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Journal of Clinical Microbiology, November 2002, p. 4045-4050, Vol. 40, No. 11
0095-1137/02/$04.00+0     DOI: 10.1128/JCM.40.11.4045-4050.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Use of Epitope Mapping To Identify a PCR Template for Protein Amplification and Detection by Enzyme-Linked Immunosorbent Assay of Bovine Herpesvirus Type 1 Glycoprotein D

Tomy Joseph,1 Japhet Lyaku,2 Robert A. Fredrickson,3 Arnost Cepica,1 and Frederick S. B. Kibenge1*

Department of Pathology and Microbiology, Atlantic Veterinary College, University of Prince Edward Island, Charlottetown, Prince Edward Island, C1A 4P3,1 Origenix Technologies Inc., Laval, Quebec, H7V 4A9,2 Diagnostic Chemicals Limited, Charlottetown, Prince Edward Island, C1E 1B0, Canada3

Received 19 October 2001/ Returned for modification 9 March 2002/ Accepted 16 August 2002

Infection with bovine herpesvirus type 1 (BHV-1) occurs worldwide and causes serious economic losses due to the deaths of animals, abortions, decreased milk production, and loss of body weight. BHV-1 is frequently found in bovine semen and is transmitted through natural service and artificial insemination. The detection of BHV-1 in bovine semen is a long-standing problem in veterinary virology which is important in disease control schemes. In the present study, ordered deletions of the full-length BHV-1 glycoprotein open reading frame were used to identify an epitope recognized by a specific monoclonal antibody (MAb). A glycoprotein D fragment containing this epitope was then amplified using an in vitro protein amplification assay developed previously (J. Zhou, J. Lyaku, R. A. Fredrickson, and F. S. Kibenge, J. Virol. Methods 79:181-189, 1999), and the resulting peptide was detected by indirect enzyme-linked immunosorbent assay (ELISA) with the specific MAb. This method detected 0.0395 50% tissue culture infective dose of BHV-1 in raw bovine semen, which was 1,000-fold more sensitive than traditional PCR. We therefore conclude that this in vitro protein amplification assay combined with ELISA has superior sensitivity for direct virus detection in clinical samples.

* Corresponding author. Mailing address: Department of Pathology and Microbiology, Atlantic Veterinary College, University of Prince Edward Island, Charlottetown, P.E.I., C1A 4P3, Canada. Phone: (902) 566-0967. Fax: (902) 566-0851. E-mail: kibenge{at}upei.ca.

Journal of Clinical Microbiology, November 2002, p. 4045-4050, Vol. 40, No. 11
0095-1137/02/$04.00+0     DOI: 10.1128/JCM.40.11.4045-4050.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.





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Antimicrob. Agents Chemother. Clin. Microbiol. Rev.
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Copyright © 2002 by the American Society for Microbiology. All rights reserved.