Immunogenicity of recombinant Omp31 from Brucella melitensis in rams and serum bactericidal activity against B. ovis.
Detergent-extracted recombinant Omp31 (rOmp31 extract) from Brucella melitensis produced in Escherichia coli was previously identified as a protective immunogen against B. ovis in mice. In this study, we evaluated the immunogenicity of rOmp31extract in rams. This immunogen was emulsified in an oil adjuvant and administered three times with 4 and 8 weeks intervals. Antibody response was measured in serum by whole B. ovis ELISA. Specific antibodies to purified rOmp31 (pET-Omp31) were detected by Western blotting and indirect ELISA. In addition, isotype specific antibodies were measured in tears. Serum bactericidal activity against B. ovis in the presence of complement was measured in vitro. Cellular immune response was explored by intradermal testing with purified rOmp31. Immunization with rOmp31 extract induced IgG specific antibodies in serum able to bind to whole B. ovis cells. Furthermore, strong inhibition in a competitive ELISA (with an Omp31-specific monoclonal antibody) suggested that a proportion of Omp31-specific antibodies were directed against a loop containing a protective epitope. Serum antibodies killed efficiently B. ovis in vitro in the presence of either guinea pig or ovine serum. Tears had both IgG and IgA antibodies to equivalent titers. Finally, immunized rams showed skin reactivity to Omp31. These data demonstrate that B. melitensis Omp31, a protective antigen identified in the mouse model, induces antibody and cellular immune mechanisms in sheep.
Laboratorio de Inmunoquímica y Biotecnología, Departamento de Sanidad Animal y Medicina Preventiva, Facultad de Ciencias Veterinarias, Universidad Nacional del Centro de la Provincia de Buenos Aires, 7000 Tandil, Argentina. silmares@vet.unicen.edu.ar
Silvia M
SM
Estein
Pablo C
PC
Cheves
María A
MA
Fiorentino
Juliana
J
Cassataro
Fernando A
FA
Paolicchi
Raúl A
RA
Bowden
D000818:Animals; D000907:Antibodies, Bacterial / Q000032:analysis* / Q000097:blood; D001425:Bacterial Outer Membrane Proteins / Q000276:immunology*; D001770:Blood Bactericidal Activity / Q000276:immunology; D015153:Blotting, Western / Q000379:methods / Q000662:veterinary; D017347:Brucella melitensis / Q000276:immunology*; D042061:Brucella ovis / Q000276:immunology*; D002006:Brucellosis / Q000276:immunology / Q000382:microbiology / Q000662:veterinary*; D003165:Complement System Proteins / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods / Q000662:veterinary; D004926:Escherichia coli; D008297:Male; D011897:Random Allocation; D011994:Recombinant Proteins / Q000276:immunology; D012756:Sheep; D012757:Sheep Diseases / Q000382:microbiology*; D013666:Tears / Q000276:immunology
D000907:Antibodies, Bacterial; D001425:Bacterial Outer Membrane Proteins; C101108:Omp31 protein, Brucella melitensis; D011994:Recombinant Proteins; D003165:Complement System Proteins