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Infection and Immunity, December 2001, p. 7851-7857, Vol. 69, No. 12
0019-9567/01/$04.00+0 DOI: 10.1128/IAI.69.12.7851-7857.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.
Evaluation of a Tetracycline-Inducible Promoter in
Staphylococcus aureus In Vitro and In Vivo and Its
Application in Demonstrating the Role of sigB in
Microcolony Formation
B. T.
Bateman,
N.
P.
Donegan,
T. M.
Jarry,
M.
Palma, and
A. L.
Cheung*
Department of Microbiology and Immunology,
Dartmouth Medical School, Hanover, New Hampshire 03755
Received 2 July 2001/Returned for modification 13 August
2001/Accepted 30 August 2001
An inducible promoter system provides a powerful tool for studying
the genetic basis for virulence. A variety of inducible systems have
been used in other organisms, including
pXyl-xylR-inducible promoter, the
pSpac-lacI system, and the arabinose-inducible
PBAD promoter, but each of these systems has limitations in
its application to Staphylococcus aureus. In this study,
we demonstrated the efficacy of a tetracycline-inducible promoter
system in inducing gene expression in S. aureus in vitro
and inside epithelial cells as well as in an animal model of infection.
Using the xyl/tetO
promoter::gfpuvr fusion carried on
a shuttle plasmid, we demonstrated that dose-dependant tetracycline
induction, as measured by bacterial fluorescence, occurred in each of
the above environments while basal activation under noninduced
conditions remained low. To ascertain how the system can be used to
elucidate the genetic basis of a pathogenic phenotype, we cloned the
sigB gene downstream of the inducible promoter.
Induction of SigB expression led to dose-dependent attachment of the
tested strain to polystyrene microtiter wells. Additionally, bacterial
microcolony formation, an event preceding mature biofilm formation,
also increased with tetracycline induction of SigB.
*
Corresponding author. Mailing address: Department of
Microbiology, Dartmouth Medical School, Hanover, NH 03755. Phone: (603) 650-1340. Fax: (603) 650-1362. E-mail:
ambrose.cheung{at}dartmouth.edu.
Infection and Immunity, December 2001, p. 7851-7857, Vol. 69, No. 12
0019-9567/01/$04.00+0 DOI: 10.1128/IAI.69.12.7851-7857.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.
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