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Infection and Immunity, July 2008, p. 3037-3044, Vol. 76, No. 7
0019-9567/08/$08.00+0     doi:10.1128/IAI.01737-07
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

Characterization of a Helicobacter hepaticus putA Mutant Strain in Host Colonization and Oxidative Stress ^,

Navasona Krishnan,¹ Alan R. Doster,² Gerald E. Duhamel,^2, and Donald F. Becker^1*

Department of Biochemistry, University of Nebraska, Lincoln, Nebraska 68588,¹ Veterinary and Biomedical Science Department, University of Nebraska, Lincoln, Nebraska 68588²

Received 27 December 2007/ Returned for modification 28 January 2008/ Accepted 28 April 2008

Helicobacter hepaticus is a gram-negative, spiral-shaped microaerophilic bacterium associated with chronic intestinal infection leading to hepatitis and colonic and hepatic carcinomas in susceptible strains of mice. In the closely related human pathogen Helicobacter pylori, L-proline is a preferred respiratory substrate and is found at significantly high levels in the gastric juice of infected patients. A previous study of the proline catabolic PutA flavoenzymes from H. pylori and H. hepaticus revealed that Helicobacter PutA generates reactive oxygen species during proline oxidation by transferring electrons from reduced flavin to molecular oxygen. We further explored the preference for proline as a respiratory substrate and the potential impact of proline metabolism on the redox environment in Helicobacter species during host infection by disrupting the putA gene in H. hepaticus. The resulting putA knockout mutant strain was characterized by oxidative stress analysis and mouse infection studies. The putA mutant strain of H. hepaticus exhibited increased proline levels and resistance to oxidative stress relative to that of the wild-type strain, consistent with proline's role as an antioxidant. The significant increase in stress resistance was attributed to higher proline content, as no upregulation of antioxidant genes was observed for the putA mutant strain. The wild-type and putA mutant H. hepaticus strains displayed similar levels of infection in mice, but in mice challenged with the putA mutant strain, significantly reduced inflammation was observed, suggesting a role for proline metabolism in H. hepaticus pathogenicity in vivo.

Published ahead of print on 5 May 2008.

Supplemental material for this article may be found at http://iai.asm.org/.

Editor: B. A. McCormick

Present address: Department of Biomedical Sciences, Section of Anatomic Pathology, College of Veterinary Medicine, Cornell University, T4 012A VRT Road, Box 11, Ithaca, NY 14853-6401.