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Infection and Immunity, July 2003, p. 3920-3926, Vol. 71, No. 7
0019-9567/03/$08.00+0     DOI: 10.1128/IAI.71.7.3920-3926.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Identification of CD8 T-Lymphocyte Epitopes in OmpB of Rickettsia conorii

Zhen Li, C. Marcela Díaz-Montero, Gustavo Valbuena, Xue-Jie Yu, Juan P. Olano, Hui-Min Feng, and David H. Walker^*

Department of Pathology, WHO Collaborating Center for Tropical Diseases, University of Texas Medical Branch at Galveston, Galveston, Texas 77555-0609

Received 6 February 2003/ Returned for modification 12 March 2003/ Accepted 12 April 2003

The 1.2-kb DNA fragment of the Rickettsia conorii outer membrane protein B gene (OmpB_451-846) was subcloned using site-specific PCR primers and expressed as six smaller fragments: OmpB_458-652, OmpB_595-744, OmpB_595-654, OmpB_645-692, OmpB_689-744, and OmpB_739-848. NCTC cells transfected with a mammalian expression vector expressing the fragments OmpB_689-744 and OmpB_739-848 stimulated immune anti-R. conorii CD8 T lymphocytes, suggesting the presence of CD8 T-lymphocyte-stimulating epitopes on these fragments. In order to further characterize the CD8 T-lymphocyte-stimulatory elements, CD8 T-lymphocyte epitopes on OmpB_689-744 and OmpB_739-848 were mapped by overlapping synthetic peptides. The ability of these synthetic peptides to stimulate immune CD8 T lymphocytes was determined by gamma interferon (IFN-) production and cell proliferation after incubation with simian virus 40-transformed murine vascular endothelial cells in the presence of a 20 µM solution of each synthetic peptide. Five synthetic peptides, SKGVNVDTV (OmpB_708-716), ANVGSFVFN (OmpB_735-743), IVSGTVGGQ (OmpB_749-757), ANSTLQIGG (OmpB_789-797), and IVEFVNTGP (OmpB_812-820), induced secretion of IFN- at significantly higher levels than the controls. Three of these five peptides, SKGVNVDTV (OmpB_708-716), ANSTLQIGG (OmpB_789-797), and IVEFVNTGP (OmpB_812-820), also stimulated the proliferation of immune CD8 T lymphocytes. Significantly higher levels of specific cytotoxic T-lymphocyte killing were observed with the same three synthetic peptides, SKGVNVDTV (OmpB_708-716), ANSTLQIGG (OmpB_789-797), and IVEFVNTGP (OmpB_812-820).

Editor: W. A. Petri, Jr.

Present address: Department of Immunology, University of Texas M. D. Anderson Cancer Center, Houston, TX 77030.

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