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Infection and Immunity, December 2004, p. 7265-7274, Vol. 72, No. 12
0019-9567/04/$08.00+0     DOI: 10.1128/IAI.72.12.7265-7274.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

Characterization of Immunodominant and Potentially Protective Epitopes of Mannheimia haemolytica Serotype 1 Outer Membrane Lipoprotein PlpE

Sahlu Ayalew,* Anthony W. Confer, and Emily R. Blackwood

Department of Veterinary Pathobiology, College of Veterinary Medicine, Oklahoma State University, Stillwater, Oklahoma

Received 22 June 2004/ Returned for modification 27 July 2004/ Accepted 11 August 2004

Mannheimia haemolytica serotype 1 (S1) is the most common bacterial isolate found in shipping fever pneumonia in beef cattle. Currently used vaccines against M. haemolytica do not provide complete protection against the disease. Research with M. haemolytica outer membrane proteins (OMPs) has shown that antibodies to one particular OMP from S1, PlpE, may be important in immunity. In a recently published work, members of our laboratory showed that recombinant PlpE (rPlpE) is highly immunogenic when injected subcutaneously into cattle and that the acquired immunity markedly enhanced resistance to experimental challenge (A. W. Confer, S. Ayalew, R. J. Panciera, M. Montelongo, L. C. Whitworth, and J. D. Hammer, Vaccine 21:2821-2829, 2003). The objective of this work was to identify epitopes of PlpE that are responsible for inducing the immune response. Western blot analysis of a series of rPlpE with nested deletions on both termini with bovine anti-PlpE hyperimmune sera showed that the immunodominant region is located close to the N terminus of PlpE. Fine epitope mapping, in which an array of overlapping 13-mer synthetic peptides attached to a derivatized cellulose membrane was probed with various affinity-purified anti-PlpE antibodies, identified eight highly reactive regions, of which region 2 (R2) was identified as the specific epitope. The R2 region is comprised of eight imperfect repeats of a hexapeptide (QAQNAP) and is located between residues 26 and 76. Complement-mediated bactericidal activity of affinity-purified anti-PlpE bovine antibodies confirmed that antibodies directed against the R2 region are effective in killing M. haemolytica.

* Corresponding author. Mailing address: Department of Veterinary Pathobiology, College of Veterinary Medicine, Oklahoma State University, Stillwater, OK 74078. Phone: (405) 744-3593. Fax: (405) 744-5275. E-mail: ayalew{at}okstate.edu.

Editor: D. L. Burns

Infection and Immunity, December 2004, p. 7265-7274, Vol. 72, No. 12
0019-9567/04/$08.00+0     DOI: 10.1128/IAI.72.12.7265-7274.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.





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