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Infection and Immunity, September 2003, p. 5130-5138, Vol. 71, No. 9
0019-9567/03/$08.00+0     DOI: 10.1128/IAI.71.9.5130-5138.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Early Induction of Humoral and Cellular Immune Responses during Experimental Mycobacterium avium subsp. paratuberculosis Infection of Calves

W. R. Waters,1* J. M. Miller,2 M. V. Palmer,1 J. R. Stabel,1 D. E. Jones,3 K. A. Koistinen,3 E. M. Steadham,3 M. J. Hamilton,4 W. C. Davis,4 and J. P. Bannantine1

Bacterial Diseases of Livestock Research Unit,1 Virus and Prion Diseases Research Unit, National Animal Disease Center, Agricultural Research Service, United States Department of Agriculture, Ames, Iowa 50010,2 Department of Veterinary Pathology, Iowa State University College of Veterinary Medicine, Ames, Iowa, 50011,3 Department of Veterinary Microbiology and Pathology, Washington State University College of Veterinary Medicine, Pullman, Washington 991644

Received 23 April 2003/ Returned for modification 7 June 2003/ Accepted 27 June 2003

Johne's disease (paratuberculosis) of cattle is widespread and causes significant economic losses for producers due to decreased production and poor health of affected animals. The chronic nature of the disease and the lack of a reproducible model of infection hinder research efforts. In the present study, instillation of Mycobacterium avium subsp. paratuberculosis into the tonsillar crypts of neonatal calves resulted in peripheral colonization as detected by antemortem culture of feces and postmortem (320 days postchallenge) culture of intestinal tissues. Antigen-specific blastogenic, gamma interferon (IFN-{gamma}), and nitric oxide responses by blood mononuclear cells from infected calves exceeded prechallenge responses beginning 194 days postchallenge. Upon in vitro stimulation with paratuberculosis antigens, CD4+ cells from infected calves proliferated, produced IFN-{gamma}, and increased expression of CD26 and CD45RO (indicative of an activated memory phenotype). Utilizing a lipoarabinomannan-based enzyme-linked immunosorbent assay, specific serum immunoglobulin was detected as early as 134 days postchallenge and generally increased after this time point. Two antigens of ~50 and ~60 kDa were particularly immunodominant early in infection, as shown by immunoblot with serum collected within 2 weeks postchallenge. Findings indicate that the intratonsillar inoculation route will prove useful as an experimental model for paratuberculosis infection. Additionally, this study confirms that mycobacteria-specific antibody is detectable early in the course of experimental Johne's disease, even preceding the development of specific cell-mediated responses.

* Corresponding author. Mailing address: United States Department of Agriculture, Agricultural Research Service, National Animal Disease Center, P.O. Box 70, Ames, IA 50010-0070. Phone: (515) 663-7756. Fax: (515) 663-7458. E-mail: rwaters{at}nadc.ars.usda.gov.

Editor: W. A. Petri, Jr.

Infection and Immunity, September 2003, p. 5130-5138, Vol. 71, No. 9
0019-9567/03/$08.00+0     DOI: 10.1128/IAI.71.9.5130-5138.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



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