non-emetic food strains, but a very low diversity among emetic isolates. Using all typing methods, cluster analysis revealed a single, distinct cluster of emetic B. cereus strains. The isolates belonging to this cluster were neither able to degrade starch nor could they ferment salicin; they did not possess the genes encoding haemolysin BL (Hbl) and showed only weak or no haemolysis. In contrast, haemolytic-enterotoxin-producing B. cereus strains showed a high degree of heterogeneity and were scattered over different clusters when different typing methods were applied. These data provide evidence for a clonal population structure of cereulide-producing emetic B. cereus and indicate that emetic strains represent a highly clonal complex within a potentially panmictic or weakly clonal background population structure of the species. It may have originated only recently through acquisition of specific virulence factors such as the cereulide synthetase gene.

The GenBank/EMBL/DDBJ accession numbers for the sequences of the internal gene fragments used for MLST and for the sporulation stage III AB genes reported in this paper are AY762151–AY762213 and AY578317–AY578349, respectively.

Full details of the strains used in this study and additional genetic relationship data are available as supplementary data with the online version of this paper at http://mic.sgmjournals.org.

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