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Applied and Environmental Microbiology, January 2001, p. 185-189, Vol. 67, No. 1
0099-2240/01/$04.00+0 DOI: 10.1128/AEM.67.1.185-189.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.
Detection of Enterotoxic Bacillus cereus and
Bacillus thuringiensis Strains by PCR Analysis
Bjarne Munk
Hansen* and
Niels Bohse
Hendriksen
Department of Microbial Ecology and
Biotechnology, National Environmental Research Institute, DK-4000
Roskilde, Denmark
Received 9 May 2000/Accepted 5 October 2000
Many strains of Bacillus cereus cause gastrointestinal
diseases, and the closely related insect pathogen B. thuringiensis has also been involved in outbreaks of diarrhea.
The diarrheal types of diseases are attributed to enterotoxins. Two
different enterotoxic protein complexes, hemolysin BL (HBL) and
nonhemolytic enterotoxin (NHE), and an enterotoxic protein, enterotoxin
T, have been characterized, and the genes have been sequenced. PCR primers for the detection of these genes were deduced and used to
detect the genes in 22 B. cereus and 41 B. thuringiensis strains. At least one gene of each of the two
protein complexes HBL and NHE was detected in all of the B. thuringiensis strains, while six B. cereus strains
were devoid of all three HBL genes, three lacked at least two of the
three NHE genes, and one lacked all three. Five different sets of
primers were used for detection of the gene (bceT) encoding
enterotoxin T. The results obtained with these primer sets indicate
that bceT is widely distributed among B. cereus
and B. thuringiensis strains and that the gene varies in
sequence among different strains. PCR with the two primer sets
BCET1-BCET3 and BCET1-BCET4 unambiguously detected the bceT gene, as confirmed by Southern analysis. The occurrence of the genes
within the two complexes is significantly associated, while neither the
occurrence of the two complexes nor the occurrence of the
bceT gene is significantly associated in the 63 strains. We
suggest an approach for detection of enterotoxin-encoding genes in
B. cereus and B. thuringiensis based on PCR
analysis with the six primer sets for the detection of genes in the HBL
and NHE operons and with the BCET1, BCET3, and BCET4 primers for the
detection of bceT. PCR analysis of the 16S-23S rRNA gene
internal transcribed spacer region revealed identical patterns for all
strains studied.
*
Corresponding author. Mailing address: Department of
Microbial Ecology, National Environmental Research Institute, P.O. Box 358, Frederiksborgvej 399, DK-4000 Roskilde, Denmark. Phone: 45 46301200. Fax: 45 46301216. E-mail: bmh{at}dmu.dk.
Applied and Environmental Microbiology, January 2001, p. 185-189, Vol. 67, No. 1
0099-2240/01/$04.00+0 DOI: 10.1128/AEM.67.1.185-189.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.
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