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Journal of Bacteriology, August 2009, p. 4798-4806, Vol. 191, No. 15
0021-9193/09/$08.00+0     doi:10.1128/JB.00212-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Contribution of Citrulline Ureidase to Francisella tularensis Strain Schu S4 Pathogenesis

Manish Mahawar, Girish S. Kirimanjeswara, Dennis W. Metzger, and Chandra Shekhar Bakshi^*

Center for Immunology and Microbial Disease, Albany Medical College, Albany, New York 12208

Received 17 February 2009/ Accepted 21 May 2009

The citrulline ureidase (CTU) activity has been shown to be associated with highly virulent Francisella tularensis strains, including Schu S4, while it is absent in avirulent or less virulent strains. A definitive role of the ctu gene in virulence and pathogenesis of F. tularensis Schu S4 has not been assessed; thus, an understanding of the significance of this phenotype is long overdue. CTU is a carbon-nitrogen hydrolase encoded by the citrulline ureidase (ctu) gene (FTT0435) on the F. tularensis Schu S4 genome. In the present study, we evaluated the contribution of the ctu gene in the virulence of category A agent F. tularensis Schu S4 by generating a nonpolar deletion mutant, the ctu mutant. The deletion of the ctu gene resulted in loss of CTU activity, which was restored by transcomplementing the ctu gene. The ctu mutant did not exhibit any growth defect under acellular growth conditions; however, it was impaired for intramacrophage growth in resting as well as gamma interferon-stimulated macrophages. The ctu mutant was further tested for its virulence attributes in a mouse model of respiratory tularemia. Mice infected intranasally with the ctu mutant showed significantly reduced bacterial burden in the lungs, liver, and spleen compared to wild-type (WT) Schu S4-infected mice. The reduced bacterial burden in mice infected with the ctu mutant was also associated with significantly lower histopathological scores in the lungs. Mice infected with the ctu mutant succumbed to infection, but they survived longer and showed significantly extended median time to death compared to that shown by WT Schu S4-infected mice. To conclude, this study demonstrates that ctu contributes to intracellular survival, in vivo growth, and pathogenesis. However, ctu is not an absolute requirement for the virulence of F. tularensis Schu S4 in mice.

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* Corresponding author. Mailing address: Center for Immunology and Microbial Disease, Albany Medical College, MC-151, 47 New Scotland Avenue, Albany, NY 12208-3479. Phone: (518) 262-6263. Fax: (518) 262-6161. E-mail: bakshis{at}mail.amc.edu

Published ahead of print on 5 June 2009.

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Journal of Bacteriology, August 2009, p. 4798-4806, Vol. 191, No. 15
0021-9193/09/$08.00+0     doi:10.1128/JB.00212-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

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