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Infection and Immunity, December 2005, p. 8079-8088, Vol. 73, No. 12
0019-9567/05/$08.00+0     doi:10.1128/IAI.73.12.8079-8088.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

Vaccination with the Recombinant Brucella Outer Membrane Protein 31 or a Derived 27-Amino-Acid Synthetic Peptide Elicits a CD4+ T Helper 1 Response That Protects against Brucella melitensis Infection

Juliana Cassataro,1,2* Silvia M. Estein,3 Karina A. Pasquevich,1,2 Carlos A. Velikovsky,2,{dagger} Silvia de la Barrera,4 Raúl Bowden,3,{ddagger} Carlos A. Fossati,2 and Guillermo H. Giambartolomei1,2

Laboratorio de Inmunogenética, Hospital de Clínicas "José de San Martín," Facultad de Medicina, Universidad de Buenos Aires, Buenos Aires, Argentina,1 Instituto de Estudios de la Inmunidad Humoral (IDEHU-CONICET), Facultad de Farmacia y Bioquímica, Universidad de Buenos Aires, Buenos Aires, Argentina,2 Laboratorio de Inmunología, Departamento de Sanidad Animal y Medicina Preventiva, Facultad de Ciencias Veterinarias, Universidad Nacional del Centro de la Provincia de Buenos Aires, Tandil, Argentina,3 Departamento de Inmunologia, Instituto de Investigaciones Hematológicas, Academia Nacional de Medicina, Buenos Aires, Argentina4

Received 8 July 2005/ Returned for modification 16 August 2005/ Accepted 4 September 2005

The immunogenicity and protective efficacy of the recombinant 31-kDa outer membrane protein from Brucella melitensis (rOmp31), administered with incomplete Freund's adjuvant, were evaluated in mice. Immunization of BALB/c mice with rOmp31 conferred protection against B. ovis and B. melitensis infection. rOmp31 induced a vigorous immunoglobulin G (IgG) response, with higher IgG1 than IgG2 titers. In addition, spleen cells from rOmp31-immunized mice produced interleukin 2 (IL-2) and gamma interferon, but not IL-10 or IL-4, after in vitro stimulation with rOmp31, suggesting the induction of a T helper 1 (Th1) response. Splenocytes from rOmp31-vaccinated animals also induced a specific cytotoxic-T-lymphocyte activity, which led to the in vitro lysis of Brucella-infected macrophages. In vitro T-cell subset depletion indicated that rOmp31 immunization elicited specific CD4+ T cells that secrete IL-2 and gamma interferon, while CD8+ T cells induced cytotoxic-T-lymphocyte activity. In vivo depletion of T-cell subsets showed that the rOmp31-elicited protection against B. melitensis infection is mediated by CD4+ T cells while the contribution of CD8+ T cells may be limited. We then evaluated the immunogenicity and protective efficacy of a known exposed region from Omp31 on the Brucella membrane, a peptide that contains amino acids 48 to 74 of Omp31. Immunization with the synthetic peptide in adjuvant did not elicit a specific humoral response but elicited a Th1 response mediated by CD4+ T cells. The peptide in adjuvant induced levels of protection similar to those induced by rOmp31 against B. melitensis but less protection than was induced by rOmp31 against B. ovis. Our results indicate that rOmp31 could be a useful candidate for the development of subunit vaccines against B. melitensis and B. ovis.

* Corresponding author. Mailing address: Laboratorio de Inmunogenética, Hospital de Clínicas "José de San Martín," Facultad de Medicina, UBA, Córdoba 2351 3er Piso Sala 4 (1120), Buenos Aires, Argentina. Phone: (54-11) 5950-8755. Fax: (54-11) 5950-8758. E-mail: jucassat{at}ffyb.uba.ar.

Editor: J. D. Clements

{dagger} Present address: Center for Advanced Research in Biotechnology, Rockville, Md.

{ddagger} Deceased.

Infection and Immunity, December 2005, p. 8079-8088, Vol. 73, No. 12
0019-9567/05/$08.00+0     doi:10.1128/IAI.73.12.8079-8088.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.



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